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| ORIGINAL ARTICLE |
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| Year : 2023 | Volume
: 3
| Issue : 1 | Page : 15 |
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Molecular detection and public health risk assessment of Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi, and Blastocystis sp. of animals in a tropical wildlife park of Hainan Island, China
Guangxu Ren1, Jiaqi Li1, Jingyan Xiong2, Xiuyi Lai3, Yuan Wang1, Sheng Lei1, Xin Lu3, Tianya He3, Yunfei Zhou1, Yun Zhang2, Gang Lv1
1 Key Laboratory of Tropical Translational Medicine of Ministry of Education; NHC Key Laboratory of Tropical Disease Control, Hainan Medical University; The University of Hong Kong Joint Laboratory of Tropical Infectious Diseases; Academician Workstation of Hainan Province; Department of Pathogen Biology, Haikou 571199, China
2 Key Laboratory of Tropical Translational Medicine of Ministry of Education, Hainan Medical University; NHC Key Laboratory of Tropical Disease Control; The University of Hong Kong Joint Laboratory of Tropical Infectious Diseases; Academician Workstation of Hainan Province, Haikou 571199, China
3 Key Laboratory of Tropical Translational Medicine of Ministry of Education, Hainan Medical University; NHC Key Laboratory of Tropical Disease Control; The University of Hong Kong Joint Laboratory of Tropical Infectious Diseases; Academician Workstation of Hainan Province; International School of public health and one health, Haikou 571199, China
| Date of Submission | 24-May-2023 |
| Date of Decision | 27-Jun-2023 |
| Date of Acceptance | 28-Jul-2023 |
| Date of Web Publication | 17-Aug-2023 |
Correspondence Address:
Gang Lv
Key Laboratory of Tropical Translational Medicine of Ministry of Education; NHC Key Laboratory of Tropical Disease Control, Hainan Medical University; The University of Hong Kong Joint Laboratory of Tropical Infectious Diseases; Academician Workstation of Hainan Province; Department of Pathogen Biology
China
Yun Zhang
Key Laboratory of Tropical Translational Medicine of Ministry of Education, Hainan Medical University; NHC Key Laboratory of Tropical Disease Control; The University of Hong Kong Joint Laboratory of Tropical Infectious Diseases; Academician Workstation of Hainan Province
China
 Source of Support: None, Conflict of Interest: None
DOI: 10.4103/2773-0344.383636
Objective: To detect the prevalence and characterize of Cryptosporidium spp., Giardia (G.) duodenalis, Enterocytozoon (E.) bieneusi and Blastocystis sp. of animals at a tropical wildlife park in Hainan Province, China, and to assess zoonotic risks and the potential threat of public health.
Methods: Fecal specimens were collected from animals of 27 species at the wildlife park in Hainan Province, China, and detected these pathogens using polymerase chain reaction (PCR) amplifications and sequencing of gene fragments based on small subunit ribosomal RNA (SSU rRNA) gene, glutamate dehydrogenase (GDH), internal transcribed spacer (ITS) and SSU rRNA gene, respectively.
Results: A total of 99 animals were studied, and 33 (33.3%) animals were found to harbor intestinal parasite, and the prevalence of Cryptosporidium spp., G. duodenalis, E. bieneusi and Blastocystis sp. were 9.1%, 2.0%, 5.1%, and 25.3%, respectively. Five Cryptosporidium species (C. parvum, C. ubiquitum, C. scrofarum, C. xiaoi and C. parvum-like), one G. duodenalis assemblages (E), four E. bieneusi ITS genotypes (CM1, HLJD-I, HNR-III and 1 novel genotype HNED-III) were detected, and seven subtypes of Blastocystis sp. (ST1, ST2, ST3, ST5, ST10, ST14 and ST15) were also identified.
Conclusions: This is the first molecular detection and public health risk assessment of four intestinal protozoa species in Hainan Tropical Wildlife Park and Botanical Garden, China. Almost all species/genotypes/subtypes of four intestinal protozoa identified in this study have the potential of zoonosis and may cause public health risks.
