Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups
Abstract Background Genus Anopheles mosquitoes are the primary vectors of human malaria, which is a serious threat to public health worldwide. To reduce the spread of malaria and identify the malaria infection rates in mosquitoes, accurate species identification is needed. Malaria re-emerged in 1993...
| Main Authors: | , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2021-06-01
|
| Series: | Malaria Journal |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12936-021-03808-w |
| _version_ | 1819034918078906368 |
|---|---|
| author | Woo Jun Bang Heung Chul Kim Jihun Ryu Hyeon Seung Lee So Youn Lee Myung Soon Kim Sung Tae Chong Terry A. Klein Kwang Shik Choi |
| author_facet | Woo Jun Bang Heung Chul Kim Jihun Ryu Hyeon Seung Lee So Youn Lee Myung Soon Kim Sung Tae Chong Terry A. Klein Kwang Shik Choi |
| author_sort | Woo Jun Bang |
| collection | DOAJ |
| description | Abstract Background Genus Anopheles mosquitoes are the primary vectors of human malaria, which is a serious threat to public health worldwide. To reduce the spread of malaria and identify the malaria infection rates in mosquitoes, accurate species identification is needed. Malaria re-emerged in 1993 in the Republic of Korea (ROK), with numbers peaking in 2004 before decreasing to current levels. Eight Anopheles species (Anopheles sinensis, Anopheles pullus, Anopheles belenrae, Anopheles lesteri, Anopheles kleini, Anopheles sineroides, Anopheles koreicus, Anopheles lindesayi) are distributed throughout Korea. Members of the Anopheles Hyrcanus group currently cannot be identified morphologically. The other species of Anopheles can be identified morphologically, except when specimens are damaged in traps. The purpose of this study was to develop a rapid and accurate method for simultaneous molecular identification of the eight Anopheles species present in the ROK. Methods Anopheles spp. used in this study were collected near/in the demilitarized zone in ROK, where most malaria cases are reported. DNA from 165 of the Anopheles specimens was used to develop a multiplex PCR assay. The internal transcribed spacer 2 (ITS2) region of each species was sequenced and analysed for molecular identification. Results DNA from a total of 165 Anopheles specimens was identified to species using a multiplex diagnostic system. These included: 20 An. sinensis, 21 An. koreicus, 17 An. lindesayi, 25 An. kleini, 11 An. lesteri, 22 An. sineroides, 23 An. belenrae, and 26 An. pullus. Each species was clearly distinguished by electrophoresis as follows: 1,112 bp for An. sinensis; 925 bp for An. koreicus; 650 bp for An. lindesayi; 527 bp for An. kleini; 436 bp for An. lesteri; 315 bp for An. sineroides; 260 bp for An. belenrae; and, 157 bp for An. pullus. Conclusion A multiplex PCR assay was developed to identify Anopheles spp. distributed in ROK. This method can be used to accurately identify Anopheles species that are difficult to identify morphologically to determine species distributions and malaria infection rates. |
| first_indexed | 2024-12-21T07:41:22Z |
| format | Article |
| id | doaj.art-0fb43a7b52b5406f81a01a6b1e298595 |
| institution | Directory Open Access Journal |
| issn | 1475-2875 |
| language | English |
| last_indexed | 2024-12-21T07:41:22Z |
| publishDate | 2021-06-01 |
| publisher | BMC |
| record_format | Article |
| series | Malaria Journal |
| spelling | doaj.art-0fb43a7b52b5406f81a01a6b1e2985952022-12-21T19:11:18ZengBMCMalaria Journal1475-28752021-06-012011710.1186/s12936-021-03808-wMultiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groupsWoo Jun Bang0Heung Chul Kim1Jihun Ryu2Hyeon Seung Lee3So Youn Lee4Myung Soon Kim5Sung Tae Chong6Terry A. Klein7Kwang Shik Choi8School of Life Sciences, BK21 FOUR KNU Creative BioResearch Groups, Kyungpook National UniversityForce Health Protection and Preventive Medicine, Medical Department Activity-Korea/65th Medical Brigade, Unit 15281School of Life Sciences, BK21 FOUR KNU Creative BioResearch Groups, Kyungpook National UniversitySchool of Life Sciences, BK21 FOUR KNU Creative BioResearch Groups, Kyungpook National UniversitySchool of Life Sciences, BK21 FOUR KNU Creative BioResearch Groups, Kyungpook National UniversityForce Health Protection and Preventive