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Purpose: Blastocystis species are widely distributed micro-eukaryote parasites found in both human and nonhuman primates. Despite having a global distribution, descriptions of Blastocystis subtype diversity in neotropical primates is largely limited to captive animals. The aim of this study was to molecularly characterize the presence of Blastocystis in free-ranging black-headed night monkeys, Aotus nigriceps, and to analyze Blastocystis heterogeneity in primates of the Parvorder Platyrrhini. Methods: We analyzed Blastocystis small sub-unit ribosomal DNA (SSUrDNA) from both A. nigriceps and Azara's night monkey, A. azarae boliviensis, in Southeastern Peru. We also included additional Blastocystis sequence from other neotropical primate studies to explore the distribution and host specificity of Blastocystis subtypes (ST) throughout the neotropics. Results: Thirteen percent of A. nigriceps samples were Blastocystis positive. Only ST8 was amplified in A. nigriceps and this partial DNA sequence was highly similar to ST8 sequence previously obtained from a human in Brazil. In our analysis of all available Blastocystis SSU sequences from primates of the Parvorder Platyrrhini, we found 15 monophyletic lineages corresponding to previously described subtypes ST1-ST10, ST12-15, and ST17. Conclusions: Blastocystis SSU sequences amplified from A. nigriceps fecal samples shared high sequence similarity to isolates found in several other neotropical primates, Alouatta palliata, A. caraya, Ateles fusciceps, and Lagothrix. Similar subtypes have been found in human and captive primates which supports the possibility of transmission when in close contact. Expanded sampling of sympatric neotropical primates in the wild will establish whether subtypes and clades are limited to taxonomic group or whether transmission occurs between overlapping species.
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Molecular characterization of Blastocystis in captive and free-ranging New World Primates, Platyrrhini
William D. Helenbrook1,2 *; Christopher M. Whipps2
1 Tropical Conservation Fund, Marietta, GA 30064, USA; 2Fish and Wildlife Disease Laboratory, State University
of New York College of Environmental Science and Forestry, Syracuse, NY, USA
*Corresponding author,,
Blastocystis species are widely distributed micro-eukaryote parasites found in both human and nonhuman primates.
Despite having a global distribution, descriptions of Blastocystis subtype diversity in neotropical primates is largely
limited to captive animals. The aim of this study was to molecularly characterize the presence of Blastocystis in free-
ranging black-headed night monkeys, Aotus nigriceps, and to analyze Blastocystis heterogeneity in primates of the
Parvorder Platyrrhini.
We analyzed Blastocystis small sub-unit ribosomal DNA (SSUrDNA) from both A. nigriceps and Azara’s night
monkey, A. azarae boliviensis, in Southeastern Peru. We also included additional Blastocystis sequence from other
neotropical primate studies to explore the distribution and host specificity of Blastocystis subtypes (ST) throughout
the neotropics.
Thirteen percent of A. nigriceps samples were Blastocystis positive. Only ST8 was amplified in A. nigriceps and this
partial DNA sequence was highly similar to ST8 sequence previously obtained from a human in Brazil. In our
analysis of all available Blastocystis SSU sequences from primates of the Parvorder Platyrrhini, we found fifteen
monophyletic lineages corresponding to previously described subtypes ST1-ST10, ST12-15, and ST17.
Blastocystis SSU sequences amplified from A. nigriceps fecal samples shared high sequence similarity to isolates
found in several other neotropical primates, Alouatta palliata, A. caraya, Ateles fusciceps, and Lagothrix. Similar
subtypes have been found in human and captive primates which supports the possibility of transmission when in
close contact. Expanded sampling of sympatric neotropical primates in the wild will establish whether subtypes and
clades are limited to taxonomic group or whether transmission occurs between overlapping species.
Keywords: Blastocystis; zoonotic disease; transmission; primates; Aotus
We would like to thank the following organizations for hosting us: Asociación para la Conservación de la Cuenca
Amazónica, Crees, ARCAmazon, and Taricaya Eco Reserve. And thank you to Jessica Suarez for her field support.
Logistical support was provided by the Tropical Conservation Fund.
Conflicts of interest
Both authors state that there is no conflict of interest.
Authors’ contributions
Study was conceived by WDH. All field and laboratory research was conducted by WDH. First draft was written by
WDH. Figure construction and revisions contributed by CMW. Both authors read and approved the final
Ethics approval
We carried out data collection and analysis in accordance with the legal requirements of Peru, and with permission
of Villa Carmen and Los Amigos Biological Field Stations, MLC, ArcAmazon, and Taricaya.
Availability of data
The data that support the findings of this study are openly available in NCBI at
Blastocystis species hereafter referred to simply as Blastocystis are some of the most common protists
found in vertebrate hosts [1-2]. Molecular characterization of these parasites has uncovered a complex assortment of
subtypes (STs) representing unique phylogenetic lineages [3]. To date, there are 17 officially accepted subtypes and
another five which are likely valid. Several others are presumably invalid, including ST18-20 and ST22 [4]. The
presence of this parasite in wildlife is of great interest because the impact on host health is still poorly understood
and the potential for zoonotic transmission is still debated [5-6].
