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PubMed2006

Global and regional distribution of HIV-1 genetic subtypes and recombinants in 2004.

OBJECTIVE: To estimate the global and regional distribution of HIV-1 subtypes and recombinants in 2004. DESIGN: A study was conducted in which molecular epidemiological data on HIV-1 subtype distribution in individual countries were combined with country-specific estimates of the number of people living with HIV. METHODS: HIV-1 subtype data were collected for 23 874 HIV-1 samples from 70 countries, which together accounted for 89% of all people living with HIV worldwide in 2004. The proportions of HIV-1 infections due to various subtypes detected in each country were combined with the number of HIV infected people in the respective countries to generate regional and global HIV-1 subtype distribution estimates. RESULTS: Subtype C accounted for 50% of all infections worldwide in 2004. Subtypes A, B, D and G accounted for 12%, 10%, 3% and 6%, respectively. The subtypes F, H, J and K together accounted for 0.94% of infections. The circulating recombinant forms CRF01_AE and CRF02_AG each were responsible for 5% of cases, and CRF03_AB for 0.1%. Other recombinants accounted for the remaining 8% of infections. All recombinant forms taken together were responsible for 18% of infections worldwide. CONCLUSION: Combining data on HIV-1 subtype distribution in individual countries with country-specific estimates of the number of people living with HIV provided a good method to generate estimates of the global and regional HIV-1 genetic diversity in 2004. The results could serve as an important resource for HIV scientists, public health officials and HIV vaccine developers.

Genetic Variation↗

PubMed2006

Characterization of a new circulating recombinant form comprising HIV-1 subtypes C and B in southern Brazil.

OBJECTIVE: To identify a new circulating recombinant form (CRF) of HIV-1 comprising two circulating subtypes in the southern region in Brazil, subtypes B and C. METHODS: A total of 152 HIV-positive patients followed at two hospitals in southern Brazil had their viral pol genes isolated by reverse transcriptase-polymerase chain reaction (PCR) from plasma. PCR products were sequenced and phylogenetically analysed using HIV-1 subtype reference sequences. Six full-length subtype C viruses from Brazil previously described as 'pure' strains were included in the analysis. Sequences suggestive of recombination were analysed by boot scanning and phylogenetic analyses of separate fragments. The common ancestry of recombinant strains was evaluated by similarity plot and informative site analyses. RESULTS: : HIV-1 subtypes commonly found in Brazil (B, C and F1) were observed. Sixty-two viruses were initially assigned as subtype C, but 15 viruses clustered in a separate internal clade. Pol from two full-length genomes of subtype C viruses grouped together with those samples. Boot scanning analysis showed that all 17 viruses had the same recombinant structure, with a 240 base pair fragment of subtype B in the middle of the reverse transcriptase pol region. Subtype B assignment of this fragment was confirmed by phylogenetic analyses using different methods of tree inference and cluster robustness tests. Mosaics were shown to have a common ancestry. CONCLUSION: As CRF_BC represents 11% of the HIV-1 viruses circulating in the southern region of the country, which borders several south American countries, the assessment of its spread is of pivotal importance to the HIV/AIDS epidemic in Brazil and Latin America.

Base Sequence↗

PubMed2006

Quantitative analysis of HIV-1 variants with the K103N resistance mutation after single-dose nevirapine in women with HIV-1 subtypes A, C, and D.

INTRODUCTION: We used a sensitive point mutation assay, LigAmp, to detect and quantify K103N-containing variants in African women who received single-dose nevirapine (NVP) to prevent mother-to-child HIV-1 transmission. METHODS: Plasma for testing was collected 6 to 8 weeks postpartum from 301 women (144 subtype A, 63 subtype C, and 94 subtype D). RESULTS: The portion of women with 0.5% or more K103N-containing variants was lowest for subtype A (60/144, 41.7%) and highest for subtype C (44/63, 69.8%; P < 0.0001). K103N was rarely detected in pre-NVP samples. In a multivariate model, K103N detection was associated with HIV-1 subtype (C > A), after adjusting for log10 delivery viral load, the number of days between NVP dosing and sample collection, age, and parity. Among women with K103N detected: (1) the median %K103N was lower for subtype A (2.2%) than C (11.7%, P = 0.0001) or D (5.5%, P = 0.04), and (2) in a multivariate linear model, higher log10 (%K103N) was associated with HIV subtype (C > A, P = 0.0001; D > A, P = 0.01; and C vs D, no difference), but not other factors. CONCLUSIONS: After administration of single-dose NVP, K103N was detected more frequently and at higher levels in women with subtypes C and D than A. Further studies are needed to evaluate the clinical significance of NVP-resistant variants in this setting.

