HIV oral self-screening test among HIV/STD/TB clinic attendees: A mixed-method pilot investigation examining merit for larger evaluation.

Background & objectives: Globally, several countries consider HIV self-test as an important element in the toolbox to end AIDS by 2030. Against this background, the present investigation was conducted to pilot test the performance of an indigenous HIV oral self-test (HIVOST) and explore its acceptability. The overall purpose was to examine if this kit could serve as a promising tool and merit future larger clinical evaluation. Methods: A concurrent mixed-method investigation was undertaken during March-October 2019. One hundred and thirty two consecutive HIV/sexually transmitted diseases/tuberculosis clinic attendees were invited for participation; of whom, 100 were enrolled, and among them, 40 provided consent for qualitative in-depth interviews. The HIVOST kit assessed for its performance served as the 'index test', which worked on the principle of lateral flow chromatography. The results of the HIVOST were interpreted independently by the study physicians and participants at 20 min. HIVOST kit performance was assessed against the HIV confirmatory blood test result based on the national algorithm (3 rapid test or 1 ELISA and 2 rapid test) serving as the 'reference'. Sensitivity, specificity, positive predictive value, negative predictive value and inter-rater agreement were estimated. The voices and concerns of the study participants were coded followed by identification of qualitative themes and ideas. Results: The sensitivity and specificity of the index test at the end of 20 min as interpreted by the participants were 83.3 per cent [95% confidence interval (CI): 69.8 to 92.5] and 98 per cent (95% CI: 89.4 to 99.5), respectively. Study physicians and participants independently interpreted HIVOST results with substantial inter-rater agreement (kappa value 0.88; 95% CI: 0.78-0.97). All HIVOST test strips were valid. Majority of the participants preferred saliva over blood for HIV self-test. 'Comfort', 'confidentiality' and 'convenience' were the perceived advantages of HIVOST. Some of the participants wished the package inserts contained 'how-to-do instructions in local languages', 'expiry date (if any)' and 'contact helpline number'. A few of them highlighted the need for a confirmatory HIV result following oral self-test. Concerns of the participants revolved around potential self-harm following HIVOST-positive result and safe disposal of kits. Interpretation & conclusions: Two major highlights of the present investigation are (i) high level of concordance in HIVOST results interpreted by participants and physicians, and (ii) encouraging level of acceptance of HIVOST. These findings and encouraging HIVOST performance statistics lend support towards large-scale clinical evaluation of this index test.

Phylogenetic Analysis of Spread of Hepatitis C Virus Identified during HIV Outbreak Investigation, Unnao, India.

An HIV outbreak investigation during 2017-2018 in Unnao District, Uttar Pradesh, India, unearthed high prevalence of hepatitis C virus (HCV) antibodies among the study participants. We investigated these HCV infections by analyzing NS5B and core regions. We observed no correlation between HIV-HCV viral loads and clustering of HCV sequences, regardless of HIV serostatus. All HCV isolates belonged to genotype 3a. Monophyletic clustering of isolates in NS5B phylogeny indicates emergence of the outbreak from a single isolate or its closely related descendants. The nucleotide substitution rate for NS5B was 6 × 10-3 and for core was 2 × 10-3 substitutions/site/year. Estimated time to most recent common ancestor of these isolates was 2012, aligning with the timeline of this outbreak, which might be attributable to unsafe injection practices while seeking healthcare. HIV-HCV co-infection underlines the need for integrated testing, surveillance, strengthening of healthcare systems, community empowerment, and molecular analyses as pragmatic public health tools.

Community-based point-of-care testing to identify new HIV infections: A cross-sectional study from Pune, India.

