Fifty faecal samples from diarrheic calves between 1 and 6 months old were collected per rectum from 5 farms around Petaling District in Selangor, Malaysia for Cryptosporidium species detection and genotyping investigation. Oocysts were purified using sedimentation and gradient centrifugation, then examined by immunofluorescence assay (IFAT). Genomic DNA was extracted from all samples and nested PCR was performed to amplify the SSU rRNA gene. Eighteen samples (36%) were positive for Cryptosporidium species by PCR. The sequence and phylogenetic analysis of 14 isolates indicated that Cryptosporidium parvum was most common (11 isolates) followed by Cryptosporidium deer-like genotype (3 isolates). The present work reports the first data on Cryptosporidium genotyping from cattle in Malaysia.
This study was conducted to reassess the risk factors for giardiasis in communities of the Orang Asli (indigenous people) in Pahang, Malaysia. Stool samples were collected from 321 individuals (2-76 years old; 160 males, 161 females). Data were collected via laboratory analysis of faecal samples and a pre-tested standard questionnaire. River water samples were tested for Giardia cysts and Cryptosporidium oocysts. The overall prevalence of G. intestinalis infection was 23.7%. Children < or =12 years old had the highest infection rate and have been identified as a high risk group (odds ratio (OR)=6.2, 95% CI 1.5-27.0, P<0.005). The risk of getting giardiasis also appeared to be significantly associated with drinking piped water (OR=5.1, 95% CI 0.06-0.7, P<0.005) and eating raw vegetables (OR=2.4, 95% CI 0.2-0.6, P<0.005). In conclusion, sociodemographic factors have always been associated with the high prevalence of Giardia infections in Malaysia. However, the present study also highlights the need to look into the possibility of other risks such as water and food transmission routes. In future, it is necessary that these two aspects be considered in control strategies.
This cross-sectional study determined the prevalence of cryptosporidiosis in HIV-infected patients using polymerase chain reaction (PCR). Stool specimens were collected from HIV infected patients who were admitted to Hospital Raja Perempuan Zainab II, Kota Bharu, Malaysia, for various indications from December 2004 to December 2005. A modified acid-fast stain was performed on the direct stool smears, then the stool specimens were further tested using nested PCR targeting the 18S rRNA gene of Cryptosporidium parvum, with a built-in internal control (IC). Out of 59 samples, 11 were positives. Nested PCR identified a total of nine samples (16%) compared to microscopy, which identified only three samples. All PCR negative results showed IC amplicons, suggesting that these samples were true negatives and were not due to inhibition of PCR. This study highlights the importance of molecular diagnosis in determining the true prevalence and epidemiology of C. parvum.
Data on soil-transmitted helminth (STH) infections and reinfection among Orang Asli (aborigine) schoolchildren and their nutritional and socioeconomic status were analyzed to investigate the pattern and the possible predictors of STH reinfection. In this longitudinal study, 120 (60 males and 60 females) Orang Asli primary schoolchildren aged 7-12 years and living in remote areas in Pos Betau, Kuala Lipis, Pahang were screened for the presence of STH using modified cellophane thick smear and Harada Mori techniques. The overall prevalence of ascariasis, trichuriasis and hookworm infections were 65.8, 97.5 and 10.8%, respectively. After complete deworming with a 3-day course of 400mg/daily of albendazole tablets, children were re-examined at 3 and 6 months from baseline. The reinfection rate, by one or more of STH species, at 3 months after deworming was high (49.5%) while 79.6% of the children were reinfected at 6 months after deworming. Logistic regression analyses showed that females, stunted children and those living in houses without toilets had significantly higher reinfection rates than others at 3 months (P<0.05). At 6 months, maternal employment status emerged as another predictor where children of working mothers had significantly higher reinfection rates (P=0.026). In conclusion, reinfection rate of STH is high and thus necessitates frequent and periodic deworming among children. Public health personnel need to re-look at the current control measures and identify innovative and integrated ways in order to reduce STH significantly in the rural communities.
