Lesbian uses of and satisfaction with mental health services: results from Boston Lesbian Health Project

Eighteen isolates representing six Fusarium species from diverse hosts and geographical origins were evaluated to determine ribosomal DNA variation using polymerase chain reaction and restriction fragment length polymorphisms. No length variation was observed for amplified 18S and 28S regions. However, amplification of the ITS region showed one isolate, a F. oxysporum, to be about 120 bp larger than the remaining 17. Restriction digestions in the 18S region revealed polymorphisms within species of F. oxysporum and F. solani. An amplified variable stretch of the 28S gene showed restriction site differences between F. avenecum, F. sambucinum and F. sporotrichioides. A large degree of polymorphism was observed both between and within species in the ITS region. Therefore, entire sequences of the ITS and the 5.8S subunit were obtained for 17 of the 18 isolates. These sequences, along with those from eight additional isolates, were analysed using PAUP to assess the occurrence of DNA sequence divergence within the ITS region. The lack of correlation between molecular-based relationships and species affinities inferred from morphology for some isolates indicates that species designation can be unreliable using morphological data alone. Possible reasons for the discordance of the sequence and morphological data are discussed.     

IgG autoantibodies to complement C1q in pediatric-onset systemic lupus erythematosus

The ribosomal internal transcribed spacer (ITS) region from individual Gyrodactylus specimens was amplified by polymerase chain reaction (PCR). The reaction amplified the entire ITS1-5.8S-ITS2 region of the ribosomal RNA gene cluster using primers that hybridize to the 3' terminus of the small subunit and the 5' terminus of the large subunit ribosomal RNA genes. The PCR products from Gyrodactylus salaris and Gyrodactylus thymalli were cloned and sequenced. The Gyrodactylus 5.8S gene was identified following comparative alignment of the G. salaris sequence and a Schistosoma 5.8S gene sequence. The ITS regions from G. salaris, G. thymalli, Gyrodactylus derjavini, and Gyrodactylus truttae were compared by restriction enzyme analysis and interspecific restriction fragment length polymorphisms were found. Gyrodactylus salaris and G. thymalli restriction fragment sizes were confirmed from sequence data. ITS amplification followed by Sau3AI digestion enables rapid and clear differentiation of G. salaris, G. derjavini, and G. truttae.     

Comparison of the 5.8s rDNA and internal transcribed spacer sequences of isolates of Leptosphaeria maculans from different pathogenicity groups

The regions coding for the 5.8s rRNA and the flanking internal transcribed spacers (ITS1 and ITS2) from nine isolates of the blackleg pathogen Leptosphaeria maculans and one isolate of Sclerotinia sclerotiorum were amplified by the polymerase chain reaction and sequenced. Five of the L. maculans isolates were highly virulent to Brassica plants, two were weakly virulent and two were isolated from the cruciferous weed Thlaspi arvense. The 5.8s DNA sequences of all L. maculans isolates were identical. However, there were major differences in both ITS1 and ITS2 sequences that correlated with the pathogenicity grouping. Phylogenetic analysis of the ITS sequences by both parsimony and maximum-likelihood methods indicated that each pathogenicity group was statistically different from each other with the weakly-virulent isolates being more closely related to the Thlaspi than to the highly-virulent isolates. The relationships of L. maculans to other fungi, based on a comparison of the 5.8s rDNA sequences, are discussed.     

