Antibacterial activity of Citrus hystrix toward Salmonella spp. infection | Enfermedades Infecciosas y Microbiología Clínica

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REIPI Pseudomonas study group" ] ] ] ] ] "idiomaDefecto" => "en" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S0213005X20302214?idApp=UINPBA00004N" "url" => "/0213005X/0000003900000006/v1_202106020853/S0213005X20302214/v1_202106020853/en/main.assets" ] "en" => array:21 [ "idiomaDefecto" => true "cabecera" => "Brief report" "titulo" => "Antibacterial activity of Citrus hystrix toward Salmonella spp. infection" "tieneTextoCompleto" => true "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "283" "paginaFinal" => "286" ] ] "autores" => array:1 [ 0 => array:4 [ "autoresLista" => "Zulvikar Syambani Ulhaq, Tenta Hartian Hendyatama, Faizanah Hameed, Dewi Santosaningsih" "autores" => array:4 [ 0 => array:4 [ "nombre" => "Zulvikar Syambani" "apellidos" => "Ulhaq" "email" => array:1 [ 0 => "zulhaq@kedokteran.uin-malang.ac.id" ] "referencia" => array:3 [ 0 => array:2 [ "etiqueta" => "a" "identificador" => "aff0005" ] 1 => array:2 [ "etiqueta" => "d" "identificador" => "aff0020" ] 2 => array:2 [ "etiqueta" => "*" "identificador" => "cor0005" ] ] ] 1 => array:3 [ "nombre" => "Tenta Hartian" "apellidos" => "Hendyatama" "referencia" => array:2 [ 0 => array:2 [ "etiqueta" => "b" "identificador" => "aff0010" ] 1 => array:2 [ "etiqueta" => "d" "identificador" => "aff0020" ] ] ] 2 => array:3 [ "nombre" => "Faizanah" "apellidos" => "Hameed" "referencia" => array:2 [ 0 => array:2 [ "etiqueta" => "c" "identificador" => "aff0015" ] 1 => array:2 [ "etiqueta" => "d" "identificador" => "aff0020" ] ] ] 3 => array:3 [ "nombre" => "Dewi" "apellidos" => "Santosaningsih" "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "d" "identificador" => "aff0020" ] ] ] ] "afiliaciones" => array:4 [ 0 => array:3 [ "entidad" => "Department of Biochemistry, Faculty of Medicine and Health Sciences, Maulana Malik Ibrahim Islamic State University of Malang, Batu, East Java, Indonesia" "etiqueta" => "a" "identificador" => "aff0005" ] 1 => array:3 [ "entidad" => "Department of Internal Medicine, Faculty of Medicine, Airlangga University, Surabaya, East Java, Indonesia" "etiqueta" => "b" "identificador" => "aff0010" ] 2 => array:3 [ "entidad" => "Batu Rakit Health Clinic, Trengganu, Malaysia" "etiqueta" => "c" "identificador" => "aff0015" ] 3 => array:3 [ "entidad" => "Department of Clinical Microbiology, Faculty of Medicine, Brawijaya University, Malang, East Java, Indonesia" "etiqueta" => "d" "identificador" => "aff0020" ] ] "correspondencia" => array:1 [ 0 => array:3 [ "identificador" => "cor0005" "etiqueta" => "⁎" "correspondencia" => "Corresponding author." ] ] ] ] "titulosAlternativos" => array:1 [ "es" => array:1 [ "titulo" => "Actividad antibacteriana de Citrus hystrix frente a la infección por Salmonella spp" ] ] "resumenGrafico" => array:2 [ "original" => 0 "multimedia" => array:7 [ "identificador" => "fig0010" "etiqueta" => "Fig. 2" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr2.jpeg" "Alto" => 1049 "Ancho" => 2508 "Tamanyo" => 217593 ] ] "descripcion" => array:1 [ "en" => "(A) Growth of S. typhimurium colonies on BSA medium. (B) Bacterial load in the ileum, liver, and spleen of S. typhimurium infected-model. Data are expressed as a mean±standard deviation (SD). (*) and (#) indicate a significant difference compared to day-1 post-infection and between two groups indicated in the graphs (p<0.05), respectively." ] ] ] "textoCompleto" => "IntroductionSalmonella spp. are gram negative bacteria that are associated with food and water-borne gastroenteritis and typhoid fever in humans.<a class="elsevierStyleCrossRef" href="#bib0080">1</a> Systemic infections caused by Salmonella enterica serotype Typhi (S. typhi) remain a public health concern in developing countries, including Indonesia due to a high incidence rate. A study at the slums of Jakarta province, estimated the incidence rate of typhoid at 148.7, 180.3, and 51.2 per 100,000 person-years in the age groups of 2–4, 5–15, and over 16 years of age, respectively.<a class="elsevierStyleCrossRef" href="#bib0085">2</a> Furthermore, a community-based case-control study conducted at the Jatinegara district, Jakarta, found that 9% and 3% of 1019 patients with fever, presented S. typhi and S. paratyphi infections, respectively.<a class="elsevierStyleCrossRef" href="#bib0090">3</a> A study reported that patients receiving appropriate therapy may reduce the case-fatality rate of typhoid fever to 1–4%.<a class="elsevierStyleCrossRef" href="#bib0095">4</a> Nonetheless, the proportion of multi-drug resistant (MDR) S. typhi tends to increase in several areas in Indonesia.<a class="elsevierStyleCrossRefs" href="#bib0085">2,5</a> Thus, MDR typhoid fever may become a serious problem and requires further appropriate treatment in the future.Citrus hytrix (Indonesian name Jeruk purut) is a member of the genus Citrus, family Rutaceae, and has been used as a key ingredient in Southeast Asian cuisines. In Indonesia, the use of Citrus is not limited to culinary purpose but also as a remedy for respiratory and gastrointestinal disorders, including Salmonella spp. infections, although the mechanism of action remains unclear. Interestingly, mice infected with S. typhimurium mimic typhoid like-disease.<a class="elsevierStyleCrossRef" href="#bib0080">1</a> Therefore, this study aims to investigate the antibacterial activity of the ethanolic peel extract of C. hystrixin vitro and in vivo in a mice model of Salmonella enterica serotype Typhimurium (S. typhimurium) infection.MethodsPlant material and extract preparationThe fruit peels of C. hystrix were collected from Malang, East Java, Indonesia. The species of Citrus was identified and confirmed by a plant taxonomist of the herbarium unit, UPT Materia Medica, Batu, East Java, Indonesia. Duplicate voucher specimens have been deposited in the herbarium unit, UPT Materia Medica, Batu, East Java, Indonesia. The extract was prepared as previously described.<a class="elsevierStyleCrossRef" href="#bib0105">6</a> Briefly, the fruit peels were shade dried and ground into a fine powder. Thirty grams of fruit peel powder were then macerated in 100ml absolute ethanol for 48h by the Soxhlet extraction apparatus. The extract was evaporated to dry at 40°C using a rotary evaporator and stored at 4°C for further use.Bacterial isolateS. typhimurium was provided by the Department of Clinical Microbiology, Faculty of Medicine, Brawijaya University and identification was confirmed by the Microbact™ Gram-negative system.Determination of the minimum inhibitory concentrationThe agar dilution method was used to determine the MIC of the ethanolic peel extract of C. hystrix, as described by Bailey and Scott's Diagnostic Microbiology.