Veerachai Watanaveeradej

A new circulating recombinant form, CRF15_01B, reinforces the linkage between IDU and heterosexual epidemics in Thailand.

HIV-1 subtype B and CRF01_AE have been in circulation in Thailand and Southeast Asia for more than a decade. Initially separated by risk group, the two strains are increasingly intermixed, and two recombinant strains of essentially reciprocal structure have been recently reported. Here we identify additional CRF_01B recombinants and provide the evidence that HIV-1 strains now pass freely between the two high-risk populations. HIV isolates that showed discordance between CRF01_AE and subtype B in multi-region genotyping assays were selected for the study. They were drawn from 3 different cohorts in Thailand representing different risk behaviors and demographic characteristics: a drug user cohort in the north, a family planning clinic attendee cohort in the southeast, and a cohort study of the mucosal virology and immunology of HIV-1 infection in Thailand. The DNA from these isolates was PCR amplified to recover the full HIV-1 genome and subjected to sequencing and phylogenetic analysis. We establish that one particular CRF_01B recombinant, with the external envelope of subtype B and the rest of the genome from CRF01_AE, is circulating widely in Thailand. Termed CRF15_01B (also referred to as CRF15), the strain was primarily heterosexually transmitted, although injecting drug use (IDU) also played a role. In aggregate data from the studies, CRF15 constituted 1.7% of all HIV-1 infections (95% confidence interval 0.5-4.4%) and was dispersed widely in the country. The previously separate heterosexual and IDU epidemics have apparently been bridged by a new CRF. The entry of CRF15 into the mainstream of the epidemic signals new complexity in the long stable molecular picture in Thailand. These recombinants must be considered in ongoing or projected efficacy evaluations of HIV-1 vaccines and antiviral therapies.

Safety and Immunogenicity of a Tetravalent Live-attenuated Dengue Vaccine in Flavivirus Naive Children

A Phase I/II observer-blind, randomized, controlled trial evaluated the safety and immunogenicity of a dengue virus (DENV) vaccine candidate in healthy Thai infants (aged 12-15 months) without measurable pre-vaccination neutralizing antibodies to DENV and Japanese encephalitis virus. Fifty-one subjects received two doses of either DENV (N = 34; four received 1/10th dose) or control vaccine (N = 17; dose 1, live varicella; dose 2, Haemophilus influenzae type b). After each vaccine dose, adverse events (AEs) were solicited for 21 days, and non-serious AEs were solicited for 30 days; serious AEs (SAEs) were recorded throughout the study. Laboratory safety assessments were performed at 10 and 30 days; neutralizing antibodies were measured at 30 days. The DENV vaccine was well-tolerated without any related SAEs. After the second dose, 85.7% of full-dose DENV vaccinees developed at least trivalent and 53.6% developed tetravalent neutralizing antibodies ≥ 1:10 to DENV (control group = 0%). This vaccine candidate, therefore, warrants continued development in this age group (NCT00322049; clinicaltrials.gov).

Japanese encephalitis virus remains an important cause of encephalitis in Thailand

BACKGROUND Japanese encephalitis virus (JEV) is endemic in Thailand and prevention strategies include vaccination, vector control, and health education. METHODS Between July 2003 and August 2005, we conducted hospital-based surveillance for encephalitis at seven hospitals in Bangkok and Hat Yai. Serum and cerebrospinal (CSF) specimens were tested for evidence of recent JEV infection by immunoglobulin M (IgM) enzyme-linked immunosorbent assay (ELISA) and a plaque reduction neutralization test (PRNT). RESULTS Of the 147 patients enrolled and tested, 24 (16%) had evidence of acute flavivirus infection: 22 (15%) with JEV and two (1%) with dengue virus. Of the 22 Japanese encephalitis (JE) cases, 10 (46%) were aged ≤ 15 years. The median length of hospital stay was 13 days; one 13-year-old child died. Ten percent of encephalitis patients enrolled in Bangkok hospitals were found to have JEV infection compared to 28% of patients enrolled in hospitals in southern Thailand (p < 0.01). Four (40%) of the 10 children with JE were reported as being vaccinated. CONCLUSIONS JEV remains an important cause of encephalitis among hospitalized patients in Thailand. The high proportion of JE among encephalitis cases is concerning and additional public health prevention efforts or expanded vaccination may be needed.

Single-dose, live-attenuated Japanese encephalitis vaccine in children aged 12-18 months: randomized, controlled phase 3 immunogenicity and safety trial.

