Livestock-associated Methicillin-resistant Staphylococcus Aureus Research Articles

Molecular Surveillance of MRSA in Raw Milk Provides Insight into MRSA Cross Species Evolution.

Methicillin-resistant Staphylococcus aureus (MRSA) in foods has been associated with severe infections in humans and animals worldwide. In the present study, the molecular characteristics of livestock-associated MRSA (LA-MRSA) and human-associated MRSA (hMRSA) isolates obtained in China, as well as MRSA isolates obtained from raw milk in 2018, were investigated. In total, 343 (20.38%; 343/1,683) S. aureus isolates were obtained from 1,683 raw milk samples from 100 dairy farms in 11 provinces across China. Among these, 49 (2.91%; 49/1,683) were mecA-positive MRSA. All LA-MRSA isolates were resistant to penicillin and highly resistant to erythromycin, sulfisoxazole, and clindamycin. Bioinformatic analysis the 49 genomes of LA-MRSA and 71 previously published hMRSA genomes isolated from Chinese individuals in 2018 indicated that blaZ, erm, ant(6)-Ia, aph(3')-III, tet(K), cat, and aph(2″)-Ia were more prevalent in MRSA from raw milk (P < 0.05) compared to hMRSA. Additionally, hMRSA isolates were more significantly associated with ST5 (P < 0.01) compared to LA-MRSA; in contrast, ST338 was more prevalent among LA-MRSA isolates (P < 0.01). Likewise, the SCCmec type II was only detected in hMRSA isolates, whereas SCCmec type V and IV were more prevalent among LA-MRSA (P < 0.01). Furthermore, core-genome phylogenetic analysis showed the endemic characteristics of LA-MRSA in local provinces, as well as the close evolutionary relationships between MRSA from cattle and humans. Finally, homology analysis of mecA and blaZ genetic contexts revealed a high possibility of horizontal transmission of MRSA resistance genes among raw milk-associated and hMRSA strains, which increases the risk for public health. IMPORTANCE Methicillin-resistant Staphylococcus aureus (MRSA) is considered a public health concern as it is resistant to multiple antibiotics, thus being in zoonotic transmission of antibiotic resistance genes. MRSA causes serious public health issues and leads to hard-to-treat infections in humans and animals; therefore, it was meaningful to determine the prevalence of MRSA in raw milk samples and investigate phenotype and genotype of antimicrobial resistance and molecular characteristics in livestock-associated MRSA (LA-MRSA) and human-associated MRSA (hMRSA) in China, which could provide a theoretical basis for preventing and controlling the spread of MRSA between livestock and humans.

Fluoroquinolone resistance does not facilitate phage Φ13 integration or excision in Staphylococcus aureus.

Prophages of the ΦSa3int family are commonly found in human-associated strains of Staphylococcus aureus where they encode factors for evading the human innate immune system. In contrast, they are usually absent in livestock-associated methicillin-resistant S. aureus (LA-MRSA) strains where the phage attachment site is mutated compared to the human strains. However, ΦSa3int phages have been found in a subset of LA-MRSA strains belonging to clonal complex 398 (CC398), including a lineage that is widespread in pig farms in Northern Jutland, Denmark. This lineage contains amino acid changes in the DNA topoisomerase IV and the DNA gyrase encoded by grlA and gyrA, respectively, which have been associated with fluoroquinolone (FQ) resistance. As both of these enzymes are involved in DNA supercoiling, we speculated that the mutations might impact recombination between the ΦSa3int phage and the bacterial chromosome. To examine this, we introduced the FQ resistance mutations into S. aureus 8325-4attBLA that carry the mutated CC398-like bacterial attachment site for ΦSa3int phages. When monitoring phage integration and release of Φ13, a well-described representative of the ΦSa3int phage family, we did not observe any significant differences between the FQ-resistant mutant and the wild-type strain. Thus our results suggest that mutations in grlA and gyrA do not contribute to the presence of the ΦSa3int phages in LA-MRSA CC398.