<a class="elsevierStyleCrossRef" href="#bib0110">7</a> Twofold serial dilutions of the extract (0.3125%, 0.625%, and 1.25%) were prepared in Mueller Hinton agar for the agar dilution assay. Ten microliters of bacterial inoculum were delivered onto the agar with the final inoculum of 106CFU/ml. Bacterial plates were incubated at 37°C and evaluated after 24h.Animal infection and tissue samplesFemale Balb/c mice between 8 and 12 weeks of age were obtained from the Faculty of Medicine, Gadjah Mada University and used in this study. This experiment was divided into two sequential approaches. First, to elucidate the initial stage of transient primary bacteremia in the animal model, thirty-five mice were infected orally with 0.5ml of 108CFU/ml S. typhimurium. Tissue collection procedures were initiated after the mice have been euthanized with isoflurane overdose (mice were placed into a chamber filled with a 5% vapor of the anesthetic isoflurane until respiration ceased, approximately within 2min). Five mice per day were sacrificed, bacterial counts in the ileum, liver, and spleen were evaluated from day 1 to day 7 post-infection (<a class="elsevierStyleCrossRef" href="#fig0005">Fig. 1</a>A). No colonization was observed in the uninfected control (data not shown). One hundred grams of the organs were homogenized and diluted in 2ml of nutrient broth. Ten-microliters of each samples were then cultured onto solid bismuth sulfite agar (BSA) and incubated at 37°C overnight. The number of S. typhimurium colonies were enumerated in CFU/g. For each sample, the quantification was done in triplicate. Second, to evaluate the antibacterial activity of the ethanolic peel extract of C. hystrixin vivo, twenty-five mice were randomly divided into five groups (5 mice each group), which were negative control, positive control, and infected mice treated with the extract orally for one day, two days and three days. The duration of S. typhimurium infection was seven days. Five mice of each infected group were sacrificed within 24h after the last administration of the extract, whereas negative and positive controls were sacrificed at the same time as corresponds to the treatment groups (<a class="elsevierStyleCrossRef" href="#fig0005">Fig. 1</a>B). The in vivo dose of the extract was quantified by converting % MIC into mg/ml, and then the result was multiplied by a total blood volume of mice.<a class="elsevierStyleCrossRef" href="#bib0115">8</a> The extract was diluted in distilled water and given orally for 0.5ml per mouse. The ileum, liver, and spleen were collected and cultured on BSA. Bacterial load was expressed as CFU/g. No colonization was observed in the negative control. The study was approved by the medical ethics committee of the Faculty of Medicine, Brawijaya University, Malang, Indonesia (Reference No. 25-KE).<elsevierMultimedia ident="fig0005"></elsevierMultimedia>Statistical analysisBacterial load was analyzed by one-way ANOVA, followed by Tukey's post hoc test using StatPlus. Significant differences were accepted when p<0.05.Results and discussionWe evaluated the growth of S. typhimurium isolated from the ileum, liver, and spleen on the BSA medium at day-1 to 7 post-infection (Supplementary Table 1). The number of bacterial colonies isolated from ileum was significantly increased at day-6 post-infection (p=0.02) (<a class="elsevierStyleCrossRef" href="#fig0010">Fig. 2</a>A) and nearly a 3-folde increased at day-7 post-infection (p=0.0001) (<a class="elsevierStyleCrossRef" href="#fig0010">Fig. 2</a>B). In this study, the colonization of S. typhimurium in the liver and spleen were found at day-7 post-infection, two more days than previously reported.<a class="elsevierStyleCrossRef" href="#bib0120">9</a> These differences may be due to individual variation of the immune response among mice model and the inoculum concentration of S. typhimurium. Together, these results suggest that bacteria might be released into the bloodstream between day-5 and 7 post-infection.<elsevierMultimedia ident="fig0010"></elsevierMultimedia>A previous report in our laboratory demonstrated that the MIC and minimum bactericidal concentration (MBC) of the ethanolic peel extract of C. hystrix against S. typhi were 0.625% and 1.25%, respectively.<a class="elsevierStyleCrossRef" href="#bib0125">10</a> In the same way, our study revealed that the MIC value of the ethanolic peel extract of C. hystrix against S. typhimurium was 0.625% (data not shown), which then was converted into 16mg (given orally in 0.5ml/day for 3 days) for in vivo experiments. The administration of the extract was given at day-7 post-infection. Our results showed that the administration of the ethanolic peel extract of C. hystrix inhibited significantly the colonization of S. typhimurium in the ileum, liver, and spleen after 24h (p=0.00008, p=0.00084, and p=0.00003, respectively) (Supplementary Table 2, <a class="elsevierStyleCrossRef" href="#fig0015">Fig. 3</a>A–D). Thus, our study suggests that daily oral administration of the ethanolic peel extract of C. hystrix at 16mg is effectively inhibiting the colonization of S. typhimuriumin vivo and might be a potential alternative treatment for Salmonella infections.<elsevierMultimedia ident="fig0015"></elsevierMultimedia>Several major constituents have been identified in C. hystrix peel extract.<a class="elsevierStyleCrossRefs" href="#bib0130">11,12</a> Interestingly, Srifuengfung et al. demonstrated that two active compounds, α-terpineol and terpinene-4-ol, were more concentrated than the other constituents.<a class="elsevierStyleCrossRef" href="#bib0135">12</a> Moreover, a previous report has shown that α-terpineol displays the most potent antibacterial activity in comparison with the other constituents.<a class="elsevierStyleCrossRef" href="#bib0140">13</a> Both Gram-positive and Gram-negative bacteria exposed to α-terpineol exhibited morphostructural alterations, mainly causing pores on the outer membrane of S. typhimurium.<a class="elsevierStyleCrossRef" href="#bib0145">14</a> Furthermore, α-terpineol is also able to induce lipid leakage of Gram-negative bacteria.