Polymerase Chain Reaction Typing Research Articles

Enterobacterial intergenic consensus sequence polymerase chain reaction as a typing method for Burkholderia (Pseudomonas) cepacia

OBJECTIVE: We developed a rapid polymerase chain reaction (PCR) method, which utilizes primers from an enterobacterial repetitive consensus sequence, to differentiate among strains of Burkholderia cepacia. We then applied this method to isolates from a device-associated pseudoinfection and also compared the discriminative typing potential with that of another PCR-based method. METHODS: A simplified lysis procedure was used to generate DNA substrate for amplification by a PCR method which utilized primer ERIC2 and a reverse complement of ERIC1R. The alternative PCR-based method employed primers which are directed to Bordetella pertussis repetitive sequences. Amplification products were visually assessed after conventional agarose gel electrophoresis and staining. RESULTS: The ERIC-based method appropriately clustered strains for the device-associated pseudo-infection. In addition, there was consistency in the definition of similar or dissimilar strains between the two PCR---based systems. CONCLUSIONS: The ERIC-based PCR typing method offers sufficient discriminatory power for it to be used for epidemiologic purposes.

Repetitive element sequence based polymerase chain reaction for typing of Brucella strains

A collection of 40 Brucella strains, including 19 clinical B.canis isolates, was analyzed using ERIC-PCR and REP-PCR. PCR amplification using primers REP 1R-I and REP 2-I provided distinct patterns for Brucella spp. All Brucella strains could be discriminated at least to the species level. ERIC-PCR, employing ERIC 1R and ERIC 2 as primers was less discriminative for Brucella, only allowing a discrimination to the genus level with occasional discrimination between individual strains. It can be concluded that rep PCR is a promising fingerprinting method for the evaluation of an outbreak.

PCR typing of DNA fragments of the two short tandem repeat (STR) systems upstream of the human myelin basic protein (MBP) gene in Danes and Greenland Eskimos

DNA from the double short tandem repeat (STR) system MBP (locus 18q23-pter) was amplified by the polymerase chain reaction (PCR) and the two polymorphic repeat systems were separated by cutting with the restriction enzyme NlaIII. The lengths of the DNA fragments of the two MBP STR systems MBP-A and MBP-B were analyzed by vertical electrophoresis in polyacrylamide gels followed by silver staining. DNA samples from 112 unrelated Danes, 140 unrelated Greenland Eskimos, and 88 Danish mother/child pairs were analyzed. The distributions of MBP phenotypes were in Hardy-Weinberg equilibrium in both the Eskimo and Danish populations. Significant differences were observed between the distribution of fragments (‘alleles’) in Greenland Eskimos and in Danes. The allele MBP-A7 was considerably more frequent in Eskimos (0.2214) than in Danes (0.0775) and also the allele MBP-B9 was considerably more frequent in Eskimos (0.225) than in Danes (0.06). Strong gametic associations were found between fragments from the MBP-A and MBP-B series in both Danes and Eskimos. Some of the associations were different in Danes and Eskimos. In the 88 Danish mother/child pairs, the segregation of the MBP genotypes were in accordance with a genetic model of co-dominant inheritance and no mutation was found. Two MBP STR regions with irregular structures were sequenced. One fragment had a single base G to A transition at position 124 in the primer binding region between the MBP-A and MBP-B regions. In the other fragment, a deletion starting at position 117 and including the primer binding region between the MBP-A and MBP-B regions was found.

Origin and interstate spread of a New York City multidrug-resistant Mycobacterium tuberculosis clone family

