Abstract
Entamoeba suis and E. polecki subtype (ST) 1 and ST3 recently have been inferred to be virulent in pigs. However, because relevant molecular epidemiological surveys have been limited, the prevalences of these species remain unknown and their pathogenicities are still controversial. We surveyed 196 fecal samples of pigs (118 of adults, 78 of piglets) at Tangerang in West Java, Indonesia, in 2017, employing PCR using porcine Entamoeba-specific primers. E. suis was the more frequently detected species, observed in 81.1% of samples, while E. polecki ST1 and ST3 were detected in 18.4% and 17.3% of samples, respectively; mixed infections (harboring 2–3 species or subtypes of Entamoeba) were confirmed in 29.3% of positive samples. Statistically significant differences in the positive rates were not seen between adult pigs and piglets, except for those of E. polecki ST3. The prevalences of Eimeria spp. and/or Cystoisospora suis (79.1%), strongyles (55.6%), and Strongyloides spp. (6.1%) were also observed morphologically in the samples. Further chronological or seasonal investigations of pigs and humans in these high-prevalence areas are needed to assess the virulence of the Entamoeba parasites, including the effects on pig productivity, and to evaluate the zoonotic impacts of these organisms.
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References
Clark CG, Kaffashian F, Tawari B, Windsor JJ, Twigg-Flesner A, Davies-Morel MC, Blessmann J, Ebert F, Peschel B, Le Van A, Jackson CJ, Macfarlane L, Tannich E (2006) New insights into the phylogeny of Entamoeba species provided by analysis of four new small-subunit rRNA genes. Int J Syst Evol Microbiol 56:2235–2239. https://doi.org/10.1099/ijs.0.64208-0
Clark CG (1995) Axenic cultivation of Entamoeba dispar Brumpt 1925, Entamoeba insolita Geiman and Wichterman 1937 and Entamoeba ranarum Grassi 1879. J Eukaryot Microbiol 42:590–593. https://doi.org/10.1111/j.1550-7408.1995.tb05912.x
Cuvertoret-Sanz M, Weissenbacher-Lang C, Lunardi M, Brunthaler R, Coma M, Weissenböck H, Segalés J (2019) Coinfection with Entamoeba polecki and Brachyspira hyodysenteriae in a pig with severe diarrhea. J Vet Diagn Investig 31:298–302. https://doi.org/10.1177/1040638719829800
Ekawasti F, Nurcahyo W, Wardhana AH, Shibahara T, Tokoro M, Sasai K, Matsubayashi M. (2020) Molecular characterization of highly pathogenic Eimeria species among beef cattle on Java Island, Indonesia. Parasitol Int. In press
Evangelopoulos A, Spanakos G, Patsoula E, Vakalis N, Legakis N (2000) A nested, multiplex, PCR assay for the simultaneous detection and differentiation of Entamoeba histolytica and Entamoeba dispar in faeces. Ann Trop Med Parasitol 94:233–240. https://doi.org/10.1080/00034980050006401
Fotedar R, Stark D, Beebe N, Marriott D, Ellis J, Harkness J (2007) Laboratory diagnostic techniques for Entamoeba species. Clin Microbiol Rev 20:511–532. https://doi.org/10.1128/CMR.00004-07
Gordon HML, Whitlock HV (1939) A new technique for counting nematode eggs in sheep faeces. J Sci Ind Res Australia 12:50–52
He GZ, Feng Y, Deng SX (2012) Evaluation of the intestinal microbial diversity in miniature pig after orally infected with Entamoeba histolytica. Parasitol Res 111:939–941. https://doi.org/10.1007/s00436-012-2841-x
Hirashima Y, Manchanayake T, Yano T, Kitahara S, Koreeda T, Kamimura S, Sasai K, Matsubayashi M, Shibahara T (2017) Development of molecular diagnostic protocols for detecting three types of Entamoeba from diarrheal and asymptomatic pigs and environmental moist soils. Parasitol Res 116:2001–2007. https://doi.org/10.1007/s00436-017-5483-1
Ito H, Hosokawa K, Kawamura M, Ito N, Abeto Y, Matsubayashi M, Sasai K, Shibahara T (2020) Swine multifocal ulcerative colitis associated with Entamoeba polecki subtype 3 and Salmonella Typhimurium. J Vet Med Sci 82:463–466. https://doi.org/10.1292/jvms.19-0682
Ji T, Cao HX, Wu R, Cui LL, Su GM, Niu C, Zhang N, Wang SK, Zhou DH (2019) Prevalence and genetic identification of three Entamoeba species in pigs in Southeastern China. Biomed Res Int 2019:2824017–2824018. https://doi.org/10.1155/2019/2824017
Joachim A, Altreuther G, Bangoura B, Charles S, Daugschies A, Hinney B, Lindsay DS, Mundt HC, Ocak M, Sotiraki S (2018) WAAVP guideline for evaluating the efficacy of anticoccidials in mammals (pigs, dogs, cattle, sheep). Vet Parasitol 253:102–119. https://doi.org/10.1016/j.vetpar.2018.02.029
Kyany’a C, Eyase F, Odundo E, Kipkirui E, Kipkemoi N, Kirera R, Philip C, Ndonye J, Kirui M, Ombogo A, Koech M, Bulimo W, Hulseberg CE (2019) First report of Entamoeba moshkovskii in human stool samples from symptomatic and asymptomatic participants in Kenya. Trop Dis Travel Med Vaccines 5:23. https://doi.org/10.1186/s40794-019-0098-4
Levine ND (1973) The Apicomplexa and the coccidia proper. Protozoan parasites of domestic animals and man, 2nd edn. Burgress Publishing Company, Minneapolis
