Evaluation of a widely used culture-based method for detection of livestock-associated meticillin-resistant Staphylococcus aureus (MRSA), Denmark and Norway, 2014 to 2016.
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ABSTRACT: We evaluated a widely used culture-based method for detection of livestock-associated meticillin-resistant Staphylococcus aureus (LA-MRSA) in samples collected from pigs and the environment inside pig stables in Denmark and Norway. Selective enrichment in tryptic soy broth containing cefoxitin and aztreonam led to a high ratio of false-negative results (26%; 57/221). On this basis, we recommend reconsidering the use of selective enrichment for detection of LA-MRSA in animal and environmental samples.
Project description:BackgroundLivestock-associated methicillin-resistant Staphylococcus aureus clonal complex 398 (LA-MRSA CC398) is causing an increasing number of skin and soft tissue infections (SSTIs) in Denmark and other European countries with industrial pig production. Yet, its impact on MRSA bloodstream infections (BSIs) has not been well studied.MethodsWe investigated the clinical epidemiology of all human cases of LA-MRSA CC398 BSI during 2010-2015. Cases of LA-MRSA CC398 BSI were compared to cases of BSI caused by other types of MRSA and cases of SSTI caused by LA-MRSA CC398. Whole-genome sequence analysis was used to assess the phylogenetic relationship among LA-MRSA CC398 isolates from Danish pigs and cases of BSI and SSTI.ResultsThe number of LA-MRSA CC398 BSIs and SSTIs increased over the years, peaking in 2014, when LA-MRSA CC398 accounted for 16% (7/44) and 21% (211/985) of all MRSA BSIs and SSTIs, corresponding to 1.2 and 37.4 cases of BSI and SSTI per 1000000 person-years, respectively. Most patients with LA-MRSA CC398 BSI had no contact to livestock, although they tended to live in rural areas. LA-MRSA CC398 caused 24.3 BSIs per 1000 SSTIs among people with no livestock contact, which is similar to the ratio observed for other types of MRSA. Whole-genome sequence analysis showed that most of the BSI and SSTI isolates were closely related to Danish pig isolates.ConclusionsThis study demonstrates that the increasing number of LA-MRSA CC398 BSIs occurred in parallel with a much larger wave of LA-MRSA CC398 SSTIs and an expanding pig reservoir.
Project description:Pigs, cattle and poultry are colonized with MRSA and the zoonotic transmission of such MRSA to humans via direct animal contact, environmental contaminations or meat are a matter of concern. Livestock-associated (LA) MRSA are mostly belonging to clonal complex (CC) 398 as defined by multilocus sequence typing. However, MRSA of other clonal lineages including CC5, CC9 and CC97 have also been detected in livestock animals in Germany. Within the framework of a Dutch-German network project (EUREGIO), 14,036 MRSA isolated from clinical and screening specimens (January 2008 - June 2012) derived from human patients in hospitals as well as general or specialized practices in a German region characterized by a high density of livestock production, were subjected to S. aureus protein A (spa) sequence typing. The prevalence of putative LA-MRSA among the human MRSA isolates was determined by analyzing the detection of livestock-indicator (LI) spa types which had already been reported in German livestock. Overall, 578 spa types were detected among the MRSA isolates. LI spa types t011, t034, t108, t1451, t2011, t571, t1456, t1250, t1255, t1580, t2970, t2346, t1344, t2576, t2330 and t2510 (all of which are indicative for LA-MRSA CC398) accounted for 18.6% of all human isolates. The LI spa types t1430 (CC9), t3992 (CC97), t002 (CC5) and t007 (CC30) were found in 0.14%, 0.01%, 1.01% and 0.04% of all human MRSA isolates, respectively. LI spa types associated with CC398 represented 23% of all MRSA from screening samples and a varying proportion among isolates from clinical specimens ranging between 0% in cerebrospinal fluid, 8% in blood cultures and 14% in deep respiratory fluids. Our findings indicate that LA-MRSA are a major cause for human infection and stress the need for close surveillance. Although LA-MRSA CC398 predominates, the occurrence of putative LA-MRSA from other clonal lineages should be monitored.