<a class="elsevierStyleCrossRef" href="#bib0150">15</a> Therefore, it is possible to speculate that the active compounds of C. hystrix peels extract might disrupt the membrane integrity of bacteria, which then influence its metabolism. However, further experiments are required to examine the molecular mechanisms of the ethanolic peel extract of C. hystrix against bacterial infections, particularly against these caused by Salmonella spp.In conclusion, the ethanolic peel extract of C. hystrix exhibits antibacterial activity against S. typhimurium both in vitro and in vivo. These findings imply that peel extract of C. hystrix could be an alternative treatment for Salmonella infections. Further studies are required to define appropriate C. hystrix dosing regimens for Salmonella infections and to assess novel treatment strategies, including combination therapies.Funding statementNone to declare.Conflict of interestNone to declare." "textoCompletoSecciones" => array:1 [ "secciones" => array:11 [ 0 => array:3 [ "identificador" => "xres1518930" "titulo" => "Abstract" "secciones" => array:4 [ 0 => array:2 [ "identificador" => "abst0005" "titulo" => "Introduction" ] 1 => array:2 [ "identificador" => "abst0010" "titulo" => "Methods" ] 2 => array:2 [ "identificador" => "abst0015" "titulo" => "Results" ] 3 => array:2 [ "identificador" => "abst0020" "titulo" => "Conclusion" ] ] ] 1 => array:2 [ "identificador" => "xpalclavsec1377453" "titulo" => "Keywords" ] 2 => array:3 [ "identificador" => "xres1518929" "titulo" => "Resumen" "secciones" => array:4 [ 0 => array:2 [ "identificador" => "abst0025" "titulo" => "Introducción" ] 1 => array:2 [ "identificador" => "abst0030" "titulo" => "Métodos" ] 2 => array:2 [ "identificador" => "abst0035" "titulo" => "Resultados" ] 3 => array:2 [ "identificador" => "abst0040" "titulo" => "Conclusión" ] ] ] 3 => array:2 [ "identificador" => "xpalclavsec1377452" "titulo" => "Palabras clave" ] 4 => array:2 [ "identificador" => "sec0005" "titulo" => "Introduction" ] 5 => array:3 [ "identificador" => "sec0010" "titulo" => "Methods" "secciones" => array:5 [ 0 => array:2 [ "identificador" => "sec0015" "titulo" => "Plant material and extract preparation" ] 1 => array:2 [ "identificador" => "sec0020" "titulo" => "Bacterial isolate" ] 2 => array:2 [ "identificador" => "sec0025" "titulo" => "Determination of the minimum inhibitory concentration" ] 3 => array:2 [ "identificador" => "sec0030" "titulo" => "Animal infection and tissue samples" ] 4 => array:2 [ "identificador" => "sec0035" "titulo" => "Statistical analysis" ] ] ] 6 => array:2 [ "identificador" => "sec0040" "titulo" => "Results and discussion" ] 7 => array:2 [ "identificador" => "sec0045" "titulo" => "Funding statement" ] 8 => array:2 [ "identificador" => "sec0050" "titulo" => "Conflict of interest" ] 9 => array:2 [ "identificador" => "xack533585" "titulo" => "Acknowledgements" ] 10 => array:1 [ "titulo" => "References" ] ] ] "pdfFichero" => "main.pdf" "tienePdf" => true "fechaRecibido" => "2020-01-24" "fechaAceptado" => "2020-05-15" "PalabrasClave" => array:2 [ "en" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Keywords" "identificador" => "xpalclavsec1377453" "palabras" => array:4 [ 0 => "Antibacterial" 1 => "Citrus hytrix" 2 => "Salmonella enterica serotype typhimurium" 3 => "Salmonella typhimurium" ] ] ] "es" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Palabras clave" "identificador" => "xpalclavsec1377452" "palabras" => array:4 [ 0 => "Antibacteriano" 1 => "Citrus hystrix" 2 => "Salmonella enterica serotipo Typhimurium" 3 => "Salmonella typhimurium" ] ] ] ] "tieneResumen" => true "resumen" => array:2 [ "en" => array:3 [ "titulo" => "Abstract" "resumen" => "IntroductionCitrus hystrix is widely used by Indonesians as a traditional medicine for gastrointestinal diseases, including Salmonella spp. infection. We investigated the antibacterial activity of the ethanolic peel extract of C. hystrix against Salmonella typhimurium. MethodsThe antibacterial activity was evaluated both in vitro and in vivo. The minimum inhibitory concentration (MIC) of the extract was determined at a concentration of 0.625% by agar dilution assay. Later, the in vivo antibacterial activity was examined by the administration of 16mg of the extract daily for three consecutive days in a mouse model infected with S. typhimurium. ResultsThe bacterial loads of S. typhimurium in the ileum, liver, and spleen decreased after 24h of administration of the extract (p=0.00008, p=0.00084, and p=0.00003, respectively). ConclusionThe ethanolic peel extract of C. hystrix shows antibacterial activity against S. typhimurium, indicating the potential of C. hystrix as an effective treatment for Salmonella spp. infection." "secciones" => array:4 [ 0 => array:2 [ "identificador" => "abst0005" "titulo" => "Introduction" ] 1 => array:2 [ "identificador" => "abst0010" "titulo" => "Methods" ] 2 => array:2 [ "identificador" => "abst0015" "titulo" => "Results" ] 3 => array:2 [ "identificador" => "abst0020" "titulo" => "Conclusion" ] ] ] "es" => array:3 [ "titulo" => "Resumen" "resumen" => "IntroducciónEl Citrus hystrix es muy utilizado por los indonesios como medicina tradicional para las enfermedades gastrointestinales, incluida la infección por Salmonella spp. Investigamos la actividad antibacteriana del extracto etanólico de piel de C. hystrix frente a Salmonella typhimurium. MétodosLa actividad antibacteriana se evaluó tanto in vitro como in vivo. La concentración mínima inhibitoria (CMI) del extracto se determinó a una concentración del 0,625% mediante un ensayo de dilución en agar. Posteriormente, se evaluó la actividad antibacteriana in vivo mediante la administración de 16mg de extracto de forma diaria durante 3 días consecutivos en un modelo murino infectado con S. typhimurium. ResultadosLa carga bacteriana de S. typhimurium en el íleo, el hígado y el bazo se redujo 24h después de la administración del extracto (p=0,00008, p=0,00084 y p=0,00003, respectivamente). ConclusiónEl extracto etanólico de piel de C. hystrix muestra actividad antibacteriana frente a S. typhimurium, lo que indica el potencial de C. hystrix como tratamiento eficaz para la infección por Salmonella spp." "secciones" => array:4 [ 0 => array:2 [ "identificador" => "abst0025" "titulo" => "Introducción" ] 1 => array:2 [ "identificador" => "abst0030" "titulo" => "Métodos" ] 2 => array:2 [ "identificador" => "abst0035" "titulo" => "Resultados" ] 3 => array:2 [ "identificador" => "abst0040" "titulo" => "Conclusión" ] ] ] ] "apendice" => array:1 [ 0 => array:1 [ "seccion" => array:1 [ 0 => array:4 [ "apendice" => "The following are the supplementary data to this article:<elsevierMultimedia ident="upi0005"></elsevierMultimedia>" "etiqueta" => "Appendix A" "titulo" => "Supplementary data" "identificador" => "sec0060" ] ] ] ] "multimedia" => array:4 [ 0 => array:7 [ "identificador" => "fig0005" "etiqueta" => "Fig. 1" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr1.jpeg" "Alto" => 1186 "Ancho" => 1591 "Tamanyo" => 114041 ] ] "descripcion" => array:1 [ "en" => "Experimental timeline. (A) Evaluation of colony counts after mice were infected with S. typhimurium; (B) Evaluation of the effectivity of Citrus peel extract against Salmonella infection." ] ] 1 => array:7 [ "identificador" => "fig0010" "etiqueta" => "Fig. 2" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr2.jpeg" "Alto" => 1049 "Ancho" => 2508 "Tamanyo" => 217593 ] ] "descripcion" => array:1 [ "en" => "(A) Growth of S. typhimurium colonies on BSA medium. (B) Bacterial load in the ileum, liver, and spleen of S. typhimurium infected-model. Data are expressed as a mean±standard deviation (SD). 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Antibacterial activity of Citrus hystrix toward Salmonella spp. infection

