Entamoeba moshkovskii and Entamoeba dispar are microscopically indistinguishable from the pathogenic species Entamoeba histolytica. Although sporadic cases of human infection with E. moshkovskii have been reported, the amoeba is still considered primarily as a free-living amoeba. A cross-sectional study was carried out among Orang Asli communities in 3 different states of Peninsular Malaysia. Fecal samples were examined by formalin-ether sedimentation and trichrome staining techniques and then single-round PCR assay was used to detect E. moshkovskii. Out of 500 fecal samples examined microscopically, 93 (18·6%) samples were positive for E. histolytica/E. dispar/E. moshkovskii complex cysts and/or trophozoites. PCR products were detected in 106 fecal samples. E. moshkovskii isolates were detected in 13 (12·3%) fecal samples. Of the 13 E. moshkovskii-positive samples, 5 were of single isolation of E. moshkovskii, 6 were also positive for E. dispar, and only 2 samples were positive for E. dispar and E. histolytica. Moreover, 3 E. moshkovskii-positive samples were collected from symptomatic individuals while the remaining 10 samples were from asymptomatic subjects. This is the first report on the identification of E. moshkovskii in Malaysia. Further studies are needed to confirm the pathogenicity of E. moshkovskii infection and determine the epidemiology among Orang Asli communities in Malaysia.
Soil-transmitted helminthiasis (STH), among the most common neglected tropical diseases, is a major public health problem in Malaysia with a possible impact on the nutritional status and school participation of rural children. This study was carried out among Aboriginal schoolchildren, living in an endemic area for STH in Malaysia, to determine the possible relationship between intestinal helminthiasis and school absenteeism. We also evaluated whether successful treatment of the infection will affect school attendance among the subjects. Stool analysis revealed that more than 90% of the subjects were infected with at least 1 helminth species, with Ascaris lumbricoides and Trichuris trichiura infections being most prevalent. Infection of moderate-to-heavy worm burdens, low level of fathers' education and anaemia were identified as the significant predictors of high absenteeism among the subjects (P<0·05). Following treatment of the infected children, it was found that school absenteeism was reduced significantly (P<0·01). In conclusion, STH continues to have significant impacts on public health, particularly in rural communities with a negatively significant effect on the school participation of Aboriginal children. A school-based de-worming programme should be introduced and incorporated in the current educational assistance targeted towards the Aboriginal communities, under the auspices of the government.
Brugia malayi is one of the parasitic worms which causes lymphatic filariasis in humans. Its geographical distribution includes a large part of Asia. Despite its wide distribution, very little is known about the genetic variation and molecular epidemiology of this species. In this study, the internal transcribed spacer 1 (ITS1) nucleotide sequences of B. malayi from microfilaria-positive human blood samples in Northeast Borneo Island were determined, and compared with published ITS1 sequences of B. malayi isolated from cats and humans in Thailand. Multiple alignment analysis revealed that B. malayi ITS1 sequences from Northeast Borneo were more similar to each other than to those from Thailand. Phylogenetic trees inferred using Neighbour-Joining and Maximum Parsimony methods showed similar topology, with 2 distinct B. malayi clusters. The first cluster consisted of Northeast Borneo B. malayi isolates, whereas the second consisted of the Thailand isolates. The findings of this study suggest that B. malayi in Borneo Island has diverged significantly from those of mainland Asia, and this has implications for the diagnosis of B. malayi infection across the region using ITS1-based molecular techniques.
Blastocystis infection has a worldwide distribution especially among the disadvantaged population and immunocompromised subjects. This study was carried out to determine the prevalence and the association of Blastocystis infection with the socio-economic characteristics among 300 primary schoolchildren, living in rural communities in Lipis and Raub districts of Pahang state, Malaysia. Stool samples were collected and examined for the presence of Blastocystis using direct smear microscopy after in vitro cultivation in Jones' medium. The overall prevalence of Blastocystis infection was found to be as high as 25.7%. The prevalence was significantly higher among children with gastrointestinal symptoms as compared to asymptomatic children (x2 =4.246; P=0.039). Univariate and multivariate analyses showed that absence of a piped water supply (OR=3.13; 95% CI=1.78, 5.46; P<0.001) and low levels of mothers' education (OR=3.41; 95% CI=1.62, 7.18; P<0.01) were the significant predictors of Blastocystis infection. In conclusion, Blastocystis is prevalent among rural children and the important factors that determine the infection were the sources of drinking water and mothers' educational level. Interventions with provision of clean water supply and health education especially to mothers are required.
