Evaluation of Immunodiagnostic Performances of Neospora caninum Peroxiredoxin 2 (NcPrx2), Microneme 4 (NcMIC4), and Surface Antigen 1 (NcSAG1) Recombinant Proteins for Bovine Neosporosis.

Bovine neosporosis is among the main causes of abortion in cattle worldwide, causing serious economic losses in the beef and dairy industries. A highly sensitive and specific diagnostic method for the assessment of the epidemiology of the disease, as well as it surveillance and management, is imperative, due to the absence of an effective treatment or vaccine against neosporosis. In the present study, the immunodiagnostic performance of Neospora caninum peroxiredoxin 2 (NcPrx2), microneme 4 (NcMIC4), and surface antigen 1 (NcSAG1) to detect IgG antibodies against N. caninum in cattle were evaluated and compared with that of the indirect fluorescent antibody test (IFAT). The results revealed that NcSAG1 had the highest sensitivity and specificity, with values of 88.4% and 80.7%, respectively, followed by NcPrx2, with a high sensitivity of 87.0% but a low specificity of 67.0%, whereas NcMIC4 showed sensitivity and specificity of 84.1% and 78.9%, respectively, when compared with IFAT. A high degree of agreement was observed for NcSAG1 (k = 0.713) recombinant protein, showing the highest diagnostic capability, followed by NcMIC4 (k = 0.64) and NcPrx2 (k = 0.558). The present study demonstrates that NcSAG1 is helpful as an antigen marker and also demonstrates the potential immunodiagnostic capabilities of NcPrx2 and NcMIC4, which could serve as alternative diagnostic markers for detecting N. caninum infection in cattle. These markers may find utility in future treatment management, surveillance, and risk assessment of neosporosis in livestock or other animal host species. Further research should be directed toward understanding the in vivo immune response differences resulting from immunization with both recombinant proteins.

Recombinant Dense Granule Protein (NcGRA4) Is a Novel Serological Marker for Neospora caninum Infection in Goats.

Neospora caninum is widely recognised as one of the most significant causes of abortion in cattle, with infections also occurring in sheep and goats. To prevent and control animal neosporosis, it is crucial to develop sensitive and specific methods for detecting N. caninum infection. Recently, several recombinant proteins have been utilised in serological assays for the diagnosis of neosporosis. In this study, we used commercial gene synthesis to produce dense granular antigen 4 (NcGRA4) recombinant protein. NcGRA4 plasmids were expressed in the Escherichia coli system and then purified. The purified recombinant protein was analysed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis. To evaluate the diagnostic potential of recombinant NcGRA4 protein, we tested 214 serum samples from goat farms via indirect enzyme-linked immunosorbent assay (iELISA) and compared the results to those from the indirect fluorescent antibody test (IFAT). Western blotting analysis revealed a single NcGRA4 band with an expected molecular weight of 32 kDa. The specific IgG against N. caninum was detected in 34.1% and 35% of samples evaluated by NcGRA4 iELISA and IFAT, respectively. The sensitivity and specificity of the NcGRA4 iELISA were 71.6% and 86.3%, respectively, when compared with the results from IFAT. Our results demonstrate that a recombinant protein that can be used to detect animal neosporosis can be produced using a synthetic NcGRA4 gene. Overall, recombinant NcGRA4 shows promise as a sensitive and specific serological marker for identifying target IgG in goat samples.

Immunoproteomics to identify species-specific antigens in Neospora caninum recognised by infected bovine sera.

