Showing papers on "Typing published in 1997"
TL;DR: A rapid method to improve the response time and provide specific bacterial strain identification for the typing of Escherichia coli O157:H7 and other gram-negative organisms in a single day is described.
Abstract: Genomic DNA patterns generated by pulsed-field gel electrophoresis are highly specific for different strains of an organism and have significant value in epidemiologic investigations of infectious-disease outbreaks. Unfortunately, time-consuming and tedious specimen processing is an inherent problem which limits the use of this powerful technology as a real-time epidemic investigational tool. Here, I describe a rapid method to improve the response time and provide specific bacterial strain identification for the typing of Escherichia coli O157:H7 and other gram-negative organisms in a single day.
509 citations
TL;DR: The results of this study support the previous finding that type II strains are most often associated with human toxoplasmosis and provide rapid assignment of T. gondii to a specific genotype that should be useful in analyzing a variety of clinical samples.
Abstract: To determine the genotypes of Toxoplasma gondii strains associated with human toxoplasmosis, we developed a sensitive approach for typing parasites grown from clinical samples by short-term in vitro culture. A newly described nested PCR assay was capable of amplifying genomic DNA from as few as five parasites in the presence of host tissues. Typing was based on DNA polymorphisms at the SAG2 locus, encoding tachyzoite surface antigen p22. Restriction fragment length polymorphisms in PCR-amplified SAG2 products were used to classify strains into one of the three major lineages of T. gondii. This approach was successfully used to determine the genotypes of 68 of 72 samples that had been previously isolated from patients with congenital, cerebral, and disseminated toxoplasmosis. Type II strains of T. gondii were found in a majority of the samples, accounting for 55 (81%) of the 68 toxoplasmosis cases. In contrast, type I and III strains were found in only 7 (10%) and 6 (9%) of the 68 cases, respectively. The results of this study support the previous finding that type II strains are most often associated with human toxoplasmosis. Nested PCR analysis at the SAG2 locus provides rapid assignment of T. gondii to a specific genotype that should be useful in analyzing a variety of clinical samples.
507 citations
TL;DR: Strain typing cannot be used to replace a sound epidemiological investigation, but serves as a useful adjunct to such investigations as an integral part of epidemiological investigations of nosocomial infections.
Abstract: Strain typing is an integral part of epidemiological investigations of nosocomial infections. Methods for distinguishing among bacterial strains have improved dramatically over the last 5 years, due mainly to the introduction of molecular technology. Although not all molecular techniques are equally effective for typing all organisms, pulsed-field gel electrophoresis is the technique currently favored for most nosocomial pathogens. Criteria to aid epidemiologists in interpreting results have been published. Nucleic acid amplification-based typing methods also are applicable to many organisms and can be completed within a single day, but interpretive criteria still are under debate. Strain typing cannot be used to replace a sound epidemiological investigation, but serves as a useful adjunct to such investigations.
495 citations
TL;DR: Strain typing is an integral part of epidemiological investigations of nosocomial infections as discussed by the authors, and it has improved dramatically over the last 5 years, due mainly to the introduction of molecular technology.
Abstract: Strain typing is an integral part of epidemiological investigations of nosocomial infections. Methods for distinguishing among bacterial strains have improved dramatically over the last 5 years, due mainly to the introduction of molecular technology. Although not all molecular techniques are equally effective for typing all organisms, pulsed-field gel electrophoresis is the technique currently favored for most nosocomial pathogens. Criteria to aid epidemiologists in interpreting results have been published. Nucleic acid amplification-based typing methods also are applicable to many organisms and can be completed within a single day, but interpretive criteria still are under debate. Strain typing cannot be used to replace a sound epidemiological investigation, but serves as a useful adjunct to such investigations.
377 citations
262 citations
TL;DR: The random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) was shown to be a convenient typing method for S. marcescens for the first time, and 73 different RAPD patterns with good reproducibility were obtained.
Abstract: Serratia marcescens has emerged in the last few years as an important nosocomial pathogen. Many methods for typing this organism have been described. In this study the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) was shown to be a convenient typing method for S. marcescens. Different combinations of primers previously used for typing other gram-negative bacilli were assessed. The combination of primer HLWL-74 and 1254 gave distinguishable patterns for different serotypes and proved to be the most satisfactory. By applying this combination to 175 isolates of S. marcescens, which could be classified into 38 groups on the basis of serotyping and phage typing, 73 different RAPD patterns with good reproducibility were obtained. This is, to our knowledge, the first application of the method to a large collection of S. marcescens representing a wide range of serotypes.
