High prevalence and low diversity of chigger infestation in small mammals found in Bangkok Metropolitan parks.

Chiggers are recognized as vectors of scrub typhus disease caused by the bacteria, Orientia tsutsugamushi. The risk of disease exposure is mainly related to chigger bites when humans or animals roam into vector-infested habitats. In big cities, urban public parks could provide areas for the animal-human interface and zoonotic pathogen transmission. The ecology and epidemiology of urban scrub typhus are still poorly understood in Thailand. Small mammals were trapped and examined for chigger infestation in urban public parks across metropolitan Bangkok, Thailand. We found a high prevalence of infestation (76.8%) with surprisingly low diversity. Two chigger species, Leptotrombidium deliense and Ascoschoengastia indica, were identified using morphological characteristics and molecular confirmation. The generalized linear model identified host intrinsic variables (i.e. body mass index) with host density, habitat composition and open field as the extrinsic factors explaining the abundance of chigger infestation. The bacteria O. tsutsugamushi was not detected in chiggers (90 chigger-pooled samples) and animal host tissues (164 spleen samples). However, the existence of chigger vectors calls for the Bangkok Metropolitan Administration and public health authorities to develop a comprehensive scrub typhus monitoring and prevention strategy in the parks and nearby communities.

A rapid polymerase chain reaction based method for identification of the Anopheles dirus sibling species.

A simple polymerase chain reaction (PCR) based method was developed to differentiate the Anopheles dirus, species A, B, C and D in Thailand using specific primers designed from species specific sequences. The PCR protocol was optimized to obtain products of 120 bp, 75 bp, 60 bp and 172 bp for species A, B, C and D, respectively. This method used a cocktail of four primer sets to identify these An. dirus sibling species. The method is very sensitive as only a small portion of mosquito was required allowing the rest of the mosquito to be used for other analyses. Specimens also kept for up to 14 years could be analyzed unambiguously from either larvae or adult. This method is advantageous over other PCR-based methods for identification of malaria vectors because it does not require any specific DNA extraction. A mosquito specimen was homogenized in 1x PCR buffer, then the supernatant directly used for PCR identification, allowing a large number of samples to be processed at the same time. It provides a simple and rapid practical method for screening An. dirus species, which is essential in malaria vector epidemiological studies in Southeast Asia.

Stable marker on flagellin gene sequences related to arabinose non-assimilating pathogenic Burkholderia pseudomallei.

Using PCR-based isolation and sequence analysis of the flagellin gene from two distinct biotypes of Burkholderia pseudomallei, a 15-bp deletion was found within the variable domain of the gene in isolates capable of assimilating arabinose (Ara+). This finding led to the development of a PCR-based method in order to differentiate and identify pathogenic B. pseudomallei for epidemiological study. A pair of specific primers was designed covering the 15-bp deletion region at the variable domain. PCR-amplification products of 176 and 191 bp in size were detected from 41 Ara+ isolates and 39 Ara - isolates of B. pseudomallei, respectively. Moreover, flagellin gene fragments of other bacterial species tested in this study were not amplified using these primers. The results suggest that the flagellin gene sequences of both B. pseudomallei biotypes in this region are stable and distinct. This method can be applied and useful for the epidemiological study of B. pseudomallei.