Tropical Biomedicine 30(4): 727–730 (2013)

Research Note

Vector and reservoir host of a case of human pahangi infection in Selangor, peninsular Malaysia

Muslim, A.1*, Fong, M.Y. 2, Mahmud, R.2 and Sivanandam, S.2 1Faculty of Medicine, Universiti Teknologi MARA (UiTM), 47000, Sungai Buloh, Selangor, Malaysia 2Department of Parasitology and TIDREC, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia *Corresponding author email: [email protected] Received 4 July 2013; received in revised form 27 August 2013; accepted 27 August 2013

Abstract. A case of human eye infection caused by was reported in 2010 in a semi rural village in Selangor, peninsular Malaysia. Our report here reveals results of investigation on the vector and host for the transmission of the infection. We conducted entomological survey and cat blood examination in the vicinity of the patient’s home. The species subalbatus was incriminated as the vector, whereas cat served as the reservoir host.

Brugia pahangi is a lymphatic filarial worm host of the B. pahangi infection in Sungai that infects cats, dogs and monkeys (Denham Buloh. & McGreevy, 1977). Mosquitoe of species The patient, a 24 year old Malay woman, Mansonia spp. have been reported to be the presented with complaints of persistent right natural vector of B. pahangi (Edeson et al., eye redness associated with mild pain and 1960a). This worm has been experimentally mild grade fever for one month. She was given transmitted to human and the infection topical antibiotic eye drops but there was no produced signs and symptoms of lymphatic improvement. During further examination filariasis, but hardly produced micro- using slit-lamp, entangled tiny transparent filaraemia (Edeson et al., 1960b). Although thread-like mobile worms were detected at there was a report on the presence of B. the subconjunctival area. The worms were pahangi microfilariae (mf) in human blood extracted and confirmed as B. pahangi by in South Kalimantan, the finding was not PCR (Muslim, 2010). On further questioning conclusive as the method used to identify the the patient said that she had four pet cats at mf relied on measuring phosphatase home. She had no history of travelling and distribution in the mf (Palmieri et al., 1985). lived with her family in a village in Sungai The first confirmed cases of human lympatic Buloh. filariasis caused by B. pahangi in Malaysia The village (3º 12' 51" North, 101º 30' 57" were reported in 2010 (Tan et al., 2011). East) is highly populated and surrounded by Shortly after that, an isolated case of human orchard farms and an oil palm estate. Houses eye infection caused by B. pahangi was are built far apart from each other. Most of reported in Sungai Buloh, a semi-rural town the people living in this area are Malays of in the state of Selangor in Peninsular Malaysia Banjar and Mendailing ethnic groups. Similar (Muslim, 2010). The present report here to other typical villages elsewhere, this describes the vector and animal reservoir village also has improper drainage system

727 and some of the houses have unsatisfactory single mosquito was 21. Overall, the total sanitary facilities. larvae recovered were 121 L3, 56 L2 and 9 L1. Blood samples were collected only from Summary of the entomological findings is three of the patient’s pet cats. Thick blood shown in Table 1. smear (TBS) on microscope slides were Larvae from the mosquitoes and mf from prepared and stained with Giemsa solution. the infected cat’s blood were subjected to Microscopy examination revealed one of cats PCR and sequencing of the PCR fragments. was positive for mf with a density of 20 mf/ The PCR amplified a region in the B. pahangi 60 µl blood. The mf were sheathed with cytochrome oxidase subunit I (COXI) gene. obvious Innenkorper body and terminal Nucleotide sequence of the PCR fragments nuclei (Figure 1). The average length of the matched a B. pahangi COXI sequence mf and Innenkorper body were 245.8 ± 6.3 (Accession Number AJ271611) in the µm and 49.5 ± 4.5 µm, respectively. These GenBank database. In addition, multiple morphometric measurements were within alignment analysis (Figure 2) revealed > 99% the range for B. pahangi mf, as reported similarity in the sequences the larvae, mf and previously by Buckley & Edeson (1956) and B. pahangi COXI AJ271611. These findings Sivanandam & Fredericks (1966). further confirmed the identity of the larvae Mosquito collection was conducted in and mf as B. pahangi. the surrounding area from 0600-0900 and Ocular infection by filarial worms has 1800-2030 hours using human landing catch previously been reported in Malaysia, in (HLC) method. The mosquitoes were which Dirofilaria spp. were implicated as identified to species level and then dissected the aetiologic agents (Dissanaike et al., 1977; for the presence of filarial larvae. Of the 381 Mak & Thanalingam, 1984; Rohela et al., mosquitoes collected, 363 were dissected. 2009). Our report here is the first record of Three species, Amigeres subalbatus (87.7%), human eye infection caused by B. pahangi. Aedes albopictus (9.9%) and Aedes aegypti In this infection, human serves as accidental (2.4%) were found in the area. Only Ar. or dead end host. Hence, B. pahangi larva subalbatus was found to be positive (5%) for does not reach adulthood or sexual maturity filarial larvae. Of these infected Ar. in the human body. The worm is present as subalbatus, 2.5% had stage 3 larvae (L3). The occult infection as no mf is present in the highest number of larvae recovered from a blood.

