Macrorestriction Analysis Research Articles

Occurrence and characteristics of Listeria monocytogenes in ready-to-eat food from retail market in the Czech Republic

The study objectives were to test ready-to-eat food from the retail market in the Czech Republic for the presence of L. monocytogenes and, based on typing methods, to investigate probable causes of contamination. A total 2180 samples of ready-to-eat food (meat, dairy, fish, delicatessen and confectionery products and fresh fruit and vegetables) were analysed qualitatively and quantitatively. L. monocytogenes isolates were characterised by serotyping and macrorestriction analysis after digestion with the restriction enzyme AscI. In 2004–2008 L. monocytogenes was most often detected in delicatessen (5.2%), meat (3.4%) and dairy products (1.8%). In the analysed samples, L. monocytogenes was mostly present at counts lower than 102 CFU/g. Only in 2004, higher counts of L. monocytogenes were found in two heat-processed meat products (103 CFU/g). The obtained macrorestriction patterns helped in tracing the source of contamination and routes of the spread of L. monocytogenes in the manufacturing plant and retail market.

PCR melting profile (PCR MP) - a new tool for differentiation of Candida albicansstrains

BackgroundWe have previously reported the use of PCR Melting Profile (PCR MP) technique based on using low denaturation temperatures during ligation mediated PCR (LM PCR) for bacterial strain differentiation. The aim of the current study was to evaluate this method for intra-species differentiation of Candida albicans strains.MethodsIn total 123 Candida albicans strains (including 7 reference, 11 clinical unrelated, and 105 isolates from patients of two hospitals in Poland) were examined using three genotyping methods: PCR MP, macrorestriction analysis of the chromosomal DNA by pulsed-field gel electrophoresis (REA-PFGE) and RAPD techniques.ResultsThe genotyping results of the PCR MP were compared with results from REA-PFGE and RAPD techniques giving 27, 26 and 25 unique types, respectively. The results showed that the PCR MP technique has at least the same discriminatory power as REA-PFGE and RAPD.ConclusionData presented here show for the first time the evaluation of PCR MP technique for candidial strains differentiation and we propose that this can be used as a relatively simple and cheap technique for epidemiological studies in short period of time in hospital.

Salmonella enterica subspecies enterica serovar Choleraesuis in a wild boar population in Germany

Salmonella (S.) enterica subspecies enterica serovar Choleraesuis, the swine-adapted serovar is found rarely in Western European countries including Germany. However, the regional laboratory of the federal state Thuringia in Germany examined diseased wild boars routinely also for the occurrence of Salmonella organisms. Between 2006 and 2008, only the serovar S. Choleraesuis was islolated from 24 animals, three strains isolated from domestic pigs were included. In order to detect a possible epidemiological context, the strains of S. Choleraesuis were characterised by macrorestriction and plasmid analysis, repetitive sequence PCR, antimicrobial testing and determining the biochemical profile. A combination of all methods enabled the identification of five epidemiological groups. Two groups were detected in the same territory but three other discriminative groups were predominant in different regions. S. Choleraesuis strains of the different epidemiological groups circulate in wild boar populations in the corresponding regions. However, it could be concluded that both natural barriers like mountains and artificial barriers like arterial roads may cause the separation of wild boar populations and as a result also the respective S. Choleraesuis organisms. The occurrence of the identical epidemiological groups in wild boars and domestic pigs indicates the possible mutual exposure of the pathogen. To avoid risks for human and domestic pig health regular inspection of meat from wildlife by official veterinarians and advice of hunters and persons who prepare and consume wild boar meat should be enhanced.