There is continued debate on the pathogenicity of Blastocystis, particularly in humans [7-8]. Subtype may
be important, with some evidence to suggest that specific parasite proteins and associated proteases may be
responsible for pathogenicity [9-10]. Certain subtypes are also more often associated with disease in humans than
others [11]. Blastocystis in wildlife and captive animals is often characterized as asymptomatic [12]. Yet researchers
continue to be interested in Blastocystis infections and genetic heterogeneity because it is considered a possible
reservoir for human infections [1]. All but one human subtype (ST9) has been found in other animals [13]. And,
ST1, ST2, and ST3 are widely found in both human and nonhuman primates [6]. We have previously characterized
the widespread occurrence of Blastocystis in sympatric human and a non-human neotropical primates but found no
evidence of zoonotic transmission [5]. For example, only ST8 was found in howler monkeys, Alouatta palliata, and
ST1-ST3 were found in humans living nearby.
There are currently eleven described night monkey species Aotus spp. that are distributed from Panama to
Argentina. Several studies have morphologically examined Blastocystis in Aotus species, but none of them have
molecularly characterized Blastocystis in any free-ranging night monkey species [13-15]. Therefore, the purpose of
our study was to i) characterize Blastocystis sequence data from free-ranging black-headed night monkeys Aotus
nigriceps and assess subtype, ii) examine the relationship of Blastocystis subtypes with other neotropical primates in
the Parvorder Platyrrhini, and iii) examine whether Blastocystis subtype is restricted to taxon or by geography.
We collected twenty-three fecal samples from free-ranging black-headed night monkeys, Aotus nigriceps,
in Southeastern Peru and a single sample from the Azara’s night monkey, Aotus azarae boliviensis. All samples
were molecularly characterized previously to confirm host species. We used primer sets modified from Babb et al.
[16] to amplify partial sequence from the cytochrome c oxidase subunit II (COII) gene of each monkey sample.
Twelve of the A. nigriceps samples were obtained from a single group at the Villa Carmen Biological Research
Station (VC: 12°53'39"S, 71°24'16"W), three samples from a single group at the Los Amigos Biological Station
(LA: 12°34'2"S, 70° 5'40.39"W), one sample from Crees - Manu Learning Centre, MLC (12°47′22″S 71°23′32″W)
in the Manu National Park Cultural Zone, and six samples from four groups at ARCAmazon - Las Piedras Amazon
Center (LPAC: 12°04′12″S 69°29′37″W). We collected a single sample from an A. azarae boliviensis group at
Taricaya Eco Reserve (12°31′08″S 68°58′48″W). Field sites are dispersed across the Madre de Dios region, with the
most distant sites approximately 266km apart. All samples are recovered from cotton or plastic netting placed below
nesting sites or under areas where groups were observed passing overhead in order to avoid soil contamination. Nets
were checked within twelve hours, at least twice a day. All samples were preserved in RNAlater™ Stabilization
Solution Qiagen Inc. Valencia, CA.
We used Quick-DNA Fecal/Soil Microbe Kits to extract parasite DNA following the protocol provided by
the manufacturer Zymo Research, Corp, Irvine, CA. Blastocystis small sub-unit ribosomal DNA SSUrDNA was
amplified from A. nigriceps fecal samples using previously published primers BH1F [17] and BHRDr [18]. DNA
was amplified by PCR in 50 μL reaction volumes in Taq 2× Master Mix New England Biolabs, Ipswich, MA, 0.25
μM of each primer, and 3 μL of template DNA using an C1000 Thermal Cycler BioRad for 40 cycles at 95 °C for 30
s, 53 °C for 60 s, and 68 °C for 60 s, preceded by an initial denaturation at 95 °C for 3 min, and followed by a final
extension at 68 °C for 7 min. Product amplification was initially evaluated by agarose gel electrophoresis. Negative
samples were run twice more with the same PCR regime.
Amplified products (356bp) were purified with E.Z.N.A. Cycle Pure Kit Omega Bio-Tek, Inc. Norcross,
Georgia, and purified PCR products sequenced in both directions using BH1F and BHRDr. The DNA Analysis
Facility at Yale ran sequences using the ABI BigDye Terminator Cycle Sequencing Ready Reaction Kit v3.1, on the
3730xl DNA Analyzer Applied Biosystems, Thermo Fischer Scientific, Inc. with a 96-capillary 50cm array.
