Transmission dynamics of methicillin-resistant Staphylococcus aureus in pigs

From the mid-2000s on, numerous studies have shown that methicillin-resistant Staphylococcus aureus (MRSA), renowned as human pathogen, has a reservoir in pigs and other livestock. In Europe and North America, clonal complex (CC) 398 appears to be the predominant lineage involved. Especially worrisome is its capacity to contaminate humans in close contact with affected animals. Indeed, the typical multi-resistant phenotype of MRSA CC398 and its observed ability of easily acquiring genetic material suggests that MRSA CC398 strains with an increased virulence potential may emerge, for which few therapeutic options would remain. This questions the need to implement interventions to control the presence and spread of MRSA CC398 among pigs. MRSA CC398 shows a high but not fully understood transmission potential in the pig population and is able to persist within that population. Although direct contact is probably the main route for MRSA transmission between pigs, also environmental contamination, the presence of other livestock, the herd size, and farm management are factors that may be involved in the dissemination of MRSA CC398. The current review aims at summarizing the research that has so far been done on the transmission dynamics and risk factors for introduction and persistence of MRSA CC398 in farms

Serological profiles in nursery piglets colonized with Staphylococcus aureus

At present, the immune response of pigs in relation to Staphylococcus aureus carriage is poorly understood. This study aimed at investigating the dynamics of the anti-staphylococcal humoral immune response in methicillin-susceptible S. aureus (MSSA)-positive piglets and at assessing the effect of the experimental introduction of a methicillin-resistant S. aureus (MRSA) Sequence Type (ST) 398 strain. Therefore, serum samples were collected at different times from 31 weaned piglets originating from four different sows. Twenty-four out of the 31 piglets were challenged with MRSA ST398. The serum samples were analysed for IgG antibodies to 39 S. aureus antigens, using a multiplex bead-based assay (xMAP technology, Luminex Corporation). Though antibody responses showed broad inter-individual variability, serological results appeared to be clustered by litter of origin. For most antigens, an age-related response was observed with an apparent increase in antibody titres directed against staphylococcal microbial surface components recognizing adhesive matrix molecules (MSCRAMMs), which have been shown to play a role in S. aureus colonization. In most animals, antibody titres directed against staphylococcal toxins or immune-modulating proteins decreased with age, possibly reflecting absence of bacterial invasion. The introduction of MRSA ST398 did not elicit a significant humoral immune reaction. This study describes, for the first time, the humoral immune response in weaned pigs colonized with S. aureus

The benefits of whole genome sequencing for foodborne outbreak investigation from the perspective of a national reference laboratory in a smaller country

Gradually, conventional methods for foodborne pathogen typing are replaced by whole genome sequencing (WGS). Despite studies describing the overall benefits, National Reference Laboratories of smaller countries often show slower uptake of WGS, mainly because of significant investments required to generate and analyze data of a limited amount of samples. To facilitate this process and incite policy makers to support its implementation, a Shiga toxin-producing Escherichia coli (STEC) O157:H7 (stx1+, stx2+, eae+) outbreak (2012) and a STEC O157:H7 (stx2+, eae+) outbreak (2013) were retrospectively analyzed using WGS and compared with their conventional investigations. The corresponding results were obtained, with WGS delivering even more information, e.g., on virulence and antimicrobial resistance genotypes. Besides a universal, all-in-one workflow with less hands-on-time (five versus seven actual working days for WGS versus conventional), WGS-based cgMLST-typing demonstrated increased resolution. This enabled an accurate cluster definition, which remained unsolved for the 2013 outbreak, partly due to scarce epidemiological linking with the suspect source. Moreover, it allowed detecting two and one earlier circulating STEC O157:H7 (stx1+, stx2+, eae+) and STEC O157:H7 (stx2+, eae+) strains as closely related to the 2012 and 2013 outbreaks, respectively, which might have further directed epidemiological investigation initially. Although some bottlenecks concerning centralized data-sharing, sampling strategies, and perceived costs should be considered, we delivered a proof-of-concept that even in smaller countries, WGS offers benefits for outbreak investigation, if a sufficient budget is available to ensure its implementation in surveillance. Indeed, applying a database with background isolates is critical in interpreting isolate relationships to outbreaks, and leveraging the true benefit of WGS in outbreak investigation and/or prevention

