Infection of Cyclospora cayetanensis in Diarrhoeal Children of Nepal.

Introduction: Cyclospora cayetanensis is a coccidian parasite that causes recurrent gastroenteritis among children living under poor sanitary condition and adults from industrialized countries who lived or traveled in endemic developing countries. Methods: A total of 1842 stool specimens from gastroenteritis patients were examined between March 2006 to February 2007 and collected various types of faecal specimens from human, animals and samples of water and green leafy vegetables. Results: Of 1842 stool specimens collected from different areas of Nepal, 146 (7.9 %) were found to be positive for Cyclospora cayetanensis. with the majority were from children 2-9 years of age. The highest rate of infections was found in the month of June; (31.6%). Cyclospora cayetanensis were found to be contaminated in green vegetables including leaves of basil, mint leaves, water sources and feces of domestic animals and these findings were presented in the paper. Conclusion: Cyclospora-like oocysts were detected in vegetables leaves, feces of domestic animals and water sources. The results suggest that water, vegetables and domestic animals are possible sources of infection in Nepal. The study also obtained more information on Cyclospora cayetanensis a coccidian parasite that infect humans especially children and causes prolonged diarrhea and life threatening infestation.

Affinity purified oval antigen for diagnosis of Opisthorchiasis viverrini.

Monoclonal antibodies (MAb) were raised against an oval antigen of the liver fluke Opisthorchis viverrini which is the causative agent of a parasitosis, i.e. opisthorchiasis in Thailand. The antibodies were used in an affinity column to purify the O. viverrini oval antigen from a crude extract of adult parasites by chromatography. The oval antigen was then used in a membrane (dot) ELISA for detecting antibodies in serum samples of parasitologically confirmed Opisthorchis viverrini infected individuals (adult parasites were found in stools after praziquantel treatment and salt purgation), as well as of individuals infected with other parasites and parasite-free controls. The MAb-based dot-ELISA using the affinity purified O. viverrini oval antigen revealed 100% sensitivity, specificity and accuracy for detecting O. viverrini infection. The test is simple, rapid and highly reproducible. Several samples can be tested at the same time without the requirement for special equipment or much increase in testing time; thus it is suitable for mass screening for O. viverrini exposure, especially in new endemic areas. Furthermore using serum specimens could increase patient and community compliance compared to the conventional parasitological survey which uses stool samples for the detection of O. viverrini ova, without treatment and subsequent salt purgation, this conventional method shows a low sensitivity and is also unpleasant to both the sample donors and the laboratory technicians which has historically shown a further negative impact on the final outcome.

Emerging pathogen Cyclospora cayetanensis infection in Nepal.

Cylospora cayetanensis, an emerging parasitic pathogen of human is being increasingly recognized throughout the world, however the means of transmission and the possibility of a reservoir host remain an enigma. A longitudinal study on cyclosporiasis in different parts of Nepal was carried out from April, 1995 until November, 2000. Fecal specimens were collected from symptomatic and asymptomatic patients. The data shows a distinct seasonality with the highest infection rates occurring during the summer and rainy season of the year. Attempts have been made to determine the sources of infection and possible reservoir hosts. Stools were examined from nearly 700 animals such as chickens, pigs, buffalos, cows, dogs, cats, monkeys, rats, mice and pigeons. In addition, vegetable farms around the Kathmandu Valley were examined during the seasonal high and low periods of transmission. C. cayetanensis-like oocysts were found in sewage water and from vegetable washings on five occasions during June, July, August, October, and November. Similarly, C. cayetanensis-like oocysts were recovered from mice, rats, chickens, and dogs. These results suggest that these sources may be important in the transmission of this parasitosis. However, further studies will be required to obtain definitive answers on transmission.

Study of Cyclospora cayetanensis in health care facilities, sewage water and green leafy vegetables in Nepal.