Sequences were screened in FinchTV 1.4.0 and edited in JalView to identify base-calling errors. We compared
forward and reverse sequence to ensure maximum sequence length and integrity. Blastocystis subtypes (ST) were
assigned using both BLASTn analysis of National Center for Biotechnology Information NCBI and the Blastocystis
18S and Sequence Typing (PubMLST) database (
We analyzed thirty-two Blastocystis SSUrDNA sequences in our phylogenetic analysis, including three A.
nigriceps isolates which were amplified in this study and other homologous Blastocystis sequences described in
neotropical primates (Table 1; Fig. 1). Blastocystis SSUrDNA sequences were obtained from the National Center for
Biotechnology Information (NCBI), deposited by nine other studies and representing eleven known primate species.
We also included sequence from several other unrepresented subtypes for reference purposes, including from hosts
not found in South America: Pan troglodytes, Gorilla gorilla, Macaca nemestrina, Cercopithecus aethiops, Canus
lupus, and Ctenodactylus gundi. Homologous sequence was not available for ST11. Some Blastocystis sequences
from neotropical primates were not included in our analysis either because they represented non-overlapping regions
of SSUrDNA, the overlapping sequence was insufficiently small, or they were identical to other sequences from the
same host species. Most samples were from captive individuals with the exception of two wild samples (e.g.
JQ74945 and MH784444: Table 1). We used MAFFT (version 7.407_1) for sequence alignment and curated with
trimAI (version 1.41) [19]. The phylogenetic tree was built using distance-based inference of phylogenetic trees with
combined PhyML + SMS programs (version 1.8.1_1). Phylogenetic trees were arranged in TreeGraph2 [20], and
final editing was carried out using Adobe Illustrator Adobe Systems Inc. San Jose, California.
Of 23 A. nigriceps samples that we screened, three were positive for Blastocystis 13.0% (Table 1). We
found a single infection at three sampled sites (VC, MLC, LA), and no infections were found at LPAC or Taricaya
(Fig. 1). A single A. azarae boliviensis sample we obtained was negative for Blastocystis. Prevalence varied from
8.3% at VC (1/12), to 100% at MLC (1/1), 33.3% at LA (1/3), 0% at LPAC (0/6) and 0% at Taricaya (0/1). All three
amplified Blastocystis sequences (MT509449-51) were identical across overlapping sequence (i.e. 451 bp in length)
and shared 99-100% identity with several other host species, including humans, dogs, and goats. A total of 28 Aotus
samples have been tested for Blastocystis including the 24 from our study, and 4 from Valenca-Barbosa et al. [13]
and only five of these were positive (17.9%), of which 20% were ST1 and 80% were ST8.
Combining our Aotus findings with nine previously published reports from neotropical primates, 160 out of
360 (47.5%) samples were Blastocystis positive (Table 1). This calculation does not include studies which fail to
specify overall sample size. Alouatta spp. were the mostly widely sampled neotropical primates (N=328) of which
one hundred fifty-one samples (46.0%) were positive with Blastocystis. Three percent (2.6%) of positive Alouatta
samples were ST1, 53.6% were positive for ST2, 1.3% were ST4 positive, 41.7% were positive with ST8 and a
single individual had ST3. Three studies recovered ten positive samples from Lagothrix lagotricha and described
ST1, ST2, ST3, and ST8. There were eight samples from Ateles all of which were positive for Blastocystis (i.e.
ST1, ST2, ST3, ST8), and a sample with multiple subtypes. Of all studies included in our analysis, only four others
have assessed Blastocystis subtypes in free-ranging neotropical primates (Table 1).
The rooted phylogenetic tree resulted in fifteen monophyletic lineages corresponding to previously
described subtypes ST1-ST10, ST12-15, and ST17 (Fig. 1). Isolates from the same subtype clustered with each other
with good bootstrap support. The variability within subtype suggests inter-species variation with one exception
ST8 sequence recovered from Alouatta palliata more closely resembled that from Lagothrix sp. than from Alouatta
caraya. Lagothrix sp. samples were recovered from laboratory animals and origin is unknown. Our newly amplified
samples clustered together with other ST8 samples, though the closest sequence similarity with a human sample
recovered in Brazil (MN585875).
We molecularly characterized three Blastocystis positive samples in A. nigriceps which were all ST8 - a
subtype quite common in neotropical primates and relatively rare in humans [1; 21]. No other studies have
molecularly characterized Blastocystis in free-ranging Aotus species, though a single zoo study of an unnamed Aotus
species found both ST1 and ST8 [13]. Other studies have analyzed the presence of Blastocystis morphologically in
Aotus species [22]; yet expanded molecular analysis of the eleven currently recognized night monkey species is
needed to determine whether our results, along with those from Valenca-Barbosa et al. (2019) [13], are broadly
applicable to other members of the genus.