Project description:Methicillin-resistant Staphylococcus aureus (MRSA) colonization of livestock animals is common and prevalence rates for pigs have been reported to be as high as 49%. Mechanisms contributing to the persistent carriage and high prevalence rates of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) strains in swine herds and production facilities have not been investigated. One explanation for the high prevalence of MRSA in swine herds is the ability of these organisms to exist as biofilms. In this report, the ability of swine LA-MRSA strains, including ST398, ST9, and ST5, to form biofilms was quantified and compared to several swine and human isolates. The contribution of known biofilm matrix components, polysaccharides, proteins and extracellular DNA (eDNA), was tested in all strains as well. All MRSA swine isolates formed robust biofilms similar to human clinical isolates. The addition of Dispersin B had no inhibitory effect on swine MRSA isolates when added at the initiation of biofilm growth or after pre-established mature biofilms formed. In contrast, the addition of proteinase K inhibited biofilm formation in all strains when added at the initiation of biofilm growth and was able to disperse pre-established mature biofilms. Of the LA-MRSA strains tested, we found ST398 strains to be the most sensitive to both inhibition of biofilm formation and dispersal of pre-formed biofilms by DNaseI. Collectively, these findings provide a critical first step in designing strategies to control or eliminate MRSA in swine herds.
Project description:Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) sequence type 9 (ST9) has emerged and disseminated in Asia. It is associated with colonization or infection in both humans and animal hosts; however, the genetic factors underpinning its adaptation to animal and human population remain to be determined. Here, we conducted a genomic analysis of 191 ST9 S. aureus genomes collected from 12 different countries, including 174 genomes retrieved from public databases and 17 sequenced in this study. In silico spa typing, staphylococcal cassette chromosome mec (SCCmec) typing, and antimicrobial resistance and virulence gene mining were conducted, and the temporal phylogenetic signal was assessed by Bayesian inference. Our results point toward a human methicillin-susceptible S. aureus (MSSA) origin of ST9 that evolved approximately 2 centuries ago. Three major genetic events occurred during ST9 host shift from human to animals: the loss of the immune evasion cluster genes (scn, chp, and sak), which were reported to contribute to virulence in human infections, the acquisition of the SaPIbov4-like element-encoding vwb gene, which is an animal-specific virulence factor responsible for the clotting of animal plasma, and the acquisition of antibiotic resistance genes, including SCCmec, quinolone resistance-determining region (QRDR) mutations, and a multidrug resistance genetic element (MDRST9). Evidence of direct transmission of animal-adapted strains to human hosts also suggest that transmission could potentially reshape the resistance and virulence genetic pool in these isolates. The rapid clonal expansion of MDR ST9 strains in mainland China and Taiwan highlights the increasing need for effective surveillance of antibiotic consumption in animal husbandry to control antimicrobial resistance spread. IMPORTANCE Staphylococcus aureus sequence type 9 (ST9) is the main LA-MRSA clone spreading in the Asian continent. It can colonize and cause mild to severe infections both in animal and humans. Previous work described its genotypic characteristics; however, the molecular history of global spread of ST9 strains remains largely unclear. We conducted a detailed analysis of genomic evolution of global ST9 strains and identified key genetic changes associated with its adaptation to specific hosts. Our results suggest that the ST9 clone originated from human-adapted strains, which lost genes related to the evasion of the immune system. The introduction of ST9 strains in animal populations was aligned with the acquisition of animal-specific virulent factors and mobile elements harboring multiple antimicrobial resistance genes, especially in isolates from mainland China and Taiwan.
Project description:The spread of livestock-associated methicillin-resistant Staphylococcus aureus clonal complex 398 (LA-MRSA CC398) within the Danish pig production system has been linked to an increased number of human infections. Yet, the population structure and transmission dynamics of this important pathogen remain poorly understood. In this study, whole-genome sequences from 371 LA-MRSA CC398 isolates collected between 2004 and 2015 were subjected to bioinformatic analyses. The isolates originated from Danish pig farms (n = 209) and people having livestock contact (n = 79). In addition, whole-genome sequence data from 82 isolates representing an international reference collection and 83 isolates from Danish patients were included in the analysis. The results demonstrated that the increasing prevalence of LA-MRSA CC398 in Danish pigs and patients was caused by clonal expansion of three dominant lineages. The results also showed that these lineages were enriched for the tetracycline resistance gene tet(K) and other determinants conferring resistance to some of the most frequently used antimicrobials in Danish pigs. The association between pig movements and the spread of LA-MRSA CC398 was assessed in a Poisson regression analysis of 17,009 pig movements into 273 farms with known LA-MRSA CC398 status. The results demonstrated that animal movements have played a critical role in the dissemination of LA-MRSA CC398 within the Danish pig production system, although other transmission routes may also have contributed. Consistent with this scenario, the genetic relatedness of isolates from different farms was positively correlated with the number of animal movements between the farms.IMPORTANCE Livestock-associated methicillin-resistant Staphylococcus aureus clonal complex CC398 (LA-MRSA CC398) is resistant to nearly all β-lactams and several non-β-lactam antimicrobials. Over the last decade, it has become widespread in pig farms across Europe and is now an important cause of human infections in countries with previously low levels of MRSA, such as the Netherlands and Denmark. The hitherto uncontrolled spread of LA-MRSA CC398 underscores an urgent need to understand its epidemiology in order to develop evidence-based interventions. This study demonstrates that pig movements between farms in combination with increased bacterial resistance to specific antibiotics and heavy metals were important drivers of the rapid spread of LA-MRSA CC398 in the Danish pig production system. These findings should be taken into consideration when researchers and policy makers evaluate and decide on actions and policies to limit the spread of LA-MRSA CC398 and other pathogens in food animals.