Biodiversity of Pneumocystis carinii hominis: typing with different DNA regions

The purpose of this study was to identify the most useful gene for the detection of biodiversity of Pneumocystis carinii hominis isolates and to compare samples from French and Italian subjects. We studied 20 bronchoalveolar lavage fluid specimens from 20 human immunodeficiency virus-infected patients (10 French and 10 Italian patients) with Pneumocystis carinii pneumonia by DNA sequencing of the thymidylate synthase (TS), 5S rRNA, large-subunit mitochondrial rRNA (mt LSU rRNA), and internal transcribed spacer (ITS1 and ITS2) genes. Thirteen of the 20 sequenced samples had the prototype TS gene sequence. Fourteen of the 20 samples showed the prototype sequence of the 5S rRNA gene, and 6 had variant sequences of the 5S rRNA gene. The mt LSU rRNA gene was sequenced for 18 of the 20 samples; all sequences were different from the prototype sequence and were classified into four groups. Thirteen of the 20 ITS1 and ITS2 sequences were analyzed, and all the sequences were found to be different from the prototype sequence and were classified into 10 groups. The internal transcribed spacer regions thus appear to be the most discriminatory region of DNA for analysis of the biodiversity of P. carinii hominis isolates.     

Intergenic transcribed spacer PCR ribotyping for differentiation of Saccharomyces species and interspecific hybrids

The taxonomy of the genus Saccharomyces has undergone significant changes recently with the use of genotypic rather than phenotypic methods for the identification of strains to the species level. The sequence of rRNA genes has been utilized for the identification of a variety of fungi to the species level. This methodology, applied to species of Saccharomyces, allows unknown Saccharomyces isolates to be assigned to the type strains. It was the aim of the present study to assess whether typing of the intergenic spacer region by using restriction fragment length polymorphisms of PCR products (intergenic transcribed spacer PCR [ITS-PCR] ribotyping) could distinguish among type strains of the 10 accepted species of Saccharomyces and further to assess if this method could distinguish strains that were interspecific hybrids. Cellular DNA, isolated after the lysis of protoplasts, was amplified by PCR using ITS1 and ITS4 primers, purified by liquid chromatography, and digested with restriction endonucleases. Ribotyping patterns using the restriction enzymes MaeI and HaeIII could distinguish all species of Saccharomyces from each other, as well as from Candida glabrata, Candida albicans, and Blastomyces dermatitidis. The only exception to this was the inability to distinguish between Saccharomyces bayanus and S. pastorianus (S. carlsbergensis). Furthermore, interspecific hybrids resulting from the mating of sibling species of Saccharomyces were shown to share the ITS-PCR ribotyping patterns of both parental species. It should now be possible, by this simple PCR-based technique, to accurately identify these strains to the species level, thereby allowing an increase in our understanding of the characteristics required by these interspecific hybrids for their particular ecological niches.     

Rapid identification and typing of Staphylococcus aureus by PCR-restriction fragment length polymorphism analysis of the aroA gene

The Staphylococcus aureus aroA gene, which encodes 5-enolpyruvylshikimate-3-phosphate synthase, was used as a target for the amplification of a 1,153-bp DNA fragment by PCR with a pair of primers of 24 and 19 nucleotides. The PCR products, which were detected by agarose gel electrophoresis, were amplified from all S. aureus strains so far analyzed (reference strains and isolates from cows and sheep with mastitis, as well as 59 isolates from humans involved in four confirmed outbreaks). Hybridization with an internal 536-bp DNA fragment probe was positive for all PCR-positive samples. No PCR products were amplified when other Staphylococcus spp. or genera were analyzed by using the same pair of primers. The detection limit for S. aureus cells was 20 CFU when the cells were suspended in saline; however, the sensitivity of the PCR was lower (5 x 10(2) CFU) when S. aureus cells were suspended in sterilized whole milk. TaqI digestion of the PCR-generated products rendered two different restriction fragment length polymorphism patterns with the cow and sheep strains tested, and these patterns corresponded to the two different patterns obtained by antibiotic susceptibility tests. Analysis of the 59 human isolates by our easy and rapid protocol rendered results similar to those of other assays.     