Keywords: Tropical wildlife park; Cryptosporidium spp.; Giardia duodenalis; Enterocytozoon bieneusi; Blastocystis sp.; zoonotic risks
How to cite this article:
Ren G, Li J, Xiong J, Lai X, Wang Y, Lei S, Lu X, He T, Zhou Y, Zhang Y, Lv G. Molecular detection and public health risk assessment of Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi, and Blastocystis sp. of animals in a tropical wildlife park of Hainan Island, China. One Health Bull 2023;3:15 |
How to cite this URL:
Ren G, Li J, Xiong J, Lai X, Wang Y, Lei S, Lu X, He T, Zhou Y, Zhang Y, Lv G. Molecular detection and public health risk assessment of Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi, and Blastocystis sp. of animals in a tropical wildlife park of Hainan Island, China. One Health Bull [serial online] 2023 [cited 2023 Sep 28];3:15. Available from: http://www.johb.info/text.asp?2023/3/1/15/383636 |
| 1. Introduction | |  |
Cryptosporidium spp., Giardia (G.) duodenalis, Enterocytozoon (E.) bieneusi and Blastocystis sp. are important zoonotic protists in humans and animals around the world, especially in tropical and subtropical regions[1],[2],[3],[4]. They can cause diarrhea or other gastrointestinal sufferings in human and animals[5],[6].
The fecal-oral route is the main route of transmission for these pathogens, and infections can also be caused by contaminated food or water[7],[8].
Based polymerase chain reaction (PCR) and sequence analysis techniques, at least 45 valid species and 120 genotypes of Cryptosporidium spp. have been described, and at least 25 species/genotypes have been detected in humans[9]. Eight distinct assemblages (A-H) of G. duodenalis have been reported, and assemblages A and B are infectious to humans and other vertebrate hosts[2]. More than 560 E. bieneusi genotypes clustered into 13 major groups in phylogenetic analysis were identified and most zoonotic genotypes were clustered into group 1 and group 2[3]. Overall, 28 Blastocystis sp. subtypes (ST1-ST17, ST21, ST23-ST32) have been identified, including zoonotic subtypes (ST1-ST10, ST12, ST14, ST16, ST23) and host adopted subtypes from specific animals (ST11, ST13, ST15, ST17, ST21, ST24-ST32)[10].
The wildlife park is a special ecological environment where a variety of species of native and imported animals live in the same natural state. Zoonoses are caused by pathogens that are naturally transmitted from animals to humans, especially workers or visitors in the park. The ongoing occurrence of zoonoses pose significant threats to public health. Moreover, pathogens can spread outside the park through environmental pollution especially by water contamination and cause public health risks[11]. The aim of this study was to detect the prevalence and molecular characterization of the four common zoonosis and opportunistic intestinal protozoa including Cryptosporidium spp., G. duodenalis, E. bieneusi and Blastocystis sp. of animals in the only wildlife park located in tropical island province of China, and to assess their zoonosis and potential public health risks.
| 2. Materials and methods | | |
2.1. Ethics statement
The protocol of this study was reviewed and approved by the Research Ethics Committee and the Animal Ethical Committee of Hainan Medical University (HYLL-2022-405). Only excreted feces of the wildlife were used for this study and during the entire procedure, no animals were injured.
2.2. Sampling area and collection
From June to July 2021, samples were collected from Hainan Tropical Wildlife Park and Botanical Garden, located in Dongshan Town Xiuying District Haikou City [Figure 1]. Hainan Tropical Wildlife Park and Botanical Garden is the first tropical wildlife park in China, covering over 100 hectares and collecting at least 2000 animals including over 100 species, with more than 500000 visitors from around world every year. The 99 fecal samples, representing 27 species were obtained from the park at the time of sampling. The fresh samples were placed in plastic bags, which were labeled, recorded and transfered to laboratory for storage at 4 °C within 24 h. | Figure 1: The location of Hainan Tropical Wildlife Park and Botanical Garden, Hainan, China.
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2.3. DNA extraction
All the fecal samples were washed with distilled water twice and centrifuged at 3000 x g for 10 min. Genomic DNA was extracted from 200 mg of faeces per sample by using the QIAamp DNA Mini Stool Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. The DNA samples was stored at -20 °C until PCR analysis was performed.
