A multilocus sequence typing (MLST) scheme has been developed for Enterococcus faecium. Internal fragments from seven housekeeping genes of 123 epidemiologically unlinked isolates from humans and livestock and 16 human-derived isolates from several outbreaks in the United States, the United Kingdom, Australia, and The Netherlands were analyzed. A total of 62 sequence types were detected in vancomycin-sensitive E. faecium (VSEF) and vancomycin-resistant E. faecium (VREF) isolates. VSEF isolates were genetically more diverse than VREF isolates. Both VSEF and VREF isolates clustered in host-specific lineages that were similar to the host-specific clustering obtained by amplified fragment length polymorphism analysis. Outbreak isolates from hospitalized humans clustered in a subgroup that was defined by the presence of a unique allele from the housekeeping gene purK and the surface protein gene esp. The MLST results suggest that epidemic lineages of E. faecium emerged recently worldwide, while genetic variation in both VREF and VSEF was created by longer-term recombination. The results show that MLST of E. faecium provides an excellent tool for isolate characterization and long-term epidemiologic analysis.Vancomycin-resistant Enterococcus faecium (VREF) has recently emerged as an important threat in U.S. hospitals (5, 24). In Europe, VREF isolates are found relatively frequently in the community and farm animals, while prevalence in hospitals is generally low (14). The latter observation was explained by the use of the glycopeptide avoparcin as an antimicrobial growth promoter in animal feeding operations.Several molecular typing schemes have been developed to study the epidemiology of VREF. Of these, pulsed-field gel electrophoresis analysis of genomic restriction fragments has been considered the "gold standard" for the study of hospital outbreaks because of its high degree of isolate differentiation (15,17,20,23). However, due to this high degree of isolate differentiation, pulsed-field gel electrophoresis typing is less suitable for determining the degree of relatedness among epidemiologically unrelated isolates. Recently, amplified fragment length polymorphism (AFLP) analysis was applied as a new method for the typing of VREF (1, 33). AFLP analysis is a robust and fast typing technique with high intra-and interexperimental reproducibilities and appears to be discriminatory enough for the recognition of hospital outbreaks (1, 32, 33). In addition, AFLP analysis has allowed the detection of associations among different E. faecium genetic lineages and different human and animal hosts (33), suggesting the existence of host-specific VREF lineages. Whether this is also true for vancomycin-sensitive E. faecium (VSEF) is not known, since VSEF isolates were not included in that study. AFLP typing also disclosed two different human-associated lineages. One lineage comprised epidemic-related isolates recovered from hospitalized patients, while isolates of the other lineage were mainly from nonhospitalized persons. Interes...

No abstract

No abstract

A multilocus sequence typing (MLST) scheme has been developed for Enterococcus faecium. Internal fragments from seven housekeeping genes of 123 epidemiologically unlinked isolates from humans and livestock and 16 human-derived isolates from several outbreaks in the United States, the United Kingdom, Australia, and The Netherlands were analyzed. A total of 62 sequence types were detected in vancomycin-sensitive E. faecium (VSEF) and vancomycin-resistant E. faecium (VREF) isolates. VSEF isolates were genetically more diverse than VREF isolates. Both VSEF and VREF isolates clustered in host-specific lineages that were similar to the host-specific clustering obtained by amplified fragment length polymorphism analysis. Outbreak isolates from hospitalized humans clustered in a subgroup that was defined by the presence of a unique allele from the housekeeping gene purK and the surface protein gene esp. The MLST results suggest that epidemic lineages of E. faecium emerged recently worldwide, while genetic variation in both VREF and VSEF was created by longer-term recombination. The results show that MLST of E. faecium provides an excellent tool for isolate characterization and long-term epidemiologic analysis.Vancomycin-resistant Enterococcus faecium (VREF) has recently emerged as an important threat in U.S. hospitals (5, 24). In Europe, VREF isolates are found relatively frequently in the community and farm animals, while prevalence in hospitals is generally low (14). The latter observation was explained by the use of the glycopeptide avoparcin as an antimicrobial growth promoter in animal feeding operations.Several molecular typing schemes have been developed to study the epidemiology of VREF. Of these, pulsed-field gel electrophoresis analysis of genomic restriction fragments has been considered the "gold standard" for the study of hospital outbreaks because of its high degree of isolate differentiation (15,17,20,23). However, due to this high degree of isolate differentiation, pulsed-field gel electrophoresis typing is less suitable for determining the degree of relatedness among epidemiologically unrelated isolates. Recently, amplified fragment length polymorphism (AFLP) analysis was applied as a new method for the typing of VREF (1, 33). AFLP analysis is a robust and fast typing technique with high intra-and interexperimental reproducibilities and appears to be discriminatory enough for the recognition of hospital outbreaks (1, 32, 33). In addition, AFLP analysis has allowed the detection of associations among different E. faecium genetic lineages and different human and animal hosts (33), suggesting the existence of host-specific VREF lineages. Whether this is also true for vancomycin-sensitive E. faecium (VSEF) is not known, since VSEF isolates were not included in that study. AFLP typing also disclosed two different human-associated lineages. One lineage comprised epidemic-related isolates recovered from hospitalized patients, while isolates of the other lineage were mainly from nonhospitalized persons. Interes...