Parasites and bacteria are reported in the faeces of birds in the current study. Fresh faecal samples of the large-billed crow (Corvus spp.) were collected from the study site at Bangsar, an urban setting in Kuala Lumpur, Malaysia. These samples were transported to laboratory and analysed for parasites and bacteria. Pre-prepared XLD agar plates were used for culturing the bacteria in the laboratory. Using the API 20ETM Test Strips, 9 different species of bacteria were identified belonging to the family Enterobacteriacea. They were Citrobacter freundii, Enterobacter cloacae, Proteus mirabilis, Klebsiella pneumoniae, Kluyvera ascorbata, Salmonella arizonae, Salmonella typhi, Shigella flexneri and Shigella sonnei. The protozoan parasites detected include Cryptosporidium spp., Cyclospora spp., Blastocystis spp., and Capillaria hepatica and Ascaris lumbricoidus ova. Environmental air samples collected on agar plates using an air sampler in the area only produced fungal colonies. Some of these pathogens found in the crows are of zoonotic importance, especially Cryptosporidium, Blastocystis, Cyclopsora, Salmonella, Shigella and Kluyvera. The finding of Kluyvera spp. in crows in our current study highlights its zoonotic potential in an urban setting.
Despite being a common skin dermatosis in the tropics, physicians in the tropics may miss the diagnosis of cutaneous larva migrans for other pruritic skin manifestation. This is especially in those who live in urban housing with no history of travel. Cutaneous larva migrans, an intensely pruritic skin pathology is mainly contracted by people with history of beach holiday or contact with moist soft sand which had been contaminated with dog or cat faeces. This article reports a patient who presented with intensely itchy papular spots over the dorsum of his foot after walking barefooted in an urban toilet soiled with cat faeces. The patient had initially seen an urban general practitioner who diagnosed the papular skin lesion as an allergic reaction, and prescribed antihistamines. The patient subsequently developed creeping skin lesions and was seen by the author who prescribed albendazole 400 mg twice daily for three days. The patient reported reduction in itching after two days of albendazole treatment and a follow up at ten days revealed a healed infection.
A cross-sectional study to determine the current prevalence of anaemia and iron deficiency anaemia (IDA) and to investigate the possible risk factors for IDA was carried out on 241 aboriginal schoolchildren (120 boys, 121 girls) aged 7-12 years and living in remote areas in Pos Betau, Pahang, Malaysia. Haemoglobin (Hb) level was measured and serum iron status was assessed by serum ferritin (SF), serum iron (SI) and total iron binding capacity measurements. Socioeconomic data were collected using pre-tested questionnaires. All children were screened for intestinal parasitic infections. Overall, 48.5% (95% CI 42.3-54.8) of children were anaemic (Hb<12 g/dl). The prevalence of IDA was 34% (95% CI 28.3-40.2), which accounted for 70.1% of the anaemia cases. The prevalence of IDA was significantly higher in females than males. Low levels of mothers' education and low household income were identified as risk factors for IDA. Severe trichuriasis also found to be associated with low SF and SI. Logistic regression confirmed low levels of mothers' education and gender as significant risk factors for IDA. Improvement of socioeconomic status and health education together with periodic mass deworming should be included in public health strategies for the control and prevention of anaemia and IDA in this population.
A survey was undertaken to investigate the prevalence of intestinal parasites from different groups of mammals housed in a zoological garden in Malaysia. A total of 197 faecal samples were collected randomly from various primates (99), hoofed mammals (70) and feline (28). It was discovered that 89.3% of feline, 54.5% of primates and 45.7% of hoofed mammals were infected with intestinal parasites. Intestinal parasites found in primates were Balantidium coli (19.2%), Cryptosporidium spp. (14.1%), hookworm (10.1%), Trichuris spp. (5.1%), Ascaris (4.0%) and Blastocystis spp. (2.0%). For hoofed mammals, hookworm had the highest prevalence (34.3%) followed by Trichuris spp. and Cryptosporidium spp. (5.7%). Meanwhile, for feline, Toxocara cati was the most prevalent (64.3%), followed by Cryptosporidium spp. (14.3%), Spirometra spp. (7.1%), and hookworm (3.6%). Animals that were infected were all asymptomatic with low parasite load. Routine monitoring of the presence of parasites in animals kept in the zoo is imperative in assisting zoo management in the formulation and implementation of preventive and control measures against the spread of infectious parasitic diseases among animals within the zoo or to humans.