This trial in 1200 JE-vaccination naïve children (age 12–18 mo) in Thailand and the Philippines aimed to demonstrate consistency of three successive industrial scale manufacturing lots of live attenuated Japanese encephalitis chimeric virus vaccine (JE-CV) and consistency between industrial scale manufacturing lots and a fourth, development lot. Children received JE-CV from one of three successive industrial scale lots produced in Thailand (n = 899), or from a fourth development lot produced in the USA (n = 199), or hepatitis A control vaccine (n = 102). Antibodies were assessed by 50% plaque reduction neutralization test (PRNT50) at screening and Day 28. Seroconversion rates (titer of < 10 at baseline and ≥ 10 on Day 28, or a four-fold rise from a baseline titer of ≥ 10) were determined per group. Lot-to-lot consistency of seroconversion rate and GMT was demonstrated between the 3 industrial scale lots, and between these lots and the US lot. Seroconversion rate on pooled data 28 d after JE-CV vaccination (Thai lots) was 95.0% [95% confidence interval (CI); 93.3–96.3]. The safety profile of JE-CV was favorable and comparable with hepatitis A vaccine. There were no serious adverse events related to vaccination. This study demonstrated the consistency of three successive industrial scale JE-CV vaccine lots, as well as consistency with a development lot. The study also demonstrated that a single dose of JE-CV is well tolerated and elicits a high protective immune response, seroconverting 95% of JE-naïve Asian children aged 12–18 mo. ClinicalTrials.gov: NCT00735644

Safety and Immunogenicity of a Rederived, Live-Attenuated Dengue Virus Vaccine in Healthy Adults Living in Thailand: A Randomized Trial

Safety and immunogenicity of two formulations of a live-attenuated tetravalent dengue virus (TDEN) vaccine produced using rederived master seeds from a precursor vaccine were tested against a placebo control in a phase II, randomized, double blind trial (NCT00370682). Two doses were administered 6 months apart to 120 healthy, predominantly flavivirus-primed adults (87.5% and 97.5% in the two vaccine groups and 92.5% in the placebo group). Symptoms and signs reported after vaccination were mild to moderate and transient. There were no vaccine-related serious adverse events or dengue cases reported. Asymptomatic, low-level viremia (dengue virus type 2 [DENV-2], DENV-3, or DENV-4) was detected in 5 of 80 vaccine recipients. One placebo recipient developed a subclinical natural DENV-1 infection. All flavivirus-unprimed subjects and at least 97.1% of flavivirus-primed subjects were seropositive to antibodies against all four DENV types 1 and 3 months post-TDEN dose 2. The TDEN vaccine was immunogenic with an acceptable safety profile in flavivirus-primed adults.

Infectious Causes of Encephalitis and Meningoencephalitis in Thailand, 2003-2005

Although many causes were identified, most remain unknown.

Metagenomics Study of Viral Pathogens in Undiagnosed Respiratory Specimens and Identification of Human Enteroviruses at a Thailand Hospital

Numerous pathogens cause respiratory infections with similar symptoms. Routine diagnostics detect only a limited number of pathogens, leaving a gap in respiratory illness etiology surveillance. This study evaluated next-generation sequencing for unbiased pathogen identification. Respiratory samples collected in Thailand, Philippines, Bhutan, and Nepal, that were negative by several molecular and immunofluorescence assays, underwent viral cultivation. Samples which demonstrated cytopathic effect in culture (N = 121) were extracted and tested by Luminex xTAG respiratory viral panel (RVP) assay and deep sequencing by Roche 454 FLX Titanium system. Using RVP assay, 52 (43%) samples were positive for enterovirus or rhinovirus and another three were positive for respiratory syncytial virus B, parainfluenza 4, and adenovirus. Deep sequencing confirmed the Luminex assay results and identified additional viral pathogens. Human enteroviruses, including Enterovirus A type 71 and 12 types of Enterovirus B (EV-B) were identified from a hospital in Bangkok. Phylogenetic and recombination analysis showed high correlation of VP1 gene-based phylogeny with genome-wide phylogeny and the frequent genetic exchange among EV-B viruses. The high number and diversity of enteroviruses in the hospital in Bangkok suggests prevalent existence. The metagenomic approach used in our study enabled comprehensive diagnoses of respiratory viruses.

