Veeranoot Nissapatorn

Abstract: Background: Plasmodium knowlesi is a simian malaria parasite that is widespread in humans in Malaysian Borneo. However, little is known about the incidence and distribution of this parasite in the Sandakan division, Malaysian Borneo. Therefore, the aim of the present epidemiological study was to investigate the incidence and distribution of P. knowlesi as well as other Plasmodium species in this division based on a most recent developed hexaplex PCR system (PlasmoNex (TM)). Methods: A total of 189 whole blood samples were collected from Telupid Health Clinic, Sabah, Malaysia, from 2008 to 2011. All patients who participated in the study were microscopically malaria positive before recruitment. Complete demographic details and haematological profiles were obtained from 85 patients (13 females and 72 males). Identification of Plasmodium species was conducted using PlasmoNex (TM) targeting the 18S ssu rRNA gene. Results: A total of 178 samples were positive for Plasmodium species by using PlasmoNex (TM). Plasmodium falciparum was identified in 68 samples (38.2%) followed by 64 cases (36.0%) of Plasmodium vivax, 42 (23.6%) cases of P. knowlesi, two (1.1%) cases of Plasmodium malariae and two (1.1%) mixed-species infections (ie, P. vivax/P. falciparum). Thirty-five PlasmoNex (TM) positive P. knowlesi samples were misdiagnosed as P. malariae by microscopy. Plasmodium knowlesi was detected in all four districts of Sandakan division with the highest incidence in the Kinabatangan district. Thrombocytopaenia and anaemia showed to be the most frequent malaria-associated haematological complications in this study. Conclusions: The discovery of P. knowlesi in Sandakan division showed that prospective studies on the epidemiological risk factors and transmission dynamics of P. knowlesi in these areas are crucial in order to develop strategies for effective malaria control. The availability of advanced diagnostic tool PlasmoNex (TM) enhanced the accuracy and accelerated the speed in the diagnosis of malaria.

Abstract: Toxoplasma gondii infects all warm-blooded animals including humans, causing serious public health problems and great economic loss in the food industry. Commonly used serological tests involve preparation of whole Toxoplasma lysate antigens from tachyzoites which are costly and hazardous. An alternative method for better antigen production involving the prokaryotic expression system was therefore used in this study. Recombinant dense granular protein, GRA2, was successfully cloned, expressed, and purified in Escherichia coli, BL21 (DE3) pLysS. The potential of this purified antigen for diagnosis of human infections was evaluated through western blot analysis against 100 human serum samples. Results showed that the rGRA2 protein has 100 and 61.5 % sensitivity towards acute and chronic infection, respectively, in T. gondii-infected humans, indicating that this protein is useful in differentiating present and past infections. Therefore, it is suitable to be used as a sensitive and specific molecular marker for the serodiagnosis of Toxoplasma infection in both humans and animals.

Abstract: Toxoplasma gondii is a public health risk in developing countries, especially those located in the tropics. Widespread infection may inflict a substantial burden on state resources, as patients can develop severe neurological defects and ocular diseases that result in lifelong loss of economic independence. We tested sera for IgG antibody from 493 eye patients in Malaysia. Overall age-adjusted seroprevalence was estimated to be 25% (95% CI: [21%, 29%]). We found approximately equal age-adjusted seroprevalence in Chinese (31%; 95% CI: [25%, 38%]) and Malays (29%; 95% CI: [21%, 36%]), followed by Indians (19%; 95% CI: [13%, 25%]). A logistic regression of the odds for T. gondii seroprevalence against age, gender, ethnicity and the occurrence of six types of ocular diseases showed that only age and ethnicity were significant predictors. The odds for T. gondii seroprevalence were 2.7 (95% CI for OR: [1.9, 4.0]) times higher for a patient twice as old as the other, with ethnicity held constant. In Malays, we estimated the odds for T. gondii seroprevalence to be 2.9 (95% CI for OR: [1.8, 4.5]) times higher compared to non-Malays, with age held constant. Previous studies of T. gondii seroprevalence in Malaysia did not explicitly adjust for age, rendering comparisons difficult. Our study highlights the need to adopt a more rigorous epidemiological approach in monitoring T. gondii seroprevalence in Malaysia. (C) 2012 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved.

Abstract: Seven stains were studied to determine the best color and contrast for staining the developmental stages of free living pathogenic Acanthamoeba and Naegleria species. The acid-fast bacilli stain (AFB) produced a blue color without contrast; trichrome-eosin and modified Field's showed various color contrasts; Giemsa, iron-hematoxylin, modified AFB and Gram produced only one color which distinguished the nucleus, nucleolus, cytoplasm, food- and water-vacuoles. The motile organs (acanthopodia, pseudopodia, lobopodia and flagella) were also clearly differentiated but produced a similar color as the cytoplasm. These motile organelles were first induced by incubating at 37 C for at least 15 minutes and then fixing with methanol in order to preserve the protruding morphology prior to staining. The trichrome-eosin and iron-hematoxylin stains showed good color contrast for detecting all three stages, the trophozoite, cyst and flagellate; Giemsa and Gram stained the trophozoite and flagellate stages; the modified Field's and modified AFB stains stained only the trophozoite stage. Depending on the purpose, all these stains (except the AFB stain) can be used to identify the developmental stages of Acanthanweba and Naegleria for clinical, epidemiological or public health use.

Abstract: Rhoptry protein 2 (ROP2) of Toxoplasma gondii is a rhoptry-secreted protein that plays a critical role in parasitophorous vacuole membrane formation during invasion. In previous studies, ROP2 has been shown to be efficient in triggering humoral and cell-mediated responses. High immunogenicity of ROP2 makes it a potential candidate for diagnosis and vaccination against toxoplasmosis. In this study, the ROP2 gene was cloned into pPICZ alpha A expression vector and extracellularly expressed in the yeast Pichia pastoris, which has numerous advantages over other expression systems for eukaryotic proteins expression. The effectiveness of the secreted recombinant ROP2 as a diagnosis agent was assessed by Western Blot with 200 human serum samples. Recombinant ROP2 reacted with toxoplasmosis-positive human serum samples and yielded an overall sensitivity of 90% and specificity of 95%. However, recombinant ROP2 is a better marker for detection of IgG (91.7%) rather than IgM (80%).

