RESUMO
OBJECTIVE: The objective of the study was to determine the sensitivity and specificity of the APTIMA human papillomavirus (AHPV) assay for high-grade cervical intraepithelial neoplasia (CIN) in women 21 years old and older with atypical squamous cells of undetermined significance (ASC-US) cytology. STUDY DESIGN: Women 21 years old and older with ASC-US cytology had colposcopy/biopsy and molecular human papillomavirus testing. Performance of the AHPV and Hybrid Capture 2 assays was compared with a clinical diagnosis of CIN grade 2, CIN grade 3, or adenocarcinoma in situ (CIN2 or greater). RESULTS: Of 939 evaluable subjects, CIN2 or greater and CIN3 or greater prevalence was 9.7% and 4.4%, respectively. AHPV sensitivity and specificity was 86.8% and 62.9% for CIN2 or greater detection and 90.2% and 60.2% for CIN3 or greater, respectively. AHPV had a similar sensitivity to Hybrid Capture 2 but a significantly higher specificity (62.9% vs 55.8%, P < .001) for CIN2 or greater detection. CONCLUSION: Among women with ASC-US cytology, detection of high-risk human papillomavirus E6/E7 oncogenic messenger ribonucleic acid is an effective triage method for colposcopy referral.
Assuntos
Carcinoma in Situ/diagnóstico , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/diagnóstico , Displasia do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/diagnóstico , Adulto , Idoso , Proteínas do Capsídeo/genética , Carcinoma in Situ/virologia , Colposcopia , DNA Viral/análise , Feminino , Humanos , Pessoa de Meia-Idade , Proteínas Oncogênicas Virais/genética , Papillomaviridae/genética , Infecções por Papillomavirus/virologia , Estudos Prospectivos , RNA Mensageiro/genética , Medição de Risco , Sensibilidade e Especificidade , Neoplasias do Colo do Útero/virologia , Esfregaço Vaginal , Adulto Jovem , Displasia do Colo do Útero/virologiaRESUMO
By using a mathematical model, the APTIMA human immunodeficiency virus type 1 (HIV-1) RNA qualitative assay was evaluated as a semiquantitative assay to distinguish HIV-1 patient samples needing quantitation from samples in which the virus was suppressed with antiretroviral therapy.
Assuntos
Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/isolamento & purificação , RNA Viral/sangue , HIV-1/genética , Humanos , Sensibilidade e Especificidade , Carga ViralRESUMO
BACKGROUND: The clinical significance of occult hepatitis B virus (HBV) infection, defined as the presence of HBV DNA in individuals with HBV core antibodies (anti-HBc) in the absence of HBV surface antigen (HBsAg), is unclear in HIV-infected patients. This information is needed to determine the importance of detecting and treating occult HBV in this population. OBJECTIVE: To determine if HIV-infected patients with occult HBV infection have an increased incidence of transaminitis. STUDY DESIGN: We performed a cohort study among randomly selected HBsAg-/anti-HBc+ HIV-infected patients in the Penn CFAR Database and Specimen Repository. HBV DNA was qualitatively detected using a transcription-mediated amplification assay. Hepatic transaminase levels, the main study outcome, were collected at 6-month intervals from the time of occult HBV determination. RESULTS: Among 97 randomly selected subjects without baseline transaminitis, 13 (13%) had occult HBV. These subjects more frequently had detectable HIV RNA. The 2-year incidence of transaminitis among HIV-infected subjects with occult HBV (50 events/100 person-years) was not significantly different from those without occult HBV (38 events/100 person-years; adjusted incidence rate ratio=1.36 [95% CI, 0.72-2.59]). CONCLUSIONS: Occult HBV did not increase the incidence of hepatic transaminitis over 2 years. Future studies should determine whether occult HBV is associated with other clinically important outcomes, particularly hepatocellular carcinoma.
