Thijs Bosch

Prevalence of methicillin-resistant Staphylococcus aureus in meat

Recently the isolation of methicillin-resistant Staphylococcus aureus (MRSA) strains from several food-producing animals has been reported. During slaughtering of MRSA-positive animals, contamination of carcasses with MRSA may occur and consequently the meat of these animals may get contaminated. The aim of this study was to estimate the prevalence of MRSA in raw meat samples from the retail trade. Samples of raw beef, pork, veal, lamb/mutton, chicken, turkey, fowl and game were collected from the retail trade. A detection method including a two-step enrichment in Mueller-Hinton broth+6.5% NaCl and phenol red mannitol broth containing ceftizoxime and aztreonam, followed by isolation on MRSA ID agar (bioMérieux) was evaluated and subsequently applied for the detection of MRSA in samples of raw meats. MRSA strains were isolated from 264 (11.9%) of 2217 samples analyzed. Isolation percentages for the meat species were: beef (10.6%), veal (15.2%), lamb and mutton (6.2%), pork (10.7%), chicken (16.0%), turkey (35.3%), fowl (3.4%) and game (2.2%). The majority (85%) of the isolated strains belonged to spa-types of pulsed-field gel electrophoresis (PFGE) non-typeable (NT)-MRSA, corresponding to the multilocus sequence type ST398, a type also recently isolated in the Netherlands from pigs. However, a smaller part of these strains were found to be of other STs, possibly of human origin. Further studies are needed to elucidate transmission routes of MRSA in relation to meat and other foods and to provide the tools for preventing the spread of MRSA. At present the high prevalence of MRSA in meat has not been shown to contribute significantly to the dissemination of MRSA to humans and the possible health hazard for consumers of the presence of MRSA in foods should be further elucidated.

Pan-European Distribution of White-Nose Syndrome Fungus (Geomyces destructans) Not Associated with Mass Mortality

Background The dramatic mass mortalities amongst hibernating bats in Northeastern America caused by “white nose-syndrome” (WNS) continue to threaten populations of different bat species. The cold-loving fungus, Geomyces destructans, is the most likely causative agent leading to extensive destruction of the skin, particularly the wing membranes. Recent investigations in Europe confirmed the presence of the fungus G. destructans without associated mass mortality in hibernating bats in six countries but its distribution remains poorly known. Methodology/Principal Findings We collected data on the presence of bats with white fungal growth in 12 countries in Europe between 2003 and 2010 and conducted morphological and genetic analysis to confirm the identity of the fungus as Geomyces destructans. Our results demonstrate the presence of the fungus in eight countries spanning over 2000 km from West to East and provide compelling photographic evidence for its presence in another four countries including Romania, and Turkey. Furthermore, matching prevalence data of a hibernaculum monitored over two consecutive years with data from across Europe show that the temporal occurrence of the fungus, which first becomes visible around February, peaks in March but can still be seen in some torpid bats in May or June, is strikingly similar throughout Europe. Finally, we isolated and cultured G. destructans from a cave wall adjacent to a bat with fungal growth. Conclusions/Significance G. destructans is widely found over large areas of the European continent without associated mass mortalities in bats, suggesting that the fungus is native to Europe. The characterisation of the temporal variation in G. destructans growth on bats provides reference data for studying the spatio-temporal dynamic of the fungus. Finally, the presence of G. destructans spores on cave walls suggests that hibernacula could act as passive vectors and/or reservoirs for G. destructans and therefore, might play an important role in the transmission process.

Prevalence of livestock-associated MRSA in broiler flocks and risk factors for slaughterhouse personnel in The Netherlands.

To determine methicillin-resistant Staphylococcus aureus (MRSA) carriage in poultry and slaughterhouse personnel, 40 Dutch broiler flocks, in six slaughterhouses and 466 personnel were sampled. Of the employees, 26 were positive (5.6%), indicating a higher risk of exposure when compared to the general Dutch population (0.1%). This risk was significantly higher for personnel having contact with live animals (5.2%) - especially hanging broilers on the slaughterline (20.0%) - than for all other personnel (1.9%). Conventional electric stunning conferred a significantly higher risk of MRSA carriage for employees than CO2 stunning (9.7% vs. 2.0%). A total of 405 broilers were sampled upon their arrival at the slaughterhouse, of which 6.9% were positive. These broilers originated from 40 Dutch slaughter flocks of which 35.0% were positive. MRSA contamination in the different compartments of slaughterhouses increased during the production day, from 8% to 35%. Of the 119 MRSA isolates, predominantly livestock-associated MRSA ST398 was found, although 27.7% belonged to ST9 (spa type t1430). There is an increased risk of MRSA carriage in personnel working at broiler slaughterhouses, particularly those having contact with live animals.

