Home / Regular Issue / JTAS Vol. 45 (3) Aug. 2022 / JTAS-2424-2022


Molecular Identification and Species Richness of Flies (Diptera) and Their Associated Bovidae Hosts at Cattle Farms in Selangor, Malaysia

Salmah Yaakop, Puteri Amira Amiruddin, Muhamad Azmi Mohammed, Aqilah Sakinah Badrulisham, Nadiatur Akmar Zulkifli and Mohd Noor Hisham Mohd Nadzir

Pertanika Journal of Tropical Agricultural Science, Volume 45, Issue 3, August 2022

DOI: https://doi.org/10.47836/pjtas.45.3.05

Keywords: Blood-sucking insect, COI, cytb, DNA barcode, fly, host, livestock, Malaysia

Published on: 8 August 2022

Flies (Diptera) play a significant role in the ecosystem as pollinators and decomposers, and they are also important vermin and disease vectors. Studies on the dipteran species are still lacking in Malaysia; therefore, the dipteran species’ biology, morphology, distribution, and abundance are necessary. The objectives of this study were to identify dipteran species using a molecular approach, determine flies’ Bovidae hosts, and investigate the diversity of the fly’s species at three different cattle farms purposively selected in Selangor, Malaysia. The fly species were identified using cytochrome oxidase subunit I (COI) (Haematopota javana, Tabanus rubidus, Tabanus fontinalis, Iranihindia martellata, Musca domestica, and Chrysomya megacephala), while another six species only up to genus level ( Haematopota sp. 1, Musca sp. 1, Asilus sp., Metopia sp., Anasillomos sp., and Ommatius sp.). In addition, two species of hosts: Bos indicus and Bos taurus, were proven to be the associated host species for the dipteran species based on molecular data of cytochrome b (cytb). However, there were no significant differences between farms in species diversity and richness (F = 1.262, df = 2, p = 0.2459 >0.05). Interestingly, the two most dominant dipteran genera collected from the cattle farms were Musca and Chrysomya. At the same time, its abundance may have been influenced by the structure of the cattle cage flooring, which serves as a breeding site and food source. These findings contribute to fundamental epidemiological data in developing control strategies for dipteran species and are of great economic and health importance to livestock production in Malaysia.

  • Adler, P. H., Cheke, R. A., & Post, R. J. (2010). Evolution, epidemiology, and population genetics of black flies (Diptera: Simuliidae). Infection, Genetics and Evolution, 10(7), 846-865. https://doi.org/10.1016/j.meegid.2010.07.003

  • Alcaide, M., Rico, C., Ruiz, S., Soriguer, R., Muñoz, J., & Figuerola, J. (2009). Disentangling vector-borne transmission networks: A universal DNA barcoding method to identify vertebrate hosts from arthropod bloodmeals. PLOS One, 4(9), e7092. https://doi.org/10.1371/journal.pone.0007092

  • Al-Shaibani, I. R. M., & Al-Mahdi, H. (2014). Seasonal abundance of flies (Diptera) in animal farms in some areas of Dhamar governorate, Yemen. Yemeni Journal of Agriculture and Veterinary Sciences, 1(2), 11-21. https://doi.org/10.12816/0007145

  • Al-Talafha, H. A., Yaakop, S., & Idris, A. B. (2017a). Two new species and seven new records of horse fly (Diptera: Tabanidae) from Malaysia, including a description of new species and modified keys. Journal of Medical Entomology, 55(1), 112-121. https://doi.org/10.1093/jme/tjx172

  • Al-Talafha, H. A., Yaakop, S., & Idris, A. B. (2017b). Taxonomic notes and new records of the genus Tabanus Linnaeus 1758 (Diptera: Tabanidae) from Malaysia. In AIP Conference Proceedings (Vol. 1784, No. 1, p. 060014). AIP Publishing LLC. https://doi.org/10.1063/1.4966852

  • Baldacchino, F., Desquesnes, M., Mihok, S., Foil, L., Duvallet, D. G., & Jittapalapong, S. (2014). Tabanids: Neglected subjects of research, but important vectors of disease agent!. Infection, Genetics and Evolution, 28, 596-615. https://doi.org/10.1016/j.meegid.2014.03.029

