Pre-existing cross-reactive antibodies to avian influenza H5N1 and 2009 pandemic H1N1 in US military personnel.

We studied cross-reactive antibodies against avian influenza H5N1 and 2009 pandemic (p) H1N1 in 200 serum samples from US military personnel collected before the H1N1 pandemic. Assays used to measure antibodies against viral proteins involved in protection included a hemagglutination inhibition (HI) assay and a neuraminidase inhibition (NI) assay. Viral neutralization by antibodies against avian influenza H5N1 and 2009 pH1N1 was assessed by influenza (H5) pseudotyped lentiviral particle-based and H1N1 microneutralization assays. Some US military personnel had cross-neutralizing antibodies against H5N1 (14%) and 2009 pH1N1 (16.5%). The odds of having cross-neutralizing antibodies against 2009 pH1N1 were 4.4 times higher in subjects receiving more than five inactivated whole influenza virus vaccinations than those subjects with no record of vaccination. Although unclear if the result of prior vaccination or disease exposure, these pre-existing antibodies may prevent or reduce disease severity.

Evaluation of loop-mediated isothermal amplification (LAMP) for malaria diagnosis in a field setting.

We used the loop-mediated isothermal amplification (LAMP) method developed by our group for malaria diagnosis with genus-specific and species-specific primers for the four human malaria parasites at a field clinic in comparison with standard microscopy. Among 110 blood samples collected from the malaria clinic in Thailand, LAMP detected 59 of 60 samples positive by microscopy (sensitivity = 98.3%) and none of the 50 microscopy-negative samples (specificity = 100%). Negative predictive value (NPV) and positive predictive value (PPV) of LAMP were 98% and 100%, respectively. These results indicate that LAMP is an effective tool for malaria diagnosis at a field clinic in a field setting.

Correlation of protection against Japanese encephalitis virus and JE vaccine (IXIARO(®)) induced neutralizing antibody titers.

Immune sera from volunteers vaccinated in a blinded Phase 3 clinical trial with JE-VAX(®) and a new Japanese encephalitis virus (JEV) vaccine (IC51 or IXIARO), were tested for the ability to protect mice against lethal JEV challenge. Sera from IXIARO vaccinated subjects were pooled into four batches based on neutralizing antibody measured by plaque reduction neutralization test (PRNT(50) titer): high (∼200), medium (∼40-50), low (∼20) and negative (<10). Pooled sera from JE-VAX(®) vaccinated subjects (PRNT(50) titer∼55) and pooled JEV antibody negative pre-vaccination sera were used as controls. Groups of ten 6- to 7-week-old female ICR mice were injected intraperitoneally with 0.5 ml of each serum pool diluted 1:2 or 1:10, challenged approximately 18 h later with a lethal dose of either JEV strain SA14 (genotype III) or strain KE-093 (genotype I) and observed for 21 days. All mice in the non-immune serum groups developed clinical signs consistent with JEV infection or died, whereas high titer sera from both IXIARO and JE-VAX(®) sera protected 90-100% of the animals. Statistical tests showed similar protection against both JEV strains SA14 and KE-093 and protection correlated with the anti-JEV antibody titer of IXIARO sera as measured by PRNT(50). Ex vivo neutralizing antibody titers showed that almost all mice with a titer of 10 or greater were fully protected. In a separate study, analysis of geometric mean titers (GMTs) of the groups of mice vaccinated with different doses of IXIARO and challenged with JEV SA14 provided additional evidence that titers≥10 were protective.

HIV type 1 molecular epidemiology among high-risk clients attending the Thai Red Cross Anonymous Clinic in Bangkok, Thailand.

Several studies have reported an increasing number of non-CRF01_AE infections in high-risk groups in Thailand suggesting a more complex HIV-1 epidemic. This study assessed the complexity of the HIV epidemic among high-risk clients tested for HIV-1 at the Thai Red Cross Anonymous Clinic (TRCAC) between July 1, 2006 and February 28, 2007. HIV-1 genotypes were determined from plasma of infected subjects (n = 401) by the multiregion hybridization assay (MHAbce, v.2). Univariate and multivariate logistic regression analyses were used to determine risk factors associated with HIV prevalence and non-CRF01_AE infection. The estimated overall HIV prevalence was 14.1%: 25.3% among men who have sex with men (MSM), 18.4% among heterosexual women, and 9.6% among heterosexual men. Among the risk factors found to be associated with HIV prevalence were age (25-29 years), risk behavior (MSM), marital status (not single), education (less than high school), and inconsistent condom use. Overall, non-CRF01_AE strains accounted for 18.9% of the infections: 25.3% among MSM and 14.8% and 20.4% among heterosexual women and men, respectively. Our results indicate a concentrated and genetically complex HIV epidemic among Thai MSM. These findings advocate for targeted intervention and prevention measures among high-risk populations in Thailand.

