First evidence of systemic infection and specific cytotoxic T lymphocyte immune response evoked by oral infection with recombinant Salmonella enterica serovar Albany-Ovalbumin in C57BL/6 mice

array:24 ["pii" => "S0325754122000529" "issn" => "03257541" "doi" => "10.1016/j.ram.2022.05.008" "estado" => "S300" "fechaPublicacion" => "2022-10-01" "aid" => "502" "copyright" => "Asociación Argentina de Microbiología" "copyrightAnyo" => "2022" "documento" => "article" "crossmark" => 1 "licencia" => "http://creativecommons.org/licenses/by-nc-nd/4.0/" "subdocumento" => "fla" "cita" => "Rev Argent Microbiol. 2022;54:282-7" "abierto" => array:3 [ "ES" => true "ES2" => true "LATM" => true ] "gratuito" => true "lecturas" => array:1 [ "total" => 0 ] "itemSiguiente" => array:19 [ "pii" => "S032575412200027X" "issn" => "03257541" "doi" => "10.1016/j.ram.2022.04.002" "estado" => "S300" "fechaPublicacion" => "2022-10-01" "aid" => "493" "copyright" => "Asociación Argentina de Microbiología" "documento" => "article" "crossmark" => 1 "licencia" => "http://creativecommons.org/licenses/by-nc-nd/4.0/" "subdocumento" => "fla" "cita" => "Rev Argent Microbiol. 2022;54:288-92" "abierto" => array:3 [ "ES" => true "ES2" => true "LATM" => true ] "gratuito" => true "lecturas" => array:1 [ "total" => 0 ] "en" => array:14 [ "idiomaDefecto" => true "cabecera" => "Brief report" "titulo" => "Emergence and clonal expansion of Klebsiella pneumoniae ST307, simultaneously producing KPC-3 and NDM-1" "tienePdf" => "en" "tieneTextoCompleto" => "en" "tieneResumen" => array:3 [ 0 => "en" 1 => "en" 2 => "es" ] "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "288" "paginaFinal" => "292" ] ] "titulosAlternativos" => array:1 [ "es" => array:1 [ "titulo" => "Emergencia y diseminación clonal de Klebsiella pneumoniae ST307 coproductora de KPC-3 y NDM-1" ] ] "contieneResumen" => array:2 [ "en" => true "es" => true ] "contieneTextoCompleto" => array:1 [ "en" => true ] "contienePdf" => array:1 [ "en" => true ] "resumenGrafico" => array:2 [ "original" => 0 "multimedia" => array:7 [ "identificador" => "fig0005" "etiqueta" => "Figure 1" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr1.jpeg" "Alto" => 322 "Ancho" => 2508 "Tamanyo" => 62712 ] ] "descripcion" => array:1 [ "en" => "Clonal relationship of carbapenemase-producing K. pneumoniae ST307 isolates. Footnote: KpNR corresponds to a non-related KPC-2 producing isolate." ] ] ] "autores" => array:1 [ 0 => array:2 [ "autoresLista" => "Daniela Cejas, Francisco Magariños, Alan Elena, Micaela Ferrara, Cecilia Ormazábal, María Valeria Yernazian, Gabriel Gutkind, Marcela Radice" "autores" => array:8 [ 0 => array:2 [ "nombre" => "Daniela" "apellidos" => "Cejas" ] 1 => array:2 [ "nombre" => "Francisco" "apellidos" => "Magariños" ] 2 => array:2 [ "nombre" => "Alan" "apellidos" => "Elena" ] 3 => array:2 [ "nombre" => "Micaela" "apellidos" => "Ferrara" ] 4 => array:2 [ "nombre" => "Cecilia" "apellidos" => "Ormazábal" ] 5 => array:2 [ "nombre" => "María Valeria" "apellidos" => "Yernazian" ] 6 => array:2 [ "nombre" => "Gabriel" "apellidos" => "Gutkind" ] 7 => array:2 [ "nombre" => "Marcela" "apellidos" => "Radice" ] ] ] ] "resumen" => array:1 [ 0 => array:3 [ "titulo" => "Highlights" "clase" => "author-highlights" "resumen" => "<ul class="elsevierStyleList" id="lis0005"><li class="elsevierStyleListItem" id="lsti0005">•Klebsiella pneumoniae ST307 resistant to carbapenems and ceftazidime/avibactam.</li><li class="elsevierStyleListItem" id="lsti0010">•KPC-3 and NDM-1 producing Klebsiella pneumoniae isolates are clonally related.</li><li class="elsevierStyleListItem" id="lsti0015">•blaNDM-1 is located in a conjugative IncA/C plasmid.</li><li class="elsevierStyleListItem" id="lsti0020">•Genetic context of blaNDM-1 presented ISKpn14 and bleMBL.</li><li class="elsevierStyleListItem" id="lsti0025">•blaKPC was located in Tn4401a.</li></ul>" ] ] ] "idiomaDefecto" => "en" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S032575412200027X?idApp=UINPBA00004N" "url" => "/03257541/0000005400000004/v1_202212150521/S032575412200027X/v1_202212150521/en/main.assets" ] "itemAnterior" => array:18 [ "pii" => "S0325754122000311" "issn" => "03257541" "doi" => "10.1016/j.ram.2022.05.005" "estado" => "S300" "fechaPublicacion" => "2022-10-01" "aid" => "497" "copyright" => "Asociación Argentina de Microbiología" "documento" => "article" "crossmark" => 1 "subdocumento" => "fla" "cita" => "Rev Argent Microbiol. 2022;54:268-81" "abierto" => array:3 [ "ES" => false "ES2" => false "LATM" => false ] "gratuito" => false "lecturas" => array:1 [ "total" => 0 ] "en" => array:14 [ "idiomaDefecto" => true "cabecera" => "Original article" "titulo" => "Human hookworms from Argentina: Differential diagnosis of Necator americanus and Ancylostoma duodenale in endemic populations from Buenos Aires and Misiones" "tienePdf" => "en" "tieneTextoCompleto" => "en" "tieneResumen" => array:3 [ 0 => "en" 1 => "en" 2 => "es" ] "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "268" "paginaFinal" => "281" ] ] "titulosAlternativos" => array:1 [ "es" => array:1 [ "titulo" => "Ancylostomideos humanos de Argentina: Diagnóstico diferencial de Necator americanus y Ancylostoma duodenale en poblaciones endémicas de Buenos Aires y Misiones" ] ] "contieneResumen" => array:2 [ "en" => true "es" => true ] "contieneTextoCompleto" => array:1 [ "en" => true ] "contienePdf" => array:1 [ "en" => true ] "resumenGrafico" => array:2 [ "original" => 0 "multimedia" => array:7 [ "identificador" => "fig0010" "etiqueta" => "Figure 2" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr2.jpeg" "Alto" => 2505 "Ancho" => 2925 "Tamanyo" => 589855 ] ] "descripcion" => array:1 [ "en" => "Heat-map distance matrix showing percentage (above diagonal) and number (below diagonal) of base differences per site based on pairwise comparison of the partial fragment of the cob gene among the sequences obtained in this study, and 4 sequences of available at the GenBank database. There was a total of 169 positions in the final dataset. Sequences obtained in the present study are in bold." ] ] ] "autores" => array:1 [ 0 => array:2 [ "autoresLista" => "Andrea Servián, Silvia A. Repetto, María Lorena Zonta, Graciela T. Navone" "autores" => array:4 [ 0 => array:2 [ "nombre" => "Andrea" "apellidos" => "Servián" ] 1 => array:2 [ "nombre" => "Silvia A." "apellidos" => "Repetto" ] 2 => array:2 [ "nombre" => "María" "apellidos" => "Lorena Zonta" ] 3 => array:2 [ "nombre" => "Graciela T." "apellidos" => "Navone" ] ] ] ] "resumen" => array:1 [ 0 => array:3 [ "titulo" => "Highlights" "clase" => "author-highlights" "resumen" => "<ul class="elsevierStyleList" id="lis0005"><li class="elsevierStyleListItem" id="lsti0005">•Microscopic and PCR methods may be used as complementary tools to diagnose hookworms.</li><li class="elsevierStyleListItem" id="lsti0010">•All hookworm infections in this study were caused by Necator americanus.</li><li class="elsevierStyleListItem" id="lsti0015">•This is the first study to provide molecular data on Necator americanus in Argentina.</li><li class="elsevierStyleListItem" id="lsti0020">•Imported hookworm infections found here highlight the importance of their assessment in non-endemic areas.</li></ul>" ] ] ] "idiomaDefecto" => "en" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S0325754122000311?idApp=UINPBA00004N" "url" => "/03257541/0000005400000004/v1_202212150521/S0325754122000311/v1_202212150521/en/main.assets" ] "en" => array:22 [ "idiomaDefecto" => true "cabecera" => "Brief report" "titulo" => "First evidence of systemic infection and specific cytotoxic T lymphocyte immune response evoked by oral infection with recombinant Salmonella enterica serovar Albany-Ovalbumin in C57BL/6 mice" "tieneTextoCompleto" => true "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "282" "paginaFinal" => "287" ] ] "autores" => array:1 [ 0 => array:4 [ "autoresLista" => "Carolina Murúa-López, Priscilla González, Stephanie Almeida-Luna, Adriana Garibay-Escobar, Maribel Aguilar-Medina, Evangelina Beltrán-López, Héctor Samuel López-Moreno" "autores" => array:7 [ 0 => array:3 [ "nombre" => "Carolina" "apellidos" => "Murúa-López" "referencia" => array:2 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] 1 => array:2 [ "etiqueta" => "1" "identificador" => "fn0005" ] ] ] 1 => array:3 [ "nombre" => "Priscilla" "apellidos" => "González" "referencia" => array:2 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] 1 => array:2 [ "etiqueta" => "1" "identificador" => "fn0005" ] ] ] 2 => array:3 [ "nombre" => "Stephanie" "apellidos" => "Almeida-Luna" "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] ] ] 3 => array:3 [ "nombre" => "Adriana" "apellidos" => "Garibay-Escobar" "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "b" "identificador" => "aff0010" ] ] ] 4 => array:3 [ "nombre" => "Maribel" "apellidos" => "Aguilar-Medina" "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] ] ] 5 => array:3 [ "nombre" => "Evangelina" "apellidos" => "Beltrán-López" "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] ] ] 6 => array:4 [ "nombre" => "Héctor Samuel" "apellidos" => "López-Moreno" "email" => array:1 [ 0 => "hslmoreno@me.com" ] "referencia" => array:2 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] 1 => array:2 [ "etiqueta" => "*" "identificador" => "cor0005" ] ] ] ] "afiliaciones" => array:2 [ 0 => array:3 [ "entidad" => "Laboratorio de Biomedicina Molecular, Posgrado en Biotecnología y Posgrado en Ciencias Biomédicas, Facultad de Ciencias Químico Biológicas, Universidad Autónoma de Sinaloa, Sinaloa, Mexico" "etiqueta" => "a" "identificador" => "aff0005" ] 1 => array:3 [ "entidad" => "Facultad de Ciencias Químico Biológicas, Universidad de Sonora, Hermosillo, Sonora, Mexico" "etiqueta" => "b" "identificador" => "aff0010" ] ] "correspondencia" => array:1 [ 0 => array:3 [ "identificador" => "cor0005" "etiqueta" => "⁎" "correspondencia" => "Corresponding author." ] ] ] ] "titulosAlternativos" => array:1 [ "es" => array:1 [ "titulo" => "Primera evidencia de infección sistémica y respuesta inmune celular citotóxica específica evocada por la infección oral en ratones C57BL/6 con Salmonella enterica serovar Albany-Ovoalbúmina recombinante" ] ] "resumenGrafico" => array:2 [ "original" => 0 "multimedia" => array:7 [ "identificador" => "fig0025" "etiqueta" => "Figure 2" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr2.jpeg" "Alto" => 1810 "Ancho" => 3175 "Tamanyo" => 213963 ] ] "descripcion" => array:1 [ "en" => "rSAO evokes an epitope-specific CTL response with a Th-1 cytokine profile invivo. (A) Percentages of specific lysis observed at effector:target ratios 1:1, 10:1 and 50:1. Data are means±SEM from three independent experiments. (B) Concentrations of cytokines IFN-γ, TNF-α, IL-2, IL-6, IL-4, IL-10 and IL-17A in sera of infected mice 7 days pi. (*) Indicates statistical significance at p≤0.05 using the Student's t test. Data are means±SEM (n=3 mice per group) from three independent experiments. rSAO and rSTO are recombinant S. Albany and S. Typhimurium that express the Ovalbumin antigen, respectively." ] ] ] "textoCompleto" => "Salmonella enterica serovar Albany (S. Albany) is classified as a non-typhoidal Salmonella. This serovar was previously isolated by our group in food and feces of captive carnivores in a zoo located in northwestern Mexico<a class="elsevierStyleCrossRef" href="#bib0105">10</a>. Afterwards, it was also associated with the death of an ocelot (Leoparduspardalis) in the same zoo<a class="elsevierStyleCrossRef" href="#bib0100">9</a>. Additionally, this serovar was also involved in a lethal case of a 65-year-old patient<a class="elsevierStyleCrossRef" href="#bib0110">11</a>. This zoonotic and lethal potential support our aim to start with the study of important aspects about the host-S. Albany relationship. Here we analyzed the in vivo infection and specific ex vivo CTL immune response using the oral administration of the recombinant S. Albany-Ovalbumin (rSAO) strain in a murine model.All the reagents used in this study were purchased from Merk (Mexico). Once isolated, S. Albany was transformed with the Ovalbumin (Ova) gene using pST13-OVA, which codifies the OmpC porin from Salmonella Typhi (gently donated by Dr. Ortiz-Navarrete, from CINVESTAV, México) to obtain our recombinant bacterium designated as rSAO. S. Typhimurium 14028 transformed with the same plasmid, designated as rSTO (also donated by Dr. Ortiz-Navarrete), and used as positive control. After the transformation, the Ova expression was confirmed by Western blotting using a homemade mouse polyclonal α-Ova diluted 1:100 as first antibody and rabbit α-mouse IgG conjugated to horse-radish peroxidase diluted 1:3000 as second antibody. The reactions were revealed following protocols previously established in our investigation group using diaminobenzidine and H2O2<a class="elsevierStyleCrossRef" href="#bib0095">8</a>. Three groups consisting of 3 C57BL/6 mice (6–8 weeks-old) each were formed. While the first group was orally infected using 106 colony-forming units (CFU) of rSAO, the second group was infected with 105CFU of rSTO, due to the fact that S. Typhimurium was lethal for C57BL/6 mice. The mice in the third group were uninfected as previously described<a class="elsevierStyleCrossRef" href="#bib0065">2</a>. Six hours post-infection (pi) the feces were collected, weighed, diluted, homogenized in phosphate buffered-saline (PBS) pH 7.2 and plated on brain heart-infusion (BHI) agar supplemented with ampicillin at 100μg/ml; 24h after incubation at 37°C, CFU/mg were determined by colony counting. The mice were monitored every day, weighed, and clinically analyzed according to the scale previously proposed for evaluating mice clinical signs in response to Salmonella infection<a class="elsevierStyleCrossRef" href="#bib0080">5</a>. Briefly, the following five clinical criteria were evaluated: general activity level, posture, tremors, fur appearance and finally change in eating habits, which was determined by weighing the food. Each of these aspects was evaluated from a scale of 0–2 for every mouse, where a score of 0 was assigned to severely sick mice, indicating a great presence of the criteria evaluated. This could be evidenced by low activity or a significant decrease in movement, a constant hunch over posture, the presence of tremors, greasy fur appearance and a