Assuntos
Alanina Transaminase/sangue , Aspartato Aminotransferases/sangue , Infecções por HIV/complicações , Hepatite B/complicações , Fígado/enzimologia , Adulto , Estudos de Coortes , DNA Viral/sangue , Feminino , Infecções por HIV/enzimologia , Hepatite B/enzimologia , Anticorpos Anti-Hepatite B/sangue , Vírus da Hepatite B/isolamento & purificação , Humanos , Fígado/virologia , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Distribuição de Poisson , Estatísticas não ParamétricasRESUMO
PURPOSE: To evaluate carcinogenic human papillomavirus (HPV) mRNA for E6 and E7 mRNA detection on clinical specimens to identify women with cervical precancer and cancer. EXPERIMENTAL DESIGN: We evaluated a prototype assay that collectively detects oncogenes E6/E7 mRNA for 14 carcinogenic HPV genotypes on a sample of liquid cytology specimens (n=531), masked to clinical data and to the presence of HPV genotypes detected by PGMY09/11 L1 consensus primer PCR assay. RESULTS: We found an increasing likelihood of testing positive for carcinogenic HPV E6/E7 mRNA with increasing severity of cytology (P(Trend) < 0.0001) and histology (P(Trend) < 0.0001), with 94% of cervical intraepithelial neoplasia grade 3 (CIN3) histology cases (46 of 49) and all five cancer cases testing positive for carcinogenic HPV E6/E7 mRNA. Overall, fewer specimens tested positive for carcinogenic HPV E6/E7 mRNA than for carcinogenic HPV DNA (P<0.0001, McNemar's chi(2) test), especially in women with Assuntos
Alphapapillomavirus/isolamento & purificação
, Biomarcadores Tumorais/análise
, Papillomavirus Humano 6/isolamento & purificação
, Lesões Pré-Cancerosas/diagnóstico
, RNA Mensageiro/análise
, Neoplasias do Colo do Útero/diagnóstico
, Alphapapillomavirus/genética
, Estudos Transversais
, DNA Viral/análise
, Feminino
, Papillomavirus Humano 6/genética
, Humanos
, Hibridização de Ácido Nucleico
, Lesões Pré-Cancerosas/patologia
, Neoplasias do Colo do Útero/patologia
RESUMO
Diagnostics developers often face challenges introducing in-vitro diagnostic (IVD) products to low- and middle-income countries (LMICs) because of difficulty in accessing robust market data, navigating policy and regulatory requirements and implementing and supporting products in healthcare systems with limited infrastructure. Best practices recommend the use of a phase-gate model with defined activities and milestones by phase to successfully move a product from concept to commercialisation. While activities for commercialisation of products in high-income countries (HICs) are well understood, the activities required for introduction of IVDs in LMICs are not. In this paper, we identify the key activities needed for IVD product development and implementation and map them to the various phases of the model, paying particular attention to those activities that might be conducted differently in LMICs.
RESUMO
BACKGROUND: The second-generation hepatitis C virus (HCV) enzyme immunoassay (EIA 2), an antibody-detection test, has high sensitivity and is one of the recommended screening tests for detecting HCV infection in the United States. However, its sensitivity among oncology patients is unknown. OBJECTIVE: Assess the EIA 2 sensitivity among a group of oncology patients at a Nebraska clinic where an HCV outbreak occurred during 2000-2001 using nucleic acid testing (NAT) and recombinant immunoblot assay (RIBA) as the gold standards. STUDY DESIGN: Serum specimens were collected from patients 16 months after transmission had stopped. We tested the specimens using EIA 2 (Abbott HCV EIA 2.0), a NAT assay based on transcription-mediated amplification (TMA) (Gen-Probe TMA assay) and RIBA (Chiron RIBA HCV 3.0 SIA). HCV infection was defined as a positive RIBA or TMA test in an oncology patient. Alanine aminotransferase (ALT) levels were determined in EIA 2-negative/TMA-positive samples. RESULTS: A total of 264 samples were included in the study. We identified 92 HCV infections, 76 of which were Abbott EIA 2 positive. Abbott EIA 2 sensitivity was 83% (76/92), lower than that reported among healthy adults (90%) (p=0.01) and poor sensitivity was associated with receipt of chemotherapy during the outbreak period (p=0.02). Only 1 (6%) of the 16 EIA 2-negative cases had elevated ALT. CONCLUSIONS: In this study, EIA 2 sensitivity among oncology patients was lower than that previously reported among immunocompetent persons. Impaired antibody production related to cancer and/or chemotherapy might explain the reduced sensitivity. These findings indicate that, when assessing HCV status in oncology patients, a NAT test should be routinely considered in addition to EIA.
