Ocular Sparganosis: The First Report of Spirometra ranarum in Thailand.

A 22-year-old Thai man from the Northeast region presented with acute eye swelling, itching, and discharge on his left eye. He was suspected of having gnathostomiasis and treated with albendazole and prednisolone for 3 weeks. Nine months later, he was treated with high-dose oral prednisolone for the preliminary and differential diagnoses with thyroid-associated orbitopathy and lymphoma. He had been administered prednisolone intermittently over a few years. Then he developed a painless movable mass at the left upper eyelid and recurrent pseudotumor oculi was suspected. The surgical removal of the mass was performed. A white pseudosegmented worm revealed a definite diagnosis of ocular sparganosis by a plerocercoid larva. Molecular diagnosis of the causative species was made based on the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. Proper technique of extraction and amplification of short fragments DNA from formalin-fixed paraffin-embedded tissue successfully identified parasite species. The result from the sequencing of the PCR-amplified cox1 fragments in this study showed 99.0% sequence homology to Spirometra ranarum. This is the first report of S. ranarum in Thailand.

Gnathostoma spinigerum: immunodepression in experimental infected mice.

Mice were infected with 8- or 25-infective worms of advanced third stage Gnathostoma spinigerum larvae (L3) which were obtained from natural infected eels. On day 14, 60 and 200 post infections (PI), spleen cells of infected mice were tested for lymphoproliferative responses in vitro against the mitogen and specific L3 somatic antigen in order to clarify the cellular immune status of the host upon this nematode infection. Reduced responsiveness to Con A was observed in infected mice. These depressed responses were more pronounced in chronically infected mice (day 200, PI) than in day 14 and day 60, PI. There was no significant difference of lymphoproliferative response between groups of high (25 L3) and low (8 L3)-infective dose in the chronic readily stage. Regarding to the L3 somatic Ag stimulation, the depressed response was obviously detected in high dose and chronic infection. Our results demonstrated that in this G. spinigerum-mouse system T-cell response is defective. The depression could be reversible and was associated with active infection because it was abolished by anthelmintic (ivermectin) treatment. This study shows the involvement of Th-2 response to this nematode in regulating T cell proliferation.

Prevalence of giardiasis and genotypic characterization of Giardia duodenalis in hilltribe children, Northern Thailand.

A cross-sectional study was performed to determine the prevalence of giardiasis in hilltribe children of 2 different remote districts (Mae-chaem and Hod), Chiang Mai, Northern Thailand from November 2006-April 2007. The overall prevalence of giardiasis was 5.2%. Genetic characterization of Giardia duodenalis isolated from these children was performed using PCR methods specific for small subunit ribosomal rRNA (SSU-rRNA) and glutamate dehydrogenase (gdh) gene. This study shows that the distribution of Giardia assemblages varied in these 2 populations. Assemblage BIV was found predominantly in children from Hod District while assemblage AII was more common in children from Mae-Chaem District. Our result showed that assemblage A was significantly associated with loose/watery stool (p = 0.001). In addition, children harbouring assemblage B had shed a significantly higher number of cysts (p = 0.019) in stools than those infected with assemblage A. Further study on the epidemiology of giardiasis especially risk factors associated with genotyping would help to understand the nature of this disease in each population.

Positivity and intensity of Gnathostoma spinigerum infective larvae in farmed and wild-caught swamp eels in Thailand.

From July 2008 to June 2009, livers of the swamp eels (Monopterus alba) were investigated for advanced third-stage larvae (AL3) of Gnathostoma spinigerum. Results revealed that 10.2% (106/1,037) and 20.4% (78/383) of farmed eels from Aranyaprathet District, Sa Kaeo Province and those of wild-caught eels obtained from a market in Min Buri District of Bangkok, Thailand were infected, respectively. The prevalence was high during the rainy and winter seasons. The infection rate abruptly decreased in the beginning of summer. The highest infection rate (13.7%) was observed in September and absence of infection (0%) in March-April in the farmed eels. Whereas, in the wild-caught eels, the highest rate (30.7%) was observed in November, and the rate decreased to the lowest at 6.3% in March. The average no. (mean±SE) of AL3 per investigated liver in farmed eels (1.1±0.2) was significantly lower (P=0.040) than those in the caught eels (0.2±0.03). In addition, the intensity of AL3 recovered from each infected liver varied from 1 to 18 (2.3±0.3) in the farmed eels and from 1 to 47 (6.3±1.2) in the caught eels, respectively. The AL3 intensity showed significant difference (P=0.011) between these 2 different sources of eels. This is the first observation that farmed eels showed positive findings of G. spinigerum infective larvae. This may affect the standard farming of the culture farm and also present a risk of consuming undercooked eels from the wild-caught and farmed eels.