Laboratory and semi-field evaluation of Mosquito Dunks against Aedes aegypti and Aedes albopictus larvae (Diptera: Culicidae).

Laboratory bioassays and semi-field studies were conducted on the efficacy and longevity of Mosquito Dunks (7,000 ITU/mg Bti) in order to determine the concentration-response relationship and the effectiveness on the potency of the Bti product against Aedes mosquito species based on the WHO protocol standard methods and to determine the longevity of release for this product against Ae. aegypti mosquito larvae in water storage containers. This bio-potency study with the late 3rd instar larvae of Ae. aegypti and Ae. albopictus was carried out according to WHO standard protocols. The six concentrations of the Bti product used in each test were replicated 4 times with 25 mosquito larvae. Probit analysis was then used to determine the LC50 and LC95 which was equated with dosages of 1.02 and 1.86 ppm for Ae. aegypti; and 0.39 and 0.84 ppm for Ae. albopictus, which reveals a potency of 382.95 and 303.74 ITU/mg, respectively. The semi-field evaluation of this product in 200-liter earthen jars against 3rd instar larvae of Ae. aegypti showed satisfactory control of greater than 80% at 11 weeks post-treatment.

Soil analysis around anopheline breeding habitats in north-western Thailand.

The epidemiology of malaria is largely dependent on its vector habitat. Each species of Anopheles larvae has a specific habitat requirement for its development. Anopheline mosquitoes are common throughout Thailand and utilize a wide variety of habitats. The dominant malaria vectors in Thailand are An. dirus, An. maculatus, and An. minimus. The relationship between soil chemical components and the particular species of anopheline in their specific aquatic habitats was studied from September 2002 to July 2003 at Ban Khun Huay, Ban Pa Dae, and Ban Tham Seau in the Mae Sot district, Tak Province, Thailand. Mapping of each habitat was performed using a Global Positioning System unit. A total count of 2,130 laboratory reared adult Anopheles were collected from 138 habitats categorized into 11 different types identified into 18 species from larval sampling in three villages. An. dirus, An. maculatus, and An. minimus were found 5.26%, 10.70%, and 55.31%, respectively, along with other minor species. Drainage and/or season seemed to be associated with the presence of An. dirus, An. maculatus, An. minimus, An. jamesii, An. sawadwongporni, and An. peditaeniatus. Chemical tests: pH, aluminum, magnesium, calcium, and ferric iron showed some associations with the presence of Anopheles. Only drainage was found to be a parameter associated with the presence of An. minimus.

Molecular characterization of hereditary persistence of fetal hemoglobin in the Karen people of Thailand.

Hereditary persistence of fetal hemoglobin (HPFH) is the condition whereby a continuously active gamma-globin gene expression leads to elevated fetal hemoglobin (Hb F) levels in adult life [Stamatoyannopoulos G, Grosveld F. Hemoglobin switching. In: Stamatoyannopoulos G, Majerus PW, Perlmutter RM, Varmus H, eds. The Molecular Basis of Blood Diseases. Philadelphia: W.B. Saunders, 2001:135-182; Wood WG. Hereditary persistence of fetal hemoglobin and delta(beta) thalassemia. In: Steinberg MH, Forget BG, Higgs DR, Nagel RL, eds. Disorders of Hemoglobin: Genetics, Pathophysiology, and Clinical Management. Cambridge: Cambridge University Press, 2001:356-388; and Weatherall DJ, Clegg JB. Hereditary persistence of fetal hemoglobin. In: Weatherall DJ, Clegg JB, eds. The Thalassaemia Syndromes. Oxford: Blackwell Scientific Publishers, 1981:450-507]. The condition is caused either by mutation of the beta- and gamma-globin genes, or the gamma-gene controlled region on other chromosomes. Several families with this condition have been reported from Vietnam, Cambodia and China, and the Southeast Asian mutation (or HPFH-6), a 27 kb deletion, was demonstrated. Here we report on a mother and her daughter of the Karen ethnic group with high levels of Hb F, living in the Suan Pueng District on the border of Thailand and Myanmar. Genotyping showed a heterozygosity for the 27 kb deletion of the beta-globin gene. Their conditions have been confirmed by gap polymerase chain reaction (PCR) with three oligonucleotide primers recently developed by Xu et al. [Xu X-M, Li Z-Q, Liu Z-Y, Zhong X-L, Zhao Y-Z, Mo Q-H. Molecular characterization and PCR detection of a deletional HPFH: application to rapid prenatal diagnosis for compound heterozygotes of this defect with beta-thalassemia in a Chinese family. Am J Hematol 2000; 65:183-188.], and a DNA sequencing method. Thus far there has been no official report of the HPFH-6 anomaly from Thailand. The compound heterozygosity of beta-thalassemia (thal) and hereditary persistence of Hb F causes the phenotype of thalassemia intermedia; in contrast, homozygotes for this anomaly show only mild microcytic anemia. Hence, genetic counseling for hereditary persistence of Hb F carriers is needed for family planning.