Abstract: This study aims to evaluate the current seroprevalence of toxoplasmosis among indigenous communities in Peninsular Malaysia and relate its association with epidemiological data. Overall seroprevalence of Toxoplasma gondii was 37.0% with 31.0% immunoglobulin (Ig) G, 1.8% IgM, and 4.2% seropositivity for both anti-Toxoplasma antibodies. Multivariate analysis showed that age above 12 years (odds ratio [OR] = 2.70, 95% confidence interval [CI] = 1.75-4.04, P < 0.001), using untreated river and mountain water supplies (OR = 1.50,95% CI = 1.01-2.40, P = 0.050), and close proximity with cats (OR = 1.40, 95% CI = 1.10-1.76, P = 0.010) were factors associated with toxoplasmosis. Given the high seroprevalence of toxoplasmosis among these communities who live in poor socioeconomic conditions, a comprehensive health surveillance program and screening should be initiated among women of childbearing age and pregnant women during the antenatal period for early diagnosis and treatment. The role of domestic cats and environmental contamination with oocyst in soil and water has to be highlighted and addressed in future prevention strategies for these communities.

Abstract: Background: In Malaysia, researchers and medical practitioners are unfamiliar with Naegleria infections. Thus little is known about the existence of pathogenic Naegleria fowleri, and the resultant primary amoebic meningoencephalitis (PAM) is seldom included in the differential diagnosis of central nervous system infections. This study was conducted to detect the presence of Naegleria species in various environmental samples. Methods/Findings: A total of 41 Naegleria-like isolates were isolated from water and dust samples. All these isolates were subjected to PCR using two primer sets designed from the ITS1-ITS2 regions. The N. fowleri species-specific primer set failed to produce the expected amplicon. The Naegleria genus-specific primers produced amplicons of 408 bp (35), 450 bp (2), 457 bp (2) or 381 bp (2) from all 41 isolates isolated from aquatic (33) and dust (8) samples. Analysis of the sequences from 10 representative isolates revealed that amplicons with fragments 408, 450 and 457 bp showed homology with nonpathogenic Naegleria species, and 381 bp showed homology with Vahlkampfia species. These results concurred with the morphological observation that all 39 isolates which exhibited flagella were Naegleria, while 2 isolates (AC7, JN034055 and AC8, JN034056) that did not exhibit flagella were Vahlkampfia species. Conclusion: To date, pathogenic species of N. fowleri have not been isolated from Malaysia. All 39 isolates that produced amplicons (408, 450 and 457 bp) from the genus-specific primers were identified as being similar to nonpathogenic Naegleria. Amplicon 408 bp from 5 representative isolates showed 100% and 99.7% identity to Naegleria philippinensis isolate RJTM (AM167890) and is thus believed to be the most common species in our environment. Amplicons 450 bp and 457 bp were respectively believed to be from 2 new species of Naegleria, since representative isolates showed lower homology and had a longer base pair length when compared to the reference species in the Genbank, Naegleria schusteri (AJ566626) and Naegleria laresi (AJ566630), respectively.

Abstract: Toxoplasma gondii is an important parasite in pregnant women. This case-controlled study assessed the seroprevalence of toxoplasmosis in 640 pregnant women in southern Thailand and identified their associated risk factors. The overall seroprevalence of toxoplasmosis was 181 (28.3%). Of this, 138 (21.6%) were positive for only anti-Toxoplasma immunoglobulin G (IgG) antibody, 43 (6.7%) were positive for both IgG and IgM antibodies, and none were positive for IgM antibody. Multivariate analysis revealed that increasing age (adjusted odds ratio [OR] = 1.64,95% confidence interval [CI] = 1.01-2.67), living outside Songkhla province (adjusted OR = 1.56, 95% CI = 1.08-2.24), parity (adjusted OR = 1.65, 95% CI = 1.01-2.68), contact with cats (adjusted OR = 1.70, 95% CI = 1.20-2.43), and drinking of unclean water (adjusted OR = 1.70, 95% CI = 1.08-2.68) were factors associated with Toxoplasma seroprevalence. On the basis of the results obtained, a health surveillance program should be initiated as a primary preventive measure for congenital toxoplasmosis and focus on educating women of the child-bearing age group to avoid contact with cats and to strictly practice personal hygiene.

Abstract: Toxoplasmosis is an important parasitic disease in immunosuppressed patients. This prospective study was conducted to determine the seroprevalence, associated risk factors and the incidence of clinically confirmed toxoplasmosis among renal patients at the University of Malaya Medical Center, Kuala Lumpur, Malaysia. We interviewed 247 renal patients, each of whom answered an epidemiological questionnaire, and collected blood samples for measurement of anti-Toxoplasma IgG and IgM antibodies by ELISA. Overall seroprevalence of latent toxoplasmosis was observed in 126 (51%) renal patients. Race (Malays), marital status (married) and primary level of education, were all factors associated with a greater chance of Toxoplasma infection. A case of clinically confirned toxoplasmosis was diagnosed in a renal transplant recipient as a result of immunosuppression. Based on the findings obtained, this preliminary study shows a high prevalence of latent toxoplasmosis in renal patients. Risk factors may have significantly contributed to Toxoplasma acquisition in these patients. We recommend further studies be carried out to monitor for trends in toxoplasmosis among immunosuppressed patients.

