Low prevalence of methicillin-resistant Staphylococcus aureus among men who have sex with men attending an STI clinic in Amsterdam: a cross-sectional study.

OBJECTIVE: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is common among men who have sex with men (MSM) in the USA. It is unknown whether this is also the case in Amsterdam, the Netherlands. DESIGN: Cross-sectional study. SETTING: Sexually transmitted infection outpatient low-threshold clinic, Amsterdam, the Netherlands. PARTICIPANTS: Between October 2008 and April 2010, a total of 211 men were included, in two groups: (1) 74 MSM with clinical signs of a skin or soft tissue infection (symptomatic group) and (2) 137 MSM without clinical signs of such infections (asymptomatic group). PRIMARY OUTCOME MEASURES: S aureus and MRSA infection and/or colonisation. Swabs were collected from the anterior nasal cavity, throat, perineum, penile glans and, if present, from infected skin lesions. Culture for S aureus was carried out on blood agar plates and for MRSA on selective chromagar plates after enrichment in broth. If MRSA was found, the spa-gene was sequenced. SECONDARY OUTCOME MEASURES: Associated demographic characteristics, medical history, risk factors for colonisation with S aureus and high-risk sexual behaviour were collected through a self-completed questionnaire. RESULTS: The prevalence of S aureus colonisation in the nares was 37%, the pharynx 11%, the perianal region 12%, the glans penis 10% and in skin lesions 40%. In multivariable analysis adjusting for age, anogenital S aureus colonisation was significantly associated with the symptomatic group (p=0.01) and marginally with HIV (p=0.06). MRSA was diagnosed in two cases: prevalence 0.9% (95% CI 0.1% to 3.4%)). Neither had CA-MRSA strains. CONCLUSIONS: CA-MRSA among MSM in Amsterdam is rare. Genital colonisation of S aureus is not associated with high-risk sexual behaviour.

Is living in a border region a risk for a high prevalence of resistance?

This study assessed the antimicrobial resistance and population structure of Staphylococcus aureus isolated from general practice (GP) patients and nursing home (NH) residents in the province of Limburg (near the border with Germany and Belgium) in comparison with those obtained in the remaining provinces of the Netherlands. A total of 617 and 418 S. aureus isolates were isolated from 2,691 to 1,351 nasal swabs from GP patients and NH residents, respectively. Quantitative antibiotic susceptibility testing was performed using a microbroth dilution method. Putative methicillin-resistant S. aureus (MRSA) isolates were tested for the presence of the mecA gene and spa typing was performed on all S. aureus isolates. No significant differences in the prevalence of resistance were found between the two groups of GP isolates, but the isolates from the NH residents showed a lower resistance for trimethoprim-sulfamethoxazole (p = 0.003) in Limburg province compared with the remaining provinces in the Netherlands. Among the isolates from NH residents in Limburg province, the prevalence of spa-CC 084 was higher (p = 0.003) and that of spa-CC 002 was lower (p = 0.01) compared with isolates from NHs in the remaining provinces of the Netherlands. We observed no differences in resistance and population structure between S. aureus isolates from GP patients in Limburg and the remaining provinces of the Netherlands, and only a few differences were observed between the NH populations. There was no higher prevalence of resistance among the GP and NH isolates from Limburg compared with the remaining provinces.

Real-time PCR to distinguish livestock-associated (ST398) from non-livestock-associated (methicillin-resistant) Staphylococcus aureus.