Amino Acid Substitution↗

PubMed2006

HIV-1 subtypes: epidemiology and significance for HIV management.

PURPOSE OF REVIEW: This review presents an update on the molecular epidemic patterns of HIV-1 infection and the effects of subtype-related genetic variability on transmission, disease progression, response to antiretroviral therapy and drug-resistance pathways. RECENT FINDINGS: The molecular epidemiology of HIV-1 infection is complex and evolving. The emergence of new variants reflects HIV-1 prevalence, subtype epidemiology and risk-behaviour patterns in different geographical areas. Evidence indicates that certain subtypes may have a transmission advantage while others display higher replicative efficiency. The molecular mechanisms underlying these differences are being identified and include both virus- and host-related factors. Although drug susceptibility varies and clinical evidence remains limited, current antiretroviral regimens appear to have comparable efficacy in patients infected with B and non-B subtypes. Subtype-related variability influences resistance pathways. However, the major treatment-associated resistance mutations seen in subtype B also confer resistance in non-B subtypes and vice versa. SUMMARY: Genetic differences among HIV-1 variants can influence the virus biological properties, susceptibility to existing and candidate antiretroviral drugs, and evolution of antiretroviral drug resistance. Further studies are required to define the impact of this variability on risk of transmission, disease outcomes, responses to antiretroviral therapy and resistance pathways. Meanwhile, plasma viral load and CD4 count remain the important predictors of disease outcome, regardless of the infecting subtype. Current antiretroviral regimens can be used reliably to treat patients with both B and non-B subtypes, and resistance interpretation algorithms provide adequate guidance. The limitations of current evidence should be acknowledged and instigate ongoing vigilance.

Anti-HIV Agents↗

PubMed2025

Phosphoproteomics delineates hepatocellular carcinoma subtypes and pinpoints therapeutic targets.

BACKGROUND AND AIMS: Only a minority of patients could benefit from systemic therapy owing to the high heterogeneity of HCC. Therefore, a deeper understanding of the pathogenesis of HCC is essential for precision therapy. Genomic and proteomic studies of HCC have enhanced our understanding of HCC. However, the phosphoproteomic characterization of HCC remains poorly understood. APPROACH AND RESULTS: We conducted an in-depth analysis of a clinical cohort of HCC using high-coverage phosphoproteomic. Effective therapeutic targets were validated using liver cancer cell lines and HCC patient-derived xenograft mouse models that correspond to the phosphoproteomic subtypes of HCC. Phosphoproteomic analysis classified HCC into 3 subtypes, A, B, and C, with increasing malignancy and correlation with clinical features, including patient prognosis, tumor staging, serum alpha-fetoprotein levels, tumor thrombus, and tumor size. Phosphoproteomic subtyping deeply reflected the biological characteristics and clinical features of patients with HCC. The profiles of HCC-dysregulated kinase activities inferred from the different phosphoproteomic subtypes consistently identify increased kinase activity related to cell proliferation. Subtype-C HCC patients showed the most significant dysregulation, indicating a potential therapeutic target. The corresponding drug, bosutinib, demonstrated efficacy in inhibiting the growth of subtype C tumors in liver cancer cell lines and HCC patient-derived xenograft mouse models representative of the phosphoproteomic HCC subtypes. CONCLUSIONS: Our study provides a comprehensive exploration of the phosphoproteomic landscape of HCC, establishing new subtypes that match clinical features and identifying potential therapeutic targets for the most malignant C subtype.

Carcinoma, Hepatocellular↗

PubMed1994

Genetic conservation within subtypes in the hepatitis B virus pre-S2 region.

