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.

Canine parasitic zoonoses in Bangkok temples.

Fecal samples were collected from 204 humans and 229 dogs from 20 different temples in Bangkok, as well as communities in the surrounding temple ground areas. Human and dog stool samples were examined for intestinal parasites including Giardia using zinc sulfate flotation and microscopy. Hookworms were the most common parasite in dogs (58.1%) followed by Trichuris (20.5%), Isospora (10%), Giardia (7.9%), Toxocara (7.4%), Dipylidium caninum (4.4%) and Spirometra (3.1%). Blastocystis hominis (5.9%) was the most common parasite in humans followed by hookworms (3.4%), Giardia (2.5%), Strongyloides (2%) and Cryptosporidium (1.5%). All samples microscopy-positive for Giardia were genotyped. The majority of Giardia isolated from the dog population was placed in Assemblage A, followed by Assemblages D, B and C, respectively, while human isolates were placed in Assemblages A and B. Therefore, dogs in temple communities posed a potential zoonotic risk to humans for transmission of hookworms, Giardia (especially Assemblage A genotypes) and Toxocara canis.

Immunohistochemical study of acute and chronic toxoplasmosis in experimentally infected mice.

Acute and chronic Toxoplasma infections were evaluated in mice using stage specific antibodies and immunocytochemistry. Mice with acute toxoplasmosis were less active, had erectile body hair and seldom took food or water resulting in weight loss. All mice died within 7 days post-inoculation. The immunohistochemical technique enhanced visualization of parasites allowing their distribution to be accurately followed. Following intraperitoneal infection, tachyzoites were initially identified on the surface of the liver and spleen. There was a rapid increase in the number of tachyzoites associated with invasion from the surrounding connective tissue into the organs with formation of inflammatory lesions in the liver. The focal inflammatory lesions showed increasing numbers of tachyzoites with the period post-inoculation. Similar increases in tachyzoites were observed for the spleen. In contrast, only a few individual tachyzoites were seen in the brain at the final time point. In chronic infections, the mice were asymptomatic but tissue cysts containing large numbers of bradyzoites were observed in all brains with the average number of 295 tissue cysts per half brain and the average cystic size of 46.02 +/- 5.08 microm. By histology and immunostaining, the tissue cysts were readily identifiable along with a mild inflammatory cell infiltration into the meninges and perivascular cuffing. Double immunocytochemical labelling confirmed the exclusive presence of tachyzoites during the acute phase and bradyzoites during the chronic phase.

In vivo culture of Cryptosporidium oocysts for laboratory use.

In the present study, we describe in vivo cultivation to produce oocysts. Seven-day-old mice were orally infected with 100,000-120,000 Cryptosporidium oocysts. On day 8 post-infection, the mice were killed by ether, and the small and large intestines collected. A simple extraction procedure was used and purified using Ficoll gradient centrifugation. After purification, the oocysts were preserved in phosphate buffered saline with antibiotic at 4 degrees C before use.

Detection of Toxolasma gondii in captive wild felids.

Toxoplasma gondii can infect all species of warm-blooded animals, including humans, and causes serious diseases in immunocompromized hosts. Live tachyzoites derived from serial passage in HeLa culture were used in the Sabin-Feldman dye test for detection of Toxoplasma gondii antibody in serum samples of 21 captive wild felids including one fishing cat (Prion nailurus viverrina), one leopard (Panthera pardus), two flat-headed cats (Prion nailurus planiceps), 6 tigers (Panthera tigris), two leopard cats (Felis bengalensis), two clouded leopards (Felis nebulosa), 3 pumas (Puma concolor), and 4 jungle cats (Felis chaus). Antibodies to Toxoplasma gondii were founded in 9 of 21 felids (42.8%). This study revealed that cell culture-derived tachyzoites can be used successfully as a source of live organisms in a gold standard Sabin-Feldman dye test, which is simpler, cheaper and less ethically sensitive than in vivo inoculation.

Cultivation of Giardia duodenalis in Mongolian gerbils.