The Occurrence of Methicillin-Resistant Staphylococcus Aureus (MRSA) Around the World and Antibiotic Therapy for Selected Infections Caused By MRSA

Abstract Staphylococcus aureus is a Gram-positive bacterium, which can cause serious bacterial infections in humans. It constitutes an important etiological factor of many diseases, for instance, soft tissue and skin infections (including skin boils and abscesses), as well as life-threatening necrotizing pneumonia or toxic shock syndrome. It is estimated that about 25–30% of people are carriers of S. aureus mainly in the anterior nostrils. A smaller percentage of people are carriers of methicillin-resistant S. aureus (MRSA). In accordance with its definition, methicillin-resistant S. aureus is resistant to almost all β-lactam antibiotics. This phenomenon is mainly caused by the presence of penicillin-binding protein in the cell wall – PBP2a, which is the product of the mecA gene, which is part of the complex called SCCmec (staphylococcal cassette chromosome mec). Methicillin-resistant S. aureus (MRSA) results in endemic in hospitals around the world and are one of the leading causes of morbidity and mortality in society. Infections initiated by hospital strains of MRSA (health care-associated MRSA, HA-MRSA) concern mainly immunocompromised patients after surgery. In addition, there are populations of acommunity-associated MRSA (CA-MRSA) strains and populations of livestock-associated MRSA (LA-MRSA) strains. The treatment of infections with MRSA etiology, after exhausting the possibilities of standard antibiotic therapy with the use of i.e. vancomycin, is based on treatment with new-generation antibiotics, such as dalbavacin.

Genome Analysis of Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus ST398 Strains Isolated from Patients with Invasive Infection.

Using genomic data, we determined the origin of MRSA ST398 isolates responsible for invasive infection in patients with no known livestock contact. We sequenced the genome of seven MSSA and four MRSA ST398 isolates from patients with invasive infections between 2013 and 2017, using the Illumina technique. Prophage-associated virulence genes and resistance genes were identified. To determine the origin of the isolates, their genome sequences were included in phylogenetic analysis also encompassing the ST398 genomes available on NCBI. All isolates carried the φSa3 prophage, but with variations in the immune evasion cluster: type C in MRSA isolates, and type B in MSSA isolates. All MSSA belonged to the spa type t1451. MRSA strains had the same SCCmec type IVa (2B) cassette and belonged to spa types t899, t4132, t1939 and t2922. All MRSA harbored the tetracycline resistance gene, tet(M). Phylogenetic analysis revealed that MSSA isolates belonged to a cluster of human-associated isolates, while MRSA isolates belonged to a cluster containing livestock-associated MRSA. We showed that the clinical isolates MRSA and MSSA ST398 have different origins. An acquisition of virulence genes by livestock-associated MRSA isolates allows them to induce an invasive infection in human.

First Isolation of Methicillin-Resistant Livestock-Associated Staphylococcus aureus CC398 and CC1 in Intensive Pig Production Farms in Argentina.

Since the mid-2000s, livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has been identified among pigs worldwide, CC398 being the most relevant LA-MRSA clone. In the present work, nasal swabs were taken from healthy pigs of different age categories (25 to 154 days) from 2019 to 2021 in four intensive farms located in three provinces of Argentina. The aim of the present study was to characterize the first LA-MRSA isolates that colonized healthy fattening pigs in Argentina in terms of their resistance phenotype and genotype and to know the circulating clones in the country. Antimicrobial susceptibility, presence of the mecA gene and PCR screening of CC398 were evaluated in all the isolates. They were resistant to cefoxitin, penicillin, tetracycline, chloramphenicol and ciprofloxacin but susceptible to nitrofurantoin, rifampicin, vancomycin and linezolid. Furthermore, 79% were resistant to clindamycin and lincomycin, 68% to erythromycin, 58% to gentamicin and 37% to trimethoprim/sulfamethoxazole. All the isolates were multidrug resistant. The clonal relation was assessed by SmaI-PFGE (pulsed-field gel electrophoresis) and a representative isolate of each PFGE type was whole genome sequenced by Illumina. MLST (multilocus sequence typing), resistance and virulence genes and SCCmec typing were performed on sequenced isolates. The isolates were differentiated in three clonal types by PFGE, and they belonged to sequence-type ST398 (58%) and ST9, CC1 (42%) by MLST. SCCmec typeV and several resistance genes detected showed complete correlation with resistance phenotypes. The present study revealed that LA-MRSA colonizing healthy pigs in Argentina belongs to CC398 and CC1, two MRSA lineages frequently associated to pigs in other countries.

