Using Molecular Diagnostics Based On Internal Transcribed Spacer 2 Sequence To Study Geographical Distribution of Holarctic Malaria Mosquitoes

Diseases like malaria claim the lives of millions of people every year. This deadly disease can result in considerable morbidity, which presently affects countries in Africa, Eurasia, and South America. Anopheline mosquitoes transmit this disease. Studies look at the identification first of the species to accurately determine their distribution. For identification, sequencing of the Internal Transcribed Spacer 2 region of the ribosomal DNA is analyzed. Although African Anophelines are very well studied species, there has been no recent significant research for Holarctic Anophelines. In particular, in North America and Eurasia, because of the eradication of malaria in the northern territories and the focus on other diseases transmitted by Aedes and Culex mosquitoes. In this study, we first look at the Holarctic species Anopheles punctipennis in North America from the Midwest to the eastern United States. Then we look at samples received from Eurasia, in particular Russia and Iran. We physically harvested 500 mosquitoes from ten breeding sites to analyze the identity and distribution of An. punctipennis. We received 110 samples from Russia and 180 samples from Iran to examine the identification and geographic distribution of An. daciae and An. persiensis. Mosquito ITS2 ribosomal DNA was extracted and amplified using polymerase chain reaction (PCR) methods. The PCR products were then sequenced and analyzed based on GenBank information obtained. An analysis by Restriction Fragment Length Polymorphism using ITS2 PCR products on An. punctipennis was conducted. Seven hundred ninety samples were processed to look at the identity and geographic distribution of Holarctic Anophlines. An. puntipennis has no current ITS2 records in GenBank. The distribution of An. daciae and An. persiensis was analyzed by ITS2 and location data. The identity and presence of a malaria vector in new areas or existing areas would prove to be vital if the disease were to re-emerge due to climate changes.