The Mongolian gerbil (Meriones unguiculatus) is susceptible to infection with Giardia duodenalis trophozoites. Each animal was orally infected with 0.5 ml Diamond's TYIS-33 culture medium containing 10(6) trophozoites. Cysts were then collected and concentrated by sucrose gradient centrifugation. G. duodenalis cysts were first observed in feces on day 5 post-infection. The characteristic of G. duodenalis infection in gerbils was intermittent cyst release. The range in the number of cysts released per gerbil for a 4-hour collection period was 0-1.5 x 10(3).

Protozoal contamination of water used in Thai frozen food industry.

This study evaluated the prevalence of contamination of water that was used for food preparation. Since protozoal cysts can be found in small numbers in water, 1,000 liters of either untreated or treated water were filtered through activated carbon block filters (1 microm nominal porosity). Identification of protozoa was performed using specific monoclonal antibodies against Giardia and Cryptosporidium parasites followed by fluorescence microscopy. Twelve of 20 untreated water samples (60%) were found to be contaminated by Giardia cysts, with an average of 53.33 cysts/1,000 liters (geometric mean 39.43), whilst 7 samples (35%) were contaminated by Cryptosporidium oocysts, with an average of 28.57 oocysts/1,000 liters (geometric mean 26.92). Three samples of untreated water (15%) were positive for both organisms. In contrast, none of the treated water samples were contaminated.

Spa, springs and safety.

Natural mineral water has long been used worldwide for bathing and health purposes. At present, Thailand is famous for health spas and natural hot springs among local people and tourists. Due to possible risks of exposure to harmful agents, we studied hazardous pollutants at 57 natural hot springs from 11 provinces in northern, central, eastern and southern Thailand. Pathogenic, free-living amebae of the genera Naegleria and Acanthamoeba, which can cause central nervous system infection, were found in 26.3% (15/57) and 15.8% (9/ 57), respectively. Dissolved radon, a soil gas with carcinogenic properties, was present in nearly all hot springs sites, with concentration ranging from 0.87-76,527 Becquerels/m3. There were 5 water samples in which radon concentration exceeded the safety limit for drinking. Legionella pneumoniphila (serogroups 1, 3, 5, 6, 7 10 and 13) were found in samples from 71.9% (41/57) of studied sites. Because spas and natural springs are popular tourist attractions, health authorities should be aware of possible hazards and provide tactful measures and guidelines to ensure safety without causing undue alarm to foreign and Thai tourists.

Free-living ameba contamination in natural hot springs in Thailand.

Thermo tolerant free-living ameba, Naegleria spp and Acanthamoeba spp contamination in natural hot springs in Thailand were carried out from 13 provinces. The temperature of hot springs water varied from 28 degrees-65 degrees C and pH from 6-8. We found that 38.2 % (26/68) of water samples were positive, Acanthamoeba was 13.2% (9/68) whilst Naegleria was 35.3% (24/68). Contamination by free-living ameba in natural hot springs may pose a significant health risk to people who use such water for recreation activities.

Comparison of five DNA extraction methods and optimization of a b1 gene nested PCR (nPCR) for detection of Toxoplasma gondii tissue cyst in mouse brain.

Toxoplasmosis, caused by Toxoplasma gondii, is an important parasitic disease worldwide. Different techniques have been developed for T. gondii detection. At present, polymerase chain reaction (PCR) has been widely used. However, PCR for identifying T. gondii remains unsatisfactory in many laboratories because of lack of standardization and variations in efficiency. In the present study, we optimized a nested PCR protocol (n-PCR) in order to compare the amplification of T. gondii DNA, after being extracted from mouse brain by five different DNA extraction methods including phenol chloroform, QIAamp DNA minikit, Genomic DNA purification kit and Chelex with or without proteinase K. All DNA extraction methods were able to extract DNA from a single tissue cyst from mouse brain. However, among the five DNA extraction methods, the Chelex without proteinase K appeared to be the most rapid and easiest.

Sensitivity and specificity of polymerase chain reaction for the detection of Toxoplasma gondii DNA added to laboratory samples.