significant decrease in appetite. Likewise, a score of 1 was assigned to moderately sick mice with a small presence of the clinical sign evaluated. Lastly, a score of 2 corresponded to healthy mice with no change in activity or no signs of sickness<a class="elsevierStyleCrossRef" href="#bib0080">5</a>. At day 7 pi, all the clinical criteria values were added. If the final value ranged from 8 to 10, the mice were considered healthy; from 4 to 7, moderately sick; and from 0 to 3, severely sick. Afterwards, the mice were euthanized and their spleens were aseptically removed, measured, and splenocytes were counted using a Neubauer chamber and Trypan Blue 0.4%. To determine systemic infection, splenocytes and bone marrow cells were lysed using 1% TritonX-100, and intracellular bacteria were plated on BHI agar supplemented with ampicillin at 100μg/ml; 24h after incubation at 37°C, CFU were determined by colony counting.The cytotoxic effector T lymphocytes were obtained by in vitro stimulation with SIINFEKL (Kb restricted OVA-peptide, synthesized by Sigma-Aldrich, Mexico) for 5 days, following standard protocols. Likewise, 1×106 RMA-S cells (also donated by Dr. Ortiz from CINVESTAV-IPN) were stained with CFSE and pulsed with SIINFEKL for 4h prior to the interactions with the cytotoxic effector T lymphocytes (E). RMA-S cells were deemed target cells (T) and effector cells were adjusted to 1:1, 10:1 and 50:1 effector:target (E:T) ratios to evaluate antigen specific lysis; any unspecific lysis was eliminated by normalizing the data with results obtained against an irrelevant peptide (TAVIRADN). The samples were assessed on FACS Accuri C6 (BD) and analyzed using FlowJo software<a class="elsevierStyleCrossRef" href="#bib0075">4</a>.Additionally, the sera of uninfected or infected rSAO or rSTO mice were collected on day 7 pi and were used to determine the cytokine profile associated with the T helper (Th) subset immune response using a CBA Mouse Cytokine Kit (BD). For Th1, the cytokines evaluated were: IFN-γ, TNF-α, IL-2 and IL-6, for Th2: IL-4 and IL-10 and for Th17: IL-17, according to the manufacturer's instructions. The data were acquired on a FACS Accuri C6 (BD) and analyzed using FlowJo software. The data obtained were analyzed using Student's t test on GraphPad Prism 6, where p≤0.05 was considered to be statistically significant.The Ova expression in our recombinant S. Albany strain was confirmed by the Western blot analysis (<a class="elsevierStyleCrossRef" href="#fig0020">Fig. 1</a>A), where it was possible to observe a clear band of approximately 45 kDa in crude extract of rSAO, consistent with commercial Ova as positive control in lane 4. The clinical findings of infected as well as uninfected mice are summarized in <a class="elsevierStyleCrossRef" href="#fig0020">Figure 1</a>B, and can also be observed in <a class="elsevierStyleCrossRef" href="#sec0025">Supplementary Figure 1</a>, where infected mice with rSAO presented a clinical score of 5, which is consistent with that of moderately sick mice, suggesting that this serovar has the capacity to cause characteristic symptoms of an infection by Salmonella. Similar results were obtained from mice infected with the positive control rSTO, while the small differences observed can be explained by the initial inoculum concentration. Both groups presented signs consistent with Salmonella infection, which was considered an initial indicator of the pathogenic potential of rSAO. Various groups have described that along with the behavioral aspects, mice tend to lose weight during infection, and we observed that mice infected with rSAO presented a total average weight loss of 16%, suggesting that the clinical signs observed were significant enough to cause a decrease in appetite in infected mice, and showing that rSAO can display its pathogenic capability (<a class="elsevierStyleCrossRef" href="#fig0020">Fig. 1</a>C). To confirm this intestinal infection, the feces collected at 6h pi were plated and the mean CFU observed in rSAO infected mice was of 1650CFU/mg, showing no statistical difference with respect to the rSTO-infected mice (<a class="elsevierStyleCrossRef" href="#fig0020">Fig. 1</a>D). These results suggest that rSAO can colonize the intestine, establishing a gastrointestinal infection in mice. Our findings are additionally supported by a similar infection study conducted recently, where it was determined that S. Albany can establish infection and cause pathological changes in tissue in the small intestine, cecum and colon<a class="elsevierStyleCrossRef" href="#bib0090">7</a>. To further assess the infection caused by rSAO, the spleens were extracted on day 7 pi and measured; the spleens from mice infected with rSAO presented a mean of 0.6 inches in length, exhibiting a statistically significant difference with respect to the spleens extracted from negative control mice (<a class="elsevierStyleCrossRef" href="#fig0020">Fig. 1</a>E). As well as a visible increase in size, it was possible to observe similar macroscopic characteristics in the spleens from mice infected with the positive control rSTO. To demonstrate the possible increased cellularity in infected mice responsible for the increase in spleen size, splenocytes were counted and although the values were not statistically significant, it was evident that there were approximately 0.5 time more cells in the spleens extracted from rSAO and rSTO infected mice when compared to spleens extracted from the non-infected control group (see <a class="elsevierStyleCrossRef" href="#fig0020">Figure 1</a>f). Once cellularity was determined, splenocytes were lysed to assess rSAO to evidence its capacity to cause a systemic infection; the mean observed for CFUs in splenocytes of rSAO infected mice was of 3.98×105 per million cells (see <a class="elsevierStyleCrossRef" href="#fig0020">Fig. 1</a>G). Parallelly, we determined the bacterial burden in total bone marrow cells to further assess the systemic infection and the mean observed was of 1.87×105 CFUs in total bone marrow cells, demonstrating the capacity of this serovar to establish a systemic infection in mice, thus highlighting its pathogenic potential (see <a class="elsevierStyleCrossRef" href="#fig0020">Fig. 1</a>H). A possible explanation, at least in part, for our data, can be related to reports that S. Typhimurium can cause systemic disease in C57BL/6 infected mice, by persisting in B lymphocytes and precursors specifically within the bone marrow<a class="elsevierStyleCrossRef" href="#bib0060">1</a>. Maybe rSAO, as well as rSTO, are capable of infecting and establishing a niche in memory B cells and thus, chronic infection. However, more detailed studies should be conducted.<elsevierMultimedia ident="fig0020"></elsevierMultimedia>CTL antigen-specific lysis demonstrated that effector cells obtained from rSAO-infected mice were able to kill RMA-S cells pulsed with SIINFEKL, but were inefficient when these target cells were pulsed with an irrelevant peptide. It is relevant to mention that although splenocytes were infected with recombinant bacteria, memory effector T CD8+ clonal expansion was clearly decreased in the irrelevant peptide pulsed group, in contrast with the greater clonal expansion observed in the group pulsed with the SIINFEKL epitope, as shown in <a class="elsevierStyleCrossRef" href="#sec0025">Supplementary Figure 3</a>. These results demonstrate that although there is small participation of the genetic construction used, it is not significant. Effector cells evoked 15.6% of specific lysis for 1:1 E:T ratio, 45.1% for 10:1 E:T ratio and 60% for 50:1 E:T ratio. Therefore, CTL from rSAO infected mice recognize this epitope and generate a Kb-restricted CTL response against S. Albany (<a class="elsevierStyleCrossRef" href="#fig0025">Fig. 2</a>A). To determine the cytokine profile evoked in vivo by rSAO, the sera of infected and uninfected mice were analyzed, and we observed a predominance of the Th1-like cytokine profile (<a class="elsevierStyleCrossRef" href="#fig0025">Fig. 2</a>B). Concentrations of IL-4 and IL-10 were 4 and 5Ypg/ml respectively. IL-6 and IL-2 were both close to 40Ypg/ml, TNF-α concentrations were of 20Ypg/ml, but the IFN-γ concentration was the predominant cytokine, reaching levels of 200Ypg/ml in rSAO-infected mice. These results were consistent with what was observed in our positive control group rSTO, where Th1 cytokines IFN-γ, IL-2, IL-6 and TNF-α were predominant, further suggesting immune response similarities evoked by both serovars. Along with the importance of the Th1 cytokine profile, it has also been reported that the presence of IL-17A along with the Th1 cytokine profile is necessary in controlling Salmonella infection. However, these findings also show that its presence is predominant in the initial stages of infection because its primary function is the recruitment of neutrophils at the initial stages of inflammation<a class="elsevierStyleCrossRef" href="#bib0065">2</a>. Because we determined the levels of all the cytokines in the final stages of infection, this could be a possible explanation as to why no statistical difference in the levels of IL-17A was observed across our experimental groups. The secretion of Th1 profile cytokines, specifically IFN-γ, has been previously described in S. Typhimurium infections, where macrophages must be activated to eliminate the intracellular bacteria. Thus, these results suggest that rSAO could establish intracellular infection, evoking protective immunity in mice, which is characterized by the secretion of the Th1 cytokine profile.<elsevierMultimedia ident="fig0025"></elsevierMultimedia>Infection by S. Typhimurium has been thoroughly characterized in C57BL/6 mice and has been used as the comparative gold standard model for other potentially pathogenic Salmonella serovars<a class="elsevierStyleCrossRef" href="#bib0070">3</a>. This was consistent with S. Albany, a serovar that had not been previously reported in our region, until its isolation by our investigation group in a zoo from northwestern Mexico, and its further association with the death of an ocelot (Leopardus pardalis) in the same zoo<a class="elsevierStyleCrossRefs" href="#bib0100">9,10</a>, as it was the first indicator of the pathogenic potential of S. Albany. In this study we have demonstrated for the first time that S. Albany has the pathogenic potential to establish a systemic infection in C57BL/6 mice, displaying the capability to evoke a specific CTL immune response. We found that mice that were orally infected with rSAO exhibited similar nociceptive signs as those infected with the positive control rSTO, suggesting that rSAO has the capability to establish a systemic infection such as rSTO. In addition to these findings, we also demonstrated that rSAO infected mice lost weight in response to the infection, which could be explained by the loss of appetite caused by the strong symptoms induced by the rSAO infection. Likewise, it could also be attributed to the high energy expenditure required to fight off the infection, suggesting that rSAO was capable of establishing a strong enough infection to cause symptoms that directly impacted on the mice appetite and therefore their weight. The capability of rSAO to establish an infection in the gastrointestinal tract was demonstrated by the bacteria isolated from the feces collected 6h pi. The number of colonies exhibited no statistical differences compared with rSTO- infected mice, suggesting that rSAO had the ability to resist stomach acidic conditions and pass through the intestinal mucosa to start infection. This also suggests that rSAO expresses a set of vital virulence factors for establishing infection such as Type III Secretion Systems, Sop and Inv proteins, as well as niche factors that would allow it to overcome bile salts, high osmolarity and competition for nutrients in the intestine; however, to confirm these molecular mechanisms, more detailed studies should be conducted. It has also been previously reported that murine S. Typhimurium infection evokes an increase in spleen size, probably due to the specific clonal expansion of resident immunity cells from this secondary immune organ<a class="elsevierStyleCrossRef" href="#bib0085">6</a>.The specific lysis observed demonstrated that this response was specific for cells harboring the SIINFEKL epitope and CTL were activated by the rSAO infection, generating a specific in vivo immune response. The high concentrations of IFN-γ and TNF-α obtained in our experimental conditions could explain the high percentages of specific lysis as well as the severity of the nociceptive signs observed in infected mice, suggesting that although the infection was strong enough to evoke an immune response, it was probably insufficient to resolve the infection; however, a more detailed study should be conducted to confirm this hypothesis.We demonstrated here, for the first time that S. Albany causes a systemic disease in C57BL/6 mice and has the same potential to infect other mammalian hosts, including humans, probably expressing a set of virulence factors, pathogenicity islands and secretion systems, although further molecular analyses are needed. Additionally, the isolation of rSAO in bone marrow cells suggest a chronic systemic infection, such as human typhoid fever. The use of recombinant S. Albany-Ovalbumin strain as an antigenic flag represents an important biotechnological tool to study in vivo immune responses. Finally, the CTL immune response evidenced here supports the possibility to manipulate the cellular immune response to eliminate this bacterium effectively.Authorship contributionCML, PG, SAL, AGE, MAM, EBL, HSLM: Investigation, experimental procedures, data analysis, writing and editing the manuscript. Also, HSLM: design the project, obtained the grant support, and validation of data.Conflicts of interestNone.Ethical responsibilitiesNone." "textoCompletoSecciones" => array:1 [ "secciones" => array:10 [ 0 => array:3 [ "identificador" => "xres1819626" "titulo" => "Abstract" "secciones" => array:1 [ 0 => array:1 [ "identificador" => "abst0005" ] ] ] 1 => array:2 [ "identificador" => "xpalclavsec1588105" "titulo" => "Keywords" ] 2 => array:2 [ "identificador" => "xpalclavsec1588104" "titulo" => "Abbreviations" ] 3 => array:3 [ "identificador" => "xres1819625" "titulo" => "Resumen" "secciones" => array:1 [ 0 => array:1 [ "identificador" => "abst0010" ] ] ] 4 => array:2 [ "identificador" => "xpalclavsec1588106" "titulo" => "Palabras clave" ] 5 => array:2 [ "identificador" => "sec0005" "titulo" => "Authorship contribution" ] 6 => array:2 [ "identificador" => "sec0010" "titulo" => "Conflicts of interest" ] 7 => array:2 [ "identificador" => "sec0015" "titulo" => "Ethical responsibilities" ] 8 => array:2 [ "identificador" => "xack641879" "titulo" => "Acknowledgements" ] 9 => array:1 [ "titulo" => "References" ] ] ] "pdfFichero" => "main.pdf" "tienePdf" => true "fechaRecibido" => "2020-11-10" "fechaAceptado" => "2022-05-02" "PalabrasClave" => array:2 [ "en" => array:2 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Keywords" "identificador" => "xpalclavsec1588105" "palabras" => array:3 [ 0 => "Cytotoxic T lymphocytes" 1 => "Salmonella enterica serovar Albany" 2 => "Recombinant bacterium" ] ] 1 => array:4 [ "clase" => "abr" "titulo" => "Abbreviations" "identificador" => "xpalclavsec1588104" "palabras" => array:10 [ 0 => "rSAO" 1 => "rSTO" 2 => "Ova" 3 => "CTL" 4 => "TNF-α" 5 => "IFN-γ" 6 => "IL" 7 => "pi" 8 => "CFU" 9 => "BHI" ] ] ] "es" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Palabras clave" "identificador" => "xpalclavsec1588106" "palabras" => array:3 [ 0 => "Linfocito T citotóxico" 1 => "Salmonella enterica serovar Albany" 2 => "Bacteria recombinante" ] ] ] ] "tieneResumen" => true "resumen" => array:2 [ "en" => array:2 [ "titulo" => "Abstract" "resumen" => "Our group isolated Salmonella enterica serovar Albany from food and feces of wild captive carnivores in a zoo from northwestern Mexico. This serovar was also associated with the death of an ocelot (Leopardus pardalis) in the same zoo. Another group associated S. Albany with the death of a human patient. It is due to this zoonotic potential that the in vivo study of the host-S. Albany relationship is critical. The recombinant S. Albany-Ovalbumin (rSAO) strain was used to analyze a murine oral infection and its specific cytotoxic T lymphocyte (CTL) response. Our results have shown for the first time that rSAO establishes a systemic infection and evokes epitope-specific lysis with a Th1-like cytokine profile in vivo." ] "es" => array:2 [ "titulo" => "Resumen" "resumen" => "Salmonella enterica serovar Albany fue aislada por nuestro grupo de investigación de alimentos contaminados y de heces de animales carnívoros en cautiverio en un zoológico del noroeste de México; posteriormente, se logró asociar a este serovar con la muerte de un ocelote (Leopardus pardalis), dentro de este mismo zoológico. Otro grupo de investigación asoció a este serovar con la muerte de un paciente. Es debido a este potencial zoonótico que el estudio in vivo de la relación hospedero-S. Albany es crítico. La cepa recombinante S. Albany-Ovoalbúmina (rSAO) fue utilizada para analizar la infección múrida, al igual que la respuesta inmune celular citotóxica específica. Nuestros resultados demuestran, por primera vez, que rSAO establece una infección sistémica y evoca lisis epítopo-específica con un perfil de citocinas tipo Th1 in vivo." ] ] "NotaPie" => array:1 [ 0 => array:3 [ "etiqueta" => "1" "nota" => "These authors contributed equally to this article." "identificador" => "fn0005" ] ] "apendice" => array:1 [ 0 => array:1 [ "seccion" => array:1 [ 0 => array:4 [ "apendice" => "The following are Supplementary data to this article:<elsevierMultimedia ident="fig0005"></elsevierMultimedia><elsevierMultimedia ident="fig0010"></elsevierMultimedia><elsevierMultimedia ident="fig0015"></elsevierMultimedia>" "etiqueta" => "Appendix A" "titulo" => "Supplementary data" "identificador" => "sec0025" ] ] ] ] "multimedia" => array:5 [ 0 => array:7 [ "identificador" => "fig0020" "etiqueta" => "Figure 1" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr1.jpeg" "Alto" => 1130 "Ancho" => 2925 "Tamanyo" => 154526 ] ] "descripcion" => array:1 [ "en" => "rSAO causes systemic infection in C57BL/6 mice. (A) Ova expression by transformed S. Albany; Lane 1: molecular weight marker, Lane 2: crude extract S. Albany WT, Lane 3: crude extract rSAO, Lane 4: positive control Ova protein fraction V. (B) Clinical findings in mice orally infected with 1×106 CFU rSAO, 1×105 rSTO and uninfected mice. (C) Weight loss observed in infected and uninfected mice at 1, 3, 5, and 7 days pi. (D) CFU per milligram of feces collected 6h pi. (E) Increase in spleen size measured in cm 7 days pi. (F) Increase in cellularity observed in infected mice spleens 7 days pi. (G) CFU per million cells in infected mice spleens at 7 days pi. (H) CFU in total bone marrow cells at 7d pi. (*) Indicates statistical significance at p≤0.05 using Student's t test. Data are means±SEM (n=3 mice per group) from three independent experiments. rSAO and rSTO are recombinant S. Albany and S. Typhimurium that express the Ovalbumin antigen, respectively." ] ] 1 => array:7 [ "identificador" => "fig0025" "etiqueta" => "Figure 2" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr2.jpeg" "Alto" => 1810 "Ancho" => 3175 "Tamanyo" => 213963 ] ] "descripcion" => array:1 [ "en" => "rSAO evokes an epitope-specific CTL response with a Th-1 cytokine profile invivo. (A) Percentages of specific lysis observed at effector:target ratios 1:1, 10:1 and 50:1. Data are means±SEM from three independent experiments. (B) Concentrations of cytokines IFN-γ, TNF-α, IL-2, IL-6, IL-4, IL-10 and IL-17A in sera of infected mice 7 days pi. (*) Indicates statistical significance at p≤0.05 using the Student's t test. Data are means±SEM (n=3 mice per group) from three independent experiments. rSAO and rSTO are recombinant S. Albany and S. Typhimurium that express the Ovalbumin antigen, respectively." ] ] 2 => array:5 [ "identificador" => "fig0005" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => false "mostrarDisplay" => true "figura" => array:1 [ 0 => array:4 [ "imagen" => "mmc1.jpeg" "Alto" => 1088 "Ancho" => 2500 "Tamanyo" => 298546 ] ] ] 3 => array:5 [ "identificador" => "fig0010" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => false "mostrarDisplay" => true "figura" => array:1 [ 0 => array:4 [ "imagen" => "mmc2.jpeg" "Alto" => 684 "Ancho" => 1500 "Tamanyo" => 138680 ] ] ] 4 => array:5 [ "identificador" => "fig0015" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => false "mostrarDisplay" => true "figura" => array:1 [ 0 => array:4 [ "imagen" => "mmc3.jpeg" "Alto" => 726 "Ancho" => 1500 "Tamanyo" => 123593 ] ] ] ] "bibliografia" => array:2 [ "titulo" => "References" "seccion" => array:1 [ 0 => array:2 [ "identificador" => "bibs0015" "bibliografiaReferencia" => array:11 [ 0 => array:3 [ "identificador" => "bib0060" "etiqueta" => "1" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "B cell precursors are targets for Salmonella infection" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:6 [ 0 => "D. Castro-Eguiluz" 1 => "R. Pelayo" 2 => "V. Rosales-Garcia" 3 => "R. Rosales-Reyes" 4 => "C. Alpuche-Aranda" 5 => "V. Ortiz-Navarrete" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:5 [ "tituloSerie" => "Microb Pathog" "fecha" => "2009" "volumen" => "47" "paginaInicial" => "52" "paginaFinal" => "56" ] ] ] ] ] ] 1 => array:3 [ "identificador" => "bib0065" "etiqueta" => "2" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Interleukin-17A is required to suppress invasion of Salmonella enterica serovar Typhimurium to enteric mucosa" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:5 [ 0 => "H. Mayuzumi" 1 => "K. Inagaki-Ohara" 2 => "C. Uyttenhove" 3 => "Y. Okamoto" 4 => "G. Matsuzaki" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:5 [ "tituloSerie" => "Immunology" "fecha" => "2010" "volumen" => "131" "paginaInicial" => "377" "paginaFinal" => "385" ] ] ] ] ] ] 2 => array:3 [ "identificador" => "bib0070" "etiqueta" => "3" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Entry route of Salmonella Typhimurium directs the type of induced immune response" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:3 [ 0 => "C. Martinoli" 1 => "A. Chiavelli" 2 => "M. Rescigno" ] ] ] ] ] "host" => array:1 [ 0 => array:2 [ "doi" => "10.1016/j.immuni.2007.10.011" "Revista" => array:6 [ "tituloSerie" => "Immunity" "fecha" => "2007" "volumen" => "27" "paginaInicial" => "975" "paginaFinal" => "984" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/18083577" "web" => "Medline" ] ] ] ] ] ] ] ] 3 => array:3 [ "identificador" => "bib0075" "etiqueta" => "4" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "The plasmacytoid dendritic cells evoke Salmonella-specific CTL effector response" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:3 [ 0 => "C. Murúa-López" 1 => "M. González-Orozco" 2 => "H.S. López-Moreno" ] ] ] ] ] "host" => array:1 [ 0 => array:2 [ "doi" => "10.7705/biomedica.v39i3.4415" "Revista" => array:6 [ "tituloSerie" => "Biomédica" "fecha" => "2019" "volumen" => "39" "paginaInicial" => "172" "paginaFinal" => "181" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/31529859" "web" => "Medline" ] ] ] ] ] ] ] ] 4 => array:3 [ "identificador" => "bib0080" "etiqueta" => "5" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Salmonella Typhimurium infections in BALB/c mice: a comparison of tissue bioluminescence, tissue cultures and mice clinical scores" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:7 [ 0 => "H. Özkaya" 1 => "B. Akcan" 2 => "G. Aydemir" 3 => "S. Aydinöz" 4 => "Y. Razia" 5 => "T. Gammon" 6 => "J. McKinney" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "New Microbiol" "fecha" => "2012" "volumen" => "35" "paginaInicial" => "53" "paginaFinal" => "59" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/22378553" "web" => "Medline" ] ] ] ] ] ] ] ] 5 => array:3 [ "identificador" => "bib0085" "etiqueta" => "6" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Infection with Salmonella enterica serovar Typhimurium leads to increased proportions of F4/80+ red pulp macrophages and decreased proportions of B and T lymphocytes in the spleen" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:5 [ 0 => "L. Rosche" 1 => "T. Aljasham" 2 => "N. Kipfer" 3 => "T. Piatkowski" 4 => "V. Konjufca" ] ] ] ] ] "host" => array:1 [ 0 => array:2 [ "doi" => "10.1371/journal.pone.0130092" "Revista" => array:5 [ "tituloSerie" => "PLoS ONE" "fecha" => "2015" "volumen" => "10" "paginaInicial" => "e0130092" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/26068006" "web" => "Medline" ] ] ] ] ] ] ] ] 6 => array:3 [ "identificador" => "bib0090" "etiqueta" => "7" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Pathogenesis of Salmonellaenterica serovar Albany in experimental infected SPF BALB/c Mice" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:5 [ 0 => "J. Sabri" 1 => "I. Al-Sultan" 2 => "K. Altaif" 3 => "S. Peter" 4 => "M. Saadh" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:5 [ "tituloSerie" => "Iraq J Vet Sci" "fecha" => "2020" "volumen" => "34" "paginaInicial" => "339" "paginaFinal" => "344" ] ] ] ] ] ] 7 => array:3 [ "identificador" => "bib0095" "etiqueta" => "8" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Reaction of Leishmania (Leishmania) mexicana antigens by sera of patients with cutaneous leishmaniasis from Sinaloa, Mexico" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:3 [ 0 => "P. Salazar-Mejía" 1 => "R. Tejeda-Aguirre" 2 => "H. López-Moreno" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:5 [ "tituloSerie" => "Salud Publica Mex" "fecha" => "2010" "volumen" => "52" "paginaInicial" => "165" "paginaFinal" => "169" ] ] ] ] ] ] 8 => array:3 [ "identificador" => "bib0100" "etiqueta" => "9" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Excreción fecal de Salmonella Albany, su aislamiento en la ración alimenticia y repercusión en el estado de salud de un ocelote (Leopardus pardalis) en cautiverio" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:5 [ 0 => "G. Silva-Hidalgo" 1 => "H.S. López-Moreno" 2 => "V.F. Ortiz-Navarrete" 3 => "F. Juárez-Barranco" 4 => "M. López-Valenzuela" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:5 [ "tituloSerie" => "Vet Mex" "fecha" => "2012" "volumen" => "43" "paginaInicial" => "59" "paginaFinal" => "69" ] ] ] ] ] ] 9 => array:3 [ "identificador" => "bib0105" "etiqueta" => "10" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Non-typhi Salmonella serovars found in Mexican zoo animals" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:7 [ 0 => "G. Silva-Hidalgo" 1 => "V. Ortiz-Navarrete" 2 => "C. Alpuche-Aranda" 3 => "J. Rendón-Maldonado" 4 => "M. López-Valenzuela" 5 => "F. Juárez-Barranco" 6 => "H.S. López-Moreno" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:4 [ "tituloSerie" => "Res Vet Sci" "fecha" => "2012" "volumen" => "93" "paginaInicial" => "11325" ] ] ] ] ] ] 10 => array:3 [ "identificador" => "bib0110" "etiqueta" => "11" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Peritonitis por Salmonella serotipo Albany en paciente sometido a diálisis peritoneal continua ambulatoria" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:4 [ 0 => "R. Tato" 1 => "M. Zamora" 2 => "M. Pulian" 3 => "M. Garcia" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "Rev Esp Quimioter" "fecha" => "2015" "volumen" => "28" "paginaInicial" => "162" "paginaFinal" => "163" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/26033003" "web" => "Medline" ] ] ] ] ] ] ] ] ] ] ] ] "agradecimientos" => array:1 [ 0 => array:4 [ "identificador" => "xack641879" "titulo" => "Acknowledgements" "texto" => "The authors would like to thank fully to Dr. Vianney Francisco Ortiz Navarrete for gently donative the S. Typhimurium 14028 and plasmidic construction pST13-OVA. Also, they extended your acknowledgment to PROFAPI-UAS for financial support with the HSLM's Grant #2015/151. And LBM team for technical assistance" "vista" => "all" ] ] ] "idiomaDefecto" => "en" "url" => "/03257541/0000005400000004/v1_202212150521/S0325754122000529/v1_202212150521/en/main.assets" "Apartado" => array:4 [ "identificador" => "37863" "tipo" => "SECCION" "en" => array:2 [ "titulo" => "Microbiología básica" "idiomaDefecto" => true ] "idiomaDefecto" => "en" ] "PDF" => "https://static.elsevier.es/multimedia/03257541/0000005400000004/v1_202212150521/S0325754122000529/v1_202212150521/en/main.pdf?idApp=UINPBA00004N&text.app=https://www.elsevier.es/" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S0325754122000529?idApp=UINPBA00004N"]