An observational study was carried out to establish the existence of intestinal microsporidiosis among patients with and without gastrointestinal symptoms in Hospital Universiti Kebangsaan Malaysia, Malaysia. A total of 893 faecal specimens from hospitalized patients were examined for microsporidia using a modification of the usual Gram-chromotrope stain technique. One hundred and sixteen (13.0%) patients were positive for microsporidia: 84 (72.4%), 27 (23.3%) and 5 (4.3%) were low, moderate and high excreters of microsporidia spores, respectively. Of the 91 patients with available medical records, microsporidiosis was commonly observed in children aged 0-6 years (26.4%) and adults aged >or=31 years (57.2%). About one-third of this infection was observed in immunocompetent individuals. Among the immunosuppressive group, microsporidia were observed to be more prevalent in patients with haematological malignancy or a combination of malignancy and diabetes mellitus. About 74% of the patients who had microsporidia in their faeces had gastrointestinal symptoms, which could be related to infections or induced by immunosuppressive therapy. The role of microsporidia in causing gastrointestinal symptoms in this population is as yet unclear.
This paper reports the occurrence of helminth and protozoan infections on small ruminants from eight farms situated in Kinta and Perak Tengah district, Perak. The results of this survey indicate that helminthiasis and coccidiosis is rampant in sheep and goat farms. Several anthelmintics have been used for the control of helminths. The smallholders depended on health and extention services from the State Veterinary Department. This survey is part of an ongoing programme by the Department of Veterinary Services to upgrade services and report the current status of parasitic diseases in the state.
Blastocystis from infected stools of a person who showed chronic symptoms of abdominal discomfort and diarrhea were examined over a 6-month period, using transmission electron microscopy, for the ultrastructural changes from vacuolar to cystic stage. The study confirms the irregular shedding phenomenon of the organism previously reported, and for the first time, records sequential changes in encystation in stools collected over a time period. The study also confirms the existence of a precystic stage which has an immature cell wall consisting of a layer of a homogenous electron-dense mass surrounding the cell which acts as a intermediatory stage between the vacuolar and cystic stage.
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.
This study was conducted to identify genotypes related risk factors of Giardia intestinalis in an Orang Asli (aboriginal) community in Pahang, Malaysia. Stool samples were collected from 321 individuals aged between 2 and 76 years old, of whom 160 were males and 161 were females. Faecal samples were processed with trichrome staining technique for the primary identification of G. intestinalis. Molecular identification was carried out by the amplification of a partial SSU rRNA gene using nested PCR. PCR products were purified and genotyped. 42 samples successfully amplified from the 76 positive faecal samples, only 1 was Assemblage A, the rest were Assemblage B. Risk analysis based on the detected genotypes of Giardia using univariate analysis and logistic regression identified three significant risk factors of giardiasis caused by assemblage B which included children =12 years (OR=13.56, 95% CI=1.79-102.64, p=0.012), females (OR=2.52, 95% CI=1.11-5.75, p=0.027) and eating fresh fruits (OR=7.78, 95% CI=1.01-60.00, p=0.049). Assemblage B infection was significantly correlated with clinical symptoms of giardiasis (OR=2.4, 95% CI=1.13-5.12, p=0.019). Females infected with Assemblage B were at higher risk of manifesting gastroenteritis signs and symptoms (OR=3.9, 95% CI=1.50-10.31, p=0.004). It has been concluded that giardiasis is still a public health problem in Orang Asli community and most commonly caused by assemblage B. The dynamic of transmission is most probably anthroponotic which is human to human either directly or indirectly through contaminated food. This route of transmission should be considered in the control strategy of the disease. Mass treatment together with health education could be the most practical intervention for reducing the infection. Those at high risk should receive more attention from public health authorities.