Organisms isolated from dogs and cats with anaerobic infections and susceptibility to selected antimicrobial agents

OBJECTIVE: To determine the prevalence of obligate anaerobic bacteria in bacterial infections in dogs and cats and susceptibility to selected antimicrobial agents. DESIGN: Case series. SAMPLE POPULATION: Specimens from 1,267 dogs and 243 cats. PROCEDURE: Standard anaerobic and aerobic bacterial culture methods were used. Anaerobic isolates were tested for susceptibility to selected antimicrobial agents. RESULTS: Obligate anaerobic bacteria were isolated from 199 (15.7%) and 69 (28.4%) specimens obtained from dogs and cats, respectively. More than half of the specimens that contained obligate anaerobic bacteria were from draining tracts (exclusively dogs), pleural fluid, abscesses, bones, the respiratory tract, or the abdominal cavity. The most commonly isolated obligate anaerobic bacteria (approx 70% of all isolates) were Bacteroides spp, Peptostreptococcus spp, Fusobacterium spp, and Porphyromonas spp. Eighty percent of the specimens that contained obligate anaerobic bacteria also contained facultative anaerobic or aerobic organisms. The organisms most commonly isolated in association with obligate anaerobic bacteria were members of the family Enterobacteriaceae (Escherichia coli was the most common), Pasteurella spp, and Staphylococcus intermedius. Ninety-seven obligate anaerobic isolates were tested for susceptibility to ampicillin, amoxicillin-clavulanic acid, chloramphenicol, clindamycin, and metronidazole. All were susceptible to amoxicillin-clavulanic acid and chloramphenicol, and most were susceptible to metronidazole. Only 71% of the Bacteroides isolates were susceptible to ampicillin, and only 83% were susceptible to clindamycin. Only 80% of the Clostridium isolates were susceptible to clindamycin, but all were susceptible to ampicillin. CLINICAL IMPLICATIONS: Data on sites and conditions from which anaerobic bacteria are commonly isolated, along with results of susceptibility testing, may be useful in designing antimicrobial treatment regimens.     

Rapid identification of clinically significant species and taxa of aerobic actinomycetes, including Actinomadura, Gordona, Nocardia, Rhodococcus, Streptomyces, and Tsukamurella isolates, by DNA amplification and restriction endonuclease analysis

A previously described PCR-restriction fragment length polymorphism (RFLP) identification schema for Nocardia that used an amplified 439-bp segment (amplicon) of the 65-kDa heat shock protein gene was evaluated for potential use with isolates of all clinically significant aerobic actinomycetes. The study included 28 reference (American Type Culture Collection) strains and 198 clinical isolates belonging to 20 taxonomic groups. Of these 198 isolates, 188 could be differentiated by this PCR-RFLP method. Amplicons from all aerobic actinomycete isolates lacked BstEII recognition sites, thereby distinguishing them from those of mycobacteria that contain one or more such sites. Of 29 restriction endonucleases, MspI plus HinfI produced RFLP patterns that differentiated 16 of the 20 taxa. A single RFLP pattern was observed for 15 of 20 taxa that included 65% of phenotypically clustered isolates. Multiple patterns were seen with Gordona bronchialis, Nocardia asteroides complex type VI, Nocardia otitidiscaviarum, Nocardia transvalensis, and Streptomyces spp. Streptomyces RFLP patterns were the most heterogeneous (five patterns among 19 isolates), but exhibited a unique HinfI fragment of > 320 bp. RFLP patterns that matched those from type strains of Streptomyces albus, Streptomyces griseus, or Streptomyces somaliensis were obtained from 14 of 19 Streptomyces isolates. Only 10 of 28 isolates of N. otitidiscaviarum failed to yield satisfactory amplicons, while only 6 of 188 (3.2%) clinical isolates exhibited patterns that failed to match one of the 21 defined RFLP patterns. These studies extended the feasibility of using PCR-RFLP analysis as a rapid method for the identification of all clinically significant species and taxa of aerobic actinomycetes.     

