Molecular genetics analysis for co-infection of Brugia malayi and Brugia pahangi in cat reservoirs based on internal transcribed spacer region 1.

This study described the diagnosis of a mixed infection of Brugia malayi and Brugia pahangi in a single domestic cat using the internal transcribed spacer 1 (ITS1) region. Following polymerase chain reaction amplification of the ITS1 region, the 580 bp amplicon was cloned, and 29 white colonies were randomly selected for DNA sequencing and phylogenetic tree construction. A DNA parsimony tree generated two groups of Brugia spp with one group containing 6 clones corresponding to B. pahangi and the other 23 clones corresponding to B. malayi. This indicated that mixed infection of the two Brugia spp, B. pahangi and B. malayi, had occurred in a single host.

The efficacy of a single-oral-dose administration of ivermectin and diethylcarbamazine on the treatment of feline Brugia malayi.

The combination of ivermectin and diethylcarbamazine (DEC) have been shown to be superior to either drug alone for the suppression of Brugia malayi in humans, but their efficacy against infection with B. malayi in cats has never been investigated. Fourteen asymptomatic microfilaremic (1-200 microfilariae/20 microl blood) cats received oral doses of ivermectin (400 microg/kg body weight) and DEC (6 mg/kg body weight) as a single treatment. A two-month post-treatment examination revealed that 87-100% of the microfilariae in each subject had been cleared, with two of the subjects being amicrofilaremic. A further reduction in microfilarial levels was observed until the final follow-up, at 8 months post-treatment, when the mean clearance rate was 99% and 12 out of the 14 subjects (86%) were amicrofilaremic. The combination of ivermectin and DEC demonstrated a microfilaricidal effect superior to that of either drug used alone, both in the initial rapid clearance of microfilariae, and in sustaining the effect for 8 months. This finding has important implications for the control of brugian lymphatic filariasis in the cat reservoir.

Determination of dengue virus serotypes in Thailand using PCR based method.

A reverse transcriptase-polymerase chain reaction (RT-PCR) and a single-tube multiplex PCR assay was modified for typing of dengue virus in different geographical areas of Thailand during 2000-2001. A set of primers (D1 and D2) was used to generate the RT-PCR product of 511 bp in size which subsequently underwent a single-tube multiplex PCR amplification using the highly specific primers for each of the dengue virus serotypes (D1, TS1, TS2, TS3 and DEN4). The PCR products of 482, 119, 290 and 392 bp in size were generated for dengue virus serotypes 1, 2, 3, and 4, respectively. Each set of specific primers showed no amplification of non-specific and non-target PCR products from human genomic DNA. The method was applied for investigation of 637 human blood samples in Thailand during 2000-2001 and found that 71, 43, 28, and 43 patients were classified as having a single infection with serotypes 1, 2, 3, and 4, respectively. Multiple infections with two or more dengue virus serotypes were also detected.

A polymerase chain reaction assay for the survey of bancroftian filariasis.

A polymerase chain reaction (PCR) assay based on a highly repeated DNA sequence found in Wuchereria bancrofti (SspI repeat) has been modified for the survey of bancroftian filariasis in expatriate workers (Myamese, Karen and Mon) from Myanmar where human filariasis is endemic. The PCR was very sensitive with the ability to detect the presence of as little as 10 pg of parasite DNA. The primers used in this PCR also showed highly specific amplification of parasite DNA without the presence of non-specific and non-target PCR products such as Brugia malayi, Plasmodium falciparum and human DNA. The primers were used to investigate filariasis in four provinces in the central and western Thailand, Samut Songkram, Ratchaburi, Nakhon Pathom and Tak during 1997-2001. Among them, Tak and Ratchaburi are the only endemic areas of bancroftian filariasis. In this field study, 1,299 human blood samples (501 from Samut Songkram, 510 from Ratchaburi, 109 from Nakhon Pathom, and 179 from Tak) were collected and screened by PCR. The result showed that 1, 2, 3, and 33 patients from Samut Songkram, Ratchaburi, Nakhon Pathom, and Tak respectively were infected with W. bancrofti. These numbers were corresponded to the prevalence rate of infection of 0.2, 0.4, 2.8, and 18.5%, respectively. The PCR was able to detect the third-stage infectious larvae (L3) from Culex quinquefasciatus, mosquito vector of the W. bancrofti, that was experimentally fed to infected patient. The PCR screening of each of field mosquito pools from two endemic areas, Ratchaburi and Tak, showed that no L3 of W. bancrofti was detected.