BACKGROUND: The Netherlands is one of the most densely populated countries in the world, with extensive livestock of pigs. In 2005, the emergence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) was a fact, with a relatively high MRSA colonisation among pig farmers. These MRSA isolates mostly belonged to sequence type 398 (ST398). Compared to hospital-associated MRSA (HA-MRSA), severe infections due to LA-MRSA and transmission between individuals are still relatively rare. Therefore, LA-MRSA may warrant less stringent containment measures than HA-MRSA in hospital settings. RESULTS: The aim of this study was to develop a rapid diagnostic tool to distinguish LA-MRSA from non-LA-MRSA in aid of infection control. Here, we show that ST398 strains can be readily detected with real-time polymerase chain reaction (PCR). Analysis of a large panel of related and unrelated microorganisms confirmed that the real-time ST398 PCR (ST398-qPCR) assay does not cross-react with other microorganisms or with non-LA-S. aureus strains. ST398-qPCR analysis of MRSA isolates collected in 2010, 2011 and 2012 at the Jeroen Bosch Hospital (n = 275) showed that an average of 78 % of MRSA belonged to sequence type ST398. CONCLUSION: We conclude that the ST398 real-time PCR is a reliable assay to detect LA-S. aureus and anticipate that the use of this assay can prevent the unnecessary closing of hospital wards, which may lead to substantial savings for the health care system.

Multi-centre evaluation of a phenotypic extended spectrum ß-lactamase detection guideline in the routine setting.

This study aimed to evaluate the routine setting performance of a guideline for phenotypic detection of extended spectrum ß-lactamases (ESBLs) in Enterobacteriaceae, recommending ESBL confirmation with Etest or combination disc for isolates with a positive ESBL screen test (i.e. cefotaxime and/or ceftazidime MIC >1 mg/L or an automated system ESBL warning). Twenty laboratories submitted 443 Enterobacteriaceae with a positive ESBL screen test and their confirmation test result (74%Escherichia coli, 12%Enterobacter cloacae, 8%Klebsiella pneumoniae, 3%Proteus mirabilis, 2%Klebsiella oxytoca). Presence of ESBL genes was used as reference test. Accuracy of local phenotypic ESBL detection was 88%. The positive predictive value (PPV) of local screen tests was 70%, and differed per method (Vitek-2: 69%, Phoenix: 68%, disc diffusion: 92%), and species (95%K. pneumoniae-27%K. oxytoca). A low PPV (3%) was observed for isolates with automated system alarm but third-generation cephalosporin MICs <2 mg/L. Local ESBL confirmation had a PPV and negative predictive value (NPV) of 93% and 90%, respectively. Compared with centrally performed confirmation tests, 7% of local tests were misinterpreted. Combination disc was more specific than Etest (91% versus 61%). Confirmation tests were not reliable for P. mirabilis and K. oxytoca (PPV 33% and 38%, respectively, although NPVs were 100%). In conclusion, performance of Etests could be enhanced by education of technicians to improve their interpretation, by genotypic ESBL confirmation of P. mirabilis and K. oxytoca isolates with positive phenotypic ESBL confirmation, and by interpreting isolates with a positive ESBL alarm but an MIC <2 mg/L for cefotaxime and ceftazidime as ESBL-negative.

Staphylococcus aureus (MSSA) and MRSA (CC398) isolated from post-mortem samples from pigs.

There are many reports on the occurrence of Livestock Associated Methicilline resistant Staphylococcus aureus (LA-MRSA, CC398) in healthy pigs. There are however, very few reports of LA-MRSA being associated with pathological lesions in pigs. With this study we try to find the answers to the questions: (1) how often is S. aureus found in post-mortem material from pigs, (2) how many of these isolates are methicillin resistant, (3) are these equally distributed over the years? Here we report the isolation of MRSA and of methicillin sensitive S. aureus (MSSA) from samples derived from post-mortem examinations at the Animal Health Service in The Netherlands in the period from 2003 through October 2008. The MSSA and MRSA described here were isolated from 159 pathological lesions and from 7 submissions of aborted foetuses derived from a total of 116 animals, representing 103 submissions coming from 92 different herds. This is approximately 0.5% of all pigs submitted for post mortem examination in those years. The proportion of pigs from which S. aureus (both MSSA and MRSA) was isolated from, did not increase over the years. MSSA (N=97) and LA-MRSA CC398 (N=18) were present mainly in (peri)arthritis in over 30% of all cases, but were also isolated from internal organs such as lung, brain, spleen, kidneys, heart, indicating septicaemia. Remarkably, one non-CC398 MRSA (ST1) was isolated in a joint and a kidney of one pig. This isolate was resistant to 5 out of 6 antimicrobials tested. There was no significant difference in the type of lesions in which LA-MRSA was found compared to MSSA. The number of antimicrobials these isolates were resistant to, increased rapidly after 2004. LA-MRSA was isolated for the first time in 2005 and then again in 2007 and 2008, suggesting that this is an emerging pathogen. However, due to changes in the panel of antimicrobials used to test S. aureus for antimicrobial susceptibility in 2005 and 2007, the possibility exists that we may have missed some MRSA isolates. LA-MRSA isolates are resistant to at least three but sometimes five out of six antimicrobials tested. All isolates were susceptible to the combination of Trimethoprim/Sulfamethaxol.