Abstract: After 30 years of the human immunodeficiency virus (HIV) epidemic, parasites have been one of the most common opportunistic infections (OIs) and one of the most frequent causes of morbidity and mortality associated with HIV-infected patients. Due to severe immunosuppression, enteric parasitic pathogens in general are emerging and are OIs capable of causing diarrhoeal disease associated with HIV. Of these, Cryptosporidium parvum and Isospora belli are the two most common intestinal protozoan parasites and pose a public health problem in acquired immunodeficiency syndrome (AIDS) patients. These are the only two enteric protozoan parasites that remain in the case definition of AIDS till today. Leismaniasis, strongyloidiasis and toxoplasmosis are the three main opportunistic causes of systemic involvements reported in HIV-infected patients. Of these, toxoplasmosis is the most important parasitic infection associated with the central nervous system. Due to its complexity in nature, toxoplasmosis is the only parasitic disease capable of not only causing focal but also disseminated forms and it has been included in AIDS-defining illnesses (ADI) ever since. With the introduction of highly active anti-retroviral therapy (HAART), cryptosporidiosis, leishmaniasis, schistosomiasis, strongyloidiasis, and toxoplasmosis are among parasitic diseases reported in association with immune reconstitution inflammatory syndrome (IRIS). This review addresses various aspects of parasitic infections in term of clinical, diagnostic and therapeutic challenges associated with HIV-infection.

Abstract: This was an eight year (2000-2007) retrospective study of tuberculosis in patients admitted to the UMMC. A total of 131 cases were analyzed. Malays constituted the most cases, (43%), followed by Chinese (22%), Indians (17%) and others (18%). The majority of cases were within the 21-60 year old age group, which constituted 69.5% of the total. Males were more commonly affected (65%). Most cases were reported among Malaysians (83%). The majority of patients were unemployed (39%), followed by housewives (10%), laborers (9%), students (8%), shop assistants (7%), and other occupations (27%) The most common presenting complaints were prolonged productive cough, night sweats, fever, anorexia, weight loss (57%), hemoptysis (34%), and undifferentiated symptoms, such as prolonged diarrhea and dysphagia (9%) Sputum was positive for acid-fast bacilli (AFB) in 89%, but only 69% of cases had abnormal chest radiographs. The majority of patients (65%) developed no complications The most common complications were pleural effusion, pneumothorax and pulmonary fibrosis. The majority of patients (82%) suffered either from diabetes mellitus, hypertension, ischemic heart disease or all 3 conditions Regarding risk factors for tuberculosis, two were HIV positive and two were intravenous drug users. The majority of the patients (85%) did not complain of any side effects from then-anti-tuberculosis treatment. Among those who did complain of side effects, the most common were nausea and vomiting (41%), drug induced hepatitis (37%), blurring of vision (11%) and skin rashes (11%). Two cases of death were reported

Abstract: This study reports the detection of Acanthamoeba and Naegleria species in 14 swimming pools around Petaling Jaya and Kuala Lumpur, Malaysia. Sampling was carried out at 4 sites (the platforms (P), wall (W), 1 meter from the wall (1) and middle (2)) of each swimming pool. These free living amoebae (FLA) were detected under light and inverted microscopes after being cultured on the surface of non-nutrient agar lawned with Escherichia coli. Acanthamoeba species were detected in higher number of culture plates from all sampling sites of all the swimming pools. While Naegleria, were detected in fewer culture plates at 3 sampling sites (absent at site P) of 8 swimming pools. This suggested that the thick double-walled cysts of Acanthamoeba were more resistant, thus remaining viable in the dry-hot areas of the platforms and in chlorinated water of the swimming pools whereas Naegleria cysts, that are fragile and susceptible to desiccation, preferred watery or moist areas for growth and proliferation. The prevalence of both FLA was highest at site W (76.2%), followed by site 1 (64.7%), lowest at site 2 (19.4%), and could be detected at all 3 sampling levels (top, middle and bottom) of these 3 sites. The surface of site W might act as a bio-film that accumulated all kinds of microbes providing sufficient requirement for the FLA to develop and undergo many rounds of life cycles as well as moving from top to bottom in order to graze food. Other factors such as human activities, the circulating system which was fixed at all swimming pools, blowing wind which might carry the cysts from surroundings and the swimming flagellate stage of Naegleria, could also contribute to the distribution of the FLA at these sampling sites. Both FLA showed highest growth (80.4%) at room temperature (25-28 degrees C) and lesser (70.0%) at 37 degrees C which might be due to the overgrowth of other microbes (E. coli, fungi, algae, etc). While at 44 degrees C, only Acanthamoeba species could survive thus showing that our swimming pools are free from potentially pathogenic Naegleria species. However, further study is needed in order to confirm the virulence levels of these amoebae isolates.

Abstract: Loop-mediated isothermal amplification (LAMP), a rapid nucleic acid amplification method, was developed for the clinical diagnosis of toxoplasmosis. Three LAMP assays based on the SAG1, SAG2, and B1 genes of Toxoplasma gondii were developed. The sensitivities and specificities of the LAMP assays were evaluated by comparison with the results of conventional nested PCR. The LAMP assays were highly sensitive and had a detection limit of 0.1 tachyzoite, and no cross-reactivity with the DNA of other parasites was observed. Blood was collected from 105 individuals to test the LAMP assays: 40 patients with active toxoplasmosis, 40 negative controls, and 25 patients with other parasitic infections. The SAG2-based LAMP (SAG2-LAMP) had a greater sensitivity (87.5%) than the SAG1-LAMP (80%), B1-LAMP (80%), and nested PCR (62.5%). All the LAMP assays and nested PCR were 100% specific. This is the first report of a study which applied the LAMP method to diagnose toxoplasmosis from human blood samples. Due to its simplicity, sensitivity, and specificity, LAMP is suggested as an appropriate method for routine diagnosis of active toxoplasmosis in humans.