ABSTRACT: The World Health Organization recommends point-of-care testing (POCT) to detect human immunodeficiency virus (HIV) infected individuals in the community. This will help improve treatment coverage through detection of HIV infection among those who are unaware of their status.This study was planned with an objective to investigate the feasibility and acceptability of POCT for HIV in the community.A community-based cross-sectional study was conducted in rural and peri-urban areas of Pune, India. These sites were selected based on the distance from the nearest HIV testing center. Testing locations were identified in consultation with the local stakeholders and grass-root health workers to identify and capture the priority population. The POCT was performed on blood samples collected by the finger-prick method.The proportion of participants seeking HIV tests for the first time was 79.6% that signifies the feasibility of POCT. The acceptability in the peri-urban and rural areas was 70.2% and 69.7%, respectively. POCT was performed at construction sites (24.9%), nearby industries (16.1%) and parking areas of long-distance trucks (8.1%) in the peri-urban area. Three newly diagnosed HIV-infected participants (0.1%) were detected from the peri-urban areas but none from the rural areas. Two of the newly diagnosed participants and their spouses were linked to care.There was a high acceptability of POCT and wider coverage of priority population with a strategy of testing at places preferable to the study population. Therefore, we believe that community-based POCT is a promising tool for improving HIV testing coverage even in low prevalence settings with the concentrated HIV epidemic.

Unsterile injection equipment associated with HIV outbreak and an extremely high prevalence of HCV-A case-control investigation from Unnao, India.

The integrated counseling and testing center (ICTC) located in the district hospital, Unnao in the northern state of Uttar Pradesh (UP), India witnessed an increased detection of HIV among its attendees in July 2017. Subsequently, health camps were organized by the UP State AIDS Control Society in the villages and townships contributing to such detection. We conducted a case-control study to identify factors associated with this increased detection; 33 cases and 125 controls were enrolled. Cases were individuals, detected HIV sero-reactive during November 2017-April 2018 from three locations namely Premganj, Karimuddinpur and Chakmeerapur in the Bangarmau block of the district of Unnao. Controls hailed from the same geographical setting and tested HIV sero-nonreactive either in health camps or at ICTC centers from where the cases were detected. Misclassification bias was avoided by confirming HIV sero-status of both cases as well as controls prior to final analysis. Study participants were interviewed on various risk practices and invasive treatment procedures. They were also tested for HIV and other bio-markers reflecting unsafe injecting and sexual exposures such as hepatitis B surface antigen (HBsAg), anti-HCV antibody (HCV Ab), anti-herpes simplex-2 Immunoglobulin G (HSV-2 IgG) and rapid plasma regain (RPR) test for syphilis. Secondary data analysis on three time points during 2015 through 2018 revealed a rising trend of HIV among attendees of the ICTCs (ICTC-Hasanganj, ICTC-Unnao district hospital and ICTC- Nawabganj) catering to the entire district of Unnao. While there was a seven fold rise of HIV among ICTC attendees of Hasanganj (χ2 value for trend 23.83; p < 0.001), the rise in Unnao district hospital was twofold (χ2 value for trend 4.37; p < 0.05) and was tenfold at ICTC-Nawabganj (χ2 value for trend 5.23; p < 0.05) indicating risk of infection prevailing throughout the district. Primary data was generated through interviews and laboratory investigations as mentioned above. The median age of cases and controls was 50 year (minimum 18 -maximum 68; IQR 31-57) and 38 year (minimum 18 -maximum 78; IQR 29-50) respectively. Thirty six percent of the cases and 47% of controls were male. A significantly higher proportion of cases (85%) had HCV Ab compared to controls (56%; OR 4.4, 95% CI 1.5-12.1); none reported injection drug use. However, cases and controls did not differ significantly regarding presence of HSV-2 IgG (6% versus 8% respectively). Neither any significant difference existed between cases and controls in terms of receiving blood transfusion, undergoing invasive surgical procedures, tattooing, tonsuring of head or skin piercing. In multivariate logistic regression model, 'unsafe injection exposure during treatment-seeking'(AOR 6.61, 95% CI 1.80-24.18) and 'receipt of intramuscular injection in last five years' (AOR 7.20, 95% CI 1.48-34.88) were independently associated with HIV sero-reactive status. The monophyletic clustering of HIV sequences from 14 cases (HIV-1 pol gene amplified) indicated a common ancestry. Availability of auto-disabled syringes and needles, empowerment of the local communities and effective regulatory practices across care settings would serve as important intervention measures in this context.

