Bieneusi Genotypes Research Articles

Multilocus Sequence Typing and Population Genetic Analysis of Enterocytozoon bieneusi: Host Specificity and Its Impacts on Public Health.

Microsporidia comprise a large class of unicellular eukaryotic pathogens that are medically and agriculturally important, but poorly understood. There have been nearly 1,500 microsporidian species described thus far, which are variable in biology, genetics, genomics, and host specificity. Among those, Enterocytozoon bieneusi is the well-known species responsible for the most recorded cases of human microsporidian affections. The pathogen can colonize a broad range of mammals and birds and most of the animals surveyed share some genotypes with humans, posing a threat to public health. Based on DNA sequence analysis of the ribosomal internal transcribed spacer (ITS) and phylogenetic analysis, several hundreds of E. bieneusi genotypes have been defined and clustered into different genetic groups with varied levels of host specificity. However, single locus-based typing using ITS might have insufficient resolution to discriminate among E. bieneusi isolates with complex genetic or hereditary characteristics and to assess the elusive reproduction or transmission modes of the organism, highlighting the need for exploration and application of multilocus sequence typing (MLST) and population genetic tools. The present review begins with a primer on microsporidia and major microsporidian species, briefly introduces the recent advances on E. bieneusi ITS genotyping and phylogeny, summarizes recent MLST and population genetic data, analyzes the inter- and intragroup host specificity at the MLST level, and interprets the public health implications of host specificity in zoonotic or cross-species transmission of this ubiquitous fungus.

Prevalence and genotypic identification of Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi in pre-weaned dairy calves in Guangdong, China

BackgroundCryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi are common enteric pathogens in humans and animals. Data on the transmission of these pathogens are scarce from Guangdong, China, which has a subtropical monsoon climate and is the epicenter for many emerging infectious diseases. This study was conducted to better understand the prevalence and identity of the three pathogens in pre-weaned dairy calves in Guangdong.MethodsThe occurrence and genetic identity of three pathogens were analyzed by polymerase chain reaction. PCR-positive products were sequenced to determine the species and genotypes. A Chi-square test was used to compare the prevalence of pathogens among sampling dates, age groups, or clinical signs.ResultsThe detection rates of Cryptosporidium spp., G. duodenalis and E. bieneusi were 24.0% (93/388), 74.2% (288/388) and 15.7% (61/388), respectively. Three Cryptosporidium species were detected, including C. bovis (n = 73), C. parvum (n = 12) and C. ryanae (n = 7); one animal had concurrence of C. bovis and C. parvum. C. parvum was the dominant species during the first two weeks of life, whereas C. bovis and C. ryanae were mostly seen at 3–9 weeks of age. Sequence analysis identified the C. parvum as subtype IIdA19G1. Assemblage E (n = 282), assemblage A (n = 1), and concurrence of A and E (n = 5) were identified among G. duodenalis-positive animals using multilocus genotyping (MLG). Altogether, 15, 10 and 17 subtypes of assemblage E were observed at the bg, gdh and tpi loci, respectively, forming 49 assemblage E MLGs. The highest detection rate of G. duodenalis was found in winter. Sequence analysis identified genotypes J (n = 57), D (n = 3) and one concurrence of J and D among E. bieneusi-positive animals. The detection rate of E. bieneusi was significantly higher in spring (38.0%; 41/108) than in summer (7.2%; 8/111) and winter (7.1%; 12/169).ConclusionsThese results indicate a common occurrence of C. parvum subtype IIdA19G1, G. duodenalis assemblage E, and E. bieneusi genotype J in pre-weaned dairy calves in Guangdong. More studies are needed to understand the unique genetic characteristics and zoonotic potential of the three enteric pathogens in the province.

Molecular Detection of Zoonotic Microsporidia in Domestic Cats in Turkey: A Preliminary Study.

This preliminary study was conducted to reveal that the molecular identity of Encephalitozoon spp. and Enterocytozoon bieneusi in indoor domestic cats' fecal samples from Turkey was screened using the PCR. Nested PCR was performed using MSP and EBITS primers.All of the amplification products were sequenced to identify the microsporidia species. Four (5.5%) and three (4.1%) genomic DNA isolates of the fecal samples from 72 indoor domestic cats showed amplification of the ITS regions of E. bieneusi and Encephalitozoon spp., respectively. Two different genotypes, D and IV, of E. bieneusi were determined in two cats each based on the ITS sequence analyses. Moreover, Encephalitozoon spp. sequence analyses revealed three isolates belonging to E. intestinalis. Thispreliminary study has provided the first molecular data on the zoonotic genotypes of E. bieneusi and E. intestinalis in cats in Turkey. Furthermore, E. bieneusi genotype IV (accession number MG727664) was submitted to GenBank for the first time in the Western Palearctic Region as hosted by a domestic cat. Additionally, E. intestinalis (accession number MG570080) was also submitted to GenBank as a valid ITS sequence for the first time as hosted by a domestic cat worldwide.