Cryptosporidium species are protozoan parasites that infect humans and a wide variety of animals. This study was aimed at identifying Cryptosporidium species and genotypes isolated from avian hosts. A total of 90 samples from 37 different species of birds were collected throughout a 3-month period from April 2008 to June 2008 in the National Zoo of Kuala Lumpur, Malaysia. Prior to molecular characterization, all samples were screened for Cryptosporidium using a modified Ziehl-Neelsen staining technique. Subsequently samples were analysed with nested-PCR targeting the partial SSU rRNA gene. Amplicons were sequenced in both directions and used for phylogenetic analysis using Neighbour-Joining and Maximum Parsimony methods. Although 9 (10%) samples were positive for Cryptosporidium via microscopy, 8 (8.9%) produced amplicons using nested PCR. Phylogenetic trees identified all the isolates as Cryptosporidium parvum. Although C. parvum has not been reported to cause infection in birds, and the role of birds in this study was postulated mainly as mechanical transporters, these present findings highlight the significant public health risk posed by birds that harbour the zoonotic species of Cryptosporidium.
Limitations with current chemotherapeutic and vaccinal control of coccidiosis caused by Eimeria species continue to prompt development of novel controls, including the identification of new drug targets. Glucose-6-phosphate isomerase (G6-PI) has been proposed as a valid drug target for many protozoa, although polymorphism revealed by electrophoretic enzyme mobility has raised doubts for Eimeria. In this study we identified and sequenced the Eimeria tenella G6-PI orthologue (EtG6-PI) from the reference Houghton strain and confirmed its position within the prevailing taxonomic hierarchy, branching with the Apicomplexa and Plantae, distinct from the Animalia including the host, Gallus gallus. Comparison of the deduced 1647 bp EtG6-PI coding sequence with the 9016 bp genomic locus revealed 15 exons, all of which obey the intron-AG-/exon/-GT-intron splicing rule. Comparison with the Weybridge and Wisconsin strains revealed the presence of 33 single nucleotide polymorphisms (SNPs) and 14 insertion/deletion sites. Three SNPs were exonic and all yielded non-synonymous substitutions. Preliminary structural predictions suggest little association between the coding SNPs and key G6-PI catalytic residues or residues thought to be involved in the coordination of the G6-PI's substrate phosphate group. Thus, the significant polymorphism from its host orthologue and minimal intra-specific polymorphism suggest G6-PI remains a valid anti-coccidial drug target.
This study was conducted to determine the genotypes of Giardia duodenalis isolated from human faecal samples at Pos Betau, Pahang, Malaysia. Faecal specimens were collected and examined for G. duodenalis cysts using Trichrome staining techniques. Molecular identification was carried out by the amplification of a region of the small subunit of the nuclear ribosomal RNA (SSU rRNA) gene using nested PCR and subsequent sequencing. The sequences from 15 isolates from G. duodenalis were subjected to phylogenetic analysis (including appropriate outgroups) using the neighbor-joining and maximum parsimony methods. The trees identified G. duodenalis assemblages A and B, with a predominance of assemblage B. The predominance of anthroponotic genotypes indicates the possibility of anthroponotic transmission of these protozoa in this Semai Pahang Orang Asli community.
Numerous studies have revealed the presence of oxidative stress in parasitic infections. However, such studies were lacking in the Malaysian population. Previously, we have provided evidence that oxidative stress is elevated in Malaysians infected with intestinal parasites. Stool examinations revealed that about 47.5% of them were infected with the polymorphic protozoa, Blastocystis hominis. However, they were found to have mixed infection with other intestinal parasites.