2.4. PCR amplification
All DNA preparations were detected for the presence of Cryptosporidium spp., G. duodenalis, E. bieneusi and Blastocystis sp. by PCR amplifications and sequencing of gene fragments based on SSU rRNA (~850 bp) gene, GDH (~530 bp) gene, ITS ~390 bp) gene and SSU rRNA (~260 bp) gene, respectively, and PCR conditions have been previously described[12],[13],[14],[15] [Table 1]. PCR reactions (25 μL) included 0.5 μL of Taq DNA polymerase (TaKaRa Bio Inc., Tokyo, Japan), 0.5 μL of each primer (20 mM), 2.5 μL of 10 × PCR buffer (TaKaRa Bio Inc., Tokyo, Japan), 2 μL of dNTPs (TaKaRa Bio Inc., Tokyo, Japan) and 2 μL of template DNA. A negative control without DNA and a positive control (rat-derived Cryptosporidium rat genotype IV DNA for Cryptosporidium spp., goat-derived assemblage E DNA for G. duodenalis, rat-derived genotype Peru 8 DNA for E. bieneusi and cattle-derived ST1 DNA for Blastocystis sp.) were included in all the PCR tests. Amplified products were electrophoresed on a 1.5% agarose gel stained with GelRed (Biotium, Inc., Hayward, CA) and visualized under UV light. | Table 1: Primers used in the characterization of the Cryptosporidium spp., G. duodenalis, E. bieneusi, and Blastocystis sp.
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2.5. Nucleotide sequencing and analysis
All appropriately sized PCR products were sequenced by Sangon Biotech Co., Ltd (Shanghai, China). Two-directional sequencing and additional sequencing were used to confirm the accuracy of the sequence. The Basic Local Alignment Search Tool (BLAST) and Clustal X were used to genotype the isolates by comparing the sequenced data published on GenBank.
2.6. Phylogenetic analysis
The phylogenetic tree was constructed using the Mega 11 (http://www.megasoftware.net/) software based on the neighbor-joining method and Kimura-2-parameter model. The reliability of the tree was assessed through bootstrap analysis with 1000 replicates.
| 3. Results | | |
3.1. Prevalence of Cryptosporidium spp., G. duodenalis, E. bieneusi and Blastocystis sp.
In this study, 33 (33.3%) animals were found to harbor intestinal parasite, and the prevalence of Cryptosporidium spp., G. duodenalis, E. bieneusi and Blastocystis sp. were 9.1%, 2.0%, 5.1%, and 25.3%, respectively [Table 2]. Co-infection with Cryptosporidium spp. and Blastocystis sp. were observed in white buffalo (Hainan) (n=1), sika deer (n=1), red-bellied squirrel (n=1), beaver rat (n=1). While co-infection with E. bieneusi and Blastocystis sp. were observed in macaques (n=2), brown bear (n=1) and Hainan eld's deer (n=1). | Table 2: Occurrence of Cryptosporidium spp., G. duodenalis, Enterocytozoon bieneusi and Blastocystis sp.
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3.2. Molecular characterization of Cryptosporidium spp., G. duodenalis, E. bieneusi and Blastocystis sp.
Five Cryptosporidium species (C. parvum, C. ubiquitum, C. scrofarum, C. xiaoi and C. parvum-like) were identified in David's deer (C. xiaoi), white buffalo (Hainan) (C. xiaoi, C. scrofarum), sika deer (C. ubiquitum), emu (C. ubiquitum) and Africa lion (C. parvum- like), red-bellied squirrel (C. parvum) and beaver rat (C. ubiquitum) in this study [Table 2]. Comparison with the sequenced data published on GenBank showed that the sequences of C. parvum (MW769926), C. scrofarum (MT071828), and C. xiaoi (LC414392) were identical to previous sequences. While the sequence of the C. parvum- like isolate (OQ993217) had 98.6% similarity with a sequence (MK956936) which was isolated from a bamboo rat from Guangxi, China. The sequence of the C. xiaoi (OQ993214) had 99.8% similarity with a sequence (LC414388) which was isolated from a capra hircus from Algeria. In this study, C. ubiquitum were isolated from a sika deer, two beaver rats and an emu, of which the emu was identified as the host for the first time. The C. ubiquitum sequences in this study were 99.8% to 100% identical to known C. ubiquitum isolates in the GenBank database. Phylogenetic reconstructions of the Cryptosporidium spp. based on SSU rRNA gene sequences from zoo animals can be seen in [Figure 2]. | Figure 2: Phylogenetic relationships among Cryptosporidium spp. based on the SSU rRNA gene. Neighbor-Joining method and genetic distances calculated with the Kimura 2-parameter model using MEGA 11. Bootstrapping with 1000 replicates was used to support the clades. The bootstrap number is shown on each node. • Known species detected.
