title: VIABILITY AND INFECTIVITY OF CRYOPRESERVED PLASMODIUM VIVAX SPOROZOITES.
abstract: Plasmodium vivax presents a great challenge to malaria control because of the ability of its dormant form in the liver, the hypnozoite, to cause relapse in otherwise fully recovered patient. Research efforts to better understand P. vivax hypnozoite biology have been hampered by the limited availability of its sporozoite form responsible for liver infection. Thus, the ability to cryopreserve and recover P. vivax sporozoites is an essential procedure. In this study, protective effects of hydroxyethyl starch (HES) alone and in combination with other cryoprotectants on P. vivax sporozoite recovery, viability and in vitro infectivity of a human liver HC-04 cell line were investigated. Sporozoites were harvested from P. vivax-infected female Anopheles mosquitoes and cryopreserved at a freezing rate of -1°C/minute to a final temperature of -80°C before being stored in a vapor phase liquid nitrogen tank. Cryopreserved sporozoites were thawed at 37°C and recovery of intact sporozoites assessed using a hemocytometer. Sporozoite viability and in vitro infectivity was measured using a gliding and an indirect immunofluorescence assay, respectively. A combination of 10% HES + 50% fetal bovine serum was the best cryopreservant compared to HES solution alone or mixed with cryopreservants such as dimethyl sulfoxide (DMSO) and sucrose. A mixture of bovine serum albumin, DMSO and sucrose in RPMI 1640 medium constituted an alternative cryopreservant. Sporozoites recovered from all cryopreservation media exhibited motility and infectivity of < 0.1% and < 0.001%, respectively. Thus, there is an urgent need for a vast improvement in cryopreservation procedures of viable and infective P. vivax sporozoites necessary for advancing research on hypnozoite biology.
authors.forename: Rapatbhorn
authors.initials: R
authors.lastname: Patrapuvich
authors.forename: Kaewta
authors.initials: K
authors.lastname: Lerdpanyangam
authors.forename: Rachaneeporn
authors.initials: R
authors.lastname: Jenwithisuk
authors.forename: Siriwan
authors.initials: S
authors.lastname: Rungin
authors.forename: Rachasak
authors.initials: R
authors.lastname: Boonhok
authors.forename: Apisak
authors.initials: A
authors.lastname: Duangmanee
authors.forename: Narathatai
authors.initials: N
authors.lastname: Yimamnuaychok
authors.forename: Jetsumon
authors.initials: J
authors.lastname: Sattabongkot
mesh_terms: D000818:Animals; D000852:Anopheles / Q000469:parasitology; D002417:Cattle; D002460:Cell Line; D002465:Cell Movement; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents; D005260:Female; D006801:Humans; D006895:Hydroxyethyl Starch Derivatives; D007303:Insect Vectors / Q000469:parasitology; D008099:Liver / Q000469:parasitology; D016780:Malaria, Vivax / Q000469:parasitology; D010966:Plasmodium vivax / Q000166:cytology* / Q000472:pathogenicity*; D012710:Serum Albumin, Bovine; D034101:Sporozoites / Q000502:physiology*; D014774:Virulence
chemical_list: D003451:Cryoprotective Agents; D006895:Hydroxyethyl Starch Derivatives; D012710:Serum Albumin, Bovine