Actividad antibacteriana de Citrus hystrix frente a la infección por Salmonella spp

Zulvikar Syambani Ulhaqa,d,

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zulhaq@kedokteran.uin-malang.ac.id

Corresponding author.

, Tenta Hartian Hendyatamab,d, Faizanah Hameedc,d, Dewi Santosaningsihd

a Department of Biochemistry, Faculty of Medicine and Health Sciences, Maulana Malik Ibrahim Islamic State University of Malang, Batu, East Java, Indonesia

b Department of Internal Medicine, Faculty of Medicine, Airlangga University, Surabaya, East Java, Indonesia

c Batu Rakit Health Clinic, Trengganu, Malaysia

d Department of Clinical Microbiology, Faculty of Medicine, Brawijaya University, Malang, East Java, Indonesia

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    "titulo" => "Antibacterial activity of <span class="elsevierStyleItalic">Citrus hystrix</span> toward <span class="elsevierStyleItalic">Salmonella</span> spp&#46; infection"
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          "en" => "<p id="spar0050" class="elsevierStyleSimplePara elsevierViewall">&#40;A&#41; Growth of <span class="elsevierStyleItalic">S&#46; typhimurium</span> colonies on BSA medium&#46; &#40;B&#41; Bacterial load in the ileum&#44; liver&#44; and spleen of <span class="elsevierStyleItalic">S&#46; typhimurium</span> infected-model&#46; Data are expressed as a mean<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>standard deviation &#40;SD&#41;&#46; &#40;&#42;&#41; and &#40;&#35;&#41; indicate a significant difference compared to day-1 post-infection and between two groups indicated in the graphs &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#60;<span class="elsevierStyleHsp" style=""></span>0&#46;05&#41;&#44; respectively&#46;</p>"
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    "textoCompleto" => "<span class="elsevierStyleSections"><span id="sec0005" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0065">Introduction</span><p id="par0005" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">Salmonella</span> spp&#46; are gram negative bacteria that are associated with food and water-borne gastroenteritis and typhoid fever in humans&#46;<a class="elsevierStyleCrossRef" href="#bib0080"><span class="elsevierStyleSup">1</span></a> Systemic infections caused by <span class="elsevierStyleItalic">Salmonella enterica</span> serotype Typhi &#40;<span class="elsevierStyleItalic">S&#46; typhi</span>&#41; remain a public health concern in developing countries&#44; including Indonesia due to a high incidence rate&#46; A study at the slums of Jakarta province&#44; estimated the incidence rate of typhoid at 148&#46;7&#44; 180&#46;3&#44; and 51&#46;2 per 100&#44;000 person-years in the age groups of 2&#8211;4&#44; 5&#8211;15&#44; and over 16 years of age&#44; respectively&#46;<a class="elsevierStyleCrossRef" href="#bib0085"><span class="elsevierStyleSup">2</span></a> Furthermore&#44; a community-based case-control study conducted at the Jatinegara district&#44; Jakarta&#44; found that 9&#37; and 3&#37; of 1019 patients with fever&#44; presented <span class="elsevierStyleItalic">S&#46; typhi</span> and <span class="elsevierStyleItalic">S&#46; paratyphi</span> infections&#44; respectively&#46;<a class="elsevierStyleCrossRef" href="#bib0090"><span class="elsevierStyleSup">3</span></a> A study reported that patients receiving appropriate therapy may reduce the case-fatality rate of typhoid fever to 1&#8211;4&#37;&#46;<a class="elsevierStyleCrossRef" href="#bib0095"><span class="elsevierStyleSup">4</span></a> Nonetheless&#44; the proportion of multi-drug resistant &#40;MDR&#41; <span class="elsevierStyleItalic">S&#46; typhi</span> tends to increase in several areas in Indonesia&#46;<a class="elsevierStyleCrossRefs" href="#bib0085"><span class="elsevierStyleSup">2&#44;5</span></a> Thus&#44; MDR typhoid fever may become a serious problem and requires further appropriate treatment in the future&#46;</p><p id="par0010" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">Citrus hytrix</span> &#40;Indonesian name Jeruk purut&#41; is a member of the genus <span class="elsevierStyleItalic">Citrus</span>&#44; family <span class="elsevierStyleItalic">Rutaceae</span>&#44; and has been used as a key ingredient in Southeast Asian cuisines&#46; In Indonesia&#44; the use of <span class="elsevierStyleItalic">Citrus</span> is not limited to culinary purpose but also as a remedy for respiratory and gastrointestinal disorders&#44; including <span class="elsevierStyleItalic">Salmonella</span> spp&#46; infections&#44; although the mechanism of action remains unclear&#46; Interestingly&#44; mice infected with <span class="elsevierStyleItalic">S&#46; typhimurium</span> mimic typhoid like-disease&#46;<a class="elsevierStyleCrossRef" href="#bib0080"><span class="elsevierStyleSup">1</span></a> Therefore&#44; this study aims to investigate the antibacterial activity of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span><span class="elsevierStyleItalic">in vitro</span> and <span class="elsevierStyleItalic">in vivo</span> in a mice model of <span class="elsevierStyleItalic">Salmonella enterica</span> serotype Typhimurium <span class="elsevierStyleItalic">&#40;S&#46; typhimurium&#41;</span> infection&#46;</p></span><span id="sec0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0070">Methods</span><span id="sec0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0075">Plant material and extract preparation</span><p id="par0015" class="elsevierStylePara elsevierViewall">The fruit peels of <span class="elsevierStyleItalic">C&#46; hystrix</span> were collected from Malang&#44; East Java&#44; Indonesia&#46; The species of <span class="elsevierStyleItalic">Citrus</span> was identified and confirmed by a plant taxonomist of the herbarium unit&#44; UPT Materia Medica&#44; Batu&#44; East Java&#44; Indonesia&#46; Duplicate voucher specimens have been deposited in the herbarium unit&#44; UPT Materia Medica&#44; Batu&#44; East Java&#44; Indonesia&#46; The extract was prepared as previously described&#46;<a