Staphylococcus aureus is a microorganism that can be found in the natural microbiota of humans and animals, mainly colonizing skin and nasopharynx.1 It has been considered as one of the most common opportunistic pathogens, able to cause infections of varying severity.2,3 In some situations, these infections are caused by S. aureus with methicillin-resistance determinants as mecA gene, which confers resistance to most β-lactams. Thus, some genetic lineages, as the clonal complexes CC5, CC22, CC30 or CC45, have been associated with hospital environments (HA-MRSA) with high capacity to cause nosocomial infections and to carry a large set of antibiotic-resistance genes.4 On the other hand, genetic lineages as CC8 or CC80 and CC30, among others, have been related to community environments (CA-MRSA), known for its high content of virulence factors but low levels of resistance.5 However, the differences between HA-MRSA and CA-MRSA isolates have become diffuse, and clonal lineages considered community associated are being related to hospital environments and vice versa.

Moreover, in the last years, other genetic lineages belonging to CC398, CC1, CC9, CC130 or CC133, known as livestock-associated MRSA (LA-MRSA), have emerged as an important zoonotic problem.6,7 They have been increasingly detected in animals or humans, either as colonizer or causing infections.6,8 In 2011, a new mecA homologue was described in the S. aureus isolate LGA251 from an epidemiological study of bovine mastitis in England.10 This new resistance determinant shared homology of 70% over the mecA and was named as mecALGA251,9 and renamed in 2012 as mecC10 and was integrated into the new chromosomal cassette SCCmec-XI.11

There is special interest to know the genetic lineages of S. aureus that are circulating in livestock environments in order to track its evolution, particularly those genetic lineages with high zoonotic potential, as CC398, CC130 or CC133. For this aim, the use of molecular techniques, such as spa-, agr-, or MLST typing, is very useful. A recent important tool for discriminating possible human or animal origin of isolates seems to be the determination of the human immune evasion cluster (IEC). The IEC constitutes a set of genes that enhances colonization by S. aureus allowing evasion of the first barriers of the human immune defenses, and increasing their invasive abilities.2,12

The objective of the present study was to perform a longitudinal study of S. aureus carriage in 11-member family of livestock farmers with different animal contact degree. The purpose was to carry out the molecular characterization of the recovered S. aureus isolates to determine the genetic lineages circulating and to study their resistance and virulence genes content as well as their IEC system.

The detection of a mecC positive MRSA strain from a skin lesion of one of the workers of the livestock farm is of great relevance, and the animal origin of this strain is highly suggested. The lack of the IEC cluster in this strain is of relevance, as it supports its potential animal origin. MRSA strains of the lineage CC130 have been detected in other studies in farm animals,17 mainly in cows and also in cow's milk.7 It is interesting to note that this farmer was working in the traditional production of cow cheese in a dairy farm close to the farm under study, where most animals were cattle. It is possible that the contact with the animals or with the milk could be in the origin of this strain. The characteristic profile of susceptibility for non-beta-lactams and the restricted virulence gene content to etd2 of our CC130 strain, agree with results of other mecC MRSA strains previously reported.18

The mecC-MRSA strain detected in our study was typed as t843-ST130-CC130, and it is one of the first cases described in farmers in Spain and the first one relating with cheese traditional making, with the potential risk that could entail its entry into the food chain. Some previous studies performed in Spain found the lineage t843/ST130 as the causative agent of fatal bacteremia in hospitalized patients or in a community patient.18–20 Other authors have identified similar genetic lineages (t1535/ST130) harboring mecC gene in wild small mammals14 or t843/ST2676 in urban wastewater,21 evidencing the spread of this lineage in different environments in Spain. Moreover, a case has recently been detected of S. aureus carrying mecC in livestock in Spain22 and in river water close to a livestock farm.23 Regarding the situation in other countries, MRSA strains typed as t843/ST130 have been described previously in humans in the European Union,10 but this lineage was also detected in different animal host species as cattle, cows, sheep or pets in UK, Denmark, Germany or France.10 Some authors suggested that MRSA mecC emerged in animals, mainly in ruminants, and subsequently made the leap to humans.9 In our case, the strain belonging to CC130 lacked the genes of the IEC system, and this fact points to an animal origin, probably being a potential case of zoonosis of LA-MRSA from cattle to human. In the same line, there are a few European studies with the objective of testing the possible zoonotic transmission between cattle and humans.24–26 In all of these studies, humans with different infections presented livestock animal contact or were linked to a farming activity. This fact supports the possible zoonotic transmission in our study.

