Microscopic and Molecular Identification of Trypanosoma lewisi among Rattus rattus and Rattus norvegicus from the District Kasur, Punjab, Pakistan
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Abstract
Background: We investigated the prevalence of Trypanosoma lewisi in Black rats (Rattus rattus) and Brown rats (R. norvegicus) using both microscopic and molecular detection methods, along with the analysis of associated risk factors.
Methods: A total of 178 rodents were trapped in Kasur district between November 2023 and November 2024, with epidemiological data and geographical coordinates recorded. Rodents were identified, euthanized and blood samples were collected. T. lewisi was confirmed through microscopy, PCR, sequencing and phylogenetic analysis. A GIS map was generated using ArcGIS 10.5.1 to illustrate geographical distribution.
Results: A total of 178 blood samples were examined by microscopically and by PCR. PCR confirmed to be either microscopically positive or negative samples. Overall prevalence of T. lewisi was 10(5.62%) and the findings were consistent across both diagnostic methods. At the species level, T. lewisi prevalence was higher in R. rattus (7.59%) compared to R. norvegicus (4.04%). Gender and the presence of ectoparasites were identified as potential risk factors.
Conclusion: These findings confirm the presence of T. lewisi in black and brown rats and serve as a baseline for further surveillance and control strategies.
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2. Griffiths J, Yeo HL, Yap G, et al. Survey of rodent-borne pathogens in Singapore reveals the circulation of Leptospira spp., Seoul hantavirus, and Rikettsia typhi. Sci Rep. 2022;12(1): 2692.
3. Hardgrove E, Zimmerman DM, von Fricken ME, et al. A scoping review of rodent-borne pathogen presence, exposure, and transmission at zoological institutions. Prev Vet Med. 2021;193:105345.
4. Kelly TR, Machalaba C, Karesh WB, et al. Implementing one health approaches to confront emerging and reemerging zoonotic disease threats: lessons from PREDICT. One Health Outlook. 2020;2:1.
5. Sobhy HM, Barghash SM, Behour TS, Razin EA. Seasonal fluctuation of trypanosomiasis in camels in North-West Egypt and effect of age, sex, location, health status and vector abundance on the prevalence. Beni-Suef University Journal of Basic and Applied Sciences. 2017;6(1):64-8.
6. Kim HJ, Han B, Lee HI, et al. Current Status of Trypanosoma grosi and Babesia microti in Small Mammals in the Republic of Korea. Animals (Basel). 2024;14(7):989.
7. Coura JR, Vinas PA. Chagas disease: a new worldwide challenge. Nature. 2010;465(7301):S6-7.
8. Samdi SM, Fajinmi AO, Kalejaye JO, et al. Prevalence of trypanosomosis in cattle at slaughter in Kaduna central abattoir. Asian J Anim Sci. 2011;5(2):162-5.
9. Magri A, Galuppi R, Fioravanti M. Autochthonous Trypanosoma spp. in European mammals: a brief journey amongst the neglected trypanosomes. Pathogens. 2021;10(3):334.
10. Ortiz PA, Garcia HA, Lima L, et al. Diagnosis and genetic analysis of the worldwide distributed Rattus-borne Trypanosoma (Herpetosoma) lewisi and its allied species in blood and fleas of rodents. Infect Genet Evol. 2018;63:380-90.
11. Salzer JS, Pinto CM, Grippi DC, et al. Impact of anthropogenic disturbance on native and invasive trypanosomes of rodents in forested Uganda. Ecohealth. 2016;13(4):698-707.
12. Kumar R, Gupta S, Bhutia WD, et al. Atypical human trypanosomiasis: Potentially emerging disease with lack of understanding. Zoonoses Public Health. 2022;69(4):259-76.
13. Jain P, Goyal V, Agrawal R. An atypical Trypanosoma lewisi infection in a 22-day-old neonate from india: an emergent zoonosis. Indian J Pathol Microbiol. 2023;66(1):199-201.
14. Shah I, Ali US, Andankar P, Joshi RR. Trypanosomiasis in an infant from India. J Vector Borne Dis. 2011; 48(2): 122-3.
15. Pumhom P, Morand S, Tran A, et al. Trypanosoma from rodents as potential source of infection in human-shaped landscapes of South-East Asia. Vet Parasitol. 2015;208(3-4):174-80.
16. Tang HJ, Lan YG, Wen YZ, et al. Detection of Trypanosoma lewisi from wild rats in Southern China and its genetic diversity based on the ITS1 and ITS2 sequences. Infect Genet Evol. 2012;12(5):1046-51.