The antigenic determinants for the main hepatitis B virus (HBV) subtypes adw, adr, ayw and ayr lie in the S (surface) polypeptide. Two amino acid residues in particular, encoded by the S gene at codon positions 122 and 160, have been postulated to determine the different antigenic subtypes. In contrast, the 165 nucleotide pre-S2 gene encodes an immunodominant region common to all subtypes that can give rise to neutralizing antibodies. We have characterized the pre-S2 gene sequences of 29 HBV strains of the three main subtypes, adw, ayw and adr. Seven base positions showed variation that was entirely subtype-specific, with six of these variations leading to subtype-specific amino acid differences. This finding affords the possibility of using pre-S2 sequences for genetic subtyping. Two ayw strains from unrelated patients infected in the Middle East had identical pre-S2 sequences with a block of 12 nucleotides deleted. A geographical correlation with subtype observed from serological results was also apparent from phylogenetic analysis of DNA identities within the pre-S2 region. The results support the concept that the main HBV subtypes truly represent families of phylogenetically different strains.

Base Sequence↗

PubMed1994

Molecular subtyping of Borrelia burgdorferi in erythema migrans and acrodermatitis chronica atrophicans.

Recently, three subtypes of Borrelia burgdorferi have been identified: Borrelia burgdorferi sensu stricto, Borrelia garinii, and the VS 461 group of Borrelia burgdorferi. These subtypes differ by nucleotide sequence variations within several Borrelia burgdorferi specific genes and most likely by their pathogenetic potential. To assess whether different subtypes of Borrelia burgdorferi might be associated with different cutaneous manifestations and clinical courses of Lyme disease, lesional skin biopsies from 35 patients with erythema migrans and 18 patients with acrodermatitis chronica atrophicans were analyzed. A Borrelia burgdorferi specific gene segment encoding a 26-kD protein with subtype specific nucleotide sequence variations was amplified by a nested polymerase chain reaction technique. For molecular subtyping, the products were transcribed into complementary RNA. Upon polyacrylamide gel electrophoresis, complementary RNA molecules separate into several metastable conformational forms resulting in patterns of bands highly specific for the nucleotide sequence of the transcribed molecules. In biopsy specimens of erythema migrans, the VS 461 subtype was detected in 28 of 35 and the Borrelia garinii subtype in six of 35 cases. In one of 35 cases of erythema migrans Borrelia burgdorferi sensu stricto as well as Borrelia garinii was detected. In contrast, in all 18 biopsies of acrodermatitis chronica atrophicans, only the VS 461 subtype was identified. This subtype is rarely found in the USA, where acrodermatitis chronica atrophicans is almost unknown. These data indicate that acrodermatitis chronica atrophicans might be closely associated with the VS 461 group of Borrelia burgdorferi.

Acrodermatitis↗

PubMed1993

Differential affinity of dihydroimidazoquinoxalines and diimidazoquinazolines to the alpha 1 beta 2 gamma 2 and alpha 6 beta 2 gamma 2 subtypes of cloned GABAA receptors.

1. In this study, we compared two series of newly discovered ligands for their selectivity to benzodiazepine sites in the alpha 1 beta 2 gamma 2 and the alpha 6 beta 2 gamma 2 subtypes of cloned gamma-aminobutyric acidA (GABAA) receptors, the latter being unique in not interacting with classical benzodiazepines. 2. The prototype compounds, U-85575 (12-chloro-5-(5-cyclopropyl-1',2',4'- oxadiazol-3'-yl)-2,3-dihydro-diimidazo [1,5-a;1,2-c]quinazoline), and U-92330 (5-acetyl-3-(5'-cyclopropyl-1',2',4'-oxadiazole-3'-yl)-7-chloro-4,5-d ihy dro [1,5-a]quinoxaline), appear to share an overlapping recognition site with classical benzodiazepines on the GABAA receptor, because their potentiation of GABA-mediated Cl- currents in both subtypes were sensitive to Ro 15-1788, a classical benzodiazepine antagonist. 3. Minor changes in the ring substituents of the drugs reduced their affinity to the alpha 6 beta 2 gamma 2 subtype more pronouncedly than to the alpha 1 beta 2 gamma 2 subtype. The diimidazoquinazoline containing a 2-methyl group which projected below the plane of the rigid ring showed a markedly lower affinity to the alpha 6 beta 2 gamma 2 subtype as compared to its stereoisomer having the methyl group above the plane of the ring. Also, the dihydroimidazoquinoxalines containing the 5-benzoyl group showed a lower affinity to the alpha 6 beta 2 gamma 2 subtype than the 5-acetyl counterpart. In particular, the 5-benzoyl analogue containing a 6-fluoro group showed no interaction with the alpha 6 beta 2 gamma 2 subtype even at the concentration of 10 microM, probably due to stabilization of the benzoyl group in the out-of-plane region by the steric and electrostatic effects of the 6-fluoro group.4. We propose that the benzodiazepine site of the alpha 6 beta 2 gamma 2 subtype shares overlapping regions with that of the alpha 1 beta 2 gamma 2 subtype, but has a sterically restricted out-of-plane region, which may be also incompatible with the 5-phenyl group of classical benzodiazepines.