Assuntos
Hepacivirus/imunologia , Anticorpos Anti-Hepatite C/sangue , Hepatite C/diagnóstico , Neoplasias/complicações , Idoso , Surtos de Doenças , Feminino , Hepatite C/epidemiologia , Hepatite C/virologia , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: This study determined the longitudinal clinical performance of a high-risk human papillomavirus (HR-HPV) E6/E7 RNA assay (Aptima HPV [AHPV]; Hologic, San Diego, CA) compared with an HR-HPV DNA assay (Hybrid Capture 2 [HC2]; Qiagen, Gaithersburg, MD) as an adjunctive method for cervical cancer screening. METHODS: Women 30 years or older with a negative result for intraepithelial lesions or malignancy cytology (n = 10,860) positive by AHPV and/or HC2 assays and randomly selected women negative by both assays were referred to colposcopy at baseline. Women without baseline cervical intraepithelial neoplasia (CIN) grade 2 or higher (CIN2+) continued into the 3-year follow-up. RESULTS: The specificity of AHPV for CIN2 or lower was significantly greater at 96.3% compared with HC2 specificity of 94.8% (P < .001). Estimated sensitivities and risks for detection of CIN2+ were similar between the two assays. After 3 years of follow-up, women negative by either human papillomavirus test had a very low risk of CIN2+ (<0.3%) compared with CIN2+ risk in women with positive AHPV results (6.3%) or positive HC2 results (5.1%). CONCLUSIONS: These results support the use of AHPV as a safe and effective adjunctive cervical cancer screening method.
Assuntos
Detecção Precoce de Câncer , Papillomaviridae/genética , Infecções por Papillomavirus/diagnóstico , RNA Mensageiro/genética , RNA Viral/isolamento & purificação , Neoplasias do Colo do Útero/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Detecção Precoce de Câncer/métodos , Feminino , Testes Genéticos , Humanos , Estudos Longitudinais , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular , Papillomaviridae/isolamento & purificação , Gravidez , RNA Viral/genética , Neoplasias do Colo do Útero/virologia , Displasia do Colo do Útero/diagnósticoRESUMO
OBJECTIVES: The Aptima human papillomavirus (HPV) 16 18/45 Genotype (GT) assay (AHPV-GT) is a qualitative E6/ E7 oncogene messenger RNA test that detects HPV 16 and a pool of HPV 18 and 45. The CLEAR (Clinical Evaluation of APTIMA mRNA) study was the pivotal, prospective, multicenter US clinical study to validate the Aptima HPV (AHPV) assays. METHODS: In this analysis, we evaluated the clinical performance of AHPV and AHPV-GT assays for detection of cervical intraepithelial neoplasia grade 2 or more severe (CIN2 +) and grade 3 (CIN3) or adenocarcinoma in situ in 912 women with atypical squamous cells of undetermined significance (ASC-US) Papanicolaou result. The AHPV-GT assay was performed on high-risk HPV (hrHPV) positives as determined by the AHPV assay. RESULTS: Overall, the percent positive for hrHPV was 38.8% (354/912), of which 34.2% (121/354) were GT positive. Among hrHPV-positive women, the risks of CIN2 + were 37.0% for HPV 16 positives, 15.9% for HPV 18/45 positives, 14.3% for other hrHPV positives, and 2.2% for AHPV negatives. The risks of CIN3 + were 20.5% for HPV 16 positives, 9.1% for HPV 18/45 positives, 4.3% for other hrHPV positives, and 0.7% for HPV negatives. CONCLUSIONS: We demonstrated that AHPV-GT is a reliable and effective test for cervical cancer risk stratification in women with an ASC-US cytology diagnosis.
Assuntos
Infecções por Papillomavirus/diagnóstico , RNA Viral/análise , Displasia do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Células Escamosas Atípicas do Colo do Útero , Feminino , Humanos , Pessoa de Meia-Idade , Infecções por Papillomavirus/complicações , Neoplasias do Colo do Útero/virologia , Esfregaço Vaginal , Adulto Jovem , Displasia do Colo do Útero/virologiaRESUMO
HIV-1 recombinants between viruses from different subtypes appear to be surprisingly common in several regions of the world. To detect such intersubtype recombinants that contain mosaic genomes, we have analyzed sequences from the integrase (IN)-coding region of the polymerase (pol) gene from 23 viruses of known envelope (env) subtype from South America and Africa. As defined by env sequences, these viral genomes included nine subtype A, four subtype B, three subtype C, and four subtype D viruses from group M, and three viruses from group O HIV-1. Mosaic genomes were common, with 7 mosaic genomes among the 20 group M isolates analyzed. Two of these isolates had mosaic IN-coding regions that were distinct, but that had recombination breakpoints at the same location, in the highly conserved polypurine track. Mosaic genomes were particularly common in the viruses from Kenya (five of nine), consistent with our previous prediction that there was a high frequency of intersubtype recombinants circulating in this country. The IN amino acid sequence was highly conserved among the several represented subtypes, including group O. Group M IN sequences shared 94% or greater amino acid sequence identity within a subtype and 91% or greater identity between subtypes. The most divergent M and O variant amino acid sequences differed by only 19%, and the known functional domains were conserved among all of the isolates. The high degree of genetic homogeneity among the virus isolates representing several subtypes indicates that a single drug targeted against IN might be effective for all HIV-1 infections.