  • Banarjee, D., Kumar, V., Maity, A., Ghosh, B., Tyagi, K., Singha, D., Kundu, S., Laskar, B. A., Naskar, A., & Rath, S. (2015). Identification through DNA barcoding of Tabanidae (Diptera) vectors of surra disease in India. Acta Tropica, 150, 52-58. https://doi.org/10.1016/j.actatropica.2015.06.023

  • Banasik, M., Stanisławska-Sachadyn, A., & Sachadyn, P. (2016). A simple modification of PCR thermal profile applied to evade persisting contamination. Journal of Applied Genetics, 57(3), 409-415. https://doi.org/10.1007/s13353-015-0336-z

  • Barrett, R. D., & Hebert, P. D. (2005). Identifying spiders through DNA barcodes. Canadian Journal of Zoology, 83(3), 481-491. https://doi.org/10.1139/z05-024

  • Boessenkool, S., Epp, L. S., Haile, J., Bellemain, E., Edwards, M., Coissac, E., Willerslev, E., & Brochmann, C. (2012). Blocking human contaminant DNA during PCR allows amplification of rare mammal species from sedimentary ancient DNA. Molecular Ecology, 21(8), 1806-1815. https://doi.org/10.1111/j.1365-294X.2011.05306.x

  • Brockerhoff, E. G., Barbaro, L., Castagneyrol, B., Forrester, D. I., Gardiner, B., Gonzalez-Olabarria, J. R., Lyver, P. O. B., Meurisse, N., Oxbrough, A., Taki, H., & Thompson, I. D. (2017). Forest biodiversity, ecosystem functioning and the provision of ecosystem services. Biodiversity and Conservation, 26, 3005-3035. https://doi.org/10.1007/s10531-017-1453-2

  • Brown, B. V. (2001). Flies, gnats, and mosquitoes. In S. Levin (Ed.), Encyclopedia of biodiversity (pp. 815-826). Academic Press.

  • Chaiwong, T., Srivoramas, T., Sueabsamran, P., Sukontason, K., Sanford, M. R., & Sukontason, K. L. (2014). The blow fly, Chrysomya megacephala, and the house fly, Musca domestica, as mechanical vectors of pathogenic bacteria in Northeast Thailand. Tropical Biomedicine, 31(2), 336-346.

  • Changbunjong, T., Sedwisi, P., Weluwanarak, T., Nitiyamatawat, E., Sariwongchan, R., & Chareonviriyaphap, T. (2018). Species diversity and abundance of Tabanus spp. (Diptera: Tabanidae) in different habitats of Thailand. Journal of Asia-Pacific Entomology, 21(1), 134-139. https://doi.org/10.1016/j.aspen.2017.11.013

  • Clark, A. R., Armstrong, K. F., Carmichael, A. E., Milne, J. R., Raghu, S., Roderick, G. K., & Yeates, D. K. (2005). Invasive phytophagous pests arising through a recent tropical evolutionary radiation: The Bactrocera dorsalis complex of fruit flies. Annual Review of Entomology, 50, 293-319. https://doi.org/10.1146/annurev.ento.50.071803.130428

  • Colless, D. H., & McAlpine, D. K. (1991). Diptera (flies). In I. D. Nauman (Ed.), The insects of Australia: A textbook for students and research workers (pp. 656-740). Melbourne University Press.