Medicine, Medical Department Activity-Korea/65th Medical Brigade, Unit 15281Force Health Protection and Preventive Medicine, Medical Department Activity-Korea/65th Medical Brigade, Unit 15281Force Health Protection and Preventive Medicine, Medical Department Activity-Korea/65th Medical Brigade, Unit 15281School of Life Sciences, BK21 FOUR KNU Creative BioResearch Groups, Kyungpook National UniversityAbstract Background Genus Anopheles mosquitoes are the primary vectors of human malaria, which is a serious threat to public health worldwide. To reduce the spread of malaria and identify the malaria infection rates in mosquitoes, accurate species identification is needed. Malaria re-emerged in 1993 in the Republic of Korea (ROK), with numbers peaking in 2004 before decreasing to current levels. Eight Anopheles species (Anopheles sinensis, Anopheles pullus, Anopheles belenrae, Anopheles lesteri, Anopheles kleini, Anopheles sineroides, Anopheles koreicus, Anopheles lindesayi) are distributed throughout Korea. Members of the Anopheles Hyrcanus group currently cannot be identified morphologically. The other species of Anopheles can be identified morphologically, except when specimens are damaged in traps. The purpose of this study was to develop a rapid and accurate method for simultaneous molecular identification of the eight Anopheles species present in the ROK. Methods Anopheles spp. used in this study were collected near/in the demilitarized zone in ROK, where most malaria cases are reported. DNA from 165 of the Anopheles specimens was used to develop a multiplex PCR assay. The internal transcribed spacer 2 (ITS2) region of each species was sequenced and analysed for molecular identification. Results DNA from a total of 165 Anopheles specimens was identified to species using a multiplex diagnostic system. These included: 20 An. sinensis, 21 An. koreicus, 17 An. lindesayi, 25 An. kleini, 11 An. lesteri, 22 An. sineroides, 23 An. belenrae, and 26 An. pullus. Each species was clearly distinguished by electrophoresis as follows: 1,112 bp for An. sinensis; 925 bp for An. koreicus; 650 bp for An. lindesayi; 527 bp for An. kleini; 436 bp for An. lesteri; 315 bp for An. sineroides; 260 bp for An. belenrae; and, 157 bp for An. pullus. Conclusion A multiplex PCR assay was developed to identify Anopheles spp. distributed in ROK. This method can be used to accurately identify Anopheles species that are difficult to identify morphologically to determine species distributions and malaria infection rates.https://doi.org/10.1186/s12936-021-03808-wAnophelesMultiplex PCR assayMalariaKorea |
| spellingShingle | Woo Jun Bang Heung Chul Kim Jihun Ryu Hyeon Seung Lee So Youn Lee Myung Soon Kim Sung Tae Chong Terry A. Klein Kwang Shik Choi Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups Malaria Journal Anopheles Multiplex PCR assay Malaria Korea |
| title | Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups |
| title_full | Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups |
| title_fullStr | Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups |
| title_full_unstemmed | Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups |
| title_short | Multiplex PCR assay for the identification of eight Anopheles species belonging to the Hyrcanus, Barbirostris and Lindesayi groups |
| title_sort | multiplex pcr assay for the identification of eight anopheles species belonging to the hyrcanus barbirostris and lindesayi groups |
| topic | Anopheles Multiplex PCR assay Malaria Korea |
| url | https://doi.org/10.1186/s12936-021-03808-w |
| work_keys_str_mv | AT woojunbang multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT heungchulkim multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT jihunryu multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT hyeonseunglee multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT soyounlee multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT myungsoonkim multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT sungtaechong multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT terryaklein multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups AT kwangshikchoi multiplexpcrassayfortheidentificationofeightanophelesspeciesbelongingtothehyrcanusbarbirostrisandlindesayigroups |