Molecular evolution of the internal transcribed spacers (ITS1 and ITS2) and phylogenetic relationships among species of the family Cucurbitaceae

Phylogenetic relationships of different members of the family Cucurbitaceae were estimated from sequences of the internal transcribed spacer (ITS1 and ITS2) regions of the nuclear ribosomal RNA genes. Twenty-six species of different genera belonging to different tribes and several subtribes were analyzed. The whole ITS regions were amplified by PCR technique and cloned, and three to five different clones of each species were sequenced; for some species PCR products were sequenced directly. ITS1 and ITS2 regions are slightly variable in length, with each length appearing genus-specific. A substitution rate of 3.62 x 10(-9) substitutions per site per year was calculated assuming 40 MYA separation time. Phylogenetic relationships inferred from ITS sequences of some species is in agreement with morphological data, but deviations to the taxonomic classification were also observed. A polyphyletic origin of the New World species must be considered. In the genus Cucurbita different "types" of ITS sequences within one species exist, possibly due to the high frequency of introgression during domestication or due to polyploidization events; in contrast, low intraspecific variability was detectable in the genus Cucumis, indicating different stages of speciation.     

Clostridial infection in children

A survey of the isolation of Clostridium spp. from 1543 specimens sent to anaerobic microbiology laboratories revealed 113 isolates from 107 specimens (7.0% of all specimens) from 96 children. The isolates comprised 43 (38%) unidentified Clostridium spp., 37 (33%) C. perfringens, 13 (12%) C. ramosum, five (4%) C. innocuum, six (5%) C. botulinum, three (3%) C. difficile, two (2%) C. butyricum, and one isolate each of C. bifermentans, C. clostridiiforme, C. limosum and C. paraputrificum. Most clostridial isolates were from abscesses (38), peritonitis (26), bacteraemia (10), and chronic otitis media (7). Predisposing or underlying conditions were present in 31 (32%) cases. These were immunodeficiency (12), malignancy (9), diabetes (7), trauma (7), presence of a foreign body (6) and previous surgery (6). The clostridia were the only bacterial isolates in 14 (15%) cases; 82 (85%) cases had mixed infection. The species most commonly isolated with clostridia were anaerobic cocci (57); Bacteroides spp. (B. fragilis group) (50), Escherichia coli (22), pigmented Prevotella or Porphyromonas spp. (18) and Fusobacterium spp. (10). Most Bacteroides and Escherichia coli isolates with clostridia were from abdominal infections and skin and soft tissue infections adjacent to the rectal area; most pigmented Prevotella and Porphyromonas isolates were from oropharyngeal, pulmonary, and head and neck sites. Antimicrobial therapy was given to all patients, in conjunction with surgical drainage in 34 (35%). Only two patients died. These data illustrate the importance of Clostridium spp. in paediatric infections.     

Three distinct genotypes within Candida parapsilosis from clinical sources

Three genetically distinct groups of Candida parapsilosis were detected among clinical isolates. These were distinguishable on the basis of isoenzyme profiles and DNA sequences of internally transcribed spacer (ITS) sequences flanking the 5.8S RNA gene. In an investigation of 45 strains, including 32 clinical isolates from Texas, C. parapsilosis group I composed the majority of the common clinical isolates. The type strain of C. parapsilosis was a member of this group. The 10 group II isolates were indistinguishable from group I strains when tested with the API 20C kit. The two group III isolates differed from those in groups I and II by being D-xylitol positive by the API 20C kit; however, isolates in all groups assimilated D-xylitol from broth. Isoenzyme profiles excluded the close relationship of any of these groups to Lodderomyces elongisporus, which is a teleomorphic yeast that has a physiological profile similar to that of C. parapsilosis. Although there were insignificant differences in the ITS2 rDNA sequences, comparisons of the ITS1 sequences revealed several differences. A sequence analysis of ITS1 in which missing bases were counted as mismatches showed the following similarities: group I versus group II, 87.7%; group I versus group III, 82.1%; group II versus group III, 84.5%. Also, the activity of secreted proteinase showed differences among the three groups, with many group I isolates having moderate to high activity. The degree of susceptibility to antifungal agents, amphotericin B, ketoconazole, and 5-fluorocytosine, could not be used to determine an isolate's group.     