Emergence of MRSA of unknown origin in the Netherlands.

The Netherlands is known for its low methicillin-resistant Staphylococcus aureus (MRSA) prevalence. Yet MRSA with no link to established Dutch risk factors for acquisition, MRSA of unknown origin (MUO), has now emerged and hampers early detection and control by active screening upon hospital admittance. We assessed the magnitude of the problem and determined the differences between MUO and MRSA of known origin (MKO) for CC398 and non-CC398. National MRSA Surveillance data (2008-2009) were analysed for epidemiological determinants and genotypic characteristics (Panton-Valentine leukocidin, spa). A quarter (24%) of the 5545 MRSA isolates registered were MUO, i.e. not from defined risk groups. There are two genotypic MUO groups: CC398 MUO (352; 26%) and non-CC398 MUO (998; 74%). CC398 MUO needs further investigation because it could suggest spread, not by direct contact with livestock (pigs, veal calves), but through the community. Non-CC398 MUO is less likely to be from a nursing home than non-CC398 MKO (relative risk 0.55; 95% CI 0.42-0.72) and Panton-Valentine leukocidin positivity was more frequent in non-CC398 MUO than MKO (relative risk 1.19; 95% CI 1.11-1.29). Exact transmission routes and risk factors for non-CC398 as CC398 MUO remain undefined.

First report of a brain abscess caused by Nocardia veterana.

Among Nocardia species causing infections, Nocardia veterana is rarely isolated and is mostly described as causing pulmonary infections. This is the first presentation of a case of brain abscess attributable to an N. veterana infection in a patient with type 2 diabetes. Prolonged antibiotic therapy with trimethoprim-sulfamethoxazole led to successful clinical recovery.

Prevalence of methicillin-resistant Staphylococcus aureus and risk factors for carriage in Dutch hospitals.

To evaluate the actual burden of methicillin-resistant Staphylococcus aureus and determine risk factors for carriage and infection, we performed a prevalence survey with a nested case-control study among inpatients in Dutch hospitals. The prevalence of carriage was 0.94 cases per 1,000 inpatients, and the prevalence of infection was 0.21 cases per 1,000 inpatients. Professional contact with livestock and a stay in a foreign hospital were associated with carriage.

Increasing trend in gonococcal resistance to ciprofloxacin in The Netherlands, 2006-8.

