Next generation sequencing reveals widespread presence of human-pathogenic Blastocystis subtypes in chickens

Authors: Maloney, Jenny; DA CUNHA, MARIA - US Department Of Agriculture (USDA); Molokin, Aleksey; CURY, MARCIA - Universidade Federal De Minas Gerais; Santin-Duran, Monica

Submitted to: Veterinary Parasitology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/21/2021
Publication Date: 4/27/2021
Citation: Maloney, J.G., Da Cunha, M., Molokin, A., Cury, M.C., Santin, M. 2021. Next generation sequencing reveals widespread presence of human-pathogenic Blastocystis subtypes in chickens. Veterinary Parasitology. 12:2219-2231. https://doi.org/10.1007/s00436-021-07170-3.
DOI: https://doi.org/10.1007/s00436-021-07170-3

Interpretive Summary: Blastocystis sp. is one of the most common intestinal parasites found in humans and animals worldwide. Extensive genetic diversity has been described for this parasite and most subtypes isolated in humans have also been isolated from animals raising the question of the importance of animal reservoirs for Blastocystis and the possibility of zoonotic transmission through either direct contact or through fecal contamination of food or water. However, reports detailing the subtypes and prevalence of Blastocystis in birds are limited. Therefore, the aim of this study was to investigate the presence of Blastocystis in chickens to explore the role of chickens in the transmission of Blastocystis. Fecal samples from 130 chickens were collected from local markets in Uberlândia and Belo Horizonte in the state of Minas Gerais, Brazil. To detect and identify subtypes of Blastocystis, a next generation amplicon sequencing protocol was used. Forty-four of the 130 (33.8%) chickens examined were positive for Blastocystis. Blastocystis subtypes ST6, ST7, ST10, ST14, ST25, and a novel subtype (ST29) were observed. Mixed infections containing multiple subtypes were common and identified in 63.6% of Blastocystis positive chickens. Molecular characterization mostly identified subtypes previously found in humans (ST6 and ST7). Thus, chickens may be a source of human infection and environmental contamination. This information will be useful to other scientists, veterinarians, and public health agencies in understanding the current status of taxonomy, epidemiology, zoonotic potential, and public health importance of Blastocystis.

Technical Abstract: Blastocystis sp. is an intestinal protist parasite commonly found in the feces of humans and animals worldwide. Blastocystis exhibits extensive genetic diversity and has been identified in humans and a variety of animals including other mammals and birds using the small subunit rRNA (SSU rRNA) gene. Blastocystis subtypes do not exhibit strict host specificity which raises the possibility of zoonotic transmission through either direct contact or through fecal contamination of food or water. However, reports detailing the subtypes and prevalence of Blastocystis in birds are limited. Therefore, the aim of this study was to investigate the presence of Blastocystis in chickens by molecular characterization of the SSU rRNA gene to explore the role of chickens in the transmission of Blastocystis. Fecal samples from 130 chickens were collected from local markets in Uberlândia and Belo Horizonte in the state of Minas Gerais, Brazil. To detect and identify subtypes of Blastocystis, a next generation amplicon sequencing protocol was used to amplify and sequence a ca. 500-bp fragment of the SSU rRNA gene. Forty-four of the 130 (33.8%) chickens examined were positive for Blastocystis. Blastocystis subtypes ST6, ST7, ST10, ST14, ST25, and a novel subtype (ST29) were observed. A Nanopore sequencing strategy was used to validate novel subtype ST29 as recommended by recently proposed guidelines to obtain the near full-length SSU rRNA gene nucleotide sequence. Mixed infections containing multiple subtypes were common and identified in 63.6% of Blastocystis positive chickens. All positive samples contained one or both potentially zoonotic subtypes ST6 and ST7, while the other four subtypes were observed more sporadically. To our knowledge, this is the first genetic characterization of Blastocystis subtypes using next generation sequencing in chickens. The prevalence of Blastocystis in chickens was high, and molecular characterization mostly identified subtypes previously found in humans. Thus, chickens may be a source of human infection and environmental contamination. Further subtyping studies are required for a better understanding of host specificity and transmission dynamics of Blastocystis in poultry as well as other animals and humans in Brazil.