Small sub-unit ribosomal Blastocystis sequence amplified from A. nigriceps fecal samples shared high
sequence similarity to isolates found in several other neotropical primates, Alouatta palliata, A. caraya, Ateles
fusciceps, and Lagothrix (Fig. 1). Similar subtypes have been found in human and captive primates which support
the possibility of transmission when in close contact [e.g. 1, 23]. However, to our knowledge, there is no evidence to
suggest that free-ranging nonhuman primates are acting as a reservoir for Blastocystis in nearby communities. All of
the subtypes described in neotropical primates ST1, ST2, ST3, ST8 have been reported in both Old World primates
Catarrhini and in humans [1]. However, distinct subtype strains are likely found in people and nonhuman primates
indicating cryptic host specificity [5, 24]. For example, ST8 is rare in people but quite common in neotropical
primates. And yet, we previously found people living sympatrically with another nonhuman primate species
(Alouatta palliata) did not share the same subtypes [5].
The presence of Blastocystis subtypes in captive animals may not represent those found in the wild for
several reasons. First, only three subtypes have been described from free-ranging neotropical primates ST1, ST2 and
ST8, whereas ST1, ST2, ST3, ST4, and ST8 have been described in captive individuals. Four different primate
species housed at the Rio Zoo all shared the same subtype (ST8) with 100% sequence similarity MG280767-68 and
MG280770-71 [13]. These sequences are most similar to those recovered from other captive neotropical primates.
Similarly, sequence characterized in a captive Aotus individual MG280729 shared 100% similarity to ST1 sequence
obtained from other captive primates and people (GenBank accession numbers KY438944 and MN396274
respectively). Similar or identical Blastocystis sequence in primates housed near each other suggests the possibility
of transmission. ST1 is found in people, though molecular testing of animals in captivity along with caretakers
would be necessary to confirm whether these sequences are identical.
Reports in humans suggest an association of Blastocystis with irritable bowel syndrome and other
gastrointestinal problems such as diarrhea [25]. However, we are not aware of any evidence of this in wild
nonhuman primates and a single report in an Old World monkey species describes the pathogenic potential of
Blastocystis [26]. None of the samples that we collected in this study particularly those that were Blastocystis
positive - would suggest gastrointestinal symptoms such as watery or loose stool. Continued monitoring and
expansion of sampling in free-ranging nonhuman primates along with general health examinations, when possible,
would help determine the pathogenic potential of this microparasite.
This study found low prevalence of Blastocystis ST8 in free-ranging black-headed night monkeys
compared to several other neotropical primate species. ST1-ST3, and ST8 varied according to host species and
whether in captivity or free-ranging. SSUrDNA sequence provided sufficient variability to distinguish phylogenetic
relationships between Blastocystis subtypes. ST8 in A. nigriceps shared high sequence similarity to other free-
ranging neotropical primate species. However, none of these primates are sympatric with A. nigriceps. Instead, ST8
is widespread, and clustering of isolates appears largely correlated to taxonomic group. Expanded sampling of
sympatric neotropical primates will establish whether subtypes and clades are limited to taxonomic group or whether
transmission occurs between species living in proximity to one another in the wild.
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Fig. 1 Phylogenetic analysis of Blastocystis amplified sequence from SSU-rDNA gene. GenBank accession numbers
and host names are shown at terminal nodes. Values at nodes represent aLRT SH-like supports. *Zoological
... ST5-ST7, ST10, ST14, ST24, ST27, and ST28 in captive wild birds (33), and ST6, ST7, ST10, ST14, ST25, and ST29 in chickens (28). In Peru, ST8 was reported in monkeys (34). In Ecuador ST8 was detected in monkeys (35), and in Colombia ST6 was found in birds, ST8 in marsupials, ST4 in howler monkeys and ST1-ST3 were detected in domesticated mammals (26) and ST1 in dogs (36). ...
... However, studies in South America about Blastocystis specifically in animals are scarce. In some South American countries, there are studies reporting different Blastocystis STs in humans and their relationship with symptomatic patients (54)(55)(56)(57)(58). Countries from the Americas where animal samples have been tested from Blastocystis include United States (16,23,29,59), Brazil (25), Mexico (31), Ecuador (35), Peru (34) and Colombia (26). In those studies, a relative specific association of STs with hosts has been found. ...
Full-text available
Blastocystis is frequently reported in fecal samples from animals and humans worldwide, and a variety of subtypes (STs) have been observed in wild and domestic animals. In Colombia, few studies have focused on the transmission dynamics and epidemiological importance of Blastocystis in animals. In this study, we characterized the frequency and subtypes of Blastocystis in fecal samples of domestic animals including pigs, minipigs, cows, dogs, horses, goats, sheep, and llama from three departments of Colombia. Of the 118 fecal samples included in this study 81.4% ( n = 96) were positive for Blastocystis using a PCR that amplifies a fragment of the small subunit ribosomal RNA ( SSU rRNA) gene. PCR positive samples were sequenced by next generation amplicon sequencing (NGS) to determine subtypes. Eleven subtypes were detected, ten previously reported, ST5 (50.7%), ST10 (47.8%), ST25 (34.3%), ST26 (29.8%), ST21 (22.4%), ST23 (22.4%), ST1 (17.9%), ST14 (16.4%), ST24 (14.9%), ST3 (7.5%), and a novel subtype, named ST32 (3.0%). Mixed infection and/or intra -subtype variations were identified in most of the samples. Novel ST32 was observed in two samples from a goat and a cow. To support novel subtype designation, a MinION based sequencing strategy was used to generate the full-length of the SSU rRNA gene. Comparison of full-length nucleotide sequences with those from current valid subtypes supported the designation of ST32. This is the first study in Colombia using NGS to molecularly characterize subtypes of Blastocystis in farm animals. A great diversity of subtypes was observed in domestic animals including subtypes previously identified in humans. Additionally, subtype overlap between the different hosts examined in this study were observed. These findings highlight the presence of Blastocystis subtypes with zoonotic potential in farm animals indicating that farm animals could play a role in transmission to humans.
