Prevalence of cryptococcal antigenemia and cost-effectiveness of a cryptococcal antigen screening program--Vietnam.

BACKGROUND: An estimated 120,000 HIV-associated cryptococcal meningitis (CM) cases occur each year in South and Southeast Asia; early treatment may improve outcomes. The World Health Organization (WHO) recently recommended screening HIV-infected adults with CD4<100 cells/mm(3) for serum cryptococcal antigen (CrAg), a marker of early cryptococcal infection, in areas of high CrAg prevalence. We evaluated CrAg prevalence and cost-effectiveness of this screening strategy in HIV-infected adults in northern and southern Vietnam. METHODS: Serum samples were collected and stored during 2009-2012 in Hanoi and Ho Chi Minh City, Vietnam, from HIV-infected, ART-naïve patients presenting to care in 12 clinics. All specimens from patients with CD4<100 cells/mm(3) were tested using the CrAg lateral flow assay. We obtained cost estimates from laboratory staff, clinicians and hospital administrators in Vietnam, and evaluated cost-effectiveness using WHO guidelines. RESULTS: Sera from 226 patients [104 (46%) from North Vietnam and 122 (54%) from the South] with CD4<100 cells/mm(3) were available for CrAg testing. Median CD4 count was 40 (range 0-99) cells/mm(3). Nine (4%; 95% CI 2-7%) specimens were CrAg-positive. CrAg prevalence was higher in South Vietnam (6%; 95% CI 3-11%) than in North Vietnam (2%; 95% CI 0-6%) (p = 0.18). Cost per life-year gained under a screening scenario was $190, $137, and $119 at CrAg prevalences of 2%, 4% and 6%, respectively. CONCLUSION: CrAg prevalence was higher in southern compared with northern Vietnam; however, CrAg screening would be considered cost-effective by WHO criteria in both regions. Public health officials in Vietnam should consider adding cryptococcal screening to existing national guidelines for HIV/AIDS care.

Natural polymorphisms of HIV-1 CRF01_AE integrase coding region in ARV-naïve individuals in Cambodia, Thailand and Vietnam: an ANRS AC12 working group study.

The HIV integrase enzyme is essential for the HIV life cycle as it mediates integration of HIV-1 proviral DNA into the infected cell's genome. Recently, the development of drugs capable of inhibiting integrase has provided major new options for HIV-infected, treatment-experienced patients with multidrug resistant virus, as well treatment-naïve patients. More than 40 amino acid substitutions within integrase have been described as associated mostly with resistance of HIV B-subtypes to currently available integrase inhibitors (INIs). We have analyzed the natural polymorphisms of the integrase coding region in 87 antiretroviral-naïve subjects (32 from Cambodia, 37 from Thailand and 18 from Vietnam) infected with CRF01_AE virus, the predominant HIV-1 strain circulating in Southeast Asia. The 864bp integrase coding region was sequenced using the ANRS consensus sequencing technique from plasma samples, and amino acid results were interpreted for drug resistance according to the ANRS (Updated July 2009, version 18) and Stanford algorithms (Version November 6, 2009). Alignment of the 87 amino acid sequences against the 2004 Los Alamos HIV-1 clade B consensus sequence showed that overall, 119 of 288 (41.3%) amino acid positions presented at least one polymorphism each. Substitutions found in >60% of study subjects occurred at: K14, A21, V31, S39, I72, T112, T124, T125, G134, I135, K136, D167, V201, L234 and S283. Also, new amino acid substitutions of as yet unknown significance were identified: E152K/H, S153F/L, N155I and E157G. None of the known integrase resistance mutations were observed, except E157Q found in one Cambodian subject (1.1%, CI 95% 0.02-6.3%). The clinical impact of this substitution on resistance of B and nonB-viruses to the licensed INI raltegravir is unclear. If this substitution is confirmed to compromise the virologic response to raltegravir, further studies will be needed to better assess the prevalence of this substitution among CRF01_AE virus.

Failure of human immunodeficiency virus enzyme immunoassay to rule out infection among polymerase chain reaction-negative Vietnamese infants at 12 months of age.

BACKGROUND: : Previous studies have demonstrated that >90% of HIV-uninfected infants serorevert, as seen in the results of enzyme immunoassay (EIA) testing by 12 months of age, making it feasible to confirm or rule out infection. We assessed the reliability of EIA in a cohort of Vietnamese infants. METHODS: : HIV-exposed, uninfected infants enrolled in a parent diagnostic and monitoring study from February 2005 through August 2006 were eligible for inclusion in a prospective cohort study of HIV-EIA performance. Testing using 2 standard assays (Genscreen HIV 1/2 version 2, Bio-Rad; Murex 1.2.0, Murex Biotech) was initiated at 12 months of age. Infants were categorized as EIA-negative (seroreverted; negative Genscreen), EIA-indeterminate (positive Genscreen, negative Murex), or EIA-positive (Genscreen and Murex positive). RESULTS: : Of 273 infants included in the study, 59 (22%) were EIA-negative at 12 months, 131 (48%) were indeterminate, and 83 (30%) were EIA-positive; specificity 21.6 (95% confidence interval: 16.6, 26.3). Infants with positive EIAs at 12 months were 74% more likely than EIA-indeterminate infants to test indeterminate or positive at 18 months (risk ratio, 1.74, 95% confidence interval: 1.15, 2.64; P = 0.03). CONCLUSIONS: : Expectations regarding infant seroreversion by standard EIAs should be reassessed to reflect potential cross-regional differences in their performance.