Abstract: Detection of Toxoplasma gondu in blood by means of the polymerase chain reaction (PCR) may facilitate early diagnosis of toxoplasmosis in different groups of patients We evaluated this approach in 42 patients presenting with ocular or psychotic diseases by comparing the sensitivity and specificity of PCR after heat treatment using a microwave oven with a standard genomic DNA extraction method for paired serum and whole blood samples. The presence of serum IgM and IgG antibodies against T. gondii was detected using a standard commercial enzyme-linked immunosorbent assay and enzyme immunoassay for IgG avidity test. Of 42 whole blood samples, PCR after microwave treatment was positive in 8 samples with a sensitivity of 73% and specificity of 100% compared to 11 samples positive by the extraction method. Although none of 42 sera samples was PCR positive by the extraction method, 7 specimens were positive after microwave treatment This is the first study to use a microwave heat treatment, which is simple, rapid and a promising alternative method, in detecting small amounts of T gondii DNA in human blood. Furthermore, irradiation of blood samples with microwaves allows incorporation of PCR into a practical tool for routine clinical assessment of patients with Toxoplasma infection

Abstract: This is a ten year (1999-2008) retrospective study of amebiasis in patients admitted to UMMC. A total of 34 cases were analyzed. The most: common were amebic liver abscess 22(65%) and the rest were amoebic dysentery 12(35%). Majority of the eases occurred among Malaysians 29(85%), with Chinese 14(41%), followed by the Malays 9(26%) and the Indians 6(18%). Foreigners made up of one Indonesian, one Pakistani and three Myanmarese and constituted 5(15%) of the total cases. Males 24(71%) were more commonly affected. Most of the cases occurred between the age group of 40-49 years, 8(23%) and 60 years and above, 8(23%). Age group of 20-50 years constituted 20(60%) of the cases. The most common clinical presentations were fever with chills and rigors 26(76%), diarrhoea 20 (59%), right hypochondrium pain 17(50%), abdominal pain 17(50%), hepatomegaly 16 (47%) and jaundice 7(20%). All were discharged well after treatment except for one case of death in a 69-year old Chinese male with amebic liver abscess.

Abstract: Toxoplasmosis has historically been considered one of the most important opportunistic infections detected in HIV/AIDS patients The prevalence rates of latent Toxoplasma infections in HIV-infected patients has been found to vary greatly from 3% to 97%, Prevalence has been found to be related to ethnicity, certain risk factors, and reactivation of toxoplasmosis. Prior to antiretroviral therapy, toxoplasmic encephalitis (TE) was the most common focal cerebral lesion detected in AIDS patients with Toxoplasma infection, occurring in approximately half of Toxoplasma-seropositive patients Other forms of dissemination have also been reported in AIDS patients in sites Such as the eves, lungs, heart and spinal cord. Anti-Toxoplasma therapy and chemoprophylaxis have shown effectiveness in reducing the incidence of TE, while noncompliance has been identified as a cause of relapse in these settings. Toxoplasmosis is one of the most common neuropathological complications found at autopsy Rapid progress in the development of highly active. antiretroviral therapy (HAART) has changed the observed patterns with TE, for which there has been a marked decrease in overall incidence Subsequently, TE has been found to be significantly associated with the so-called "neurological immune restoration inflammatory syndrome" (NIRIS). Toxoplasma screening programs are recommended for all newly diagnosed HIV-positive patients. Chemoprophylaxis Should be considered in HIV-infected patients who have a CD4 < 200 cells/mm(3), particularly in settings where resources are limited and there is not access to HAART. TE remains a cause of morbidity and mortality among AIDS patients.

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Abstract: A total of 693 HIV/AIDS patients were recruited in this retrospective and descriptive study during April 2003 to December 2004, Hospital Kuala Lumpur, Malaysia. It was found that both genders were shown to be within the same range of age (M = 18-79 vs F = 18-73), while a median age was slightly higher in male (36 years) than female (31 years). The majority of both genders were significantly shown in the age group of 25 to 34 years, but a higher rate was evident in females (42) (p = 0.001). It was even observed that male patients were mainly Chinese (44.7) and single (57.3) (p = 0.001), while the females were Chinese (40.6) and married (83.3) (p = 0.000). However, the highest numbers of both genders were manifested to be heterosexuals (M = 47.6 vs F = 86.2), followed by intravenous drug users (M = 39.3 vs F = 6.5) (p = 0.000). The range of CD4 cell count was 0 to 1799 with a median of 230 cells/mm3. The level of CD4 cell count of < 200 cells/mm3 was significantly found in males (48.5) and between 200 to 499 and �500 cells/mm3 were 36.2 and 26.8 in females, respectively (p = 0.001). Overall, the Toxoplasma seroprevalence was 43.85 where seropositive of anti-Toxoplasma antibody relatively higher in males (50) than in females (37.7) (p = 0.015). Toxoplasma seropositivity was thoroughly evident among Malays (168; 51.1), Intravenous Drug Users (IDUs) and patients with toxoplasmic encephalitis (TE) (p = 0.001). Furthermore, anti-retroviral therapy including HAART was more significantly found in patients with 9 seropositive Toxoplasma when compared to 2 seronegative and unknown serostatus patients (p = 0.025). Seventeen AIDS-related toxoplasmic encephalitis patients were diagnosed at the time of this study, depicting hemiparesis as the most common neurological manifestation in 11 (64.7) patients, followed by headache and seizure in 6 (35.3) and 3 (17.7) patients, respectively. Interestingly, CT scan finding showed mass like structure with multiple (58.8), ring enhancing lesions (100), in parietal region (58.8) and edema (29.4) in these patients. Overall, the treatment outcome showed that 13 (76.5) patients had completed treatment with maintenance, whereas, 3 (17.6) patients were lost to follow up and 1 (6) patient was transferred to another hospital. No relapse or death case was reported during the time of this study.