17. Kamaruzaman INA, Ting HW, Mokhtar MAM, et al. First case report on molecular detection of Trypanosoma lewisi in an urban rat in Kelantan, Malaysia. An accidental finding. J Adv Vet Anim Res. 2021;8(3):435-9.
18. Tanthanathipchai N, Mitsuwan W, Chaisiri K, et al. Trypanosoma lewisi in blood of Rattus Rattus complex residing in human settlements, Nakhon Si Thammarat, Thailand: Microscopic and molecular investigations. Comp Immunol Microbiol Infect Dis. 2023;98:102010.
19. Wardhana AH, Putra FL, Yudhana A, et al. Detection of Trypanosoma lewisi from rodents residing in the densely populated residential regions along the coastal areas of Banyuwangi Sub District, Indonesia. Open Vet J. 2024;14(8):1808-18.
20. Andleeb S, Mahmood T, Khalid A, et al. Hexavalent chromium induces testicular dysfunction in small Indian mongoose (Herpestes javanicus) inhabiting tanneries area of Kasur District, Pakistan. Ecotoxicol Environ Saf. 2018;148:1001-9.
21. Sobia M, Mirza IS, Nuzhat S, et al. Prevalence and characterization of Trypanosoma species from livestock of Cholistan desert of Pakistan. Trop Biomed. 2018;35(1):140-48.
22. Thrusfield M. Veterinary Epidemiology. 4th ed. John Wiley & Sons. Edinburgh: UK; 2018. p. 887.
23. Wei Z, Du Y, Zhang J, et al. Prevalence and indicators of tooth wear among Chinese adults. PLoS One. 2016;11(9):e0162181.
24. Khan W, Nisa NN, Ilahi I, et al. Distribution of commensal rodents in rain-fed and irrigated areas of Swat district, Khyber Pakhtunkhwa, Pakistan. Braz J Biol. 2021;82:e236499.
25. Saeed U, Ali S, Latif T, et al. Prevalence and spatial distribution of animal brucellosis in central Punjab, Pakistan. Int J Environ Res Public Health. 2020;17(18):6903.
26. Herbreteau V, Jittapalapomg S, Rerkamnuaychoke W, et al. Protocols for Field and Laboratory Rodent Studies. Bangkok: Kasetsart University Press; 2011. p. 46.
27. Setiati N, Auliya R, Partaya P, et al. Types of rats and their parasites that potential to transmit disease in Tugu District, Semarang City. J Biol Biol Edu. 2021;13(3):363-8.
28. Milocco C, Kamyingkird K, Desquesnes M, et al. Molecular Demonstration of Trypanosoma evansi and Trypanosoma lewisi DNA in Wild Rodents from Cambodia, Lao PDR and Thailand. Transbound Emerg Dis. 2013;60(1):17-26.
29. Hoang DT, Chernomor O, Von Haeseler A, et al. UFBoot2: improving the ultrafast bootstrap approximation. Mol Biol Evol. 2018;35(2):518-22.
30. Saeed U, Ali S, Khan TM, et al. Seroepidemiology and the molecular detection of animal brucellosis in Punjab, Pakistan. Microorganisms. 2019;7(10):449.
31. Babyesiza WS, Katakweba A, Fornuskova A, et al. Trypanosome diversity in small mammals in Uganda and the spread of Trypanosoma lewisi to native species. Parasitol Res. 2023;123(1):54.
32. Karbowiak G, Rychlik L, Nowakowski W, Wita I. Natural infections of small mammals with blood parasites on the borderland of boreal and temperate forest zones. Acta Theriol. 2005;50(1):31-42.
33. Zhang K, Fu Y, Li J, Zhang L. Public health and ecological significance of rodents in Cryptosporidium infections. One Health. 2021;14:100364.
34. Schwan TG, Lopez JE, Safronetz D, et al. Fleas and trypanosomes of peridomestic small mammals in sub-Saharan Mali. Parasit Vectors. 2016;9(1):541.
35. Shafiyyah SC, Jamaiah I, Rohela M, et al. Prevalence of intestinal and blood parasites among wild rats in Kuala Lumpur, Malaysia. Trop Biomed. 2012;29(4):544-50.
36. Winterhoff ML, Achmadi AS, Roycroft EJ, et al. Native introduced trypanosome parasites in endemic and introduced murine rodents of Sulawesi. J Parasitol. 2020;106(5):523-36.
37. Molee PW, Sakulsak N, Saengamnatdej S. Detection of Trypanosoma spp. in Bandicota indica from the Thai-Myanmar border area, Mae Sot District Tak Province, Thailand. Asian Pac J Trop Med. 2019;12(10):457-62.