This study is the first report on the occurrence of Giardia and Cryptosporidium (oo)cysts in recreational rivers water from Malaysia. It was carried out in water samples at two rivers, 'Sungai Congkak' and 'Sungai Batu', located in Selangor State. The occurrence of both Giardia lamblia and Cryptosporidium parvum (oo)cysts was higher in Sungai Congkak (50% or 15/30 and 10% or 3/30 respectively) than Sungai Batu (16% or 5/30 and 3.3% or 1/30 respectively). The mean density of cysts/L was 0.72 in Sungai Congkak and 0.023 in Sungai Batu, and that of oocysts/L was 0.023 in Sungai Congkak and 0.0033 in Sungai Batu, showing that the occurrence of Giardia was higher and more frequent than Cryptosporidium in both rivers. Sungai Congkak also showed higher faecal coliforms count (ranging from 0.48x10³ to 73x10³ CFU/100 mL) than Sungai Batu (0.41x10³ to 16x10³ CFU/100 mL). On the other hand, the Giardia and Cryptosporidium (oo)cysts and faecal coliforms were more concentrated at the downstream station, followed by midstream and upstream stations which might be due to human factors where settlements and recreation areas were located around and between midstream and downstream stations. The (oo)cysts and faecal coliforms also increased during public holidays due to the significantly higher number of visitors (bathers) compared with the week days. All the parameters (physical, faecal coliforms and rainfall) did not show consistent significant correlation (based on r values of Pearson correlation analysis) with both protozoa, therefore these parameters are not suitable as indicator for the presence of Giardia and Cryptosporidium (oo)cysts in both rivers.
Microsporidia are ubiquitous parasites thought to be closely related to fungi. Their presence in the environment means that humans are frequently exposed to infection. Stool samples were collected from 151 indigenous villagers from the eastern state of Pahang in 2005. The samples were concentrated with water-ether sedimentation, stained with modified trichrome stain and examined under oil-immersion microscopy. Thirty-two specimens (21.2%) were positive for microsporidia. Microsporidia were observed as ovoid or rounded ovoid shapes measuring approximately 1mum, with a bright pink outline containing a central or posterior vacuole. PCR amplification with specific primers on microscopy-positive specimens amplified Encephalitozoon intestinalis DNA from five of the ten specimens used.
Entamoeba histolytica causes about 50 million infections worldwide with a death rate of over 100,000 annually. In endemic developing countries where resources are limited, microscopic examinations based on Wheatley trichrome staining is commonly used for diagnosis of intestinal amoebiasis. Other than being a time-consuming method, it must be performed promptly after stool collection as trophozoites disintegrate rapidly in faeces. The aim of this study was to compare the efficacies of Eosin-Y, Wheatley trichrome and Iodine stains in delineating the diagnostic features of the parasite, and subsequently to determine the suitable microscopy observation period for detection of erythrophagocytic and non-erythrophagocytic trophozoites spiked in semi-solid stool sample. Wheatley trichrome staining technique was performed using the standard method while the other two techniques were performed on the slides by mixing the respective staining solution with the spiked stool sample. One million of axenically cultured non-erythrophagocytic E. histolytica and erythrophagocytic E. histolytica were separately spiked into 2 g of fresh semisolid faeces. Percentage viability of the trophozoites in the spiked stool sample was determined at 30 minute intervals for eight hours using the 0.4% Trypan blue exclusion method. The results showed that Eosin-Y and Wheatley trichrome stained the karyosome and chromatin granules better as compared to Iodine stain. The percentage viability of non-erythrophagocytic trophozoites decreased faster than the erythrophagocytic form in the first 5 hours and both dropped to ~10% in the 6th hour spiked sample. In conclusion, Eosin-Y staining technique was found to be the easiest to perform, most rapid and as accurate as the commonly used Wheatley trichrome technique; Eosin-Y stained slide sealed with DPX could also be kept as a permanent record. A period not exceeding 6 hours after stool collection was found to be the most suitable in order to obtain good microscopy results of viable trophozoites.