To determine whether isolates of Mycobacterium tuberculosis from New York and elsewhere that are resistant to four or more primary antimicrobial agents and responsible for widespread disease in the 1990s represent a newly emerged clone or a heterogeneous array of unrelated organisms. New York City area and selected locations in the United States. M tuberculosis isolates from 1953 patients in New York and multidrug-resistant isolates from six patients from other US communities. Convenience sample of all M tuberculosis strains (M tuberculosis isolates resistant to rifampin, streptomycin, isoniazid, and ethambutol, and sometimes ethionamide, kanamycin, capreomycin, or ciprofloxacin) submitted to the Public Health Research Institute Tuberculosis Center since 1991 and samples submitted to the Centers for Disease Control and Prevention from throughout the United States. The samples submitted were representative of the New York City strains of M tuberculosis. Characterization of resistant M tuberculosis strains studied by IS6110 and polymorphic GC-rich repetitive sequence (PGRS) hybridization patterns, multiplex polymerase chain reaction (PCR) analysis, and automated DNA sequencing of genes containing mutations associated with resistance to rifampin (rpoB), isoniazid (katG and inhA locus), and streptomycin (strA and rrs). Multidrug-resistant M tuberculosis isolates were recovered from 253 New York City patients and had the same or closely allied IS6110 and PGRS patterns, multiplex PCR type, and gene mutations associated with resistance to rifampin, isoniazid, and streptomycin. Isolates with these same molecular characteristics were recovered from patients in Florida and Nevada, health care workers in Atlanta, Ga, and Miami, Fla, and an individual who recently moved from New York City to Denver, Colo, and caused disease or skin test conversion in at least 12 people in a nursing home environment. The results document the molecular origin and spread of progeny of a closely related family of multidrug-resistant M tuberculosis strains that have recently shared a common ancestor and undergone clonal expansion. The multidrug-resistant phenotype in these organisms arose by sequential acquisition of resistance-conferring mutations in several genes, most likely as a consequence of antibiotic selection of randomly occurring mutants in concert with inadequately treated infections. Dissemination of these difficult-to-treat bacteria throughout New York City and to at least four additional US cities has adverse implications for tuberculosis control in the 21st century.

Origin and Interstate Spread of a New York City Multidrug-Resistant Mycobacterium tuberculosis Clone Family

<h3>Objective.</h3> —To determine whether isolates of<i>Mycobacterium tuberculosis</i>from New York and elsewhere that are resistant to four or more primary antimicrobial agents and responsible for widespread disease in the 1990s represent a newly emerged clone or a heterogeneous array of unrelated organisms. <h3>Setting.</h3> —New York City area and selected locations in the United States. <h3>Patients.</h3> —<i>M tuberculosis</i>isolates from 1953 patients in New York and multidrug-resistant isolates from six patients from other US communities. <h3>Design.</h3> —Convenience sample of all<i>M tuberculosis</i>strains (<i>M tuberculosis</i>isolates resistant to rifampin, streptomycin, isoniazid, and ethambutol, and sometimes ethionamide, kanamycin, capreomycin, or ciprofloxacin) submitted to the Public Health Research Institute Tuberculosis Center since 1991 and samples submitted to the Centers for Disease Control and Prevention from throughout the United States. The samples submitted were representative of the New York City strains of<i>M tuberculosis</i>. <h3>Main Outcome Measure.</h3> —Characterization of resistant<i>M tuberculosis</i>strains studied by IS<i>61 10</i>and polymorphic GC-rich repetitive sequence (PGRS) hybridization patterns, multiplex polymerase chain reaction (PCR) analysis, and automated DNA sequencing of genes containing mutations associated with resistance to rifampin (<i>rpoB</i>), isoniazid (<i>katG</i>and<i>inhA</i>locus), and streptomycin (<i>strA</i>and<i>rrs</i>). <h3>Results.</h3> —Multidrung-resistant<i>M tuberculosis</i>isolates were recovered from 253 New York City patients and had the same or closely allied IS<i>6110</i>and PGRS patterns, multiplex PCR type, and gene mutations associated with resistance to rifampin, isoniazid, and streptomycin. Isolates with these same molecular characteristics were recovered from patients in Florida and Nevada, health care workers in Atlanta, Ga, and Miami, Fla, and an individual who recently moved from New York City to Denver, Colo, and caused disease or skin test conversion in at least 12 people in a nursing home environment. <h3>Conclusions.</h3> —The results document the molecular origin and spread of progeny of a closely related family of multidrug-resistant<i>M tuberculosis</i>strains that have recently shared a common ancestor and undergone clonal expansion. The multidrug-resistant phenotype in these organisms arose by sequential acquisition of resistance-conferring mutations in several genes, most likely as a consequence of antibiotic selection of randomly occurring mutants in concert with inadequately treated infections. Dissemination of these difficult-to-treat bacteria throughout New York City and to at least four additional US cities has adverse implications for tuberculosis control in the 21st century. (<i>JAMA</i>. 1996;275:452-457)

Primer Master: a new program for the design and analysis of PCR primers.