INTRODUCTION: Rapid development of Neisseria gonorrhoeae resistance to several antibiotics in recent years threatens treatment and prevention. Targeted surveillance of new resistance patterns and insight into networks and determinants are essential to control this trend. METHODS: Since the Gonococcal Resistance to Antimicrobials Surveillance (GRAS) project was implemented within the Dutch national sexually transmitted infection (STI) surveillance network in July 2006, participating STI centres have collected a culture from each gonorrhoea patient. Isolates were tested for susceptibility to penicillin, tetracycline, ciprofloxacin and cefotaxime using Etest. Logistic regression was used to determine risk factors for ciprofloxacin resistance. RESULTS: Between July 2006 and July 2008, prevalence of resistance to penicillin was 10%, to tetracycline 22% and to ciprofloxacin 42%. Resistance to cefotaxime was not found, although minimum inhibitory concentrations higher than 0.125 mg/l drifted upward (p<0.05). Ciprofloxacin resistance rose from 35% in 2006 to 46% in 2008 (p<0.05), despite 2003 guidelines naming cefotaxime as first-choice therapy. In men, ciprofloxacin resistance was higher in men having sex with men (MSM) than in heterosexual men (adjusted OR 2.0, 95% CI : 1.5 to 2.6). In women, it was higher in commercial sex workers (adjusted OR 25.0, 95% CI 7.7 to 78.2) and women aged over 35 years (adjusted OR 8.2, 95% CI 3.0 to 22.7) than in other women. CONCLUSION: Ciprofloxacin resistance in The Netherlands is increasing, and is particularly found in MSM, older women, and female sex workers. No resistance to current first-choice therapy was found, but alertness to potential clinical failures is essential. By merging epidemiological and microbiological data in GRAS, specific high-risk transmission groups can be identified and policy adjusted when needed.

Methicillin-resistant Staphylococcus aureus in horses and horse personnel: an investigation of several outbreaks.

At the Veterinary Microbiological Diagnostic Center, the Netherlands, the percentage of methicillin-resistant Staphylococcus aureus (MRSA) isolates found in equine clinical samples increased from 0% in 2002 to 37% in 2008. MRSA of spa-type t064, belonging to MLST ST8 and spa-types t011 and t2123, both belonging to the livestock-associated MLST ST398, predominated. During an outbreak of post-surgical MRSA infections in horses at a veterinary teaching hospital in 2006/2007, MRSA isolates of spa-type t2123 were cultured from 7 horses and 4/61 personnel which indicated zoonotic transmission. After intervention the outbreak stopped. However, another outbreak occurred in 2008, where 17 equine MRSA isolates of spa-type t011 (n=12), t2123 (n=4), and t064 (n=1) were found. This time, 16/170 personnel were positive for MRSA with spa-type t011 (n=11) and t2123 (n=5). Personnel in close contact with horses were more often MRSA-positive (15/106) than those without (1/64). Screening of horses upon admission showed that 9.3% were MRSA-positive predominantly with spa-type t011. Weekly cross-sectional sampling of all hospitalized horses for 5 weeks showed that 42% of the horses were MRSA-positive at least once, again predominantly with spa-type t011, which suggests that nosocomial transmission took place. Fifty-three percent of the environmental samples were MRSA-positive, including samples from students' and staff members' rooms, and all were spa-type t011. This indicates that humans contribute to spreading the organism. Culturing of samples employing high-salt pre-enrichment performed better than a comparable method without pre-enrichment. Our results show that nosocomial transmission occurs in equine clinics and suggests that personnel play a role in the transmission.

Short term micro-evolution and PCR-detection of methicillin-resistant and -susceptible Staphylococcus aureus sequence type 398.

Micro-evolutionary analysis of 70 ST398 isolates by pulsed-field gel electrophoresis (PFGE) using Cfr9I revealed three sub-clones with abundant inter- and intra-sub-clone heterogeneity in spa- and SCCmec-types. In addition, we developed two specific PCRs for the detection of Staphylococcus aureus sequence type 398 (ST 398) isolates with 100% specificity and high sensitivity.

Molecular characterisation of PFGE non-typable methicillin-resistant Staphylococcus aureus in The Netherlands, 2007.

In 2007 in The Netherlands, 30% of all human isolates of methicillin-resistant Staphylococcus aureus (MRSA) sent to the National Institute for Public Health and the Environment could not be typed by pulsed-field gel electrophoresis (non-typable (NT)-MRSA). Molecular characterisation of the NT-MRSA isolates revealed 27 different spa types and two distinct SCCmec types, type IV and V. All NT-MRSA isolates were closely related based on spa and multi-locus sequence typing and belonged to the ST398 lineage. The rapid increase of NT-MRSA (ST398) isolates over the last years shows the importance of this relatively new clonal lineage.