Bacterial pathogens and their antimicrobial susceptibility in Kumasi, Ghana

Between January, 1994 and June, 1996 a survey of bacterial isolates from clinical specimens and their antimicrobial susceptibility was performed at the Komfo Anokye Teaching Hospital, Microbiology Department, Kumasi, Ghana. A total of 11,380 bacterial isolates were cultured from eight different specimens. The sites of origin were wounds 32.2%, urine 28.1%, ear, nose and throat 3.6%, sputum 2.5% and aspirates 2.5%. Gram-negative bacteria accounted for 7955 (69.9%) isolates, the main species were Escherichia coli 47.1%, Pseudomonas spp. 16.8%, Proteus spp 14.6%, Klebsiella spp 10.2%, Neisseria gonorrhoeae 4.2%, Gram-positive bacteria contributed 3425 ((30.1%) of isolates, with Staphylococcus aureus 54.6% being the most predominant followed by Coagulase negative Staphylococcus 18.1%, Streptococcus pneumoniae 13.7% and Beta-haemolytic streptococci 4.1%. Escherichia coli showed 88% and 82% resistance to ampicillin and cotrimoxazole respectively with 78% being susceptible to gentamicin. Cefuroxime resistance in Gram-negative bacilli was 5%. As much as 30.6% and 21.7% of Streptococcus pneumoniae isolates were resistant to Penicillin and chloramphenicol respectively. Ten per cent of Staphylococcus aureus strains were susceptible to penicillin and 18% were resistant to flucloxacillin.     

A novel toxinotyping scheme and correlation of toxinotypes with serogroups of Clostridium difficile isolates

Two hundred nineteen Clostridium difficile isolates from 22 serogroups were screened for changes in the genes coding for toxin B (tcdB) and toxin A (tcdA). Parts of the toxin genes were amplified, and the PCR fragments were checked for length polymorphisms and cut with several restriction enzymes to monitor restriction fragment length polymorphisms (RFLPs). For 47 strains (21%), differences in the toxin genes were found compared to the toxin genes of reference strain VPI 10,463. Polymorphisms were usually observed in both toxin genes. RFLPs were more commonly found in the tcdB gene, in which a single restriction enzyme could give up to five different patterns. Restriction sites seemed to be less heterogeneous in the tcdA gene, in which for most enzymes only two different RFLPs were recognized. However, deletions were observed in tcdA, and four new types of shortened tcdA genes are described. According to the changes in their toxin genes, variant strains could be divided into 10 groups (toxinotypes I to X). A toxinotype was characterized by similar patterns of changes in the toxin genes and in other regions of the pathogenicity locus and also similar pulsed-field gel electrophoresis patterns. Variant toxinotypes were found in 9 of the 22 serogroups studied, and some toxinotypes were clearly associated with specific serogroups. Toxinotype VIII is characteristic for all strains of serogroup F. Other serogroups in which variant toxinotypes were commonly found are A1, A15, E, and X. Testing of variability in C. difficile toxin genes not only might be useful as a molecular typing system but also could have implications in diagnostics and pathogenesis.     