Characterization of major capsid protein (L1) variants of Human papillomavirus type 16 by cervical neoplastic status in Indian women: Phylogenetic and functional analysis.

The etiological role of infection with Human papillomavirus type 16 (HPV16) in cervical cancer is well established. HPV16 variants, classified based on less than 10% nucleotide variations in the major capsid (L1 ORF) are known to contribute to persistent infection leading to cancer development. L1 protein forms the cornerstone of HPV structure and antigenicity. In the present study, HPV16 L1 variants were characterized by cervical lesion grade and variations in sequences were correlated to structure and function. The L1 gene was analyzed in 152 HPV16 positive cervical samples obtained from Indian women using polymerase chain reaction-directed sequencing. Phylogenetic analysis was carried out for lineage typing. Sixty-one SNPs were detected in L1 genes resulting in 20 nonsynonymous amino acid substitutions of which N56T, N92T, L158F, V178G, N181I, K236T, K443Q, K454T, and K475R are reported in Indian isolates for the first time. The substitutions N181T, T353P, and T389S were significantly associated with high-grade cervical disease. The predominance of lineage A (A1-A4, 84.96%) was observed among the isolates, while the D3 sublineage showed significant association with high-grade cervical lesions. No evidence for recombination and the positive selection was obtained. These substitutions, when mapped on three-dimensional structure, revealed that 11 and 4 substitutions are part of experimentally validated B- and T-cell epitopes, of which T266A and N285T were common to both types of epitopes and may impact HPV vaccine efficacy. The variants identified through this study have the potential to serve as translational leads for designing diagnostic probes and vaccines.

Polymorphisms in Toll-Like Receptors (TLRs)-7 and 9 Genes in Indian Population with Progressive and Nonprogressive HIV-1 Infection.

The polymorphisms in Toll-like receptor (TLR) 7 and 9 genes are shown to influence HIV-1 infection. We studied HIV-1-infected Indian individuals for presence and association of TLR7 and 9 gene polymorphism with different disease outcomes. Genomic DNA from 65 HIV-infected individuals (35 long-term nonprogressors and 30 progressors) and 89 uninfected healthy donors was isolated, amplified, and sequenced for the reported polymorphisms in TLR7 [Gln11Leu (A/T); rs179008] and TLR9 (1635A/G; rs352140) genes. Of these, only the reported TLR9 single-nucleotide polymorphism [SNP; p = .017, odds ratio (OR) = 0.20] and its allele A frequency (p = .038, OR = 0.41) were found to be associated with slow disease progression. Of the new SNPs observed (three TLR7 and two TLR9), the TLR7 rs2074109 G allele showed less likely association with HIV-1 acquisition (p = .019, OR = 0.27). These findings indicate that TLR7 SNP (rs2074109) could be one of the factors for predisposition to HIV-1 and TLR9 1635A/G genotype and allele might have a role in HIV-1 disease progression in Indian population.

Seroprevalence of spotted fever group and typhus group rickettsiae in individuals with acute febrile illness from Gorakhpur, India.

BACKGROUND: The aim of this study was to estimate the seroprevalence of spotted fever group (SFG) and typhus group (TG) rickettsiae among individuals with acute febrile illness (AFI) in the scrub typhus endemic district of Gorakhpur in India. This district is one of the worst affected by annual seasonal acute encephalitis syndrome (AES) outbreaks. METHODS: Antibodies against SFG and TG rickettsiae and the associated risk factors were determined in 294 individuals presenting with an AFI, encountered during a community-based survey conducted during the AES outbreak period October-November 2016. RESULTS: Respective IgM and IgG seropositivity was 13.6% and 36.7% for SFG, and 7.1% and 15.3% for TG. SFG IgM positivity was significantly higher among females, while IgG positivity was significantly higher among individuals ≥45 years of age. IgM and IgG seropositivity for TG rickettsiae were significantly higher in individuals involved in outdoor activities and housewives, but did not differ according to age group, sex, or educational status. CONCLUSION: The study results present serological evidence of SFG and TG rickettsiosis, in addition to scrub typhus, among individuals with AFI in Gorakhpur region and indicate the need to explore their roles as potential causes of AES in the region.

