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MONITORING CHLOROQUINE RESISTANCE USING PLASMODIUM FALCIPARUM PARASITES ISOLATED FROM WILD MOSQUITOES IN TANZANIA

Monitoring antimalarial drug resistance is a useful epidemiologic tool and provides early detection of resistance foci. Using DNA extracted from the head/thorax of wild mosquitoes collected from Bagamoyo Coastal Tanzania, samples infected by Plasmodium falciparum (N = 89, in 2002 and N = 249 in 2004) were screened by nested polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) assay for mutations at Pfcrt76 and Pfmdr1-86 associated with chloroquine (CQ) resistance. The majority of isolates were of single infection (71%), and the prevalence of mutant alleles of Pfcrt76 decreased from 64.5% in 2002 to 16% in 2004; likewise, mutant Pfmdr1-86 alleles decreased from 46.6% to 2.7%. Overall, there was a decline of mutant isolates by a factor of 17 and 4 for Pfmdr1 and Pfcrt, respectively. In contrast, isolates with wild-type alleles increased significantly from < 20% in 2002 to 67.6% for Pfcrt76 and 83.5% for Pfmdr1-86 in 2004. This observation suggest a biologic trend of decrease of CQ mutants and a subsequent increase of CQ susceptible parasites in circulation after the discontinued use of CQ in 2001 as a first-line drug in Tanzania. High prevalence of susceptible P. falciparum found in circulation not only supports other reports of a decline of mutant parasites after a reduction of drug selection pressure but suggests that the fitness cost is high in mutant parasites. Typing parasite isolates from infected mosquitoes, an alternative means of data collection, has the potential to increase the spatial and temporal coverage, and this approach is practical in highly endemic regions of Africa.

Received May 15, 2006. Accepted for publication September 6, 2006.

Acknowledgments: The authors thank the Bagamoyo village community for allowing us to collect mosquitoes in their houses; the field team of young men from Bagamoyo who were involved in sampling; the Department of Parasitology and Medical Entomology staff, especially S. Kungulilo and Dr D. Gasarasi; the staff of Department of Vector Ecology and Environment for contributing to this study, and Dr M. Alfrangis of CMP Denmark for providing DNA of different strains of parasites used as positive controls in this study. We are grateful to Drs K. Hirayama, A. Kukuchi, and S. Nakazana at the Institute of Tropical Medicine, Nagasaki University, for providing some of the positive controls and valuable technical advice.

Financial support: This study was supported by the JSPS COE to E.A.T.

Disclaimer: The authors declare that they have no competing interests. The funding sources had no role in study design, data collection, analysis, and interpretation. They had no role in writing this report.

^* Address correspondence to Emmanuel A. Temu, Department of Vector Ecology and Environment, Institute of Tropical Medicine, Nagasaki University, 1-12-4 Sakamoto, Nagasaki-shi 852-8352, Japan. E-mail: emmatemu{at}net.nagasaki-u.ca.jp

Authors’ addresses: Emmanuel A Temu, Nabuko Tuno, Hitoshi Kawada, and Masahiro Takagi, Department of Vector Ecology and Environment, Institute of Tropical Medicine, Nagasaki University, 1-12-4 Sakamoto, Nagasaki-shi 852-8523, Japan, Telephone: 95-849-7809, Fax: 95-849-7812, E-mail: emmatemu{at}net.nagasaki-u.ac.jp, tuno{at}net.nagasaki-u.ac.jp, vergiss{at}net.nagasaki-u.ac.jp, and mstakagi{at}net.nagasaki-u.ac.jp. Issac Kimani, Ministry of Health, PO Box 30016, Nairobi, Kenya, Telephone: 254-020-717077, E-mail: isaackimanidr{at}yahoo.com. Japhet N. Minjas, Department of Parasitology and Medical Entomology, School of Public Health and Social Sciences, Muhimbili University College of Health Sciences, PO Box 65015, Dar es Salaam, Tanzania, Telephone: 255-22-215-3371, Fax: 255-022-215-0465, E-mail: jminjas{at}muchs.ac.tz.

Reprint requests: Masahiro Takagi, Vector Ecology and Environment, Institute of Tropical Medicine, Nagasaki University, 1-12-4 Sakamoto, Nagasaki-shi 852-8523, Japan. E-mail: mstakagi{at}net.nagasaki-u.ac.jp.