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Nucleotide sequence and phylogenetic analysis based on GDH gene identified one G. duodenalis assemblages as assemblage E in a sika deer and a Hainan eld's deer was identified in present study [Figure 3]. The isolates shared 99.6%-99.8% identity with the sequence (MK645786) which isolated from Tan sheep in Ningxia, China. | Figure 3: Phylogenetic relationships among G. duodenalis based on GDH gene. Neighbor-Joining method and genetic distances calculated with the Kimura 2-parameter model using MEGA 11. Bootstrapping with 1000 replicates was used to support the clades. The bootstrap number is shown on each node. • Known assemblages detected.
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Four E. bieneusi genotypes were identified that including three known genotypes named CM1 (n=2), HLJD-I (n=1) and HNR-III (n=1), and one novel genotypes (HNED-III). The novel genotypes HNED-III (OQ674806) from Hainan eld's deer had the largest similarity with genotype CGC2 (MK559495) with one base difference at positions 45 (A→G). Phylogenetic analysis of E. bieneusi showed that genotypes CM1 and HNR-III were clustered in group 1, whereas genotype HLJD-I and HNED-III were clustered into group 2 [Figure 4]. | Figure 4: Phylogenetic relationships among E. bieneusi based on ITS gene. Neighbor-Joining method and genetic distances calculated with the Kimura 2-parameter model using MEGA 11. Bootstrapping with 1000 replicates was used to support the clades. The bootstrap number is shown on each node. • Known genotypes detected; ∆Novel genotype detected.
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A total of 7 Blastocystis sp. subtypes were identified as ST1, ST2, ST3, ST5, ST10, ST14 and ST15. The most prevalent Blastocystis sp. subtype was ST10 (36%) followed by ST5 (28%). Phylogenetic analyses showed that the sequences of the 7 subtypes obtained in this study clustered with their reference subtypes which obtained from other animals or humans into one branch, with high bootstrap values [Figure 5]. | Figure 5: Phylogenetic relationships among Blastocystis based on the SSU rRNA gene. Neighbor-Joining method and genetic distances calculated with the Kimura 2-parameter model using MEGA 11. Bootstrapping with 1000 replicates was used to support the clades. The bootstrap number is shown on each node. • Known sequence detected, δ Novel sequence detected.
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| 4. Discussion | | |
In the present study, the infection rate of Cryptosporidium spp. in the wildlife park was 9.1%, which was higher than the rates (2.3%) reported in petting zoos in Canada[16], zoos in Henan, China (2.8%)[17], zoo mammals in Bangladesh (3.5%)[18], birds in Beijing and Harbin, China (1.9%)[19], non-human primates in France, Germany, and Spain (0.9%)[20], but lower than zoo felines in Harbin, China (30.4%)[21].
The infection rate of G. duodenalis was 2.0%, which was similar to those in Zhengzhou zoo, China (2.5%)[22], lower than petting zoos in Canada (14.8%)[16], zoo mammals in Bangladesh (5.5%)[18], non-human primates in France, Germany, and Spain (18.1%)[20], while higher than zoos in Henan, China (0.5%)[17].
The E. bieneusi prevalence was 5.1%, which was higher than non-human primates in France, Germany, and Spain (0.9%)[20], lower than in previous studies conducted in the great apes in Senegal, the Republic of the Congo and France (7.9%)[23], in Zhengzhou zoo, China (15.8%)[22], in zoos in Henan, China (20.8%)[17].