Assuntos
Variação Genética , Integrase de HIV/química , Integrase de HIV/genética , HIV-1/classificação , HIV-1/enzimologia , África Oriental , Sequência de Aminoácidos , Brasil , DNA Viral/análise , Infecções por HIV/virologia , HIV-1/genética , Humanos , Dados de Sequência Molecular , Filogenia , Recombinação Genética , Análise de Sequência de DNARESUMO
This paper describes a comprehensive study of hepatitis C virus (HCV) and human immunodeficiency virus type 1 (HIV-1) genotype sensitivity of the transcription-mediated amplification (TMA)-based HIV-1/HCV assay, developed and manufactured by Gen-Probe Incorporated (San Diego, CA) for screening human plasma specimens in blood bank settings. The TMA HIV-1/HCV assay is a qualitative, in vitro nucleic acid testing system used for initial screening. HIV-1 and HCV discriminatory assays are used to distinguish between HIV-1 and HCV infection or co-infection. In this study, multiple unique specimens representing HCV genotypes 1-6 were tested at various dilutions. The results show that the TMA HIV-1/HCV assay and the TMA HCV discriminatory assay have similar HCV genotype sensitivity, as both assays detected all six genotypes at 100 copies/ml and nearly all replicates tested at 30 copies/ml. Similarly, numerous unique specimens representing HIV-1 group M subtypes (A-G), HIV-1 group N, and group O specimens were tested at various dilutions. The TMA HIV-1/HCV assay and the TMA HIV-1 discriminatory assay were found to have similar HIV-1 subtype sensitivity; all variants at 100 copies/ml and nearly all at 30 copies/ml were detected. These results indicate that the TMA assays meet the sensitivity requirements for blood screening in blood banks worldwide.
Assuntos
Variação Genética , Infecções por HIV/virologia , HIV-1/genética , Hepatite C/virologia , Kit de Reagentes para Diagnóstico , Genótipo , Infecções por HIV/sangue , HIV-1/classificação , HIV-1/isolamento & purificação , Hepacivirus/classificação , Hepacivirus/genética , Hepacivirus/isolamento & purificação , Hepatite C/sangue , Humanos , Técnicas de Amplificação de Ácido Nucleico , Sensibilidade e Especificidade , Transcrição GênicaRESUMO
BACKGROUND: Human papillomavirus (HPV) DNA testing is widely used in conjunction with Papanicolaou (Pap) testing in cervical cancer screening programs to improve the detection of high-grade lesions. While HPV DNA test sensitivity is good, an improvement in specificity is desired. Detection of HPV mRNA may improve specificity. The APTIMA HPV Assay detects the mRNA of 14 high-risk HPV types in liquid-based cytology specimens. OBJECTIVE: To evaluate APTIMA HPV Assay performance for detection of high-risk HPV and high-grade cervical intraepithelial neoplasia (CIN) compared to Qiagen's Hybrid Capture 2 HPV DNA (HC2) test. STUDY DESIGN: Liquid Pap specimens were collected from 800 women referred to colposcopy and tested with the APTIMA HPV Assay and the HC2 test. Complete results were available for 753 subjects. A subset of samples (n = 393) were typed using Roche's Linear Array HPV Genotyping Test. RESULTS: Sensitivity and specificity for detection of high-risk HPV were >92% and 99% for the APTIMA HPV Assay and 93% and 82% for the HC2 test. Clinical sensitivity and specificity were 91% and >55% for detection of CIN 2+, and 98% and 53% for detection of CIN 3+ for the APTIMA HPV Assay; values for the HC2 test were 95% and 47% for CIN 2+, and 99% and 44% for CIN 3+. CONCLUSIONS: The APTIMA HPV Assay is sensitive and very specific for detection of high-risk HPV. The APTIMA HPV Assay had similar clinical sensitivity for disease detection but higher clinical specificity than the HC2 test, which may improve patient management and reduce the cost of care.