38. Garcia HA, Rangel CJ, Ortıiz PA, et al. Zoonotic trypanosomes in rats and fleas of Venezuelan slums. Ecohealth. 2019; 16(3):523-33.
39. Dobigny G, Poirier P, Hima K, et al. Molecular survey of rodent-borne Trypanosoma in Niger with special emphasis on T. lewisi imported by invasive black rats. Acta Trop. 2011;117(3):183-8.
40. Mohammed ES, El Kady AM, Youseef AG, Hassan AA. Distribution pattern of Trypanosoma lewisi in (Rattus norvegicus) in Egypt. Biomed J. 2018;1(4):6636-9.
41. Cassan C, Diagne CA, Tatard C, et al. Leishmania major and Trypanosoma lewisi infection in invasive and native rodents in Senegal. PLoS Neg Trop Dis. 2018;12(6):e0006615.
42. Pumhom P, Pognon D, Yangtara S, et al. Molecular prevalence of Trypanosoma spp. in wild rodents of Southeast Asia: influence of human settlement habitat. Epidemiol Infect. 2014;142(6):1221-30.
43. Ghersi BM, Peterson AC, Gibson NL, et al. In the heart of the city: Trypanosoma cruzi infection prevalence in rodents across New Orleans. Parasit Vectors. 2020;13(1):577.
44. Feng AY, Himsworth CG. The secret life of the city rat: a review of the ecology of urban Norway and black rats (Rattus norvegicus and Rattus rattus). Urban Ecosys. 2014;17(1):149-62.
45. Allam A, Ibrahim ES, Kazienga A, Al Zahrani F. Detection of parasites of animal health importance carried by wild rats collected from Jeddah city, Saudi Arabia. Adv Anim Vet Sci. 2019;7(10):921-8.
46. Linardi PM, Botelho JR. Prevalence of Trypanosoma lewisi in Rattus norvegicus from Belo Horizonte, State of Minas Gerais, Brazil. Mem Inst Oswaldo Cruz. 2002;97(3):411-4.
47. Katakweba AA. The prevalence of haemoparasites in rodents and shrews trapped from domestic and peridomestic houses in Morogoro Municipality, Tanzania. A Hidden Public Health Threat. Tanz Vet J. 2018;36:75¬-82.
48. Benacer D, Zain SNM, Amran F, et al. Isolation and molecular characterization of Leptospira interrogans and Leptospira borgpetersenii Isolates from the urban rat populations of Kuala Lumpur, Malaysia. Am J Trop Med Hyg. 2013;88(4):704-9.
49. Beldomenico PM, Begon M. Disease spread, susceptibility and infection intensity: vicious circles? Trends Ecol Evol. 2010;25(1):21-7.
2. Griffiths J, Yeo HL, Yap G, et al. Survey of rodent-borne pathogens in Singapore reveals the circulation of Leptospira spp., Seoul hantavirus, and Rikettsia typhi. Sci Rep. 2022;12(1): 2692.
3. Hardgrove E, Zimmerman DM, von Fricken ME, et al. A scoping review of rodent-borne pathogen presence, exposure, and transmission at zoological institutions. Prev Vet Med. 2021;193:105345.
4. Kelly TR, Machalaba C, Karesh WB, et al. Implementing one health approaches to confront emerging and reemerging zoonotic disease threats: lessons from PREDICT. One Health Outlook. 2020;2:1.
5. Sobhy HM, Barghash SM, Behour TS, Razin EA. Seasonal fluctuation of trypanosomiasis in camels in North-West Egypt and effect of age, sex, location, health status and vector abundance on the prevalence. Beni-Suef University Journal of Basic and Applied Sciences. 2017;6(1):64-8.
6. Kim HJ, Han B, Lee HI, et al. Current Status of Trypanosoma grosi and Babesia microti in Small Mammals in the Republic of Korea. Animals (Basel). 2024;14(7):989.
7. Coura JR, Vinas PA. Chagas disease: a new worldwide challenge. Nature. 2010;465(7301):S6-7.
8. Samdi SM, Fajinmi AO, Kalejaye JO, et al. Prevalence of trypanosomosis in cattle at slaughter in Kaduna central abattoir. Asian J Anim Sci. 2011;5(2):162-5.
9. Magri A, Galuppi R, Fioravanti M. Autochthonous Trypanosoma spp. in European mammals: a brief journey amongst the neglected trypanosomes. Pathogens. 2021;10(3):334.
10. Ortiz PA, Garcia HA, Lima L, et al. Diagnosis and genetic analysis of the worldwide distributed Rattus-borne Trypanosoma (Herpetosoma) lewisi and its allied species in blood and fleas of rodents. Infect Genet Evol. 2018;63:380-90.