The polymerase chain reaction (PCR) is one of the most rapidly expanding methods in molecular biology. Although the range of applications of PCR is very broad, the key points for successful performance remain the careful selection of optimal primers and the proper determination of the temperature conditions of the reaction. The program presented here, 'Primer Master', has been developed to assist in solving these problems for two major types of PCR: the amplification of the appropriate fragments of several related, but not identical, sequences and the specific amplification of a certain fragment of only one nucleotide sequence where amplification of any other sequence needs to be prevented. Unlike other programs developed for the design of PCR primers, 'Primer Master' undertakes an automatic search and selection of the optimal primers and primer pairs for both types of reaction. The wide applicability of'Primer Master' is determined by two main automatic search regimes: 'Universal' and 'Specific'. The 'Universal' regime searches and selects primers and primer pairs which amplify the designated fragments of all required sequences (consensus primers) and allows a 'combined search', where the universal primers for a particular group of sequences must not amplify any sequence from another group. The number of sequences allowed is 2 to 100, with a maximum length of 65 000 nucleotides. The 'Specific' regime is designed to select the primers and primer pairs which amplify the fragment of a specific nucleotide sequence from all those the user wishes to consider. The total number of sequences to be tested is unlimited, and sequences up to 32000 nucleotides in length are allowed. 'Primer Master' also incorporates a number of special features designed to overcome problems common to PCR. Within the 'Specific' regime it may be necessary to find primers from one of several related sequences. However, if the sequences span different positions then there is the risk that the primers chosen may include regions absent from

Investigation of an outbreak of Campylobacter upsaliensis in day care centers in Brussels: analysis of relationships among isolates by phenotypic and genotypic typing methods.

An outbreak of Campylobacter upsaliensis in four Brussels day care centers (A, B-1, B-2, and C) affected 44 children. Diarrhea was the major symptom. From January 1991 to June 1992, the outbreak strain was isolated from 3, 1, and 21 (of 68) children in centers A, B-1, and B-2, respectively, and from 19 of 22 children in center C, IgG, IgM, and IgA antibodies were detected by Western blotting of serum specimens of 9 of 10 and 13 of 16 children in centers B-2 and C, respectively. Strains were typed by biotyping, DNA restriction-based and antibiotic susceptibility typing, whole cell protein and plasmid analysis, restriction fragment length polymorphism (RFLP), and polymerase chain reaction (PCR). On the basis of RFLP and PCR typing, the strains could be divided into two strongly related clonal variants: One was isolated only from the children of center A and the second only from children in the other day care centers.

Arab Population Data on the PCR-Based Loci: HLA-DQA1, LDLR, GYPA, HBGG, D7S8, Gc, and D1S80

Allele and genotype frequencies for seven polymerase chain reaction (PCR)-based DNA genetic markers were determined in an Arab sample population. The loci analyzed were HLA-DQA1, LDLR, GYPA, HBGG, D7S8, Gc and D1S80. Results were obtained from the first six loci using the AmpliType HLA-DQ alpha DNA and AmpliType PM PCR Amplification and Typing Kits. The VNTR locus D1S80 PCR product was analyzed by polyacrylamide electrophoresis and silver staining. All loci meet Hardy-Weinberg expectations. The frequency data can be used in forensic analyses and paternity tests to estimate the frequency of a DNA profile in the Arab population.

Chinese population data on three tetrameric short tandem repeat loci — HUMTHO1, TPOX, and CSF1PO — derived using multiplex PCR and manual typing

Allele and genotype frequencies for three tetrameric short tandem repeat (STR) loci were determined in a Chinese sample population using multiplex polymerase chain reaction (PCR), electrophoresis of the PCR products in DNA sequencing gels and subsequent detection by silver staining. The loci are HUMTHO1, TPOX, and CSF1PO. All loci meet Hardy-Weinberg expectations. In addition, there is no evidence for association of alleles among the three loci. The allelic frequency data can be used in human identity testing to estimate the frequency of a multiple STR locus DNA profile in the Chinese population.

Chinese population data on the PCR-based loci HLA-DQ alpha, low-density-lipoprotein receptor, glycophorin A, hemoglobin gamma G, D7S8, and group-specific component.

Allele and genotype frequencies for six polymerase chain reaction (PCR)-based DNA genetic markers were determined in a Chinese sample population. The loci are HLA-DQ alpha, low-density-lipoprotein receptor, glycophorin A, hemoglobin gamma G, D7S8, and group-specific component. These results were obtained using the AmpliType HLA-DQ alpha forensic DNA amplification and typing kit and the AmpliType PM PCR amplification and typing kit. All loci met Hardy-Weinberg expectations and there was no evidence for association of alleles between the loci. The frequency data can be used in forensic analyses and paternity tests to estimate the frequency of a DNA profile in the Chinese population.

