Malaysia is a non-endemic country for hepatitis E virus (HEV) infection. However, seroprevalence as high as 50% among samples of aboriginal people were reported over two decades ago. A total of 207 samples collected from seven aboriginal villages in rural settlements across two states in Malaysia were analysed for anti-HEV IgG and IgM by an enzyme-linked immunoassay. Following the detection of anti-HEV seroprevalence, we organized health outreach to inform and educate the community. Qualitative interviews were conducted with individuals tested positive for anti-HEV antibodies. Data derived from interviews and observations were used to investigate possible lifestyle behaviours associated with HEV infection. Anti-HEV IgG was detected in six samples (5.9%) from the village of Dusun Kubur. Qualitative inquiry and observation study revealed poor dietary and household hygiene, contaminated food and water, contact with animal faeces, unsanitary and domestic waste disposal, and wildlife reservoirs could be the contributing factors for transmission and acquisition of HEV infection. Investigation during health outreach is important to provide insights for future empirical research and implementation for improvement of lifestyle behaviours among the aborigines. Managing the risk of HEV infection in the aborigines may reduce the risk of HEV transmission to the local communities.
An accurate diagnosis for toxoplasmosis is crucial for pregnant women as this infection may lead to severe sequelae in the fetus. The value of IgG avidity assay as a tool to determine acute and chronic toxoplasmosis during pregnancy was evaluated in Universiti Kebangsaan Malaysia Medical Centre (UKMMC). In this study, 281 serum samples from 281 pregnant women in various trimesters were collected. These samples were assayed using specific anti-Toxoplasma IgM and IgG antibodies, followed by IgG avidity test. The overall seroprevalence of toxoplasmosis in pregnant women was 35.2% (33.5% for anti-Toxoplasma IgG and 1.8% for both anti-Toxoplasma IgG and IgM antibodies). Of 5 (1.8%) serum samples positive for IgM ELISA, 4 had high-avidity antibodies, suggesting past infection and one sample with borderline avidity index. Two samples with low avidity were from IgM negative serum samples. The IgG avidity assay exhibited an excellent specificity of 97.6% and a negative predictive value (NPV) of 95.6%. The study also demonstrated no significant correlation between avidity indexes of the sera with IgG (r=0.12, p=0.24) and IgM (r=-0.00, p=0.98), suggesting the complementary needs of the two tests for a better diagnosis outcome. These findings highlight the usefulness of IgG avidity assay in excluding a recently acquired toxoplasmosis infection in IgM-positive serum sample.
In this study we have cloned unreported gene fragments of Toxoplasma gondii GRA7 and SAG1 and expressed the corresponding recombinant proteins, followed by evaluation of their usefulness for the serological diagnosis of toxoplasmosis. Both recombinant proteins were expressed efficiently in insoluble form, purified by single step Ni-NTA affinity chromatography and their antigenicity to detect toxoplasma specific IgG antibodies were determined by immunoblotting. A total of 60 serum samples from three groups of individuals based on their anti-toxoplasma antibody profiles were tested, namely (I) IgM+, IgG+ (n=20), (II) IgM-, IgG+ (n=20) and (III) IgM-, IgG- (n=20). Both recombinant proteins exhibited high sensitivity (100%) with sera from Group I. rGRA7 and rSAG1 reacted 40% and 80% respectively with Group II sera. The specificity of the recombinant proteins based on reactivities with Group III sera were 100% and 80% with rGRA7 and rSAG1 respectively. Thus rGRA7 was found to be better at discriminating probable acute from chronic phases of toxoplasmosis, and it also showed higher specificity.