Assuntos
Colo do Útero/virologia , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/virologia , RNA Mensageiro/análise , RNA Viral/análise , Kit de Reagentes para Diagnóstico , Displasia do Colo do Útero/diagnóstico , Feminino , Humanos , Papillomaviridae/genética , RNA Mensageiro/genética , RNA Viral/genética , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Human papillomavirus (HPV) testing has improved the sensitivity for the detection of cervical pre-cancer and cancer as compared to Pap testing. Several HPV tests are commercially available and most target the DNA from 13 or 14 high-risk HPV types. The APTIMA HPV Assay however, detects HPV E6/E7 mRNA from 14 high-risk types of HPV: 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68. OBJECTIVE: To determine the analytical performance characteristics of the APTIMA HPV Assay. STUDY DESIGN: Analytical sensitivity, analytical specificity, reproducibility, and the effect of potentially interfering substances was determined for the APTIMA HPV Assay on both the DTS (semi-automated) and TIGRIS DTS (fully automated) systems. RESULTS: The 95% detection limit for both systems was between 17 and 488 copies/reaction, depending on the HPV type. The assay did not cross-react with normal flora and opportunistic organisms that may be found in cervical samples, or low-risk HPV types. Spermicides, anti-fungal and anti-itch medications, whole blood, glacial acetic acid, and most lubricants did not interfere with assay performance. Those lubricants containing polyquaternium 15 did interfere with assay performance. Inter-instrument, inter-operator, inter-lot, and inter-run signal variability were <10% for >99% of the data. Intra-run variability was <15%, except for those samples with concentrations at or below the 95% detection limit of the assay. CONCLUSIONS: Based upon the analytical sensitivity, analytical specificity, and low variability, the APTIMA HPV Assay showed excellent performance and robustness.
Assuntos
Perfilação da Expressão Gênica/métodos , Proteínas Oncogênicas Virais/biossíntese , Papillomaviridae/patogenicidade , Infecções por Papillomavirus/virologia , RNA Mensageiro/biossíntese , RNA Viral/biossíntese , Kit de Reagentes para Diagnóstico , Neoplasias do Colo do Útero/diagnóstico , Automação , Colo do Útero/virologia , Feminino , Humanos , Proteínas Oncogênicas Virais/genética , RNA Mensageiro/genética , RNA Viral/genética , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: The APTIMA HPV Assay (AHPV) is designed to detect HPV E6/E7 mRNA from 14 high-risk types in Cytyc PreservCyt liquid-based cytology specimens. OBJECTIVES: To compare AHPV analytical sensitivity for RNA and DNA; to compare the sensitivity of AHPV and Hybrid Capture 2 (HC2) assays for HPV DNA detection; to compare assay performance with and without sample denaturation; to compare assay results with cytology. STUDY DESIGN: Analytical sensitivity of AHPV for detecting E6/E7 RNA was assessed by spiking samples with various quantities of HPV 16 E6/E7 in vitro RNA transcript or HPV 16-positive SiHa cells. AHPV and HC2 analytical sensitivity for HPV 16 DNA was evaluated by spiking samples with various quantities of a plasmid vector containing cloned HPV 16 DNA, or with purified SiHa cell genomic DNA containing integrated HPV 16 genome. Samples were tested using standard AHPV and HC2 protocols. Endocervical samples from 568 women were collected in Digene Specimen Transport Medium. Non-denatured and denatured samples were tested in AHPV and denatured samples with HC2. Assay results were compared to each other, and to cytology. RESULTS: AHPV had substantially higher (2 4 log(10)) analytical sensitivity for HPV 16 RNA than for HPV 16 DNA. AHPV also had substantially lower (3 log(10)) analytical sensitivity for HPV 16 DNA compared to HC2. The overall agreement between assay results in clinical specimens was 94.2%, but AHPV had fewer positives than HC2 (48.4% positive agreement). In denatured samples, the number of samples testing positive in AHPV increased two-fold, yielding a positive agreement rate of 88.7%. When assay results were compared with cytology, AHPV had fewer positives in samples with normal or ASC-US diagnoses than did HC2. CONCLUSIONS: AHPV is much more efficient at detecting HPV 16 RNA than DNA. Selective capture, amplification, and detection of HPV RNA by AHPV may improve the specificity of molecular HPV testing.