class="elsevierStyleCrossRef" href="#bib0105"><span class="elsevierStyleSup">6</span></a> Briefly&#44; the fruit peels were shade dried and ground into a fine powder&#46; Thirty grams of fruit peel powder were then macerated in 100<span class="elsevierStyleHsp" style=""></span>ml absolute ethanol for 48<span class="elsevierStyleHsp" style=""></span>h by the Soxhlet extraction apparatus&#46; The extract was evaporated to dry at 40<span class="elsevierStyleHsp" style=""></span>&#176;C using a rotary evaporator and stored at 4<span class="elsevierStyleHsp" style=""></span>&#176;C for further use&#46;</p></span><span id="sec0020" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0080">Bacterial isolate</span><p id="par0020" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">S&#46; typhimurium</span> was provided by the Department of Clinical Microbiology&#44; Faculty of Medicine&#44; Brawijaya University and identification was confirmed by the Microbact&#8482; Gram-negative system&#46;</p></span><span id="sec0025" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0085">Determination of the minimum inhibitory concentration</span><p id="par0025" class="elsevierStylePara elsevierViewall">The agar dilution method was used to determine the MIC of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix&#44;</span> as described by Bailey and Scott&#39;s Diagnostic Microbiology&#46;<a class="elsevierStyleCrossRef" href="#bib0110"><span class="elsevierStyleSup">7</span></a> Twofold serial dilutions of the extract &#40;0&#46;3125&#37;&#44; 0&#46;625&#37;&#44; and 1&#46;25&#37;&#41; were prepared in Mueller Hinton agar for the agar dilution assay&#46; Ten microliters of bacterial inoculum were delivered onto the agar with the final inoculum of 10<span class="elsevierStyleSup">6</span><span class="elsevierStyleHsp" style=""></span>CFU&#47;ml&#46; Bacterial plates were incubated at 37<span class="elsevierStyleHsp" style=""></span>&#176;C and evaluated after 24<span class="elsevierStyleHsp" style=""></span>h&#46;</p></span><span id="sec0030" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0090">Animal infection and tissue samples</span><p id="par0030" class="elsevierStylePara elsevierViewall">Female Balb&#47;c mice between 8 and 12 weeks of age were obtained from the Faculty of Medicine&#44; Gadjah Mada University and used in this study&#46; This experiment was divided into two sequential approaches&#46; First&#44; to elucidate the initial stage of transient primary bacteremia in the animal model&#44; thirty-five mice were infected orally with 0&#46;5<span class="elsevierStyleHsp" style=""></span>ml of 10<span class="elsevierStyleSup">8</span><span class="elsevierStyleHsp" style=""></span>CFU&#47;ml <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#46; Tissue collection procedures were initiated after the mice have been euthanized with isoflurane overdose &#40;mice were placed into a chamber filled with a 5&#37; vapor of the anesthetic isoflurane until respiration ceased&#44; approximately within 2<span class="elsevierStyleHsp" style=""></span>min&#41;&#46; Five mice per day were sacrificed&#44; bacterial counts in the ileum&#44; liver&#44; and spleen were evaluated from day 1 to day 7 post-infection &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46; 1</a>A&#41;&#46; No colonization was observed in the uninfected control &#40;data not shown&#41;&#46; One hundred grams of the organs were homogenized and diluted in 2<span class="elsevierStyleHsp" style=""></span>ml of nutrient broth&#46; Ten-microliters of each samples were then cultured onto solid bismuth sulfite agar &#40;BSA&#41; and incubated at 37<span class="elsevierStyleHsp" style=""></span>&#176;C overnight&#46; The number of <span class="elsevierStyleItalic">S&#46; typhimurium</span> colonies were enumerated in CFU&#47;g&#46; For each sample&#44; the quantification was done in triplicate&#46; Second&#44; to evaluate the antibacterial activity of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span><span class="elsevierStyleItalic">in vivo</span>&#44; twenty-five mice were randomly divided into five groups &#40;5 mice each group&#41;&#44; which were negative control&#44; positive control&#44; and infected mice treated with the extract orally for one day&#44; two days and three days&#46; The duration of <span class="elsevierStyleItalic">S&#46; typhimurium</span> infection was seven days&#46; Five mice of each infected group were sacrificed within 24<span class="elsevierStyleHsp" style=""></span>h after the last administration of the extract&#44; whereas negative and positive controls were sacrificed at the same time as corresponds to the treatment groups &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46; 1</a>B&#41;&#46; The <span class="elsevierStyleItalic">in vivo</span> dose of the extract was quantified by converting &#37; MIC into mg&#47;ml&#44; and then the result was multiplied by a total blood volume of mice&#46;<a class="elsevierStyleCrossRef" href="#bib0115"><span class="elsevierStyleSup">8</span></a> The extract was diluted in distilled water and given orally for 0&#46;5<span class="elsevierStyleHsp" style=""></span>ml per mouse&#46; The ileum&#44; liver&#44; and spleen were collected and cultured on BSA&#46; Bacterial load was expressed as CFU&#47;g&#46; No colonization was observed in the negative control&#46; The study was approved by the medical ethics committee of the Faculty of Medicine&#44; Brawijaya University&#44; Malang&#44; Indonesia &#40;Reference No&#46; 25-KE&#41;&#46;</p><elsevierMultimedia ident="fig0005"></elsevierMultimedia></span><span id="sec0035" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0095">Statistical analysis</span><p id="par0035" class="elsevierStylePara elsevierViewall">Bacterial load was analyzed by one-way ANOVA&#44; followed by Tukey&#39;s <span class="elsevierStyleItalic">post hoc</span> test using StatPlus&#46; Significant differences were accepted when <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#60;<span class="elsevierStyleHsp" style=""></span>0&#46;05&#46;</p></span></span><span