Statistics

Brief report

Primera evidencia de infección sistémica y respuesta inmune celular citotóxica específica evocada por la infección oral en ratones C57BL/6 con Salmonella enterica serovar Albany-Ovoalbúmina recombinante

Carolina Murúa-Lópeza,1, Priscilla Gonzáleza,1, Stephanie Almeida-Lunaa, Adriana Garibay-Escobarb, Maribel Aguilar-Medinaa, Evangelina Beltrán-Lópeza, Héctor Samuel López-Morenoa,

Corresponding author

hslmoreno@me.com

Corresponding author.

a Laboratorio de Biomedicina Molecular, Posgrado en Biotecnología y Posgrado en Ciencias Biomédicas, Facultad de Ciencias Químico Biológicas, Universidad Autónoma de Sinaloa, Sinaloa, Mexico

b Facultad de Ciencias Químico Biológicas, Universidad de Sonora, Hermosillo, Sonora, Mexico

Read

879

Times
was read the article

209

Total PDF

670

Total HTML

Share statistics

 array:24 [
  "pii" => "S0325754122000529"
  "issn" => "03257541"
  "doi" => "10.1016/j.ram.2022.05.008"
  "estado" => "S300"
  "fechaPublicacion" => "2022-10-01"
  "aid" => "502"
  "copyright" => "Asociaci&#243;n Argentina de Microbiolog&#237;a"
  "copyrightAnyo" => "2022"
  "documento" => "article"
  "crossmark" => 1
  "licencia" => "http://creativecommons.org/licenses/by-nc-nd/4.0/"
  "subdocumento" => "fla"
  "cita" => "Rev Argent Microbiol. 2022;54:282-7"
  "abierto" => array:3 [
    "ES" => true
    "ES2" => true
    "LATM" => true
  ]
  "gratuito" => true
  "lecturas" => array:1 [
    "total" => 0
  ]
  "itemSiguiente" => array:19 [
    "pii" => "S032575412200027X"
    "issn" => "03257541"
    "doi" => "10.1016/j.ram.2022.04.002"
    "estado" => "S300"
    "fechaPublicacion" => "2022-10-01"
    "aid" => "493"
    "copyright" => "Asociaci&#243;n Argentina de Microbiolog&#237;a"
    "documento" => "article"
    "crossmark" => 1
    "licencia" => "http://creativecommons.org/licenses/by-nc-nd/4.0/"
    "subdocumento" => "fla"
    "cita" => "Rev Argent Microbiol. 2022;54:288-92"
    "abierto" => array:3 [
      "ES" => true
      "ES2" => true
      "LATM" => true
    ]
    "gratuito" => true
    "lecturas" => array:1 [
      "total" => 0
    ]
    "en" => array:14 [
      "idiomaDefecto" => true
      "cabecera" => "<span class="elsevierStyleTextfn">Brief report</span>"
      "titulo" => "Emergence and clonal expansion of <span class="elsevierStyleItalic">Klebsiella pneumoniae</span> ST307&#44; simultaneously producing KPC-3 and NDM-1"
      "tienePdf" => "en"
      "tieneTextoCompleto" => "en"
      "tieneResumen" => array:3 [
        0 => "en"
        1 => "en"
        2 => "es"
      ]
      "paginas" => array:1 [
        0 => array:2 [
          "paginaInicial" => "288"
          "paginaFinal" => "292"
        ]
      ]
      "titulosAlternativos" => array:1 [
        "es" => array:1 [
          "titulo" => "Emergencia y diseminaci&#243;n clonal de <span class="elsevierStyleItalic">Klebsiella pneumoniae</span> ST307 coproductora de KPC-3 y NDM-1"
        ]
      ]
      "contieneResumen" => array:2 [
        "en" => true
        "es" => true
      ]
      "contieneTextoCompleto" => array:1 [
        "en" => true
      ]
      "contienePdf" => array:1 [
        "en" => true
      ]
      "resumenGrafico" => array:2 [
        "original" => 0
        "multimedia" => array:7 [
          "identificador" => "fig0005"
          "etiqueta" => "Figure 1"
          "tipo" => "MULTIMEDIAFIGURA"
          "mostrarFloat" => true
          "mostrarDisplay" => false
          "figura" => array:1 [
            0 => array:4 [
              "imagen" => "gr1.jpeg"
              "Alto" => 322
              "Ancho" => 2508
              "Tamanyo" => 62712
            ]
          ]
          "descripcion" => array:1 [
            "en" => "<p id="spar0020" class="elsevierStyleSimplePara elsevierViewall">Clonal relationship of carbapenemase-producing <span class="elsevierStyleItalic">K&#46; pneumoniae</span> ST307 isolates&#46;</p> <p id="spar0025" class="elsevierStyleSimplePara elsevierViewall">Footnote&#58; KpNR corresponds to a non-related KPC-2 producing isolate&#46;</p>"
          ]
        ]
      ]
      "autores" => array:1 [
        0 => array:2 [
          "autoresLista" => "Daniela Cejas, Francisco Magari&#241;os, Alan Elena, Micaela Ferrara, Cecilia Ormaz&#225;bal, Mar&#237;a Valeria Yernazian, Gabriel Gutkind, Marcela Radice"
          "autores" => array:8 [
            0 => array:2 [
              "nombre" => "Daniela"
              "apellidos" => "Cejas"
            ]
            1 => array:2 [
              "nombre" => "Francisco"
              "apellidos" => "Magari&#241;os"
            ]
            2 => array:2 [
              "nombre" => "Alan"
              "apellidos" => "Elena"
            ]
            3 => array:2 [
              "nombre" => "Micaela"
              "apellidos" => "Ferrara"
            ]
            4 => array:2 [
              "nombre" => "Cecilia"
              "apellidos" => "Ormaz&#225;bal"
            ]
            5 => array:2 [
              "nombre" => "Mar&#237;a Valeria"
              "apellidos" => "Yernazian"
            ]
            6 => array:2 [
              "nombre" => "Gabriel"
              "apellidos" => "Gutkind"
            ]
            7 => array:2 [
              "nombre" => "Marcela"
              "apellidos" => "Radice"
            ]
          ]
        ]
      ]
      "resumen" => array:1 [
        0 => array:3 [
          "titulo" => "Highlights"
          "clase" => "author-highlights"
          "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><p id="spar0005" class="elsevierStyleSimplePara elsevierViewall"><ul class="elsevierStyleList" id="lis0005"><li class="elsevierStyleListItem" id="lsti0005"><span class="elsevierStyleLabel">&#8226;</span><p id="par0005" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">Klebsiella pneumoniae</span> ST307 resistant to carbapenems and ceftazidime&#47;avibactam&#46;</p></li><li class="elsevierStyleListItem" id="lsti0010"><span class="elsevierStyleLabel">&#8226;</span><p id="par0010" class="elsevierStylePara elsevierViewall">KPC-3 and NDM-1 producing <span class="elsevierStyleItalic">Klebsiella pneumoniae</span> isolates are clonally related&#46;</p></li><li class="elsevierStyleListItem" id="lsti0015"><span class="elsevierStyleLabel">&#8226;</span><p id="par0015" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">bla</span><span class="elsevierStyleInf">NDM-1</span> is located in a conjugative IncA&#47;C plasmid&#46;</p></li><li class="elsevierStyleListItem" id="lsti0020"><span class="elsevierStyleLabel">&#8226;</span><p id="par0020" class="elsevierStylePara elsevierViewall">Genetic context of <span class="elsevierStyleItalic">bla</span><span class="elsevierStyleInf">NDM-1</span> presented IS<span class="elsevierStyleItalic">Kpn14</span> and <span class="elsevierStyleItalic">bleMBL</span>&#46;</p></li><li class="elsevierStyleListItem" id="lsti0025"><span class="elsevierStyleLabel">&#8226;</span><p id="par0025" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">bla</span><span class="elsevierStyleInf">KPC</span> was located in Tn<span class="elsevierStyleItalic">4401a</span>&#46;</p></li></ul></p></span>"
        ]
      ]
    ]
    "idiomaDefecto" => "en"
    "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S032575412200027X?idApp=UINPBA00004N"
    "url" => "/03257541/0000005400000004/v1_202212150521/S032575412200027X/v1_202212150521/en/main.assets"
  ]
  "itemAnterior" => array:18 [
    "pii" => "S0325754122000311"
    "issn" => "03257541"
    "doi" => "10.1016/j.ram.2022.05.005"
    "estado" => "S300"
    "fechaPublicacion" => "2022-10-01"
    "aid" => "497"
    "copyright" => "Asociaci&#243;n Argentina de Microbiolog&#237;a"
    "documento" => "article"
    "crossmark" => 1
    "subdocumento" => "fla"
    "cita" => "Rev Argent Microbiol. 2022;54:268-81"
    "abierto" => array:3 [
      "ES" => false
      "ES2" => false
      "LATM" => false
    ]
    "gratuito" => false
    "lecturas" => array:1 [
      "total" => 0
    ]
    "en" => array:14 [
      "idiomaDefecto" => true
      "cabecera" => "<span class="elsevierStyleTextfn">Original article</span>"
      "titulo" => "Human hookworms from Argentina&#58; Differential diagnosis of <span class="elsevierStyleItalic">Necator americanus</span> and <span class="elsevierStyleItalic">Ancylostoma duodenale</span> in endemic populations from Buenos Aires and Misiones"
      "tienePdf" => "en"
      "tieneTextoCompleto" => "en"
      "tieneResumen" => array:3 [
        0 => "en"
        1 => "en"
        2 => "es"
      ]
      "paginas" => array:1 [
        0 => array:2 [
          "paginaInicial" => "268"
          "paginaFinal" => "281"
        ]
      ]
      "titulosAlternativos" => array:1 [
        "es" => array:1 [
          "titulo" => "Ancylostomideos humanos de Argentina&#58; Diagn&#243;stico diferencial de <span class="elsevierStyleItalic">Necator americanus</span> y <span class="elsevierStyleItalic">Ancylostoma duodenale</span> en poblaciones end&#233;micas de Buenos Aires y Misiones"
        ]
      ]
      "contieneResumen" => array:2 [
        "en" => true
        "es" => true
      ]
      "contieneTextoCompleto" => array:1 [
        "en" => true
      ]
      "contienePdf" => array:1 [
        "en" => true
      ]
      "resumenGrafico" => array:2 [
        "original" => 0
        "multimedia" => array:7 [
          "identificador" => "fig0010"
          "etiqueta" => "Figure 2"
          "tipo" => "MULTIMEDIAFIGURA"
          "mostrarFloat" => true
          "mostrarDisplay" => false
          "figura" => array:1 [
            0 => array:4 [
              "imagen" => "gr2.jpeg"
              "Alto" => 2505
              "Ancho" => 2925
              "Tamanyo" => 589855
            ]
          ]
          "descripcion" => array:1 [
            "en" => "<p id="spar0025" class="elsevierStyleSimplePara elsevierViewall">Heat-map distance matrix showing percentage &#40;above diagonal&#41; and number &#40;below diagonal&#41; of base differences per site based on pairwise comparison of the partial fragment of the <span class="elsevierStyleItalic">cob</span> gene among the sequences obtained in this study&#44; and 4 sequences of available at the GenBank database&#46; There was a total of 169 positions in the final dataset&#46; Sequences obtained in the present study are in bold&#46;</p>"
          ]
        ]
      ]
      "autores" => array:1 [
        0 => array:2 [
          "autoresLista" => "Andrea Servi&#225;n, Silvia A&#46; Repetto, Mar&#237;a Lorena Zonta, Graciela T&#46; Navone"
          "autores" => array:4 [
            0 => array:2 [
              "nombre" => "Andrea"
              "apellidos" => "Servi&#225;n"
            ]
            1 => array:2 [
              "nombre" => "Silvia A&#46;"
              "apellidos" => "Repetto"
            ]
            2 => array:2 [
              "nombre" => "Mar&#237;a"
              "apellidos" => "Lorena Zonta"
            ]
            3 => array:2 [
              "nombre" => "Graciela T&#46;"
              "apellidos" => "Navone"
            ]
          ]
        ]
      ]
      "resumen" => array:1 [
        0 => array:3 [
          "titulo" => "Highlights"
          "clase" => "author-highlights"
          "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><p id="spar0005" class="elsevierStyleSimplePara elsevierViewall"><ul class="elsevierStyleList" id="lis0005"><li class="elsevierStyleListItem" id="lsti0005"><span class="elsevierStyleLabel">&#8226;</span><p id="par0005" class="elsevierStylePara elsevierViewall">Microscopic and PCR methods may be used as complementary tools to diagnose hookworms&#46;</p></li><li class="elsevierStyleListItem" id="lsti0010"><span class="elsevierStyleLabel">&#8226;</span><p id="par0010" class="elsevierStylePara elsevierViewall">All hookworm infections in this study were caused by <span class="elsevierStyleItalic">Necator americanus</span>&#46;</p></li><li class="elsevierStyleListItem" id="lsti0015"><span class="elsevierStyleLabel">&#8226;</span><p id="par0015" class="elsevierStylePara elsevierViewall">This is the first study to provide molecular data on <span class="elsevierStyleItalic">Necator americanus</span> in Argentina&#46;</p></li><li class="elsevierStyleListItem" id="lsti0020"><span class="elsevierStyleLabel">&#8226;</span><p id="par0020" class="elsevierStylePara elsevierViewall">Imported hookworm infections found here highlight the importance of their assessment in non-endemic areas&#46;</p></li></ul></p></span>"
        ]
      ]
    ]
    "idiomaDefecto" => "en"
    "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S0325754122000311?idApp=UINPBA00004N"
    "url" => "/03257541/0000005400000004/v1_202212150521/S0325754122000311/v1_202212150521/en/main.assets"
  ]
  "en" => array:22 [
    "idiomaDefecto" => true
    "cabecera" => "<span class="elsevierStyleTextfn">Brief report</span>"
    "titulo" => "First evidence of systemic infection and specific cytotoxic T lymphocyte immune response evoked by oral infection with recombinant <span class="elsevierStyleItalic">Salmonella enterica</span> serovar Albany-Ovalbumin in C57BL&#47;6 mice"
    "tieneTextoCompleto" => true
    "paginas" => array:1 [
      0 => array:2 [
        "paginaInicial" => "282"
        "paginaFinal" => "287"
      ]
    ]
    "autores" => array:1 [
      0 => array:4 [
        "autoresLista" => "Carolina Mur&#250;a-L&#243;pez, Priscilla Gonz&#225;lez, Stephanie Almeida-Luna, Adriana Garibay-Escobar, Maribel Aguilar-Medina, Evangelina Beltr&#225;n-L&#243;pez, H&#233;ctor Samuel L&#243;pez-Moreno"
        "autores" => array:7 [
          0 => array:3 [
            "nombre" => "Carolina"
            "apellidos" => "Mur&#250;a-L&#243;pez"
            "referencia" => array:2 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">a</span>"
                "identificador" => "aff0005"
              ]
              1 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">1</span>"
                "identificador" => "fn0005"
              ]
            ]
          ]
