Experimental Study On Infection And Transmission Of Babesia Microti With Three Ticks

To study the capacity of Haemaphysalis longicornis, Boophilus microplus, Rhipicephalus sanguineus as the vector of Babesia microti,we study the infection and transmission of B. microti by gamogenetic H. longicornis and parthenogenetic H. longicornis, Boophilus microplus, Rhipicephalus sanguineus in laboratory conditions,aims to provide scientific basis for human babesiosis control and provention.Therefore,we conduct our researches in3parts.Firstly we study that can B. microti be infected or not by different strains of H. longicornis, Boophilus microplus, Rhipicephalus sanguineus larvaand the infection difference between them in laboratory conditions.Then we study that can B. microti be transmitted or not by H. longicornis nymph to the susceptible host,Balb/c mice.At the end,we study can B. microti be trans-stadially transmitted or transovarian transmission or not by H.longicornis during its lifecycle. AT first,the B. microti positive mice with high infection rate should be acquired through intraperitoneally injecting Balb/c mice with red blood cells infected with B. microti.Then these B. microti positive Balb/c mice were bited by different strains of H. longicornis, Boophilus microplus, Rhipicephalus sanguineus larve in the same condiions.while larve were engorged,they were kept in constant temperature and humidity chamber for inoculation.During the peroid of inoculation,randomly sampled larve for detecting infection rate of B.microti at the1th,3th,5th,7th,9th,11th,13th and20th day.we considered the day when larve was engorged to drop from the mice as the first day.The tick which were sampled at13th and20th had inoculated to nymph.The result of this part of research implied that both gamogenetic H. longicornis and parthenogenetic H. longicornis, Boophilus microplus, Rhipicephalus sanguineus could be infected by B. microti in laboratory conditions,and the infection rate between them had no difference statistically.lt also revealed that the infection rate had a declining trend as time went on.Besides, B. microti could be detected in nymphal ticks by nested PCR, implying that B. microti may can be trans-stadially transmit during the inoculation period from larval ticks to nymphal ticks.Of course,there could be another situation that B. microti detected in nymph may be dead,or gene fragments of B. microti.In the second part of research,the nymphal ticks which were inoculated was our studying objects which were inoculated from larval ticks of part one research.Negative Balb/c mice were bited by these nymph to study these nymph’s ability of transmit B. microti to mice.In this study,3mice were bited by60gamogenetic nymph and2mice were bited by40parthenogenetic, each mouse was bited by20nymph.After these5mice were bited for56h,the ticks were removed from mice and kept in75%ethanol for detecting B. microti by nested PCR.These mice were numbered.These mice were detected the infection of B. micrtoi every week through Giemsa-stained examination of thin blood smears,nested PCR and FTA cards detection method.The result revealed that the infection rate of different strains of H. longicornis had no difference.fi. microti could be detected through nested PCR and FTA cards detection method in one mice at the second week which were bited by gamogenetic nymph,and B. microti could be detected in the Giemsa-stained blood smears.This result nidicated that B.microti could be transmitted to mice by H.longicornis nymph.B. microti could not be detected in other four mice in four weeks,but at the seventh week,these5mice were detected again,6. microti was detected in2mice including one that was acertain at the second week,another mouse in which B.microti was detected B elongde to the group of mice bited by parthenogenetic H. longicornis.Through the two results,we acertained that both gamogenetic nymph and parthenogenetic mice could transmit B. microti to mice and also provided us a new clue to study the vector of B. microti in wild area.Through the study of infection and transmission of B. microti by H.longicornis,,we constructed the third part research,in order to search that H. longicornis nymph could trans-stadially transmit and transovarian transmit6. microti or not.So New Zealand rabbits were bited by nymph of second part research, then the engorged nymph hacthed to adult ticks.New Zealand rabbits were bited by adult ticks again.When adult ticks spawned, B. microti was detected in adult ticks through nested PCR.34adult ticks were detected by nested PCR,B.microti was detected in6 adult ticks with a infection rate of6.67%.It indicated that nymph could trans-stadially transmit B. microti to adult ticks.Then B. microti was detected in the larva inoculated from the eggs of positive adult ticks,a total of300larva were random samples and were detected by nested PCR.Among them,none was detected as B. microti positive.This result indicated that B. microti could not be transovarian transmitted by H. longicornis adult ticks.Through3parts of research,we found that H. longicornis, Boophilus microplus, Rhipicephalus sanguineus larval ticks could be infected by B. microti and B. microti could be trans-stadially transmitted to nymphal ticks in laboratory conditions.B. microti could be transmitted to susceptible Balb/c mice and trans-stadially transmitted to adult ticks by H. longicornis nymphal ticks.B. microti could also be transovarian transmitted to larva by H. longicornis.There was few research about the vector of B. microti in China,our research provided a new clue to acertain the vector of B. microti in China wild area and a meaningful mechanism for human babesiosis control and provention.