... Interestingly, ST1-ST3 distribution was independent of NHP group or geographical association, ST5 was seen only in apes and old world monkeys, and ST8 primarily in species native to Asia or South America (85). Subsequent surveys have confirmed (13,53,82,84) or extended these preliminary molecular data. For instance, ST7 has been described in cynomolgus monkeys in Italy (50), ST13 in langurs and golden snub-nosed monkeys in Bangladesh (80) and China (83), ST17 in squirrel monkeys in China (86), and ST19 in a rhesus macaque in the latter country (83). ...
Full-text available
We assessed the occurrence, genetic diversity, and zoonotic potential of four protozoan ( Cryptosporidium spp., Entamoeba histolytica, Entamoeba dispar, Giardia duodenalis ), one stramenopile ( Blastocystis sp.), one microsporidia ( Enterocytozoon bieneusi ), and two ciliate ( Balantioides coli, Troglodytella abrassarti ) intestinal parasite or commensal protist species in captive non-human primates (NHP) and their zookeepers from six European zoological gardens in France ( n = 1), Germany ( n = 1), and Spain ( n = 4). Faecal samples from NHP ( n = 454) belonging to 63 species within 35 genera and humans ( n = 70) were collected at two sampling periods in each participating institution between October 2018-August 2021. Detection and species identification was accomplished by PCR and Sanger sequencing of the ssu rRNA and/or ITS genes. Sub-genotyping analyses using specific markers were conducted on isolates positive for G. duodenalis ( gdh, bg, tpi ) and Cryptosporidium spp. ( gp60 ). Overall, 41.0% (186/454) and 30.0% (21/70) of the faecal samples of NHP and human origin tested positive for at least one intestinal protist species, respectively. In NHP, Blastocystis sp. was the most prevalent protist species found (20.3%), followed by G. duodenalis (18.1%), E. dispar (7.9%), B. coli and T. abrassarti (1.5% each), and Cryptosporidium spp. and E. bieneusi (0.9% each). Occurrence rates varied largely among NHP host species, sampling periods, and zoological institutions. The predominant protist species found in humans was Blastocystis sp. (25.7%), followed by Cryptosporidium spp. (2.9%), E. dispar (1.4%), and G. duodenalis (1.4%). Sequencing of PCR-positive amplicons in human and/or NHP confirmed the presence of Cryptosporidium in six isolates ( C. hominis : 66.7%, C. parvum : 33.3%), G. duodenalis in 18 isolates (assemblage A: 16.7%, assemblage B: 83.3%), Blastocystis in 110 isolates (ST1:38.2%, ST2:11.8%, ST3: 18.2%, ST4: 9.1%, ST5: 17.3%, ST8: 2.7%, ST13: 0.9%), and E. bieneusi in four isolates (CM18: 75.0%, Type IV: 25.0%). Zoonotic transmission events involving Blastocystis ST1–ST4 were identified in four zoological institutions. Zoonotic transmission of C. hominis was highly suspected, but not fully demonstrated, in one of them. Monitoring of intestinal protist species might be useful for assessing health status of captive NHP and their zookeepers, and to identify transmission pathways of faecal-orally transmitted pathogens.