In-house HIV-1 RNA real-time RT-PCR assays: principle, available tests and usefulness in developing countries.

The principle of currently available licensed HIV-1 RNA assays is based on real-time technologies that continuously monitor the fluorescence emitted by the amplification products. Besides these assays, in-house quantitative (q) real-time reverse transcription (RT)-PCR (RT-qPCR) tests have been developed and evaluated particularly in developing countries, for two main reasons. First, affordable and generalized access to HIV-1 RNA viral load is urgently needed in the context of expected universal access to prevention and antiretroviral treatment programs in these settings. Second, since many non-B subtypes, circulating recombinant forms and unique recombinant forms circulate in these areas, in-house HIV-1 RNA RT-qPCR assays are ideal academic tools to thoroughly evaluate the impact of HIV-1 genetic diversity on the accuracy of HIV-1 RNA quantification, as compared with licensed techniques. To date, at least 15 distinct in-house assays have been designed. They differ by their chemistry and the HIV-1 target sequence (located in gag, Pol-IN or LTR gene). Analytical performances of the tests that have been extensively evaluated appear at least as good as (or even better than) those of approved assays, with regard to HIV-1 strain diversity. Their clinical usefulness has been clearly demonstrated for early diagnosis of pediatric HIV-1 infection and monitoring of highly active antiretroviral therapy efficacy. The LTR-based HIV-1 RNA RT-qPCR assay has been evaluated by several groups under the auspices of the Agence Nationale de Recherches sur le SIDA et les hépatites virales B et C. It exists now as a complete standardized commercial test.

Complete genome analysis of hepatitis C virus subtypes 6t and 6u.

Hepatitis C virus (HCV) genomes exhibit high nucleotide sequence diversity. In this study, we performed complete genome sequence analysis of 11 HCV genotype 6 samples from Vietnam and Thailand. We identified nine HCV complete genomes belonging to subtypes 6a (D9), 6e (D42 and D88), 6f (TH52), 6i (TH24), 6l (D33), 6n (TH22 and TH31) and 6o (D85). Phylogenetic analysis of the core/E1 and NS5B regions from unclassified genotype 6 isolates from Asian immigrants in Canada revealed that two other viruses (D49 and D83) could be classified as novel candidates of HCV subtypes 6t and 6u.

Characterization of hepatitis C virus deletion mutants circulating in chronically infected patients.

Hepatitis C virus (HCV) has a linear positive-stranded RNA genome of approximately 9,600 nucleotides in length and displays a high level of sequence diversity caused by high mutation rates and recombination. However, when we performed long distance reverse transcription-PCRs on HCV RNA isolated from serum of chronic HCV patients, not only full-length HCV genomes but also HCV RNAs which varied in size from 7,600 to 8,346 nucleotides and contained large in-frame deletions between E1 and NS2 were amplified. Carefully designed control experiments indicated that these deletion mutants are a bona fide natural RNA species, most likely packaged in virions. Moreover, deletion mutants were detected in sera of patients infected with different HCV genotypes. We observed that 7/37 (18.9%) of genotype 1, 5/43 (11.6%) of genotype 3, and 4/13 (30.7%) of genotype 6 samples contained HCV deletion mutant genomes. These observations further exemplify HCV's huge genetic diversity and warrant studies to explore their biological relevance.

Identification of a naturally occurring recombinant genotype 2/6 hepatitis C virus.

Hepatitis C viruses (HCVs) display a high level of sequence diversity and are currently classified into six genotypes and an increasing number of subtypes. Most likely, this heterogeneity is caused by genetic drift; evidence for recombination is scarce. To study the molecular heterogeneity of HCV in Vietnam, we analyzed 58 HCV RNA-positive sera from Vietnamese blood donors by sequence analysis of the CORE and NS5B regions. Phylogenetic analyses revealed the presence of genotype 1 (38%), genotype 2 (10.3%), and genotype 6 viruses (51.7%). All samples showed concordant results except for two (D3 and D54). Sample D54 was a mixed infection of genotype 2i and 6h viruses. Whole-genome analysis and bootscan analysis of sample D3, on the other hand, revealed a recombinant virus with genotype 2i and genotype 6p sequences at the 5' and 3' ends, respectively. The crossover point was located between nucleotide positions 3405 to 3464 (numbering according to prototype strain HCV-H, M67463) at the NS2/NS3 junction. The identification of this naturally occurring recombinant virus strengthens the concept that recombination may play a role in HCV epidemiology and evolution. Furthermore, the location of the recombination breakpoint may be relevant for constructing infectious chimeric viruses.