Toxoplasmosis can cause serious disease in immunocompromised patients and to congenitally infected foetuses. Appropriate laboratory investigations in potential cases of acute Toxoplasma infection are important. Excretory secretory antigen (ESA) is immunogenic during both human and experimental infections, therefore is considered as a good candidate for investigation into new infection markers. In this study, ESA was prepared from in vitro cultures of Toxoplasma gondii to identify T. gondii ESA antigenic component(s) that is/are most reactive with serum samples from probable acute cases of toxoplasmosis. Serum samples were obtained from several categories of individuals with the following Toxoplasma serology: Group I: IgM+ IgG+ (low IgG avidity) or IgM+ IgG- from sera of patients who had clinical query of toxoplasmosis (n=35). Group II: IgM- IgG+ (high IgG avidity) from chronically infected individuals (n=30). Group III: normal/healthy individuals with anti-Toxoplasma IgMIgG- (n=20). Group IV: individuals with other infections who had anti-Toxoplasma IgM- IgG- (n=10). The ESA was subjected to SDS-PAGE, followed by Western blot analysis using the above sera and probed with peroxidase conjugated anti-human IgM and IgA antibodies. The blots were then developed using chemiluminescence substrate. The selected antigenic band was excised from the gel after two dimensional electrophoresis and sent for mass spectrometry analysis using MALDI TOF-TOF. The most promising antigenic band was a 10 kDa protein which showed sensitivity of 80% in both IgM and IgA blots, and specificity of 96.7% with sera from other infections and healthy controls. The two best identifications for the 10 kDa band were ubiquitin (ribosomal protein CEP52 fusion protein) and polyubiquitin.
There is a need for identification of new infection markers against common Leptospira isolates in Malaysia. To achieve this goal, seven-day-old cultures of Leptospira interrogans serogroup Icterohemorrhagiae (L44) and Leptospira interrogans serogroup Javanica (L55) were used for antigen preparation by sequential extraction method using 40 mM Tris, 8M Urea and 2M thiourea. Immunoblot analysis of the antigens were performed using serum samples from 46 local patients with confirmed acute leptospirosis, 28 patients with other infections and 14 healthy controls. The patients serum samples used in this study contained heterologous antibody against a number of different leptospira serovars. A strong IgM reactivity to a broad diffuse band of 10-15 kDa was observed. Combining results using L44 and L55 antigens showed sensitivity of 80.4% and specificity of 95.2% for detection of leptospirosis. Proteinase K and periodate treatment indicated that the band is likely to be lipopolysaccharide (LPS) in nature. This study showed that the 10-15 kDa antigen could potentially be useful for serodiagnosis of acute leptospirosis in Malaysia.
The detection of leptospires in patient blood in the first week of the disease using PCR provides an early diagnostic tool. PCR using two sets of primers (G1/G2 and B64-I/B64-II) tested with samples seeded with 23 leptospiral strains from pathogenic and non-pathogenic strains was able to amplify leptospiral DNA from pathogenic strains only. Of the 39 antibody negative samples collected from patients suspected for leptospirosis, only 1 sample (2.6%) was PCR positive. Using LSSP-PCR, the G2 primers allowed the characterization of Leptopira species to 10 different genetic signatures which may have epidemiological value in determining species involved in outbreaks. Leptospiral outer membrane proteins from three strains were purified and reacted against patients sera and gave rise to different profiles for pathogenic and non-pathogenic strains. Lymphocytes of mice injected with OMPs proliferated and released IFN(-3) when stimulated in vitro using Leptospira OMP as antigens. This suggests that an immune response could be established using leptospiral OMPs as a putative vaccine. OMPs were also used in a Dot-ELISA to detect antibodies against Leptospira pathogens in humans.