Project description:Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has been reported to colonize and cause infections in animals as well as in humans. LA-MRSA isolates have only recently been identified in patients admitted to Kuwait hospitals. This study was conducted to characterize LA-MRSA isolates obtained from patients admitted to Kuwait hospitals. A total of 202 (7.1%) of 2,823 MRSA isolates obtained from clinical samples in 2016 and 2017 in 11 public Kuwait hospitals were assigned to lineages previously known to be associated with livestock. They were characterized using antibiogram, spa typing, and DNA microarray for the assignment of clonal complexes (CCs) and detection of antibiotic resistance and virulence determinants. Identification as putative LA-MRSA clones was based on the molecular definition inferred from DNA microarray. The LA-MRSA isolates consisted of CC96 (N = 31), CC97 (N = 169), and CC398 (N = 2). Isolates belonging to CC96 and CC398 were resistant to erythromycin and clindamycin mediated by erm(A) and erm(C). CC97 isolates were multiresistant to gentamicin, kanamycin, erythromycin, clindamycin, tetracycline, chloramphenicol, fusidic acid, trimethoprim, and ciprofloxacin and harbored aacA-aphD, erm(A), erm(C), msr(A), tet(K), cat, fusC, and dfrS1. In total, 35 spa types were identified among the isolates. CC398 isolates consisted of t899 and t034. Ten spa types were identified among CC96 with t11822 (N = 13) as the most prevalent. CC97 consisted of 26 spa types with most belonging to t267 (N = 73) followed by t359 (N = 39). CC398 was composed of CC398-MRSA-IV and CC398-MRSA-V (PVL+). CC96 belonged to CC96-MRSA-IV and CC96-MRSA-IV (PVL+) Central Asian caMRSA/WA MRSA-119. CC97 consisted of six strains including CC97-MRSA-V (fusC +), CC97-MRSA-IV WA MRSA-54/63, CC97-MRSA-V, CC97-MRSA-(V+fus), CC97-MRSA-(mec VI+fus), and CC97-MRSA (mecV/VT+fus+ccrAB2). Whereas CC96 and CC97 isolates were identified in 2016 and 2017, CC398 isolates were detected only in 2016. This study identified four LA-MRSA clones among MRSA isolated from patients in Kuwait hospitals in 2016-2017 with CC97-MRSA-V (fusC +) as the dominant clone. The presence of LA-MRSA with different genetic backgrounds suggests its independent acquisition from different sources.
Project description:Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has been reported to colonize and cause infections in animals as well as in humans. LA-MRSA isolates have only recently been identified in patients admitted to Kuwait hospitals. This study was conducted to characterize LA-MRSA isolates obtained from patients admitted to Kuwait hospitals. A total of 202 (7.1%) of 2,823 MRSA isolates obtained from clinical samples in 2016 and 2017 in 11 public Kuwait hospitals were assigned to lineages previously known to be associated with livestock. They were characterized using antibiogram, spa typing, and DNA microarray for the assignment of clonal complexes (CCs) and detection of antibiotic resistance and virulence determinants. Identification as putative LA-MRSA clones was based on the molecular definition inferred from DNA microarray. The LA-MRSA isolates consisted of CC96 (N = 31), CC97 (N = 169), and CC398 (N = 2). Isolates belonging to CC96 and CC398 were resistant to erythromycin and clindamycin mediated by erm(A) and erm(C). CC97 isolates were multiresistant to gentamicin, kanamycin, erythromycin, clindamycin, tetracycline, chloramphenicol, fusidic acid, trimethoprim, and ciprofloxacin and harbored aacA-aphD, erm(A), erm(C), msr(A), tet(K), cat, fusC, and dfrS1. In total, 35 spa types were identified among the isolates. CC398 isolates consisted of t899 and t034. Ten spa types were identified among CC96 with t11822 (N = 13) as the most prevalent. CC97 consisted of 26 spa types with most belonging to t267 (N = 73) followed by t359 (N = 39). CC398 was composed of CC398-MRSA-IV and CC398-MRSA-V (PVL+). CC96 belonged to CC96-MRSA-IV and CC96-MRSA-IV (PVL+) Central Asian caMRSA/WA MRSA-119. CC97 consisted of six strains including CC97-MRSA-V (fusC +), CC97-MRSA-IV WA MRSA-54/63, CC97-MRSA-V, CC97-MRSA-(V+fus), CC97-MRSA-(mec VI+fus), and CC97-MRSA (mecV/VT+fus+ccrAB2). Whereas CC96 and CC97 isolates were identified in 2016 and 2017, CC398 isolates were detected only in 2016. This study identified four LA-MRSA clones among MRSA isolated from patients in Kuwait hospitals in 2016-2017 with CC97-MRSA-V (fusC +) as the dominant clone. The presence of LA-MRSA with different genetic backgrounds suggests its independent acquisition from different sources.