Genomic Diversity of Methicillin-Resistant Staphylococcus aureus CC398 Isolates Collected from Diseased Swine in the German National Resistance Monitoring Program GERM-Vet from 2007 to 2019.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) clonal complex 398 (CC398) isolates (n = 178) collected in the national resistance monitoring program GERM-Vet from diseased swine in Germany from 2007 to 2019 were investigated for their genomic diversity with a focus on virulence and antimicrobial resistance (AMR) traits. Whole-genome sequencing was followed by molecular typing and sequence analysis. A minimum spanning tree based on core-genome multilocus sequence typing was constructed, and antimicrobial susceptibility testing was performed. Most isolates were assigned to nine clusters. They displayed close phylogenetic relationships but a wide molecular variety, including 13 spa types and 19 known and four novel dru types. Several toxin-encoding genes, including eta, seb, sek, sep, and seq, were detected. The isolates harbored a wide range of AMR properties mirroring the proportions of the classes of antimicrobial agents applied in veterinary medicine in Germany. Multiple novel or rare AMR genes were identified, including the phenicol-lincosamide-oxazolidinone-pleuromutilin-streptogramin A resistance gene cfr, the lincosamide-pleuromutilin-streptogramin A resistance gene vga(C), and the novel macrolide-lincosamide-streptogramin B resistance gene erm(54). Many AMR genes were part of small transposons or plasmids. Clonal and geographical correlations of molecular characteristics and resistance and virulence genes were more frequently observed than temporal relations. In conclusion, this study provides insight into population dynamics of the main epidemic porcine LA-MRSA lineage in Germany over a 13-year-period. The observed comprehensive AMR and virulence properties, most likely resulting from the exchange of genetic material between bacteria, highlighted the importance of LA-MRSA surveillance to prevent further dissemination among swine husbandry facilities and entry into the human community. IMPORTANCE The LA-MRSA-CC398 lineage is known for its low host specificity and frequent multiresistance to antimicrobial agents. Colonized swine and their related surroundings represent a considerable risk of LA-MRSA-CC398 colonization or infection for occupationally exposed people through which such isolates might be further disseminated within the human community. This study provides insight into the diversity of the porcine LA-MRSA-CC398 lineage in Germany. Clonal and geographical correlations of molecular characteristics and resistance and virulence traits were detected and may be associated with the spread of specific isolates through livestock trade, human occupational exposure, or dust emission. The demonstrated genetic variability underlines the lineage's ability to horizontally acquire foreign genetic material. Thus, LA-MRSA-CC398 isolates have the potential to become even more dangerous for various host species, including humans, due to increased virulence and/or limited therapeutic options for infection control. Full-scale LA-MRSA monitoring at the farm, community, and hospital level is therefore essential.

Assessment of control measures against livestock-associated methicillin-resistant Staphylococcus aureus in a farrow-to-finish pig herd using infectious disease modelling

Pigs are considered to be the main reservoir for livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), which is a zoonotic opportunistic pathogen. As LA-MRSA is an occupational hazard, there is an incentive to control its spread in pig herds. Currently, knowledge about effective control measures which do not require culling the whole herd are limited, and the control strategies against LA-MRSA vary between countries. This study uses a stochastic compartment model to simulate possible control measures for LA-MRSA in a farrow-to-finish pig herd. The aims of the study were to (1) extend a previously published disease spread model with additional management and control measures; (2) use the extended model to study the effect of the individual LA-MRSA control measures on the within-herd LA-MRSA prevalence; (3) evaluate the effect of control measures when they are implemented in combinations. From the individual control measures tested in the study, thorough cleaning was found to be most effective in reducing the LA-MRSA prevalence in the herd. When the different control measures were combined, cleaning together with disease surveillance had the largest impact on reducing the LA-MRSA and a higher chance of causing disease elimination. The results of the study showed that achieving disease elimination once LA-MRSA had been introduced in the herd was challenging but was more likely when control measures were introduced early during the outbreak. This emphasises the importance of early detection of the pathogen and subsequent rapid implementation of LA-MRSA control measures.

Biosecurity and Hygiene Procedures in Pig Farms: Effects of a Tailor-Made Approach as Monitored by Environmental Samples.