Toxoplasma gondii is a protozoan parasite that is capable of causing a zoonotic disease, known as toxoplasmosis. Vertical transmission of T. gondii from the mother to the fetus, during pregnancy may cause severe complications to the developing fetus. This current study aimed to determine the seroprevalence and investigate the associated risk factors of Toxoplasma infection in pregnant women (n=219) visiting the antenatal clinic at UMMC. While the elevated level of anti-Toxoplasma IgG and IgM antibodies indicates the presence of infection, it fails to differentiate between a past and a recent infection. Thus, the study also demonstrates the usefulness of IgG avidity in validating the timing of infection. The serum samples were tested for the presence of anti-Toxoplasma IgG and IgM antibodies by ELISA test, and the seropositive samples for both anti-Toxoplasma IgG and IgM antibodies were further evaluated by IgG avidity. The results showed that the overall prevalence of T. gondii seropositivity was 34.7%. Of these, 30.6% (67/219) were positive for anti-Toxoplasma IgG antibody only, 2.3% (5/219) were positive for anti-Toxoplasma IgM only, and the remaining 1.8% (4/219) was positive for both anti-Toxoplasma IgG and IgM antibodies. All of the pregnant women who were positive for both anti-Toxoplasma IgG and IgM antibody were found to have past infection when evaluated by IgG avidity. In this study, Malay ethnicity and the number of existing previous children were significantly associated with T. gondii seropositivity (p<0.05). Based on these findings, information and education on the transmission and prevention of congenital toxoplasmosis are very crucial as a public health effort towards a healthier society.
A study was undertaken to evaluate the relevance of detecting IgM and IgG antibodies in diagnosis of canine leptospirosis in Kerala, a southern state of India, which is endemic for the disease. A total of 205 blood (35 from healthy vaccinated, 30 from healthy unvaccinated and 140 from diseased dogs) and 151 urine samples (11 from healthy vaccinated and 140 from diseased dogs) were collected from three districts of Kerala, Thrissur, Palakkad and Kozhikode with high incidence of leptospirosis. Recombinant LipL41 protein was used as antigen and IgG and IgM based ELISAs were standardized. The results were compared with the gold standard test, microscopic agglutination test (MAT). The MAT positive samples (146 samples) were divided into those having titre >1:800 and those between 1:100 and 1:400 in view that the former constituted the acute cases. It was found that IgM ELISA was more specific and sensitive in detecting acute cases (MAT >1:800) whereas IgG ELISA was less specific. In case of seroprevalence studies (MAT titre 1:100 to 1: 400), IgG ELISA was found to be more sensitive and specific than IgM ELISA. Receiver operating characteristic curves when plotted, revealed the accuracy of IgM ELISA in acute leptospirosis. Many samples were positive for both IgG and IgM antibodies. Polymerase Chain Reaction (PCR) targeting lipl41 gene was standardized and urine and blood samples from the same dogs were tested. PCR was found to be the specific test for the early detection of leptospires in blood even before seroconversion. However, PCR analysis of the urine samples was found to be insensitive. Hence, it can be concluded that the diagnostic strategies should be modified, and a combination of serological and molecular tests is recommended in endemic areas rather than simple detection of IgM or IgG antibodies, for the early detection of acute clinical cases of leptospirosis.
Leptospirosis is a worldwide zoonotic disease caused by spirochetes of the genus Leptospira. The clinical manifestation of leptospirosis is non-specific and frequently misdiagnosed as other illnesses. The aim of this study was to compare the diagnostic accuracies of two commercial tests for early diagnosis of Leptospira species: the IgM latex agglutination test (IgM LAT) and the IgM enzyme-linked immunosorbent assay (IgM ELISA). A total of 140 serum samples were obtained from patients suspected of leptospirosis at the Universiti Kebangsaan Malaysia Medical Centre (UKMMC). These serum samples were tested for the presence of Leptospira sp. using IgM LAT, IgM ELISA and MAT. From Table 1, IgM LAT showed 21% (n = 29) positive, 18% (n = 25) inconclusive and 61% (n = 86) negative, while IgM ELISA showed 6% (n = 8) positive, 6% (n = 8) inconclusive, 88% (n = 124) negative and MAT showed 11% (n = 16) positive, 47% (n = 65) inconclusive, 42% (n = 59) negative. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of IgM LAT were 68.8%, 57.6%, 30.6% and 87.2% respectively, while for IgM ELISA they were 37.5%, 89.8%, 50% and 84.1%, respectively as compared to MAT (Table 2). The results showed that IgM LAT had higher sensitivity but lower specificity compared to IgM ELISA. In conclusion, IgM LAT can be useful as an early screening test for early diagnosis of Leptospira sp., while IgM ELISA is a suitable method for reducing false negative detection of Leptospira sp. As both tests show moderate percentages (~65%) in accuracy, an additional test is required for better detection of Leptospira sp.