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Little information is currently available on the occurrence and genetic diversity of pathogenic and commensal protist species in captive and semi-captive non-human primates (NHP) resident in zoological gardens or sanctuaries in low- and medium-income countries. In this molecular-based study, we prospectively collected individual faecal samples from apparently healthy NHP at the Abidjan Zoological Garden (AZG) in Côte d’Ivoire, the Tacugama Sanctuary (TS) in Sierra Leone, and the Quistococha Zoological Garden (QZG) in Peru between November 2018 and February 2020. We evaluated for the presence of pathogenic (Cryptosporidium spp., Entamoeba histolytica, Giardia duodenalis, Blastocystis sp., Enterocytozoon bieneusi, Balantioides coli) and commensal (Entamoeba dispar, Troglodytella abrassarti) protist species using PCR methods and Sanger sequencing. Giardia duodenalis was the most prevalent species found (25.9%, 30/116), followed by Blastocystis sp. (22.4%, 26/116), and E. dispar (18.1%, 21/116). We detected E. bieneusi (4.2%, 1/24) and T. abrassarti (12.5%, 3/24) only on NHP from AZG. Cryptosporidium spp., E. histolytica, and B. coli were undetected at the three sampling sites investigated here. Sequence analyses revealed the presence of zoonotic sub-assemblages BIII (n = 1) in AZG and BIV (n = 1) in TS within G. duodenalis. We identified Blastocystis subtype ST3 (100%, 6/6) in AZG, ST1 (80.0%, 12/15), ST2 (6.7%, 1/15), and ST3 (13.3%, 2/15) in TS, and ST2 (80.0%, 4/5) and ST3 (20.0%, 1/5) in QZG. The only E. bieneusi isolate detected here was identified as zoonotic genotype CAF4. Our PCR-based data indicate that potentially pathogenic protist species including G. duodenalis, Blastocystis sp., E. bieneusi, and B. coli are present at variable rates in the three NHP populations investigated here. The identification of zoonotic genotypes within these species indicates that human-NHP transmission is possible, although the extent and directionality of these events need to be elucidated in future molecular surveys.
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Blastocystis is a genetically diverse microbial eukaryote thriving in the gut of humans and other animals. While Blastocystis has been linked with gastrointestinal disorders, its pathogenicity remains controversial. Previous reports have suggested that one out of six humans could be carrying Blastocystis in their gut, while the numbers could be even higher in animals. Most studies on Blastocystis are either exclusively targeting the organism itself and/or the associated prokaryotic microbiome, while co-occurrence of other microbial eukaryotes has been mainly ignored. Herein, we aimed to explore presence and genetic diversity of Blastocystis along with the commonly occurring eukaryotes Cryptosporidium, Eimeria, Entamoeba and Giardia in the gut of asymptomatic animals from two conservation parks in the United Kingdom. Building upon a previous study, a total of 231 fecal samples were collected from 38 vertebrates, which included 12 carnivorous and 26 non-carnivorous species. None of the animals examined herein showed gastrointestinal symptoms. The barcoding region of the small subunit ribosomal RNA was used for subtyping of Blastocystis. Overall, 47% of animal species were positive for Blastocystis. Twenty six percent of samples carried more than one subtypes, including the newly identified hosts Scottish wildcat, bongo and lynx. Fifty three percent of samples carried at least another microbial eukaryote. Herewith, we discuss potential implications of these findings and the increasingly blurred definition of microbial parasites.
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Background: Blastocystis is a stramenopile of worldwide significance due to its capacity to colonize several hosts. Based on its high level of genetic diversity, Blastocystis is classified into global ribosomal subtypes (STs). The aim of this study was to conduct a summary of Blastocystis STs and depict their distribution throughout North and South America; we did this by assembling maps and identifying its most common 18S alleles based on diverse studies that had been reported all over the continent and whose Blastocystis-positive samples were obtained from numerous hosts. Results: Thirty-nine articles relating to nine countries from the American continent were considered, revealing that ST1 (33.3%), ST2 (21.9%), ST3 (37.9%), ST4 (1.7%), ST5 (0.4%), ST6 (1.2%), ST7 (1%), ST8 (0.7%), ST9 (0.4%), ST12 (0.3%), Novel ST (1.1%) and Mixed STs (0.2%) occurred in humans. The STs in other animal hosts were ST1 (6.5%), ST2 (6.5%), ST3 (4.7%), ST4 (7.2%), ST5 (15.9%), ST6 (17.3%), ST7 (3.6%), ST8 (20.6%), ST10 (9%), ST14 (3.6%), ST17 (1.1%) and Novel ST (4%). The countries that presented the most abundant variety of studies reporting STs were the USA with 14 STs, Brazil with 9 STs and Colombia with 8 STs. Additionally, new variants had been described in the last few years, which have increased the prevalence of these subtypes in the countries studied, such as Novel ST (1.1%) and Mixed STs (0.2%) in humans and Novel ST (4%) in animals. Conclusions: This summary updates the epidemiological situation on the distribution of Blastocystis STs in North and South America and will augment current knowledge on the prevalence and genetic diversity of this protozoan.
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The enteric protist Blastocystis is one of the most frequently reported parasites infecting both humans and many other animal hosts worldwide. A remarkable genetic diversity has been observed in the species, with 17 different subtypes (STs) on a molecular phylogeny based on small subunit RNA genes (SSU rDNA). Nonetheless, information regarding its distribution, diversity and zoonotic potential remains still scarce, especially in groups other than primates. In Brazil, only a few surveys limited to human isolates have so far been conducted on Blastocystis STs. The aim of this study is to determine the occurrence of Blastocystis subtypes in non-human vertebrate and invertebrate animal groups in different areas of the state of Rio de Janeiro, Brazil. A total of 334 stool samples were collected from animals representing 28 different genera. Blastocystis cultivated samples were subtyped using nuclear small subunit ribosomal DNA (SSU rDNA) sequencing. Phylogenetic analyses and BLAST searches revealed six subtypes: ST5 (28.8%), ST2 (21.1%), ST1 and ST8 (19.2%), ST3 (7.7%) and ST4 (3.8%). Our findings indicate a considerable overlap between STs in humans and other animals. This highlights the importance of investigating a range of hosts for Blastocystis to understand the eco-epidemiological aspects of the parasite and its host specificity.