Cells, Cultured↗

PubMed1994

Effects of GABA and various allosteric ligands on TBPS binding to cloned rat GABA(A) receptor subtypes.

1. [35S]t-butylbicyclophosphorothionate (TBPS) is a high affinity ligand for the picrotoxin site of GABA(A) receptors. Here we examined TBPS binding to the cloned receptors made of alpha 1, alpha 3 or alpha 6 in combination with beta 2 or beta 2 and gamma 2 subunits, in the presence of GABA and several allosteric ligands (diazepam, methyl 6,7-dimethoxy-4-methyl-beta-carboline-3-carboxylate (DMCM), 3 alpha,21-dihydroxy-5 alpha-pregnan-20-one (5 alpha-THDOC), pentobarbitone and Zn). The cloned receptors were transiently expressed in SF-9 insect cells by infecting with recombinant baculoviruses. 2. In alpha beta subtypes, GABA at nanomolar concentrations enhanced TBPS binding but inhibited binding at micromolar concentrations. Half maximal GABA concentrations for enhancement or inhibition of TBPS binding were correlated with high and low affinity GABA binding sites, respectively, in individual subtypes. The maximal enhancement of binding also varied according to the alpha isoform (alpha 3 beta 2 >> alpha 1 beta 2). In alpha beta gamma subtypes, TBPS binding was unaffected by GABA at nanomolar concentrations, but was inhibited by GABA at micromolar concentrations. Addition of gamma 2 thus appeared to abolish conformational coupling between high affinity GABA sites and TBPS sites, and also altered low affinity GABA sites; in particular, the half maximal GABA concentration for inhibition of TBPS binding changed from > 100 (alpha 6 beta 2) to 1 microM (alpha 6 beta 2 gamma 2). 3. Allosteric ligands also altered TBPS binding to sensitive GABA(A) receptor subtypes. For instance,diazepam only in the alpha 1 beta2 gamma 2 and alpha 3 beta 2 gamma 2 subtypes, and 5 alpha-THDOC in all the subtypes enhanced TBPS binding in the absence of GABA, and intensified the inhibitory effect of GABA. Pentobarbitone exhibited only the latter effect in all the subtypes we examined.4. DMCM and Zn, inhibitors of GABA-induced Cl currents in alpha beta gamma and alpha beta subtypes, respectively,produced opposite effects to agonists, decreasing TBPS binding in the absence of GABA and attenuating(or eliminating in the case of Zn) the inhibitory effect of GABA on TBPS binding.5. These results show that GABA binding sites and their conformational coupling with TBPS sites are differentially affected by the alpha isoform (particularly alpha 6 as compared to alpha l or alpha 3) and by quaternary interactions involving the gamma 2 subunit. Moreover, changes in TBPS binding by allosteric ligands include not only direct (allosteric) effects on TBPS sites but also indirect effects via GABA sites, and are consistent with their known subtype selectivity and functionality from previous studies.

Allosteric Regulation↗

PubMed1996

The distribution and density of receptor subtypes for endothelin-1 in peripheral lung of the rat, guinea-pig and pig.