225 citations
111 citations
TL;DR: The RAPD technique revealed intraserotype-specific variations, undetectable by the combination of several phenotypic typing methods, among the ETEC strains analyzed, showing that RAPD typing represents a useful tool for population genetics as well as for epidemiological studies of ETEC.
Abstract: The genetic diversity of 47 enterotoxigenic Escherichia coli (ETEC) strains of serotypes O6:H16, O27:H7, O29:H21, O128ac:H12, and O153:H45, previously isolated from diarrheic patients in Brazil over a period of 15 years, was investigated by random amplification of polymorphic DNA (RAPD). Informative band arrays were obtained with three 10-mer primers with G+C contents of 50, 60, and 70%. Based on the combination of the band profiles generated by the three primers 22 RAPD types were detected, and 5 major clonal clusters, each one with at least 80% identical bands, were established. The clonal clusters corresponded to strains having the same serotype which, in most cases, also had the same virulence factors (colonization factors and toxin types) and outer membrane protein and lipopolysaccharide sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles. The results suggested a correlation between phenotypic properties and genetic relatedness of ETEC isolates of human origin and indicated that a reduced number of clonally related strains are found in areas of ETEC endemicity in Brazil. Moreover, the RAPD technique revealed intraserotype-specific variations, undetectable by the combination of several phenotypic typing methods, among the ETEC strains analyzed. These results show that RAPD typing represents a useful tool for population genetics as well as for epidemiological studies of ETEC.
102 citations
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01 Jan 1997
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89 citations
TL;DR: RS-PCR is a rapid, inexpensive technique that is highly reproducible and almost as discriminatory as PFGE for typing MRSA isolates and should be useful in the local investigation of MRSA outbreaks.
Abstract: Analysis of sequences in the fragments of the 16S-23S rRNA intergenic spacer region by the ribosome spacer PCR (RS-PCR) can differentiate strains of methicillin-resistant Staphylococcus aureus (MRSA). We compared this technique with pulsed-field gel electrophoresis (PFGE) for typing MRSA strains and its application during an investigation of an outbreak. A total of 180 isolates of MRSA collected from various hospital laboratories within the United Kingdom and elsewhere were typed by PFGE and RS-PCR. PFGE identified 17 different types among the 180 strains examined, and RS-PCR generated 13 different types. PFGE could detect minor genetic variations among the isolates and could identify the variants which were not discriminated by RS-PCR. Four unique strain types detected by PFGE were not detected by RS-PCR. When applied to typing the outbreak-related strains from the vascular surgery unit at the General Infirmary at Leeds, the results of RS-PCR were identical to those of PFGE. Our results have shown that RS-PCR is a rapid, inexpensive technique that is highly reproducible and almost as discriminatory as PFGE for typing MRSA isolates and should be useful in the local investigation of MRSA outbreaks.
TL;DR: A total of 98 isolates of group A streptococci of T1M1 serotype isolated in Finland during 1988-1995 from bacteremic, pharyngeal, and pyogenic infections were studied by molecular means.
Abstract: A total of 98 isolates of group A streptococci of T1M1 serotype isolated in Finland during 1988-1995 from bacteremic, pharyngeal, and pyogenic infections were studied by molecular means The typing techniques used included restriction endonuclease analysis, ribotyping, and random amplified polymorphic DNA analysis Representatives of each observed genotype were also examined for carriage of the speA gene encoding for pyrogenic exotoxin A All of the serotype T1M1 isolates studied were considered to be of a single clonal origin, and 66% were identical by all three genotyping techniques used Among the remaining 34% of closely related isolates, 13 different genotypes were detected All isolates studied carried the speA gene No evidence was found of correlation of the genomic type to the severity of the infection or the time of isolation
TL;DR: Comparisons between SBT of DRB1/3/4/5 alleles performed in two laboratories each using a different strategy for SBT concluded that nor the method of sequencing nor the sequencing machine influences typing results.
Abstract: Sequence-based typing (SBT) has become an important tool in the identification of HLA alleles. In this study a comparison was made between SBT of DRB 1/3/4/5 alleles performed in two laboratories each using a different strategy for SBT. The laboratories in Utrecht and in Maastricht performed direct sequencing of PCR amplified genomic DNA from 30 selected samples. Primers and conditions for PCR amplification were different. Sequencing was either performed with T7 polymerase, using internal sequencing primers, or with cycle sequencing using an M13 tailed system. Two different automated DNA sequencers were used; the ALFexpress from Pharmacia and Applied Biosystems 373 A. We concluded that nor the method of sequencing nor the sequencing machine influences typing results. However the PCR reaction used for generating template DNA is the most critical step. Different primers and different conditions can lead to false negative reactions. The fact that these false negative reactions can occur with different alleles in different combinations but not in all, implicates that extensive quality control is needed to assure correct typing results.