Clonal diversity of the glutamate dehydrogenase gene in Giardia duodenalis from Thai isolates: evidence of genetic exchange or mixed infections?

BACKGROUND: The glutamate dehydrogenase gene (gdh) is one of the most popular and useful genetic markers for the genotypic analysis of Giardia duodenalis (syn. G. lamblia, G. intestinalis), the protozoan that widely causes enteric disease in humans. To determine the distribution of genotypes of G. duodenalis in Thai populations and to investigate the extent of sequence variation at this locus, 42 fecal samples were collected from 3 regions of Thailand i.e., Central, Northern, and Eastern regions. All specimens were analyzed using PCR-based genotyping and recombinant subcloning methods. RESULTS: The results showed that the prevalence of assemblages A and B among these populations was approximately equal, 20 (47.6%) and 22 (52.4%), respectively. Sequence analysis revealed that the nucleotide diversity of assemblage B was significantly greater than that in assemblage A. Among all assemblage B positive specimens, the allelic sequence divergence within isolates was detected. Nine isolates showed mixed alleles, ranged from three to nine distinct alleles per isolate. Statistical analysis demonstrated the occurrence of genetic recombination within subassemblages BIII and BIV was likely. CONCLUSION: This study supports increasing evidence that G. duodenalis has the potential for genetic exchange.

Screening for intestinal parasitic infections among Myanmar migrant workers in Thai food industry: a high-risk transmission.

The impact of intestinal parasitic infections on public health has been neglected. Millions of Myanmar natives have migrated to work in Thailand. We performed a study of intestinal parasitic infections in Myanmar-migrants working in the Thai food industry. A total of 338 Myanmar migrant workers in a food plant at Samut Sakhon Province, Thailand, were recruited for this study. 284 (84%) returned requested stool samples. Samples were examined for intestinal parasites by means of simple smear, formalin-ether concentration, Locke-Egg-Serum medium, and Harada-Mori culture methods. We found parasites in 177 (62.3%) migrants (29 of 46 males; 148 of 238 females). The majority (89.3%) were infected with parasites transmitted by fecal-oral route, including Blastocystis hominis (41.5%), Trichuris trichiura (22.2%), Giardia lamblia (14.1%), and Ascaris lumbricoides (1.8%). Mixed infections were common (40.7%). The highest prevalence (73.3%) was found among migrants from Kohsong city, Myanmar. This high parasite infection rate in Myanmar migrant workers is an obvious public health hazard.

FlindersTechnology Associates (FTA) filter paper-based DNA extraction with polymerase chain reaction (PCR) for detection of Pneumocystis jirovecii from respiratory specimens of immunocompromised patients.

We evaluated the diagnostic value of Flinders Technology Associates (FTA) filter paper together with polymerase chain reaction (PCR) for detection of Pneumocystis jirovecii (carinii) from induced sputum (IS) and bronchoalveolar lavage fluid (BALF) samples. The study involved 162 patients with clinical diagnosis of pneumocystis pneumonia (PcP) of human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS) patients and other immunocompromised patients. P. jirovecii cysts or trophozoites were detected in IS and BALF by cytological method. The mitochondrial 5S ribosomal ribonucleic acid (rRNA) gene of P. jirovecii was amplified from these samples by using FTA filters together with a one-step PCR method (FTA-PCR). With the FTA-PCR method, the sensitivity and specificity of the test compared to microscopic examination were 67% and 90% for IS, while they were 67% and 91% for BALF, respectively. The sensitivity and specificity of the FTA-PCR test was also comparable to PCR with the conventional deoxyribonucleic acid (DNA) extraction method. We concluded that FTA-PCR is useful to detect P. jirovecii in noninvasive IS.

Prevalence of intestinal parasitic infections among school children in the central region of Thailand.