In livestock, the importance of hygiene management is gaining importance within the context of biosecurity. The aim of this study was to monitor the implementation of biosecurity and hygiene procedures in 20 swine herds over a 12-month period, as driven by tailor-made plans, including training on-farm. The measure of adenosine triphosphate (ATP) environmental contents was used as an output biomarker. The presence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) and extended-spectrum β-lactamase producing Escherichia coli (ESBL-E. coli) was also investigated as sentinels of antibiotic resistance. A significant biosecurity improvement (p = 0.006) and a reduction in the ATP content in the sanitised environment (p = 0.039) were observed. A cluster including 6/20 farms greatly improved both biosecurity and ATP contents, while the remaining 14/20 farms ameliorated them only slightly. Even if the ESBL-E. coli prevalence (30.0%) after the hygiene procedures significantly decreased, the prevalence of LA-MRSA (22.5%) was unaffected. Despite the promising results supporting the adoption of tailor-made biosecurity plans and the measure of environmental ATP as an output biomarker, the high LA-MRSA prevalence still detected at the end of the study underlines the importance of improving even more biosecurity and farm hygiene in a one-health approach aimed to preserve also the pig workers health.

Survival of livestock-associated methicillin-resistant Staphylococcus aureus CC398 on different surface materials

BackgroundZoonotic livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is widely spread in pig herds in many countries. However, the knowledge regarding the survival of LA-MRSA in the pig farm environment is currently limited. The aim of this study was to assess the survival of LA-MRSA on different surface materials found in the farm environment. The study investigated the survival of two different LA-MRSA strains belonging to the clonal complex (CC) 398 on four different surfaces: stainless steel, polypropylene plastic, K30 concrete and commercial concrete disk coupons. The survival of the bacteria over time was determined by the viable count method and, where possible, fitting a model to the observed data by using nonlinear least squares method to calculate the half-life ({t}_{1/2}) for different strain and material combinations.ResultsThe study showed that the half-life of the bacteria was longer on polypropylene plastic ({t}_{1/2}=11.08–15.78 days) than on stainless steel ({t}_{1/2}=2.45–7.83 days). On these materials, both LA-MRSA strains survived through the 14 week observation period. The bacterial decay was fastest on the concrete surfaces, where LA-MRSA became undetectable after 3–9 weeks.ConclusionsThe survival of LA-MRSA in the pig farm environment may be affected by different surface materials. A more frequent sampling protocol (< 7 days) is needed to determine the half-life on concrete surfaces.

Genomics Insight into cfr-Mediated Linezolid-Resistant LA-MRSA in Italian Pig Holdings

The cfr genes encode for a 23S rRNA methyltransferase, conferring a multiresistance phenotype to phenicol, lincosamide, oxazolidinone, pleuromutilin, and streptogramin A antibiotics. These genes have been described in staphylococci, including methicillin-resistant Staphylococcus aureus (MRSA). In this study, we retrospectively performed an in-depth genomic characterisation of three cfr-positive, multidrug-resistant (MDR) livestock-associated (LA) MRSA clonal complexes (CCs) 1 and 398 detected in different Italian pig holdings (2008-2011) during population studies on Italian livestock (2008-2014). We used a combined Illumina and Oxford Nanopore Technologies (ONT) whole genome sequencing (WGS) approach on two isolates (the 2008 CC1 and the 2010 CC398 isolates, but not the 2011 CC1 isolate). Interestingly, the three isolates presented different cfr variants, with only one displaying a linezolid-resistant phenotype. In isolate 2008 CC1, the cfr gene was identified within a Tn558 composite transposon-like structure flanked by IS elements located on a novel 44,826 bp plasmid. This represents the first report of CC1 LA-MRSA harbouring the cfr gene in its functional variant. Differently, cfr was chromosomally located in isolate 2010 CC398. Our findings have significant public health implications, confirm the need for the continuous genomic surveillance of cfr-positive zoonotic LA-MRSA, and backdate cfr presence in LA-MRSA from Italian pigs to at least 2008.

High-Throughput Mutagenesis Reveals a Role for Antimicrobial Resistance- and Virulence-Associated Mobile Genetic Elements in Staphylococcus aureus Host Adaptation.