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Figure 1. Brugia pahangi microfilaria in the blood of infected cat (40X). The sheath (SH) and Innenkorper body (IK) are arrowed

728 Table 1. Result of entomological findings in the Kg. Merbau Sempak

Mosquito species Total Dissection results

POSITIVE Total mosquitoes infected: 18 (5.0%) Caught 334 with L3: 9 (2.5 %) Dissected 334 with L2: 7 with L1: 2 Aedes aegypti Caught 9 NEGATIVE Dissected 9 Aedes albopictus Caught 38 NEGATIVE Dissected 20 Grand Total Caught 381 Dissected 363

Figure 2. Multiple sequence alignment of fragments from PCR on filarial larvae and mf in infected cat blood, and B. pahangi COXI (GenBank Accession Number AJ271611). Asterisks* indicate nucleotide identity among the sequences

729 Tan et al. (2011) reported several cases Dissanaike, A., Ramalingam, S., Fong, A., of zoonotic B. pahangi lymphatic filariasis Pathmayokan, S., Thomas, V. & Kan, S. in a suburbia of Kuala Lumpur City, Malaysia. (1977). Filaria in the vitreous of the eye Microfilaria-positive domestic cats were the of man in peninsular Malaysia. The source of the zoonotic infections in the American Journal of Tropical Medicine suburbia. The vector implicated was Ar. and Hygiene 26: 1143. subalbatus as only adult females of this Edeson, J., Wharton, R. & Laing, A. (1960a). A mosquito species were infected with B. preliminary account of the transmission, pahangi larvae. Our findings in the present maintenance and laboratory vectors of study further underline the role of Ar. Brugia pahangi. Transactions of the subalbatus and cat in the transmission of Royal Society of Tropical Medicine and zoonotic B. pahangi in Malaysia. Hygiene 54(5): 439-449. Armigeres subalbatus is widely Edeson, J., Wilson, T., Wharton, R. & Laing, A. distributed in Malaysia. Previously, this (1960b). Experimental transmission of species has never been considered a and B. pahangi to man. medically important mosquito in Malaysia Transactions of the Royal Society of as compared to other mosquitoes such as Tropical Medicine and Hygiene 54(3): Culex, Aedes, Mansonia and Anopheles spp. 229-234. Armigeres subalbatus has been observed in Mak, J.W. & Thanalingam, V. (1984). Human the laboratory to be an efficient vector for B. infection with Dirofilaria sp.(Nematoda: pahangi. It can rapidly produce high number Filaroidea), probably D. repens. Tropical of B. pahangi L3 larvae after a bloodmeal on Biomedicine 1: 109-113. infected cat (Edeson et al., 1960a). Thus, with Muslim, A. (2010). Filariasis in Kuala Lumpur a capacity to naturally harbour high number and Selangor: Entomological, para- of B. pahangi larvae, and being a human biter sitological and molecular studies. which thrives in areas of human habitations, Dissertation (M.Med.Sc). University of Ar. subalbatus should now be considered a Malaya, Kuala Lumpur. medically important mosquito species in Palmieri, J., Ratiwayanto, S., Masbar, S., Malaysia. Tirtokusumo, S., Rusch, J. & Marwoto, H. (1985). Evidence of possible natural Acknowledgements. This study was infections of man with Brugia pahangi supported by the University of Malaya in South Kalimantan (Borneo), Indonesia. Research Grant (RG333/11HTM) and Tropical and Geographical Medicine University of Malaya Postgraduate Research 37(3): 239. Fund No. PS 158/2008C. The authors are very Rohela, M., Jamaiah, I., Hui, T., Mak, J., Ithoi, grateful to Mrs. Hidaitul Masalaina, Mohd I. & Amirah, A. (2009). Dirofilaria causing Aidil and Romano Ngui for their assistance. eye infection in a patient from Malaysia. Southeast Asian Journal of Tropical Medicine and Public Health 40(5): 914- REFERENCES 918. Sivanandam, S. & Fredericks, H. (1966). The Buckley, J. & Edeson, J. (1956). On the adult “Innenkorper” in differentiation between morphology of Wuchereria sp.(malayi?) the microfilanriae of Brugia pahangi and from a monkey (Macaca irus) and from B. malayi (sub-periodic form). The cats in Malaya, and on Wuchereria Medical journal of Malaya 20(4): 337- pahangi n. sp. from a dog and a cat. 338. Journal of Helminthology 30(1): 1-20. Tan, L.H., Fong, M.Y., Mahmud, R., Muslim, A., Denham, D. & McGreevy, P. (1977). Brugian Lau, Y.L. & Kamarulzaman, A. (2011). filariasis: epidemiological and experi- Zoonotic Brugia pahangi filariasis in a mental studies. Advances in Para- suburbia of Kuala Lumpur City, Malaysia. sitology 15: 243-309. Parasitology international 60(1): 111- 113.

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