Typing of Campylobacter jejuni Isolates from Dogs by Use of Multilocus Sequence Typing and Pulsed-Field Gel Electrophoresis

Campylobacter is a major cause of human gastroenteritis worldwide. Risk of Campylobacter infection in humans has been associated with many sources, including dogs. This study aimed to investigate whether C. jejuni carried by dogs could potentially be a zoonotic risk for humans and if there were common sources of C. jejuni infection for both humans and dogs. Multilocus sequence typing (MLST) together with macrorestriction analysis of genomic DNA using SmaI and pulsed-field gel electrophoresis (PFGE) were both used to analyze 33 C. jejuni isolates obtained from various dog populations, including those visiting veterinary practices and from different types of kennels. MLST data suggested that there was a large amount of genetic diversity between dog isolates and that the majority of sequence types found in isolates from these dogs were the same as those found in isolates from humans. The main exception was ST-2772, which was isolated from four samples and could not be assigned to a clonal complex. The most commonly identified clonal complex was ST-45 (11 isolates), followed by ST-21 (4 isolates), ST-508 (4 isolates), and ST-403 (3 isolates). The profiles obtained by macrorestriction PFGE were largely in concordance with the MLST results, with a similar amount of genetic diversity found. The diversity of sequence types found within dogs suggests they are exposed to various sources of C. jejuni infection. The similarity of these sequence types to C. jejuni isolates from humans suggests there may be common sources of infection for both dogs and humans. Although only a small number of household dogs may carry C. jejuni, infected dogs should still be considered a potential zoonotic risk to humans, particularly if the dogs originate from kennelled or hunt kennel dog populations, where the prevalence may be higher.

Analysis of Slovenian MRSA strains with susceptibility patterns suggestive of CA-MRSA

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) differs from healthcare-associated MRSA (HA-MRSA) in its molecular and microbiological characteristics. Six Slovenian regional public health institutes and the National Institute of Public Health took part in monitoring CA-MRSA infections. S. aureus isolates resistant to oxacillin and susceptible to > or = two of the four antibiotics ciprofloxacin, erythromycin, clindamycin or gentamicin were defined as CA-MRSA and further analyzed. The presence of the gene for Panton-Valentine leukocidin (PVL) was confirmed using PCR, the type of staphylococcal cassette chromosome (SCCmec) using multiplex PCR, and macrorestriction analysis of chromosomal DNA using pulsed-field gel electrophoresis (PFGE). A total of 31 strains from 31 patients were analyzed during a period of 21 months: 23 specimens were sent from hospitals, six from primary care, two from a long-term care facility. All 31 isolates contained the gene mecA. Sixteen (51.6%) isolates were identified as SCCmec type IV, three isolates were PVL positive. Using PFGE, the CA-MRSA strains were classified into 15 similarity groups. Results of antibiotic susceptibility showed there were five resistance types among the 31 strains. Simultaneous resistance against ciprofloxacin and gentamicin was often associated with the presence of SCCmec type I, strongly resembling HA-MRSA. PVL-positive strains of CA-MRSA have been isolated in Slovenia only rarely. We will continue to monitor strains of MRSA in order to obtain the complete microbiological and epidemiological features.

Molecular Characterization ofStenotrophomonas maltophiliaIsolates from Cystic Fibrosis Patients and the Hospital Environment

To ascertain whether cystic fibrosis (CF) patients are colonized or infected with unique or multiple strains of Stenotrophomonas maltophilia; to understand whether some strains colonize or infect more than 1 patient, indicating clonal spread; and to explore the molecular heterogeneity of hospital water isolates and their correlation with clinical isolates. The regional CF center of Policlinico "Umberto I" of Rome, Italy. The study was carried out on a random sample of S. maltophilia isolates (n = 110) collected from CF patients (n = 50) during the period 2002-2005 and on 24 water isolates obtained during a monitoring program in the first 6 months of 2005. Home environmental samplings were not performed. All isolates, which were recovered from cultures of specimens obtained in both inpatient and outpatient settings, were genotyped with DNA macrorestriction analysis with the restriction enzyme XbaI and pulsed-field gel electrophoresis. One-third of the patients with repeated episodes of S. maltophilia infection or colonization hosted more than 1 strain. A potential transmission, defined as the isolation of the same strain in 2 or more patients, occurred 5 times, showing a frequency of potential transmission episodes slightly higher than previously reported. Water, taps, and sinks of the different rooms of the CF center tended to be persistently colonized with the same strain of S. maltophilia, with no correlation between clinical and water-associated isolates. The study does not provide sufficient data to conclude definitively that isolation of colonized or infected CF patients and control of hospital water systems contamination would be beneficial infection control measures. Epidemiologic analytical studies that correlate the presence of S. maltophilia with clinical outcomes are strongly needed.