Methicillin-resistant Staphylococcus aureus (MRSA) clonal-complex 398 (CC398) is the dominant livestock-associated (LA) MRSA lineage in European livestock and an increasing cause of difficult-to-treat human disease. LA-CC398 MRSA evolved from a diverse human-associated methicillin-sensitive population, and this transition from humans to livestock was associated with three mobile genetic elements (MGEs). In this study, we apply transposon-directed insertion site sequencing (TraDIS), a high-throughput transposon mutagenesis approach, to investigate genetic signatures that contribute to LA-CC398 causing disease in humans. We identified 26 genes associated with LA-CC398 survival in human blood and 47 genes in porcine blood. We carried out phylogenetic reconstruction on 1,180 CC398 isolates to investigate the genetic context of all identified genes. We found that all genes associated with survival in human blood were part of the CC398 core genome, while 2/47 genes essential for survival in porcine blood were located on MGEs. Gene SAPIG0966 was located on the previously identified Tn916 transposon carrying a tetracycline resistance gene, which has been shown to be stably inherited within LA-CC398. Gene SAPIG1525 was carried on a phage element, which in part, matched phiSa2wa_st1, a previously identified bacteriophage carrying the Panton-Valentine leucocidin (PVL) virulence factor. Gene deletion mutants constructed in two LA-CC398 strains confirmed that the SAPIG0966 carrying Tn916 and SAPIG1525 were important for CC398 survival in porcine blood. Our study shows that MGEs that carry antimicrobial resistance and virulence genes could have a secondary function in bacterial survival in blood and may be important for host adaptation. IMPORTANCE CC398 is the dominant type of methicillin-resistant Staphylococcus aureus (MRSA) in European livestock and a growing cause of human infections. Previous studies have suggested MRSA CC398 evolved from human-associated methicillin-sensitive Staphylococcus aureus and is capable of rapidly readapting to human hosts while maintaining antibiotic resistance. Using high-throughput transposon mutagenesis, our study identified 26 and 47 genes important for MRSA CC398 survival in human and porcine blood, respectively. Two of the genes important for MRSA CC398 survival in porcine blood were located on mobile genetic elements (MGEs) carrying resistance or virulence genes. Our study shows that these MGEs carrying antimicrobial resistance and virulence genes could have a secondary function in bacterial survival in blood and may be important for blood infection and host adaptation.

High Prevalence of Livestock-Associated Methicillin-Resistant Staphylococcus aureus in Hungarian Pig Farms and Genomic Evidence for the Spillover of the Pathogen to Humans

Livestock-Associated Methicillin-ResistantStaphylococcus aureus (LA-MRSA) strains of clonal complex (CC) 398 are widely disseminated in pigs and are considered emerging pathogens in human medicine. To investigate the prevalence, genetic characteristics, and zoonotic potential of the pathogen in pig production settings, dust samples were collected from 40 pig operations in Hungary, along with nasal swabs of attending veterinarians and other swine professionals (n = 27) in 2019. MRSA isolates were further characterized by performing whole-genome sequencing and susceptibility testing. The whole-genome sequences of 14 human-derived LA-MRSA clinical isolates from the same year were also included in the study. The proportion of positive farms was 83% (33/40), and 70% (19/27) of the swine professionals carried the pathogen. All but one MRSA strain belonged to CC398, including the human clinical isolates. The core genome multilocus sequence typing (cgMLST) analysis revealed clusters of closely-related isolates of both environmental and human origin with a pairwise allelic distance of ≤24, and both cgMLST and single nucleotide polymorphism (SNP) analyses suggest recent transmission events between the farm environment and humans. Four human clinical isolates harboured the immune-evasion gene cluster, of which one was considered to be closely related to farm isolates. Half of the swine-related strains showed decreased susceptibility to eight or more antimicrobials, and along with human isolates, they carried eight different types of multidrug-resistance genes, including cfr. The results showed a dramatic increase in the occurrence of LA-MRSA in the swine industry in Hungary, compared with the 2% prevalence reported by the European Food Safety Authority baseline study in 2008. The wide range of antimicrobial resistance of the strains, accompanied by the emergence of the pathogen in humans — both asymptomatic carriers and diseased — call for revision of the risk posed by LA-MRSA to the public health.