TL;DR: Sequence-based typing of the DQB1 gene proved a reliable typing strategy for assignment of the different D QB1 alleles after intensive selection of primers and test conditions.
Abstract: The high degree of polymorphism of the HLA genes at the nucleotide sequence level has proven sequence-based typing a major typing strategy. For DRB1 the allelic variability is predominantly present in the second exon and by DNA sequencing of exon 2 all hitherto known DRB1 alleles can be detected. For the associated genes DRB3, DRB4 and DRB5 the situation is slightly different. Allelic differences are not limited to exon 2 and the sequence of exon 3 and sometimes exon 4 is needed for complete subtyping. Oligonucleotides to amplify the exons needed for subtyping of DRB3, DRB4 and DRB5 were designed. Gene-specific products were generated to make simultaneous detection of alleles in heterozygous combinations possible. In this way 238 individuals were fully typed for their DRB3, 4 and 5 subtypes. Additional samples were typed for only one of the genes. All samples had been previously typed by PCR-SSP. Concordant typing results were obtained for all individuals tested. The DRB3 alleles typed for included *0101, *0201, *0202 and *0301, for DRB4 they were *01011, *0102 and *0103 and for DRB5 *0101, *0102, *0103, *0105, *0201, *0202 and *0203. All alleles were easily detected by the protocol described except for DRB5*0201. Sequencing of exon 3 and 4 of the DRB5*0201 allele showed this allele to be a sequencing error and the sequences obtained were identical to the exon 2, 3 and 4 sequences of DRB5*0202. Two new alleles were identified in the samples studied, DRB4*0105 and DRB3*0207. Sequence based typing has been recognized as a valuable tool for HLA typing of DRB1, DQB1 and DPB1 since several years. It is shown to be a superior typing method as well in the detection of the different DRB3, 4 and 5 subtypes.
TL;DR: PCR-based analysis of inter-IS256 spacer polymorphisms was developed for typing of methicillin-resistant Staphylococcus aureus and S. epidermidis strains and showed complete typeability and good discriminatory power, and is proposed as an efficient molecular typing assay for epidemiological studies of MRSA.
Abstract: IS256 elements are present in multiple copies in the staphylococcal genome, either flanking the transposon Tn4001 or independent of it. PCR-based analysis of inter-IS256 spacer polymorphisms was developed for typing of methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus epidermidis strains. Using SmaI macrorestriction analysis resolved by pulsed-field gel electrophoresis (PFGE) as the reference method for MRSA typing, excellent reproducibility (100%), discriminatory power (97%), and in vivo stability were observed. Good concordance of the results with those of other molecular typing methods was found for two MRSA collections. Inter-IS256 PCR analysis of a U.S. collection of MRSA strains (n = 36), previously characterized by 15 typing methods, showed more limited discrimination. Agreement was 78% with PFGE analysis and 83% with ribotyping (HindIII). Analysis of a second set of Belgian MRSA strains (n = 17), categorized into two widespread epidemic clones by PFGE analysis, showed 65% agreement. For typing of S. epidermidis strains (n = 26), inter-IS256 PCR showed complete typeability (100%) and good discriminatory power (85%). Inter-IS256 PCR analysis is proposed as an efficient molecular typing assay for epidemiological studies of MRSA or S. epidermidis isolates.
TL;DR: Multitarget typing of P. carinii hominis by PCR-SSCP should allow investigation of strain diversity and thus be useful for future epidemiological studies.
Abstract: To better investigate Pneumocystis carinii f. sp. hominis epidemiology, we have developed a molecular typing method. Because of the limited genetic variability of the P. carinii hominis genome, a multitarget approach was used. Four variable regions of the genome were amplified by PCR, polymorphism in each region was assessed by the single-strand conformation polymorphism (SSCP) technique, and the results for the four regions of each patient were combined. Bronchoalveolar lavage specimens collected from 11 patients were examined. Four patients were probably infected by a single strain, since their specimens yielded simple SSCP patterns (two bands corresponding to one allele). The combinations of these patterns were unique, suggesting that the strains which infected these patients were different. For the other seven patients, complex patterns were found (three or four bands corresponding to two alleles). The presence of more than one allele of a region in a patient is likely to be due to coinfection. Polymorphism was also assessed by sequencing, which revealed variations at nucleotide positions previously reported to vary. About half of the observed alleles had already been reported by laboratories in different countries. Multitarget typing of P. carinii hominis by PCR-SSCP should allow investigation of strain diversity and thus be useful for future epidemiological studies.