Cyclospora cayetanensis, a newly emerging parasite, is endemic in Nepal. A total of 2,123 stool specimens were collected from 3 health care facilities based on clinical symptoms during the period between 1995 to October, 1998. Out of these specimens, cayetanensis oocysts were found in 632 (29.8%). To identify possible sources for Cyclospora infection, drinking water, sewage water, green-leafy vegetables including fecal samples of various animals were collected and examined. The vegetable leaves were washed in distilled water then the washings, sewage water and drinking water were centrifuged and the sediment were examined microscopically. As a result, oocyst of Cyclospora were identified in sewage water and vegetable washings on four different occasions in June, August, October and November. The positive results were also confirmed as C. cayetanensis by development of 2 sporocysts after 2 week incubation period in potassium dichromate. A survey of 196 domestic animals from the same areas demonstrated that two chickens were positive for Cyclospora-like organism and others were negative. Although further studies are needed to clarify the direct link between Cyclospora infection and these sources, the results suggest that sewage water, green leafy vegetables are possible sources of infection and chickens could be possible reservoir host of Cyclospora in Nepal.

Recent advances in epidemiological field techniques in filariasis.

There have been very few advances in epidemiological field techniques for filariasis in recent years. Advances in other areas of study of filariasis have been published and a summary of some of those which have some application to field technology will be given. Studies on vectors involving host influences on the development of filariae and the use of DNA probes for vector speciation were reported as well as the development of techniques that yielded species-specific identification of larvae in mosquito vectors. Experiments on animal transmission potentials also have been described. Mathematical models are becoming popular for various biological systems. It is postulated that models will enable workers to make predictions on prevalence of diseases and effect of control measures. A model has been proposed for bancroftian filariasis transmission dynamics and frequency distribution of microfilariae in blood samples. Other statistical methodologies have been published relating to age and sex distribution and frequency distribution and one suggested that females have increased resistance to filariasis, especially in the reproductive years. Advances in diagnostic technologies are highly relevant to the epidemiology of filariasis. The development of antibody and especially antigen detection systems using monoclonal antibodies, DNA probes, and polymerase chain reactions, etc, will make a significant impact on the identification of infected persons. Findings along other lines may also have application to field epidemiology. The quantitative blood count (QBC) acridine orange hematocrit tube has also been found to be of value in diagnosis. Elevated levels of ecdysteroids found in other filarial infections should be tested in cases of lymphatic filariasis.

Capillariasis philippinensis: a fish-borne parasitic zoonosis.

Fish from lagoons in Northern Luzon, Philippines, have been shown experimentally to be intermediate hosts of Capillaria philippinensis. Eggs ingested by the fish hatch in the intestines and the larvae double in size in 3 weeks. When fed to monkeys, Mongolian gerbils and birds, the larvae develop into adults and reproduce. Fish from the lagoons or purchased from the market were fed to gerbils and the animals developed patent infections, demonstrating natural transmission of the parasite. Philippine and Thailand populations have the habit of eating uncooked freshwater and brackish water fish and some have been experimentally infected with C. philippinensis. These countries report the largest number of cases of intestinal capillariasis. Fish-eating birds are now considered natural hosts for the parasite, fish the intermediate hosts, and humans accidentally become infected by eating infected fish.

Symptoms associated with diarrhoeal illness at San Lazaro Hospital, Manila in 1983 and 1984.

The clinical features associated with various agents of diarrhoeal disease were studied using 2,836 patients admitted to San Lazaro Hospital, Manila. Three general patient groups were considered including single pathogen isolations, "multiple pathogen" isolations, and "no pathogen" isolations. In general, symptoms of diarrhoeal illness were found to be non-specific. However, Shigella flexneri. Vibrio parahemolyticus, and rotavirus were significantly associated with a number of prominent symptoms and could sometimes be predictably diagnosed on clinical grounds, especially when age of the patient was considered. Clinical diagnosis cannot be considered an adequate substitute for laboratory methods; other enteric pathogens can sometimes present with the same symptoms. When appropriate laboratory testing is unavailable, as is often the case in developing countries, symptomatologic diagnosis may be of limited value for the organisms mentioned.

Biotechnological methods on the study of nematodes.