Pseudomonas aeruginosain a neonatal intensive care unit: molecular epidemiology and infection control measures

BackgroundPseudomonas aeruginosa, a non-fermentative, gram-negative rod, is responsible for a wide variety of clinical syndromes in NICU patients, including sepsis, pneumonia, meningitis, diarrhea, conjunctivitis and skin infections. An increased number of infections and colonisations by P. aeruginosa has been observed in the neonatal intensive care unit (NICU) of our university hospital between 2005 and 2007.MethodsHand disinfection compliance before and after an educational programme on hand hygiene was evaluated. Identification of microrganisms was performed using conventional methods. Antibiotic susceptibility was evaluated by MIC microdilution. Genotyping was performed by PFGE analysis.ResultsThe molecular epidemiology of Pseudomonas aeruginosa in the NICU of the Federico II University hospital (Naples, Italy) and the infection control measures adopted to stop the spreading of P. aeruginosa in the ward were described. From July 2005 to June 2007, P. aeruginosa was isolated from 135 neonates and caused severe infections in 11 of them. Macrorestriction analysis of clinical isolates from 90 neonates identified 20 distinct genotypes, one major PFGE type (A) being isolated from 48 patients and responsible for 4 infections in 4 of them, four other distinct recurrent genotypes being isolated in 6 to 4 patients. Seven environmental strains were isolated from the hand of a nurse and from three sinks on two occasions, two of these showing PFGE profiles A and G identical to two clinical isolates responsible for infection. The successful control of the outbreak was achieved through implementation of active surveillance of healthcare-associated infections in the ward together with environmental microbiological sampling and an intense educational programme on hand disinfection among the staff members.ConclusionP. aeruginosa infections in the NICU were caused by the cross-transmission of an epidemic clone in 4 neonates, and by the selection of sporadic clones in 7 others. An infection control programme that included active surveillance and strict adherence to hand disinfection policies was effective in controlling NICU-acquired infections and colonisations caused by P. aeruginosa.

Epidemic multidrug-resistant Acinetobacter baumannii related to European clonal types I and II in Rome (Italy)

The molecular epidemiology and the genetic basis of antibiotic resistance in 88 multidrug-resistant (MDR) Acinetobacter baumannii strains isolated during 18 months from infected patients in seven intensive care units (ICUs) in Rome were investigated. Random amplified polymorphic DNA and macrorestriction analysis identified two predominant clonal types, genetically related to the European epidemic clones I (type 2) and II (type 1), accounting for 98.9% of A. baumannii ICU isolates. Type 1 was isolated from all ICUs under survey. Class 1 integrons of 2.2 and 2.5 kb were detected in type 1 and type 2 isolates, respectively. The integron structures were similar to those previously determined for epidemic A. baumannii strains from various European countries, and suggestive of integron rearrangement/exchange among isolates related to the European epidemic clones I and II. Carbapenem resistance was associated with the presence of the blaOXA-58 gene in type 1 isolates. The results indicate that the A. baumannii type 1 clone has a high potential of spreading among hospitals.