Bovine neosporosis is a disease of concern due to its global distribution and significant economic impact through massive losses in the dairy and meat industries. To date, there is no effective chemotherapeutic drug or vaccine to prevent neosporosis. Control of this disease is therefore dependent on efficient detection tests that may affect treatment management strategies. This study was conducted to identify the specific immunoreactive proteins of Neospora caninum tachyzoites recognised by sera from cattle infected with N. caninum, Toxoplasma gondii, Cryptosporidium parvum, Babesia bovis and B. bigemina, and by sera from uninfected cattle using two-DE dimensional gel electrophoresis (2-DE) combined with immunoblot and mass spectrometry (LC-MS/MS). Among 70 protein spots that reacted with all infected sera, 20 specific antigenic spots corresponding to 14 different antigenic proteins were recognised by N. caninum-positive sera. Of these immunoreactive antigens, proteins involved in cell proliferation and invasion process were highly immunogenic, including HSP90-like protein, putative microneme 4 (MIC4), actin, elongation factor 1-alpha and armadillo/beta-catenin-like repeat-containing protein. Interestingly, we discovered an unnamed protein product, rhoptry protein (ROP1), possessing strong immunoreactivity against N. caninum but with no data on function available. Moreover, we identified cross-reactive antigens among these apicomplexan parasites, especially N. caninum, T. gondii and C. parvum. Neospora caninum-specific immunodominant proteins were identified for immunodiagnosis and vaccine development. The cross-reactive antigens could be evaluated as potential common vaccine candidates or drug targets to control the diseases caused by these apicomplexan protozoan parasites.

Title: L'immunoprotéomique pour identifier chez Neospora caninum les antigènes spécifiques de l'espèce reconnus par les sérums de bovins infectés. Abstract: La néosporose bovine est une maladie préoccupante en raison de sa distribution mondiale et de son impact économique important par d'énormes pertes dans les industries laitières et de la viande. À ce jour, il n'existe aucun médicament chimiothérapeutique ou vaccin efficace pour prévenir la néosporose. Par conséquent, le contrôle de cette maladie dépend de tests de détection efficaces qui affecteraient les stratégies de gestion du traitement. Cette étude a été menée pour identifier les protéines immunoréactives spécifiques des tachyzoïtes de Neospora caninum reconnues par les sérums de bovins infectés par N. caninum, Toxoplasma gondii, Cryptosporidium parvum, Babesia bovis et B. bigemina et par les sérums de bovins non infectés, à l'aide d'un gel d'électrophorèse bidimensionnel (2DE) combiné à l'immunoblot et à la spectrométrie de masse (LC-MS/MS). Parmi 70 spots protéiques ayant réagi avec tous les sérums infectés, 20 spots antigéniques spécifiques correspondant à 14 protéines antigéniques différentes ont été reconnus par les sérums positifs à N. caninum. Parmi ces antigènes immunoréactifs, les protéines impliquées dans la prolifération cellulaire et le processus d'invasion étaient hautement immunogènes, notamment la protéine de type HSP90, le micronème putatif 4 (MIC4), l'actine, le facteur d'élongation 1-alpha et la protéine à répétition de type armadillo/bêta-caténine. Fait intéressant, nous avons découvert un produit protéique sans nom, la protéine de rhoptries (ROP1), possédant une forte immunoréactivité contre N. caninum mais sans données disponibles sur sa fonction. De plus, nous avons identifié des antigènes à réaction croisée parmi ces parasites apicomplexes, en particulier N. caninum, T. gondii et C. parvum. Des protéines immunodominantes spécifiques de Neospora caninum ont été identifiées pour l'immunodiagnostic et le développement de vaccins. Les antigènes à réaction croisée pourraient être évalués comme candidats vaccins communs potentiels ou comme cibles médicamenteuses pour contrôler les maladies causées par ces parasites protozoaires apicomplexes.

Development and evaluation of indirect enzyme-linked immunosorbent assay using recombinant dense granule antigen 7 protein for the detection of Toxoplasma gondii infection in cats in Thailand.