Assuntos
Colo do Útero/virologia , DNA Viral/análise , Papillomaviridae/genética , Infecções por Papillomavirus/virologia , RNA Mensageiro/análise , RNA Viral/análise , Kit de Reagentes para Diagnóstico , Neoplasias do Colo do Útero/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , DNA Viral/genética , Feminino , Humanos , Pessoa de Meia-Idade , RNA Mensageiro/genética , RNA Viral/genética , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Occult hepatitis B virus (HBV) is defined by the presence of HBV DNA in individuals with HBV core antibodies (anti-HBc) but without HBV surface antigen (HBsAg). The prevalence of occult HBV in HIV-infected patients remains controversial, and the risk factors and clinical significance are unknown. OBJECTIVES: To determine the prevalence, risk factors, and clinical significance of occult HBV among HIV-infected patients. Hypothesized risk factors include chronic hepatitis C virus (HCV), CD4 count < 200 cells/mm, HIV RNA level >1000 copies/mL, and lack of use of anti-HBV antiretrovirals. METHODS: We examined randomly selected HBsAg/anti-HBc HIV-infected patients in the Penn Center for AIDS Research Adult/Adolescent Database and Specimen Repository. HBV DNA was qualitatively detected using a transcription-mediated amplification assay. Risk factors and transaminases were ascertained at the time sera were collected. RESULTS: A total of 699 HBsAg/anti-HBc HIV-infected patients were identified. Of 179 randomly selected subjects, 17 (10%; 95% confidence interval [CI]: 5% to 14%) had occult HBV. Differences in the prevalence of HBV surface antibody (anti-HBs) between those with (7 [41%]) and without (94 [58%]) occult HBV were not statistically significant (P = 0.3). An HIV RNA level >1000 copies/mL (adjusted odds ratio [OR] = 4.88, 95% CI: 1.01 to 30.26) and the absence of chronic HCV (adjusted OR = 0.26, 95% CI: 0.05 to 0.95) were associated with occult HBV. Occult HBV did not increase the risk of transaminitis (adjusted OR = 0.42, 95% CI: 0.12 to 1.45). CONCLUSIONS: Occult HBV occurred in a sizable proportion of HIV-infected patients and was associated with detectable HIV and the absence of chronic HCV. It did not increase the risk of transaminitis. The presence of anti-HBs does not rule out occult HBV. Future studies should examine the long-term clinical implications of occult HBV in HIV-infected patients.
Assuntos
Infecções por HIV/complicações , Vírus da Hepatite B/isolamento & purificação , Hepatite B/epidemiologia , Adulto , DNA Viral/sangue , Feminino , HIV-1/isolamento & purificação , Hepacivirus/isolamento & purificação , Hepatite B/fisiopatologia , Hepatite B/virologia , Anticorpos Anti-Hepatite B/sangue , Antígenos de Superfície da Hepatite B/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Técnicas de Amplificação de Ácido Nucleico , Philadelphia , Prevalência , RNA Viral/análise , Fatores de Risco , Estatística como Assunto , Transaminases/sangue , Carga ViralRESUMO
The PROCLEIX West Nile virus assay (WNV assay) is a qualitative nucleic acid test based on transcription-mediated amplification (TMA). The assay was used under an investigational protocol in the United States to screen blood donations for West Nile virus (WNV) RNA starting in the summer of 2003, and was licensed by the FDA in December 2005 for use on the PROCLEIX System, also known as the enhanced semi-automated system (eSAS). Performance characteristics for the assay were determined on both eSAS and the fully automated PROCLEIX TIGRIS (TIGRIS) System. Detection of both lineage 1 and lineage 2 WNV was demonstrated on both systems. For lineage 1, the 95% detection limit was 8.2 copies/ml for eSAS and 9.8 copies/ml for the TIGRIS system. For lineage 2, > or =95% detection was seen at > or =30 copies/ml on both systems. The overall specificity of the assay was >99.9% in fresh and frozen plasma specimens. Reproducibility studies on the TIGRIS system yielded > or =99.1% agreement with expected results for the 3-member panel tested (0, 30, and 100 copies/ml). The WNV assay exhibited robust performance in cadaveric specimens and specimens representing various donor and donation conditions, including specimens from different plasma collection tubes that were subjected to multiple freeze/thaw cycles; specimens with elevated levels of endogenous substances; specimens containing other viruses and microorganisms; and specimens from patients with autoimmune and other diseases. Overall, these studies demonstrate high sensitivity, specificity, and reproducibility of the WNV assay on both the semi-automated and automated systems.