Staphylococcus aureus and MRSA in Livestock: Antimicrobial Resistance and Genetic Lineages.

Animal production is associated with the frequent use of antimicrobial agents for growth promotion and for the prevention, treatment, and control of animal diseases, thus maintaining animal health and productivity. Staphylococcus aureus, in particular methicillin-resistant S. aureus (MRSA), can cause a variety of infections from superficial skin and soft tissue infections to life-threatening septicaemia. S. aureus represents a serious public health problem in hospital and community settings, as well as an economic and animal welfare problem. Livestock-associated MRSA (LA-MRSA) was first described associated with the sequence (ST) 398 that was grouped within the clonal complex (CC) 398. Initially, LA-MRSA strains were restricted to CC398, but over the years it has become clear that its diversity is much greater and that it is constantly changing, a trend increasingly associated with multidrug resistance. Therefore, in this review, we aimed to describe the main clonal lineages associated with different production animals, such as swine, cattle, rabbits, and poultry, as well as verify the multidrug resistance associated with each animal species and clonal lineage. Overall, S. aureus ST398 still remains the most common clone among livestock and was reported in rabbits, goats, cattle, pigs, and birds, often together with spa-type t011. Nevertheless, a wide diversity of clonal lineages was reported worldwide in livestock.

Antibiotic resistance and molecular typing of clinical Staphylococcus aureus isolates from Malaysian military hospital

Objective: To determine the antibiotic resistance profile (ARP) of Staphylococcus (S.) aureus isolates and molecular typing of the methicillin-resistant S. aureus (MRSA) isolates from Tuanku Mizan Armed Forces Hospital (TMAFH), Kuala Lumpur. Methods: The ARP and presence of the pvl gene were determined for 209 S. aureus isolates from clinical specimens. Of these, 123 were methicillin-susceptible S. aureus (MSSA) isolates and 86 were MRSA isolates. All MRSA isolates were characterized using SCCmec typing and spa typing. Descriptive analysis was performed to compare the demographic data with the phenotypic and genotypic variables of the S. aureus isolates. Results: No vancomycin-intermediate and -resistant S. aureus (VISA and VRSA, respectively) were detected among the study isolates. The MSSA isolates showed low resistance rates to all tested antibiotics, were commonly invasive (28/42, 66.7%), and mostly harboured pvl (35/42, 83.3%). Meanwhile, MRSA isolates showed high resistance to penicillin (86/86, 100%), ampicillin (86/86, 100%), sulbactam/ampicillin (86/86, 100%), cefuroxime (81/86, 94.19%), cefoperazone (76/86, 88.37%), azithromycin (56/86, 65.12%), and erythromycin (54/86, 62.79%). The majority of MRSA isolates were of SCCmec type IVh (65/86, 75.58%), spa type t032 (55/85, 63.95%), and grouped into spaCC-t022 (66/85, 77.65%). The t032 type was found to be associated with resistance traits to azithromycin and erythromycin (P<0.05). We also found several spa types that are typically associated with hospital-, community-, and livestock-associated MRSA co-existing in our MRSA population. Conclusions: This study reflected the consistent absence of VISA and VRSA and corroborated the clonal shifting of MRSA isolates in the Malaysian MRSA isolates.

Livestock-associated methicillin-resistant Staphylococcus aureus epidemiology, genetic diversity, and clinical characteristics in an urban region.

While Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), defined as CC398, is a well-known pathogen among those working with livestock, there are indications that LA-MRSA prevalence among the general population is increasing. However, the clinical impact in urban areas remains unknown. The aim of this study was to assess the genetic epidemiology and clinical characteristics of LA-MRSA in an urban area with a limited livestock population. In this retrospective study, we evaluated LA-MRSA strains that were collected between 2014 and 2018 from patients who received clinical care in a single urban area in Netherlands. Patient files were assessed for livestock exposure data, clinical findings, and contact tracing information. Next-generation sequencing (NGS) analysis in combination with wgMLST was conducted to assess genetic diversity and relatedness and to detect virulence and resistance genes. LA-MRSA strains were cultured from 81 patients, comprising 12% of all the MRSA strains found in seven study laboratories between 2014 and 2018. No livestock link was found in 76% of patients (n = 61), and 28% of patients (n = 23) had an infection, mostly of the skin or soft tissue. Contact tracing had been initiated in 14 cases, leading to the identification of two hospital transmissions: a cluster of 9 cases and one of 2 cases. NGS data were available for 91% (n = 75) of the patients. wgMLST confirmed the clusters detected via contact tracing (n = 2) and identified 5 additional clusters without a known epidemiological link. Relevant resistance and virulence findings included the PVL virulence gene (3 isolates) and tetracycline resistance (79 isolates). LA-MRSA may cause a relevant burden of disease in urban areas. Surprisingly, most infections in the present study occurred in the absence of a livestock link, suggesting inter-human transmission. These findings and the presence of PVL and other immune evasive complex virulence genes warrant future surveillance and preventative measures.