id="sec0040" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0100">Results and discussion</span><p id="par0040" class="elsevierStylePara elsevierViewall">We evaluated the growth of <span class="elsevierStyleItalic">S&#46; typhimurium</span> isolated from the ileum&#44; liver&#44; and spleen on the BSA medium at day-1 to 7 post-infection &#40;Supplementary Table 1&#41;&#46; The number of bacterial colonies isolated from ileum was significantly increased at day-6 post-infection &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;02&#41; &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>A&#41; and nearly a 3-folde increased at day-7 post-infection &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;0001&#41; &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>B&#41;&#46; In this study&#44; the colonization of <span class="elsevierStyleItalic">S&#46; typhimurium</span> in the liver and spleen were found at day-7 post-infection&#44; two more days than previously reported&#46;<a class="elsevierStyleCrossRef" href="#bib0120"><span class="elsevierStyleSup">9</span></a> These differences may be due to individual variation of the immune response among mice model and the inoculum concentration of <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#46; Together&#44; these results suggest that bacteria might be released into the bloodstream between day-5 and 7 post-infection&#46;</p><elsevierMultimedia ident="fig0010"></elsevierMultimedia><p id="par0045" class="elsevierStylePara elsevierViewall">A previous report in our laboratory demonstrated that the MIC and minimum bactericidal concentration &#40;MBC&#41; of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> against <span class="elsevierStyleItalic">S&#46; typhi</span> were 0&#46;625&#37; and 1&#46;25&#37;&#44; respectively&#46;<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">10</span></a> In the same way&#44; our study revealed that the MIC value of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> against <span class="elsevierStyleItalic">S</span>&#46; <span class="elsevierStyleItalic">typhimurium</span> was 0&#46;625&#37; &#40;data not shown&#41;&#44; which then was converted into 16<span class="elsevierStyleHsp" style=""></span>mg &#40;given orally in 0&#46;5<span class="elsevierStyleHsp" style=""></span>ml&#47;day for 3 days&#41; for <span class="elsevierStyleItalic">in vivo</span> experiments&#46; The administration of the extract was given at day-7 post-infection&#46; Our results showed that the administration of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> inhibited significantly the colonization of <span class="elsevierStyleItalic">S</span>&#46; <span class="elsevierStyleItalic">typhimurium</span> in the ileum&#44; liver&#44; and spleen after 24<span class="elsevierStyleHsp" style=""></span>h &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;00008&#44; <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;00084&#44; and <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;00003&#44; respectively&#41; &#40;Supplementary Table 2&#44; <a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a>A&#8211;D&#41;&#46; Thus&#44; our study suggests that daily oral administration of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> at 16<span class="elsevierStyleHsp" style=""></span>mg is effectively inhibiting the colonization of <span class="elsevierStyleItalic">S</span>&#46; <span class="elsevierStyleItalic">typhimurium</span><span class="elsevierStyleItalic">in vivo</span> and might be a potential alternative treatment for <span class="elsevierStyleItalic">Salmonella</span> infections&#46;</p><elsevierMultimedia ident="fig0015"></elsevierMultimedia><p id="par0050" class="elsevierStylePara elsevierViewall">Several major constituents have been identified in <span class="elsevierStyleItalic">C&#46; hystrix</span> peel extract&#46;<a class="elsevierStyleCrossRefs" href="#bib0130"><span class="elsevierStyleSup">11&#44;12</span></a> Interestingly&#44; Srifuengfung et al&#46; demonstrated that two active compounds&#44; &#945;-terpineol and terpinene-4-ol&#44; were more concentrated than the other constituents&#46;<a class="elsevierStyleCrossRef" href="#bib0135"><span class="elsevierStyleSup">12</span></a> Moreover&#44; a previous report has shown that &#945;-terpineol displays the most potent antibacterial activity in comparison with the other constituents&#46;<a class="elsevierStyleCrossRef" href="#bib0140"><span class="elsevierStyleSup">13</span></a> Both Gram-positive and Gram-negative bacteria exposed to &#945;-terpineol exhibited morphostructural alterations&#44; mainly causing pores on the outer membrane of <span class="elsevierStyleItalic">S</span>&#46; <span class="elsevierStyleItalic">typhimurium</span>&#46;<a class="elsevierStyleCrossRef" href="#bib0145"><span class="elsevierStyleSup">14</span></a> Furthermore&#44; &#945;-terpineol is also able to induce lipid leakage of Gram-negative bacteria&#46;<a class="elsevierStyleCrossRef" href="#bib0150"><span class="elsevierStyleSup">15</span></a> Therefore&#44; it is possible to speculate that the active compounds of <span class="elsevierStyleItalic">C&#46; hystrix</span> peels extract might disrupt the membrane integrity of bacteria&#44; which then influence its metabolism&#46; However&#44; further experiments are required to examine the molecular mechanisms of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> against bacterial infections&#44; particularly against these caused by <span class="elsevierStyleItalic">Salmonella</span> spp&#46;</p><p id="par0055" class="elsevierStylePara elsevierViewall">In conclusion&#44; the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> exhibits antibacterial activity against <span class="elsevierStyleItalic">S&#46; typhimurium</span> both <span class="elsevierStyleItalic">in vitro</span> and <span class="elsevierStyleItalic">in vivo</span>&#46; These findings imply that peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> could be an alternative treatment for <span class="elsevierStyleItalic">Salmonella</span> infections&#46; Further studies are required to define appropriate <span class="elsevierStyleItalic">C&#46; hystrix</span> dosing regimens for <span class="elsevierStyleItalic">Salmonella</span> infections and to assess novel treatment strategies&#44; including combination therapies&#46;</p></span><span id="sec0045" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0105">Funding statement</span><p id="par0060" class="elsevierStylePara elsevierViewall">None to declare&#46;</p></span><span id="sec0050" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0110">Conflict of interest</span><p id="par0065" class="elsevierStylePara elsevierViewall">None to declare&#46;</p></span></span>"