Concerning Blastocystis sp., the prevalence was 25.3% which was similar to the studies conducted in Poland (26.9%)[24], non-human primates in France, Germany, and Spain (20.3%)[20], in a zoo in Slovakia (20.8%)[25], higher than the captive wildlife species in Sichuan, China (15.7%)[26], zoo animals in Hangzhou, Dalian, and Suzhou cities, China (6.0%)[27], non-human primates in Harbin (7.0%)[28], mammalian wildlife species in Bangladesh (15.5%)[29], zoo animals in Henan, China (19.1%)[17], while lower than golden monkeys in Shijiazhuang, China (48.7%)[30], wild mammals in Brazil (51.7%)[31], zoo animals in Australia (42.1%)[32], non-human primates in Spain (45.1%)[33], mammalian and non-mammalian species in French (32.3%)[34], great apes in Senegal, the Republic of the Congo, and France (97.4%)[23].
According to the literature, all five Cryptosporidium species (C. parvum, C. ubiquitum, C. scrofarum, C. xiaoi and C. parvum-like) identified in present study had zoonotic potential. C. parvum was one of the two Cryptosporidium responsible for most cases of cryptosporidiosis in humans[35]. C. ubiquitum, previously identified as the Cryptosporidium cervine genotype, were commonly detected in domestic and wild ruminants, rodents, carnivores, and primates[36]. It has been found in humans worldwide, primarily in industrialized nations[37]. C. scrofarum appeared to be the main pig-adapted species[38], but human infection was reported in an immunocompetent man[39]. C. xiaoi was most common species in sheep but it was identified in two children with HIV/AIDS in Ethiopia and had zoomotic potential[40]. C. parvum- like was described in immunocompetent and immunosuppressed individuals[41]. Therefore, the results above indicate that the animals in the wildlife with those Cryptosporidium species may transmit it to other animals and humans.
For G. duodenalis, assemblages A and B were considered to be zoonotic and have been found in various mammals including humans. Assemblage E was mostly identified in hoofed mammals (such as cattle, sheep, goats, and pigs), but it has been reported in at least 57 people in several countries, indicating the zoonotic potencial[2].
Four E. bieneusi genotypes including CM1, HNR-III, HLJD-1 and HNED-III were detected in this study. Genotype CM1 was only found in NHPs and dog at present[42]. HNR-III was first reported in wild rats in Hainan, China[43]. HLJD-1 was first reported in sika deer[44]. HNED-III is a novel genotype. Although no human infection of these four genotypes was reported, phylogenetic analysis showed they were clustered into group 1 and group 2, which was composed of most zoonotic genotypes of E. bieneusi, indicating the zoonotic potential.
Blastocystis ST1-ST9 and ST12 have been detected in humans, and ST1-ST4 are the most common, comprising over 90% of all STs identified in humans globally[45].
In this study, seven Blastocystis sp. subtypes were detected from 10 animal species including ST1 (n=1), ST2 (n=4), ST3 (n=1), ST5 (n=7), ST10 (n=9), ST14 (n=1), and ST15 (n=2), all subtypes have zoonotic potential except ST15 from two rodents. The presence of zoonotic Blastocystis subtypes indicative those animals represent a potential threat to other animals and humans.
| 5. Conclusions | | |
This is the first molecular detection and public health risk assessment on four intestinal protozoa in a tropical wildlife park of Hainan Island, China. Almost all species/genotypes/subtypes of four intestinal protozoa identified in this study have zoonotic potential and may cause public health risks. These findings extend the host range for four intestinal protozoa and also provide important data support for prevention and control of zoonoses. Further studies should be made to reveal the genetic characteristics and assess the zoonotic risks of these parasites.
Conflict of interest statement
The authors declare that there is no conflict of interest.
Acknowledgements
We would like to thank the staff at the tropical wildlife park in Hainan Province, and the support of the Hainan Tropical Infectious Diseases Biobank.
Funding
This work was supported by the National Natural Science Foundation of China (82060375), Research Project of Hainan Academician Innovation Platform (YSPTZX202004), Major Science and Technology Program of Hainan Province (ZDKJ202003), Hainan Talent Development Project (SRC200003), High level talents project of Hainan Natural Science Foundation (822RC695), the Scientific Research Foundation of the Higher Education Institutions of Hainan Province, China (Hnky2021-44), the Hainan Medical University Talent Development Project (XRC2021002), Graduate student innovation grant of Hainan Medical University (HYYB2020-04, HYYS2021A20, HYYB2022A25).