Toxoplasma gondii (T. gondii) is a zoonotic infection that may be transmitted to human beings either by consumption of raw or uncooked meat or by ingesting oocysts. Toxoplasma organisms can cross blood placenta barrier and may result in congenital toxoplasmosis. About 80% of immunocompetent individuals do not show any clinical manifestations and are silent carriers of this disease. Pregnant women especially in highly prevalent areas are recommended to be screened for this disease in order to prevent the potential vertical transmission. To our knowledge no such study has been conducted in this region of Saudi Arabia. This study attempted to carry out two objectives: first, to find out the seroprevalence of T. gondii infection in pregnant women attending prenatal care services in our hospital; second, to find out risk factors associated with T. gondii seroprevalence in our patients. It was carried out in Teaching Hospital in Al-Kharj over a period of one year. All 306 pregnant women attending antenatal clinic were involved in the study. A pretested selfexplanatory questionnaire was filled out by the patients and their sera were collected to be tested for IgG and/or IgM against T. gondii. The results were then statistically analyzed using SPSS software and p-value was calculated using Pearson Chi Square test. Out of the 306 blood samples tested, 99 (32.4%) were seropositive for specific anti T. gondii IgG antibodies and 3(1%) were seropositive for IgM. This show that seroprevalence of T. gondii antibodies was high among pregnant women and the prevalence showed a significant association with age. The study recommends conducting educational programs to raise awareness among women about risk factors and precautions to be taken.
Toxoplasma gondii is an obligate intracellular protozoan parasite that causes toxoplasmosis in humans. To date, little is known about T. gondii infection among the indigenous community, particularly in East Malaysia. This study was conducted to determine the status of T. gondii infection and to investigate associated risk factors among the indigenous community of Sarawak, East Malaysia. The sociodemographic data was obtained using a pretested questionnaire. A serological test was done to detect the presence of specific IgM and IgG antibodies against T. gondii in serum samples. A nested polymerase chain reaction (PCR) was used to determine acute infection among seropositive individuals. The overall seroprevalence of T. gondii infection was 50% (95% CI = 43.3 - 56.7). From this subset, 40.1%, 5.7%, and 4.2% were positive for anti-T. Gondii IgG antibodies, IgM, and both IgG and IgM, respectively. Four seropositive samples were amplified through PCR. None of the pregnant women tested positive for T. gondii infection based on the serological and PCR assays. A significant association was found between age, low monthly household income, unemployment, usage of untreated water and close contact with T. gondii seropositive cats. These results provide basic information on T. gondii infection and may be useful for policymakers to initiate prevention and control programs, especially amongst pregnant women and women of childbearing age in the indigenous community.
Toxoplasma gondii is a world-widely spread zoonotic parasite. However, scarce knowledge is known about the prevalence of T. gondii infection in people in Hubei province, China. This study herein was to perform epidemiological investigation of T. gondii infection in people in this region. A total 12527 blood samples were obtained during 2015-2018, and were assayed for T. gondii antibodies of IgG and IgM, respectively by employing an indirect hemagglutination test (IHA). The results discovered that the prevalence of T. gondii in people was 2.44% and 6.1%, respectively based on antibodies of IgG and IgM, respectively. The prevalence was ranged from 0.3% to 5.4% during 2015-2018 based on IgM antibodies. For genders, the prevalence was 0.7% and 2.6% in males and females, respectively based on IgM antibodies. In different years, the prevalence was ranged from 4.9% to 14.0% based on IgG antibodies. The prevalence of T. gondii was 4.9% and 6.6% in males and femalesy based on IgG antibodies. The current results may be helpful for the implementation of preventive measures against Toxoplasma infection among people living in this region.