Unusual tumour-like pathologies caused by mysterious cells termed 'X-cells' have been reported from numerous fish groups worldwide. After nearly 100 years of research, the tumour-like growths have recently been shown to be caused by a protozoan parasite. In the present study, histopathology and small subunit ribosomal DNA (SSU rDNA) sequences are used to assess whether the X-cell parasite infecting Atlantic dab Limanda limanda L. is distinct from the X-cell parasite infecting Japanese flounder and goby, and to determine their systematic position within the protists. SSU rDNA from Scottish dab was 89.3% and 86.7% similar to Japanese X-cell sequences from flounder and goby respectively, indicating that the parasite infecting dab in the Atlantic is distinct from the Pacific species. Histological studies revealed significant gill pathology and demonstrated the precise location of the parasites within the gill tissues using specific in situ hybridization probes. Phylogenetic analyses showed that the X-cell parasites from Scotland and Japan form a monophyletic group within the Myzozoa, and are basal alveolates. However, ultrastructure of X-cells from dab fails to confirm this systematic placement.
Matched MeSH terms: Fish Diseases/parasitology*; Flatfishes/parasitology*; Protozoan Infections, Animal/parasitology*
Oxidative stress has been implicated as an important pathogenic factor in the pathophysiology of various life-threatening diseases such as cancer, cardiovascular diseases and diabetes. It occurs when the production of free radicals (generated during aerobic metabolism, inflammation, and infections) overcome the antioxidant defences in the body. Although previous studies have implied that oxidative stress is present in serum of patients with parasitic infection there have been no studies confirming oxidative stress levels in the Malaysian population infected with intestinal parasites. Three biochemical assays namely hydrogen peroxide (H2O2), lipid peroxidation (LP) and advanced oxidative protein product (AOPP) assays were carried out to measure oxidative stress levels in the urine of human subjects whose stools were infected with parasites such as Blastocystis hominis, Ascaris, Trichuris, hookworm and microsporidia. The levels of H2O2, AOPP and LP were significantly higher (P<0.001, P<0.05 and P<0.05 respectively) in the parasite-infected subjects (n=75) compared to the controls (n=95). In conclusion, the study provides evidence that oxidative stress is elevated in humans infected by intestinal parasites. This study may influence future researchers to consider free radical-related pathways to be a target in the interventions of new drugs against parasitic infection and related diseases.
The protozoan parasite Toxoplasma gondii produces a family of microneme proteins that are thought to play diverse roles in aiding the parasite's intracellular existence. Among these, TgMIC2 has a putative function in parasite adhesion to the host cell to initiate the invasion process. The invasion process may be localized and inhibited by monoclonal antibodies against the protein(s) involved. Here we report on the construction of a phage-displayed single-chain variable fragment (scFv) library from mice immunized with whole T. gondii parasites. The library was subsequently panned against recombinant TgMIC2 (rpTgMIC2) and 2 different groups of antibody clones were obtained, based on fingerprinting and sequencing data. The expressed recombinant scFv antibody was able to recognize rpTgMIC2 in a Western blot detection experiment. These results show that the phage display technology allows quick and effective production of monoclonal antibodies against parasite antigens. By panning the scFv-displayed library, we should be able to obtain a plethora of multi-functional scFv antibodies towards T. gondii proteins.
Cryptosporidium is a protozoan parasite of humans and animals and has a worldwide distribution. The parasite has a unique epidemiology in Middle Eastern countries where the IId subtype family of Cryptosporidium parvum dominates. However, there has been no information on Cryptosporidium species in Yemen. Thus, this study was conducted in Yemen to examine the distribution of Cryptosporidium species and subtype families. Fecal samples were collected from 335 patients who attended hospitals in Sana'a city. Cryptosporidium species were determined by PCR and sequence analysis of the 18 s rRNA gene. Cryptosporidium parvum and C. hominis subtypes were identified based on sequence analysis of the 60 kDa glycoprotein (gp60) gene. Out of 335 samples, 33 (9.9%) were positive for Cryptosporidium. Of them, 97% were identified as C. parvum whilst 1 case (3%) was caused by C. hominis. All 7 C. parvum isolates subtyped belonged to the IIaA15G2R1 subtype. The common occurrence of the zoonotic IIa subtype family of C. parvum highlights the potential occurrence of zoonotic transmission of cryptosporidiosis in Yemen. However, this postulation needs confirmation with future molecular epidemiological studies of cryptosporidiosis in both humans and animals in Yemen.