1. Quantitative autoradiographic studies were conducted to determine the distributions and densities of endothelin-A (ETA) and ETB receptor subtypes in peripheral lung alveolar wall tissue of the rat, guinea-pig and pig, with a view to assessing the potential suitability of these tissues as models for investigations of ET receptor function in human alveolar tissue. 2. High levels of specific [125I]-ET-1 binding were detected in peripheral lung components from all three species tested. In mature porcine alveolar wall tissue, specific binding increased in a time-dependent manner to a plateau, consistent with the previously described pseudo-irreversible binding of this ligand to a finite population of specific binding sites. 3. [125I]-ET-1 was associated specifically with both ETA and ETB binding site subtypes in alveolar wall tissue of foetal pig lung as early as 36 days gestation, raising the possibility of a functional role for ET-1 in lung development. In addition, both ETA and ETB binding site subtypes were detected in alveolar wall tissue and in peripheral airway smooth muscle of mature lung parenchyma from all three species. However, the binding subtype proportions differed in these tissues. For example, in porcine peripheral bronchial smooth muscle, ETA sites apparently predominated, whereas ETB sites constituted the major subtype detected in alveolar wall in this species. These data suggest significant shifts in ET receptor subtype expression at different levels in the respiratory tract. 4. ET binding site subtype proportions in the alveolar wall also differed markedly between species. In rat lung alveoli, ETA and ETB sites were detected in similar proportions (52 +/- 3% and 43 +/- 5% respectively). In contrast, in guinea-pig peripheral lung, ETB binding sites clearly predominated, constituting approximately 80% of total specific binding, with ETA sites accounting for only 12%. Porcine alveolar wall tissue also contained a mixture of these ET receptor subtypes, with ETA and ETB binding comprising 23 +/- 3% and 65 +/- 1% respectively of the total population of specific binding sites detected. These latter proportions are similar to values previously obtained in human peripheral lung tissue, suggesting that porcine lung might be a useful model of the human peripheral lung in subsequent studies of the functions of these pulmonary ET receptor subtypes.

Animals↗

PubMed1996

Subtypes of alcohol-dependent men: a typology based on relative genetic and environmental loading.

Using scales that distinguish between relative genetic and environmental loading, cluster analysis was used to identify three subtypes of alcohol dependence in Caucasian men from the Epidemiologic Catchment Area study (n = 911). Although all subjects met DSM-III criteria for alcohol dependence, only the severe subtype showed evidence of substantial genetic influence. When compared on a range of clinical characteristics, the mild subtype (53% of the sample) was typically least adversely affected and the severe subtype (17%) most affected, with the dyssocial subtype (30%) falling between. Severe subtype subjects had significantly greater comorbid drug dependence and were at least four times more likely than mild subjects to have sought treatment for alcohol problems. Ratio of genetic scale score to total symptom count (genetic ratio) was highest for the severe subtype (mean = 0.37), and negatively correlated with age of first alcohol problem (rs = -0.16) and years between first intoxication and first problem (rs = 0.19). No significant correlations were found between these clinical features and genetic ratio for the mild or dyssocial subtypes. Use of these scales and subtypes may improve our ability to detect specific gene effects in genetic linkage studies and to identify environmental influences in behavioral and epidemiological studies.

Adult↗

PubMed1996

A molecular perspective of the genetic relationships of G-protein coupled melatonin receptor subtypes.

Successful cloning of melatonin receptors from various target tissues in the past few years has increased our understanding of the molecular signal transduction mechanisms of G-protein coupled melatonin receptors, of which three subtypes (MEL-1A, MEL-1B, and MEL-1C) have been reported in different vertebrates. Based upon melatonin receptor sequences available in the Genbank database, we have performed phylogenetic analyses of the nucleotide and encoded amino acid sequences of G-protein-coupled melatonin receptors, and determined the range of amino acid identities between melatonin receptors of the same and different subtypes. Besides the three well-known subtypes, a potential novel subtype of MEL-1D, as exemplified by unique separation of Xenopus X2.0 sequence (Genbank accession No. U31826) from the others in the protein phylogenetic tree, possibly exists. In addition, one of the chicken brain melatonin receptor sequences has been identified as belonging to the MEL-1B subtype. Our analyses showed that melatonin receptors of the same subtype and different subtypes are likely to share > or = 75% and < 65% amino acid identities, respectively. Phylogenetic analysis based on amino acid comparisons will be needed to determine the subtype status of any pair of melatonin receptor sequences that exhibit > or = 65% to < 75% amino acid identity. Despite the usefulness of genetic relatedness in the subtype classification of G-protein-coupled melatonin receptors, functional correlation of molecular structure may ultimately prove the most comprehensive approach in melatonin receptor classification.

Amino Acid Sequence↗

PubMed1999

Genetic variation and susceptibilities to protease inhibitors among subtype B and F isolates in Brazil.