Spread of multidrug-resistant Proteus mirabilis isolates producing an AmpC-type β-lactamase: epidemiology and clinical management

A remarkable increase in Proteus mirabilis strains producing acquired AmpC-type β-lactamases (CBLs) has been observed at Ospedale di Circolo e Fondazione Macchi (Varese, Italy) over the last few years. The epidemiology and treatment outcome of infections associated with this unprecedented spread are reported. From 2004–2006, 2070 P. mirabilis isolates were investigated. Extended-spectrum β-lactamases (ESBLs) and CBL resistance determinants were identified by gene amplification and direct sequencing. Clonal relatedness was evaluated by macrorestriction analysis. Overall, 43 CBL-positive isolates were obtained from hospitalised ( n = 22) and non-hospitalised ( n = 21) patients (median age 78.8 years). The prevalence of CBL-positive isolates increased from 0.3% in 2004 to 4.6% in 2006, whereas that of ESBL-positive isolates remained constant (ca. 10%). CBL-positive isolates were multidrug-resistant and carried the CMY-16 determinant. All but two isolates were genetically identical or closely related. Retrospective analysis of clinical records revealed that the majority of CMY-16-positive isolates were associated with urinary tract infections. Treatment with amikacin or carbapenems was consistently effective, whereas piperacillin/tazobactam produced a clinical response in seven of nine cases. This is the first report of a rapid spread of CBL-positive P. mirabilis strains endowed with remarkable antimicrobial resistance. Practical methods for CBL detection are needed for the appropriate management of related infections.

Clustering and Risk Factors of Methicillin-Resistant Staphylococcus aureus Carriage in Two Italian Long-Term Care Facilities

Methicillin-resistant Staphylococcus aureus (MRSA) is a well-recognized agent of health care-associated infections in long-term care facilities, but few data about the circulation of MRSA in this setting in Italy are available. The aim of the study is to determine the prevalence and risk factors for MRSA carriage in nursing home residents in Vicenza (northeastern Italy). A point prevalence survey was conducted in two long-term care facilities (subdivided into 15 wards) from 12 June 2006 to 6 July 2006. Anterior nasal swabs were obtained from residents and laboratory screening for MRSA was performed; full antibiotic susceptibility was assessed in MRSA isolates. Macrorestriction analysis of chromosomal DNA was carried out by pulsed field gel electrophoresis (PFGE). For each subject, demographic data, length of stay, dependency, cognitive function, presence of medical devices, comorbidities, current and previous antibiotic treatment, previous hospital admission and presence of infection were assessed on the day of sample collection. Factors that were found to be significantly associated with MRSA carriage at univariate analysis were introduced into multilevel logistic regression models in order to estimate the odds ratios (OR) with 95% confidence intervals (CI) for the risk of MRSA colonization, taking into account the clustering of patients within wards. Nasal swabs were obtained in 551 subjects; overall 43 MRSA carriers were detected (7.8%; CI = 5.7-10.4%). The rate of nasal carriers was very similar in the two institutions, and varied from 0% (0/36) to 18% (7/39) between wards. Only two out of 15 wards were found to have no MRSA carriers; overall, three pairs of colonized roommates were detected. Upon multilevel logistic regression, the risk of MRSA carriage was increased in patients with cancer (OR = 6.4; CI = 2.5-16.4), in those that had undergone recent hospitalization (OR = 2.2; CI = 1.0-4.4), and it reached OR = 4.0 (CI = 1.7-9.9) in those with three or more antibiotic treatments in the previous year; about 10% of the variability in MRSA carriage could be attributed to differences between wards. Pulsed field gel electrophoresis analysis permitted the definition of six clusters; two of these comprised 78.6% of the studied isolates and were quite similar, with one being more strongly represented among subjects hospitalized in the previous 12 months. All of the MRSA strains were resistant to ciprofloxacine; nevertheless, the majority were susceptible to most other non-betalactam antibiotics. The study suggests that nursing homes are a significant reservoir for MRSA. Statistical and PFGE analyses indicate a scenario where MRSA seems to be endemic and individual risk factors, namely recent hospitalizations and repeated antibiotic treatments, play a major role in the selection of drug-resistant organisms. Infection control measures should be coordinated among different health care settings, and the appropriate use of antibiotics has emerged as an important issue for improving the quality of care.