No abstract

No abstract

SUMMARYSamples of DNA from a panel ofGiardiaisolated from humans and animals in Europe and shown previously to consist of 2 major genotypes–‘Polish’ and ‘Belgian’–have been compared with human-derived Australian isolates chosen to represent distinct genotypes (genetic groups I–IV) defined previously by allozymic analysis. Homologous 0·52 kilobase (kb) segments of 2 trophozoite surface protein genes (tsa417 and tsp11, both present in isolates belonging to genetic groups I and II) and a 1·2 kb segment of the glutamate dehydrogenase (gdh) gene were amplified by the polymerase chain reaction (PCR) and examined for restriction fragment length polymorphisms (RFLPs). Of 21 ‘Polish’ isolates that were tested, all yieldedtsa417-like andtsp11-like PCR products that are characteristic of genetic groups I or II (15 and 6 isolates respectively) in a distinct assemblage ofG. intestinalisfrom Australia (Assemblage A). Conversely, most of the 19 ‘Belgian’ isolates resembled a second assemblage of genotypes defined in Australia (Assemblage B) which contains genetic groups III and IV. RFLP analysis ofgdhamplification products showed also that ‘Polish’ isolates-were equivalent to Australian Assemblage A isolates (this analysis does not distinguish between genetic groups I and II) and that ‘Belgian’ isolates were equivalent to Australian AssemblageB isolates. Comparison of nucleotide sequences determined for a 690 base-pair portion of thegdhPCR products revealed ≥ 99·0% identity between group I and group II (Assemblage A/‘Polish’) genotypes, 88·3–89·7% identity between Assemblage A and Assemblage B genotypes, and ≥ 98·4% identity between various Assemblage B/‘Belgian’ genotypes. The results confirm that theG. duodenalisisolates examined in this study (inclusive ofG. intestinalisfrom humans) can be divided into 2 major genetic clusters: Assemblage A (= ‘Polish’ genotype) containing allozymically defined groups I and II, and Assemblage B (= ‘Belgian’ genotype) containing allozymically defined groups III and IV and other related genotypes.

Isolates of the protozoan parasite Cryptosporidium parvum have been differentiated into 2 genotypes: genotype 'H', which is associated only with human infections, and genotype 'C', which is associated with both human and animal infections. To date, the analysis of polymorphisms of genes and of the small subunit ribosomal DNA have revealed no heterogeneity within the 2 genotypes. In the present study, a locus containing simple sequence repeats (microsatellites) was PCR amplified and sequenced from 94 C. parvum isolates, which were collected from humans (immunocompetent and immunocompromized individuals, outbreak and single cases) and from several animal hosts in 3 continents. The analysis revealed that genotype 'H' can be further differentiated into 2 subgenotypes, and genotype 'C' can be further differentiated into 4 subgenotypes. The 6 subgenotypes differ in terms of expansions/contractions of the microsatellite repeats and by point mutations. Some subgenotypes showed a wide geographical distribution, whereas others were restricted to specific regions. Therefore, microsatellites are informative markers for more defined studies on the epidemiology, the transmission routes, and the population structure of this parasite.