  • Courtney, G. W., Pape, T., Skevington, J. H., & Sinclair, B. J. (2017). Biodiversity of Diptera. In R. G. Foottit & P. H. Adler (Eds.), Insect biodiversity: Science and society (2nd ed., pp. 229-278). John Wiley & Sons. https://doi.org/10.1002/9781118945568.ch9

  • Cywinska, A., Hannan, M. A., Kevan, P. G., Roughley, R. E., Iranpour, M., & Hunter, F. F. (2010). Evaluation and identification of new haplomorphs in Canadian deerflies and horseflies. Medical and Veterinary Entomology, 24(4), 382-410. https://doi.org/10.1111/j.1365-2915.2010.00896.x

  • della Torre, A., Costantini, C., Besansky, N. J., Caccone, A., Petrarca, V., Powell, J. R., & Coluzzi, M. (2002). Speciation within Anopheles gambiae - The glass is half full. Science, 298(5591), 115-117. https://doi.org/10.1126/science.1078170

  • Ernieenor, F. C. L., Yaakop, S., Ahamad., M., & Md Nor, S. (2015). Molecular identification of blood meal sources of ticks (Acari, Ixodidae) using cytochrome b gene as a genetic marker. ZooKeys, (478), 27-43. https://doi.org/10.3897/zookeys.478.8037

  • Erwanas, A. I., Masrin, A., Chandrawathani, P., Jamnah, O., Premaalatha, B., & Ramlan, M. (2015). Vectors of veterinary importance in Malaysia: A survey of biting flies in relation to trypanosomiasis in Perak. Malaysian Journal of Veterinary Research, 6, 89-96.

  • Ferraz, A. C. P., Proença, B., Gadelha, B. Q., Faria, L. M., Barbalho, M. G. M., Aguiar-Coelho, V. M., & Lessa, C. S. S. (2010). First record of human myiasis caused by association of the species Chrysomya megacephala (Diptera: Calliplioridae), Sarcophaga (Liopygia) ruficornis (Diptera: Sarcophagidae), and Musca domestica (Diptera: Muscidae). Journal of Medical Entomology, 47(3), 487-490.

  • Foil, L. D. (1989). Tabanids as vectors of disease agents. Parasitology Today, 5(3), 88-96. https://doi.org/10.1016/0169-4758(89)90009-4

  • Folmer, O., Black, M. & Hoeh, W. (1994). DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Molecular Marine Biology and Biotechnology, 3(5), 294–299.

  • Gerhardt, R. R., & Hribar, L. J. (2019). Flies (Diptera). In G. Mullen & L. Durden (Eds.), Medical and veterinary entomology (3rd ed., pp. 171-190). Academic Press. https://doi.org/10.1016/b978-0-12-814043-7.00011-x

  • Halim, M., Aman-Zuki, A., & Yaakop, S. (2017). DNA barcoding and relationships of eight ladybugs species (Coleoptera: Coccinellidae) that infesting several crops from Peninsular Malaysia. Journal of Asia-Pacific Entomology, 20(3), 814-820. https://doi.org/10.1016/j.aspen.2017.05.009

  • Halim, M., Aman-Zuki, A., Syarifah Zulaikha, S. A., Muhaimin, A. M. D., Atikah, A. R., Mazmira, M. M. M., Md-Zain, B. M., & Yaakop, S. (2018). Exploring the abundance and DNA barcode information of eight parasitoid wasps species (Hymenoptera), the natural enemies of the important pest of oil palm, bagworm, Metisa plana (Lepidoptera: Psychidae) toward the biocontrol approach and it’s application. Journal of Asia-Pacific Entomology, 21(4), 1359-1365. https://doi.org/10.1016/j.aspen.2018.10.012

  • Hall, M. J., Wall, R. L., & Stevens, J. R. (2016). Traumatic myiasis: A neglected disease in a changing world. Annual Review of Entomology, 61, 159-176. https://doi.org/10.1146/annurev-ento-010715-023655

  • Hammer, Ø., Harper, D. A. T., & Ryan, P. D. (2001). PAST: Paleontological statistics software package for education and data analysis. Palaeontologia Electronica, 4(1), 9.