The genetic variation of the human immunodeficiency virus type 1 (HIV-1) protease gene (prt) permits the classification of HIV-1 strains into five distinct protease subtypes, which follow the gag subtyping patterns. The susceptibilities of non-B-subtype strains to protease inhibitors (PIs) and other antiretroviral drugs remain largely unknown. Subtype F is the main non-B strain contributing to the Brazilian epidemic, accounting for 15 to 20% of these infections. In this work, we report the findings on 81 isolates from PI-naive Brazilian patients collected between 1993 and 1997. In addition, the relevant PI resistance mutations and their phenotypes were determined in vitro for 15 of these patients (B = 9 and F = 6). Among these, the subtype F samples evidenced high sensitivities in vitro to ritonavir and indinavir, with MICs at which 50 and 90% of the isolates are inhibited similar to those of both the Brazilian and the U.S. subtype B isolates. Analysis of the 81 Brazilian prt sequences demonstrated that the subtype F consensus sequence differs from the U.S. and Brazilian subtype B consensus in eight positions (I15V, E35D, M36I, R41K, R57K, Q61N, L63P, and L89M). The frequency of critical PI resistance substitutions (amino acid changes D30N, V82A/F/T, I84V, N88D, and L90M) among Brazilian isolates is very low (mean, 2.5%), and the associated secondary substitutions (amino acid positions 10L, 20K, 36M, 46M, 48G, 54I, 63P, 71A, and 77A) are infrequent. These observations document the relative rarity of resistance to PIs in the treatment of patients infected with HIV-1 subtype F in South America.

Amino Acid Sequence↗

PubMed2003

Analysis of linear B-cell epitopes of the nucleoprotein of ebola virus that distinguish ebola virus subtypes.

Ebola virus consists of four genetically distinguishable subtypes. We developed monoclonal antibodies (MAbs) to the nucleoprotein (NP) of Ebola virus Zaire subtype and analyzed their cross-reactivities to the Reston and Sudan subtypes. We further determined the epitopes recognized by these MAbs. Three MAbs reacted with the three major subtypes and recognized a fragment containing 110 amino acids (aa) at the C-terminal extremity. They did not show specific reactivities to any 10-aa short peptides in Pepscan analyses, suggesting that these MAbs recognize conformational epitope(s) located within this region. Six MAbs recognized a fragment corresponding to aa 361 to 461 of the NP. Five of these six MAbs showed specific reactivities in Pepscan analyses, and the epitopes were identified in two regions, aa 424 to 430 and aa 451 to 455. Three MAbs that recognized the former epitope region cross-reacted with all three subtypes, and one that recognized the same epitope region was Zaire specific. One MAb, which recognized the latter epitope region, was reactive with Zaire and Sudan subtypes but not with the Reston subtype. These results suggest that Ebola virus NP has at least two linear epitope regions and that the recognition of the epitope by MAbs can vary even within the same epitope region. These MAbs showing different subtype specificities might be useful reagents for developing an immunological system to identify Ebola virus subtypes.

Amino Acid Sequence↗

PubMed2001

Nested restriction site-specific PCR to detect and type hepatitis C virus (HCV): a rapid method to distinguish HCV subtype 1b from other genotypes.

Genotypic differentiation of hepatitis C virus (HCV) has become an integral part of clinical management and epidemiologic studies of hepatitis C infections. Thus, it is extremely important in areas such as the Czech Republic, where current instrumentation and kits for assessing HCV infection are too costly for widespread use. We describe a new and relatively inexpensive method called nested restriction site-specific PCR (RSS-PCR) that generates a "fingerprint" pattern to represent an HCV genotype without the use of restriction endonucleases and that specifically differentiates HCV genotype 1b from the other HCV genotypes. The RSS-PCR method was applied directly to serum samples from patients with hepatitis C from the Czech Republic and from patients with known HCV genotypes from the United States. The method was validated by comparison of the subtype determined by RSS-PCR to the subtype determined from analysis of the 5' noncoding region (NC) or the nonstructural protein gene (NS5b) nucleotide sequence of HCV in these clinical samples. From 75 Czech samples containing HCV RNA, three distinct RSS-PCR patterns were observed; 54 were predicted to contain subtype 1b, 19 were predicted to contain subtype 1a, and 2 were predicted to contain subtype 3a. Among 54 samples predicted to contain HCV genotype 1b, all were confirmed by their 5' NC or NS5b sequences to be subtype 1b. Thus, both the sensitivity and specificity of the RSS-PCR test for the differentiation of HCV subtype 1b from the others were 100%. While the assay described here was designed to specifically differentiate HCV subtype 1b from the other HCV genotypes, the RSS-PCR method can be modified to differentiate any HCV genotype or subtype of interest. Its simplicity and speed may provide new opportunities to study the epidemiology of HCV infections and the relationship between HCV genotypes and clinical outcome by more laboratories throughout the world.