11. Salzer JS, Pinto CM, Grippi DC, et al. Impact of anthropogenic disturbance on native and invasive trypanosomes of rodents in forested Uganda. Ecohealth. 2016;13(4):698-707.
12. Kumar R, Gupta S, Bhutia WD, et al. Atypical human trypanosomiasis: Potentially emerging disease with lack of understanding. Zoonoses Public Health. 2022;69(4):259-76.
13. Jain P, Goyal V, Agrawal R. An atypical Trypanosoma lewisi infection in a 22-day-old neonate from india: an emergent zoonosis. Indian J Pathol Microbiol. 2023;66(1):199-201.
14. Shah I, Ali US, Andankar P, Joshi RR. Trypanosomiasis in an infant from India. J Vector Borne Dis. 2011; 48(2): 122-3.
15. Pumhom P, Morand S, Tran A, et al. Trypanosoma from rodents as potential source of infection in human-shaped landscapes of South-East Asia. Vet Parasitol. 2015;208(3-4):174-80.
16. Tang HJ, Lan YG, Wen YZ, et al. Detection of Trypanosoma lewisi from wild rats in Southern China and its genetic diversity based on the ITS1 and ITS2 sequences. Infect Genet Evol. 2012;12(5):1046-51.
17. Kamaruzaman INA, Ting HW, Mokhtar MAM, et al. First case report on molecular detection of Trypanosoma lewisi in an urban rat in Kelantan, Malaysia. An accidental finding. J Adv Vet Anim Res. 2021;8(3):435-9.
18. Tanthanathipchai N, Mitsuwan W, Chaisiri K, et al. Trypanosoma lewisi in blood of Rattus Rattus complex residing in human settlements, Nakhon Si Thammarat, Thailand: Microscopic and molecular investigations. Comp Immunol Microbiol Infect Dis. 2023;98:102010.
19. Wardhana AH, Putra FL, Yudhana A, et al. Detection of Trypanosoma lewisi from rodents residing in the densely populated residential regions along the coastal areas of Banyuwangi Sub District, Indonesia. Open Vet J. 2024;14(8):1808-18.
20. Andleeb S, Mahmood T, Khalid A, et al. Hexavalent chromium induces testicular dysfunction in small Indian mongoose (Herpestes javanicus) inhabiting tanneries area of Kasur District, Pakistan. Ecotoxicol Environ Saf. 2018;148:1001-9.
21. Sobia M, Mirza IS, Nuzhat S, et al. Prevalence and characterization of Trypanosoma species from livestock of Cholistan desert of Pakistan. Trop Biomed. 2018;35(1):140-48.
22. Thrusfield M. Veterinary Epidemiology. 4th ed. John Wiley & Sons. Edinburgh: UK; 2018. p. 887.
23. Wei Z, Du Y, Zhang J, et al. Prevalence and indicators of tooth wear among Chinese adults. PLoS One. 2016;11(9):e0162181.
24. Khan W, Nisa NN, Ilahi I, et al. Distribution of commensal rodents in rain-fed and irrigated areas of Swat district, Khyber Pakhtunkhwa, Pakistan. Braz J Biol. 2021;82:e236499.
25. Saeed U, Ali S, Latif T, et al. Prevalence and spatial distribution of animal brucellosis in central Punjab, Pakistan. Int J Environ Res Public Health. 2020;17(18):6903.
26. Herbreteau V, Jittapalapomg S, Rerkamnuaychoke W, et al. Protocols for Field and Laboratory Rodent Studies. Bangkok: Kasetsart University Press; 2011. p. 46.
27. Setiati N, Auliya R, Partaya P, et al. Types of rats and their parasites that potential to transmit disease in Tugu District, Semarang City. J Biol Biol Edu. 2021;13(3):363-8.
28. Milocco C, Kamyingkird K, Desquesnes M, et al. Molecular Demonstration of Trypanosoma evansi and Trypanosoma lewisi DNA in Wild Rodents from Cambodia, Lao PDR and Thailand. Transbound Emerg Dis. 2013;60(1):17-26.
29. Hoang DT, Chernomor O, Von Haeseler A, et al. UFBoot2: improving the ultrafast bootstrap approximation. Mol Biol Evol. 2018;35(2):518-22.
30. Saeed U, Ali S, Khan TM, et al. Seroepidemiology and the molecular detection of animal brucellosis in Punjab, Pakistan. Microorganisms. 2019;7(10):449.
31. Babyesiza WS, Katakweba A, Fornuskova A, et al. Trypanosome diversity in small mammals in Uganda and the spread of Trypanosoma lewisi to native species. Parasitol Res. 2023;123(1):54.