Isolation and Characterization of Intestinal Escherichia coli Clones from Wild Boars in Germany

Our understanding of the composition of Escherichia coli populations in wild boars is very limited. In order to obtain insight into the E. coli microflora of wild boars, we studied E. coli isolates from the jejunums, ileums, and colons of 21 wild boars hunted in five geographic locations in Germany. Ten isolates per section were subjected to clonal determination using pulsed-field gel electrophoresis. One representative isolate per clone was further investigated for virulence traits, phylogenetic affiliation, and antimicrobial susceptibility. Macrorestriction analysis of 620 isolates revealed a range of clone diversity among the sections and animals, with up to 9 and 16 different clones per section and animal, respectively. Most of the clones for a given animal were shared between two adjacent intestinal sections. The overall highest clonal diversity was observed within the colon. While the astA gene was present in a large number of clones, other virulence genes and hemolytic ability were detected only sporadically. Clones of all four ECOR groups dominated the intestinal sections. Phylogenetic analysis and the occurrence of virulence genes correlated with the isolation frequencies for clones. All E. coli clones from wild boars were susceptible to all antimicrobial agents tested. In conclusion, though several parameters (including an animal-specific and highly diverse E. coli clone composition, the simultaneous occurrence of single clones in two adjacent intestinal sections of a given animal, and a higher E. coli diversity in the large intestine than in the small intestine) of E. coli populations of wild boars were similar to those of previously described E. coli populations of conventionally reared domestic pigs, our data also indicate possible differences, as seen for the E. coli diversity in the large intestine, the occurrence of certain virulence genes and phylogenetic groups, and antimicrobial susceptibilities.

Outbreak by meropenem-resistant Pseudomonas aeruginosa producing IMP-6 metallo-β-lactamase in a Korean hospital

Pseudomonas aeruginosa isolates containing the bla IMP-6 gene were recovered from 5 patients hospitalized at a tertiary-care hospital in Korea. The bla IMP-6 gene was in a class 1 integron containing 5 different insert gene cassettes. All of the isolates showed identical pattern in SpeI macrorestriction analysis.

Multilocus sequence typing, pulsed-field gel electrophoresis, and fla short variable region typing of clonal complexes of Campylobacter jejuni strains of human, bovine, and poultry origins in Luxembourg.

Campylobacter jejuni is the most common cause of bacterial gastroenteritis in Luxembourg, with a marked seasonal peak during summer. The majority of these infections are thought to be sporadic, and the relative contribution of potential sources and reservoirs is still poorly understood. We monitored human cases from June to September 2006 (n = 124) by molecular characterization of isolates with the aim of rapidly detecting temporally related cases. In addition, isolates from poultry meat (n = 36) and cattle cecal contents (n = 48) were genotyped for comparison and identification of common clusters between veterinary and human C. jejuni populations. A total of 208 isolates were typed by sequencing the fla short variable region, macrorestriction analysis resolved by pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST). We observed a high diversity of human strains during a given summer season. Poultry and human isolates had a higher diversity of sequence types than isolates of bovine origin, for which clonal complexes CC21 (41.6%) and CC61 (18.7%) were predominant. CC21 was also the most common complex found among human isolates (21.8%). The substantial concordance between PFGE and MLST results for this last group of strains suggests that they are clonally related. Our study indicates that while poultry remains an important source, cattle could be an underestimated reservoir of human C. jejuni cases. Transmission mechanisms of cattle-specific strains warrant further investigation.