Cost-effectiveness in establishing hemophilia carrier detection and prenatal diagnosis services in a developing country with limited health resources.

The cost-effectiveness of carrier detection and prenatal diagnosis for hemophilia at the International Hemophilia Training Center, Bangkok, Thailand was studied. From 1991 to 2002, 209 females from 124 families with hemophilia A and B were included. There were 180 hemophilia A carriers and 29 hemophilia B carriers which could be classified into 78 obligate and 131 possible carriers. The phenotypic analysis for possible carriers involved the determination of levels of factor VIII or IX clotting activity (FVIII:C, FIX:C) and the ratio of FVIII:C and von Willebrand factor antigen. The result revealed that 49 females (37.4%) were diagnosed as carriers, 65 females (49.6%) were normal and 17 females (13%) were undetermined. Additional genotypic analysis was provided to 46 families with 74 females with obligate, proven or undetermined carriers within the reproductive life. The polymorphisms associated with factor VIII and IX genes were used including Bcl I for the factor VIII gene and combined use of Mse I, Sal I, Nru I, Hha I and Dde I for the factor IX gene. The informative rate was 59.4% (44/74). Consequently, 12 prenatal diagnoses for fetus at risk were performed. Sex determination was initially determined and followed by the diagnosis of hemophilia through informative gene tracking and/or the measurement of fetal levels of FVIII:C or FIX:C. The result revealed that 3 male fetuses were affected. The total cost of carrier detection and prenatal diagnosis that the families had to pay in the government hospital was 238,600 Baht (US dollars 5,965). It was compared to the estimated cost of minimal replacement therapy using lyophilized cryoprecipitate for the survival time of 30 years in one patient with hemophilia of 1,012,500 Baht (US dollars 25,312.5). The cost of prevention was much less than the replacement therapy. In conclusion, it is cost-effective to establish the service for carrier detection and prenatal diagnosis for hemophilia especially in developing countries with limited health resources.

Nutritional support in pediatric patients undergoing bone marrow transplantation.

BACKGROUND: Children undergoing bone marrow transplantation (BMT) are prone to develop severe gastrointestinal (GI) complications and metabolic imbalance which consequently impair their nutritional status. Nutritional support is an important adjunctive treatment during BMT. OBJECTIVE: To assess GI complications, metabolic complications and nutritional outcome of children undergoing BMT with nutritional support intervention. METHOD: Retrospective study of 20 children (median age 6.8 years, 11 males) undergoing BMT at Ramathibodi Hospital from March 1995 to July 2000 was conducted. Their medical records were reviewed. RESULTS: The patients underwent autologous (n = 9) and allogenic BMT (n = 11). Median z-scores of weight for age, height for age and weight for height were 0.06 +/- 1.93, -0.55 +/- 1.18 and 0.48 +/- 1.94, respectively. Nineteen patients had vomiting for 9.8 +/- 5.5 days. Eighteen patients developed diarrhea for 9.6 +/- 7.2 days. The durations of vomiting and diarrhea, as a percentage of total hospital days, were 33.5 +/- 16.3 per cent and 30.4 +/- 17.0 per cent, respectively. There were no differences between the patients with autologous and allogenic BMT regarding these durations. All patients needed enteral and/or parenteral nutrition support for 21.0 +/- 7.7 days except for one patient who could take adequate oral intake. The duration of enteral nutrition support was not significantly different between the groups but the duration of parenteral nutrition support was significantly longer in the allogenic group. Metabolic complications were hypokalemia, hypophosphatemia and one case of arrhythmia secondary to hypomagnesemia. All patients developed febrile neutropenia but none developed catheter-related sepsis. The length of hospital stay was 30.5 +/- 10.2 days. The median z-score of weight for height on the day of discharge was 1.08 +/- 2.03. CONCLUSION: Children undergoing BMT usually have GI symptoms of vomiting, diarrhea and mucositis as well as metabolic imbalances such as hypokalemia, hypophosphatemia and hypomagnesemia. Despite these complications, their nutritional status could be restored by proper nutritional support.