Abstract: A total of 255 patients including 179 (70.2) of non-HIV and 76 (29.8) HIV-infected patients were recruited in this descriptive study. The subjects was significantly found to be male Chinese (157; 61.6 vs 74; 47.1) followed by female Malays (98; 38.4 vs 35; 35.7) (p<0.05). The majority of subjects (124; 48.6) were in the age group of 21-39 years, however, no statistical difference was found between the various age groups (p>0.05). Overall seroprevalence of latent Toxoplasma infection was 82/183 (44.8) being; 3 (3.7) positive for IgM, 74 (90.2) for IgG, and 5 (6.1) for IgG and IgM antibodies. The prevalence was more relatively found in the Chinese (28; 15.3) and Malays (27; 14.8) than others (p<0.05). While, 23/76 (30.3) of HIV-positive patients were shown Toxoplasma seropositivity. The majority of these subjects (138/181; 76.2) were significantly asymptomatic (p = 0.000), while the others were clinically evident cases of toxoplasmosis. Of this, 37 patients were included in differential diagnosis relating to ocular diseases and only 4 patients were confirmed as having ocular toxoplasmosis. Toxoplasmic encephalitis (TE) was based on presumptive diagnosis, particularly found in 5 patients with AIDS. Seventeen patients were clinically diagnosed as having malaria being; 8 for P. vivax, 4 for P. falciparum, 3 for P. malariae, and 2 for mixed infections. All cases resolved satisfactorily after treatment with antimalarial drugs. Other important emerging parasitic diseases were also detected in these patients including amebiasis (2), blastocystosis (1), cryptosporidiosis (1), filariasis (1), and giardiasis (2) during the time of this study.

Abstract: This retrospective study was carried out to determine the prevalence of malaria among patients admitted to Hospital Tengku Ampuan Rahimah, Klang, Malaysia, from January 2004 to May 2006. A total of 37 malaria cases were analyzed. Most cases occurred among foreigners, 81% (30 cases), while Malaysians constituted 19% (7 cases). Among foreigners, Indians constituted the majority, 40.5% (15 cases). Among Malaysians, most cases occurred among Malays, 16% (6 cases), followed by Indians, 3% (1 case). Males, 89% (33 cases), were more commonly affected. The majority of cases were within the 20-39 year age group (84%). Most cases occurred among laborers (24%). Two species of malaria parasites were reported, of which Plasmodium vivax constituted the most, 70% (26 cases), followed by Plasmodium falciparum, 30% (11 cases). In this study, 40.5% (15 cases) developed chloroquine resistance: six cases of P. falciparum, and nine cases of P. vivax. The most common complications were thrombocytopenia, 65% (24 cases), and anemia, 54% (20 cases), followed by jaundice, 32% (12 cases), and hepatosplenomegaly, 22% (8 cases). There were no reported deaths. This new source of malaria coming from foreigners must be given serious attention, as it has great potential of increasing malaria cases in urban Malaysia.

Abstract: Attempts to address the significant impact of HAART on medical variables on the Malaysian HIV/AIDS population have yet to be evaluated. This study aims to analyze the proportions of AIDS-defining illnesses (ADIs) before and after HAART. A retrospective study was carried out on 128 new cases of HIV infected patients who first commenced HAART in 2004 at the national HIV reference center. Before commencement of HAART, 76 clinical episodes of ADIs were recorded in 52 patients. Most common being pulmonary Mycobacterium tuberculosis (28.9%), PCP (27.6%) and disseminated and extrapulmonary Mycobacterium tuberculosis (11.8%). During HAART, 8 clinical episodes of ADIs were documented in 7 patients with a median time of onset of 10 weeks after initiation of HAART (range, 4-36 weeks). The median CD4 count at the time of the commencement of HAART for these patients was 11 cells/mm(3). ADIs reported include PCP (2 episodes), disseminated and extrapulmonary Mycobacterium tuberculosis (2 episodes), extrapulmonary cryptococcosis (1 episode), esophageal candidiasis (1 episode), recurrent pneumonia (1 episode) and disseminated or extrapulmonary histoplasmosis (1 episode). Three (37.5%) of these occurred despite a reduction of viral load by at least 2 log(10) and an increased in the CD4 cell count. In conclusion, ADIs can still present after the initiation of successful HAART especially in those with CD4 counts below 100 cells/mm(3). In Malaysia, ADIs are the major causes of HIV/AIDS associated morbidity and mortality, thus increased awareness on the management of these illnesses is warranted especially in the months following HAART.

Abstract: Cryptosporidium and Giardia are two important pathogenic parasites that have caused many waterborne outbreaks which affected hundreds of thousands of people. Contamination from effluent discharged by sewage treatment plants have been implicated in previous waterborne outbreaks of Cryptosporidium and Giardia. This study evaluated the reduction of Cryptospotidium and Giardia (oo)cysts in two sewage treatment plants (STPA and STPB) in Malaysia which employed different treatment processes for a period of a year. Raw sewage influents and treated sewage effluents were concentrated by repeated centrifugation, subjected to sucrose density flotation and concentrated to a minimal volume depending upon the levels of contaminating debris. Cryptosporidium oocysts and Giardia cysts were enumerated using epifluorescence microscopy. The parasite concentrations in raw sewage were 18-8480 of Giardia cysts/litre and 1-80 of Cryptosporidium oocysts/litre. In treated sewage, the concentration of parasites ranged from 1-1462 cysts/litre and 20-80 oocysts/ litre for Giardia and Cryptosporidium respectively. Statistical analysis showed that sewage treatment process which employed extended aeration could reduce the concentration of Cryptosporidium and Giardia (oo)cysts significantly but treatment process which encompasses aerated lagoon could only reduce the concentration of Giardia cysts but not Cryptosporidium oocysts significantly. This phenomenon is of great concern in areas whereby effluent of sewage treatment plants is discharged into the upstream of rivers that are eventually used for abstraction of drinking water. Therefore, it is important that wastewater treatment authorities rethink the relevance of Cryptosporidium and Giardia contamination levels in wastewater and watersheds and to develop countermeasures in wastewater treatment plants. Further epidemiological studies on the occurrence and removal of pathogenic organisms from excreta and sewage are also recommended, in order that the public health risks can be defined and the most cost effective sewage treatment options developed.