Molecular and phenotypic characterization of genotypic Candida albicans subgroups and comparison with Candida dubliniensis and Candida stellatoidea

There have been increased reports of the isolation of unusual genotypic groups of Candida albicans (groups C and D) based on a well-defined genotypic method; this method uses cellular DNA digested with the EcoRI enzyme and the restriction fragment length polymorphisms (RFLPs) generated by agarose gel electrophoresis. The aim of the present study was to use additional molecular tools to characterize these unusual strains and to compare them with authentic strains of C. dubliniensis, a recently delineated species, and type I C. stellatoidea. The RFLPs of PCR products generated from the intergenic transcribed spacer (ITS) region did not differentiate among C. albicans genotypes A, B, and C and type I C. stellatoidea. However, this method did differentiate the C. albicans genotype D strains, which were identical to C. dubliniensis. The RFLPs generated by HaeIII digestion of the PCR products of the V3 region of the 25S rRNA gene (rDNA) could differentiate the same groups as RFLP analysis of the PCR amplicon of the ITS region. C. albicans genotype B isolates have been shown to have a transposable intron in the 25S rDNA, whereas genotype A isolates do not; C. dubliniensis strains also have an intron that is larger than that in genotype B C. albicans strains but that is in the same location. PCR designed to span this region resulted in a single product for C. albicans genotype A (450 bp), B (840 bp), type 1 C. stellatoidea (840 bp), and C. dubliniensis (1,080 bp), whereas the C. albicans genotype C isolates had two major products (450 and 840 bp). All C. albicans genotype D isolates gave a PCR product identical to that given by C. dubliniensis. These results indicate that those strains previously designated C. albicans genotype D are in fact C. dubliniensis, that no differences were found between type 1 C. stellatoidea and C. albicans genotype B strains, and that the C. albicans genotype C strains appear to have the transposable intron incompletely inserted throughout the ribosomal repeats in their genomes. The results of the antifungal susceptibility testing of 105 of these strains showed that, for fluconazole, strains of C. dubliniensis were significantly more susceptible than strains of each of the C. albicans genotypes (genotypes A, B, and C). The flucytosine susceptibility results indicated that strains of C. albicans genotype A were significantly less susceptible than either C. albicans genotype B or C. albicans genotype C strains. These results indicate that there is a correlation between the Candida groups and antifungal susceptibility.     

A comparison of three measures of estimating premorbid intellectual level in dementia of the Alzheimer type

Forty-four patients receiving intensive care were studied prospectively to assess the utility of serial rectal swab cultures and clinical correlates of resistance for Pseudomonas aeruginosa, Enterobacter spp., Citrobacter spp., Morganella morganii, and Serratia marcescens strains resistant to ceftazidime or imipenem. Strains of Pseudomonas aeruginosa, Enterobacter spp., Citrobacter spp., or Morganella morganii were found in 26 of 44 (59%) patients: 17 (65%) in clinical sites (11 with concomitant rectal isolates) and nine (35%) in a rectal site only. Of 49 total isolates, 13 (26.5%) were resistant: 10 (20.4%) to ceftazidime and three (6.1%) to imipenem. Surveillance rectal swabs from 27 patients without a clinical isolate identified two patients with resistant organisms (15% of all resistant isolates). The majority of resistance to ceftazidime or imipenem among Pseudomonas or Enterobacter can be detected by the use of clinical specimens alone.     

Isolation and identification of Aeromonas species from human stools

One thousand and three diarrhoeal stool samples were processed in our laboratory during the period 1996/1997 for the presence of enteric pathogens especially Aeromonas spp., which has emerged as a new agent causing diarrhoea. Ampicillin sheep blood agar was found to be the best medium for the isolation of Aeromonas spp. from stool specimens. Enteric pathogens were found in 200 (20%) stools, of which Aeromonas spp. was the second commonest pathogen isolated amounting to 21% of isolates. This study clearly indicates that Aeromonas spp. must be looked for in every diarrhoeal stool samples, specially in children below 10 years of age. Isolation and identification is cost effective and easy, if the given protocol is observed.     