          1 => array:3 [
            "nombre" => "Priscilla"
            "apellidos" => "Gonz&#225;lez"
            "referencia" => array:2 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">a</span>"
                "identificador" => "aff0005"
              ]
              1 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">1</span>"
                "identificador" => "fn0005"
              ]
            ]
          ]
          2 => array:3 [
            "nombre" => "Stephanie"
            "apellidos" => "Almeida-Luna"
            "referencia" => array:1 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">a</span>"
                "identificador" => "aff0005"
              ]
            ]
          ]
          3 => array:3 [
            "nombre" => "Adriana"
            "apellidos" => "Garibay-Escobar"
            "referencia" => array:1 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">b</span>"
                "identificador" => "aff0010"
              ]
            ]
          ]
          4 => array:3 [
            "nombre" => "Maribel"
            "apellidos" => "Aguilar-Medina"
            "referencia" => array:1 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">a</span>"
                "identificador" => "aff0005"
              ]
            ]
          ]
          5 => array:3 [
            "nombre" => "Evangelina"
            "apellidos" => "Beltr&#225;n-L&#243;pez"
            "referencia" => array:1 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">a</span>"
                "identificador" => "aff0005"
              ]
            ]
          ]
          6 => array:4 [
            "nombre" => "H&#233;ctor Samuel"
            "apellidos" => "L&#243;pez-Moreno"
            "email" => array:1 [
              0 => "hslmoreno@me.com"
            ]
            "referencia" => array:2 [
              0 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">a</span>"
                "identificador" => "aff0005"
              ]
              1 => array:2 [
                "etiqueta" => "<span class="elsevierStyleSup">&#42;</span>"
                "identificador" => "cor0005"
              ]
            ]
          ]
        ]
        "afiliaciones" => array:2 [
          0 => array:3 [
            "entidad" => "Laboratorio de Biomedicina Molecular&#44; Posgrado en Biotecnolog&#237;a y Posgrado en Ciencias Biom&#233;dicas&#44; Facultad de Ciencias Qu&#237;mico Biol&#243;gicas&#44; Universidad Aut&#243;noma de Sinaloa&#44; Sinaloa&#44; Mexico"
            "etiqueta" => "a"
            "identificador" => "aff0005"
          ]
          1 => array:3 [
            "entidad" => "Facultad de Ciencias Qu&#237;mico Biol&#243;gicas&#44; Universidad de Sonora&#44; Hermosillo&#44; Sonora&#44; Mexico"
            "etiqueta" => "b"
            "identificador" => "aff0010"
          ]
        ]
        "correspondencia" => array:1 [
          0 => array:3 [
            "identificador" => "cor0005"
            "etiqueta" => "&#8270;"
            "correspondencia" => "Corresponding author&#46;"
          ]
        ]
      ]
    ]
    "titulosAlternativos" => array:1 [
      "es" => array:1 [
        "titulo" => "Primera evidencia de infecci&#243;n sist&#233;mica y respuesta inmune celular citot&#243;xica espec&#237;fica evocada por la infecci&#243;n oral en ratones C57BL&#47;6 con <span class="elsevierStyleItalic">Salmonella enterica</span> serovar Albany-Ovoalb&#250;mina recombinante"
      ]
    ]
    "resumenGrafico" => array:2 [
      "original" => 0
      "multimedia" => array:7 [
        "identificador" => "fig0025"
        "etiqueta" => "Figure 2"
        "tipo" => "MULTIMEDIAFIGURA"
        "mostrarFloat" => true
        "mostrarDisplay" => false
        "figura" => array:1 [
          0 => array:4 [
            "imagen" => "gr2.jpeg"
            "Alto" => 1810
            "Ancho" => 3175
            "Tamanyo" => 213963
          ]
        ]
        "descripcion" => array:1 [
          "en" => "<p id="spar0030" class="elsevierStyleSimplePara elsevierViewall">rSAO evokes an epitope-specific CTL response with a Th-1 cytokine profile <span class="elsevierStyleItalic">in</span><span class="elsevierStyleItalic">vivo</span>&#46;</p> <p id="spar0035" class="elsevierStyleSimplePara elsevierViewall">&#40;A&#41; Percentages of specific lysis observed at effector&#58;target ratios 1&#58;1&#44; 10&#58;1 and 50&#58;1&#46; Data are means<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>SEM from three independent experiments&#46; &#40;B&#41; Concentrations of cytokines IFN-&#947;&#44; TNF-&#945;&#44; IL-2&#44; IL-6&#44; IL-4&#44; IL-10 and IL-17A in sera of infected mice 7 days pi&#46; &#40;&#42;&#41; Indicates statistical significance at <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#8804;<span class="elsevierStyleHsp" style=""></span>0&#46;05 using the Student&#39;s <span class="elsevierStyleItalic">t</span> test&#46; Data are means<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>SEM &#40;<span class="elsevierStyleItalic">n</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>3 mice per group&#41; from three independent experiments&#46; rSAO and rSTO are recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany and <span class="elsevierStyleItalic">S</span>&#46; Typhimurium that express the Ovalbumin antigen&#44; respectively&#46;</p>"
        ]
      ]
    ]
    "textoCompleto" => "<span class="elsevierStyleSections"><p id="par0020" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">Salmonella enterica</span> serovar Albany &#40;<span class="elsevierStyleItalic">S</span>&#46; Albany&#41; is classified as a non-typhoidal <span class="elsevierStyleItalic">Salmonella</span>&#46; This serovar was previously isolated by our group in food and feces of captive carnivores in a zoo located in northwestern Mexico<a class="elsevierStyleCrossRef" href="#bib0105"><span class="elsevierStyleSup">10</span></a>&#46; Afterwards&#44; it was also associated with the death of an ocelot &#40;<span class="elsevierStyleItalic">Leopardus</span><span class="elsevierStyleItalic">pardalis</span>&#41; in the same zoo<a class="elsevierStyleCrossRef" href="#bib0100"><span class="elsevierStyleSup">9</span></a>&#46; Additionally&#44; this serovar was also involved in a lethal case of a 65-year-old patient<a class="elsevierStyleCrossRef" href="#bib0110"><span class="elsevierStyleSup">11</span></a>&#46; This zoonotic and lethal potential support our aim to start with the study of important aspects about the host-<span class="elsevierStyleItalic">S</span>&#46; Albany relationship&#46; Here we analyzed the <span class="elsevierStyleItalic">in vivo</span> infection and specific <span class="elsevierStyleItalic">ex vivo</span> CTL immune response using the oral administration of the recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany-Ovalbumin &#40;rSAO&#41; strain in a murine model&#46;</p><p id="par0025" class="elsevierStylePara elsevierViewall">All the reagents used in this study were purchased from Merk &#40;Mexico&#41;&#46; Once isolated&#44; <span class="elsevierStyleItalic">S</span>&#46; Albany was transformed with the Ovalbumin &#40;Ova&#41; gene using pST13-OVA&#44; which codifies the OmpC porin from <span class="elsevierStyleItalic">Salmonella</span> Typhi &#40;gently donated by Dr&#46; Ortiz-Navarrete&#44; from CINVESTAV&#44; M&#233;xico&#41; to obtain our recombinant bacterium designated as rSAO&#46; <span class="elsevierStyleItalic">S&#46;</span> Typhimurium 14028 transformed with the same plasmid&#44; designated as rSTO &#40;also donated by Dr&#46; Ortiz-Navarrete&#41;&#44; and used as positive control&#46; After the transformation&#44; the Ova expression was confirmed by Western blotting using a homemade mouse polyclonal &#945;-Ova diluted 1&#58;100 as first antibody and rabbit &#945;-mouse IgG conjugated to horse-radish peroxidase diluted 1&#58;3000 as second antibody&#46; The reactions were revealed following protocols previously established in our investigation group using diaminobenzidine and H<span class="elsevierStyleInf">2</span>O<span class="elsevierStyleInf">2</span><a class="elsevierStyleCrossRef" href="#bib0095"><span class="elsevierStyleSup">8</span></a>&#46; Three groups consisting of 3 C57BL&#47;6 mice &#40;6&#8211;8 weeks-old&#41; each were formed&#46; While the first group was orally infected using 10<span class="elsevierStyleSup">6</span> colony-forming units &#40;CFU&#41; of rSAO&#44; the second group was infected with 10<span class="elsevierStyleSup">5</span><span class="elsevierStyleHsp" style=""></span>CFU of rSTO&#44; due to the fact that <span class="elsevierStyleItalic">S</span>&#46; Typhimurium was lethal for C57BL&#47;6 mice&#46; The mice in the third group were uninfected as previously described<a class="elsevierStyleCrossRef" href="#bib0065"><span class="elsevierStyleSup">2</span></a>&#46; Six hours post-infection &#40;pi&#41; the feces were collected&#44; weighed&#44; diluted&#44; homogenized in phosphate buffered-saline &#40;PBS&#41; pH 7&#46;2 and plated on brain heart-infusion &#40;BHI&#41; agar supplemented with ampicillin at 100<span class="elsevierStyleHsp" style=""></span>&#956;g&#47;ml&#59; 24<span class="elsevierStyleHsp" style=""></span>h after incubation at 37<span class="elsevierStyleHsp" style=""></span>&#176;C&#44; CFU&#47;mg were determined by colony counting&#46; The mice were monitored every day&#44; weighed&#44; and clinically analyzed according to the scale previously proposed for evaluating mice clinical signs in response to <span class="elsevierStyleItalic">Salmonella</span> infection<a class="elsevierStyleCrossRef" href="#bib0080"><span class="elsevierStyleSup">5</span></a>&#46; Briefly&#44; the following five clinical criteria were evaluated&#58; general activity level&#44; posture&#44; tremors&#44; fur appearance and finally change in eating habits&#44; which was determined by weighing the food&#46; Each of these aspects was evaluated from a scale of 0&#8211;2 for every mouse&#44; where a score of 0 was assigned to severely sick mice&#44; indicating a great presence of the criteria evaluated&#46; This could be evidenced by low activity or a significant decrease in movement&#44; a constant hunch over posture&#44; the presence of tremors&#44; greasy fur appearance and a significant decrease in appetite&#46; Likewise&#44; a score of 1 was assigned to moderately sick mice with a small presence of the clinical sign evaluated&#46; Lastly&#44; a score of 2 corresponded to healthy mice with no change in activity or no signs of sickness<a class="elsevierStyleCrossRef" href="#bib0080"><span class="elsevierStyleSup">5</span></a>&#46; At day 7 pi&#44; all the clinical criteria values were added&#46; If the final value ranged from 8 to 10&#44; the mice were considered healthy&#59; from 4 to 7&#44; moderately sick&#59; and from 0 to 3&#44; severely sick&#46; Afterwards&#44; the mice were euthanized and their spleens were aseptically removed&#44; measured&#44; and splenocytes were counted using a Neubauer chamber and Trypan Blue 0&#46;4&#37;&#46; To determine systemic infection&#44; splenocytes and bone marrow cells were lysed using 1&#37; TritonX-100&#44; and intracellular bacteria were plated on BHI agar supplemented with ampicillin at 100<span class="elsevierStyleHsp" style=""></span>&#956;g&#47;ml&#59; 24<span class="elsevierStyleHsp" style=""></span>h after incubation at 37<span class="elsevierStyleHsp" style=""></span>&#176;C&#44; CFU were determined by colony counting&#46;</p><p id="par0030" class="elsevierStylePara elsevierViewall">The cytotoxic effector T lymphocytes were obtained by <span class="elsevierStyleItalic">in vitro</span> stimulation with SIINFEKL &#40;K<span class="elsevierStyleSup">b</span> restricted OVA-peptide&#44; synthesized by Sigma-Aldrich&#44; Mexico&#41; for 5 days&#44; following standard protocols&#46; Likewise&#44; 1<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleSup">6</span> RMA-S cells &#40;also donated by Dr&#46; Ortiz from CINVESTAV-IPN&#41; were stained with CFSE and pulsed with SIINFEKL for 4<span class="elsevierStyleHsp" style=""></span>h prior to the interactions with the cytotoxic effector T lymphocytes &#40;E&#41;&#46; RMA-S cells were deemed target cells &#40;T&#41; and effector cells were adjusted to 1&#58;1&#44; 10&#58;1 and 50&#58;1 effector&#58;target &#40;E&#58;T&#41; ratios to evaluate antigen specific lysis&#59; any unspecific lysis was eliminated by normalizing the data with results obtained against an irrelevant peptide &#40;TAVIRADN&#41;&#46; The samples were assessed on FACS Accuri C6 &#40;BD&#41; and analyzed using FlowJo software<a class="elsevierStyleCrossRef" href="#bib0075"><span class="elsevierStyleSup">4</span></a>&#46;</p><p id="par0035" class="elsevierStylePara elsevierViewall">Additionally&#44; the sera of uninfected or infected rSAO or rSTO mice were collected on day 7 pi and were used to determine the cytokine profile associated with the T helper &#40;Th&#41; subset immune response using a CBA Mouse Cytokine Kit &#40;BD&#41;&#46; For Th1&#44; the cytokines evaluated were&#58; IFN-&#947;&#44; TNF-&#945;&#44; IL-2 and IL-6&#44; for Th2&#58; IL-4 and IL-10 and for Th17&#58; IL-17&#44; according to the manufacturer&#39;s instructions&#46; The data were acquired on a FACS Accuri C6 &#40;BD&#41; and analyzed using FlowJo software&#46; The data obtained were analyzed using Student&#39;s <span class="elsevierStyleItalic">t</span> test on GraphPad Prism 6&#44; where <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#8804;<span class="elsevierStyleHsp" style=""></span>0&#46;05 was considered to be statistically significant&#46;</p><p id="par0040" class="elsevierStylePara elsevierViewall">The Ova expression in our recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany strain was confirmed by the Western blot analysis &#40;<a class="elsevierStyleCrossRef" href="#fig0020">Fig&#46; 1</a>A&#41;&#44; where it was possible to observe a clear band of approximately 45 kDa in crude extract of rSAO&#44; consistent