TL;DR: RAPD typing was found to be a simple, rapid, and effective method for the epidemiological investigation of this outbreak, and performance of typing by this method was simpler and less time-consuming than that of Typing by PFGE.
Abstract: A cluster of methicillin-resistant Staphylococcus aureus (MRSA) infections among patients on an intensive care unit (ICU) was detected by routine infection control surveillance. In the period from 5 January to 22 June 1995, 10 patients on the ICU and a further 6 patients (5 on one ward that had received colonized patients transferred from the ICU) were affected by MRSA strains with the same antibiotic susceptibility patterns. Seven (44%) of these 16 colonized patients developed MRSA bacteremia. MRSA isolates with the same characteristics were also found on the hands of one member of the ICU staff. The isolates were untypeable by phage typing, but 15 of 17 outbreak strains analyzed genetically had identical randomly amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) profiles. A single strain of MRSA that was nontypeable by phage typing and that was isolated on the ICU on 1 January and six nontypeable and epidemiologically unrelated MRSA isolates all had RAPD profiles distinct from that of the outbreak strain. Implementation of strict infection control measures stopped the further spread of MRSA on the ICU, the affected general ward, and seven other wards that received MRSA carriers from the ICU. Although nontypeable by phage typing and not previously recognized as an epidemic strain, this strain of MRSA was readily transmissible and highly virulent. RAPD typing was found to be a simple, rapid, and effective method for the epidemiological investigation of this outbreak, and performance of typing by this method was simpler and less time-consuming than that of typing by PFGE. RAPD typing may have more general application for the study of S. aureus infections in hospitals.
TL;DR: N nosocomial outbreaks of C. difficile diarrhea in five hospitals were associated with a single genetic lineage as resolved by multiple strain typing systems.
Abstract: In a collaborative study by three laboratories, arbitrarily primed polymerase chain reaction (AP-PCR), HindIII restriction enzyme analysis (REA), and pulsed-field gel electrophoresis (PFGE) using SmaI were compared for typing of Clostridium difficile. The study included 30 isolates from nosocomial outbreaks in six geographically disparate hospitals and 15 isolates from sporadic cases of C. difficile diarrhea. REA distinguished a total of 23 types representing 10 groups; AP-PCR performed at Deaconess Hospital resolved 19 types; AP-PCR performed at the Centers for Disease Control resolved 15 types. Thirty isolates exhibited degradation of larger sized fragments during processing and therefore were nontypeable by PFGE; among the remaining 15 isolates, PFGE resolved 11 types. Outbreak isolates in five different hospitals represented REA group J and constituted a single AP-PCR strain. In summary, nosocomial outbreaks of C. difficile diarrhea in five hospitals were associated with a single genetic lineage as resolved by multiple strain typing systems.
TL;DR: This study showed that A. junii is capable of causing a serious, though non-fatal infection in neonates and it is concluded that for a better understanding of the role of the various Acinetobacter genomic species in human pathology, identification of acinetobacters according to the recent taxonomy is imperative.
TL;DR: The data refute the idea of a clonal origin for fluconazole-resistant strains among HIV-positive patients and demonstrate that at least two different molecular methods are needed for Candida albicans typing and that there is a great deal of strain variation within the species, irrespective of place of origin or antifungal resistance patterns.
Abstract: Candida albicans strain delineation by karyotyping. NotI restriction pattern analysis, hybridization with specific probe 27A, and PCR fingerprinting with the phage M13 core sequence were performed with 30 isolates from the oral cavities of 30 human immunodeficiency virus (HIV)-infected patients and 8 reference strains. Within the panel of clinical isolates, 20 were geographically related, although 10 isolates were susceptible to fluconazole and 10 isolates were resistant to fluconazole. The remaining isolates used in this study were fluconazole resistant and geographically unrelated. A composite DNA type was defined for each of the strains as the combination of types obtained by the four molecular methods. By this procedure, a great diversity of DNA types was found among isolates from the oropharynges of HIV-infected individuals with oral candidiasis. This diversity was not reduced when isolates were evaluated on the basis of whether they came from the same geographical locale and whether they were fluconazole resistant. These data refute the idea of a clonal origin for fluconazole-resistant strains among HIV-positive patients. Karyotyping was the least discriminatory method, yielding 19 DNA types among the 38 strains analyzed. Conversely, hybridization with the 27A probe showed a unique DNA pattern for each of the strains examined in this study. Our results demonstrate that at least two different molecular methods are needed for Candida albicans typing and that there is a great deal of strain variation within the species, irrespective of place of origin or antifungal resistance patterns.