Background and Aim: Toxoplasma gondii is recognized as a zoonosis causing toxoplasmosis in animals globally. Cat is a definitive host of T. gondii and sheds oocyst through feces, which can infect human beings and animals through contaminated food ingestion. A precise diagnostic test is essential to prevent T. gondii infection in both humans and animals. This study aimed to develop and evaluate the pETite-dense granule antigen 7(GRA7)-based indirect enzyme-linked immunosorbent assay (ELISA) to detect T. gondii infection in cats. Materials and Methods: T. gondii-GRA7 was cloned and expressed in the Expresso®small ubiquitin-related modifier (SUMO) T7 Cloning and Expression System. The recombinant pETite-GRA7 was purified using HisTrap affinity chromatography and confirmed using Western blot analysis. The recombinant protein was used to develop and evaluate the indirect ELISA for T. gondii infection detection. In total, 200 cat sera were tested using pETite-GRA7-based indirect ELISA and indirect fluorescent antibody test (IFAT). The statistical analysis based on Kappa value, sensitivity, specificity, positive predictive value, negative predictive value, χ 2 test, and receiver operating characteristic (ROC) curve was used to evaluate the performance of the test. Results: A 606 bp GRA7 polymerase chain reaction (PCR) product was obtained from T. gondii RH strain genomic DNA. The gene was cloned into the pETite™ vector and transformed to HI-Control Escherichia coli BL21 (DE3) for protein expression. Approximately 35 kDa of recombinant pETite-GRA7 was observed and Western blot analysis showed positive bands against anti-6-His antibody and positive-T. gondii cat serum. A sample of 0.5 µg/mL of pETite-GRA7 was subjected to indirect ELISA to detect T. gondii infection in the cat sera. The results showed sensitivity and specificity of pETite-GRA7-based indirect ELISA at 72% and 96%, respectively. An acceptable diagnostic performance was characterized by high concordant results (94%) and substantial agreement (Kappa value=0.65) with IFAT. The seroprevalence levels of ELISA and IFAT were 10% and 9%, respectively, and were not significantly (p>0.05) different. The expected performance of ELISA at different cutoff points using the ROC curve analysis revealed 89% sensitivity and 92% specificity at the cutoff value of 0.146, with a high overall assay accuracy (area under the curve=0.94). Conclusion: In this study, the pETite™ vector, N-terminal 6xHis SUMO fusion tag, was used to improve the solubility and expression level of GRA7. The recombinant pETite-GRA7 showed enhanced protein solubility and purification without special condition requirements. This pETite-GRA7-based indirect ELISA showed high concordant results and substantial agreement with IFAT. ELISA revealed an acceptable sensitivity and specificity. These initial data obtained from cats' sera demonstrated that pETite-GRA7-based indirect ELISA could be a useful method for local serological diagnosis of T. gondii infection in cats in Thailand.

Molecular Detection and Genetic Diversity of Tick-Borne Pathogens in Goats from the Southern Part of Thailand.

Tick-borne hemoprotozoan and rickettsial diseases affect the health and productivity of small ruminants in tropical and subtropical regions. Despite the large population of goats in the southern part of Thailand, there is limited information on the prevalence of tick-borne pathogens. In this study, polymerase chain reaction was used to detect the presence of Theileria spp., T. ovis, T. orientalis, Babesia ovis, Anaplasma ovis, and A. marginale in 262 goats from three provinces in the southern part of Thailand. In this investigation, Theileria spp. and A. ovis were detected while T. ovis, B. ovis, and A. marginale were not detected. Overall infection rates of Theileria spp. and A. ovis were 10.3% and 1.5%, respectively. The co-infections of two parasites was observed in 1.5% of goats. Sequence analysis showed the presence of T. luwenshuni and T. orientalis in the goat samples. This study is the first to use the molecular detection of T. orientalis in Thai goats, and presents genetic characterization using the major piroplasm surface protein (MPSP) gene. In the phylogenetic analysis, the T. orientalis MPSP sequence was classified as type 7. The A. ovis major surface protein 4 (MSP4) gene sequences shared high identities and similarity with each other and clustered with isolates from other regions. This study provides information about the prevalence and genetic diversity of tick-borne pathogens in goats in the study area, and is expected to be valuable for the development of effective control measures to prevent disease in animals in Thailand.