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 ST's, 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.

An outbreak of non-typeable MRSA within a residential care facility.

In a household setting within a residential care facility for visually and intellectually disabled people, a resident (index case) was diagnosed with dermal abscesses caused by a methicillin-resistant Staphylococcus aureus (MRSA) which was non-typeable by standard pulsed-field gel electrophoresis. All residents and staff in contact with the index case (a total of 200 people) were screened for MRSA.

Experiences with the Dutch Working Party on antibiotic policy (SWAB).

The Dutch Working Party on Antibiotic Policy (Stichting Werkgroep AntibioticaBeleid, SWAB) was founded in 1996 as an initiative of the Society for Infectious Diseases, the Dutch Society for Medical Microbiology, and the Dutch Association of Hospital Pharmacists. Its primary goal is to contribute to the containment of antimicrobial resistance and the expanding costs incurred for the use of antibiotics. SWAB is the Intersectoral Coordinating Mechanism (ICM) for the Netherlands, and it is at present the National Antimicrobial Resistance (AMR) Focal Point. It coordinates the national surveillance of antibiotic resistance, in collaboration with the National Institute for Public Health and the Environment(RIVM), coordinates the surveillance of the use of antibiotics,and runs a guideline development programme. Information about consumption of antimicrobial agents and antimicrobial resistance among medically important bacteria is presented annually in NethMap. Over the past decade, outpatient consumption of antibiotics has risen only slightly, but in the hospital setting there was an overall significant increase in antibiotic use, due mainly to the steady reduction in the average length of patient hospital stays. In 2006 we introduced our electronic national antibiotic guide 'SWAB-ID' for the antibiotic treatment and prophylaxis of common infectious diseases in hospitals.

Transmission of methicillin-resistant Staphylococcus aureus strains between different kinds of pig farms.

The main objective of the present study was to investigate if different kinds of pig farms, like farrowing farms and rearing farms, play a role in the transmission of methicillin-resistant Staphylococcus aureus (MRSA) to Dutch finishing farms. Twelve farrowing farms, 11 finishing farms, 6 farrow-to finish farms, 1 rearing farm and 1 centre for artificial insemination were included. Screening of 310 pigs from these 31 farms showed 35 pigs (11%) to carry MRSA in their nares. On 7 of the 31 (23%) investigated farms colonized pigs were found, including 3 finishing farms, 3 farrowing farms and 1 farrow-to-finish farm. The use of standard antimicrobial medication of the pigs seemed to be a risk factor for MRSA carriage. Screening of the pigs on six farms supplying pigs for the MRSA positive farms revealed that the pigs on all but one farm were MRSA positive. Genotyping revealed that all MRSA strains were non-typeable by PFGE using the SmaI restriction enzyme and had multilocus sequence type (MLST) ST398. Different spa-types were found including t011, t108, t567, t899 and t1939, but the spa-types on epidemiologically related farms were identical indicating that MRSA are transmitted between farms through the purchase of colonized pigs. Two SCCmec types were found among the MRSA: type IV and type V. SCCmec type V was predominant. On two farms MRSA isolates with ST398, the same spa-type but with different SCCmec types (IV and V) were found, suggesting that different SCCmec elements have been inserted into MSSA with the same genotype. All MRSA strains were resistant to tetracycline, but additional resistances to erythromycin, lincomycin, kanamycin and gentamicin were also found. All MRSA isolates were negative for the exfoliative toxin genes (eta and etb), PVL toxin genes (lukF and lukS), toxic shock syndrome gene (tst-1), and the leukotoxin genes (lukE, lukD, lukM, lukF').

High prevalence of methicillin resistant Staphylococcus aureus in pigs.