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The study of parasites is of great relevance to primatology given their ecological significance and their effects on primate demography, behavior, and evolution. Moreover, assessing the vulnerability of endangered species to parasitic infections is important in developing appropriate conservation strategies. We conducted an intensive bibliographical search to synthesize the available information about the parasites of Neotropical primates. We analyzed the host and parasite taxonomic coverage of the available studies, examined the advantages and disadvantages of the diagnostic techniques employed, identified information gaps that need to be addressed, and recommend future directions in the parasitological research of Neotropical primates. Researchers have reported 276 parasite taxa, including endo- and ectoparasites, in 21 of the 22 genera of Neotropical primates. Of these, 42 parasite species have also been reported in humans, although this number may be inaccurate owing to misidentification. The parasites of 50% of Neotropical primate species are completely unknown, and 32% of the parasites recorded in these hosts have not been identified to the species level. Information regarding ectoparasites is particularly limited. We need to develop methods that enhance parasite diagnosis accuracy when using noninvasive samples, and the incorporation of molecular techniques in routine procedures should be a priority in parasitological studies of Neotropical primates. An integrative approach in which veterinarians, primatologists, and parasitologists collaborate in the identification and treatment of parasites of Neotropical primates is essential to achieve significant progress in this field. © 2018 Springer Science+Business Media, LLC, part of Springer Nature
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Background Blastocystis is a group of cosmopolitan gastrointestinal parasite of humans and a wide variety of animals. These anaerobic protozoans include more than 17 specific small-subunit ribosomal RNA subtypes, of which nine are found in humans with a variable geographical distribution. Until now, no study has described the Blastocystis subtypes present in Saudi Arabia. Methods In total, 1,262 faecal samples were collected from patients with gastrointestinal complaints and asymptomatic individuals visiting two major hospitals. All samples were analysed by F1/R1 diagnostic PCR, microscopy and culture methods. The subtypes of Blastocystis sp. isolates were determined by the sequenced-tagged site (STS)-based method. ResultsOne-hundred-thirty-three positive cases were detected by F1/R1 diagnostic PCR, of which 122 were also positive by the culture method and 83 by direct microscopy. The sensitivities of direct microscopy and the culture method were 62% and 92%, respectively. Subtype (ST3) was the most prevalent (80.5%), followed by ST1 (14.5%) and ST2 (5%). ST4, ST5, ST6 and ST7 were not detected in this study. ST3 infections were significantly predominant (P < 0.05) among symptomatic patients. Conclusions To our knowledge, this study provides the first run-through information on Blastocystis sp. epidemiology in Makkah city, revealing a rather moderate prevalence of 10.5% and the presence of three subtypes, ST1, ST2, and ST3. ST3 was the most predominant, particularly among symptomatic patients.
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Blastocystis sp. is a common intestinal parasite infecting humans and a wide range of animals worldwide. It exhibits an extensive genetic diversity and 17 subtypes (STs) have thus far been identified in mammalian and avian hosts. Since several STs are common to humans and animals, it was proposed that a proportion of human infections may result from zoonotic transmission. However, the contribution of each animal source to human infection remains to be clarified. Therefore, the aim of this study was to expand our knowledge of the epidemiology and host specificity of this parasite by performing the largest epidemiological survey ever conducted in animal groups in terms of numbers of species screened. A total of 307 stool samples from 161 mammalian and non-mammalian species in two French zoos were screened by real-time PCR for the presence of Blastocystis sp. Overall, 32.2% of the animal samples and 37.9% of the species tested were shown to be infected with the parasite. A total of 111 animal Blastocystis sp. isolates were subtyped, and 11 of the 17 mammalian and avian STs as well as additional STs previously identified in reptiles and insects were found with a varying prevalence according to animal groups. These data were combined with those obtained from previous surveys to evaluate the potential risk of zoonotic transmission of Blastocystis sp. through the comparison of ST distribution between human and animal hosts. This suggests that non-human primates, artiodactyls and birds may serve as reservoirs for human infection, especially in animal handlers. In contrast, other mammals such as carnivores, and non-mammalian groups including reptiles and insects, do not seem to represent significant sources of Blastocystis sp. infection in humans. In further studies, more intensive sampling and screening of potential new animal hosts will reinforce these statements and expand our understanding of the circulation of Blastocystis sp. in animal and human populations.