Immunogenicity of a live-attenuated human rotavirus RIX4414 vaccine with or without buffering agent

Aim: The lyophilized form of the human rotavirus RIX4414 vaccine (Rotarix™) is usually reconstituted with a liquid calcium carbonate (CaCO3) buffer and administered orally. However, errors in the reconstitution could occur (e.g. reconstituted with water instead of CaCO3 buffer) or the buffer might be temporarily unavailable in few instances. This study was conducted to evaluate the immunogenicity of the RIX4414 vaccine if the vaccine was reconstituted with other agents (e.g. water) instead of CaCO3 buffer. Methods: Healthy infants aged 6–12 weeks, received two oral doses of the RIX4414 vaccine/placebo, reconstituted either with injectable water or CaCO3 buffer according to a 0, 2 month schedule. Seroconversion rates in terms of anti-rotavirus Immunoglobulin A (anti-RV IgA) antibody levels (cut off: ≥ 20U/ml by ELISA) and vaccine take were calculated 2 months post-Dose 2. Solicited and unsolicited symptoms reported during the 15- and 31-day follow-up period after each dose and serious adverse events (SAEs) reported during the entire study period were recorded. Results: There was no statistical difference detected between RIX4414 vaccine reconstituted with buffer or water in vaccine take or in seroconversion rate. The anti-RV IgA seroconversion rate 2 months post-Dose 2 was 84.7% (95% CI: 78.1–90.0) for the group with buffer and 78.6% (95% CI: 71.2–84.8) for the group with water. Solicited and unsolicited symptoms reported were similar across groups. No vaccine related SAEs were reported. Conclusion: Administration of RIX4414 vaccine in the absence of CaCO3 buffer was shown to be well tolerated and immunogenic in Thai infants.

Antibody response to hepatitis B vaccine in infants of HIV-positive mothers.

Hepatitis B remains an important health problem with more than 1500 cases of acute infection occurring each year. HIV-infected adults may be at increased risk of exposure to hepatitis B infection and both diseases can be transferred from mothers to infants. HIV-infected children have inadequate responses to T-cell-dependent vaccines. Suboptimal responses to recombinant DNA hepatitis B virus vaccination were reported in persons with impaired humoral or cellular immunity. We compared hepatitis B vaccine seroresponse rates between HIV-infected and uninfected infants born to HIV positive mothers and further evaluated whether boosters with double dosage could improve immunogenicity in the nonresponders. Full-term infants born to HIV-positive mothers who had no hepatitis B surface antigen (HBsAg) were enrolled. All subjects were born at Phramongkutklao Hospital from January 1996 to March 1998. Written informed consent was obtained and these infants had scheduled clinic visits at 12469 and 12 months of age for routine health care. (excerpt)

Long-Term Immunogenicity of a Single Dose of Japanese Encephalitis Chimeric Virus Vaccine in Toddlers and Booster Response 5 Years After Primary Immunization.

Background: Japanese encephalitis (JE) is an important mosquito-borne viral disease that is endemic in Asia, Western Pacific countries and Northern Australia. Although there is no antiviral treatment, vaccination is effective in preventing this disease. Methods: We followed a cohort of 596 children for 5 years after primary vaccination at 12–18 months of age with JE chimeric virus vaccine (JE-CV; IMOJEV) in a multicenter, phase III trial in Thailand and the Philippines to assess antibody persistence and safety. At the end of the 5 years, a subgroup of 85 participants, at 1 site in Thailand, was followed after administration of a JE-CV booster vaccination. JE antibody titers were measured annually after primary vaccination and 28 days after booster vaccination using a 50% plaque reduction neutralization test. Seroprotection was defined as a JE-CV neutralizing antibody titer ≥10 (1/dil). Kaplan–Meier survival analysis was used to estimate the proportion of participants maintaining protective JE-CV neutralizing antibody titers. Results: At 1, 2, 3, 4 and 5 years after vaccination with JE-CV, 88.5%, 82.9%, 78.2%, 74.0% and 68.6% of the participants followed remained seroprotected. Geometric mean titers in the subgroup assessed after receipt of a booster dose increased from 61.2 (95% confidence interval: 43.8–85.7) pre-booster to 4951 (95% confidence interval: 3928–6241) 28 days post-booster, with all participants seroprotected. There were no safety concerns identified. Conclusions: Protective immune responses persisted for at least 5 years after a JE-CV primary immunization in the majority of participants. JE-CV booster induced a robust immune response even after a 5-year interval.