Molecular Characterization of Human-Colonizing Streptococcus agalactiae Strains Isolated from Throat, Skin, Anal Margin, and Genital Body Sites

Streptococcus agalactiae carriage was evaluated by sampling four body sites in a group of 249 healthy individuals including both sexes and a wide range of ages; the aims were to study the population structure of colonizing strains by multilocus sequence typing (MLST) and to evaluate their diversity by serotyping, SmaI macrorestriction analysis, and PCR screening for genetic markers of highly virulent clones for neonates. The prevalences of carriage were 27% in women and 32% in men. The major positive body site was the genital tract (23% in women and 21% in men); skin, throats, and anal margins were also positive in 2%, 4%, and 14%, respectively. These human-colonizing strains belonged mostly to serotypes III (24%), Ia (21%), V (18%), and Ib (17%). Twenty-three sequence types (STs) were identified. The MLST characteristics of the strains isolated from a single anatomic site-genital (vagina [women] or from a sample of the first urination after arising from a night's sleep [men]), throat, skin, or anal margin-suggest a body site colonization specificity for particular STs: strains of STs 2, 10, 19, and 196 were isolated only from genital sites; strains of STs 1, 8, and 23 were isolated more frequently from throat florae; and strains recovered only from anal margin samples were more closely related to strains isolated from throats than to those from genital sites. Most strains of STs 1, 8, and 23-STs that are increasingly described as being responsible for adult infections-did not carry any markers of strains virulent for neonates, suggesting that the virulence of these strains is probably associated with other genetic determinants. In addition, the genetic diversities of the strains varied between STs: STs 2, 8, 10, 23, and 196 were the most diverse; STs 1 and 19 were more homogeneous; and ST 17 strains formed three distant groups.

Molecular analysis of multiresistant porcine Salmonella enterica subsp. enterica serovar Bredeney isolates from Southern Brazil: identification of resistance genes, integrons and a group II intron

The relationships of 83 porcine Salmonella enterica subsp. enterica serovar Bredeney isolates obtained at two slaughterhouses in Southern Brazil were analysed by XbaI and BlnI macrorestriction analysis, plasmid profiling and determination of antimicrobial resistance patterns. Twenty-nine XbaI and 30 BlnI macrorestriction patterns were identified. The 72 plasmid-bearing isolates exhibited 20 different plasmid profiles. Multiresistance was detected in 49 isolates (59%), of which 39 isolates showed at least resistance to sulfonamides, tetracyclines, chloramphenicol, streptomycin, kanamycin and/or ampicillin. A representative subset of 12 isolates was chosen for identification of resistance genes, their localisation and transferability. The sulfonamide resistance genes sul1, sul2 and sul3, the tetracycline resistance genes tet(A) and tet(B), the phenicol resistance genes catA1 and floR, the streptomycin resistance gene strA, the kanamycin resistance gene aphA1 and the ampicillin resistance gene bla TEM were detected and found to be located most frequently on plasmids. In addition, class 1 and 2 integrons with the cassette arrangements dfrA21/ bla OXA-129/ aadA1 and dfrA1/ sat1/ aadA1, respectively, were detected. A group II intron was found to be inserted into the 59-base element of an aadA1 gene cassette in a class 1 integron. This study revealed a wide genomic variety among the S. Bredeney isolates, and the high number of multiresistant isolates may point towards the risks that these S. Bredeney isolates can represent to human health.

Relationship between Escherichia coli Strains Causing Acute Cystitis in Women and the Fecal E. coli Population of the Host