32. Karbowiak G, Rychlik L, Nowakowski W, Wita I. Natural infections of small mammals with blood parasites on the borderland of boreal and temperate forest zones. Acta Theriol. 2005;50(1):31-42.
33. Zhang K, Fu Y, Li J, Zhang L. Public health and ecological significance of rodents in Cryptosporidium infections. One Health. 2021;14:100364.
34. Schwan TG, Lopez JE, Safronetz D, et al. Fleas and trypanosomes of peridomestic small mammals in sub-Saharan Mali. Parasit Vectors. 2016;9(1):541.
35. Shafiyyah SC, Jamaiah I, Rohela M, et al. Prevalence of intestinal and blood parasites among wild rats in Kuala Lumpur, Malaysia. Trop Biomed. 2012;29(4):544-50.
36. Winterhoff ML, Achmadi AS, Roycroft EJ, et al. Native introduced trypanosome parasites in endemic and introduced murine rodents of Sulawesi. J Parasitol. 2020;106(5):523-36.
37. Molee PW, Sakulsak N, Saengamnatdej S. Detection of Trypanosoma spp. in Bandicota indica from the Thai-Myanmar border area, Mae Sot District Tak Province, Thailand. Asian Pac J Trop Med. 2019;12(10):457-62.
38. Garcia HA, Rangel CJ, Ortıiz PA, et al. Zoonotic trypanosomes in rats and fleas of Venezuelan slums. Ecohealth. 2019; 16(3):523-33.
39. Dobigny G, Poirier P, Hima K, et al. Molecular survey of rodent-borne Trypanosoma in Niger with special emphasis on T. lewisi imported by invasive black rats. Acta Trop. 2011;117(3):183-8.
40. Mohammed ES, El Kady AM, Youseef AG, Hassan AA. Distribution pattern of Trypanosoma lewisi in (Rattus norvegicus) in Egypt. Biomed J. 2018;1(4):6636-9.
41. Cassan C, Diagne CA, Tatard C, et al. Leishmania major and Trypanosoma lewisi infection in invasive and native rodents in Senegal. PLoS Neg Trop Dis. 2018;12(6):e0006615.
42. Pumhom P, Pognon D, Yangtara S, et al. Molecular prevalence of Trypanosoma spp. in wild rodents of Southeast Asia: influence of human settlement habitat. Epidemiol Infect. 2014;142(6):1221-30.
43. Ghersi BM, Peterson AC, Gibson NL, et al. In the heart of the city: Trypanosoma cruzi infection prevalence in rodents across New Orleans. Parasit Vectors. 2020;13(1):577.
44. Feng AY, Himsworth CG. The secret life of the city rat: a review of the ecology of urban Norway and black rats (Rattus norvegicus and Rattus rattus). Urban Ecosys. 2014;17(1):149-62.
45. Allam A, Ibrahim ES, Kazienga A, Al Zahrani F. Detection of parasites of animal health importance carried by wild rats collected from Jeddah city, Saudi Arabia. Adv Anim Vet Sci. 2019;7(10):921-8.
46. Linardi PM, Botelho JR. Prevalence of Trypanosoma lewisi in Rattus norvegicus from Belo Horizonte, State of Minas Gerais, Brazil. Mem Inst Oswaldo Cruz. 2002;97(3):411-4.
47. Katakweba AA. The prevalence of haemoparasites in rodents and shrews trapped from domestic and peridomestic houses in Morogoro Municipality, Tanzania. A Hidden Public Health Threat. Tanz Vet J. 2018;36:75¬-82.
48. Benacer D, Zain SNM, Amran F, et al. Isolation and molecular characterization of Leptospira interrogans and Leptospira borgpetersenii Isolates from the urban rat populations of Kuala Lumpur, Malaysia. Am J Trop Med Hyg. 2013;88(4):704-9.
49. Beldomenico PM, Begon M. Disease spread, susceptibility and infection intensity: vicious circles? Trends Ecol Evol. 2010;25(1):21-7.
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| Issue | Vol 21 No 1 (2026) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijpa.v21i1.21638 | |
| Keywords | ||
| Trypanosoma lewisi Rattus rattus Rattus norvegicus Pakistan | ||
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How to Cite
1.
Saeed M, Iqbal A, Ali S, Akbar H, Sana S, Saeed U. Microscopic and Molecular Identification of Trypanosoma lewisi among Rattus rattus and Rattus norvegicus from the District Kasur, Punjab, Pakistan. Iran J Parasitol. 2026;21(1):94-104.