A study of the anterior adhesive apparatus (head organs) of Bravohollisia gussevi Lim, 1995 was carried out using light and electron microscopy. The anterior adhesive apparatus or head organs in B. gussevi comprise 6 circular openings or apertures in the antero-lateral region, associated pits lined with specialized microvillous tegument that differ from the general body tegument, a bundle of ducts, and uninucleate gland cells located lateral to the pharynx. The uninucleate glands of the anterior adhesive apparatus (head organs) comprise 2 types of cells, one kind of cell producing rod-like bodies (S1) and the other oval bodies (S2). The S1 bodies are filled with numerous, less electron-dense vesicles in an electron-dense matrix, while S2 bodies have no vesicles but contain a more homogeneous electron-dense matrix. Interlinking band-like structures were observed between S1 bodies. Similar band-like structures were found between S2 bodies. The formation of S1 bodies was followed by transmission electron microscopy. However, the formation of S2 bodies was unclear and could not be resolved. Uniciliated structures were also observed around the openings of the anterior adhesive apparatus. Each uniciliated structure is usually associated with an opening of a gland cell producing granular, electron-dense, secretory bodies, which differ from the secretions produced by the lateral gland cells of the anterior adhesive apparatus.
Matched MeSH terms: Fish Diseases/parasitology*; Gills/parasitology; Perciformes/parasitology*; Trematode Infections/parasitology
Plasmodium knowlesi a simian malaria parasite is currently affecting humans in Southeast Asia. Malaysia has reported the most number of cases and P. knowlesi is the predominant species occurring in humans. The vectors of P. knowlesi belong to the Leucosphyrus group of Anopheles mosquitoes. These are generally described as forest-dwelling mosquitoes. With deforestation and changes in land-use, some species have become predominant in farms and villages. However, knowledge on the distribution of these vectors in the country is sparse. From a public health point of view it is important to know the vectors, so that risk factors towards knowlesi malaria can be identified and control measures instituted where possible. Here, we review what is known about the knowlesi malaria vectors and ascertain the gaps in knowledge, so that future studies could concentrate on this paucity of data in-order to address this zoonotic problem.
The microsporidian genus Nosema is characterized by development in direct control with host cell cytoplasm, diplokaryotic nuclei throughout development and disporous sporogony. The genus Vairimorpha exhibits the same features plus an octoporous sporogony producing uninucleate spores in a sporophorous vesicle. A microsporidium from diamondback moth, Plutella xylostella, falls between Nosema and Vairimorpha in that it initiates but fails to complete the octosporous sequence in this host. The name Vairimorpha imperfecta n.sp. is proposed. Merogony is mainly by formation of buds from multinucleate meronts, the buds remaining attached in chains. Diplokaryotic spores measure 4.3 x 2.0 microns (fresh) and have 15.5 coils of the polar tube in 1 rank. The octosporous sporogony is aborted owing to irregular formation of nuclear spindles, incomplete cytoplasmic fission and bizarre deposition of electron-dense episporontal secretions. Phylogenetic analyses of the sequences of the small subunit rRNA genes of V. imperfecta and of several Nosema and Vairimorpha spp. place V. imperfecta in a clade with Nosema spp. from Lepidoptera rather than in the clade containing the more typical species of Vairimorpha. It is suggested that the ancestors of the Vairimorpha/Nosema complex of species exhibited both disporous and octosporous sporogonies, as does the type species of Vairimorpha, Vairimorpha necatrix. It would follow that true Nosema spp. have lost the ability to express an octosporous sequence and that V. imperfecta is in the process of losing it. It is proposed that the genera Nosema and Vairimorpha be placed in the same family Nosematidae Labbé 1899, rather than in separate families and orders as at present.