Data availability statement
The representative nucleotide sequences obtained in this study were deposited in the GenBank database under the following accession numbers: OQ993214 to OQ993217 for Cryptosporidium spp.; OQ999329 and OQ999330 for G. duodenalis, OQ674806 for E. bieneusi; OQ992504 for Blastocystis sp.
Authors' contributions
Ren GX was responsible for literature collection and writing the draft of this article and performed experiments, software and formal analysis. Li JQ and Xiong JY contributed to acquisition of samples and performed experiments. Lai XY, Wang Y, Lei S, Lu X, He TY and Zhou YF provided administrative, technical support and constructive discussion. As the corresponding authors, Lv G and Zhang Y controlled the content of the whole review. All authors contributed to the article and approved the submitted version.
| References | | |
| 1. | Ryan U, Fayer R, Xiao LH. Cryptosporidium species in humans and animals: Current understanding and research needs. Parasitology 2014; 141(13): 1667-1685.  |
| 2. | Cai W, Ryan U, Xiao LH, et al. Zoonotic giardiasis: An update. Parasitology Res 2021; 120(12): 4199-4218. |
| 3. | Li W, Xiao LH. Ecological and public health significance of Enterocytozoon bieneusi. One Health 2021; 12: 100209. doi: 10.1016/j.onehlt.2020.100209. |
| 4. | Deng L, Wojciech L, Gascoigne N R J, Peng GN, Tan KSW. New insights into the interactions between Blastocystis, the gut microbiota, and host immunity. PLoS Pathog 2021; 17(2): e1009253. |
| 5. | Jiang YY, Liu L, Yuan ZY, liu AQ, Cao JP, Shen YJ. Molecular identification and genetic characteristics of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in human immunodeficiency virus/acquired immunodeficiency syndrome patients in Shanghai, China. Parasit Vectors 2023; 16(1): 53. |
| 6. | Asghari A, Sadeghipour Z, Hassanipour S, Abbasali Z, Parikhani HE, Hashemzaei M, et al. Association between Blastocystis sp. infection and immunocompromised patients: A systematic review and meta-analysis. Environ Sci Pollut Res Int 2021; 28(43): 60308-60328. |
| 7. | Ryan UM, Feng YY, Fayer R, Xiao LH. Taxonomy and molecular epidemiology of Cryptosporidium and Giardia-a 50 year perspective (1971-2021). Int J Parasitol 2021; 51(13-14): 1099-1119. |
| 8. | Li S, Wang P, Zhu XQ, Zou Y, Chen XQ. Prevalence and genotypes/subtypes of Enterocytozoon bieneusi and Blastocystis sp. in different breeds of cattle in Jiangxi Province, southeastern China. Infect Genet Evol 2022, 98:105216. |
| 9. | Ryan U, Zahedi A, Feng Y, Xiao LH. An update on zoonotic Cryptosporidium species and genotypes in humans. Animals (Basel), 2021; 11(11): 3307. |
| 10. | Jinatham V, Maxamhud S, Popluechai S, Tsaousis A, Gentekaki E. Blastocystis One Health approach in a rural community of northern Thailand: Prevalence, subtypes and novel transmission routes. Front Microbiol 2021; 12: 746340. |
| 11. | Roberts M, Dobson A, Restif O, Wells K. Challenges in modelling the dynamics of infectious diseases at the wildlife-human interface. Epidemics 2021; 37: 100523. |
| 12. | Ryan U, Xiao LH, Read C, Zhou L, Lal AA, Pavlasek I. Identification of novel Cryptosporidium genotypes from the Czech Republic. Appl Environ Microbiol 2003; 69(7): 4302-4307. |
| 13. | Cacciò SM, Beck R, Lalle M, Marinculic A, Pozio E. Multilocus genotyping of Giardia duodenalis reveals striking differences between assemblages A and B. Int J Parasitol 2008; 38(13): 1523-1531. |