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          "secciones" => array:5 [
            0 => array:2 [
              "identificador" => "sec0015"
              "titulo" => "Plant material and extract preparation"
            ]
            1 => array:2 [
              "identificador" => "sec0020"
              "titulo" => "Bacterial isolate"
            ]
            2 => array:2 [
              "identificador" => "sec0025"
              "titulo" => "Determination of the minimum inhibitory concentration"
            ]
            3 => array:2 [
              "identificador" => "sec0030"
              "titulo" => "Animal infection and tissue samples"
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            4 => array:2 [
              "identificador" => "sec0035"
              "titulo" => "Statistical analysis"
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          "titulo" => "Results and discussion"
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          "identificador" => "sec0050"
          "titulo" => "Conflict of interest"
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          "identificador" => "xack533585"
          "titulo" => "Acknowledgements"
        ]
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          "titulo" => "References"
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      ]
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    "fechaRecibido" => "2020-01-24"
    "fechaAceptado" => "2020-05-15"
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        0 => array:4 [
          "clase" => "keyword"
          "titulo" => "Keywords"
          "identificador" => "xpalclavsec1377453"
          "palabras" => array:4 [
            0 => "Antibacterial"
            1 => "Citrus hytrix"
            2 => "<span class="elsevierStyleItalic">Salmonella enterica</span> serotype typhimurium"
            3 => "<span class="elsevierStyleItalic">Salmonella typhimurium</span>"
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          "palabras" => array:4 [
            0 => "Antibacteriano"
            1 => "<span class="elsevierStyleItalic">Citrus hystrix</span>"
            2 => "<span class="elsevierStyleItalic">Salmonella enterica</span> serotipo <span class="elsevierStyleItalic">Typhimurium</span>"
            3 => "<span class="elsevierStyleItalic">Salmonella typhimurium</span>"
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      "en" => array:3 [
        "titulo" => "Abstract"
        "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0010">Introduction</span><p id="spar0005" class="elsevierStyleSimplePara elsevierViewall"><span class="elsevierStyleItalic">Citrus hystrix</span> is widely used by Indonesians as a traditional medicine for gastrointestinal diseases&#44; including <span class="elsevierStyleItalic">Salmonella</span> spp&#46; infection&#46; We investigated the antibacterial activity of the ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> against <span class="elsevierStyleItalic">Salmonella typhimurium&#46;</span></p></span> <span id="abst0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0015">Methods</span><p id="spar0010" class="elsevierStyleSimplePara elsevierViewall">The antibacterial activity was evaluated both <span class="elsevierStyleItalic">in vitro</span> and <span class="elsevierStyleItalic">in vivo</span>&#46; The minimum inhibitory concentration &#40;MIC&#41; of the extract was determined at a concentration of 0&#46;625&#37; by agar dilution assay&#46; Later&#44; the <span class="elsevierStyleItalic">in vivo</span> antibacterial activity was examined by the administration of 16<span class="elsevierStyleHsp" style=""></span>mg of the extract daily for three consecutive days in a mouse model infected with <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#46;</p></span> <span id="abst0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0020">Results</span><p id="spar0015" class="elsevierStyleSimplePara elsevierViewall">The bacterial loads of <span class="elsevierStyleItalic">S&#46; typhimurium</span> in the ileum&#44; liver&#44; and spleen decreased after 24<span class="elsevierStyleHsp" style=""></span>h of administration of the extract &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;00008&#44; <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;00084&#44; and <span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;00003&#44; respectively&#41;&#46;</p></span> <span id="abst0020" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0025">Conclusion</span><p id="spar0020" class="elsevierStyleSimplePara elsevierViewall">The ethanolic peel extract of <span class="elsevierStyleItalic">C&#46; hystrix</span> shows antibacterial activity against <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#44; indicating the potential of <span class="elsevierStyleItalic">C&#46; hystrix</span> as an effective treatment for <span class="elsevierStyleItalic">Salmonella</span> spp&#46; infection&#46;</p></span>"
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          3 => array:2 [
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        "titulo" => "Resumen"