Molecular identification of Pentatrichomonas hominis in animals in central and western Thailand.

BACKGROUND: Pentatrichomonas hominis inhabits the digestive tracts of several vertebrates, such as humans, monkeys, pigs, dogs, cats and rats. This protozoan was originally considered a commensal of the digestive tract but has subsequently been identified as a potential zoonotic parasite and a causative agent of diarrhoea. Molecular techniques are considered more sensitive and specific to detect P. hominis. This study aimed to determine the presence and genetic diversity of P. hominis in animals in Thailand. A total of 403 faecal samples were collected from 119 cats, 55 dogs, 73 goats, 35 monkeys, 55 cattle and 66 pigs, and the presence of P. hominis was determined using the nested polymerase chain reaction method. Sequence analysis of small-subunit ribosomal RNA genes was used to determine the genotype of the organism. RESULTS: Twenty-six samples (26/403, 6.45%) were positive for P. hominis. The highest prevalence was found in cats (21/119; 17.65%), followed by cattle (3/55; 5.45%) and dogs (2/55; 3.64%). Seven out of 26 nucleotides demonstrated 100% sequence identity with existing sequences; additionally, 16 novel sequence patterns were identified. All nucleotide sequences of P. hominis-positive samples were shown in the same branch with the previously described P. hominis sequences found in humans, dogs and goat. CONCLUSION: This is the first study on P. hominis infections in animals in Thailand. Our findings revealed that the prevalence of P. hominis was significantly higher in cats than in cattle and dogs. Cats were the main reservoir host; however, P. hominis can infect several kinds of animals. Therefore, the proper waste management of animals is necessary to reduce and prevent infection in the community.

Development and evaluation of recombinant GRA8 protein for the serodiagnosis of Toxoplasma gondii infection in goats.

BACKGROUND: The development of sensitive and specific methods for detecting Toxoplasma gondii infection is critical for preventing and controlling toxoplasmosis in humans and other animals. Recently, various recombinant proteins have been used in serological tests for diagnosing toxoplasmosis. The production of these antigens is associated with live tachyzoites obtained from cell cultures or laboratory animals for genomic extraction to amplify target genes. Synthetic genes have gained a key role in recombinant protein production. For the first time, we demonstrated the production of the recombinant protein of the T. gondii dense granular antigen 8 (TgGRA8) gene based on commercial gene synthesis. Recombinant TgGRA8 plasmids were successfully expressed in an Escherichia coli system. The recombinant protein was affinity-purified and characterized via sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Furthermore, the diagnostic potential of the recombinant protein was assessed using 306 field serum samples from goats via indirect enzyme-linked immunosorbent assay (iELISA) and the latex agglutination test (LAT). RESULTS: Western blotting using known positive serum samples from goats identified a single antigen at the expected molecular weight of TgGRA8 (27 kDa). iELISA illustrated that 15.40% of goat samples were positive for T. gondii-specific IgG antibodies. In addition, TgGRA8 provided high sensitivity and specificity, with significant concordance (91.83) and kappa values (0.69) compared with the results obtained using LAT. CONCLUSION: Our findings highlight the production of a recombinant protein from a synthetic TgGRA8 gene and the ability to detect T. gondii infection in field samples. The sensitivity and specificity of TgGRA8 demonstrated that this protein could be a good serological marker for detecting specific IgG in goat sera.

Exposure to Toxoplasma gondii in Asian Elephants (Elephas maximus indicus) in Thailand.