Li WC, Geng JZ, Chen C, Qian L, Zhang T, Liu JL, Luo JX, Gu YF (2018) First report on the occurance of intestinal Entamoeba spp. in pigs in China. Acta Trop 185:385–390. https://doi.org/10.1016/j.actatropica.2018.06.020
Matsubayashi M, Kanamori K, Sadahiro M, Tokoro M, Abe N, Haritani M, Shibahara T (2015a) First molecular identification of Entamoeba polecki in a piglet in Japan and implications for aggravation of ileitis by coinfection with Lawsonia intracellularis. Parasitol Res 114:3069–3073. https://doi.org/10.1007/s00436-015-4509-9
Matsubayashi M, Murakoshi N, Komatsu T, Tokoro M, Haritani M, Shibahara T (2015b) Genetic identification of Entamoeba polecki subtype 3 from pigs in Japan and characterisation of its pathogenic role in ulcerative colitis. Infect Genet Evol 36:8–14. https://doi.org/10.1016/j.meegid.2015.08.030
Matsubayashi M, Sasagawa Y, Aita T, Tokoro M, Haritani M, Shibahara T (2016) First report of mixed Entamoeba polecki (ST 1) and E. suis infection in piglets shedding abnormal feces by histopathological and molecular surveys. Acta Parasitol 61:665–670. https://doi.org/10.1515/ap-2016-0093
Matsubayashi M, Suzuta F, Terayama Y, Shimojo K, Yui T, Haritani M, Shibahara T (2014) Ultrastructural characteristics and molecular identification of Entamoeba suis isolated from pigs with hemorrhagic colitis: implications for pathogenicity. Parasitol Res 113:3023–3028. https://doi.org/10.1007/s00436-014-3965-y
Neal RA (1966) Experimental studies on Entamoeba with reference to speciation. Adv Parasitol 4:1–51. https://doi.org/10.1016/s0065-308x(08)60447-9
Petri WA Jr, Singh U (1999) Diagnosis and management of amebiasis. Clin Infect Dis 29:1117–1125. https://doi.org/10.1086/313493
Schuster FL, Visvesvara GS (2004) Amebae and ciliated protozoa as causal agents of waterborne zoonotic disease. Vet Parasitol 126:91–120. https://doi.org/10.1016/j.vetpar.2004.09.019
Shimokawa C, Kabir M, Taniuchi M, Mondal D, Kobayashi S, Ali IK, Sobuz SU, Senba M, Houpt E, Haque R, Petri WA Jr, Hamano S (2012) Entamoeba moshkovskii is associated with diarrhea in infants and causes diarrhea and colitis in mice. J Infect Dis 206:744–751. https://doi.org/10.1093/infdis/jis414
Stensvold CR, Lebbad M, Victory EL, Verweij JJ, Tannich E, Alfellani M, Legarraga P, Clark CG (2011) Increased sampling reveals novel lineages of Entamoeba: consequences of genetic diversity and host specificity for taxonomy and molecular detection. Protist 162:525–541. https://doi.org/10.1016/j.protis.2010.11.002
Stensvold CR, Winiecka-Krusnell J, Lier T, Lebbad M (2018) Evaluation of a PCR method for detection of Entamoeba polecki, with an overview of its molecular epidemiology. J Clin Microbiol 56:pii:e00154–pii:e00118. https://doi.org/10.1128/JCM.00154-18
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739. https://doi.org/10.1093/molbev/msr121
Tuda J, Feng M, Imada M, Kobayashi S, Cheng X, Tachibana H (2016) Identification of Entamoeba polecki with unique 18S rRNA gene sequences from Celebes crested macaques and pigs in Tangkoko nature reserve, North Sulawesi, Indonesia. J Eukaryot Microbiol 63:572–577. https://doi.org/10.1111/jeu.12304
Verweij JJ, Polderman AM, Clark CG (2001) Genetic variation among human isolates of uninucleated cyst-producing Entamoeba species. J Clin Microbiol 39:1644–1646. https://doi.org/10.1128/JCM.39.4.1644-1646.2001
Wylezich C, Belka A, Hanke D, Beer M, Blome S, Höper D (2019) Metagenomics for broad and improved parasite detection: a proof-of-concept study using swine faecal samples. Int J Parasitol 49:769–777. https://doi.org/10.1016/j.ijpara.2019.04.007
Acknowledgments
The authors gratefully acknowledge Mrs. Rika Sekiguchi and Mrs. Noriko Asama for their help in the fecal and molecular examinations.
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This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology (JSPS) (Grant Nos. 16H05803 and 16K08025 to M.M., and No. 16K15049 to K.S.).
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All experiments were carried out by examinations of fecal samples without using live vertebrate animals. Thus, ethical approval for animal experimentation was not necessary. Field hygienic surveys of pig farms were conducted by veterinarians who were employed as civil servants by prefectural governments; all of these personnel belonged to the Livestock Hygiene Service Centre. All examinations performed in this study took place with the permission of the farm owners and were conducted as part of government affairs. No animals were sacrificed for the purposes of this study. No human participants were involved in this study.
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Wardhana, A.H., Sawitri, D.H., Ekawasti, F. et al. Occurrence and genetic identifications of porcine Entamoeba, E. suis and E. polecki, at Tangerang in West Java, Indonesia. Parasitol Res 119, 2983–2990 (2020). https://doi.org/10.1007/s00436-020-06806-0
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DOI: https://doi.org/10.1007/s00436-020-06806-0