Diversity and Dissemination of Methicillin-Resistant Staphylococcus aureus (MRSA) Genotypes in Southeast Asia.

Methicillin-resistant Staphylococcus aureus (MRSA) is a successful pathogen that has achieved global dissemination, with high prevalence rates in Southeast Asia. A huge diversity of clones has been reported in this region, with MRSA ST239 being the most successful lineage. Nonetheless, description of MRSA genotypes circulating in the Southeast Asia region has, until now, remained poorly compiled. In this review, we aim to provide a better understanding of the molecular epidemiology and distribution of MRSA clones in 11 Southeast Asian countries: Singapore, Malaysia, Thailand, Vietnam, Cambodia, Lao People's Democratic Republic (PDR), Myanmar, Philippines, Indonesia, Brunei Darussalam, and Timor-Leste. Notably, while archaic multidrug-resistant hospital-associated (HA) MRSAs, such as the ST239-III and ST241-III, were prominent in the region during earlier observations, these were then largely replaced by the more antibiotic-susceptible community-acquired (CA) MRSAs, such as ST22-IV and PVL-positive ST30-IV, in recent years after the turn of the century. Nonetheless, reports of livestock-associated (LA) MRSAs remain few in the region.

Review of pork and pork products as a source for transmission of methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is an opportunistic bacterium that can cause infection in animals and humans. Recently, MRSA from food-producing or farm animals has been identified as livestock-associated MRSA (LA-MRSA). The spread of LA-MRSA is particularly found in pork and pork products because LA-MRSA has been widely known to infect pigs. The most common type of LA-MRSA identified in pork and pork products is the clonal complex LA-MRSA 398 (LA-MRSA CC398). The MRSA strains on the surface of pork carcasses can be spread during the handling and processing of pork and pork products through human hands, cutting tools, and any surface that comes into direct contact with pork. Food infection is the main risk of MRSA in pork and pork products consumed by humans. Antibiotics to treat food infection cases due to MRSA infection include vancomycin and tigecycline. The spread of MRSA in pork and pork products is preventable by appropriately cooking and cooling the pork and pork products at temperatures above 60°C and below 5°C, respectively. It is also necessary to take other preventive measures, such as having a clean meat processing area and disinfecting the equipment used for processing pork and pork products. This review aimed to explain epidemiology, transmission, risk factors, diagnosis, public health consequences, treatment of food poisoning, and preventing the spread of MRSA in pork and pork products.

Low-level colonization of methicillin-resistant Staphylococcus aureus in pigs is maintained by slowly evolving, closely related strains in Finnish pig farms

BackgroundOver the past two decades, livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has become widely prevalent in pig production in Europe. The carriage status of LA-MRSA is known to vary among individual pigs, but bacterial load in pigs has rarely been studied. We assessed the quantity of LA-MRSA in nasal and skin samples of pigs and investigated the genetic diversity of the strains together with sequenced strains from national surveillance and pathology samples from the Finnish Food Authority. On two farms with assumed MRSA-positive status, farm 1 and farm 2, 10 healthy pigs were sampled three times during 2 weeks from the nares and skin (study A). On farm 1, 54 additional pigs were sampled and from confirmed MRSA-positive animals, 10 were randomly selected and transported to a clean, controlled environment for further sampling (study B). From the samples taken on farms 1 and 2 and in the controlled environment, MRSA was isolated both by direct plating and enrichment on selective media. spa types, multilocus sequence types, staphylococcal cassette chromosome mec types, resistance and virulence genes were determined. Core genome multilocus sequence typing (cgMLST) analysis was performed, including the sequences deriving from the surveillance/pathology samples from the Finnish Food Authority.ResultsAll pigs on farm 1 carried LA-MRSA in the nares at all three time points and five pigs on farm 2 at one time point. Nasal quantity varied between 10 and 103 CFU/swab and quantity on the skin between 10 and 102 CFU/swab. In the controlled environment, MRSA was detected in at least one of the nasal samples from each animal. spa type t034 was predominant. cgMLST showed one cluster with minimum allele differences between 0 and 11.ConclusionsThe study shows predominantly low-level carriage (< 103 CFU/swab) of LA-MRSA on farms. In the controlled environment we observed a decline in nasal carriage but constant skin carriage. cgMLST showed that strains of spa type t034 are closely related at the national level.