Recently methicillin resistant Staphylococcus aureus (MRSA) was isolated from pigs and pig farmers in The Netherlands. In order to assess the dissemination of MRSA in the Dutch pig population, we screened 540 pigs in 9 slaughterhouses, where a representative portion of Dutch pigs (63%) was slaughtered in 2005. We found 209 (39%) of the pigs to carry MRSA in their nares. Forty-four of 54 groups of 10 consecutive pigs (81%), each group from a different farm, and all slaughterhouses were affected. All MRSA isolates belonged to 1 clonal group, showing Multi-Locus Sequence Type 398 and closely related spa types (mainly t011, t108 and t1254). Three types of the Staphylococcal Chromosome Cassette (SCCmec) were found: III (3%), IVa (39%) and V (57%). All 44 tested isolates (1 isolate per group) were resistant to tetracycline, reflecting the high and predominant use of tetracyclines in pig husbandry. Twenty-three percent of the isolates were resistant to both erythromycin and clindamycin and 36% to kanamycin, gentamicin and tobramycin but only a single isolate was resistant to co-trimoxazole and none to ciprofloxacin and several other antibiotics. The percentage of MRSA positive pigs was significantly different among slaughterhouses and among groups within slaughterhouses, indicating a high prevalence of MRSA in pigs delivered from the farms as well as cross contamination in the slaughterhouses.

Multiple cases of familial transmission of community-acquired methicillin-resistant Staphylococcus aureus.

The worldwide emergence of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) can have severe public health implications. Familial transmissions of CA-MRSA in The Netherlands were investigated. Among the families studied, two clusters of CA-MRSA could be identified. This report demonstrates that family members can serve as reservoirs of CA-MRSA which may become a serious problem in containing the spread of MRSA.

Emergence of virulent methicillin-resistant Staphylococcus aureus strains carrying Panton-Valentine leucocidin genes in The Netherlands.

Methicillin-resistant Staphylococcus aureus (MRSA) strains carrying the Panton-Valentine leucocidin (PVL) genes have been reported worldwide and are a serious threat to public health. The PVL genes encode a highly potent toxin which is involved in severe skin infections and necrotizing pneumonia, even in previously healthy individuals. We assessed the prevalence of PVL-positive MRSA in The Netherlands for two periods of time: (i) 1987 through 1995 and (ii) 2000 and 2002, and determined their characteristics by using multilocus sequence typing and staphylococcal chromosome cassette (SCCmec) typing. It was found that up to 15% of all MRSA isolates detected in The Netherlands harbored the PVL genes. Most PVL-positive MRSA isolates were obtained from severe soft tissue infections in relatively young individuals. The first PVL-positive MRSA described in The Netherlands, isolated in 1988, was a single-locus variant of the "Berlin" epidemic MRSA clone. The 20 PVL-positive MRSA isolates studied in 2000 and 2002 consisted of five different sequence types (STs) that belonged to four clonal complexes. One of the STs, ST80, is considered to be a widespread European clone and was the most predominant ST (60%) in this study, while ST37 had never been found to be associated with PVL-positive MRSA. Most isolates harbored SCCmec type IV, a supposed marker for community-acquired MRSA. The number and type of virulence-associated genes varied among the different STs.

Panton-Valentine leukocidin positive MRSA in 2003: the Dutch situation.

Analysis of methicillin-resistant Staphylococcus aureus (MRSA) isolates in the Netherlands in 2003 revealed that 8% of the hospital isolates carried the loci for Panton-Valentine leukocidin (PVL). Molecular subtyping showed that most Dutch PVL-MRSA genotypes corresponded to well-documented global epidemic types. The most common PVL-MRSA genotypes were sequence type ST8, ST22, ST30, ST59 and ST80. MRSA with ST8 increased in the Netherlands from 1% in 2002 to 17% in 2003. It is emphasised that PVL-MRSA might not only emerge in the community, but also in the hospital environment.