Toxoplasma gondii is the causative agent of toxoplasmosis in humans and various animal species worldwide. In Thailand, seroprevalence studies on T. gondii have focused on domestic animals, and information on infections in Asian elephants (Elephas maximus indicus) is scarce. This study was conducted to determine the seroprevalence of T. gondii infection in archival sera collected from 268 elephants living in Thailand. The serum samples were analyzed for anti-T. gondii immunoglobulin G antibodies using the latex agglutination test (LAT) and indirect enzyme-linked immunosorbent assay (iELISA) based on T. gondii lysate antigen (TLA-iELISA) and recombinant T. gondii dense granular antigen 8 protein (TgGRA8-iELISA). The prevalence of antibodies against T. gondii was 45.1% (121/268), 40.7% (109/268), and 44.4% (119/268) using LAT, TLA-iELISA, and TgGRA8-iELISA, respectively. Young elephants had a higher seropositivity rate than elephants aged >40 years (odds ratio = 6.6; p < 0.001; 95% confidence interval: 2.9-15.4). When LAT was used as the reference, TLA-iELISA and TgGRA8-iELISA showed a substantial (κ = 0.69) and moderate (κ = 0.42) agreement, respectively. Although our findings suggest the widespread exposure of Asian elephants to T. gondii in Thailand, the source of infection was not investigated. Therefore, investigation of the predisposing factors associated with toxoplasmosis is necessary to identify the potential risk factors for infection.

Identification of Enterocytozoon bieneusi in goats and cattle in Thailand.

BACKGROUND: Enterocytozoon bieneusi has been increasingly reported to infect domestic animals and humans, with human infections primarily reported as zoonotic in origin. The aim of the present study was to determine the presence and genotype of E. bieneusi in humans and domestic animals in central Thailand by testing stool samples of 200 apparently healthy humans, 73 goats, 60 cattle and 65 pigs using nested-PCR/ sequence analysis based on the ITS region of SSU rRNA genes. RESULTS: E. bieneusi tested positive in 2 (1%) of the 200 stool samples collected from humans and 56 (28.3%) of the 198 stool samples collected from domestic animals. The highest prevalence of E. bieneusi was observed in pigs (39/65, 60%), followed by goats (14/73, 19.2%) and cattle (3/60, 5%). Seven novel E. bieneusi genotypes were identified, which were named GoatAYE1-4 and PigAYE1-3 and clustered in either zoonotic Group 1 or Group 2. Moreover, eleven previously described E. bieneusi genotypes were also identified (O, D, H, SX1, CHC8, CHG3, CS-10, SHZC1, LW1, WildBoar5, and EbpC). All novel genotypes exhibited zoonotic potential from a phylogenetic analysis of ITS region. CONCLUSION: Our data showed that the prevalence of E. bieneusi is low in apparently healthy individuals and higher in pigs than cattle and goats. This study provides baseline data useful for controlling and preventing E. bieneusi infection in farm communities, where pigs and goats appear to be the major reservoir of E. bieneusi. The results of our study support the view that E. bieneusi is a zoonotic pathogen that should be considered a potential public health threat.

Blastocystis infection and subtype distribution in humans, cattle, goats, and pigs in central and western Thailand.

Blastocystis is a common intestinal pathogen of humans and a variety of animals, with various host-specific subtypes. The aim of this study was to determine the prevalence and subtype distribution of Blastocystis in humans and domestic animals, Thailand. 113 stool samples were collected from pigs, goats, and cattle in Ayutthaya Province (AP; central Thailand) and 218 stool samples were collected from pigs, dogs, cats, chickens, and humans in Kanchanaburi Province (KP; western Thailand). Blastocystis was detected by nested PCR targeting the SSU rRNA gene. Subtypes were identified by DNA sequencing, and phylogenetic analysis was conducted. The overall prevalence of Blastocystis in animals was 76.1% (86/113) and 11.88% (12/101) in AP and KP, respectively, and the prevalence in humans was 12.82% (15/117) in KP. The prevalence of Blastocystis in the AP and KP pigs were 87.88% (29/33) and 20.37% (11/54), respectively. Blastocystis ST5 was the most abundant in pigs in both areas while Blastocystis ST10 and ST12 were most frequently found in cattle and goats. In addition, low percentage of Blastocystis ST1 and Blastocystis ST14 were found in pigs and goats, respectively. In this study, Blastocystis ST3, followed by ST2 and ST1 were predominantly found in humans. In conclusion, pigs may be a natural host of Blastocystis and this ST may be the pig-adapted ST in the studied areas, in this study.