OBJECTIVE: Survey the prevalence of intestinal parasitic infections in 14 primary schools of Central Thailand. MATERIAL AND METHOD: A cross sectional survey of the prevalence of intestinal parasitic infections in 14 primary schools of Central Thailand that included Ang Thong, Ayudthaya, and Suphanburi provinces was conducted One thousand and thirty seven children with age ranging from 3-12 years (540 males and 497 females) were examined for intestinal parasites by the formalin-ether concentration technique. RESULT: The overall prevalence was 4.24%. The pathogenic parasites found were Giardia lamblia (1.25%), Enterobius vermicularis (0.19%), Trichuris trichiura and Hookworm (0.19% each), respectively. The common non-pathogenic protozoa found were Entamoeba coli (1.64%) and Endolimax nana (0.48%). The infection rate of Blastocystis hominis was 0.19%. CONCLUSION: Interestingly, the area studied which is low and flat and contains rice and water chestnut fields and small rivers, did not demonstrate Fasciolopsis buski as highly found in the past five decades. However, Parasittic infections are still public health problems in Thailand Close monitoring and control of parasitic infections is needed

Pneumocystis jiroveci in HIV/AIDS patients: detection by FTA filter paper together with PCR in noninvasive induced sputum specimens.

OBJECTIVES: To detect P. jiroveci (previously named P. carinii) by PCR using FTA filter paper to extract the DNA, from noninvasive induced sputum samples of HIV/AIDS patients. MATERIAL AND METHOD: Fifty two HIV/AIDS patients suspected of Pneumocystis jiroveci pneumonia (PJP) in King Chulalongkorn Memorial Hospital were recruited. Both cytological method and PCR with FTA filter paper technique were performed to detect P jiroveci from each specimen. RESULTS: The detectability rate of P. jiroveci infection was 21%. The PCR with FTA filter paper method was 4 folds much more sensitive than Giemsa staining technique. P. jiroveci was detected in 18% of the HIV/AIDS patients in spite of receiving standard PJP prophylaxis. CONCLUSION: Detection of P. jiroveci by using FTA filter paper together with PCR in induced sputum samples could detect more cases of P. jiroveci infection than by using cytological method. DNA extraction using the FTA filter paper was more rapid and convenient than other extraction methods. The causes of failure of PJP prophylaxis should be evaluated.

Prevalence of parasitism among students of the Karen hill-tribe in Mae Chame district, Chiang Mai province, Thailand.

Infection caused by intestinal parasites is still a common health problem in a poor-hygiene population especially for children in developing countries. A cross-sectional study was conducted among 781 Karen students (age: 3 to 19, males: 325, females: 456) to determine the current status of intestinal parasitic infections in a mountainous area in the North of Thailand. The study was drawn from three schools in the Doi Inthanon region, in Mae Chame district of Chiang Mai province, from December 2002 to June 2003. The techniques used for the diagnosis were: stool concentration by using the 'formalin-ether' technique and perianal region examination by using the 'Scotch-tape' technique. The average rate of intestinal parasites for the group tested using the stool concentration technique was 42.06% (male: 46.87%, females: 38.82%); and 22.66% (males: 28.92%, females: 18.20%) when using the Scotch-tape technique.Among helminth-infected individuals, enterobiasis was found at the highest prevalence (15.49%). Other common infections were ascariasis (9.78%), trichuriasis (5.90%) and hookworm infection (2.20%). Strongyloildiasis was found only in 0.92%. For protozoa infection, the major cause is the non-pathogenic species "Entamoeba coli" (27.68%). The other non-pathogenic protozoa (Endolimax nana, Chilomastix meslini and Iodamoeba butschlii) had a low prevalence from ranged 0.18%-4.79%. The prevalence of pathogenic Giardia lamblia infection was 2.21%. Entamoeba histolytica infection was found in only one case. Based on the two techniques used, the results from the Scotch-tape provided a higher sensitivity for the detection of Taenia spp. and Enterobius vermicularis eggs. Drug treatment was given to all the infected students. School-based health education should be implemented in order to prevent and control the infections.

Platelet parameters in subjects infected with hookworm.