PCR typing of DNA fragments of the short tandem repeat (STR) system HUMTH01 in Danes and Greenland Eskimos

DNA from the short tandem repeat (STR) system HUMTH01 was amplified by the polymerase chain reaction (PCR) and analyzed by vertical electrophoresis in polyacrylamide gels followed by silver staining. DNA samples from 100 unrelated Danes, 147 unrelated Greenland Eskimos, and 89 Danish mother/child pairs were analyzed. Significant differences were observed between the distribution of fragments (‘alleles’), whereby allele number 7 was considerably more frequent in Eskimos (0.687) than in Danes (0.201). The distributions of HUMTH01 phenotypes were in Hardy-Weinberg equilibrium in both the Eskimo and Danish populations. In the 89 Danish mother/child pairs, the segregation of the HUMTH01 genotypes was in accordance with the genetic model of co-dominant inheritance and no mutations were found. In a blind trial, DNA samples from 40 unrelated Danes were investigated by one of us in Copenhagen, at the FBI Forensic Science Research and Training Center (FSRTC), Quantico, and at the Institut für Rechtsmedizin, Münster, Germany. Concordant HUMTH01 types were found in 39 out of 40 individuals. The allele which led to discrepant typing results was assigned type 9.3 in two laboratories and type 10 in one laboratory.

PCR typing of the locus D17S30 (YNZ22 VNTR) in an Italian population sample.

A sample of 202 subjects living in 2 Italian provinces (Ancona and Parma) was tested for YNZ22 polymorphism by the polymerase chain reaction (PCR). After amplification, the phenotypes were separated by agarose gel electrophoresis, stained with ethidium bromide and identified by comparison with a molecular weight marker. No heterogeneity was found between the 2 populations. Alleles, pooled in 4 groups to calculate the Hardy-Weinberg (H-W) equilibrium, showed good accordance between observed and expected values. The power of discrimination (PD) was 0.95 and the chance of exclusion was 0.69. The allele comparison with previous studies on Caucasians showed no significant difference.

Comparison of nucleotide sequences among hantaviruses belonging to the same serotype: an analysis of amplified DNA by thermal cycle sequencing

The hantavirus genus, belonging to the bunyaviridae family, is comprised of at least four serologically distinct types: Hantaan, Seoul, Puumala and Prospect Hill. Previously, we reported the use of the polymerase chain reaction (PCR) for grouping hantavirus isolates by using four sets of primers specific to each serotype. Our PCR typing results agreed with those of serological typing. The present study makes use of thermal cycle sequencing to sequence PCR-amplified DNA products in order to determine the level of similarity among members of the same serotype. We show that members of Hantaan and Seoul serotypes are over 92% homologous, irrespective of their host and geographical origin. Puumala sequences show a degree of homology ranging from 80 to 98%. Despite the variation in sequence at the nucleotide level, amino acids show an even higher level of conservation.

PCR detection and typing of genital papillomavirus in a new brunswick population

We have used a broad range of primers for HPV detection, using the polymerase chain reaction (PCR) so as to compare PCR typing of HPV with the results of cytological diagnosis in a New Brunswick population referred to the out-patient clinic of the Saint John Regional Hospital. The primers selected were found to be capable of amplification with high efficiency, therefore we did not perform further hybridization analysis for specific identification of HPV types. Amplification of selected fragments for detection of HPV 6, 11, 16, 18, 31 and 33 was obtained from cervical swabs collected from 154 patients. Microscopic examination was performed in duplicate samples and the results compared with the DNA-typing analysis. HPV of any of the above types was detected in 43 out of 154 patients. Among these, 32 patients showed single or multiple infections with "high-risk" HPV strains 16, 18, 31 or 33. Cytologically normal or atypical samples with any of the HPV types tested amounted to 17%, but increased to 56% in patients with CIN I, and to 100% in patients with CIN II or III. Prevalence of "high-risk" types alone increased from 15% and 10%, for normal and atypical cases respectively, to 48% for CIN I, 75% for CIN II and 100% for CIN III. Our results indicate that HPV detection and typing by this simple procedure can be a valuable indicator of cancer progression and thus can help to identify individuals at high risk in pre-malignant stages of the disease.