The fact whether Blastocystis hominis can invade has always been in question. Apart from a few sporadic studies such as that done on gnotobiotic guinea pigs which showed surface invasion and mucosal inflammation of the host's intestine caused by B. hominis infection, no real documentation of invasion has been proven. Studies have shown that hyaluronidase is secreted during the penetration into the host's skin and gut by nematode parasites. Hyaluronidase activity in protozoa namely Entamoeba histolytica has also been described previously. This study attempts to determine hyaluronidase in urine samples of B. hominis-infected rats. The presence of hyaluronidase in urine provides an indirect evidence of invasion by B. hominis into colonic epithelium causing the degradation of extracellular matrix proteins namely hyaluronic acid (HA). HA is depolymerized by hyaluronidase which may be used by organisms to invade one another. In this study, the levels of urinary hyaluronidase of Sprague-Dawley rats infected with B. hominis were monitored for 30 days. Hyaluronidase levels in the infected rats were significantly higher on days 28 and 30 compared to the day before inoculation (P < 0.01 and P < 0.05, respectively). During this stage, parasitic burden in infected stools was also at a high level. Proinflammatory cytokines, interleukin-6 and interleukin-8, were also significantly higher (P < 0.05) in the serum of infected rats. The study demonstrates that since no other pathogen was present and that amoeboid forms of the parasites have been shown to exist previously, the elevated levels of hyaluronidase in this preliminary finding suggests that the organism is capable of having invasion or penetration activity in the hosts' intestine.
Bovine lungworm Dictyocaulus viviparus is highly endemic in temperate regions. However, the occurrence of the lungworm has not been reported in any South East Asian country. The main aim of the present study was to detect the presence of lungworm in cattle in peninsular Malaysia and to examine the morphology of the parasite. A cross-sectional study was carried out in which 602 animals from four large scale government cattle farms and one dairy smallholder farm were sampled. In addition, 283 lungs from 11 abattoirs around the country were examined. Faecal samples were examined using the Baermann technique while post-mortem examination was performed on the lungs. Approximately 5% of faecal samples and 1% of lungs were positive for lungworm. Based on the morphology of adult lungworm, eggs and first stage larvae, Malaysian bovine lungworms were D. viviparus.
In order to obtain basic data on parasitic infections of Bornean orangutans, Pongo pygmaeus morio (Owen, 1837), in Danum Valley, Sabah, Malaysia, fecal examinations were conducted. Based on a total of 73 fecal samples from 25 individuals, cysts of Entamoeba coli, Entamoeba spp., and Chilomastix mesnili, cysts and trophozoites of Balantidium coli, and eggs of Trichuris sp. or spp., unknown strongylid(s), Strongyloides fuelleborni, and an unknown oxyurid, plus a rhabditoid larva of Strongyloides sp., were found. Mature and immature worms of Pongobius hugoti Baruš et al., 2007 and Pongobius foitovae n. sp. (Oxyuridae: Enterobiinae) were recovered from fecal debris and described. Pongobius foitovae is readily distinguished from P. hugoti by having a much longer esophageal corpus, a longer and distally hooked spicule in males, and a more posteriorly positioned vulva in female. Presence of plural species of non- Enterobius pinworms is a remarkable feature of the orangutan-pinworm relationship, which may reflect speciation process of the orangutans, host switching, and coevolution by pinworms.
A cross-sectional study was carried out to identify species and determine the prevalence of Cryptosporidium sp. shedding in pre-weaned and post-weaned dairy calves and to identify management factors that may be contributing to disease. A total of 240 calf faecal samples were collected from 16 farms in two districts in Johor, Malaysia, and screened by PCR. The overall Cryptosporidium prevalence was 27.1%. The prevalence of Cryptosporidium species in pre-weaned calves was 32.4% for C. parvum, 26.5% for C. bovis, followed by C. andersoni (20.6%), C. ryanae (11.8%) and mixed sp. (8.8%). The prevalence of Cryptosporidium species in post-weaned calves was 35% for C. bovis followed by C. andersoni and C. ryanae (30% each) and mixed sp. (5%). Subtyping analysis of 8 of the 11 C. parvum isolates at the gp60 locus identified five isolates as IIdA15G1, one as IIa18A3R1 and two isolates as IIa17G2R1. Management factors that increased the risk of Cryptosporidium infection included having other cattle farms close by, feeding calves with saleable milk, keeping pre-weaned calves in pens with slatted floors and keeping post-weaned calves in pens with a sand floor.