Hookworm is an important intestinal parasite infection with high prevalence in tropical countries. In addition to iron deficiency anaemia, the human hookworm appears to have evolved a number of complementary strategies to overcome the host's haemostatic processes. These include the inhibition of blood coagulation, platelet aggregation and mediator release, and the secretion of fibrinogenolytic enzymes. In this work, we studied the platelets in the subjects with detection of hookworm infection from screening stool examination comparing to those without. One hundred subjects were included, studied for stool parasite and platelet parameters. Of our 100 subjects, hookworms were identified in stools of six cases, giving an infection rate equal to 6%. The platelet parameters of the subjects showed a statistically significant lowering of mean platelet volume (MPV) in the subjects with hookworm infection. Also, we detected a non-significant lower platelet count in subjects with hookworm infections. Of interest, the alteration in platelet size, identified by platelet parameters, has never been documented in the literature. This observation might be a pathological process or might be an accidental finding in our study.

Intestinal parasitic infestations among children in an orphanage in Pathum Thani province.

Infection caused by intestinal parasites is still a common health problem especially in children from developing countries. Orphans are a group of underprivileged population in society. To evaluate the intestinal parasitic infections in children in an orphanage in Pathum Thani province, Thailand, stool samples were collected during a cross-sectional study in April 2001. Examination for intestinal parasites were performed by using simple smear, formalin-ether concentration, Boeck and Drbohlav's Locke-Egg-Serum (LES) medium culture and special staining (modified acid-fast and modified trichrome) techniques. A total of 106 pre-school orphans (60 males and 46 females), aged 10.0-82.0 months, were recruited for the study. There were 86 individuals (81.1%), 45 males and 41 females, infected with at least one parasite. Interestingly, most of the parasites identified were protozoa. Blastocystis hominis was found at the highest prevalence (45.2%). The infections caused by Giardia lamblia was 37.7 per cent and Entamoeba histolytica was 3.7 per cent. Other non-pathogenic protozoa found were Trichomonas hominis (39.6%), Entamoeba coli (18.8%), and Endolimax nana (3.7%). The only one case of helminth parasite detected was Strongyloides stercoralis (0.9%). The sensitivity for detection of B. hominis and T. hominis was increased by the LES culture technique. No history of diarrhea symptoms were recorded among these orphans. However, during the investigation, stools of all infected cases were noted for six characteristics including formed, soft, loose, mucous, loose-watery and watery. The present study emphasized the problems of protozoan infections among these orphans. Health educations as well as routine surveillance is necessary in order to control the infections.

Study of specific IgG subclass antibodies for diagnosis of Gnathostoma spinigerum.

Gnathostoma spinigerum infection is endemic in Thailand and many Asian countries. Current diagnosis is the skin test and enzyme-linked immunosorbent assay (ELISA) for IgG antibody against the G. spinigerum third-stage larvae (L3), but cross-reactivity is common. We evaluated the sensitivity and specificity of anti-G. spinigerum L3 IgG subclass antibodies for diagnosis of 43 patients with gnathostomiasis. The majority of patients with gnathostomiasis (91%) had eosinophilia. While the anti-G. spinigerum L3 IgG1 antibody provided the highest sensitivity (98%), the anti-G. spinigerum L3 IgG2 antibody had the highest specificity (88%). The ELISA that detected anti-G. spinigerum L3 IgG1 antibody could be a reliable laboratory screening test, while anti-G. spinigerum L3 IgG2 antibody could be used to confirm the diagnosis.

Preparation of Gnathostoma protein by ultrafiltration method using Nanosep membrane.

We report our experience with Gnathostoma protein preparation by the ultrafiltration method. Crude antigen was sonicated and ultrafiltrated using the Nanosep 100 K membrane. SDS-PAGE electrophoresis showed protein bands at 43, 41, 24, 22, 21, 19.5 kDa. Use of the ultrafiltration method can provide specific protein (24 kDa), similar to the non-ultrafiltration method, with the other 5 non-specific proteins. Using the non-ultrafiltration method, there were more (20) non-specific protein. The ultrafiltration method can be an alternative method for the preparation of protein, which can provide better results than non-ultrafiltration.

Comparison of methods for identification of Pneumocystis carinii in bronchoalveolar lavage fluid.