Antibiotic susceptibility pattern of Neisseria gonorrhoeae strains isolated from five cities in India during 2013-2016.

PURPOSE: Emergence of multidrug resistance in Neisseria gonorrhoeae, an STI of public health significance is the biggest challenge to gonorrhoea control. Monitoring for antimicrobial resistance is essential for the early detection of emergent drug resistance patterns. METHODOLOGY: One hundred and twenty four N. gonorrhoeae strains were isolated between September 2013-August 2016 [82-New Delhi, 3-Pune, 3-Mumbai, 20-Secunderabad and 16-Hyderabad] to determine antimicrobial susceptibility and to compare the CLSI disc diffusion method with Etest for these strains. The results of the two methods were compared by using kappa statistics. RESULTS: Ninety eight percent [CI: 96.2-100] of isolates were resistant to ciprofloxacin, 52 % [CI: 43.2-60.8] to penicillin, 56 % [CI: 47.2-64.7] to tetracycline and 5 % [CI: 1.2-8.8] to azithromycin. All the strains were susceptible to spectinomycin, ceftriaxone and cefixime except for two strains which showed decreased susceptibility to ceftriaxone and cefixime. Kappa scores for penicillin, azithromycin, ciprofloxacin, ceftriaxone and cefixime showed that the CLSI method had high agreement with Etest while tetracycline had substantial agreement. CONCLUSION: Our data suggest that the disc diffusion method which is both cost effective and more feasible, can effectively be used routinely for monitoring antibiotic susceptibility in N. gonorrhoeae, in limited resource countries like India. We demonstrate the emergence of decreased susceptibility to ceftriaxone and cefixime and threshold levels of resistance to azithromycin in India. This underscores the importance of maintaining continued surveillance for antibiotic resistance in N. gonorrhoeae and a potential requirement for strategic change in guidelines in the not so distant future.

Rates and determinants of incidence and clearance of cervical HPV genotypes among HIV-seropositive women in Pune, India.

BACKGROUND: Several studies in recent years have documented the genotype-specific prevalence of HPV infection and wide diversity and multiplicity of HPV genotypes among HIV-seropositive women. Yet, information on changes in HPV genotype-specific incidence and clearance rates over time, and their correlation with clinical or immunologic factors among HIV-seropositive women is scarce. OBJECTIVES: We conducted a prospective study to investigate the incidence and clearance rates of cervical HPV genotypes among HIV-seropositive women in India and expand the evidence base in this area of research. STUDY DESIGN: Cervical samples were collected from n=215 HIV-seropositive women in Pune, India who underwent two screening visits separated by a median of 11-months (interquartile range: 8-18 months). HPV genotypes were determined by Roche Linear Array HPV assay. Individual genotype-specific and carcinogenicity-grouping-specific HPV incidence and clearance rates were calculated and the associations between incidence/clearance and age and HIV-related metrics were explored. RESULTS: Incidence and clearance rates for 'any HPV' and 'carcinogenic HPV' genotypes were 11.1 and 18.3, and 6.7 and 33.8, per 100 person-years, respectively. Incidence and clearance rates for HPV genotypes of alpha-9 species (HPV16, HPV31, HPV33, HPV35, HPV52 and HPV58) and alpha-7 species (HPV18, HPV39, HPV45, HPV59 and HPV68) were 5.8 and 2.04, and 32.1 and 53.5, per 100 person-years, respectively. Clearance of any HPV type was associated with increasing age of participants (odds ratio: 1.08, 95%CI: 1.004-1.17), although the association marginally lost its statistical significance when adjusted for CD4 counts and antiretroviral therapy status. CONCLUSIONS: Genotype-specific clearance rates of HPV were higher than corresponding incidence rates. The suggestion of a positive associations of increasing age with HPV clearance points to the need for etiologic studies on age-related hormonal changes on clearance of cervical HPV infection.

Cognitive Function among HIV Infected Children in Pune.