Previous epidemiological assessments of the prevalence versus special-pathogenicity hypothesis for urinary tract infection (UTI) pathogenesis in women may have been confounded by underlying host population differences between women with UTI and healthy controls and have not considered the clonal complexity of the fecal Escherichia coli population of the host. In the present study, 42 women with acute uncomplicated cystitis served as their own controls for an analysis of the causative E. coli strain and the concurrent intestinal E. coli population. Clonality among the urine isolate and 30 fecal colonies per subject was assessed by repetitive-element PCR and macrorestriction analysis. Each unique clone underwent PCR-based phylotyping and virulence genotyping. Molecular analysis resolved 109 unique clones (4 urine-only, 38 urine-fecal, and 67 fecal-only clones). Urine clones exhibited a significantly higher prevalence of group B2 than fecal-only clones (69% versus 10%; P < 0.001) and higher aggregate virulence scores (mean, 6.2 versus 2.9; P < 0.001). In multilevel regression models for predicting urine clone status, significant positive predictors included group B2, 10 individual virulence traits, the aggregate virulence score, fecal dominance, relative fecal abundance, and (unique to the present study) a pauciclonal fecal sample. In summary, within the fecal E. coli populations of women with acute cystitis, pauciclonality, clonal dominance, virulence, and group B2 status are closely intertwined. Phylogenetic group B2 status and/or associated virulence factors may promote fecal abundance and pauciclonality, thereby contributing to upstream steps in UTI pathogenesis. This relationship suggests a possible reconciliation of the prevalence and special-pathogenicity hypotheses.

Efficient removal of attached biofilm in a naturally contaminated colonoscope using detachment-promoting agents

We would like to report a successful ‘last chance’ operation during which a highly contaminated colonoscope was effectively treated using an anti-biofilm procedure based on a synergistic combination of detachment-promoting agents (DPAs). Various authors have described the considerable difficulties presented by the removal of attached bacteria from endoscope channels. 1 Spach D.H. Silverstein F.E. Stamm W.E. Transmission of infection by gastrointestinal endoscopy and bronchoscopy. Ann Intern Med. 1993; 118: 117-128 Crossref PubMed Scopus (437) Google Scholar , 2 Struelens M.J. Rost F. Deplano A. et al. Pseudomonas aeroginosa and Enterobacteriaceae bacteremia after biliary endoscopy: an outbreak investigation using DNA macrorestriction analysis. Am J Med. 1993; 95: 489-498 Abstract Full Text PDF PubMed Scopus (136) Google Scholar , 3 Birnie G.G. Quigley E.M. Clements G.B. Follet E.A. Watkinson G. Endoscopic transmission of hepatitis B virus. Gut. 1983; 24: 171-174 Crossref PubMed Scopus (183) Google Scholar We agree with Vickery et al. who considered that the persistence of this attached biomass on endoscope channels is often not due to inadequate compliance with reprocessing protocols, but usually caused by the low efficiency of current reprocessing protocols that do not totally eliminate attached micro-organisms. 4 Vickery K. Pajkos A. Cossart Y. Removal of biofilms from endoscopes: evaluation of detergent efficiency. Am J Infect Control. 2004; 32: 170-176 Abstract Full Text Full Text PDF PubMed Scopus (106) Google Scholar We previously demonstrated that the use of a cleaning agent according to the Pronetron® procedure, prior to disinfection in compliance with French guidelines, totally detached a preformed biofilm from the inner surface of an operating channel. 5 Anonymous. Circulaire DHOS/E2/DGS/SD5C/2003/N°591 relative aux modalités de traitement manuel pour la désinfection des endoscopes non autoclavables dans les lieux de soins. Ministère de la santé, de la famille et des personnes handicapées. Paris: 2003. Google Scholar , 6 Marion K. Freney J. James G. Bergeron E. Renaud F.N.R. Costerton J.W. Using an efficient biofilm detaching agent: an essential step for the improvement of endoscope reprocessing protocols. J Hosp Infect. 2006; 64: 136-142 Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar This corrective procedure consists of the sequential action of two DPA-based products: a multi-enzymatic solution (Pronetron α) that digests the anchorage points of the micro-organisms to the support, and an enriched detergent solution (Pronetron β Plus) that detaches the whole biomass from the support.