with commercial Ova as positive control in lane 4&#46; The clinical findings of infected as well as uninfected mice are summarized in <a class="elsevierStyleCrossRef" href="#fig0020">Figure 1</a>B&#44; and can also be observed in <a class="elsevierStyleCrossRef" href="#sec0025">Supplementary Figure 1</a>&#44; where infected mice with rSAO presented a clinical score of 5&#44; which is consistent with that of moderately sick mice&#44; suggesting that this serovar has the capacity to cause characteristic symptoms of an infection by <span class="elsevierStyleItalic">Salmonella</span>&#46; Similar results were obtained from mice infected with the positive control rSTO&#44; while the small differences observed can be explained by the initial inoculum concentration&#46; Both groups presented signs consistent with <span class="elsevierStyleItalic">Salmonella</span> infection&#44; which was considered an initial indicator of the pathogenic potential of rSAO&#46; Various groups have described that along with the behavioral aspects&#44; mice tend to lose weight during infection&#44; and we observed that mice infected with rSAO presented a total average weight loss of 16&#37;&#44; suggesting that the clinical signs observed were significant enough to cause a decrease in appetite in infected mice&#44; and showing that rSAO can display its pathogenic capability &#40;<a class="elsevierStyleCrossRef" href="#fig0020">Fig&#46; 1</a>C&#41;&#46; To confirm this intestinal infection&#44; the feces collected at 6<span class="elsevierStyleHsp" style=""></span>h pi were plated and the mean CFU observed in rSAO infected mice was of 1650<span class="elsevierStyleHsp" style=""></span>CFU&#47;mg&#44; showing no statistical difference with respect to the rSTO-infected mice &#40;<a class="elsevierStyleCrossRef" href="#fig0020">Fig&#46; 1</a>D&#41;&#46; These results suggest that rSAO can colonize the intestine&#44; establishing a gastrointestinal infection in mice&#46; Our findings are additionally supported by a similar infection study conducted recently&#44; where it was determined that <span class="elsevierStyleItalic">S</span>&#46; Albany can establish infection and cause pathological changes in tissue in the small intestine&#44; cecum and colon<a class="elsevierStyleCrossRef" href="#bib0090"><span class="elsevierStyleSup">7</span></a>&#46; To further assess the infection caused by rSAO&#44; the spleens were extracted on day 7 pi and measured&#59; the spleens from mice infected with rSAO presented a mean of 0&#46;6 inches in length&#44; exhibiting a statistically significant difference with respect to the spleens extracted from negative control mice &#40;<a class="elsevierStyleCrossRef" href="#fig0020">Fig&#46; 1</a>E&#41;&#46; As well as a visible increase in size&#44; it was possible to observe similar macroscopic characteristics in the spleens from mice infected with the positive control rSTO&#46; To demonstrate the possible increased cellularity in infected mice responsible for the increase in spleen size&#44; splenocytes were counted and although the values were not statistically significant&#44; it was evident that there were approximately 0&#46;5 time more cells in the spleens extracted from rSAO and rSTO infected mice when compared to spleens extracted from the non-infected control group &#40;see <a class="elsevierStyleCrossRef" href="#fig0020">Figure 1</a>f&#41;&#46; Once cellularity was determined&#44; splenocytes were lysed to assess rSAO to evidence its capacity to cause a systemic infection&#59; the mean observed for CFUs in splenocytes of rSAO infected mice was of 3&#46;98<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleSup">5</span> per million cells &#40;see <a class="elsevierStyleCrossRef" href="#fig0020">Fig&#46; 1</a>G&#41;&#46; Parallelly&#44; we determined the bacterial burden in total bone marrow cells to further assess the systemic infection and the mean observed was of 1&#46;87<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleSup">5</span> CFUs in total bone marrow cells&#44; demonstrating the capacity of this serovar to establish a systemic infection in mice&#44; thus highlighting its pathogenic potential &#40;see <a class="elsevierStyleCrossRef" href="#fig0020">Fig&#46; 1</a>H&#41;&#46; A possible explanation&#44; at least in part&#44; for our data&#44; can be related to reports that <span class="elsevierStyleItalic">S&#46;</span> Typhimurium can cause systemic disease in C57BL&#47;6 infected mice&#44; by persisting in B lymphocytes and precursors specifically within the bone marrow<a class="elsevierStyleCrossRef" href="#bib0060"><span class="elsevierStyleSup">1</span></a>&#46; Maybe rSAO&#44; as well as rSTO&#44; are capable of infecting and establishing a niche in memory B cells and thus&#44; chronic infection&#46; However&#44; more detailed studies should be conducted&#46;</p><elsevierMultimedia ident="fig0020"></elsevierMultimedia><p id="par0045" class="elsevierStylePara elsevierViewall">CTL antigen-specific lysis demonstrated that effector cells obtained from rSAO-infected mice were able to kill RMA-S cells pulsed with SIINFEKL&#44; but were inefficient when these target cells were pulsed with an irrelevant peptide&#46; It is relevant to mention that although splenocytes were infected with recombinant bacteria&#44; memory effector T CD8<span class="elsevierStyleSup">&#43;</span> clonal expansion was clearly decreased in the irrelevant peptide pulsed group&#44; in contrast with the greater clonal expansion observed in the group pulsed with the SIINFEKL epitope&#44; as shown in <a class="elsevierStyleCrossRef" href="#sec0025">Supplementary Figure 3</a>&#46; These results demonstrate that although there is small participation of the genetic construction used&#44; it is not significant&#46; Effector cells evoked 15&#46;6&#37; of specific lysis for 1&#58;1 E&#58;T ratio&#44; 45&#46;1&#37; for 10&#58;1 E&#58;T ratio and 60&#37; for 50&#58;1 E&#58;T ratio&#46; Therefore&#44; CTL from rSAO infected mice recognize this epitope and generate a K<span class="elsevierStyleSup">b</span>-restricted CTL response against <span class="elsevierStyleItalic">S&#46;</span> Albany &#40;<a class="elsevierStyleCrossRef" href="#fig0025">Fig&#46; 2</a>A&#41;&#46; To determine the cytokine profile evoked <span class="elsevierStyleItalic">in vivo</span> by rSAO&#44; the sera of infected and uninfected mice were analyzed&#44; and we observed a predominance of the Th1-like cytokine profile &#40;<a class="elsevierStyleCrossRef" href="#fig0025">Fig&#46; 2</a>B&#41;&#46; Concentrations of IL-4 and IL-10 were 4 and 5<span class="elsevierStyleMonospace">Y</span>pg&#47;ml respectively&#46; IL-6 and IL-2 were both close to 40<span class="elsevierStyleMonospace">Y</span>pg&#47;ml&#44; TNF-&#945; concentrations were of 20<span class="elsevierStyleMonospace">Y</span>pg&#47;ml&#44; but the IFN-&#947; concentration was the predominant cytokine&#44; reaching levels of 200<span class="elsevierStyleMonospace">Y</span>pg&#47;ml in rSAO-infected mice&#46; These results were consistent with what was observed in our positive control group rSTO&#44; where Th1 cytokines IFN-&#947;&#44; IL-2&#44; IL-6 and TNF-&#945; were predominant&#44; further suggesting immune response similarities evoked by both serovars&#46; Along with the importance of the Th1 cytokine profile&#44; it has also been reported that the presence of IL-17A along with the Th1 cytokine profile is necessary in controlling <span class="elsevierStyleItalic">Salmonella</span> infection&#46; However&#44; these findings also show that its presence is predominant in the initial stages of infection because its primary function is the recruitment of neutrophils at the initial stages of inflammation<a class="elsevierStyleCrossRef" href="#bib0065"><span class="elsevierStyleSup">2</span></a>&#46; Because we determined the levels of all the cytokines in the final stages of infection&#44; this could be a possible explanation as to why no statistical difference in the levels of IL-17A was observed across our experimental groups&#46; The secretion of Th1 profile cytokines&#44; specifically IFN-&#947;&#44; has been previously described in <span class="elsevierStyleItalic">S&#46;</span> Typhimurium infections&#44; where macrophages must be activated to eliminate the intracellular bacteria&#46; Thus&#44; these results suggest that rSAO could establish intracellular infection&#44; evoking protective immunity in mice&#44; which is characterized by the secretion of the Th1 cytokine profile&#46;</p><elsevierMultimedia ident="fig0025"></elsevierMultimedia><p id="par0050" class="elsevierStylePara elsevierViewall">Infection by <span class="elsevierStyleItalic">S</span>&#46; Typhimurium has been thoroughly characterized in C57BL&#47;6 mice and has been used as the comparative gold standard model for other potentially pathogenic <span class="elsevierStyleItalic">Salmonella</span> serovars<a class="elsevierStyleCrossRef" href="#bib0070"><span class="elsevierStyleSup">3</span></a>&#46; This was consistent with <span class="elsevierStyleItalic">S&#46;</span> Albany&#44; a serovar that had not been previously reported in our region&#44; until its isolation by our investigation group in a zoo from northwestern Mexico&#44; and its further association with the death of an ocelot &#40;<span class="elsevierStyleItalic">Leopardus pardalis</span>&#41; in the same zoo<a class="elsevierStyleCrossRefs" href="#bib0100"><span class="elsevierStyleSup">9&#44;10</span></a>&#44; as it was the first indicator of the pathogenic potential of <span class="elsevierStyleItalic">S</span>&#46; Albany&#46; In this study we have demonstrated for the first time that <span class="elsevierStyleItalic">S</span>&#46; Albany has the pathogenic potential to establish a systemic infection in C57BL&#47;6 mice&#44; displaying the capability to evoke a specific CTL immune response&#46; We found that mice that were orally infected with rSAO exhibited similar nociceptive signs as those infected with the positive control rSTO&#44; suggesting that rSAO has the capability to establish a systemic infection such as rSTO&#46; In addition to these findings&#44; we also demonstrated that rSAO infected mice lost weight in response to the infection&#44; which could be explained by the loss of appetite caused by the strong symptoms induced by the rSAO infection&#46; Likewise&#44; it could also be attributed to the high energy expenditure required to fight off the infection&#44; suggesting that rSAO was capable of establishing a strong enough infection to cause symptoms that directly impacted on the mice appetite and therefore their weight&#46; The capability of rSAO to establish an infection in the gastrointestinal tract was demonstrated by the bacteria isolated from the feces collected 6<span class="elsevierStyleHsp" style=""></span>h pi&#46; The number of colonies exhibited no statistical differences compared with rSTO- infected mice&#44; suggesting that rSAO had the ability to resist stomach acidic conditions and pass through the intestinal mucosa to start infection&#46; This also suggests that rSAO expresses a set of vital virulence factors for establishing infection such as Type III Secretion Systems&#44; Sop and Inv proteins&#44; as well as niche factors that would allow it to overcome bile salts&#44; high osmolarity and competition for nutrients in the intestine&#59; however&#44; to confirm these molecular mechanisms&#44; more detailed studies should be conducted&#46; It has also been previously reported that murine <span class="elsevierStyleItalic">S&#46;</span> Typhimurium infection evokes an increase in spleen size&#44; probably due to the specific clonal expansion of resident immunity cells from this secondary immune organ<a class="elsevierStyleCrossRef" href="#bib0085"><span class="elsevierStyleSup">6</span></a>&#46;</p><p id="par0055" class="elsevierStylePara elsevierViewall">The specific lysis observed demonstrated that this response was specific for cells harboring the SIINFEKL epitope and CTL were activated by the rSAO infection&#44; generating a specific <span class="elsevierStyleItalic">in vivo</span> immune response&#46; The high concentrations of IFN-&#947; and TNF-&#945; obtained in our experimental conditions could explain the high percentages of specific lysis as well as the severity of the nociceptive signs observed in infected mice&#44; suggesting that although the infection was strong enough to evoke an immune response&#44; it was probably insufficient to resolve the infection&#59; however&#44; a more detailed study should be conducted to confirm this hypothesis&#46;</p><p id="par0060" class="elsevierStylePara elsevierViewall">We demonstrated here&#44; for the first time that <span class="elsevierStyleItalic">S&#46;</span> Albany causes a systemic disease in C57BL&#47;6 mice and has the same potential to infect other mammalian hosts&#44; including humans&#44; probably expressing a set of virulence factors&#44; pathogenicity islands and secretion systems&#44; although further molecular analyses are needed&#46; Additionally&#44; the isolation of rSAO in bone marrow cells suggest a chronic systemic infection&#44; such as human typhoid fever&#46; The use of recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany-Ovalbumin strain as an antigenic flag represents an important biotechnological tool to study <span class="elsevierStyleItalic">in vivo</span> immune responses&#46; Finally&#44; the CTL immune response evidenced here supports the possibility to manipulate the cellular immune response to eliminate this bacterium effectively&#46;</p><span id="sec0005" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0030">Authorship contribution</span><p id="par0065" class="elsevierStylePara elsevierViewall">CML&#44; PG&#44; SAL&#44; AGE&#44; MAM&#44; EBL&#44; HSLM&#58; Investigation&#44; experimental procedures&#44; data analysis&#44; writing and editing the manuscript&#46; Also&#44; HSLM&#58; design the project&#44; obtained the grant support&#44; and validation of data&#46;</p></span><span id="sec0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0035">Conflicts of interest</span><p id="par0070" class="elsevierStylePara elsevierViewall">None&#46;</p></span><span id="sec0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0040">Ethical responsibilities</span><p id="par0075" class="elsevierStylePara elsevierViewall">None&#46;</p></span></span>"