Validation strategy of a bioinformatics whole genome sequencing workflow for Shiga toxin-producing Escherichia coli using a reference collection extensively characterized with conventional methods

Whole genome sequencing (WGS) enables complete characterization of bacterial pathogenic isolates at single nucleotide resolution, making it the ultimate tool for routine surveillance and outbreak investigation. The lack of standardization, and the variation regarding bioinformatics workflows and parameters, however, complicates interoperability among (inter)national laboratories. We present a validation strategy applied to a bioinformatics workflow for Illumina data that performs complete characterization of Shiga toxin-producing Escherichia coli (STEC) isolates including antimicrobial resistance prediction, virulence gene detection, serotype prediction, plasmid replicon detection and sequence typing. The workflow supports three commonly used bioinformatics approaches for the detection of genes and alleles: alignment with blast+, kmer-based read mapping with KMA, and direct read mapping with SRST2. A collection of 131 STEC isolates collected from food and human sources, extensively characterized with conventional molecular methods, was used as a validation dataset. Using a validation strategy specifically adopted to WGS, we demonstrated high performance with repeatability, reproducibility, accuracy, precision, sensitivity and specificity above 95 % for the majority of all assays. The WGS workflow is publicly available as a ‘push-button’ pipeline at https://galaxy.sciensano.be. Our validation strategy and accompanying reference dataset consisting of both conventional and WGS data can be used for characterizing the performance of various bioinformatics workflows and assays, facilitating interoperability between laboratories with different WGS and bioinformatics set-ups

Transforming Shiga toxin-producing Escherichia coli surveillance through whole genome sequencing in food safety practices

IntroductionShiga toxin-producing Escherichia coli (STEC) is a gastrointestinal pathogen causing foodborne outbreaks. Whole Genome Sequencing (WGS) in STEC surveillance holds promise in outbreak prevention and confinement, in broadening STEC epidemiology and in contributing to risk assessment and source attribution. However, despite international recommendations, WGS is often restricted to assist outbreak investigation and is not yet fully implemented in food safety surveillance across all European countries, in contrast to for example in the United States.MethodsIn this study, WGS was retrospectively applied to isolates collected within the context of Belgian food safety surveillance and combined with data from clinical isolates to evaluate its benefits. A cross-sector WGS-based collection of 754 strains from 1998 to 2020 was analyzed.ResultsWe confirmed that WGS in food safety surveillance allows accurate detection of genomic relationships between human cases and strains isolated from food samples, including those dispersed over time and geographical locations. Identifying these links can reveal new insights into outbreaks and direct epidemiological investigations to facilitate outbreak management. Complete WGS-based isolate characterization enabled expanding epidemiological insights related to circulating serotypes, virulence genes and antimicrobial resistance across different reservoirs. Moreover, associations between virulence genes and severe disease were determined by incorporating human metadata into the data analysis. Gaps in the surveillance system were identified and suggestions for optimization related to sample centralization, harmonizing isolation methods, and expanding sampling strategies were formulated.DiscussionThis study contributes to developing a representative WGS-based collection of circulating STEC strains and by illustrating its benefits, it aims to incite policymakers to support WGS uptake in food safety surveillance

Preliminary study of the effect of sow washing, as performed on the farm, on livestock-associated methicillin-resistant Staphylococcus aureus skin status and strain diversity

Washing sows (n = 12 per herd) on four Belgian pig farms positive for methicillin-resistant Staphylococcus aureus (MRSA) had no significant effect on MRSA status of the sow's skin (P = .32) or nares (P = 1.00). In 64% of cases, the same strain was detected before and after washing