Genotype↗

PubMed2001

Prevalence of human immunodeficiency virus type 1 (HIV-1) non-B subtypes in foreigners living in Madrid, Spain, and comparison of the performances of the AMPLICOR HIV-1 MONITOR version 1.0 and the new automated version 1.5.

Plasma specimens collected in 1999 from 32 human immunodeficiency virus type 1 (HIV-1)-infected foreigners living in Madrid, Spain, were examined for the presence of non-B subtypes. Furthermore, plasma viremia was quantified using two different AMPLICOR HIV-1 MONITOR tests, version 1.0 and the new upgraded and automated version 1.5 (COBAS). Most patients came from Africa, where they most likely had acquired HIV-1 infection through sexual contact. HIV-1 genetic subtyping was based on the phylogenetic analysis of the protease gene. Twenty-two subtype B, six subtype G, two subtype C, one subtype A, and one D subtype infection were found. Overall, non-B subtypes represented 31.25% of the study population. Irrespective of the HIV-1 variant, viral load values above the detection limit (200 HIV RNA copies/ml) increased from 56.2 to 71.9% for results obtained using MONITOR version 1.0 and COBAS, respectively. Moreover, significant differences in viral load values (>0.5 logs) were recognized in up to 37.5% of samples. In summary, COBAS seemed to be more reliable for testing plasma viral load in HIV-infected immigrants living in Spain, one third of whom carried non-B subtypes.

Adult↗

PubMed2002

Rapid detection of human immunodeficiency virus type 1 subtype e infection by PCR.

The CRF01_AE (subtype E) strain of human immunodeficiency virus type 1 (HIV-1), originally reported in Thailand, spread rapidly to and showed prevalence in several countries in Southeast Asia, including Taiwan. This strain was also found in other regions of the world. Based on sequence analysis of the vpu gene, a nested PCR assay including an outer primer pair and a subtype E-specific inner primer pair was developed in this study for rapid detection of subtype E viruses. It was tested with 397 HIV-1-positive samples of known subtypes. For these samples, the sensitivity of detection of subtype E viruses was 100% (127 of 127), and the specificity was 97.8% (264 of 270). Although six samples of either subtype A or G showed a positive PCR, most of the cross-reactivity could be reduced by raising the annealing temperature from 54 degrees C to 63 degrees C. When tested with 195 HIV-positive samples of unknown subtypes, the assay had a sensitivity of 98.0% and a specificity of 98.6%. This is a simple, convenient, and sensitive method for rapid detection of subtype E viruses, especially in regions in which viruses of subtypes B and E are predominant.

DNA, Viral↗

PubMed2003

Segregation of human immunodeficiency virus type 1 subtypes by risk factor in Australia.

The aim of this study was to determine which human immunodeficiency virus type 1 (HIV-1) subtypes were circulating in Australia and to correlate the subtypes with risk factors associated with the acquisition of HIV-1 infection. DNA was extracted from peripheral blood mononuclear cells, and HIV-1 env genes were amplified and subtyped using heteroduplex mobility analysis, with selected samples sequenced and phylogenetic analysis performed. The HIV-1 env subtypes were determined for 141 samples, of which 40 were from female patients and 101 were from male patients; 13 samples were from children. Forty-seven patients were infected by homosexual or bisexual contact, 46 were infected through heterosexual contact, 21 were infected from injecting drug use (IDU), 13 were infected by vertical transmission, 8 were infected from nosocomial exposure, and 6 were infected by other modes of transmission, including exposure to blood products, ritualistic practices, and two cases of intrafamilial transmission. Five subtypes were detected; B (n = 104), A (n = 5), C (n = 17), E (CRF01_AE; n = 13), and G (n = 2). Subtype B predominated in HIV-1 acquired homosexually (94% of cases) and by IDU (100%), whereas non-subtype B infections were mostly seen in heterosexually (57%) or vertically (22%) acquired HIV-1 infections and were usually imported from Africa and Asia. Subtype B strains of group M viruses predominate in Australia in HIV-1 transmitted by homosexual or bisexual contact and IDU. However, non-B subtypes have been introduced, mostly acquired via heterosexual contact.

Adult↗