18S rDNA sequences from 4 isolates of Babesia gibsoni originating from Japan, Malaysia and Sri Lanka were compared with a previously published, 0.5 kb portion of the 18S rDNA from a B. gibsoni isolate from California, USA, and with the corresponding 18S rDNA sequences of other Babesia spp. Distance, parsimony and maximum likelihood analyses showed almost identical genotypes among the small canine Babesia from Asia, but an unexpectedly distant genetic relationship to that from the USA. While the American isolate segregated together with B. equi, the Asian isolates showed a close relationship to B. divergens and B. odocoilei. These results indicate that small Babesia of dogs originating from North America and Asia belong to different, genetically distantly related species.
Matched MeSH terms: Babesiosis/parasitology*; Dog Diseases/parasitology*
This study examines the persistence of predisposition to Ascaris lumbricoides and Trichuris trichiura during repeated chemotherapy in an urban community in Kuala Lumpur, Malaysia. Significant predisposition was observed over 2 periods of reinfection with and without age-standardization of data. Analysis of different age groups indicated that predisposition was most strongly detectable in the younger age classes. The intensities of infection with both parasites were strongly correlated at each cycle of intervention, suggesting that individuals were similarly predisposed to both species.
The ascaridoid nematode of cats from Kuala Lumpur, Malaysia, previously identified morphologically as Toxocara canis, was characterized using a molecular approach. The nuclear ribosomal DNA (rDNA) region spanning the first internal transcribed spacer (ITS-1), the 5.8S gene and the second internal transcribed spacer (ITS-2) was amplified and sequenced. The sequences for the parasite from Malaysian cats were compared with those for T. canis and T. cati. The sequence data showed that this taxon was genetically more similar to T. cati than to T. canis in the ITS-1, 5.8S and ITS-2. Differences in the ITS-1 and ITS-2 sequences between the taxa (9.4-26.1%) were markedly higher than variation between samples within T. canis and T. cati (0-2.9%). The sequence data demonstrate that the parasite from Malaysian cats is neither T. canis nor T. cati and indicate that it is a distinct species. Based on these data, PCR-linked restriction fragment length polymorphism (RFLP) and single-strand conformation polymorphism (SSCP) methods were employed for the unequivocal differentiation of the Toxocara variant from T. canis and T. cati. These methods should provide valuable tools for studying the life-cycle, transmission pattern(s) and zoonotic potential of this parasite.
Nucleotide sequences were obtained for the second internal transcribed spacer of the ribosomal gene repeat and for part of the mitochondrial-cytochrome c oxidase subunit I gene from geographical isolates of Paragonimus westermani from Japan, China, Korea, Taiwan, the Philippines, peninsular Malaysia and Thailand. Sequences were obtained from several other species of Paragonimus for comparative purposes. Two groups were recognized within P. westermani: an NE group (China, Japan, Korea, Taiwan) which was relatively uniform and included both diploid and triploid forms, and a southern group (Malaysia, Thailand, Philippines), members of which were genetically distant from one another. According to both ITS2 and COI data, genetic distances among P. westermani isolates equalled or exceeded those between some distinct species of Paragonimus. The ITS2 sequences were conserved relative to COI sequences. Substitutions among the latter may be approaching saturation within the genus Paragonimus.
This cross-sectional study aimed to determine the prevalence and risk factors of S. stercoralis infection among 1142 Orang Asli primary schoolchildren in six different states of Peninsular Malaysia. Fecal samples were examined using direct smear, formalin-ether sedimentation (FES), agar plate culture (APC) and PCR techniques. Overall, 15.8% of the children were found to be infected with S. stercoralis. The prevalence was 0.2, 1.3, 15.2 and 13.7% by direct smear, FES, APC and PCR, respectively. Multivariate analysis showed that an age of >10 years, being male, belonging to a Proto-Malay tribe, belonging to the Senoi tribe, indiscriminate defecation, using an unimproved water source for drinking water and not wearing shoes when outside were the significant risk factors of infection among these children. In conclusion, we provide new evidence on the occurrence of S. stercoralis in Malaysia to show that there is a relatively high prevalence of infection among Orang Asli schoolchildren. Therefore, the use of specific methods for detecting S. stercoralis should be considered when screening these children for intestinal parasites. Moreover, prevention and control measures specific to S. stercoralis should be integrated into the intestinal parasitic infections control programme in Malaysia.