    "textoCompletoSecciones" => array:1 [
      "secciones" => array:10 [
        0 => array:3 [
          "identificador" => "xres1819626"
          "titulo" => "Abstract"
          "secciones" => array:1 [
            0 => array:1 [
              "identificador" => "abst0005"
            ]
          ]
        ]
        1 => array:2 [
          "identificador" => "xpalclavsec1588105"
          "titulo" => "Keywords"
        ]
        2 => array:2 [
          "identificador" => "xpalclavsec1588104"
          "titulo" => "Abbreviations"
        ]
        3 => array:3 [
          "identificador" => "xres1819625"
          "titulo" => "Resumen"
          "secciones" => array:1 [
            0 => array:1 [
              "identificador" => "abst0010"
            ]
          ]
        ]
        4 => array:2 [
          "identificador" => "xpalclavsec1588106"
          "titulo" => "Palabras clave"
        ]
        5 => array:2 [
          "identificador" => "sec0005"
          "titulo" => "Authorship contribution"
        ]
        6 => array:2 [
          "identificador" => "sec0010"
          "titulo" => "Conflicts of interest"
        ]
        7 => array:2 [
          "identificador" => "sec0015"
          "titulo" => "Ethical responsibilities"
        ]
        8 => array:2 [
          "identificador" => "xack641879"
          "titulo" => "Acknowledgements"
        ]
        9 => array:1 [
          "titulo" => "References"
        ]
      ]
    ]
    "pdfFichero" => "main.pdf"
    "tienePdf" => true
    "fechaRecibido" => "2020-11-10"
    "fechaAceptado" => "2022-05-02"
    "PalabrasClave" => array:2 [
      "en" => array:2 [
        0 => array:4 [
          "clase" => "keyword"
          "titulo" => "Keywords"
          "identificador" => "xpalclavsec1588105"
          "palabras" => array:3 [
            0 => "Cytotoxic T lymphocytes"
            1 => "<span class="elsevierStyleItalic">Salmonella</span>  <span class="elsevierStyleItalic">enterica</span> serovar Albany"
            2 => "Recombinant bacterium"
          ]
        ]
        1 => array:4 [
          "clase" => "abr"
          "titulo" => "Abbreviations"
          "identificador" => "xpalclavsec1588104"
          "palabras" => array:10 [
            0 => "rSAO"
            1 => "rSTO"
            2 => "Ova"
            3 => "CTL"
            4 => "TNF-&#945;"
            5 => "IFN-&#947;"
            6 => "IL"
            7 => "pi"
            8 => "CFU"
            9 => "BHI"
          ]
        ]
      ]
      "es" => array:1 [
        0 => array:4 [
          "clase" => "keyword"
          "titulo" => "Palabras clave"
          "identificador" => "xpalclavsec1588106"
          "palabras" => array:3 [
            0 => "Linfocito T citot&#243;xico"
            1 => "<span class="elsevierStyleItalic">Salmonella</span>  <span class="elsevierStyleItalic">enterica</span> serovar Albany"
            2 => "Bacteria recombinante"
          ]
        ]
      ]
    ]
    "tieneResumen" => true
    "resumen" => array:2 [
      "en" => array:2 [
        "titulo" => "Abstract"
        "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><p id="spar0010" class="elsevierStyleSimplePara elsevierViewall">Our group isolated <span class="elsevierStyleItalic">Salmonella enterica</span> serovar Albany from food and feces of wild captive carnivores in a zoo from northwestern Mexico&#46; This serovar was also associated with the death of an ocelot &#40;<span class="elsevierStyleItalic">Leopardus pardalis</span>&#41; in the same zoo&#46; Another group associated <span class="elsevierStyleItalic">S&#46;</span> Albany with the death of a human patient&#46; It is due to this zoonotic potential that the <span class="elsevierStyleItalic">in vivo</span> study of the host-<span class="elsevierStyleItalic">S</span>&#46; Albany relationship is critical&#46; The recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany-Ovalbumin &#40;rSAO&#41; strain was used to analyze a murine oral infection and its specific cytotoxic T lymphocyte &#40;CTL&#41; response&#46; Our results have shown for the first time that rSAO establishes a systemic infection and evokes epitope-specific lysis with a Th1-like cytokine profile <span class="elsevierStyleItalic">in vivo</span>&#46;</p></span>"
      ]
      "es" => array:2 [
        "titulo" => "Resumen"
        "resumen" => "<span id="abst0010" class="elsevierStyleSection elsevierViewall"><p id="spar0015" class="elsevierStyleSimplePara elsevierViewall"><span class="elsevierStyleItalic">Salmonella enterica</span> serovar Albany fue aislada por nuestro grupo de investigaci&#243;n de alimentos contaminados y de heces de animales carn&#237;voros en cautiverio en un zool&#243;gico del noroeste de M&#233;xico&#59; posteriormente&#44; se logr&#243; asociar a este serovar con la muerte de un ocelote &#40;<span class="elsevierStyleItalic">Leopardus pardalis</span>&#41;&#44; dentro de este mismo zool&#243;gico&#46; Otro grupo de investigaci&#243;n asoci&#243; a este serovar con la muerte de un paciente&#46; Es debido a este potencial zoon&#243;tico que el estudio <span class="elsevierStyleItalic">in vivo</span> de la relaci&#243;n hospedero-<span class="elsevierStyleItalic">S</span>&#46; Albany es cr&#237;tico&#46; La cepa recombinante <span class="elsevierStyleItalic">S</span>&#46; Albany-Ovoalb&#250;mina &#40;rSAO&#41; fue utilizada para analizar la infecci&#243;n m&#250;rida&#44; al igual que la respuesta inmune celular citot&#243;xica espec&#237;fica&#46; Nuestros resultados demuestran&#44; por primera vez&#44; que rSAO establece una infecci&#243;n sist&#233;mica y evoca lisis ep&#237;topo-espec&#237;fica con un perfil de citocinas tipo Th1 <span class="elsevierStyleItalic">in vivo</span>&#46;</p></span>"
      ]
    ]
    "NotaPie" => array:1 [
      0 => array:3 [
        "etiqueta" => "1"
        "nota" => "<p class="elsevierStyleNotepara" id="npar0005">These authors contributed equally to this article&#46;</p>"
        "identificador" => "fn0005"
      ]
    ]
    "apendice" => array:1 [
      0 => array:1 [
        "seccion" => array:1 [
          0 => array:4 [
            "apendice" => "<p id="par0090" class="elsevierStylePara elsevierViewall">The following are Supplementary data to this article&#58;<elsevierMultimedia ident="fig0005"></elsevierMultimedia><elsevierMultimedia ident="fig0010"></elsevierMultimedia><elsevierMultimedia ident="fig0015"></elsevierMultimedia></p>"
            "etiqueta" => "Appendix A"
            "titulo" => "Supplementary data"
            "identificador" => "sec0025"
          ]
        ]
      ]
    ]
    "multimedia" => array:5 [
      0 => array:7 [
        "identificador" => "fig0020"
        "etiqueta" => "Figure 1"
        "tipo" => "MULTIMEDIAFIGURA"
        "mostrarFloat" => true
        "mostrarDisplay" => false
        "figura" => array:1 [
          0 => array:4 [
            "imagen" => "gr1.jpeg"
            "Alto" => 1130
            "Ancho" => 2925
            "Tamanyo" => 154526
          ]
        ]
        "descripcion" => array:1 [
          "en" => "<p id="spar0020" class="elsevierStyleSimplePara elsevierViewall">rSAO causes systemic infection in C57BL&#47;6 mice&#46;</p> <p id="spar0025" class="elsevierStyleSimplePara elsevierViewall">&#40;A&#41; Ova expression by transformed <span class="elsevierStyleItalic">S</span>&#46; Albany&#59; Lane 1&#58; molecular weight marker&#44; Lane 2&#58; crude extract <span class="elsevierStyleItalic">S</span>&#46; Albany WT&#44; Lane 3&#58; crude extract rSAO&#44; Lane 4&#58; positive control Ova protein fraction V&#46; &#40;B&#41; Clinical findings in mice orally infected with 1<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleSup">6</span> CFU rSAO&#44; 1<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleSup">5</span> rSTO and uninfected mice&#46; &#40;C&#41; Weight loss observed in infected and uninfected mice at 1&#44; 3&#44; 5&#44; and 7 days pi&#46; &#40;D&#41; CFU per milligram of feces collected 6<span class="elsevierStyleHsp" style=""></span>h pi&#46; &#40;E&#41; Increase in spleen size measured in cm 7 days pi&#46; &#40;F&#41; Increase in cellularity observed in infected mice spleens 7 days pi&#46; &#40;G&#41; CFU per million cells in infected mice spleens at 7 days pi&#46; &#40;H&#41; CFU in total bone marrow cells at 7d pi&#46; &#40;&#42;&#41; Indicates statistical significance at <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#8804;<span class="elsevierStyleHsp" style=""></span>0&#46;05 using Student&#39;s <span class="elsevierStyleItalic">t</span> test&#46; Data are means<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>SEM &#40;n<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>3 mice per group&#41; from three independent experiments&#46; rSAO and rSTO are recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany and <span class="elsevierStyleItalic">S</span>&#46; Typhimurium that express the Ovalbumin antigen&#44; respectively&#46;</p>"
        ]
      ]
      1 => array:7 [
        "identificador" => "fig0025"
        "etiqueta" => "Figure 2"
        "tipo" => "MULTIMEDIAFIGURA"
        "mostrarFloat" => true
        "mostrarDisplay" => false
        "figura" => array:1 [
          0 => array:4 [
            "imagen" => "gr2.jpeg"
            "Alto" => 1810
            "Ancho" => 3175
            "Tamanyo" => 213963
          ]
        ]
        "descripcion" => array:1 [
          "en" => "<p id="spar0030" class="elsevierStyleSimplePara elsevierViewall">rSAO evokes an epitope-specific CTL response with a Th-1 cytokine profile <span class="elsevierStyleItalic">in</span><span class="elsevierStyleItalic">vivo</span>&#46;</p> <p id="spar0035" class="elsevierStyleSimplePara elsevierViewall">&#40;A&#41; Percentages of specific lysis observed at effector&#58;target ratios 1&#58;1&#44; 10&#58;1 and 50&#58;1&#46; Data are means<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>SEM from three independent experiments&#46; &#40;B&#41; Concentrations of cytokines IFN-&#947;&#44; TNF-&#945;&#44; IL-2&#44; IL-6&#44; IL-4&#44; IL-10 and IL-17A in sera of infected mice 7 days pi&#46; &#40;&#42;&#41; Indicates statistical significance at <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#8804;<span class="elsevierStyleHsp" style=""></span>0&#46;05 using the Student&#39;s <span class="elsevierStyleItalic">t</span> test&#46; Data are means<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>SEM &#40;<span class="elsevierStyleItalic">n</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>3 mice per group&#41; from three independent experiments&#46; rSAO and rSTO are recombinant <span class="elsevierStyleItalic">S</span>&#46; Albany and <span class="elsevierStyleItalic">S</span>&#46; Typhimurium that express the Ovalbumin antigen&#44; respectively&#46;</p>"
        ]
      ]
      2 => array:5 [
        "identificador" => "fig0005"
        "tipo" => "MULTIMEDIAFIGURA"
        "mostrarFloat" => false
        "mostrarDisplay" => true
        "figura" => array:1 [
          0 => array:4 [
            "imagen" => "mmc1.jpeg"
            "Alto" => 1088
            "Ancho" => 2500
            "Tamanyo" => 298546
          ]
        ]
      ]
      3 => array:5 [
        "identificador" => "fig0010"
        "tipo" => "MULTIMEDIAFIGURA"
        "mostrarFloat" => false
        "mostrarDisplay" => true
        "figura" => array:1 [
          0 => array:4 [
            "imagen" => "mmc2.jpeg"
            "Alto" => 684
            "Ancho" => 1500
            "Tamanyo" => 138680
          ]
        ]
      ]
      4 => array:5 [
        "identificador" => "fig0015"
        "tipo" => "MULTIMEDIAFIGURA"
        "mostrarFloat" => false
        "mostrarDisplay" => true
        "figura" => array:1 [
          0 => array:4 [
            "imagen" => "mmc3.jpeg"
            "Alto" => 726
            "Ancho" => 1500
            "Tamanyo" => 123593
          ]
        ]
      ]
    ]
    "bibliografia" => array:2 [
      "titulo" => "References"
      "seccion" => array:1 [
        0 => array:2 [
          "identificador" => "bibs0015"
          "bibliografiaReferencia" => array:11 [
            0 => array:3 [
              "identificador" => "bib0060"
              "etiqueta" => "1"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "B cell precursors are targets for <span class="elsevierStyleItalic">Salmonella</span> infection"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:6 [
                            0 => "D&#46; Castro-Eguiluz"
                            1 => "R&#46; Pelayo"
                            2 => "V&#46; Rosales-Garcia"
                            3 => "R&#46; Rosales-Reyes"
                            4 => "C&#46; Alpuche-Aranda"
                            5 => "V&#46; Ortiz-Navarrete"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:5 [
                        "tituloSerie" => "Microb Pathog"
                        "fecha" => "2009"
                        "volumen" => "47"
                        "paginaInicial" => "52"
                        "paginaFinal" => "56"
                      ]
                    ]
                  ]
                ]
              ]
            ]
            1 => array:3 [
              "identificador" => "bib0065"
              "etiqueta" => "2"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Interleukin-17A is required to suppress invasion of <span class="elsevierStyleItalic">Salmonella enterica</span> serovar Typhimurium to enteric mucosa"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:5 [
                            0 => "H&#46; Mayuzumi"
                            1 => "K&#46; Inagaki-Ohara"