Loss of human leukocyte antigen expression on colorectal tumor cell lines: implications for anti-tumor immunity and immunotherapy.

A system devised for tissue typing the human leukocyte antigen-A (HLA-A) locus from genomic DNA by the polymerase chain reaction (PCR) has been used to investigate abnormalities of HLA expression in a panel of 30 cell lines derived from colorectal adenocarcinomas, by comparison of the HLA-A locus genotype with surface expression of HLA. Eleven cell lines gave single HLA-A locus specificity on PCR typing, suggesting that loss of HLA alleles is a common abnormality. In one of these cell lines the loss of an HLA-A locus allele was confirmed by comparison with DNA from a lymphoblastoid B cell line derived from the same patient. In three cell lines, loss of expression of an HLA-A locus determinant was observed in spite of the presence of the relevant allele in genomic DNA. Three cell lines showed absent HLA expression associated with failure to express beta 2-microglobulin. These data indicate that at least three independent mechanisms were involved in the loss of HLA expression on the colorectal tumor cell lines.

Southern blot and dot blot hybridisation compared to PCR for the detection of human papillomavirus DNA in biopsies of the uterine cervix from women with dysplasia

The aim of this study was to compare the sensitivity of Southern blot (SB) and dot blot (DB) hybridisation with polymerase chain reaction (PCR) for the detection of HPV in cervical biopsies from samples with differing histology. One hundred and forty seven women with cervical dysplasia had biopsies performed; one sample was analyzed for HPV DNA from types 6 11 , 16, and 18 by SB, DB and PCR (L1 consensus primer and type specific probes) while an adjacent sample was examined histologically. The histology of the samples was normal in 40 (27%), squamous metaplasia in 25 (17%), inflammation 2 (1%) HPV infection 24 (17%), cervical intraepithelial neoplasia (CIN) grade I in 11 (7%), CIN II in 18 (12%), CIN III in 22 (15%), while 5 (3%) had invasive cancer. The number of biopsies positive for HPV DNA from types 6 11 , 16, and 18, using the different hybridisation methods was 56 (38%) by dot blot, 57 (39%) by Southern blot hybridisation and 66 (45%) by PCR. When the L1 consensus primer was used 100 (68%) specimens were positive by PCR. The sensitivity of SB and DB hybridisation, as compared with PCR (type specific probes 6 11 , 16, 18) was greater in biopsies with abnormal histology (histological grades of HPV infection and greater, as a group) (sensitivity of SB 83%, DB 74%) than those with normal and metaplastic change (as a group) (sensitivity of SB 44%, DB 35%) ( P < 0.005 for SB and DB) (inflammation excluded from analysis). This study demonstrated that the sensitivity of SB and DB hybridisation, relative to PCR is greater in samples with abnormal histology than in samples with normal histology.

Genetic basis of human complement C8 beta deficiency.

The eighth component of human complement (C8) is a serum protein consisting of three chains (alpha, beta, and gamma) and encoded by three different genes, C8A, C8B, and C8G. C8A and C8B are closely linked on chromosome 1p, whereas C8G is located on chromosome 9q. In the serum the beta subunit is non-covalently bound to the disulfide-linked alpha-gamma subunit. Patients with C8 beta deficiency suffer from recurrent neisserial infections such as meningitis. Exon-specific polymerase chain reaction (PCR) amplification with primer pairs from the flanking intron sequences was used to amplify all 12 C8B exons separately. No difference regarding the exon sizes was observed in a C8 beta-deficient patient compared with a normal person. Therefore, direct sequence analysis of all exon-specific PCR products from normal and C8 beta-deficient individuals was carried out. As a cause for C8 beta deficiency, we found a single C-T exchange in exon 9 leading to a stop codon. An allele-specific PCR system was designed to detect the normal and the deficiency allele simultaneously. Using this approach as well as PCR typing of the Taql polymorphism located in intron 11, five families with 7 C8 beta-deficient members were investigated. The mutation was not found to be restricted to one of the two Taql RFLP alleles. The mutant allele was observed in all families investigated and can therefore be regarded as a major cause of C8 beta deficiency in the Caucasian population. In addition, two C8 beta-deficient patients were found to be heterozygous for the C-T exchange. The molecular basis of the alleles without this point mutation also causing deficiency has not yet been defined.