Assuntos
Técnicas de Amplificação de Ácido Nucleico/métodos , Vírus do Nilo Ocidental/isolamento & purificação , Bioensaio , Humanos , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: It is important to characterize viral dynamics in early hepatitis C virus (HCV) infection to further our understanding of viral pathogenesis and the potential for secondary transmission in acute infection through blood transfusion or other routes. STUDY DESIGN AND METHODS: Serial units given by 77 source plasma donors who had evolved from HCV RNA-negative to HCV RNA-positive by nucleic acid amplification technology (NAT) screening with 512-unit pool-NAT or were followed from RNA detection to antibody conversion were tested by individual NAT and quantitative RNA assays. RESULTS: During the ramp-up phase when exponential growth occurs, HCV viral load doubled every 10.8 hours (95% confidence interval [CI], 9.9-12.0). Intermittent viremia was observed before the ramp-up phase in 37 of 50 panels with the earliest detectable viremic bleed occurring 63 days before the estimated onset of ramp-up. The plateau phase or high-titer viremic period that occurs between ramp-up and seroconversion was estimated to last 56.3 days (95% CI, 44.8-67.8). CONCLUSIONS: Intermittent low-level HCV viremia can occur as much as 2 months before the periods of exponential increase in viral load and the high-titer plateau-phase viremia that usually precede seroconversion. Animal inoculation studies are in progress to evaluate if transfusion of low-level viremic plasma can transmit HCV infection.
Assuntos
Hepacivirus/genética , Hepatite C Crônica/diagnóstico , Técnicas de Amplificação de Ácido Nucleico/métodos , RNA Viral/sangue , Viremia/diagnóstico , Doadores de Sangue , Hepacivirus/isolamento & purificação , Hepatite C Crônica/sangue , Hepatite C Crônica/transmissão , Estudos Retrospectivos , Testes Sorológicos/métodos , Fatores de Tempo , Carga Viral , Viremia/sangue , Viremia/transmissão , Replicação ViralRESUMO
Since 1980, the Transfusion-transmitted Viruses Study (TTVS) (1974-1980) has continued to maintain its computerized database and stored sera to enable ongoing study of new transfusion events since the 1970s. Most recently, we have used this resource to study parameters of acute hepatitis C virus (HCV) infection among 94 donor-recipient pairs in which there was transmission. In addition, frequent recipient observations permitted further characterization of the early phase of the infection's course. Donor RNA load ranged from 3.7 to 3,160,000 IU/mL. Onset of recipient viremia was judged from a total of 67 sera collected during the 4th through 8th days posttransfusion; only 2 of the 67 sera were still RNA nonreactive by that time. The recipients' latent periods to an alanine aminotransferase (ALT) elevation of > or =90 IU/L ranged from 6 to 112 days (median, 46 days) and was shorter with higher donor RNA levels. Descriptors of the recipient's illness showed several strongly positive and negative correlations. The latent period tended to be shorter in the 37% of cases that were clinically overt. Attributes of donors with genotypes 1 and non-1 and subtypes 1a and 1b did not differ significantly. Recipients with genotype 1 strains had shorter latent intervals than non-1 strains. On multivariate analysis, latent period was significantly associated (negatively) only with the highest ALT level during the first 120 days of follow-up (P = .014). In conclusion, host factors are more important determinants of acute HCV infection dynamics than virus-associated factors.