During the past several years, a great many advances have been made in obtaining a better understanding of the biology of nematode parasites. Much of the information obtained has been with the use of tools developed in the expanding field of biotechnology, or more specifically, the use of biochemistry, genetics or molecular biology. Advances are being made on speciation of parasites, their function and pathogenesis. Some nematode parasites indigenous to Southeast Asia have been studied, but much more should be done. There is a large group of parasites in Southeast Asia that could be studied. In the past, emphasis has been on human nematodes, but there are many interesting parasites of lower animals that should be investigated. Furthermore, future parasitologists must first be trained in basic parasitology and then be exposed to new technology in order to apply these tools to further our knowledge of round worms.

Biotechnology research on cestodes in the Philippines, Malaysia, and Indonesia.

There are essentially no reports on the use of modern biotechnological methods on the study of cestode parasites in the Philippines, Indonesia or Malaysia. The only recent reports of cestode studies in these countries have been on reports of new species in animals and on prevalence rates of cestode parasites in humans; Taenia solium and cysticercosis, Taenia saginata and Hymenolepis nana, etc. Reports on the use of biotechnology has emanated from outside the area on cestodes of humans and animals, and some of these methods could be used to study cestodes in this part of the world.

Albendazole in the treatment of intestinal capillariasis.

Albendazole or Zentel a benzimidazole derivative, was used to treat intestinal capillariasis in hospitalized patients seen in Northern Luzon in the Philippines. In initial studies, four patients responded satisfactorily to the drug in dosages of 400 mg/day in divided doses for 20 days. This is the same dosage used for mebendazole, which has been the drug of choice in treating the parasitosis. In further studies, 16 patients with Capillaria philippinensis, were treated with 400 mg/day of the drug for 10 days, and 15 were completely cured. The one treatment failure was found to be passing eggs of the parasite three months later, and was retreated with the standard mebendazole regimen of 400 mg/day for 20 days. Side effects attributed to albendazole were not reported in any of the patients. Albendazole, 400 mg/day in two divided doses, is now the recommended drug of choice in Northern Luzon in the Philippines.

Development of Brugia malayi in Mongolian gerbils previously exposed to Wuchereria bancrofti.

Twelve Mongolian gerbils, Meriones unguiculatus, were infected with 100 third-stage larvae of Wuchereria bancrofti. One month later these animals, along with 4 control animals, were given 100 third-stage larvae of Brugia malayi. Eleven of the 12 experimental animals and the 4 controls survived, and 8 of the experimental animals and all of the controls demonstrated microfilaremia after 3 months. The animals were killed at 6-months post-infection and examined for parasites. One W. bancrofti larva was found in one of the experimental animals, and 15% of the B. malayi given were recovered as adults from the testes, viscera, and carcass. Thirty-eight percent of the worms given to the controls were recovered from the testes, viscera, and pelt. The worms from the experimental animals also appeared to be smaller. This study suggests that gerbils are able to develop partial resistance to Brugia malayi following a previous infection with Wuchereria bancrofti.

Attempt to culture Wuchereria bancrofti in vitro.

Third-stage larvae of Wuchereria bancrofti recovered from laboratory raised Aedes togoi and Anopheles maculatus fed on a human volunteer were recovered by mass dissection methods and introduced into in vitro culture. LLC-MK2 cells were used as feeder cells, and the culture medium consisted of RPMI-1640 buffered with Hepes and sodium bicarbonate and supplemented with human AB serum. The third-stage larvae molted as early as 12 days and those surviving had all molted by 16 days. The fourth-stage parasites averaged in length from 1.4 mm to a maximum of 1.8 mm. Some larvae remained alive in culture as long as 40 days and while the worms were distorted in fixation, possible primodial cells of a spicule could be visualized in the rectal region. The cuticle also appeared to be separating in the posterior end. Although complete development was not achieved, it seems that with a continuing effort, success could be obtained using this culture system with feeder cells.

Prevalence of Trichomonas vaginalis in some Filipino women.

Vaginal specimens obtained from 1,284 hospitality girls and 87 expectant mothers were examined for Trichomonas vaginalis by first examining material collected from vaginal swabs and after incubation in Feinberg and Whittington culture medium. Twenty-four percent of the specimens examined, shortly after the cotton-tip swab was placed into 1 ml culture medium, were positive by direct microscopic examination and 37% positive following 3 to 5 days incubation at 37 degrees C. Only 3 of the specimens from mothers were positive after the first examination and 4 positive after culture. These high prevalence rates are expected among hospitality girls and are the highest rates thus far reported from the Philippines. These results provide convincing evidence of the value of using cultural methods in determining prevalence rates for Trichomonas vaginalis infections in females.

Leptospirosis in the Philippines.

Leptospirosis was first isolated in the Philippines from one of the human cases of Weil's Disease in 1932 and from then up to 1970 a total of 65 cases were studied sporadically. From 1971 up to 1973 a total of 390 cases were studied out of which 82 (21.02%) were found positive by isolation and serological examination. An outbreak of leptospirosis was followed up from 1976 to 1983 in a penal farm in Sablayan, Mindoro. At the time of the outbreak, 31% of the sera from patients and those with complaints reacted to one or more antigen pools with 40% of these reactors reacting to only one antigen pool. Only one percent of prisoners and residents without complaints reacted to one or more antigen pools, with 80% of these reactors reacting to pool 3 alone. Three and half years later, a first follow-up was done in which 9.07% reacted to one or more pools. Of these, 42% reacted to pool 2 and 82% to pool 3 alone or in combination with other pool. Ninety five percent of those reacting to Pool 2 reacted to L. pyrogenes and 88% of those that reacted to Pool 3 reacted to L. autumnalis. More than three years later after the first follow-up, a second follow up was done. This time 2.29% reacted to one or more pools. Most reacted specifically to L. autumnalis. The residents outside the penal farm were all non-reactive. Sampling from another penal colony and in four agricultural areas revealed no significant reactivity.(ABSTRACT TRUNCATED AT 250 WORDS)

Studies on Echinostoma ilocanum in the Philippines.

Echinostoma ilocanum is widespread among the Philippine Islands but only 3% of over 30,000 stools were found to be positive for eggs. The prevalence rates were highest in Ilocana populations of northern Luzon; 11% of over 7,000 stools examined, however, rates as high as 44% were found. Pila luzonica is eaten uncooked by some people and metacercariae from these snails developed into adults in a number of laboratory animals. Rats, mice and hamsters were the most susceptible with patent infections developing as early as 7 days and lasting for 328 days. There seems to be little disease in humans and animals infected with this parasite.

Attempts to modify treatment of intestinal capillariasis.

Attempts to reduce the standard treatment of intestinal capillariasis of 200 mg mebendazole twice daily for 20 days were not successful. Two of 13 patients given mebendazole at single daily dosages of 500 mg per day for 16 days relapsed 3 to 6 months after treatment. Attempts to reduce the treatment to 14 days were even less satisfactory in that 4 of 8 patients treated experienced relapses 2 to 9 months later. Since intestinal capillariasis can be fatal it is recommended to continue the use of the standard schedule of 400 mg mebendazole daily for 20 days for new cases and 30 days for relapsed cases until a more effective drug is available.

A comparative clinical study of mefloquine and chloroquine in the treatment of vivax malaria.

Forty-six patients with Plasmodium vivax malaria were evaluated for their clinical and parasitological response to the two single doses of mefloquine in comparison with the standard 3-day treatment with chloroquine. The patients were randomly distributed into 3 groups. Group 1 with 13 patients were treated with a single dose 250 mg mefloquine, Group 2 with 15 patients were treated with a single dose 500 mg mefloquine and Group 3 with 18 cases were treated with the standard 3-day treatment course of 1,500 mg chloroquine. All patients in the three treatment group showed similar parasite, clearance time, fever clearance time and clearance time of signs and symptoms. There were no statistically significant differences between any of the groups. The age and weight, as well as the initial parasite count of the three groups were comparable. This study shows that mefloquine at the dosages used were as effective as the 3-day standard treatment with 1,500 mg chloroquine.