Seven of the spa types identified in this study were ascribed to CC30, a lineage frequently found as colonizer of human microbiota,13,27 and it is also considered one of the most frequent S. aureus lineages detected in human infections.3 Strains of skin origin of lineage CC30, recovered from the individual of case 4, harbored the tsst-1 gene encoding the toxic shock syndrome toxin (TSST). It is interesting to remark that this person lived in the farm and often showed superficial and difficult healing wounds. The presence of relevant virulence factors (typical of community-associated S. aureus) and the low content in resis-tance determinants have been previously reported by other authors in MRSA-CC30 strains.28 Interestingly one MSSA strain typed as t11719-ST1333-CC30 showed a multidrug-resistance phenotype (penicillin-erythromycin-clindamycin-tetracycline-fusidic acid-mupirocin) with the detection of several resistance determinants (Table 2). This genetic lineage has been previously reported in canine samples from Japan29 and the absence of the genes of IEC system points to a possible animal origin of the strain.13

In the same line, detection of one MSSA strain typed as t2678-ST133-CC133 is remarkable because in previous studies it has been detected in small ruminants such as goats or sheep.30 The detection of gene lukM, prevalent in ungulates, ruminants and bovine mastitis,7 and the absence of IEC genes, supports the idea of the possible animal origin of the strain. Interestingly, some authors have suggested a possible ancestral human origin of the genetic lineage CC133 and have hypothesized about the possibility of jumping to ruminants few centuries ago.31

MSSA strains detected in five members of the family (belonging to animal-contact degree groups I and L) were assigned to hospital-associated clonal complexes CC22, CC45 and CC121.4 Therefore, some authors have postulated about the ability of MSSA to serve as recipient of the SCCmec cassette, subsequently losing any of its components, as the mecA gene.27 Moreover, the lineage t2155-ST121-CC121 found in one of our MSSA strains, has also been detected among MRSA implicated in skin and soft tissues infections.32 Interestingly, this strain was isolated from a skin lesion of an individual who worked sporadically in the farm (Case 6). In the same line, many studies have associated the MRSA-ST22 genetic lineage with frequent infections in Europe. This lineage in its methicillin-resistance form was called EMRSA-15.33 In our case methicillin-susceptible CC22 clade was detected.

As for the single case (case 2) where colonization was maintained by the same CC9 clone of S. aureus over the period studied, it is interesting to note that this genetic lineage was initially associated with farm animals, mainly pigs in Asia.34 Similarly CC9 has been detected in meat samples in Spain35 and in bloodstream infections in France,36 with similar patterns of resistance, particularly resistance to erythromycin, but the difference is of methicillin resistance in our case. This MSSA profile and the presence of IEC type-B, points to the possible human origin of this clone and reveals its great capacity for colonization in different environments. Furthermore, the presence of the gene encoding the exfoliatin A (eta) on the CC9 strains of this individual is noteworthy. Previous studies have suggested a high association between the CC9 and its ability to produce exfoliative toxins.27

We report the detection of the lineage ST130/CC130 containing the novel methicillin-resistance-determinant mecC as well as other livestock-associated S. aureus lineages (as CC133 or CC9) in nasal or skin samples of a family of livestock farmers in Spain. The mecC-CC130 strain was isolated from the superficial skin lesion of a farmer which had daily contact with farm animals, mainly cattle, and with traditional cheese production. Cows have been postulated as the main reservoir of this emerging genetic lineage. This fact highlights the zoonotic potential of this emergent resistant microorganism and other similar animal-related clades, with the risk that this entails for human health. More studies should be performed in the future to assess the real risk for colonization and infection of farmers in relation to MRSA-mecC clones and the different reservoirs that may exist in nature.

The authors declare no conflict of interest.