Prognostic significance of polymerase chain reaction detected human papillomavirus of tumors and lymph nodes in surgically treated stage IB cervical cancer

This study describes the prognostic role of polymerase chain reaction detected human papillomavirus (HPV) in Stage IB cervical cancer patients treated with radical hysterectomy and pelvic and paraaortic node dissection. All tumors were confined to the cervix and all margins and nodes were disease free. Twenty-one patients were analyzed: 6 patients recurred within 20 months of initial therapy, while 15 had no evidence of disease with a minimum follow-up of 36 months. Polymerase chain reaction (PCR) was performed on paraffin-block tissue of the hysterectomy specimen cervical tumor and lymph nodes. Oligonucleotide probes for HPV types 6, 11, 16, 18, 31, 33, and 35 were used with consensus primers for uncharacterized HPV types created from an L1 constant region. Control tissues were run with each tumor sample to assure against contamination. HPV type confirmation was performed using diagnostic restriction sites. HPV was detected in all cervical tumors. Recurring tumors were infected with multiple types of HPV in all 6 tumors versus only 5 of 15 nonrecurring tumors being multiply infected ( P = 0.023). No tumor had HPV 6 or 11, and the incidence of HPV 16, 31, 33, and 35 was not significantly different for recurrent versus nonrecurrent groups. HPV 18 was found in 5 of 6 recurring cancers versus 1 of 15 nonrecurring tumors ( P = 0.0029). PCR typing of the histologically negative nodes that had been obtained at radical hysterectomy was done in all 6 recurring patients and in 6 nonrecurring patients. The recurrent patients had a significantly higher incidence of lymph nodes positive for HPV DNA (71%) than the nonrecurring patients (35%) ( P = 0.0047). These observations suggest that HPV 18 cervical cancer patients, those with infections of multiple types, and those with HPV DNA in histologically negative lymph nodes may be at increased risk for recurrence.

Rapid diagnosis of Miller-Dieker syndrome and isolated lissencephaly sequence by the polymerase chain reaction

Probe YNZ22 (D17S5) is a highly polymorphic, variable number tandem repeat (VNTR) marker previously shown to be deleted in all patients with the Miller-Dieker syndrome (MDS) but not in patients with isolated lissencephaly sequence (ILS). Primers were constructed to the unique sequence flanking the polymorphic, repetitive region of YNZ22 for amplification by the polymerase chain reaction (PCR). Analysis of 118 normal individuals revealed 12 alleles (differing in copy number of a 70-bp repeat unit) ranging in size from 168 to 938 bp. A retrospective study of eight MDS and six ILS patients was consistent with Southern blot analysis in all cases except one. In the latter, a very large allele (12 copies of the repeat unit) in a patient and her mother failed to amplify on initial attempts, but was successfully amplified by reducing the concentration of genomic DNA used in the reaction. Prospective studies on two MDS and five ILS patients were successfully performed and confirmed in all cases by Southern blot analysis. From the total sample, restriction fragment length polymorphism (RFLP) analysis was fully informative in four of ten MDS patients and showed a deletion in all four cases. Nine of eleven ILS patients were heterozygous and therefore not deleted for YNZ22. Development of primers for additional polymorphic markers in the Miller-Dieker region will lead to a rapid PCR-based diagnostic approach for all MDS and ILS patients. PCR typing of YNZ22 will also facilitate use of this marker in other applications, including genetic linkage, paternity and forensic studies, and analysis of loss of heterozygosity in tumors.

High Prevalence of HTLV-II Among Intravenous Drug Abusers: PCR Confirmation and Typing

The polymerase chain reaction (PCR) was used to confirm the presence of human T-cell lymphotropic viruses (HTLV) in intravenous drug users (IVDU) whose sera were reactive by immunofluorescence assay (IFA) for HTLV-1/-II antibody. Peripheral blood mononuclear cells from 41 IFA-positive and 19 IFA-negative individuals were analyzed. HTLV sequences were detected in 39/41 IFA-positive samples; 36 were HTLV-II positive and 3 were HTLV-I positive. Two IFA antibody-positives were negative by both PCR and by enzyme immunoassay (EIA). One IFA and EIA antibody-negative sample was positive for HTLV-II by PCR. This study indicates a high prevalence of HTLV-II among IVDUs and further demonstrates the feasibility of using PCR to differentiate between HTLV-I and -II.