| 14. | Buckholt MA, Lee JH, Tzipori S. Prevalence of Enterocytozoon bieneusi in swine: An 18-month survey at a slaughterhouse in Massachusetts. Appi Environ Microbiol 2002; 68(5): 2595-2599. |
| 15. | Menounos P G, Spanakos G, Tegos N, et al. Direct detection of Blastocystis sp. in human faecal samples and subtype assignment using single strand conformational polymorphism and sequencing. Mol Cell Probes 2008; 22(1): 24-29. |
| 16. | Conrad CC, Stanford K, Narvaez-Bravo C, Neumann NF, Munns K, Tymensen L, et al. Zoonotic fecal pathogens and antimicrobial resistance in Canadian petting zoos. Microorganisms 2018; 6(3): 70. |
| 17. | Zhang KH, Zheng SG, Wang YL, Wang K, Wang YX, Gazizova A, et al. Occurrence and molecular characterization of Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi, and Blastocystis sp. in captive wild animals in zoos in Henan, China. BMC Vet Res 2021; 17(1): 332. |
| 18. | Karim MR, Li JQ, Rume FI, Sumon SMR, Selim ASM, Hoda N, et al. Occurrence and molecular characterization of Cryptosporidium spp. and Giardia duodenalis among captive mammals in the Bangladesh National Zoo. Parasitol Int 2021; 84: 102414. |
| 19. | Lu YX, Pu TC, Ma BH, Wang LX, Zhou MC, Chen Y, et al. A survey of Cryptosporidium prevalence among birds in two zoos in China. Peer J 2022; 10: e12825. |
| 20. | Köster PC, Martínez-Nevado E, González A, Abelló-Poveda MT, Fernández-Bellon H, de la Riva-Fraga M, et al. Intestinal protists in captive Non-human primates and their handlers in six european zoological gardens. Molecular evidence of zoonotic transmission. Front Vet Sci 2021; 8: 819887. |
| 21. | Hussain S, Li X, Bukhari SM, Zhou M, Ahmad S, Javid A, et al. Cross-genera amplification and identification of Colpodella sp. with Cryptosporidium primers in fecal samples of zoo felids from northeast China. Braz J Biol 2021; 83: e247181. |
| 22. | Li JQ, Qi M, Chang YK, Wang RJ, Li TY, Dong HJ, et al. Molecular Characterization of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in Captive Wildlife at Zhengzhou Zoo, China. J Eukary Microbiol 2015; 62(6): 833-839. |
| 23. | Menu E, Davoust B, Mediannikov O, Akiana J, Mulot B, Diatta G, et al. Occurrence of ten protozoan enteric pathogens in three non-human primate populations. Pathogens 2021; 10(3): 280. |
| 24. | Rudzinska M, Kowalewska B, Waleron M, Kalicki M, Sikorska K, Szostakowska B. Molecular characterization of Blastocystis from animals and their caregivers at the Gdańsk Zoo (Poland) and the assessment of zoonotic transmission. Biology (Basel), 2021; 10(10): 984. |
| 25. | Danišová O, Valenčáková A, Kandráčová P, Tomko M, Sučik M. First report of Blastocystis spp. subtypes in zoo animals in Slovakia, Central Europe. Ann Agric Environ Med 2022; 29(1): 149-151. |
| 26. | Deng L, Yao JX, Chen SY, He TM, Chai YJ, Zhou ZY, et al. First identification and molecular subtyping of Blastocystis sp. in zoo animals in southwestern China. Parasit Vectors 2021; 14(1): 11. |
| 27. | Li XD, Zou Y, Pan J, Li QL, Zeng Z, Meng YM, et al. Prevalence and subtypes of Blastocystis sp. infection in zoo animals in three cities in China. Parasitol Res 2020; 119(2): 465-471. |
| 28. | Zhu WN, Wei ZY, Li Q, Lin YC, Yang H, Li W. Prevalence and subtype diversity of Blastocystis in human and nonhuman primates in North China. Parasitol Res 2020; 119(8): 2719-2725. |
| 29. | Li JQ, Karim MR, Li DF, Rahaman Sumon SMM, Siddiki SHMF, Rume FI, et al. Molecular characterization of Blastocystis sp. in captive wildlife in Bangladesh National Zoo: Non-human primates with high prevalence and zoonotic significance. Int J Parasitol Parasites Wildl 2019; 10: 314-320. |