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Although the presence of cryptic host specificity has been documented in Blastocystis, differences in infection rates and high genetic polymorphism within and between populations of some subtypes (ST) have impeded the clarification of the generalist or specialist specific-ity of this parasite. We assessed the genetic variability and host specificity of Blastocystis spp. in wild howler monkeys from two rainforest areas in the southeastern region of Mexico. Fecal samples of 225 Alouatta palliata (59) and A. pigra (166) monkeys, belonging to 16 syl-vatic sites, were analyzed for infection with Blastocystis ST using a region of the small sub-unit rDNA (SSUrDNA) gene as a marker. Phylogenetic and genetic diversity analyses were performed according to the geographic areas where the monkeys were found. Blastocystis ST2 was the most abundant (91.9%), followed by ST1 and ST8 with 4.6% and 3.5%, respectively ; no association between Blastocystis ST and Alouatta species was observed. SSUrDNA sequences in GenBank from human and non-human primates (NHP) were used as ST references and included in population analyses. The haplotype network trees exhibited different distributions: ST1 showed a generalist profile since several haplotypes from different animals were homogeneously distributed with few mutational changes. For ST2, a major dispersion center grouped the Mexican samples, and high mutational differences were observed between NHP. Furthermore, nucleotide and haplotype diversity values, as well as migration and genetic differentiation indexes, showed contrasting values for ST1 and ST2. These data suggest that ST1 populations are only minimally differentiated, while ST2 populations in humans are highly differentiated from those of NHP. The host generalist and specialist specificities exhibited by ST1 and ST2 Blastocystis populations indicate distinct adaptation processes. Because ST1
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This study characterizes Blastocystis species infections in humans and mantled howler monkeys, Alouatta palliata aequatorialis, living in close proximity to one another in northwestern Ecuador. Blastocystis species were identified from 58 of 96 (60.4 %) mantled howler monkey fecal samples, and 44 of 55 human fecal samples (81.5 %) by polymerase chain reaction. Using single-stranded conformation polymorphism, we were able to efficiently separate and sequence subtypes (STs) within mixed samples without the need for cloning. Blastocystis ST1, ST2, and ST3 were found in people, and two individuals were infected with more than one subtype. All monkey samples were ST8. The lack of shared subtypes between humans and monkeys suggests that no Blastocystis transmission occurs between these species in spite of close proximity in some instances. Based on analysis of demographic data from a questionnaire given to human participants, individuals who boiled their water before consumption were significantly less likely to be infected with Blastocystis (44.4 %) compared to those who did not (93.8 %) (p = 0.002). No other risk factors were significant, although hunters, females, individuals living in large families, and those living closer to forested habitat tended to have a higher proportion of Blastocystis infections.
Blastocystis is a genetically diverse intestinal protist colonizing both human and non-human hosts. By 2013, 17 subtypes had been acknowledged. Since then, nine more subtypes have been proposed. We argue that several recently proposed subtypes are invalid. We also revisit recommendations regarding the requirements for annotating sequences as new subtypes.
The enteric protist Blastocystis is one of the most commonly parasite reported in humans and a variety of animal hosts worldwide. Regarding genetic diversity, at least 17 subtypes (STs) have been identified in mammals and birds, with eight of them (ST1-8) infecting both humans and animals. Recently, isolates from wild mammalian species have been genetically characterized, however data is still scarce, mainly in Latin America. Here, we aimed to verify the occurrence and genetic diversity of Blastocystis in captive wild mammals kept in one zoo and in two units of protection and conservation in southeastern Brazil. A total of 78 fecal samples (14 pooled and 64 individual samples) were recovered from 102 wild mammals of 35 species included in the following orders: Primates, Carnivora, Artiodactyla, Pilosa, Rodentia and Marsupialia. Zoo and units staff were invited to participated but only 16 fecal samples could be screened. Based on the sequence analyses of SSUrDNA gene, out of 29 PCR products from animal samples, 51.7% (15/29) were successfully sequenced and five Blastocystis subtypes were identified as follows: ST1 (2/15; 13.3%), ST2 (2/15; 13.3%), ST3 (4/15; 26.6%), ST5 (2/15; 13.3%) and ST8 (5/14; 33.3%). Only four isolates from humans were sequenced and identified as ST1 (2 isolates), ST2 and ST3. It was observed that Blastocystis infecting non-human primates belong to ST1 and ST2 and mainly to ST3 and ST8, artiodactyls ST5, carnivores ST1 and ST5 and rodents ST1. In addition, this present study reports some interesting findings: (1) 63% (12/19) of Blastocystis isolates from animals and employees belonged to the potentially zoonotic subtypes ST1-ST3; (2) most of these isolates displayed high identity with publicly available DNA sequences from non-human primates and humans, including primate handlers; (3) Blastocystis ST5 was found infecting the northern tiger cat, a native South American felid and one of the species facing a high risk of extinction in Brazil.