                            2 => "C&#46; Uyttenhove"
                            3 => "Y&#46; Okamoto"
                            4 => "G&#46; Matsuzaki"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:5 [
                        "tituloSerie" => "Immunology"
                        "fecha" => "2010"
                        "volumen" => "131"
                        "paginaInicial" => "377"
                        "paginaFinal" => "385"
                      ]
                    ]
                  ]
                ]
              ]
            ]
            2 => array:3 [
              "identificador" => "bib0070"
              "etiqueta" => "3"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Entry route of <span class="elsevierStyleItalic">Salmonella</span> Typhimurium directs the type of induced immune response"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:3 [
                            0 => "C&#46; Martinoli"
                            1 => "A&#46; Chiavelli"
                            2 => "M&#46; Rescigno"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:2 [
                      "doi" => "10.1016/j.immuni.2007.10.011"
                      "Revista" => array:6 [
                        "tituloSerie" => "Immunity"
                        "fecha" => "2007"
                        "volumen" => "27"
                        "paginaInicial" => "975"
                        "paginaFinal" => "984"
                        "link" => array:1 [
                          0 => array:2 [
                            "url" => "https://www.ncbi.nlm.nih.gov/pubmed/18083577"
                            "web" => "Medline"
                          ]
                        ]
                      ]
                    ]
                  ]
                ]
              ]
            ]
            3 => array:3 [
              "identificador" => "bib0075"
              "etiqueta" => "4"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "The plasmacytoid dendritic cells evoke <span class="elsevierStyleItalic">Salmonella</span>-specific CTL effector response"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:3 [
                            0 => "C&#46; Mur&#250;a-L&#243;pez"
                            1 => "M&#46; Gonz&#225;lez-Orozco"
                            2 => "H&#46;S&#46; L&#243;pez-Moreno"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:2 [
                      "doi" => "10.7705/biomedica.v39i3.4415"
                      "Revista" => array:6 [
                        "tituloSerie" => "Biom&#233;dica"
                        "fecha" => "2019"
                        "volumen" => "39"
                        "paginaInicial" => "172"
                        "paginaFinal" => "181"
                        "link" => array:1 [
                          0 => array:2 [
                            "url" => "https://www.ncbi.nlm.nih.gov/pubmed/31529859"
                            "web" => "Medline"
                          ]
                        ]
                      ]
                    ]
                  ]
                ]
              ]
            ]
            4 => array:3 [
              "identificador" => "bib0080"
              "etiqueta" => "5"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "<span class="elsevierStyleItalic">Salmonella</span> Typhimurium infections in BALB&#47;c mice&#58; a comparison of tissue bioluminescence&#44; tissue cultures and mice clinical scores"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:7 [
                            0 => "H&#46; &#214;zkaya"
                            1 => "B&#46; Akcan"
                            2 => "G&#46; Aydemir"
                            3 => "S&#46; Aydin&#246;z"
                            4 => "Y&#46; Razia"
                            5 => "T&#46; Gammon"
                            6 => "J&#46; McKinney"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:6 [
                        "tituloSerie" => "New Microbiol"
                        "fecha" => "2012"
                        "volumen" => "35"
                        "paginaInicial" => "53"
                        "paginaFinal" => "59"
                        "link" => array:1 [
                          0 => array:2 [
                            "url" => "https://www.ncbi.nlm.nih.gov/pubmed/22378553"
                            "web" => "Medline"
                          ]
                        ]
                      ]
                    ]
                  ]
                ]
              ]
            ]
            5 => array:3 [
              "identificador" => "bib0085"
              "etiqueta" => "6"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Infection with <span class="elsevierStyleItalic">Salmonella enterica</span> serovar Typhimurium leads to increased proportions of F4&#47;80&#43; red pulp macrophages and decreased proportions of B and T lymphocytes in the spleen"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:5 [
                            0 => "L&#46; Rosche"
                            1 => "T&#46; Aljasham"
                            2 => "N&#46; Kipfer"
                            3 => "T&#46; Piatkowski"
                            4 => "V&#46; Konjufca"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:2 [
                      "doi" => "10.1371/journal.pone.0130092"
                      "Revista" => array:5 [
                        "tituloSerie" => "PLoS ONE"
                        "fecha" => "2015"
                        "volumen" => "10"
                        "paginaInicial" => "e0130092"
                        "link" => array:1 [
                          0 => array:2 [
                            "url" => "https://www.ncbi.nlm.nih.gov/pubmed/26068006"
                            "web" => "Medline"
                          ]
                        ]
                      ]
                    ]
                  ]
                ]
              ]
            ]
            6 => array:3 [
              "identificador" => "bib0090"
              "etiqueta" => "7"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Pathogenesis of <span class="elsevierStyleItalic">Salmonella</span><span class="elsevierStyleItalic">enterica</span> serovar Albany in experimental infected SPF BALB&#47;c Mice"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:5 [
                            0 => "J&#46; Sabri"
                            1 => "I&#46; Al-Sultan"
                            2 => "K&#46; Altaif"
                            3 => "S&#46; Peter"
                            4 => "M&#46; Saadh"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:5 [
                        "tituloSerie" => "Iraq J Vet Sci"
                        "fecha" => "2020"
                        "volumen" => "34"
                        "paginaInicial" => "339"
                        "paginaFinal" => "344"
                      ]
                    ]
                  ]
                ]
              ]
            ]
            7 => array:3 [
              "identificador" => "bib0095"
              "etiqueta" => "8"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Reaction of <span class="elsevierStyleItalic">Leishmania</span> &#40;<span class="elsevierStyleItalic">Leishmania</span>&#41; <span class="elsevierStyleItalic">mexicana</span> antigens by sera of patients with cutaneous leishmaniasis from Sinaloa&#44; Mexico"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:3 [
                            0 => "P&#46; Salazar-Mej&#237;a"
                            1 => "R&#46; Tejeda-Aguirre"
                            2 => "H&#46; L&#243;pez-Moreno"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:5 [
                        "tituloSerie" => "Salud Publica Mex"
                        "fecha" => "2010"
                        "volumen" => "52"
                        "paginaInicial" => "165"
                        "paginaFinal" => "169"
                      ]
                    ]
                  ]
                ]
              ]
            ]
            8 => array:3 [
              "identificador" => "bib0100"
              "etiqueta" => "9"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Excreci&#243;n fecal de <span class="elsevierStyleItalic">Salmonella</span> Albany&#44; su aislamiento en la raci&#243;n alimenticia y repercusi&#243;n en el estado de salud de un ocelote &#40;<span class="elsevierStyleItalic">Leopardus pardalis</span>&#41; en cautiverio"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:5 [
                            0 => "G&#46; Silva-Hidalgo"
                            1 => "H&#46;S&#46; L&#243;pez-Moreno"
                            2 => "V&#46;F&#46; Ortiz-Navarrete"
                            3 => "F&#46; Ju&#225;rez-Barranco"
                            4 => "M&#46; L&#243;pez-Valenzuela"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:5 [
                        "tituloSerie" => "Vet Mex"
                        "fecha" => "2012"
                        "volumen" => "43"
                        "paginaInicial" => "59"
                        "paginaFinal" => "69"
                      ]
                    ]
                  ]
                ]
              ]
            ]
            9 => array:3 [
              "identificador" => "bib0105"
              "etiqueta" => "10"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Non-typhi <span class="elsevierStyleItalic">Salmonella</span> serovars found in Mexican zoo animals"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:7 [
                            0 => "G&#46; Silva-Hidalgo"
                            1 => "V&#46; Ortiz-Navarrete"
                            2 => "C&#46; Alpuche-Aranda"
                            3 => "J&#46; Rend&#243;n-Maldonado"
                            4 => "M&#46; L&#243;pez-Valenzuela"
                            5 => "F&#46; Ju&#225;rez-Barranco"
                            6 => "H&#46;S&#46; L&#243;pez-Moreno"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:4 [
                        "tituloSerie" => "Res Vet Sci"
                        "fecha" => "2012"
                        "volumen" => "93"
                        "paginaInicial" => "11325"
                      ]
                    ]
                  ]
                ]
              ]
            ]
            10 => array:3 [
              "identificador" => "bib0110"
              "etiqueta" => "11"
              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "Peritonitis por <span class="elsevierStyleItalic">Salmonella</span> serotipo Albany en paciente sometido a di&#225;lisis peritoneal continua ambulatoria"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => false
                          "autores" => array:4 [
                            0 => "R&#46; Tato"
                            1 => "M&#46; Zamora"
                            2 => "M&#46; Pulian"
                            3 => "M&#46; Garcia"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:1 [
                    0 => array:1 [
                      "Revista" => array:6 [
                        "tituloSerie" => "Rev Esp Quimioter"
                        "fecha" => "2015"
                        "volumen" => "28"
                        "paginaInicial" => "162"
                        "paginaFinal" => "163"
                        "link" => array:1 [
                          0 => array:2 [
                            "url" => "https://www.ncbi.nlm.nih.gov/pubmed/26033003"
                            "web" => "Medline"
                          ]
                        ]
                      ]
                    ]
                  ]
                ]
              ]
            ]
          ]
        ]
      ]
    ]
    "agradecimientos" => array:1 [
      0 => array:4 [
        "identificador" => "xack641879"
        "titulo" => "Acknowledgements"
        "texto" => "<p id="par0080" class="elsevierStylePara elsevierViewall">The authors would like to thank fully to Dr&#46; Vianney Francisco Ortiz Navarrete for gently donative the <span class="elsevierStyleItalic">S&#46;</span> Typhimurium 14028 and plasmidic construction pST13-OVA&#46; Also&#44; they extended your acknowledgment to <span class="elsevierStyleGrantSponsor" id="gs1">PROFAPI-UAS</span> for financial support with the HSLM&#39;s Grant &#35;<span class="elsevierStyleGrantNumber" refid="gs1">2015&#47;151</span>&#46; And LBM team for technical assistance</p>"
        "vista" => "all"
      ]
    ]
  ]
  "idiomaDefecto" => "en"
  "url" => "/03257541/0000005400000004/v1_202212150521/S0325754122000529/v1_202212150521/en/main.assets"
  "Apartado" => array:4 [
    "identificador" => "37863"
    "tipo" => "SECCION"
    "en" => array:2 [
      "titulo" => "Microbiolog&#237;a b&#225;sica"
      "idiomaDefecto" => true
    ]
    "idiomaDefecto" => "en"
  ]
  "PDF" => "https://static.elsevier.es/multimedia/03257541/0000005400000004/v1_202212150521/S0325754122000529/v1_202212150521/en/main.pdf?idApp=UINPBA00004N&text.app=https://www.elsevier.es/"
  "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S0325754122000529?idApp=UINPBA00004N"
]

Article information

ISSN: 03257541
Original language: English

DOI:10.1016/j.ram.2022.05.008

The statistics are updated each day

Year/Month	Html	Pdf	Total

2024 November	6	1	7
2024 October	27	16	43
2024 September	53	5	58
2024 August	41	7	48
2024 July	26	7	33
2024 June	24	4	28
2024 May	22	8	30
2024 April	42	6	48
2024 March	20	8	28
2024 February	24	9	33
2024 January	17	2	19
2023 December	36	13	49
2023 November	28	4	32
2023 October	29	6	35
2023 September	18	1	19
2023 August	24	4	28
2023 July	14	4	18
2023 June	24	6	30
2023 May	54	10	64
2023 April	41	2	43
2023 March	37	14	51
2023 February	18	12	30
2023 January	11	6	17
2022 December	34	10	44
2022 November	0	4	4
2022 October	0	3	3
2022 September	0	11	11
2022 August	0	11	11
2022 July	0	8	8
2022 June	0	7	7

Show all

Indexed in:

Follow us:

Statistics

Indexed in:

Follow us:

Statistics

Subscribe to our newsletter