Unexpected sequence types in livestock associated methicillin-resistant Staphylococcus aureus (MRSA): MRSA ST9 and a single locus variant of ST9 in pig farming in China

In October 2008 nine farrow-to-finish pig farms were visited in Shuangliu County in Sichuan Province, China. One farm was empty for one month but not cleaned after depopulation. Dust samples were collected at each farm and analysed for the presence of methicillin-resistant Staphylococcus aureus (MRSA). Dust samples from four farms were also analysed for the presence of methicillin-susceptible S. aureus (MSSA). On 5/9 farms MRSA was isolated and on 2/4 farms MSSA was isolated. On two farms, including the empty farm, no MRSA or MSSA could be detected. All MRSA isolates (n=43) belonged to spa type t899. MSSA isolates belonged to spa type t899 (n=12) and spa type t034 (n=2). From 4/9 farms the MRSA isolates of spa type t899 were assigned to multilocus sequence type (MLST) ST9 whereas on one farm the MRSA spa type t899 isolates belonged to a single locus variant of MLST ST9 (ST1376). MSSA isolates with spa type t899 belonged to MLST ST9 and the MSSA with spa type t034 belonged to MLST ST398. This is the first report on MRSA in pig farms in China and the first time that MRSA ST9 and a single locus variant of ST9 are detected in pig farms. This study shows that livestock associated MRSA is not restricted to clonal lineage ST398 as found in Europe and Northern America in commercial pigs but that other MRSA lineages are able to spread in livestock as well. The study confirms that livestock may act as a reservoir for MRSA.

Occurrence of methicillin-resistant Staphylococcus aureus in rats living on pig farms

In The Netherlands, MRSA ST398 has emerged in hospitals and human carriers have been associated with exposure to pigs and cattle. High prevalences of MRSA ST398 in pigs and pig farmers have been determined and the transmission routes of MRSA on pig farms need to be elucidated. In the south of the Netherlands, in recent years, the black rat (Rattus rattus) has emerged as a prominent rodent on livestock farms. From March till May 2008, a survey on MRSA in rats living on livestock farms in the south of The Netherlands and the north of Belgium was conducted. In total, 40 black rats (R. rattus) and 3 brown rats (Rattus norvegicus) were collected on 12 farms including five pig farms, five poultry farms, one mixed pig and veal farm and one goat farm. MRSA ST398 was detected in black rats captured at two of the five pig farms as well as in a black rat living on the mixed pig and veal farm. From one black rat captured at another pig farm MRSA ST 97 was isolated. Considering the behaviour of rats on livestock farms, it is concluded that rats might play a role in the spread and persistence of MRSA on pig farms.

PFGE diversity within the methicillin-resistant Staphylococcus aureus clonal lineage ST398.

BackgroundLivestock has recently been identified as a new reservoir of methicillin-resistant Staphylococcus aureus (MRSA). Most isolates belong to ST398 and are non-typeable with PFGE using Sma I, making it difficult to study transmission and outbreaks. Therefore, a new PFGE using Cfr 9I, a neoschizomer of Sma I was optimized and evaluated to investigate ST398 isolates.ResultsAfter optimizing and evaluating the Cfr 9I PFGE, clear and reproducible banding patterns were obtained from all previously non-typeable MRSA (NTSma I-MRSA) isolates. The PFGE patterns of ST398 isolates showed more diversity than with spa-typing and/or MLST. The PFGE results showed diversity within and between the two most prevalent spa-types of NTSma I-MRSA (t011 and t108). No match was found, when comparing banding patterns of the NTSma I-MRSA with 700 different PFGE types, obtained with Sma I digestion, in our database of more than 4000 strains. Furthermore, possible transmission among veterinarians and their family members was investigated and an outbreak of ST398 MRSA in a residential care facility was confirmed with the Cfr 9I PFGE.ConclusionsThe adjusted PFGE can be used as a method for selecting important and distinct ST398 isolates for further research. The adjustments in the PFGE protocol using Cfr 9I are easy to implement to study the ST398 clonal lineage in laboratories which already have a PFGE facility.

