We report the natural co-infection of a single Anopheles mosquito with Plasmodium vivax Grassi & Feletti phenotypes VK210 and VK247. In total, 8,452 anopheline mosquitoes collected between June 1999 and July 2001 were tested by ELISA for the presence of circumsporozoite (CS) protein to VK210, VK247, and P. falciparum (Welch) (PF). A total of 29 species was represented; however, the predominant species tested were A. minimus Theobald (4,632), A. sawadwongporni Rattanarithikul & Green (1,248), A. maculatus Theobald (1,201), A. campestris Reid (478), and A. barbirostris Van der Wulp (391). A total of 17 positive mosquitoes was identified by ELISA, and included the following: A. minimus infected with VK210 (5), PF (3), and both VK210 and VK247 (1), A. maculatus infected with VK210 (1), VK247 (1), and both VK210 and VK247 (1), A. campestris infected with VK210 (2), A. sawadwongporni infected with VK247 (1) and PF (1), and A. hodgkini Reid infected with VK247 (1). This is the first report of a single mosquito naturally infected with both VK210 and VK247.
As part of an on-going malaria surveillance effort conducted by the U.S. Forces Korea, Republic of Korea (ROK), a total of 28,286 anopheline mosquitoes was tested for the presence of Plasmodium vivax circumsporozoite (CS) protein. Mosquitoes were collected (using a variety of light and baited traps) from 29 locations throughout the ROK (the majority were collected near the de-militarized zone), identified to species, and tested by enzyme-linked immunosorbent assay for the presence of P. vivax 210 and P. vivax 247 CS protein. Recent evidence suggests that characters used to separate Anopheles sinensis Wiedemann from An. lesteri Baisas & Hu are unreliable; therefore, the data have been analyzed by grouping these two species. A total of 25,365 Anopheles sinensis/lesteri, 2,890 An. yatsushiroensis Miyazaki, and 31 An. sineroides Yamada was tested. Of these, one pool of 10 An. sinensis/lesteri collected on 9 September 1999 at Camp Howze and one pool of nine An. sinensis/ lesteri collected on 13 September 1999 at Camp Bonifas were positive for P. vivax 247.
We evaluated the performance of the VecTest Malaria Antigen Panel (V-MAP) assay for the detection of Plasmodium falciparum and P. vivax (variants 210 and 247) circumsporozoite protein in anopheline mosquitoes in Thailand. The V-MAP assay is a rapid, one-step procedure using a 'dipstick' wicking test strip. The circumsporozoite (CS) ELISA was used as the reference standard. Mosquitoes evaluated in the study included field-collected specimens (n = 930) and laboratory-reared specimens that had been fed on blood collected from patients with and without Plasmodium gametocytes (n = 4,110) or on cultured P. falciparum gametocytes (n = 262). Field-collected mosquitoes were triturated individually or in pools of 2-5 and tested using 613 V-MAP assays. Laboratory-reared specimens were tested individually using 4,372 V-MAP assays. Assay performance depended on the species of Plasmodium and the number of sporozoites used as the cut-off. For P. falciparum, optimal performance was achieved using a cut-off of 150 sporozoites (sensitivity = 100%, specificity = 99.2%, and accuracy = 0.99). For P. vivax variant 210, optimal performance was also achieved using a cut-off of 150 sporozoites (sensitivity = 94.8%, specificity = 94.5%, and accuracy = 0.95). We were unable to develop a standard-curve for the CS-ELISA using P. vivax variant 247 because of a lack of sporozoites; however, using a cut-off of 30 pg P. vivax 247 antigen (mosquitoes with less than this amount of antigen were considered negative), assay performance (sensitivity = 94.3%, specificity = 99.2%, and accuracy = 0.99) was comparable to that achieved for P. falciparum and P. vivax 210. These results clearly demonstrate that the V-MAP assay performs at an acceptable level and offers practical advantages for field workers needing to make rapid surveys of malaria vectors.
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