Microbial diversity in a hydrocarbon- and chlorinated-solvent-contaminated aquifer undergoing intrinsic bioremediation

A culture-independent molecular phylogenetic approach was used to survey constituents of microbial communities associated with an aquifer contaminated with hydrocarbons (mainly jet fuel) and chlorinated solvents undergoing intrinsic bioremediation. Samples were obtained from three redox zones: methanogenic, methanogenic-sulfate reducing, and iron or sulfate reducing. Small-subunit rRNA genes were amplified directly from aquifer material DNA by PCR with universally conserved or Bacteria- or Archaea-specific primers and were cloned. A total of 812 clones were screened by restriction fragment length polymorphisms (RFLP), approximately 50% of which were unique. All RFLP types that occurred more than once in the libraries, as well as many of the unique types, were sequenced. A total of 104 (94 bacterial and 10 archaeal) sequence types were determined. Of the 94 bacterial sequence types, 10 have no phylogenetic association with known taxonomic divisions and are phylogenetically grouped in six novel division level groups (candidate divisions WS1 to WS6); 21 belong to four recently described candidate divisions with no cultivated representatives (OP5, OP8, OP10, and OP11); and 63 are phylogenetically associated with 10 well-recognized divisions. The physiology of two particularly abundant sequence types obtained from the methanogenic zone could be inferred from their phylogenetic association with groups of microorganisms with a consistent phenotype. One of these sequence types is associated with the genus Syntrophus; Syntrophus spp. produce energy from the anaerobic oxidation of organic acids, with the production of acetate and hydrogen. The organism represented by the other sequence type is closely related to Methanosaeta spp., which are known to be capable of energy generation only through aceticlastic methanogenesis. We hypothesize, therefore, that the terminal step of hydrocarbon degradation in the methanogenic zone of the aquifer is aceticlastic methanogenesis and that the microorganisms represented by these two sequence types occur in syntrophic association.     

Differentiation of Helicobacter pylori strains directly from gastric biopsy specimens by PCR-based restriction fragment length polymorphism analysis without culture

Recent studies have shown the usefulness of PCR-based restriction fragment length polymorphism (RFLP) analysis for differentiating Helicobacter pylori strains isolated by culture. For this study, a PCR-based RFLP assay was developed for directly typing H. pylori strains from gastric biopsy specimens. Nineteen gastric biopsy specimens obtained from patients undergoing endoscopy for gastrointestinal complaints were cultured for isolation of H. pylori. Genomic DNA preparations from these gastric biopsy specimens and the corresponding H. pylori isolates were tested by our PCR-based RFLP assay. The 1,179-bp H. pylori DNA fragments amplified by the PCR assay were digested with the restriction enzymes HhaI, MboI, and AluI and analyzed by agarose gel electrophoresis. HhaI, MboI, and AluI digestion produced 11, 10, and 6 distinguishable digestion patterns, respectively, from the 19 H. pylori isolates tested and generated 13, 11, and 6 different patterns, respectively, from the 19 gastric biopsy specimens. The patterns from 13 of the 19 gastric biopsy specimens matched those of the H. pylori isolates from the corresponding patients. The patterns from the remaining six biopsy specimens appeared to represent infection by two strains of H. pylori; the pattern of one strain was identical to that of the isolate from the corresponding patient. By combining all the restriction enzyme digestion patterns obtained by using HhaI, MboI, and AluI, we observed 19 distinct RFLP patterns from the 19 specimens. The results suggest that the PCR-based RFLP analysis method may be useful as a primary technique to identify and distinguish H. pylori strains directly from gastric biopsy specimens without culture of the organisms.     

Molecular diagnostics of clinical strains of filamentous Basidiomycetes

Eleven clinical and veterinary strains of filamentous Basidiomycetes were compared with 15 reference strains representing the orders Aphyllophorales and Agaricales. The methods used were restriction analysis of small subunit (18S) (SSU) rDNA and internal transcribed spacers (ITS) 1 and 2 and variable domain 9 (V9)-ITS1 sequencing. Six strains were found to belong to the teleomorph genera Schizophyllum or Coprinus, whereas five could not be identified unequivocally. A rapid diagnostic overview is obtained with HaeIII and HinfI digestion of the ITS region.