Abstract: The substantial influx of foreign-born persons including immigrant population into the community becomes one of the postulated reasons to be elucidated relating to this so-called �a disease without border� in Malaysia. A total of 425 TB patients, including Malaysians and foreigners, were treated at the Institute of Respiratory Medicine from May to December 2003. TB was found relatively more often in foreign laborers and Malaysians with nonspecific occupations. Tuberculin skin test (TST) was used to screen for latent TB infection and a higher positive rate of TST was found in foreign compared to local patients. Chest X-ray findings showed a higher rate of abnormalities consistent with PTB, found in the majority of both groups. Lymph node biopsy and sputum culture were used significantly to detect the presence of M. tuberculosis and confirm the diagnosis of TB. EHRZ+B6 was significantly the most commonly used anti-tubercular drug regimen, found in both local and foreign patients. Foreign patients were more significantly associated with non-compliance to anti-tubercular therapy. Hepatitis was one of the most common adverse drug reactions found in local patients. The presence of a greater number of illegal migrants, who are highly mobile within the country as well as across borders, and who do not undergo any health screening, further complicates the national tuberculosis control program in the future.

Abstract: A total of 136 patients, 67 HIV, 69 diabetes mellitus (DM) with or without (+/-) end-stage renal disease (ESRD), were registered for tuberculosis treatment at the National Tuberculosis Center (NTBC) from May to December, 2003. Ages ranged from 21-78 years (median 57.7 years) in TB/DM patients, and 21-62 (mean 37.6 +/- 8.3 years) in TB/HIV patients. TB was significantly found in younger and single HIV patients, but in older and married DM patients (p<0.05). Male patients in both groups were strongly associated with TB, while females more commonly had TB with DM (p<0.05). The majority of these patients were Malays, unemployed, and resided in Kuala Lumpur territory; however, no statistically significant difference was found between the 2 groups. Smoking, IVDUs and hepatitis C virus (HCV) infection were more significantly found in TB/HIV patients and further analysis showed that pulmonary TB was strongly associated with HCV infection in these patients (p<0.05). Pulmonary TB (62; 89.9) was the most common type found in both groups and was a markedly more common disease location in TB/DM patients, while extrapulmonary TB (21; 31.3) and miliary TB (14; 21) were significantly higher in TB/HIV patients. Cough with or without sputum, fever and loss of appetite and/or weight were common clinical presentations in both groups. Nevertheless, fever (54; 80.6) and lymphadenopathy (17; 25.4) were significantly related to TB/HIV patients (p<0.05). Interestingly, the presence of BCG vaccination and positive tuberculin skin test were stronger in TB/HIV (27; 40.3) and TB/DM (20; 29) patients, respectively (p<0.05). Overall, regular 6-, 9- and 12-months' anti-tubercular therapy (ATT) were routine practice, and EHRZ+B6 was the most common regimen used. The highest percentage of patients with treatment success were in both groups with 6 months' ATT; however, a significantly higher percentage was found in TB/DM (24; 34.8) than TB/HIV (13; 19.4) (p<0.05). A success rate of 15 (21.7) was noted for TB/DM patients with 9 months' ATT, which was similar to both groups with the 12-month regimen. A higher percentage failure rate (lost to follow-up) was seen in TB/HIV (19; 28.4) patients. Nine patients were reported to have anti-tubercular-drug side-effects, such as drug-induced hepatitis, blurred vision, and skin rash. No cases of drug resistance or death were notified among these patients.

Abstract:

Abstract: A total of 136 patients, 67 HIV, 69 diabetes mellitus (DM) with or without (±) end-stage renal disease (ESRD), were registered for tuberculosis treatment at the National Tuberculosis Center (NTBC) from May to December, 2003. Ages ranged from 21-78 years (median 57.7 years) in TB/DM patients, and 21-62 (mean 37.6 ± 8.3 years) in TB/HIV patients. TB was significantly found in younger and single HIV patients, but in older and married DM patients (p<0.05). Male patients in both groups were strongly associated with TB, while females more commonly had TB with DM (p<0.05). The majority of these patients were Malays, unemployed, and resided in Kuala Lumpur territory; however, no statistically significant difference was found between the 2 groups. Smoking, IVDUs and hepatitis C virus (HCV) infection were more significantly found in TB/HIV patients and further analysis showed that pulmonary TB was strongly associated with HCV infection in these patients (p<0.05). Pulmonary TB (62; 89.9%) was the most common type found in both groups and was a markedly more common disease location in TB/DM patients, while extrapulmonary TB (21; 31.3%) and miliary TB (14; 21%) were significantly higher in TB/HIV patients. Cough with or without sputum, fever and loss of appetite and/or weight were common clinical presentations in both groups. Nevertheless, fever (54; 80.6%) and lymphadenopathy (17; 25.4%) were significantly related to TB/HIV patients (p<0.05). Interestingly, the presence of BCG vaccination and positive tuberculin skin test were stronger in TB/HIV (27; 40.3%) and TB/DM (20; 29%) patients, respectively (p<0.05). Overall, regular 6-, 9- and 12-months� anti-tubercular therapy (ATT) were routine practice, and EHRZ+B6 was the most common regimen used. The highest percentage of patients with treatment success were in both groups with 6 months� ATT; however, a significantly higher percentage was found in TB/DM (24; 34.8%) than TB/HIV (13; 19.4%) (p<0.05). A success rate of 15 (21.7%) was noted for TB/DM patients with 9 months� ATT, which was similar to both groups with the 12-month regimen. A higher percentage failure rate (lost to follow-up) was seen in TB/HIV (19; 28.4%) patients. Nine patients were reported to have anti-tubercular-drug side-effects, such as drug-induced hepatitis, blurred vision, and skin rash. No cases of drug resistance or death were notified among these patients.

Abstract: Diphyllobothriasis was first reported in Malaysia in 2002. We are reporting a second case of diphyllobothriasis in Malaysia. The patient was a 37-year-old Chinese male seen at the outpatient clinic with a complaint of passing strands of white color flat worm in his stool. He had no other abdominal complaints. Laboratory and physical examinations were normal. Diphyllobothrium latum was confirmed by examination of the gravid proglottids passed out and the typical operculated eggs expelled from the ruptured proglottids. The patient had a history of eating raw fish. He was treated with a single dose of praziquantel.