Development of a loop-mediated isothermal amplification technique and comparison with quantitative real-time PCR for the rapid visual detection of canine neosporosis.

BACKGROUND: Dogs are the definitive hosts of Neospora caninum and play an important role in the transmission of the parasite. Despite the high sensitivity of existing molecular tools such as quantitative real-time PCR (qPCR), these techniques are not suitable for use in many countries because of equipment costs and difficulties in implementing them for field diagnostics. Therefore, we developed a simplified technique, loop-mediated isothermal amplification (LAMP), for the rapid visual detection of N. caninum. METHODS: LAMP specificity was evaluated using a panel containing DNA from a range of different organisms. Sensitivity was evaluated by preparing 10-fold serial dilutions of N. caninum tachyzoites and comparing the results with those obtained using qPCR. Assessment of the LAMP results was determined by recognition of a colour change after amplification. The usefulness of the LAMP assay in the field was tested on 396 blood and 115 faecal samples from dogs, and one placenta from a heifer collected in Lopburi, Nakhon Pathom, Sa Kaeo, and Ratchaburi provinces, Thailand. RESULTS: Specificity of the LAMP technique was shown by its inability to amplify DNA from non-target pathogens or healthy dogs. The detection limit was the equivalent of one genome for both LAMP and qPCR. LAMP and qPCR detected positive N. caninum infection in 15 of 396 (3.8%) blood samples; LAMP detected 9/115 (7.8%) positive faecal samples, while qPCR detected 5/115 (4.3%) positive faecal samples. The placental tissue was shown to be positive by both techniques. Agreement between LAMP and qPCR was perfect in blood samples (kappa value, 1.00) and substantial in faecal samples (kappa value, 0.697). CONCLUSIONS: This is the first known LAMP assay developed for the amplification of N. caninum. The technique effectively and rapidly detected the parasite with high sensitivity and specificity and was cost-effective. This assay could be used in the field to confirm the diagnosis of canine or bovine neosporosis.

Subtype distribution of Blastocystis in Thai-Myanmar border, Thailand.

Blastocystis sp. is a common zoonotic intestinal protozoa which has been classified into 17 subtypes (STs). A cross-sectional study was conducted to determine the prevalence and subtype distribution of Blastocystis in villagers living on the Thai-Myanmar border, where the risk of parasitic infection is high. A total of 207 stool samples were collected and DNA was extracted. PCR and sequencing using primers targeting small-subunit ribosomal RNA (SSU rRNA) gene were performed. The prevalence of Blastocystis infection was 37.2% (77/207). ST3 (19.8%; 41/207) was the predominant subtype, followed by ST1 (11.6%; 24/207), ST2 (5.3%; 11/207), and ST4 (0.5%; 1/207). A phylogenetic tree was reconstructed using the maximum likelihood (ML) method based on the Hasegawa-Kishino-Yano + G + I model. The percentage of bootstrapped trees in which the associated taxa clustered together was relatively high. Some sequences of Blastocystis positive samples (TK18, 39, 46, 71, and 90) were closely related to animals (pig and cattle) indicating zoonotic risks. Therefore, proper health education in parasitic prevention for the villagers should be promoted to improve their personal hygiene. Further longitudinal studies are required to monitor the prevalence of parasitic infections after providing health education and to investigate Blastocystis ST in animals living in these villages.

Gastrointestinal parasites of dogs and cats in a refuge in Nakhon Nayok, Thailand.