TL;DR: This investigation identified 71 RAPD types and the three families of strains defined by multilocus enzyme electrophoresis analysis, which contain most of the cerebrospinal fluid isolates, were also identified by clustering analysis of RAPD data.
Abstract: A collection of 54 unrelated Streptococcus agalactiae strains isolated from cerebrospinal fluid samples from neonates and 60 unrelated strains isolated from carriers that had been previously studied by multilocus enzyme electrophoresis (R. Quentin, H. Huet, F.-S. Wang, P. Geslin, A. Goudeau, and R. K. Selander, J. Clin. Microbiol. 33:2576-2581, 1995) were characterized by randomly amplified polymorphic DNA (RAPD) assay. Four primers, 5'AGGGGGTTCC3', 5'AACGCGCAAC3', 5'GCATCAATCT3', and 5'AGTCGGGTGG3', named OPS16, AP42, A4, and OPS11, respectively, were selected from 29 primers tested. This investigation identified 71 RAPD types. The three families of strains defined by multilocus enzyme electrophoresis analysis, which contain most of the cerebrospinal fluid isolates, were also identified by clustering analysis of RAPD data. Each of these three groups exhibits specific RAPD patterns or fragments. The discriminatory power of the RAPD typing method was also evaluated. The simplest typing scheme was obtained by the combination of RAPD typing done with primers AP42 and OPS11 and serotyping (index of discrimination, 0.97).
TL;DR: Antibiotyping and AluI restriction fragment length polymorphism analysis of the coagulase gene were able to define clones in the same way as DNA macrorestriction analysis (SmaI), but both techniques presented disadvantages, making neither of them useful as a single typing method.
Abstract: Forty-three methicillin-resistant Staphylococcus aureus (MRSA) isolates with known genetic and epidemiological relatedness and different degrees of transmission were analyzed by antibiotyping, protein A gene polymorphism analysis, and coagulase gene polymorphism analysis. The three typing systems were evaluated for their performance and convenience to define clones and to discriminate between epidemic MRSA (EMRSA) and sporadic MRSA (SMRSA). Antibiotyping and AluI restriction fragment length polymorphism analysis of the coagulase gene were able to define clones in the same way as DNA macrorestriction analysis (SmaI). However, both techniques presented disadvantages, making neither of them useful as a single typing method. Protein A gene polymorphism analysis appeared to be of no value for clonal analysis. None of the three typing methods was able to differentiate between EMRSA and SMRSA.
TL;DR: The SBT approach described here uses a locus specific amplification of DNA from exon 1 to exon 5 to identify new alleles directly and an identical new "HLA-A*0103" was identified in two Caucasian samples.
TL;DR: The results of this study show the extreme genomic diversity of H. pylori and indicate that infection by H.pylori in the United States does not appear to be limited to a small number of RFLP types.
Abstract: The molecular typing of 81 pretreatment Helicobacter pylori isolates and the comparison of 18 pretreatment-posttreatment pairs is described by restriction fragment length polymorphism (RFLP) of the ureC gene. The results of our study show the extreme genomic diversity of H. pylori and indicate that infection by H. pylori in the United States does not appear to be limited to a small number of RFLP types.
TL;DR: This project addressed the question whether molecular typing for HLA class I also increases the efficacy of HLA matching in kidney transplantation and found that 183 DNA class I compatible transplants had a 15% higher one-year graft survival rate than 32 transplants for which DNA typing revealed a class I incompatibility.