Assuntos
Hepacivirus/imunologia , Hepatite C/imunologia , Carga Viral , Adulto , Feminino , Humanos , Masculino , RNA Viral/isolamento & purificaçãoRESUMO
BACKGROUND: Screening donors for human immunodeficiency virus (HIV) and hepatitis C virus (HCV) RNA is primarily performed on minipools (MPs) with one of two commercial nucleic acid amplification tests (NAT; Roche Molecular Systems; or Gen-Probe/Chiron). We compared these assays with respect to detection of RNA in early HIV and HCV infection. STUDY DESIGN AND METHODS: Twelve HIV plasma donor panels (116 serial samples) and 12 HCV panels (180 serial samples) were selected to optimally represent early viremia. Initial testing was performed in singlicate or triplicate on separately coded aliquots, both neat and at dilutions corresponding to MP screening (1:16 for Gen-Probe; 1:24 for Roche); 20 additional replicates were performed when discordant results were observed. Odds ratios (ORs) comparing detection of RNA by different assays were derived with logistic regression models. Differences in window-period closure and yields of assays in MP or individual-donation (ID) format were estimated. RESULTS: Differences in detection rates between Roche and Gen-Probe NAT assays were small and only observed with samples with very-low-level viremia. ORs for detecting RNA by the Gen-Probe versus the Roche assay were significant for HIV if conducted on MPs (1.8; 95% confidence interval [CI], 1.3-2.5) but not neat (1.0; 95% CI, 0.72-1.4). Odds of detecting HCV RNA were higher if the Gen-Probe assay was conducted either neat (2.3; 95% CI, 1.6-3.2) or on MPs (4.0; 95% CI, 2.8-5.8). These differences translated to <1 day window-period closure and Assuntos
Infecções por HIV/diagnóstico
, HIV/genética
, Hepacivirus/genética
, Hepatite C/diagnóstico
, Técnicas de Amplificação de Ácido Nucleico/métodos
, Armazenamento de Sangue/métodos
, HIV/isolamento & purificação
, Infecções por HIV/sangue
, Hepacivirus/isolamento & purificação
, Hepatite C/sangue
, Humanos
, Licenciamento
, Programas de Rastreamento/métodos
, RNA Viral/sangue
, RNA Viral/isolamento & purificação
, Sensibilidade e Especificidade
, Viremia/sangue
, Viremia/diagnóstico
RESUMO
While the present generation of serology-based assays has significantly decreased the number of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) infections acquired by transfusion, the possibility of infected donations escaping detection still exists. The average seronegative viremic window duration during which immunological assays are unable to detect the virus is estimated to be between 16 and 22 days for HIV-1 and approximately 70 days for HCV. Significant reduction of detection window duration was demonstrated using a nucleic acid amplification assay, the Procleix HIV-1/HCV Assay, which utilizes transcription-mediated amplification technology to simultaneously detect HIV-1 and HCV RNAs. For 26 commercially available HIV-1 seroconversion panels tested, specimens were reactive in the HIV-1/HCV assay at the same time as or earlier than in serological assays. Overall, the HIV-1/HCV assay was able to reduce the detection window duration by an average of 14 days and 6 days compared to tests relying on recognition of HIV-1 antibody and p24 antigen, respectively. For 24 commercially available HCV seroconversion panels tested, the specimens were reactive in the HIV-1/HCV assay at an earlier blood sampling date than in serological assays, reducing the detection window duration by an average of 26 days. Similar results were obtained in testing the HIV-1 and HCV seroconversion panels in the virus-specific HIV-1- and HCV-discriminatory assays, respectively. In conclusion, the HIV-1/HCV assay and corresponding discriminatory assays significantly reduced detection window durations compared to immunoassays.
Assuntos
Soropositividade para HIV/diagnóstico , HIV-1/isolamento & purificação , Hepacivirus/isolamento & purificação , Hepatite C/diagnóstico , Técnicas de Amplificação de Ácido Nucleico , HIV-1/genética , Hepacivirus/genética , Hepatite C/transmissão , Humanos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Fatores de Tempo , Reação TransfusionalRESUMO
The risk of hepatitis B virus (HBV) transmission by transfusion in sub-Saharan Africa is considered to be relatively low, and testing of blood donors is often not done or is done relatively poorly. To re-examine this attitude, we identified HBV chronically infected blood donors from a major hospital in Ghana with a range of hepatitis B surface antigen (HBsAg) assays. Test efficacy was estimated using HBV DNA as a gold standard, and the risk of HBV infection in blood recipients was estimated for different testing strategies. Particle agglutination, dipstick, and enzyme immunoassay (EIA) HBsAg screening detected 54%, 71%, and 97% of HBV infectious donors, respectively. The risk of HBV transmission to recipients less than 10 years old ranged between 1:11 and 1:326 with blood unscreened and screened by EIA, respectively. For older recipients, the risk decreased a further 4-fold because of the high frequency of natural exposure to HBV. A total of 98% of HBsAg-confirmed positive samples contained HBV DNA. HBV DNA load was less than 1 x 10(4) IU/mL in 75% of HBsAg-reactive samples, most of them anti-HBe reactive. Approximately 0.5% of HBsAg-negative but anti-HBc-positive samples contained HBV DNA. The use of sensitive HBsAg tests is critical to prevent transfusion transmission of HBV infection to young children in a population with a 15% prevalence of chronic HBV infection in blood donors. However, this will not have much effect on the prevalence of this infection unless other strategies to protect children from infection are also advanced in parallel.