Toxoplasma gondii, a ubiquitous pathogen that infects nearly all warm-blooded animals and humans, can cause severe complications to the infected people and animals as well as serious economic losses and social problems. Here, one local strain (TgPIG-WH1) was isolated from an aborted pig fetus, and the genotype of this strain was identified as ToxoDB #3 by the PCR RFLP typing method using 10 molecular markers (SAG1, SAG2, alternative SAG2, SAG3, BTUB, GRA6, L358, PK1, C22-8, C29-2 and Apico). A comparison of the virulence of this isolate with other strains in both mice and piglets showed that TgPIG-WH1 was less virulent than type 1 strain RH and type 2 strain ME49 in mice, and caused similar symptoms to those of ME49 such as fever in piglets. Additionally, in piglet infection with both strains, the TgPIG-WH1 caused a higher IgG response and more severe pathological damages than ME49. Furthermore, TgPIG-WH1 caused one death in the 5 infected piglets, whereas ME49 did not, suggesting the higher virulence of TgPIG-WH1 than ME49 during piglet infection. Experimental infections indicate that the virulence of TgPIG-WH1 relative to ME49 is weaker in mice, but higher in pigs. This is probably the first report regarding a ToxoDB #3 strain from pigs in Hubei, China. These data will facilitate the understanding of genetic diversity of Toxoplasma strains in China as well as the prevention and control of porcine toxoplasmosis in the local region.
Toxoplasmosis is a zoonotic disease caused by Toxoplasma gondii that is prevalent in humans and animals. This study was aimed to determine the seroprevalence of T. gondii infection among hemato-oncology patients and its association with sociodemographic and behavioural characteristics. This cross-sectional study was conducted at the Hospital Universiti Sains Malaysia (USM) involving 56 blood samples from hemato-oncology patients. Anti-T. gondii IgG and IgM antibodies and IgG avidity were determined using enzyme-linked immunosorbent assays (ELISA). The association of T. gondii exposure, sociodemographic, and behavioural characteristics were assessed by a questionnaire and face-to-face interviews. Twenty-eight (50%) patients were seropositive for T. gondii antibodies, where 27 (48.21%) patients were IgG+/IgM- and one patient (1.79%) was IgG+/IgM+ with high avidity index, indicating infection of more than 20 weeks. A univariate analysis showed that age, gender, ethnicity, marital status, educational level, employment status, stem cell transplant, blood transfusion, close contact with cats, water supply, and consumption of undercooked meat were not significantly associated with Toxoplasma seropositivity (p < 0.05). Our study has demonstrated, for the first time, the serological evidence of T. gondii exposure among hemato-oncology patients in Hospital USM. Our findings indicated that latent toxoplasmosis was relatively prevalence among our patients. Therefore, serological screening tests should be considered for immunocompromised patients as well as the implementation of health education programmes to encourage a healthy lifestyle and the consumption of healthy food among them.
Understanding of antibody kinetics against SARS-CoV-2 and its vaccines is rapidly evolving. This study aims to (1) determine post-vaccination seroprevalence; (2) compare antibody levels between vaccine types and various clinical/demographic determinants; and (3) determine post-vaccination antibody concentrations against time. This is a retrospective cross-sectional study involving 148 healthcare employees all over Malaysia. IgG Spike (RBD), IgM Spike and IgG Nucleocapsid concentration medians were compared using Mann-Whitney U or Kruskal-Wallis tests. Chi Square and Spearman correlation coefficient tests were performed to identify variables associated with antibody titers. A scatter plot of IgG Spike (RBD) against time from last vaccine dose was also plotted. At 1-month post-vaccination, all employees successfully seroconverted regardless of vaccine type, health status and COVID- 19 history. Comirnaty, convalescent, female or Malay vaccinees had significantly higher IgG Spike (RBD) titers compared to their respective counterparts. No correlation was found between age and IgG Spike (RBD) levels. Concentration of all three antibodies waned with time post-vaccination, with IgM Spike and IgG Nucleocapsid waning faster than IgG Spike (RBD).
