Ethiopian Plasmodium vivax hypnozoites formation dynamics and their susceptibility to reference antimalarial drugs

Abstract

One of the key obstacles to malaria elimination is largely attributed to Plasmodium vivax's ability to form resilient hypnozoites in the host liver that cause relapsing infections. As a result, interruption of P. vivax transmission is difficult. P. vivax transmission occurs in Duffy-positive individuals and have been mainly thought to be absent in Africa. However, increasing studies using molecular tools detected P. vivax among Duffy-negative individuals in various African countries. Studies on the African P. vivax has been severely limited because most of malaria control program focus mainly on falciparum malaria. In addition, there is a scarcity of laboratory infrastructures to overcome the biological obstacles posed by P. vivax. Herein, we established field transmission of Ethiopian P. vivax for routine sporozoite supply followed by liver stage infection in Mali. Furthermore, we evaluated local P. vivax hypnozoites and schizonts susceptibilities to reference antimalarial drugs. The study enabled the assessment of local African P. vivax hypnozoite production dynamics. Our data displayed the ability of the African P. vivax to produce hypnozoite forms ex-vivo at different rates per field isolate. We report that while tafenoquine (1µM) potently inhibited both hypnozoites and schizont forms; atovaquone (0.25µM) and the phosphatidylinositol-4-OH kinase (PI4K)-specific inhibitor KDU691 (0.5µM) showed no activity against hypnozoites forms. Unlike hypnozoites forms, P. vivax schizont stages were fully susceptible to both atovaquone (0.25µM) and the (PI4K)-specific inhibitor KDU691 (0.5µM). Together, the data revealed the importance of the local platform for further biological investigation and implementation of drug discovery program on the African P. vivax clinical isolates.

Keywords: African; Drug; Hypnozoite; Invitro; Plasmodium vivax; Susceptibility.

Fig. 1

Fed mosquitoes using P. vivax gametocytes positive blood samples yielded oocytes and sporozoites production. (a)P. vivax sexual and asexual blood stages, (b)P. vivax infected mosquitoes, (c)P. vivax oocyst stages and (d)P. vivax sporozoite stages from salivary glands

Fig. 2

Slow growing referred as hypnozoites and normally developing pre-erythrocytic forms referred as schizonts. Two types of hepatic parasites can be clearly distinguished from days 5 post-infection, in in vitro cultured human primary hepatocytes infected with P. vivax sporozoites. The representative photomicrographs were made on cultures fixed on Day 5 post-infection. The parasite and host nuclei are stained with DAPI (blue), while the parasites are labelled by an antibody specific to the HSP70 of the parasite (green). (a) Slow growing referred as hypnozoites and normally developing pre-erythrocytic forms referred as schizonts are clearly distinguishable; (b) schizonts versus hypnozoites counts for different P. vivax field isolated parasites

Fig. 3

Drugs impact against distinct heptatrienes forms (schizonts versus hypnozoite) of P. vivax field isolated parasites. Atovaquone, tafenoquine and KDU691. DMSO is the control not treated