Molecular characterization of vancomycin-resistant Enterococcus spp. clinical isolates from Hungary and Serbia

The objectives of this work were to collect and characterize vancomycin-resistant Enterococcus faecium (VREF) clinical isolates from Hungary and Serbia and to analyse their genetic relatedness. VREF isolates were initially typed by PFGE. A selection of VREF isolates representing all participating hospitals was further examined by multiple-locus variable-number tandem repeat analysis (MLVA) and multilocus sequence typing (MLST). VanB VREF isolates (n=18) recovered from blood, urine and faecal cultures at a Budapest hospital between August 2003 and December 2004 were molecularly characterized. Macrorestriction analysis of the isolates revealed their monoclonal relatedness. A cluster of infections caused by 2 distinct VanA VREF clones recovered from 6 departments was identified in a Belgrade hospital in Serbia. The vanA resistance determinant was transferable by in vitro conjugation experiments. We also identified 2 vanA-positive E. gallinarum blood culture isolates in this Belgrade hospital. Molecular typing of representative VREF isolates from Hungary and Serbia by MLVA and MLST revealed that all tested isolates belonged to MLST complex CC17 and the corresponding MLVA cluster 1. Our results extend the documented occurrence of CC17 to a new region in Europe.

Methicillin-resistant Staphylococcus aureus and Staphylococcus pseudintermedius detected in the BfT-GermVet monitoring programme 2004-2006 in Germany

During recent years, methicillin-resistant Staphylococcus aureus (MRSA) strains from animals have become the focus of various studies. In the present study, coagulase-positive staphylococci obtained from pigs, dogs and cats suffering from acute infections, which had been collected in the BfT-GermVet monitoring programme in Germany and phenotypically identified as oxacillin-resistant, were characterized. The staphylococci were comparatively investigated for their resistance phenotypes and genotypes. Resistance genes were identified by PCR. MRSA strains were further characterized by SmaI macrorestriction analysis and spa typing to assess their genomic relationships. Among the 248 strains tested, 7 strains (5 porcine S. aureus and 2 canine Staphylococcus pseudintermedius) carried the resistance gene mecA. Gentamicin resistance was based on the presence of the gene aacA/aphD while three different tetracycline resistance genes, tet(K), tet(L) and tet(M), alone or in combinations, were detected. The single macrolide/lincosamide-resistant strain carried an erm(A) gene. All MRSA strains proved to be non-typeable by SmaI macrorestriction analysis and exhibited the spa types t011 (four strains) or t034 (one strain). Based on their spa types and their non-respondence to SmaI digestion, all five porcine MRSA strains resembled MRSA strains of multilocus sequence type ST398, previously detected among pigs in neighbouring countries such as The Netherlands or Denmark. The results of this study indicate that such strains are also involved in defined disease conditions of pigs from various parts of Germany.

Increasing rates of vancomycin resistance among Enterococcus faecium isolated from German hospitals between 2004 and 2006 are due to wide clonal dissemination of vancomycin-resistant enterococci and horizontal spread of vanA clusters

Results of national and international surveillance studies revealed increasing rates of vancomycin-resistant Enterococcus faecium (VREF) among German hospital patients since 2003. To investigate the molecular background of vanA-type glycopeptide resistance, 51 clinical VREF isolated between 2004 and 2006 and originating from 19 German hospitals representing 10 Federal States have been investigated. Isolates were characterised by multi-locus sequence typing (MLST), SmaI macrorestriction analysis in pulsed-field gel electrophoresis (PFGE), and multiple-locus variable-number tandem repeat analysis (MLVA). Phylogenetic relatedness between strains was identified using BioNumerics and eBURST software. Distribution of virulence markers esp and hyl Efm was investigated by PCR. The structure of the vanA gene clusters was investigated by PCR, long-template PCR, sequencing and Southern hybridisations. The 51 VREF were rather diverse constituting different strain types, different virulence markers and vanA clusters. Within this diversity we found supportive data for a dissemination of related – already vancomycin-resistant – E. faecium among various hospitals and Federal States and for spread of identical vanA gene clusters among clonally different strain types within single hospitals. In conclusion, the increase in the rates of VREF among German hospital patients within the last 2 years might be rather complex and due to different molecular events and scenarios.