  • Hebert, P. D., Cywinska, A., Ball, S. L., & DeWaard, J. R. (2003). Biological identifications through DNA barcodes. Proceedings of the Royal Society of London. Series B: Biological Sciences, 270(1512), 313-321. https://doi.org/10.1098/rspb.2002.2218

  • Hemmatinezhad, B., Ommi, D., Hafshejani, T. T., & Khamesipour, F. (2015). Molecular detection and antimicrobial resistance of Pseudomonas aeruginosa from houseflies (Musca domestica) in Iran. Journal of Venomous Animals and Toxins including Tropical Diseases, 21, 18. https://doi.org/10.1186/s40409-015-0021-z

  • Hussein, M., Pillai, V. V., Goddard, J. M., Park, H. G., Kothapalli, K. S., Ross, D. A., Ketterings, Q. M., Brenna, J. T., Milstein, M. B., Marquis, H., Johnson, P. A., Nyrop, J. P., & Selvaraj, V. (2017). Sustainable production of housefly (Musca domestica) larvae as a protein-rich feed ingredient by utilizing cattle manure. PLOS One, 12(2), e0171708. https://doi.org/10.1371/journal.pone.0171708

  • International Atomic Energy Agency. (2009). Selection and breeding of cattle and buffalo in Asia: Strategies and criteria for improved breeding. IAEA.

  • Issa, R. (2019). Musca domestica acts as transport vector hosts. Bulletin of the National Research Centre, 43, 73. https://doi.org/10.1186/s42269-019-0111-0

  • Khamesipour, F., Lankarani K. B., Honarvar B., & Kwenti, T. E. (2018). A systematic review of human pathogens carried by the housefly (Musca domestica L.). BMC Public Health, 18, 1049. https://doi.org/10.1186/s12889-018-5934-3

  • Khan, H. A., Shad, S. A., & Akram, W. (2012). Effect of livestock manures on the fitness of house fly, Musca domestica L. (Diptera: Muscidae). Parasitology Research, 111(3), 1165-1171. https://doi.org/10.1007/s00436-012-2947-1

  • Khofar P. N., Kurahashi, H., Zainal, N. S., Isa, M. S., & Heo, C. C. (2019). Diversity of coprophilic Diptera associated with buffalo dung in Selangor, Malaysia. Journal of Wildlife and Parks, 34(1), 63-80.

  • Kocher, T. D., Thomas, W. K., Meyer, A., Edwards, S. V., Paabo, S., Villablanca, F. X., & Wilson, A. C. (1989). Dynamics of mitochondrial DNA evolution in animals: Amplification and sequencing with conserved primers. Proceedings of the National Academy of Sciences of the United States of America, 86(16), 6196-6200. https://doi.org/10.1073%2Fpnas.86.16.6196

  • Kurahashi, H., Benjaphong, N., & Omar, B. (1997). Blow flies (Insecta: Diptera: Calliphoridae) of Malaysia and Singapore. Raffles Bulletin of Zoology, 45, 88-91.

  • Lee, P. S., Sing, K. W., & Wilson, J. J. (2015). Reading mammal diversity from flies: The persistence period of amplifiable mammal mtDNA in blowfly guts (Chrysomya megacephala) and a new DNA mini-barcode target. PLOS One, 10(4), e0123871. https://doi.org/10.1371/journal.pone.0123871

  • Lysyk, J., & Axtell, R. C. (1986). Field evaluation of three methods for monitoring populations of house flies (Musca domestica) (Diptera: Muscidae) and other filth flies in three types of poultry housing systems. Journal of Economic Entomology, 79(1), 136-151. https://doi.org/10.1093/jee/79.1.144

  • Marshall, S. A. (2012). Flies: The natural history and diversity of Diptera. Firefly Books Press Ltd.

  • Molaei, G., Andreadis, T. G., Armstrong, P. H., & Diuk-Wasser, M. (2008). Host-feeding patterns of potential mosquito vectors in Connecticut, USA: Molecular analysis of bloodmeals from 23 species of Aedes, Anopheles, Culex, Coquillettidia, Psorophora, and Uranotaenia. Journal of Medical Entomology, 45(6), 1143-1151. https://doi.org/10.1093/jmedent/45.6.1143

  • Morinière, J., Balke, M., Doczkal, D., Geiger, M. F., Hardulak, L. A., Haszprunar, G., Hausmann, A., Hendrich, L., Regalado, L., Rulik, B., Schmidt, S., Wägele, J. W., & Hebert, P. D. N. (2019). A DNA barcode library for 5,200 German flies and midges (Insecta: Diptera) and its implications for metabarcoding‐based biomonitoring. Molecular Ecology Resources, 19(4), 900-928. https://doi.org/10.1111/1755-0998.13022