We collected fecal samples from 500 dogs and 300 cats from an animal refuge in Nakhon Nayok Province, Thailand to test for gastrointestinal protozoa and helminths using a formalin-ether concentration technique. The overall prevalence of parasites in stool from dogs was 36.2% (181/500), 35.7% (177/500) had helminths and 2.8% (14/500) had protozoa. The helminths were: hookworm (30.6%), Trichuris vulpis (16.0%), Toxocara canis (6.6%), Hymenolepis diminuta (1.2%), Spirometra mansoni (0.6%), and Dipylidium caninum (0.2%). Giardia duodenalis (2.8%) was found in the stool of dogs. The overall prevalence of parasites in stool from cats was 44.3% (133/300), 43.3% (130/300) were helminths and 6.0% (18/300) were protozoa. The helminths were hookworm (34.7%), T. cati (9.7%), S. mansoni (4.0%), Platynosomum fastosum (2.7%), Strongyloides sp (0.7%), and Echinostoma sp (0.3%). Two species of protozoa, Isospora sp (5.7%) and G. duodenalis (0.3%) were found in the stool of cats. Two percent of dogs and 5.0% of cats had mixed protozoan and helminthic infections. Dogs with double, triple, and quadruple helminthic infections were found at rates of 22.0%, 2.8%, and 0.2%, respectively. Cats with double and triple helminthic infections were found at rates of 9.7% and 1.0%, respectively. Quadruple helminthic infections were not found in cats, and double protozoan infections were not found in either dogs or cats.

Is Sabin-Feldman dye test using T. gondii tachyzoites from animal inoculation still the best method for detecting Toxoplasma gondii antibodies?

Although the Sabin-Feldman dye test is the gold standard for detecting Toxoplasma antibodies in human, it is performed only in reference laboratories because live virulent T. gondii are used for the test. We collected 210 human serum samples and tested them by the dye test using in vivo tachyzoites (conventional method) then compared these results with three other methods: a dye test using cell culture-derived T. gondii tachyzoites and indirect immunofluorescent antibody tests (IFAT) using in vivo and in vitro tachyzoites. We found the conventional dye test detected the highest percent of cases (4.3%), followed by the IFAT using parasites from mice (3.8%), then the dye test and the IFAT using cell culture tachyzoites (both 2.8%). Agreement with the dye test when using mouse and cell culture derived tachyzoites was 96.7%. Using in vivo tachyzoites for the dye test and the IFAT gave 94.3% agreement, while using in vitro tachyzoites gave 94.8% agreement. When compared with the conventional dye test, the IFAT had 75% sensitivity and 100% specificity. The T. gondii tachyzoites obtained from cell culture had a lower virulence, as indicated by a three times longer survival period in the inoculated mice. We favor the conventional dye test as the gold standard for Toxoplasma antibody detection. In vitro tachyzoites can be used routinely in the dye test but false negative results may occur in some cases. The IFAT, using either in vivo or in vitro tachyzoites, are alternatives for laboratories where provision of live tachyzoites is limited.

Seroprevalence of Toxoplasma gondii antibody in Vietnamese villagers.

Toxoplasmosis caused by Toxoplasma gondii is a protozoan infection found worldwide. It usually produces non-specific symptoms, but in pregnant women and immunocompomised individuals, it may cause severe and fatal illness. Many serological studies have been done in various parts of the world, but information is lacking for Vietnam. A seroprevalence study of T gondii antibodies in Vietnamese villagers (n = 650) was performed using the Sabin-Feldman dye test. The average seroprevalence was 4.19% (95% CI = 1.78-4.62), including 6.36% (95% CI = 3.22-11.09), 4.73% (95% CI = 1.92-9.50) and 1.09% (95% CI = 0.23-3.15) from Nghe An, Lao Cai and Tien Giang provinces, respectively. This study confirmed the low prevalence of toxoplasmosis in Vietnam similar to other countries in the region. Further studies are necessary in order to provide a complete picture for the country.