Abstract: DNA typing for HLA class II improves the typing quality and this was shown previously to be relevant for kidney graft survival. In this project we addressed the question whether molecular typing for HLA class I also increases the efficacy of HLA matching in kidney transplantation. 215 HLA-A,-B,-DR zero-mismatched donor/recipient pairs as defined by serological typing were selected. Retrospective HLA-A and HLA-B typing was performed both by the PCR-SSP and the PCR-SSOP method. DNA typing for HLA-A revealed discrepant results to serology in 5.7% of the donors and 2.8% of the recipients. HLA-B typing discrepancies were found in 6.6% of the donors and 5.6% of the recipients. 10.4% of the donors and 6.5% of the recipients showed either an HLA-A or an HLA-B discrepancy Nearly one-third of the HLA-A discrepancies affected A19 splits. The most common reason for HLA-A discrepancies was the erroneous assignment of serological blanks, whereas HLA-B errors were caused mainly by the assignment of incorrect specificities. DNA typing allowed the definition of HLA-A and -B split specificities in all 118 "splitable" cases for which only broad specificities were reported based on serological typing. A total of 183 DNA class I compatible transplants had a 15% higher one-year graft survival rate than 32 transplants for which DNA typing revealed a class I incompatibility
TL;DR: Frequency information, based on a population of 5,000 individuals, has been established using a combination of molecular and serological typing data and PCR-SSOP detected the presence of a second HLA-A allele in over 10% of individuals who had been previously homozygous.
Abstract: A medium resolution PCR-SSOP typing method, using 26 digoxigenin labelled probes, has been established for the identification of HLA-A alleles. The system is capable of discriminating all of the serologically defined specificities except for eight heterozygous combinations which are however rare in Caucasians. The method has been applied to 1,838 individuals on the local bone marrow registry who either had only one detectable HLA-A antigen, or a HLA-A antigen whose presence had been queried using the serological technique or a broad HLA-A specificity assigned by the serological technique. In all but one case the serologically assigned antigens were detected with the PCR-SSOP method. In addition, PCR-SSOP detected the presence of a second HLA-A allele in over 10% of individuals who had been previously homozygous. Frequency information, based on a population of 5,000 individuals, has been established using a combination of molecular and serological typing data.
TL;DR: AP-PCR analysis provides a simple and practical approach to typing P. aeruginosa that is more discriminatory than traditional serotyping scheme and it is suggested that maximum discrimination can be achieved by a combination of both methods.
Abstract: Arbitrary primed PCR (AP-PCR) analysis was compared with serotyping as a means of high-resolution typing of Pseudomonas aeruginosa Seventy-four isolates from 3 different hospitals and 18 reference strains were studied Serotyping provided good index of discrimination, although eleven isolates could not be serotyped Genomic DNA was amplified with a single 10 nucleotide primer (sequence 5′-AGG GGT CTT G-3′) The strains were genetically diverse and 61 different AP-PCR profiles of 2–7 bands between 03 and 24 kb were obtained AP-PCR profiles were not consistently associated with serotypes, but they clearly subtyped strains of the same serotype Numerical analysis of AP-PCR patterns defined 7 groups at the 55% similarity level, and identified predominant strains in each hospital The results show that AP-PCR analysis provides a simple and practical approach to typing P aeruginosa that is more discriminatory than traditional serotyping scheme We suggest that maximum discrimination can be achieved by a combination of both methods
TL;DR: Typing systems for DNA profiling are predominantly selected according to criteria related to the robustness for typing of (potentially degraded) forensic specimens, the degree of genetic polymorphism, and the amenability to standardisation as a basis to obtain reproducible results.
TL;DR: The results suggest that seven lung transplant patients with necrotizing aspergillosis, bronchitis asper gillosis and bronchial colonization were infected by multiple strain genotypes, whereas three patients with invasive asperGillosis seem to have been infected by a single strain.
Abstract: A PCR typing method has been developed and tested to investigate the polymorphism of clinical strains of Aspergillus fumigatus. Firstly, the DNA fragments from random amplified polymorphic DNA (RAPD) patterns of nine epidemiologically and geographically non-related monosporal strains of A. fumigatus were cloned and sequenced. The pairs of five sequence-specific DNA primers (SSDP), characteristic of the 5′ and 3′ extremities of the RAPD products, were then used in high stringency PCR to type 43 clinical strains of A. fumigatus from 13 patients, according to the presence or absence of a single amplified band. This original approach, which uses the advantages of PCR, has made it possible to overcome the difficulties resulting from the low stringency amplification. The SSDP analysis of 51 A. fumigatus strains (9 unrelated monosporal strains and 43 clinical strains from 13 patients) can be classed into 22 different types with a high reproducibility and a high level of discrimination (D=0.96). The results suggest that seven lung transplant patients with necrotizing aspergillosis, bronchitis aspergillosis and bronchial colonization were infected by multiple strain genotypes, whereas three patients with invasive aspergillosis seem to have been infected by a single strain.