OBJECTIVE: To assess cognitive function of school-aged, perinatally acquired human immunodeficiency virus (HIV)-infected, antiretroviral therapy (ART) naïve children in Pune. METHODS: HIV infected ART naïve children (n = 50) between 6 and 12 y attending outpatient clinic were administered the Indian Child Intelligence Test (ICIT) after their clinical evaluation. Age and gender matched HIV uninfected children (n = 50) were enrolled as controls. The test battery included measures of perception of form (Gestalt), concept formation, memory span, learning and remembering, associative memory span, transformation of visual field, quantitative concepts, spatial visualization and visual motor coordination. The information on adaptive behavior was obtained from the care giver and a brief interview with the child using Vineland Social Maturity Scale (VSMS). RESULTS: Mean age of the children was 9.3 y (SD: 1.3) and mean education was 4.3 y (SD: 1.3). The mean CD4 count of HIV infected children was 588 cells/mm(3) (SD: 324.4). There was no significant difference in the overall ICIT scores between HIV infected and uninfected children (p 0.209). The adaptive behavior (VSMS) scores were significantly less in HIV infected children as compared to others (p 0.028). CONCLUSIONS: There was no significant difference in the overall cognitive abilities in HIV infected children using ICIT as compared to HIV uninfected children, although significant difference in the adaptive behavior between the two groups was observed. It is important to conduct larger prospective studies using standardized test batteries that are inclusive of multiple cognitive domains sensitive to HIV effects in Indian children.

Evaluation of a cost effective in-house method for HIV-1 drug resistance genotyping using plasma samples.

OBJECTIVES: Validation of a cost effective in-house method for HIV-1 drug resistance genotyping using plasma samples. DESIGN: The validation includes the establishment of analytical performance characteristics such as accuracy, reproducibility, precision and sensitivity. METHODS: The accuracy was assessed by comparing 26 paired Virological Quality Assessment (VQA) proficiency testing panel sequences generated by in-house and ViroSeq Genotyping System 2.0 (Celera Diagnostics, US) as a gold standard. The reproducibility and precision were carried out on five samples with five replicates representing multiple HIV-1 subtypes (A, B, C) and resistance patterns. The amplification sensitivity was evaluated on HIV-1 positive plasma samples (n = 88) with known viral loads ranges from 1000-1.8 million RNA copies/ml. RESULTS: Comparison of the nucleotide sequences generated by ViroSeq and in-house method showed 99.41±0.46 and 99.68±0.35% mean nucleotide and amino acid identity respectively. Out of 135 Stanford HIVdb listed HIV-1 drug resistance mutations, partial discordance was observed at 15 positions and complete discordance was absent. The reproducibility and precision study showed high nucleotide sequence identities i.e. 99.88±0.10 and 99.82±0.20 respectively. The in-house method showed 100% analytical sensitivity on the samples with HIV-1 viral load >1000 RNA copies/ml. The cost of running the in-house method is only 50% of that for ViroSeq method (112$ vs 300$), thus making it cost effective. CONCLUSIONS: The validated cost effective in-house method may be used to collect surveillance data on the emergence and transmission of HIV-1 drug resistance in resource limited countries. Moreover, the wide applications of a cost effective and validated in-house method for HIV-1 drug resistance testing will facilitate the decision making for the appropriate management of HIV infected patients.

Correlates of sex initiation among school going adolescents in Pune, India.

OBJECTIVE: To identify the correlates of sex initiation among school going adolescents in Pune, India. METHODS: A study among 910 school going adolescents was conducted in five schools and one junior college in and around Pune, Maharashtra (India) between 2003-2006. Case control analysis (n = 205) was performed among 41 cases who reported ever having sex and 164 controls matched for gender, location and type of school. Correlates of sex initiation were identified using conditional logistic regression. RESULTS: Adolescents studying in vernacular schools, accessing pornography and having unfriendly relationship with parents had higher likelihood of sex initiation. Adolescents who reported sexual abuse, Sexually Transmitted Diseases (STDs) symptoms, smoking habit and those who had not read scientific literature on reproductive and sexual health were more likely to have initiated sex early. In the multivariate model, unfriendly relationship with parents and reported symptoms of STDs were found to be independently associated with 'early initiation of sex' among school going adolescents in this study. CONCLUSIONS: Premarital sexual activity, both consensual and non-consensual, was reported indicating a need for school based adolescent reproductive and sexual health education (ARSHE) programs in Maharashtra, India. The program in India should focus on specialized interventions for young adolescents. Routine health check-ups and probing on symptoms of STDs, non-consensual sex and other risky practices should be implemented in schools.