  • Morita, S. I., Bayless, K. M., Yeates, D. K., & Wiegmann, B. M. (2016). Molecular phylogeny of the horse flies: A framework for renewing tabanid taxonomy. Systematic Entomology, 41(1), 56-72. https://doi.org/10.1111/syen.12145

  • Mullens, B. A. (2009). Horse flies and deer flies. In G. Mullen & L. Durden (Eds.), Medical and veterinary entomology (pp. 264-276). Academic Press.

  • Newton, B. (2006, December 14). Robber flies. https://www.uky.edu/Ag/CritterFiles/casefile/insects/flies/robber/robber.htm

  • Nihei, S., & de Carvalho, C. J. B. (2009). The Muscini flies of the world (Diptera: Muscidae): Identification key and generic diagnoses. Zootaxa, 1976(1), 1-24. https://doi.org/10.11646/zootaxa.1976.1.1

  • Papanastasis, V. P., Bautista, S., Chouvardas, D., Mantzanas, K., Papadimitriou, M., Mayor, A. G., Koukioumi, P., Papaioannou, A., & Vallejo, R. V. (2017). Comparative assessment of goods and services provided by grazing regulation and reforestation in degraded Mediterranean rangelands. Land Degradation and Development, 28(4), 1178-1187. https://doi.org/10.1002/ldr.2368

  • Pape, T. (2009). Economic importance of Diptera. In B. V. Brown, A. Borkent, J. M. Cumming, D. M. Wood, N. E. Woodley, & M. A. Zumbado (Eds.), Manual of Central American Diptera (pp. 65-77). CRC Press.

  • Patton, W. S. (1922). Notes on the species of the genus Musca, Linnaeus – Part I. Bulletin of Entomological Research, 12(4), 411-426. https://doi.org/10.1017/S0007485300045065

  • Phasuk, J., Tharawoot, T., Beaver, R. A., & Jittapalapong, S. (2011). Seasonal abundance of Tabanidae (Diptera) on dairy farms in Saraburi Province, Thailand. Thai Journal of Agricultural Science, 44(3), 175-181.

  • Renaud, A. K., Savage, J., & Adamowicz, S. J. (2012). DNA barcoding of Northern Nearctic Muscidae (Diptera) reveals high correspondence between morphological and molecular species limits. BMC Ecology, 12, 24. https://doi.org/10.1186/1472-6785-12-24

  • Rivera, J., & Currie, D. C. (2009). Identification of Nearctic black flies using DNA barcodes (Diptera: Simuliidae). Molecular Ecology Resources, 9(S1), 224-236. https://doi.org/10.1111/j.1755-0998.2009.02648.x

  • Schumann, H. (1992). Systematische Gliederung der Ordnung Diptera mit besonderer Beru ̈cksichtigung der in Deutschland vorkommenden Familien [Systematic classification of the order Diptera with special attention to the families found in German country]. Deutsche Entomologische Zeitschrift, 39, 103-116.

  • Sharanya, B., & Zuha, R. M. (2019). A preliminary geometric morphometrics assessment of two forensically important blow fly larvae in Malaysia, Chrysomya megacephala (Fabricius) and Chrysomya rufifacies (Macquart) (Diptera: Calliphoridae). Serangga, 24(1), 70-79.

  • Shariff, S., Ibrahim, N. J., Md-Zain, B. M., Idris, A. B., Suhana, Y., Roff, M. N., & Yaakop, S. (2014). Multiplex PCR in determination of Opiinae parasitoids of fruit flies, Bactrocera sp., infesting star fruit and guava. Journal of Insect Science, 14, 7.

  • Singh, D., & Bharti, M. (2000). Forensically important blow flies (Diptera: Calliphoridae) of Punjab (India). Uttar Pradesh Journal of Zoology, 20(3), 249-251.

