Background: The genus Onchocerca Diesing, 1841 includes species of medical importance, such as O. volvulus (Leuckart, 1893), which causes river blindness in the tropics. Recently, zoonotic onchocercosis has been reported in humans worldwide. In Japan, O. dewittei japonica Uni, Bain & Takaoka, 2001 from wild boars is a causative agent for this zoonosis. Many filarioid nematodes are infected with Wolbachia endosymbionts which exhibit various evolutionary relationships with their hosts. While investigating the filarial fauna of Borneo, we discovered an undescribed Onchocerca species in the bearded pig Sus barbatus Müller (Cetartiodactyla: Suidae). Methods: We isolated Onchocerca specimens from bearded pigs and examined their morphology. For comparative material, we collected fresh specimens of O. d. dewittei Bain, Ramachandran, Petter & Mak, 1977 from banded pigs (S. scrofa vittatus Boie) in Peninsular Malaysia. Partial sequences of three different genes (two mitochondrial genes, cox1 and 12S rRNA, and one nuclear ITS region) of these filarioids were analysed. By multi-locus sequence analyses based on six genes (16S rDNA, ftsZ, dnaA, coxA, fbpA and gatB) of Wolbachia, we determined the supergroups in the specimens from bearded pigs and those of O. d. dewittei. Results: Onchocerca borneensis Uni, Mat Udin & Takaoka n. sp. is described on the basis of morphological characteristics and its genetic divergence from congeners. Molecular characteristics of the new species revealed its close evolutionary relationship with O. d. dewittei. Calculated p-distance for the cox1 gene sequences between O. borneensis n. sp. and O. d. dewittei was 5.9%, while that between O. d. dewittei and O. d. japonica was 7.6%. No intraspecific genetic variation was found for the new species. Wolbachia strains identified in the new species and O. d. dewittei belonged to supergroup C and are closely related. Conclusions: Our molecular analyses of filarioids from Asian suids indicate that the new species is sister to O. d. dewittei. On the basis of its morphological and molecular characteristics, we propose to elevate O. d. japonica to species level as O. japonica Uni, Bain & Takaoka, 2001. Coevolutionary relationships exist between the Wolbachia strains and their filarial hosts in Borneo and Peninsular Malaysia.

Taxonomists frequently identify specimen from various populations based on the morphological characteristics and molecular data. This study looks into another invasive process in identification of house shrew (Suncus murinus) using image analysis and machine learning approaches. Thus, an automated identification system is developed to assist and simplify this task. In this study, seven descriptors namely area, convex area, major axis length, minor axis length, perimeter, equivalent diameter and extent which are based on the shape are used as features to represent digital image of skull that consists of dorsal, lateral and jaw views for each specimen. An Artificial Neural Network (ANN) is used as classifier to classify the skulls of S. murinus based on region (northern and southern populations of Peninsular Malaysia) and sex (adult male and female). Thus, specimen classification using Training data set and identification using Testing data set were performed through two stages of ANNs. At present, the classifier used has achieved an accuracy of 100% based on skulls’ views. Classification and identification to regions and sexes have also attained 72.5%, 87.5% and 80.0% of accuracy for dorsal, lateral, and jaw views, respectively. This results show that the shape characteristic features used are substantial because they can differentiate the specimens based on regions and sexes up to the accuracy of 80% and above. Finally, an application was developed and can be used for the scientific community. This automated system demonstrates the practicability of using computer-assisted systems in providing interesting alternative approach for quick and easy identification of unknown species.

The filarial nematodes Wuchereria bancrofti (Cobbold, 1877), Brugia malayi (Brug, 1927) and B. timori Partono, Purnomo, Dennis, Atmosoedjono, Oemijati & Cross, 1977 cause lymphatic diseases in humans in the tropics, while B. pahangi (Buckley & Edeson, 1956) infects carnivores and causes zoonotic diseases in humans in Malaysia. Wuchereria bancrofti, W. kalimantani Palmieri, Pulnomo, Dennis & Marwoto, 1980 and six out of ten Brugia spp. have been described from Australia, Southeast Asia, Sri Lanka and India. However, the origin and evolution of the species in the Wuchereria-Brugia clade remain unclear. While investigating the diversity of filarial parasites in Malaysia, we discovered an undescribed species in the common treeshrew Tupaia glis Diard & Duvaucel (Mammalia: Scandentia). We examined 81 common treeshrews from 14 areas in nine states and the Federal Territory of Peninsular Malaysia for filarial parasites. Once any filariae that were found had been isolated, we examined their morphological characteristics and determined the partial sequences of their mitochondrial cytochrome c oxidase subunit 1 (cox1) and 12S rRNA genes. Polymerase chain reaction (PCR) products of the internal transcribed spacer 1 (ITS1) region were then cloned into the pGEM-T vector, and the recombinant plasmids were used as templates for sequencing. Malayfilaria sofiani Uni, Mat Udin & Takaoka, n. g., n. sp. is described based on the morphological characteristics of adults and microfilariae found in common treeshrews from Jeram Pasu, Kelantan, Malaysia. The Kimura 2-parameter distance between the cox1 gene sequences of the new species and W. bancrofti was 11.8%. Based on the three gene sequences, the new species forms a monophyletic clade with W. bancrofti and Brugia spp. The adult parasites were found in tissues surrounding the lymph nodes of the neck of common treeshrews. The newly described species appears most closely related to Wuchereria spp. and Brugia spp., but differs from these in several morphological characteristics. Molecular analyses based on the cox1 and 12S rRNA genes and the ITS1 region indicated that this species differs from both W. bancrofti and Brugia spp. at the genus level. We thus propose a new genus, Malayfilaria, along with the new species M. sofiani.

Objective: To evaluate the prevalence and divergence of genetically identified Leptospira spp. in the population of Rattus rattus. Methods: A total of 130 rats were used in this study. The infection within the rats were screened using polymerase chain reaction (PCR)-based diagnosis, with Leptospira genus-specific 16S rRNA primer and pathogenic Leptospira spp. specific LipL32 primer, on both kidney and liver tissues of Rattus rattus to detect the presence of potential Leptospira spp. Results: Out of 130 rats studied, 51 (39.23%) individuals were positive for leptospiral DNA. Basic Local Alignment Search Tool (BLAST) and phylogenetic analysis revealed that both pathogenic Leptospira interrogans and Leptospira borgpetersenii were predominantly identified. Phylogenetically, both genes disclosed similar clustering patterns of tree topologies between the two species. Although both genes were conserved, LipL32 gene portrayed higher nucleotide divergence (5.80%) compared to the 16S rRNA gene (0.60%). Minimum-spanning network displayed several haplotypes that are unique to each species, suggesting a higher degree of subdivision between both species. As for prevalence surveillance, both adult and subadult rats were susceptible to the infection, in which males were the most susceptible. Kidney was notable as the favourable organ for colonisation of leptospires. Rats captured from fresh markets were highly infected with Leptospira spp. (54.28%) compared to those from housing areas (26.47%). Conclusions: Rattus rattus represents an important asymptomatic transmitter of pathogenic leptospires, and hence is of public health concerns.