Dengue infection is a major public health problem affecting millions of people living in tropical countries. With no suitable vaccines and specific antiviral drugs, treatment for dengue is usually symptomatic and supportive. Early diagnosis and recognition of severe disease is therefore crucial for better management of the patient. Two-dimension electrophoresis was used to identify disease-associated proteins that can be used for diagnosis and as drug targets for treatment. Two markers, identified by mass spectrometry analysis as alpha1-antitrypsin and NS1 proteins were found to be upregulated in dengue fever (DF; n=10) and dengue haemorrhagic fever (DHF; n=10) patients compared with healthy individuals (n=8). Both alpha1-antitrypsin and NS1 proteins were overexpressed two-fold in DHF patients compared with DF patients. Our study suggests that alpha1-antitrypsin and NS1 protein could be used as biomarkers as early indicators of DHF risk among patients with suspected dengue infection.
Accurate and timely diagnosis of dengue virus is important for early detection of dengue virus infection. In this study, the usefulness of the dengue NS1 antigen test was evaluated as a routine, rapid diagnostic test for dengue virus infection. A total of 208 sera from patients suspected of having dengue virus infection were collected and tested for dengue antibody, dengue genome and dengue NS1 antigen. Dengue antibody test, dengue PCR test and dengue antigen test were able to detect dengue virus infection from Days 1 to 8 in 72.8, 52.8 and 44.0% of samples, respectively. Of the 208 sera tested, 69.2% (144/208) of the acute sera were positive for dengue virus infection based on IgM antibody, IgG antibody, NS1 antigen and PCR tests. Thirty-two point two percent of the samples (67/208) were found positive for dengue NS1 antigen, 38.5% (80/208) were PCR positive, 40.9% (85/208) were IgM positive and 36.1% (75/208) were IgG positive for dengue virus. The results reveal the detection rate of dengue virus infection was similar for PCR and dengue antibody (65.9%) and for NS1 antigen and dengue antibody (62.0%) combinations. Therefore, the dengue NS1 antigen test can be used to complement the current antibody test used in peripheral laboratories. Thus, the combination of the NS1 antigen and antibody tests could increase the diagnostic efficiency for early diagnosis of dengue infection.
Detection of Toxoplasma gondii 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.
In Malaysia, the two dose measles - mumps - rubella (MMR) vaccine was introduced in the Expanded Program on Immunization in 2002. The Ministry of Health then initiated a measles elimination strategy which included enhanced case-based surveillance with laboratory testing of all suspected cases. The objective of our study was to analyse national measles laboratory data from 2004 to 2008 to study the impact of the nationwide strategy on measles case incidence. Blood samples collected from suspected measles cases during the acute stage of the illness were investigated for measles specific IgM. The estimated incidence of measles ranged from 22.3 cases (in 2004) to 2.27 cases (in 2006) per 100,000 population. During this time, the measles vaccination coverage was above 85%. Laboratory confirmed measles cases dropped from 42.2% in 2004, when sporadic outbreaks were reported, to 3.9% in 2007. Screening for measles IgG levels in 2008 showed that 82.8% of those > 7 years old had adequate immunity. The measles control strategy appears to have been successful in reducing the incidence of measles. Continuing high vaccination coverage rates and ongoing measles surveillance are necessary to achieve our goal of measles elimination.
This was a retrospective study of patients with dengue infection in pregnancy from year 2000 till 2004. Data were analyzed by looking at the presentation, complications of patient and fetus, and pregnancy outcomes. There was a total of 16 cases with an increasing trend (0.12% in 2003 vs 0.25% in 2004). The mean age of patients was 30.19+/-6.85 years. Fifty percent of patients were multiparae and in their third trimester. The average gestation was 24.44 weeks with 7.5 days being the average duration of ward admission. Tourniquet test was positive in 62.5% of patients. Dengue serology IgM was positive in 50% whereas dengue serology IgG were positive in 68.8% of patients. There were three cases of maternal death. One patient presented as missed abortion. Preterm deliveries happened in 50.0% of the women. There were 4 premature babies, one in-utero fetal death, and two fetuses which suffered acute fetal distress. Three babies required intensive care. One unrelated fetal anomaly resulted in early neonatal death.