  • Skevington, J. H., & Dang, P. T. (2002). Exploring the diversity of flies (Diptera). Biodiversity, 3(4), 3-27. https://doi.org/10.1080/14888386.2002.9712613

  • Slama, D., Haouas, N., Mezhoud, H., Babba, H., & Chaker, E. (2015). Blood meal analysis of Culicoides (Diptera: ceratopogonidae) in central Tunisia. PLOS One, 10(3), e0120528. https://doi.org/10.1371/journal.pone.0120528

  • Ssymank, A., Kearns, C. A., Pape, T., & Thompson, F. C. (2008). Pollinating flies (Diptera): A major contribution to plant diversity and agricultural production. Biodiversity, 9(1-2), 86-89. https://doi.org/10.1080/14888386.2008.9712892

  • Sukontason, K. L., Sanit, S., Klong-Klaew, T., Tomberlin, J. K., & Sukontason, K. (2014). Sarcophaga (Liosarcophaga) dux (Diptera: Sarcophagidae): A flesh fly species of medical importance. Biological Research, 47, 14. https://doi.org/10.1186/0717-6287-47-14

  • Tahir, H. M., Noor, A., Mehmood, S., Sherawat, S. M., & Qazi, M. A. (2018). Evaluating the accuracy of morphological identification of insect pests of rice crops using DNA barcoding. Mitochondrial DNA Part B: Resources, 3(2), 1220-1224. https://doi.org/10.1080/23802359.2018.1532334

  • Tan, S. H., Mohd Aris, E., Kurahashi, H., & Mohamed, Z. (2010). A new record of Iranihindia martellata (Senior-White, 1924) (Diptera: Sarcophagidae) from Peninsular Malaysia and female identification using both morphology and DNA-based approaches. Tropical Biomedicine, 27(2), 287-293.

  • Taylor, D. B., Moon, R. D., & Mark, D. R. (2012). Economic impact of stable flies (Diptera: Muscidae) on dairy and beef cattle production. Journal of Medical Entomology, 49(1), 198-209. https://doi.org/10.1603/ME10050

  • Townzen, J. S., Brower, A. V., & Judd, D. D. (2008). Identification of mosquito bloodmeals using mitochondrial cytochrome oxidase subunit I and cytochrome b gene sequences. Medical and Veterinary Entomology, 22(4), 386-393. https://doi.org/10.1111/j.1365-2915.2008.00760.x

  • Veer, V. (1999). Notes on Tabanidae (Diptera) that are Surra vectors or pestiferous with description of a new species of Tabanus from India. Oriental Insects, 33(1), 247-266. https://doi.org/10.1080/00305316.1999.10433793

  • Wang, X., Wang, W., Gao, Q., Wang, X., Lei, C., & Zhu, F. (2018). Chrysomya megacephala larvae feeding favourably influences manure microbiome, heavy metal stability and greenhouse gas emissions. Microbial Biotechnology, 11(3), 498-509. https://doi.org/10.1111/1751-7915.13253

  • Werszko, J., Szewczyk, T., Steiner-Bogdaszewska, Ż., Laskowski, Z., & Karbowiak, G. (2019). Molecular detection of Anaplasma phagocytophilum in blood-sucking flies (Diptera: Tabanidae) in Poland. Journal of Medical Entomology, 56(3), 822-827. https://doi.org/10.1093/jme/tjy217

  • Ya’cob, Z., Dawood, M. M., Jainih, L., Takaoka, H., & Azirun, M. S. (2020). Review of Malaysian black flies (Diptera: Simuliidae): Note on new record on Simulium kalimantanense from Sabah. Journal of Tropical Biology and Conservation, 17, 11-18.

  • Ya’cob, Z., Takaoka, H., Pramual, P., Low, V. L., & Sofian-Azirun, M. (2016). Distribution pattern of black fly (Diptera: Simuliidae) assemblages along an altitudinal gradient in Peninsular Malaysia. Parasites Vectors, 9, 219. https://doi.org/10.1186/s13071-016-1492-7

ISSN 0128-7702

e-ISSN 2231-8534

Article ID


Download Full Article PDF

Share this article

Recent Articles