HPV genotype distribution in cervical intraepithelial neoplasia among HIV-infected women in Pune, India.

BACKGROUND: The distribution of HPV genotypes, their association with rigorously confirmed cervical precancer endpoints, and factors associated with HPV infection have not been previously documented among HIV-infected women in India. We conducted an observational study to expand this evidence base in this population at high risk of cervical cancer. METHODS: HIV-infected women (N = 278) in Pune, India underwent HPV genotyping by Linear Array assay. Cervical intraepithelial neoplasia (CIN) disease ascertainment was maximized by detailed assessment using cytology, colposcopy, and histopathology and a composite endpoint. RESULTS: CIN2+ was detected in 11.2% while CIN3 was present in 4.7% participants. HPV genotypes were present in 52.5% (146/278) and 'carcinogenic' HPV genotypes were present in 35.3% (98/278) HIV-infected women. 'Possibly carcinogenic' and 'non/unknown carcinogenic' HPV genotypes were present in 14.7% and 29.5% participants respectively. Multiple (≥ 2) HPV genotypes were present in half (50.7%) of women with HPV, while multiple 'carcinogenic' HPV genotypes were present in just over a quarter (27.8%) of women with 'carcinogenic' HPV. HPV16 was the commonest genotype, present in 12% overall, as well as in 47% and 50% in CIN2+ and CIN3 lesions with a single carcinogenic HPV infection, respectively. The carcinogenic HPV genotypes in declining order of prevalence overall included HPV 16, 56, 18, 39, 35, 51, 31, 59, 33, 58, 68, 45 and 52. Factors independently associated with 'carcinogenic' HPV type detection were reporting ≥ 2 lifetime sexual partners and having lower CD4+ count. HPV16 detection was associated with lower CD4+ cell counts and currently receiving combination antiretroviral therapy. CONCLUSION: HPV16 was the most common HPV genotype, although a wide diversity and high multiplicity of HPV genotypes was observed. Type-specific attribution of carcinogenic HPV genotypes in CIN3 lesions in HIV-infected women, and etiologic significance of concurrently present non/unknown carcinogenic HPV genotypes await larger studies.

Predictors of retention among men attending STI clinics in HIV prevention programs and research: a case control study in Pune, India.

BACKGROUND: Retention is critical in HIV prevention programs and clinical research. We studied retention in the three modeled scenarios of primary prevention programs, cohort studies and clinical trials to identify predictors of retention. METHODOLOGY/PRINCIPAL FINDINGS: Men attending Sexually Transmitted Infection (STI) clinics (n = 10, 801) were followed in a cohort study spanning over a ten year period (1993-2002) in Pune, India. Using pre-set definitions, cases with optimal retention in prevention program (n = 1286), cohort study (n = 940) and clinical trial (n = 896) were identified from this cohort. Equal number of controls matched for age and period of enrollment were selected. A case control analysis using conditional logistic regression was performed. Being employed was a predictor of lower retention in all the three modeled scenarios. Presence of genital ulcer disease (GUD), history of commercial sex work and living away from the family were predictors of lower retention in primary prevention, cohort study and clinical trial models respectively. Alcohol consumption predicted lower retention in cohort study and clinical trial models. Married monogamous men were less likely to be retained in the primary prevention and cohort study models. CONCLUSIONS/SIGNIFICANCE: Predicting potential drop-outs among the beneficiaries or research participants at entry point in the prevention programs and research respectively is possible. Suitable interventions might help in optimizing retention. Customized counseling to prepare the clients properly may help in their retention.