Pneumocystis carinii pneumonitis in one of the most common life-threatening opportunistic infections in patients with AIDS. The definitive diagnosis of this infection can be established only by demonstration of the organism in clinical specimens. This study was a comparison of methods that provide easy recognition of the organism which is readily available, simple and can be performed rapidly in laboratory-diagnosis. Bronchoalveolar lavage fluids obtained from 35 AIDS patients suspected of having Pneumocystis carinii pneumonitis were examined by three staining methods for the presence of Pneumocystis carinii. With Giemsa stains, P. carinii could be identified in 18 cases (51.4%). Three developmental stages: "cyst", "sporozoite" and "trophozoite" were seen. The contrast of organisms against host cells was not outstanding in these stains. Toluidine blue O stains provided easy recognition of the organisms, with marked contrast between the cysts and host cells. 21 cases (60%) were positive in these stains, but the intracystic structures and trophozoites could not be identified. It was suggested that the clinical specimen should be stained first with toluidine blue O which is more rapid and permits easy recognition of the cyst clusters. If the sporozoites and trophozoites had to be identified, Giemsa stains can be made. In addition, with the methenamine silver nitrate stains, 21 cases (60%) were positive. They revealed the morphology as seen with toluidine blue O but the cost of material may make it unavailable in many laboratories especially with the budgetary restraints of developing countries.

A prevalence of Cryptosporidium infections among Thai HIV-infected patients.

The prevalence of Cryptosporidium in 156 HIV-infected Thai patients who had acute diarrheal illness at Bamrasnaradura Infectious Diseases Hospital, was studied. This cross-sectional study was performed from March to August in year 2001. The patients ranged in age from 1 month-65 years old. A stool sample from each subject was stained to find the oocysts by modified Ziehl Nelson carbolfuchsin staining. According to the present study, a diagnosis of Cryptosporidium parvum infection was found in 20 patients (11 males and 9 females). The prevalence of cryptosporidiosis in the present series was 12.8 per cent (10.0% in males and 19.1% in females). This infection rate between males and females was not significantly different. Comparing this prevalence to a report in the previous 5 years in the same hospital, the same high rate can be seen.

Prevalence and intensity of third stage Gnathostoma spinigerum larvae in swamp eels sold in three large markets in Bangkok, Thailand.

Gnathostoma spinigerum is a common human tissue parasite in Thailand. The swamp eel is the major intermediate or paratenic host for this parasite. The high prevalence of the infective third stage larvae (L3) of this parasite in the rainy season has been noted in previous studies. During June 1999 (rainy season), we performed a cross-sectional survey of the prevalence and the intensity of G. spinigerum L3s in the livers of swamp eels that were obtained from three large Bangkok markets (Klong Toey, Pran Nok, and Tevej). Of a total of 785 livers, G. spinigerum L3s were found in 97: an infection rate of 12%. The prevalence rates in Klong Toey, Pran Nok, and Tevej markets were 13%, 10% and 14% respectively. There was no significant difference in the proportion of infected eels between the markets (p > 0.05). The intensity of L3 burden in the livers ranged from 1-17 larvae, with an average of 2.60 +/- 0.24 larvae (mean +/- SE). In this study a high prevalence of G. spinigerum L3s was found; there was no significant difference in the intensity of larvae in swamp eels between markets. These findings emphasize the importance of public health education: people need to be aware of the dangers of consuming raw or undercooked food.

Parasitic infections in Thai workers that pursue overseas employment: the need for a screening program.

Stool examination is a requirement of the screening program for Thai workers seeking overseas employment. We report on the prevalence of intestinal parasitic infections among these workers; the stool examination results of 2,213 Thai workers who visited the Out-patients Department of the King Chulalongkorn Memorial Hospital between September 2000 and January 2001 were analysed. One hundred and thirty-five (6.1%) were found to be infected with 1 or 2 species of parasite. There were 54 (40%), 47 (34.8%), and 25 (18.5%) cases infected with nematodes, platyhelminths and protozoa, respectively. The commonest parasite was Opisthorchis viverrini (28.9%). Most nematode infections were due to Strongyloides stercoralis (19.2%); most protozoal infections were due to Giardia lamblia (17.0%). Mixed infections were found in 9 cases. Not unexpectedly, the rate of intestinal parasitic infection among Thai workers remains high. Screening for the parasitic infections among these workers can help to decrease their rate of rejection by the countries in which overseas employment is sought.