We studied the sensitivity and specificity of PCR to detect T. gondii DNA by aliquoting various concentration of tachyzoites into laboratory specimens from 60 positive and 10 negative buffy-coat samples. We were able to detect the specific gene from purified DNA samples containing as few as 0.25 parasites per 100,000 human leukocytes. These results had an impressive initial 100% specificity but later it decreased because of false-negative data.

PCR technique for detecting Toxoplasma gondii in animal amniotic fluid.

The goal of diagnosing congenital toxoplasmosis is early detection of maternofetal transmission, for early treatment to prevent unwanted sequelae. Polymerase chain reaction (PCR) is a method used recently for detecting toxoplasmosis during pregnancy. Amniotic fluid is a the clinical specimen used, since it provides a rapid, simple and safe method to obtain accurate results. The advantages of the PCR technique are high sensitivity, specificity and positive predictive value compared with other laboratory methods. To determine the sensitivity, specificity and lower detection limits in our laboratory, amplification of the B1 gene by nested PCR was performed on Toxoplasma gondii tachyzoites added to animal amniotic fluid samples. From 48 samples, our technique detected T. gondii in 30 out of 41 positive samples, and gave negative results for all the negative samples. The sensitivity for this nested PCR was 73%, the specificity was 100%, and the efficiency of the test was 77.1%. The nested PCR technique is recommended as a diagnostic method for detecting T. gondii in suspected congenital toxoplasmosis animals.

Toxoplasma gondii antibody in Thai cats and their owners.

Humans are thought to acquire Toxoplasma infection by three major routes: ingesting food and water contaminated with oocysts from cat excreta, consumption of under-cooked infected meat, and transplacental transfer. Congenital clinical toxoplasmosis in the newborn indicating definite transplacental transmission had been reported in Thailand, whilst studies concerning infection due to the other two routes were inconclusive. Since the way domestic cats live and eat and also the eating behavior of Thai people differ from those in the West, we conducted a sero-epidemiological study of T. gondii in cats and their owners in Bangkok metropolitan area. Among 327 humans, the prevalence of Toxoplasma antibody was 6.4% and in 315 cats it was 7.3%. These relatively low prevalence rates may result from the predominantly well-cooked fish and rice diet of stray cats, which congregate in temples where they are fed. Toxoplasma antibody seropositive was associated with living in close proximity to seropositivity cats [OR (95% CI) = 5.43 (1.28-23.04); p=0.01]. Risks were increased in and around temples, particularly if courtyards were of earth or grass, suggesting ground temperature was an important determinant of oocyst survival.

Malaria and enterobiasis among Karen Long-neck tribe in Mae Hong Son Province.

In Thailand, Mae Hong Son Province is highly endemic for malaria. Knowing this, the local Health Department has introduced a program to educate local residents about the risk factors, in particular the dangers and symptoms of malaria. This study was conducted to evaluate these efforts, by determining the number of malaria infections in a segment of the population, and also by testing for enterobiasis among a group of its children. Two villages in Mae Hong Son Province were chosen for this purpose with a combined population of about 300. Of these, 195 were screened for malaria. Two subjects were diagnosed positive for malaria by microscopy. One of these two villages was chosen to screen for Enterobius vermicularis infection in children as well. Out of 69 stool samples, five (7%) showed infection with E. vermicularis: three with a low number of eggs (1-50), and two with a high number of eggs (>100). Compared with infection rates in similar studies, the results of this study indicate that the Health Department's efforts are meeting with relative success. The low prevalence of infection indicates that the villagers are using the information they have received to help combat infection.

Pathologic study of acute toxoplasmosis in experimental animals.

We studied the pathology of acute toxoplasmosis in experimental mice inoculated with RH strain tachyzoites of Toxoplasma gondii. All died from severe disseminated toxoplasmosis involving the liver, spleen and pancreas. Pathological features of acute toxoplasmosis in susceptible mice could be regarded as an excellent model for acute reactivation of Toxoplasma in the immunosuppressed host.