| 30. | Ma L, Zhang YB, Qiao HX, Li S, Wang HQ, Zhang NN. Cockroach as a vector of Blastocystis sp. is Risk for golden monkeys in zoo. Korean J Parasitol 2020; 58(5): 583-587. |
| 31. | Oliveira-Arbex AP, David ÉB, Tenório MDS, Cicchi PJP, Patti M, Coradi ST, et al. Diversity of Blastocystis subtypes in wild mammals from a zoo and two conservation units in southeastern Brazil. Infect Genet Evol 2020; 78: 104053. |
| 32. | Parkar U, Traub RJ, Vitali S, Elliot A, Levecke B, Robertson I, et al. Molecular characterization of Blastocystis isolates from zoo animals and their animal-keepers. Vet Parasitol 2010; 169(1-2): 8-17. |
| 33. | Köster PC, Dashti A, Bailo B, Muadica AS, Maloney JG, Santín M, et al. Occurrence and genetic diversity of protist parasites in captive non-human primates, zookeepers, and free-living sympatric rats in the Cordoba Zoo Conservation Centre, Southern Spain. Animals (Basel) 2021; 11(3): 700. |
| 34. | Cian A, El Safadi D, Osman M, Moriniere R, Gantois N, Benamrouz-Vanneste S, et al. Molecular epidemiology of Blastocystis sp. in various animal groups from two French zoos and evaluation of potential zoonotic risk. PLoS One 2017; 12(1): e0169659. |
| 35. | Yang X, Guo YQ, Xiao LH, Feng YY. Molecular epidemiology of human Cryptosporidiosis in low-and middle-income countries. Clin Microbiol Rev 2021; 34(2): e00087-19. |
| 36. | Fayer R, Santín M, Macarisin D. Cryptosporidium ubiquitum n. sp. in animals and humans. Vet Parasitol 2010; 172(1-2): 23-32. |
| 37. | Tang Y, Li N, Song MX, Roelling DM, Feng YY, Xiao LH. Development of a multilocus sequence typing tool for high-resolution subtyping and genetic structure characterization of Cryptosporidium ubiquitum. Infect Genet Evol 2016; 45: 256-261. |
| 38. | KváčM, Kestřánová M, Pinková M, Květoňová D, Kalinová J, Wagnerová P, et al. Cryptosporidium scrofarum n. sp. ( Apicomplexa: Cryptosporidiidae) in domestic pigs (Sus scrofa). Vet Parasitol 2013; 191(3-4): 218-227. |
| 39. | Kvác M, Kvetonová D, Sak B, Ditrich O. Cryptosporidium pig genotype II in immunocompetent man. Emerg Infect Dis 2009; 15(6): 982-983. |
| 40. | Adamu H, Petros B, Zhang GQ, Kassa H, Amer S, Ye JB, et al. Distribution and clinical manifestations of Cryptosporidium species and subtypes in HIV/AIDS patients in Ethiopia. PLoS Negl Trop Dis 2014; 8(4): e2831. |
| 41. | Chacín-Bonilla L. Importance of the different species and genotypes of Cryptosporidium in public health. Invest Clin 2002; 43(2): 67-69. |
| 42. | Zhao W, Zhou HH, Jin HR, Sun LJ, Li PY, Liu MC, et al. Genotyping of Enterocytozoon bieneusi among captive long-tailed macaques (Macaca fascicularis) in Hainan Province: High genetic diversity and zoonotic potential. Acta Trop 2020; 201: 105211. |
| 43. | Zhao W, Zhou HH, Yang L, Ma TM, Zhou JG, Liu HJ, et al. Prevalence, genetic diversity and implications for public health of Enterocytozoon bieneusi in various rodents from Hainan Province, China. Parasit Vectors 2020; 13(1): 438. |
| 44. | Zhao W, Zhang WZ, Wang RJ, Liu WS, Liu AQ, Yang D, et al. Enterocytozoon bieneusi in sika deer (Cervus nippon) and red deer (Cervus elaphus): Deer specificity and zoonotic potential of ITS genotypes. Parasit Res 2014; 113(11): 4243-4250. |
| 45. | Popruk S, Adao DEV, Rivera WL. Epidemiology and subtype distribution of Blastocystis in humans: A review. Infect Genet Evol 2021; 95: 105085. |
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]
[Table 1], [Table 2]
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