        "resumen" => "<span id="abst0025" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0035">Introducci&#243;n</span><p id="spar0025" class="elsevierStyleSimplePara elsevierViewall">El <span class="elsevierStyleItalic">Citrus hystrix</span> es muy utilizado por los indonesios como medicina tradicional para las enfermedades gastrointestinales&#44; incluida la infecci&#243;n por <span class="elsevierStyleItalic">Salmonella</span> spp&#46; Investigamos la actividad antibacteriana del extracto etan&#243;lico de piel de <span class="elsevierStyleItalic">C&#46; hystrix</span> frente a <span class="elsevierStyleItalic">Salmonella typhimurium&#46;</span></p></span> <span id="abst0030" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0040">M&#233;todos</span><p id="spar0030" class="elsevierStyleSimplePara elsevierViewall">La actividad antibacteriana se evalu&#243; tanto <span class="elsevierStyleItalic">in vitro</span> como <span class="elsevierStyleItalic">in vivo</span>&#46; La concentraci&#243;n m&#237;nima inhibitoria &#40;CMI&#41; del extracto se determin&#243; a una concentraci&#243;n del 0&#44;625&#37; mediante un ensayo de diluci&#243;n en agar&#46; Posteriormente&#44; se evalu&#243; la actividad antibacteriana <span class="elsevierStyleItalic">in vivo</span> mediante la administraci&#243;n de 16<span class="elsevierStyleHsp" style=""></span>mg de extracto de forma diaria durante 3 d&#237;as consecutivos en un modelo murino infectado con <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#46;</p></span> <span id="abst0035" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0045">Resultados</span><p id="spar0035" class="elsevierStyleSimplePara elsevierViewall">La carga bacteriana de <span class="elsevierStyleItalic">S&#46; typhimurium</span> en el &#237;leo&#44; el h&#237;gado y el bazo se redujo 24<span class="elsevierStyleHsp" style=""></span>h despu&#233;s de la administraci&#243;n del extracto &#40;p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;00008&#44; p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;00084 y p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;00003&#44; respectivamente&#41;&#46;</p></span> <span id="abst0040" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0050">Conclusi&#243;n</span><p id="spar0040" class="elsevierStyleSimplePara elsevierViewall">El extracto etan&#243;lico de piel de <span class="elsevierStyleItalic">C&#46; hystrix</span> muestra actividad antibacteriana frente a <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#44; lo que indica el potencial de <span class="elsevierStyleItalic">C&#46; hystrix</span> como tratamiento eficaz para la infecci&#243;n por <span class="elsevierStyleItalic">Salmonella</span> spp&#46;</p></span>"
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          "en" => "<p id="spar0045" class="elsevierStyleSimplePara elsevierViewall">Experimental timeline&#46; &#40;A&#41; Evaluation of colony counts after mice were infected with <span class="elsevierStyleItalic">S&#46; typhimurium</span>&#59; &#40;B&#41; Evaluation of the effectivity of <span class="elsevierStyleItalic">Citrus</span> peel extract against <span class="elsevierStyleItalic">Salmonella</span> infection&#46;</p>"
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          "en" => "<p id="spar0050" class="elsevierStyleSimplePara elsevierViewall">&#40;A&#41; Growth of <span class="elsevierStyleItalic">S&#46; typhimurium</span> colonies on BSA medium&#46; &#40;B&#41; Bacterial load in the ileum&#44; liver&#44; and spleen of <span class="elsevierStyleItalic">S&#46; typhimurium</span> infected-model&#46; Data are expressed as a mean<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>standard deviation &#40;SD&#41;&#46; &#40;&#42;&#41; and &#40;&#35;&#41; indicate a significant difference compared to day-1 post-infection and between two groups indicated in the graphs &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#60;<span class="elsevierStyleHsp" style=""></span>0&#46;05&#41;&#44; respectively&#46;</p>"
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          "en" => "<p id="spar0055" class="elsevierStyleSimplePara elsevierViewall">Bacterial load after the oral administration of <span class="elsevierStyleItalic">Citrus</span> peel extract&#46; The colonies were evaluated every 24<span class="elsevierStyleHsp" style=""></span>h for three days&#46; &#40;A&#41; Growth of <span class="elsevierStyleItalic">S&#46; typhimurium</span> colonies on BSA medium&#59; &#40;B&#8211;D&#41; Bacterial load in the ileum&#44; liver&#44; and spleen of <span class="elsevierStyleItalic">S&#46; typhimurium</span> infected-model&#44; respectively&#46; Data are expressed as a mean<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>standard deviation &#40;SD&#41;&#46; &#40;&#42;&#41; and &#40;&#35;&#41; indicates a significant difference compared to control and between two groups indicated in the graphs &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#60;<span class="elsevierStyleHsp" style=""></span>0&#46;05&#41;&#44; respectively&#46;</p>"
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                            0 => "I&#46;N&#46;P&#46; Ajithkumar"
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Article information

ISSN: 0213005X
Original language: English

DOI:10.1016/j.eimc.2020.05.016

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Year/Month	Html	Pdf	Total

2024 November	3	2	5
2024 October	29	13	42
2024 September	54	23	77
2024 August	43	17	60
2024 July	34	10	44
2024 June	41	8	49
2024 May	31	17	48
2024 April	35	45	80
2024 March	54	17	71
2024 February	65	15	80
2024 January	70	25	95
2023 December	51	109	160
2023 November	66	22	88
2023 October	79	54	133
2023 September	57	5	62
2023 August	29	3	32
2023 July	33	45	78
2023 June	32	8	40
2023 May	92	3	95
2023 April	57	1	58
2023 March	57	5	62
2023 February	38	12	50
2023 January	39	10	49
2022 December	43	8	51
2022 November	72	27	99
2022 October	82	18	100
2022 September	100	43	143
2022 August	50	31	81
2022 July	23	13	36
2022 June	19	21	40
2022 May	28	20	48
2022 April	27	30	57
2022 March	19	11	30
2022 February	30	18	48
2022 January	35	12	47
2021 December	35	12	47
2021 October	2	2	4
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