How to survive pig farming: Mechanism of SCCmec element deletion and metabolic stress adaptation in livestock-associated MRSA.

Previous research on methicillin susceptible Staphylococcus aureus (MSSA) belonging to livestock-associated (LA-) sequence type (ST) 398, isolated from pigs and their local surroundings, indicated that differences between these MSSA and their methicillin resistant predecessors (MRSA) are often limited to the absence of the staphylococcal cassette chromosome mec (SCCmec) and few single nucleotide polymorphisms. So far, our understanding on how LA-MRSA endure the environmental conditions associated with pig-farming as well as the putative impact of this particular environment on the mobilisation of SCCmec elements is limited. Thus, we performed in-depth genomic and transcriptomic analyses using the LA-MRSA ST398 strain IMT38951 and its methicillin susceptible descendant. We identified a mosaic-structured SCCmec region including a putative replicative SCCmecVc which is absent from the MSSA chromosome through homologous recombination. Based on our data, such events occur between short repetitive sequences identified within and adjacent to two distinct alleles of the large cassette recombinase genes C (ccrC). We further evaluated the global transcriptomic response of MRSA ST398 to particular pig-farm associated conditions, i.e., contact with host proteins (porcine serum) and a high ammonia concentration. Differential expression of global regulators involved in stress response control were identified, i.e., ammonia-induced alternative sigma factor B-depending activation of genes for the alkaline shock protein 23, the heat shock response and the accessory gene regulator (agr)-controlled transcription of virulence factors. Exposure to serum transiently induced the transcription of distinct virulence factor encoding genes. Transcription of genes reported for mediating the loss of methicillin resistance, especially ccrC, was not significantly different compared to the unchallenged controls. We concluded that, from an evolutionary perspective, bacteria may save energy by incidentally dismissing a fully replicative SCCmec element in contrast to the induction of ccr genes on a population scale. Since the genomic SCCmec integration site is a hot-spot of recombination, occasional losses of elements of 16 kb size may restore capacities for the uptake of foreign genetic material. Subsequent spread of resistance, on the other hand, might depend on the autonomous replication machinery of the deleted SCCmec elements that probably enhance chances for reintegration of SCCmec into susceptible genomes by mere multiplication.

Antimicrobial Resistance and Molecular Characterization of Methicillin-Resistant Staphylococcus aureus from Two Pig Farms: Longitudinal Study of LA-MRSA.

Pigs were identified as the most important reservoir of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), mostly belonging to the emergent zoonotic clonal complex (CC) 398. Here, we investigated the presence of MRSA in sows and piglets over a period of several months in two pig farms (intensive farm A and family-run farm B). Isolates underwent antimicrobial susceptibility testing, PCR characterization and spa typing. We collected 280 samples, namely 206 nasal swabs from pigs and 74 environmental samples from pig housings at 12 consecutive time points. A total of 120/161 (74.5%) and 75/119 (63.0%) samples were MRSA-positive in farms A and B, respectively. All isolates harbored mecA but lacked mecC and PVL-encoding genes. The identified spa types (t571, t034, t1250 and t898 in farm A, t1451 and t011 in farm B) were indicative of CC398. Antimicrobial resistance patterns (all multidrug resistant in farm A, 57.2% in farm B) depended on the farm, suggesting the impact of farm size and management practices on the prevalence and characteristics of MRSA. Due to the intermittent colonization of pigs and the high contamination of their immediate environment, MRSA status should be determined at the farm level when considering preventive measures or animal trade between farms.