Dynamics and Determinants of Staphylococcus aureus Carriage in Livestock Veterinarians: A Prospective Cohort Study

BACKGROUND Since 2003, a new clade of methicillin-resistant Staphylococcus aureus (MRSA) belonging to clonal complex (CC) 398 and associated with animal husbandry has emerged in the Netherlands. The purpose of this study was to determine the dynamics of carriage in persons with direct contact to livestock. METHODS A 2-year prospective cohort study was performed in which the anterior nares and oropharynx of 137 livestock veterinarians were sampled for the presence of S. aureus every 4 months during the first year and again 1 year later. All S. aureus isolates were genotyped by staphylococcal protein A (spa) typing and with multilocus variable-number tandem repeat analysis (MLVA). RESULTS The mean prevalence of MRSA CC398 carriage was 44% (range, 42%-46%), and for S. aureus the prevalence was 72% (range, 69%-75%). Thirty-two veterinarians (23%) were always carrying MRSA CC398 and 18 of those (56%, 13% of all veterinarians) had identical MLVA types at all sampling moments. CONCLUSIONS A high proportion of veterinarians had persistent MRSA CC398 carriage during the 2-year study period, indicating that this variant may colonize humans for prolonged periods. Furthermore, prevalence of S. aureus carriage was extremely high, indicating that MRSA CC398 is not replacing the susceptible strains, but comes on top of it.

Meticillin-resistant Staphylococcus aureus epidemiology and transmission in a Dutch hospital

The application of the search and destroy (S&D) policy in Scandinavian and Dutch hospitals is associated with low rates of meticillin-resistant Staphylococcus aureus (MRSA). The objective of our study was to describe the MRSA epidemiology and transmission in a Dutch hospital. This descriptive study was performed in a teaching hospital with approximately 40,000 admissions per year. In this hospital the MRSA S&D policy has been applied for several decades. MRSA epidemiology was studied during the years 2001 to 2006. The transmission rate in this hospital was determined using (1) patients history, (2) relation in time and place to other patients or healthcare workers (HCWs), and (3) molecular typing (pulsed-field gel electrophoresis and Spa). Ninety-five persons were identified as MRSA carriers, namely 82 patients and 13 HCWs. The annual MRSA incidence increased more than three-fold during the study period, which was entirely caused by animal-related MRSA. Twenty-three percent of the patients acquired MRSA in a foreign hospital, 26% via animals, 16% by nosocomial transmission, 4% in another Dutch healthcare institution, 10% in the community via a known MRSA-positive person, and in 22% the source was unknown. For HCWs, 69% of MRSA was due to nosocomial transmission, 15% was related to working in a foreign hospital and in 15% HCWs became colonised via an MRSA-positive partner or relative. The transmission rate of 0.30 (22 secondary cases from 73 index cases) indicates that the spread of MRSA was under control during the study period, and so the S&D policy should be continued.

Outbreak of Methicillin-Resistant Staphylococcus aureus ST398 in a Dutch Nursing Home

We describe an outbreak of methicillin-resistant Staphylococcus aureus (MRSA) ST398 in a nursing home in the Netherlands. Seven residents and 4 healthcare workers were identified with MRSA ST398, but 2 of the healthcare workers carried other strains. This study demonstrates that MRSA ST398 can spread in nursing homes.

Skin Lesions in European Hibernating Bats Associated with Geomyces destructans, the Etiologic Agent of White-Nose Syndrome

White-nose syndrome (WNS) has claimed the lives of millions of hibernating insectivorous bats in North America. Its etiologic agent, the psychrophilic fungus Geomyces destructans, causes skin lesions that are the hallmark of the disease. The fungal infection is characterized by a white powdery growth on muzzle, ears and wing membranes. While WNS may threaten some species of North American bats with regional extinction, infection in hibernating bats in Europe seems not to be associated with significant mortality. We performed histopathological investigations on biopsy samples of 11 hibernating European bats, originating from 4 different countries, colonized by G. destructans. One additional bat was euthanized to allow thorough examination of multiple strips of its wing membranes. Molecular analyses of touch imprints, swabs and skin samples confirmed that fungal structures were G. destructans. Additionally, archived field notes on hibernacula monitoring data in the Harz Mountains, Germany, over an 11-year period (2000–2011) revealed multiple capture-recapture events of 8 banded bats repeatedly displaying characteristic fungal colonization. Skin lesions of G. destructans-affected hibernating European bats are intriguingly similar to the epidermal lesions described in North American bats. Nevertheless, deep invasion of fungal hyphae into the dermal connective tissue with resulting ulceration like in North American bats was not observed in the biopsy samples of European bats; all lesions found were restricted to the layers of the epidermis and its adnexae. Two bats had mild epidermal cupping erosions as described for North American bats. The possible mechanisms for any difference in outcomes of G. destructans infection in European and North American bats still need to be elucidated.