Abstract: . In this retrospective study, we investigated 263 foreign patients who were diagnosed as having tuberculosis at the National Tuberculosis Center (NTBC) from January 2001 to December 2002. The age range was 14-72 years, with a mean of 33.3 ± 9.95 years. The study subjects were predominantly males (60%) and females comprised 40%, where the greater impact of tuberculosis was observed in the young and active ones (up to 34 years of age), than middle-age (up to 54). A significantly higher percentage of these patients were from the Southeast Asian countries (87%) and particularly occurred in single male (47.5%) and married female (71.4%) patients (p<0.05). We also found that tuberculosis was significantly higher in female (50.5%) and male (64%) with smoking laborers (p<0.05). Fever (70%), cough (90.5%) and BCG vaccination status showed a significantly higher percentage in male patients (p<0.05), whereas lymphadenopathy (22%) was found in a significantly higher percentage in females (p<0.05). Overall, pulmonary disease (94.3%) occurred more commonly in males and the pleura (3.2%) was the most common site of disseminated tuberculosis. By contrast, the lymph node (11.4%) and miliary (4.8%) forms were the more common extrapulmonary tuberculosis in females. More males had higher percentage of treatment completed at � 6 (38%) and � 9 (13.3%) months in pulmonary tuberculosis, whilst, more females showed higher percentage of treatment completed (8.7%) in extrapulmonary tuberculosis. Surprisingly, more women showed noncompliance to the anti-tubercular therapy than their counterpart in this study

Abstract: This retrospective study was conducted at the National Tuberculosis Center (NTBC) where 252 HIV-positive patients coexisting with tuberculosis (TB/HIV) were examined. We found that patients with pulmonary (PTB) and extrapulmonary tuberculosis (EPT) had similar mean age. A higher sex ratio between male to female (10.7:1) was observed in patients with PTB. The other characteristics of patients with pulmonary and extrapulmonary tuberculosis were not statistically different from each other. Cough (88%) and hemoptysis were the most common presenting symptoms, significantly related to patients with PTB. Lymphadenopathy (33.5%) was the most common sign in patients with EPT. The majority of patients with pulmonary and extrapulmonary tuberculosis had CD4 cell counts of less than 200 cells/mm3 (range 0-1,179 with a median of 57 cells/mm3). Lung (89%) and miliary (55.6%) forms were the most frequent disease locations in patients with PTB and EPT, respectively. A higher percentage of patients with PTB (42%) were treated successfully with short-course (6 months) therapy, whereas in patients with EPT (43%) needed a longer period (9 months) for successful treatment. Of the patients who defaulted treatment, a higher proportion (87%) had PTB. No MDR-TB or relapse cases were found in this study.

Abstract: During a 2-year retrospective study, 195 non-HIV-infected patients with extrapulmonary tuberculosis (EPT) were diagnosed at the National Tuberculosis Center, Kuala Lumpur, representing 10 of all patients with tuberculosis. Their mean age was 39 (�±SD) 14 years old (range 14-81). The largest age group was 25-34 years, while 78.5 were less than 50 years of age. The subjects were mainly female (50.3), Malay (49.2), married (61.5), resided in Kuala Lumpur (51.0), and were unemployed (50.3). Regarding risk factors, they were smokers and/or alcohol users (21.0), and injecting drug users (IDUs) (5.1); they also had history of tuberculosis (3.6) and contact with TB patients (9.2). Lymphadenopathy was the most common sign (45.6) shown in the medical records. 42 of x-ray findings (chest, spine, and hip) showed signs consistent with tuberculosis, while histopathology was the most useful diagnostic tool (52.3) and lymph node was the most frequent specimen used (35.0) in this study. The three main sites of involvement were lymph nodes (42.6), miliary and disseminated (19.5), and pleura (12.8). The outcome of this study showed 72.8 of these patients had completed treatment for at least 6 months, whilst, only 4.6 of patients were still undergoing treatment, and unfortunately, 22.6 of them showed non-adherence to anti-tubercular therapy at a duration of less than 6 months. However, no MDR-TB or death cases were reported or registered in this study.

Abstract: The seroprevalence of toxoplasmosis among 505 of human immunodeficiency virus (HIV)/AIDS patients was 226 (44.8%; 95% CI 42.64-51.76): 27 (47.4%) and 199 (44.4%) showed Toxoplasma seropositivity with and without toxoplasmic encephalitis (TE), respectively (P<0.05). The majority of these patients were in the 25-34 age group (44 versus 39%), male (86 versus 76%), and Chinese (49 versus 53%), though no statistical significance was found between the two. Significant differences between these two groups were noted, however, in terms of marital status, occupation, and present address. The heterosexual exhibited the most frequent behavior at risk for HIV infection, and accounted for 51 and 59% of patients with and without TE, respectively. Only 17/260 (6.5%) and 1/137 (0.7%) of them later acquired TE after receiving primary chemoprophylaxis (cotrimoxazole) and antiretroviral therapy including HAART (P<0.05). Fifty-seven (11.3%) out of those 505 patients were diagnosed with AIDS-related TE. The most common clinical manifestation was headache (56%). The computed tomography scan findings showed most lesions to be multiple (96.4%), hypodense (66.7%), and in the parietal region (39.3%). Twenty-seven (47.4%) patients had chronic (latent) Toxoplasma infection as evidenced by seropositivity for anti-Toxoplasma (IgG) antibody. At the time of diagnosis, the range of CD4 cell count was from 0-239 with a median of 25 cells/cumm. We also found that a CD4 count of less than 100 cells/cumm was significantly associated with development of TE (P<0.05). Clinical outcomes showed that among those who survived, 21 (36.8%), 16 (28.1%), and 2 (3.5%) of patients had completed treatment, transferred out, and were lost to follow up, respectively. Unfortunately, 18 (31.6%) of the cases were officially pronounced dead. Overall, 7 (12.3%) patients were detected as recurrent TE in this study.