Methicillin-resistant Staphylococcus aureus (MRSA) ST398 in pig farms and multispecies farms

During the last few years, methicillin-resistant Staphylococcus aureus (MRSA) ST398 has been isolated frequently from livestock, especially from pigs and to a lesser extent from cattle and poultry. To gain insight into the distribution of this bacterium in pig farms versus multispecies farms, 30 Belgian farms (10 pig, 10 pig/poultry and 10 pig/cattle farms) were screened for the presence of MRSA. On each farm, 10 nasal swabs were taken from pigs. When present, cattle (n = 10) were sampled in the nares and poultry (n = 10) in the nares, earlobes and cloaca. A selection of the obtained isolates were further characterized using multilocus sequence typing (MLST), spa typing, SCCmec typing, pulsed field gel electrophoresis (PFGE), multiple-locus variable-number tandem repeat analysis (MLVA) and antimicrobial susceptibility testing. On 26 of 30 farms, MRSA was isolated from pigs. Furthermore, MRSA was also isolated from poultry and cattle on one pig/poultry and five pig/cattle farms, respectively. All tested MRSA isolates belonged to ST398. Eight spa types (t011, t034, t567, t571, t1451, t2974, t3423 and t5943) were detected, among which t011 predominated. SCCmec cassettes type IVa and V were present in 20% and 72% of the isolates, respectively. When combining the results of the two remaining typing methods, PFGE and MLVA, eighteen genotypes were obtained of which one genotype predominated (56% of the positive farms). All MRSA isolates were resistant to tetracycline. Resistance to trimethoprim, aminoglycosides, macrolides, lincosamides, fluoroquinolones and chloramphenicol was also observed. In conclusion, there was no effect of the farm type on the MRSA status of the pigs. A statistically significant difference was observed when comparing the pig/poultry or the pig/cattle MRSA status on the multispecies farms. Additionally, a wide variety of MRSA ST398 strains was found within certain farms when combining different typing methods

Species and staphylococcal cassette chromosome mec (SCCmec) diversity among methicillin-resistant non-Staphylococcus aureus staphylococci isolated from pigs.

While methicillin-resistant Staphylococcus aureus (MRSA) ST398 is known to be widespread in pig farms, few studies have investigated the species diversity and SCCmec types of methicillin-resistant non-S. aureus staphylococci (MRNAS) residing in the nose of pigs. We examined nasal swab samples of 200 pigs originating from 10 Belgian pig farms previously found positive for MRSA ST398. Suspected staphylococcal isolates were subjected to a 16S rRNA-mecA-nuc PCR. Confirmed MRNAS were genotypically identified to the species level and investigated with a SCCmec typing PCR. MRNAS (n=72) were detected on all 10 farms and were carried by 29.5% of the pigs. Seven MRNAS species were found: Staphylococcus epidermidis (38.9%), Staphylococcus sciuri (18.1%), Staphylococcus pasteuri (18.1%), Staphylococcus rostri (12.5%), Staphylococcus warneri (8.3%), Staphylococcus haemolyticus (2.7%) and Staphylococcus hominis (1.4%). SCCmec cassettes were of type IVa (29.2%), type IVc (25%), type III (22.2%), type V (5.6%) or could not be assigned to any of the known types (NT types) (18.1%). Five distinct NT types were found. The predominance of methicillin-resistant S. epidermidis (MRSE) in our samples is remarkable, as MRSE is mainly associated with humans. The finding of three different SCCmec elements (IVa, V, NT type 1) in MRNAS that also prevail or predominate in MRSA ST398 shows that MRNAS might be an important SCCmec reservoir for MRSA in pigs. Yet, the occurrence of multiple other SCCmec types illustrates that further studies are required to understand the presence and spread of SCCmec in methicillin-resistant staphylococci from animals.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe