Epidemiology and molecular genotyping of echinostome metacercariae in Filopaludina snails in Lamphun Province,Thailand

Objective:To analyze the prevalence of echinostome metacercariae in Filopaludina dorliaris(F.dorliaris)and Filopaludina martensi martensi(F.martensi martensi)and genotype variation of echinostome metacercariae by using random amplified polymorphic DNA(RAPD)analysis.Methods:Filopaludina sp.snails were collected from eight localities of Lamphun Province,Northern Thailand and examined for echinostome metacercariae.RAPD-PCR was used lo analyze genotype variation of echinostome metacercariae.Results:A total of 3 226 F.dorliaris and F.martensi martensi snails were collected from eight localities.The overall prevalences of echinostome metacercariae in F.dorliaris and F.martensi martensi were 40.89%and 36.27%,while the intensity of infection was 20.37 and 12.04,respectively.The dendrogram constructed base on RAPD profiles,4 well supported domains were generated;(i)group of metacercariae from Ban Hong,Mae Ta,Meaung,Pa Sang,Toong Hua Chang,and Weang Nong that were clustered in the group of E.revolutum,(ii)Ban Thi,(iii)Lee,and(iv)3 adults of an out group.Condusioiis:This research demonstrated RAPD profiling has been a useful tool to detect DNA polymorphisms to determine genetic relationship between echinostome metacercariae in Lamphun Province,Northern Thailand.

IN VIVO AND IN VITRO ENCYSTMENT OF ECHINOCHASMUS LILIPUTANUS CERCARIAE AND BIOLOGICAL ACTIVITY OF THE METACERCARIAE

Aim To explore if Echinochasmus liliputanus cercariae can develop into metacercariae both invivo and in vitro and biological activity of the metacercariae,then to determine the effects of silver ni-trate on eereariae encystation in vitro.Methods Cereariae of Echinochasmus liliputanus from Bellamyaaeruginosa snails treated with or without silver nitrate were used to infected goldfish,the second inter-mediate host,or to incubated in many different solutions for 24 h to record the encystation rates.Themetacercariae formed both in vivo and in vitro were then used to infected New Zealand rabbits to testtheir infectivity to its definitive hosts or to exeyst in 0.1% sodium deoxycholate excystation medium at37℃ for 1 h.Results In vivo encystment of cercariae occurred in the gills of goldfish.However,thecercariae were also able to encyst in vitro in Locke’s solution,NaCl solution,artificial gastric juice orhuman gastric juice with eneystation rates of 74.28 %,44.94 %,8.37 % and 10.79 %.0.7×-1.2×Locke’s or 0.7%-1.2% NaCl solution was shown to be appropriate for in vitro encystment to occurwithin 24 hour,however,full- strength Locke’s solution was shown to be optimal.The one- day- oldencysted metacereariae formed in vivo showed 88.53 % exeystation when treated in 0.1% sodium de-oxycholate exeystation medium at 37℃ for 1h.The metacercariae formed in vitro,however,showed88.60% and 84.95% excystation for normal and abnormal ones respectively.While abnormal cysts atroom temperature usually die within 10 days,about 70% normal cyst,both in vivo and in vitro,canstill excyst after stored in 0.5×Locke’s at 4℃ for 3 mouths.Cysts formed in vivo and in vitro were e-qually infective to rabbits.1 uM silver nitrate had a dramatic effect on the cercariae encysting in vitro.When treated with silver nitrate,the cereariae encystation rates decreased to 16.25% in Locke’s solu-tion and 6.69% in NaCl solution,however,the encystment was largely restored wben the cercariaewere washed to remove Ag+.Conclusion The finding of E.liliputanus cercariae encysting in vitro,especially in human gastric juice,might be helpful in elucidating mechanisms of the definitive hosts in-fected by the cercariae directly.The encystment of the cercariae in vitro could be inhibited when thecercariae were treated with 1 uM silver nitrate.As silver nitrate binds to the papillae,especially to theciliated papillae,on the cercarial surface,it is suggested that papillar chemoreceptors may be involved inencystment of the cercariae.

Comments on human eurytremiasis in Brazil

Eurytremiasis is an important parasitic disease of cattle that was recently suggested to be a neglected and emerging human disease in Brazil. Based on a misinterpretation of the life cycle of the parasite, it was suggested that a great number of people could be infected with this fluke in the country. In the present letter, aspects of the life cycle ofEurytrema spp. are revisited and clarified. The mechanism of transmission previously reported for the few accidental human cases involved the ingestion of raw or undercooked insects(grasshoppers and crickets) harboring the infective metacercariae. In reality, the zoonotic potential of Eurytrema species is extremely low, and human eurytremiasis is not, and probably never will be, a zoonotic disease in countries where entomophagy is not a common food habit.

Occurrence of Clinostomum complanatum in Aphanius dispar(Actinoptrygii:Cyprinodontidae) collected from Mehran River,Hormuzgan Province,South of Iran

Objective:To investigate the possibility of Aphanius dispar(A.dispar) acting as intermediate host for Clinostomum complanalum(C.complanalum),from Mehran River,Hormuzgan Province of Iran.Methods:During a biological study of A.dispar in Mehran River,Hormuzgan Province,South of Iran,a total of 97 fish specimens were collected in 24 January 2006.Results: 4 specimens(4.12%) including 1 male and 3 female were found infected with C.complanalum metacercaria.These metacercaria were coiled in the epiderm on the body surface of infected fishes.The infection is known as yellow spot disease.The parasite abundance,intensity and prevalence were 0.03%,1.25%and 4.12%,respectively.The infection was higher in females than males.Conclusions:To the best of our knowledge,this is the first report on occurrence of C. complanatum metacercaria in A.dispar in Iran.

Incidence and histopathology of encysted progenetic metacercaria of Clinostomum complanatum(Digenea:Clinostomidae) in Channa punctatus and its development in experimental host

Objective:To study the incidence of encysted progenetic metacercariae of Clinostomum complanatum(C. complanatum) in Channa punctatus(C. punctatus), associated histopathology and the experimental infection to laboratory chicken to obtain ovigerous adult worms.Methods:Live C. punctatus were brought from local fish market of Aligarh, India, dissected and examined on a monthly basis for the presence of C. complanatumcysts. For histochemistry, infected tissue sections with attached cysts were processed for haematoxylene and eosin staining. Cysts were aseptically fed to 4 day old leghorn chicken to obtain adult worms. Mechanically excysted metacercaria and the ovigerous adult worms were stained in carmine to prepare permanent slides.Results:One year survey for the infection of encysted progenetic metacercaria of C.complanatuminC. punctatusrevealed the prevalence, intensity and abundance of 24.7%, 2.27and 0.608, respectively. Histopathology showed heavy infiltration of immune cells at the site of cyst attachment and some tissue damage was also evident. Following feeding to experimental chicken, about 41.07% of the encysted metacercariae were able to excyst and migrate back to bucco-pharyngeal region where they tenaciously attached and fed on blood, and transformed into ovigerous adult worms from 62 hours onwards of post infection.Conclusions:The parasite is potentially pathogenic to the host, and the availability of a suitable intermediate host can be a contributing factor for the occurrence of C. complanatummetacercaria either in the excysted or encysted form, indicating loose host specificity and zoonotic potential.

Experimental observation of effects of triclabendazole on Paragonimus westermani infection in dogs

Objective To observe the therapeutic effects of triclabendazole in the treatment of dogs with Paragonimiasis westermani. Methods Six dogs were experimentally infected, each with 100 metacercariae of Paragonimus westermani intraperitoneally and divided into untreated and treated groups, 3 dogs were treated orally with triclabendazole 100 mg/kg·d 1 for 2 consecutive days on day 170 after infection. Stool egg count was done by Stoll method. All dogs were killed on day 38 after treatment and the number of worm cysts and worms in the lungs were examined by naked eye and microscopically. Results Stool eggs became negative from day 7 to day 14 after starting chemotherapy in treated dogs. The numbers of worm cysts in the lungs of untreated dogs were 18, 24 and 24 on necropsy,while those in the treated dogs were 10, 7 and 4, respectively. The numbers of adult worms in the untreated dogs were 38, 51 and 42, while in the treated dogs only 2 small adult worms were found in one dog and no worm was found in the other two dogs. The mean worm reduction rate was 98.5%. Conclusion Triclabendazole is highly effective against Paragonimus westermani in experimentally infected dogs.