Project description:Before an efficient control strategy for livestock-associated methicillin resistant Staphylococcus aureus (LA-MRSA) in pigs can be decided upon, it is necessary to obtain a better understanding of how LA-MRSA spreads and persists within a pig herd, once it is introduced. We here present a mechanistic stochastic discrete-event simulation model for spread of LA-MRSA within a farrow-to-finish sow herd to aid in this. The model was individual-based and included three different disease compartments: susceptible, intermittent or persistent shedder of MRSA. The model was used for studying transmission dynamics and within-farm prevalence after different introductions of LA-MRSA into a farm. The spread of LA-MRSA throughout the farm mainly followed the movement of pigs. After spread of LA-MRSA had reached equilibrium, the prevalence of LA-MRSA shedders was predicted to be highest in the farrowing unit, independent of how LA-MRSA was introduced. LA-MRSA took longer to spread to the whole herd if introduced in the finisher stable, rather than by gilts in the mating stable. The more LA-MRSA positive animals introduced, the shorter time before the prevalence in the herd stabilised. Introduction of a low number of intermittently shedding pigs was predicted to frequently result in LA-MRSA fading out. The model is a potential decision support tool for assessments of short and long term consequences of proposed intervention strategies or surveillance options for LA-MRSA within pig herds.
Project description:Farm animals have been identified as an emerging reservoir for transmission of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) to humans. The low incidence of MRSA in humans and farm animals in Norway has led to the implementation of a national strategy of surveillance and control of LA-MRSA aiming to prevent livestock becoming a domestic source of MRSA to humans. In 2015, MRSA clonal complex 1 spa-type t177 was identified in nine Norwegian pig herds in two neighboring counties. An outbreak investigation was undertaken, and measures of control through eradication were imposed. We performed a register-based cohort study including pig herds and MRSA-positive persons in Norway between 2008 and 2016 to investigate the livestock-association of MRSA CC1, the transmission of the outbreak strain to humans before and after control measures, and the effect of control measures imposed. Data from the Norwegian Surveillance System of Communicable Diseases were merged with data collected through outbreak investigations for LA-MRSA, the National Registry and the Norwegian Register for Health Personnel. Whole-genome sequencing was performed on isolates from livestock and humans identified through contact tracing, in addition to t177 and t127 isolates diagnosed in persons in the same counties. It is likely that a farm worker introduced MRSA CC1 to a sow farm, and further transmission to eight fattening pig farms through trade of live pigs confirmed the potential for livestock association of this MRSA type. The outbreak strain formed a distinct phylogenetic cluster which in addition to the pig farms included one sheep herd and five exposed persons. None of the investigated isolates from possible cases without direct contact to the MRSA positive farms were phylogenetically related to the outbreak strain. Moreover, isolates of t177 or t127 from healthcare and community-acquired cases were not closely related to the outbreak cluster. Eradication measures imposed were effective in eliminating MRSA t177 from the positive pig holdings, and the outbreak strain was not detected in the national pig population or in persons from these counties after control measures.
Project description:Staphylococcus aureus clonal complex 398 (CC398) isolates cluster into two distinct phylogenetic clades based on single-nucleotide polymorphisms (SNPs) revealing a basal human clade and a more derived livestock clade. The scn and tet(M) genes are strongly associated with the human and the livestock clade, respectively, due to loss and acquisition of mobile genetic elements. We present canonical single-nucleotide polymorphism (canSNP) assays that differentiate the two major host-associated S. aureus CC398 clades and a duplex PCR assay for detection of scn and tet(M). The canSNP assays correctly placed 88 S. aureus CC398 isolates from a reference collection into the human and livestock clades and the duplex PCR assay correctly identified scn and tet(M). The assays were successfully applied to a geographically diverse collection of 272 human S. aureus CC398 isolates. The simple assays described here generate signals comparable to a whole-genome phylogeny for major clade assignment and are easily integrated into S. aureus CC398 surveillance programs and epidemiological studies.