Abstract: Three hundred and one sera of HIV/AIDS patients were tested for anti-Toxoplasma IgG antibody by ELISA technique. The seroprevalence of toxoplasmosis was 41.2% (95%CI: 35.5-46.9) in HIV/AIDS patients. The seroprevalence was significantly higher in the Malay (57.9%) than the Chinese (38.7%), followed by the Indian patients (29.6%) (p<0.05). No possible risk factor, such as contact with cats, consumption of uncooked meat, and history of blood transfusions was found to have any significant association with the presence of anti-Toxoplasma antibody in the study sample (p > 0.05). Multivariate analysis was employed to find any association between Toxoplasma seroprevalence and a single subject having single or multiple risk factors. It was found that the association was not statistically significant (p>0.05). Among the HIV/AIDS study samples, 124 (41.2%) samples were found to have positive anti-Toxoplasma antibody, the association between the presence of anti-Toxoplasma antibody and CD4 cell count was determined but no statistically significant association was found (p>0.05). During the study period, only one case of active CNS toxoplasmosis was registered and the diagnostic criteria included: clinical presentations, CT scan finding, serological evidence of anti-Toxoplasma IgG antibody, and respose to anti-Toxoplasma therapy.

Abstract: We retrospectively reviewed 419 HIV/AIDS patients in Hospital Kuala Lumpur from 1994 to 2001. In the male group, the age range was 20-74, with a mean age 3 7 years, while in the female group it was 17-63, with a mean age of 33 years. With regard to age group, it was found that the preponderant age group was 25-34 years. The majority of male subjects were Chinese (52.5%), single (56.3%), and unemployed (55.1%), whereas the females were Malay (42.3%), married (79.5%), and non-laborer (64.1%). Also, both groups resided in Kuala Lumpur and had heterosexual contact as the leading cause of HIV transmission. More than half of the patients had CD4 cell counts of <200 cells/cumm. We found that the acquisition of HIV infection via intravenous drug use (IDU) was directly related to the incidence of tuberculosis infection (P < 0.05). Further analysis showed HIV-related tuberculosis with IDU was also dependently correlated with occupational status (unemployed) (P < 0.05). The four main AIDS-defining diseases include tuberculosis (48%), Pneumocystis carinii pneumonia (13%), toxoplasmic encephalitis (11%), and cryptococcal meningitis (7%); in addition, 53% of these patients were found to have CD4 cell counts of less than 200 cells/cumm at the time of diagnosis.

Abstract: Four hundred and six AIDS patients were recruited in this retrospective study. The seroprevalence of toxoplasmosis among 406 AIDS patients was 208 (51.2%). Their age ranged from 17 to 74 years with a median of 35 years. The majority of patients were males 172 (82.6%), Malays 99 (47.5%), single 109 (52.4%), unemployed 99 (47.6%) and heterosexual with commercial sex workers (CSW) 97 (46.6%) as the risk marker to HIV infection. Thirty-one (14.9%) of 208 AIDS-related toxoplasmosis were diagnosed as active toxoplasmic encephalitis. The most common clinical manifestation was headache (67.7%). The CT scan findings showed most lesions to be multiple (87.5%), hypodense (66.7%), and in frontal region (41.7%). Twenty-two (71%) patients had chronic (latent) Toxoplasma infection as evidenced by seropositivity for anti-Toxoplasma (IgG) antibody. They were statistically significant in the association between CD4 count and toxoplasmic encephalitis (P = 0.019; OR = 2.6; 95% CI = 1.14-6.02). After the initial six weeks of anti-TE therapy, relapsing toxoplasmic encephalitis was detected in 9.7% in this study.

Abstract: The gene encoding the excretory-secretory antigen TES-120 of dog ascarid worm Toxocara canis was cloned into the bacterium Escherichia coli. The specificity of the recombinant TES- 120 antigen produced by the bacterium was investigated.A total of 45 human serum samples from patients infected with different helminthes and protozoa, including 8 cases of toxocariasis, were tested against the recombinant antigens in immunoblot assays. The results from the assays revealed that the recombinant TES-120 antigen reacted with sera from toxocariasis patients only. This highly specific recombinant TES-120 antigen can potentially be used for the development of an inexpensive serodiagnostic assay for human toxocariasis.

Abstract: The objective of this study was to characterize the polypeptides associated with cysts of Blastocystis hominis. This form is believed to be infective and plays a role in parasite resistance to anti-B. hominis drugs currently used for treatment of Blastocystis associated diarrhea. Cysts were induced through in vitro culture of the parasite in complete medium supplemented with bacterial extract with trypticase, metronidazole or doxycycline. SDS-PAGE analysis showed almost similar polypeptide patterns of parasite extracts obtained from in vitro cultured parasites before and after exposure with the three supplements. Polypeptide bands at 76, 58.5, 48, 45, 40, 38, 32, 25 and 22 kDa were constantly seen in all antigenic preparations and no specific cyst-associated polypeptide was present. However, on immunoblot analysis, 3 out of 16 blastocystosis human sera identified a cyst-associated polypeptide at 60 kDa in all parasite extracts prepared from cultures with the three supplements. In addition, there were associated morphological changes detected in these parasites stained with acridine orange and observed under fluorescence microscopy. Metronidazole induced cyst forms (reddish cells) as early as 12 hours post-exposure; more cyst production (with stronger immunoblot bands) occurred after 24 hours exposure. However, cysts rupture with release and destruction of B. hominis daughters cells occurred after 48 hours exposure. Doxycycline induced less cyst-like forms at 24 hours (weaker 60 kDa band) and less destruction of the cysts (60 kDa band still present at 72 hours post exposure). Bacterial extract and trypticase also induced cysts at 12 hours with increasing numbers up to 72 hours exposure (corresponding increase in intensity of 60 kDa band from samples harvested at 12 to 72 hours post exposure) without any sign of deleterious effect on the parasite.