First report and multilocus genotyping of Enterocytozoon bieneusi from Tibetan pigs in southwestern China.

Enterocytozoon bieneusi is a common intestinal pathogen in a variety of animals. While E. bieneusi genotypes have become better-known, there are few reports on its prevalence in the Tibetan pig. This study investigated the prevalence, genetic diversity, and zoonotic potential of E. bieneusi in the Tibetan pig in southwestern China. Tibetan pig feces (266 samples) were collected from three sites in the southwest of China. Feces were subjected to PCR amplification of the internal transcribed spacer (ITS) region. Enterocytozoon bieneusi was detected in 83 (31.2%) of Tibetan pigs from the three different sites, with 25.4% in Kangding, 56% in Yaan, and 26.7% in Qionglai. Prevalence varies according to age group, from 24.4% (age 0–1 years) to 44.4% (age 1–2 years). Four genotypes of E. bieneusi were identified: two known genotypes EbpC (n = 58), Henan-IV (n = 24) and two novel genotypes, SCT01 and SCT02 (one of each). We compare our results with a compilation of published results on the host range and geographical distribution of E. bieneusi genotypes in China. Phylogenetic analysis showed these four genotypes clustered to group 1 with zoonotic potential. Multilocus sequence typing (MLST) analysis of three microsatellites (MS1, MS3, MS7) and one minisatellite (MS4) was successful in 47, 48, 23 and 47 positive specimens and identified 10, 10, 5 and 5 genotypes at four loci, respectively. This study indicates the potential danger of E. bieneusi to Tibetan pigs in southwestern China, and offers basic advice for preventing and controlling infections.

Molecular detection of Enterocytozoon bieneusi in alpacas (Vicugna pacos) in Xinjiang, China.

Enterocytozoon bieneusi, an obligate intracellular pathogen, can infect a wide variety of hosts. This study aimed to determine the prevalence and molecular characteristics of E. bieneusi in alpacas (Vicugna pacos) in China. A total of 185 alpaca fecal samples were collected from five herds in Tacheng, Wensu, Hejing, Qinghe, and Nilka counties in Xinjiang Uygur Autonomous Region. Enterocytozoon bieneusi was detected by nested PCR of the internal transcribed spacer (ITS) region. Twenty-eight fecal samples (15.1%, 28/185) were positive for E. bieneusi, with the highest prevalence in alpacas from Qinghe (42.9%, 15/35). Four E. bieneusi genotypes were identified, which included two known (P and ALP3) and two novel (ALP7 and ALP8) genotypes. Genotype ALP3 was the dominant genotype (57.1%, 16/28), followed by genotypes P (32.1%, 9/28), ALP7 (7.1%, 2/28), and ALP8 (2.6%, 1/28). Phylogenetic analysis revealed that three genotypes (P, ALP7, and ALP3) clustered into group 1, whereas genotype ALP8 clustered into group 8. This is the first report of E. bieneusi infection and genetic diversity in alpacas from Xinjiang, China.

Host-adapted Cryptosporidium and Enterocytozoon bieneusi genotypes in straw-colored fruit bats in Nigeria.

Few data are available on the distribution and human infective potential of Cryptosporidium and Enterocytozoon bieneusi genotypes in bats. In this preliminary study, we collected 109 fecal specimens during April–July 2011 from a colony of straw-colored fruit bats (Eidolon helvum) in an urban park (Agodi Gardens) of Ibadan, Nigeria, and analyzed for Cryptosporidium spp., Giardia duodenalis and E. bieneusi using PCR targeting the small subunit rRNA gene, triosephosphate isomerase gene, and ribosomal internal transcribed spacer, respectively. Genotypes of these enteric parasites were determined by DNA sequencing of the PCR products. Altogether, 6 (5.5%), 0 and 16 (14.7%) specimens were positive for Cryptosporidium spp., G. duodenalis, and E. bieneusi, respectively. DNA sequence analysis of the PCR products indicated the presence of two novel Cryptosporidium genotypes named as bat genotype XIV (in 5 specimens) and bat genotype XV (in 1 specimen) and one known E. bieneusi genotype (Type IV in 1 specimen) and two novel E. bieneusi genotypes (Bat1 in 13 specimens and Bat2 in 2 specimens). In phylogenetic analysis of DNA sequences, the two novel Cryptosporidium genotypes were genetically related to Bat genotype II previously identified in fruit bats in China and Philippines, whereas the two novel E. bieneusi genotypes were genetically related to Group 5, which contains several known genotypes from primates. With the exception of Type IV, none of the Cryptosporidium and E. bieneusi genotypes found in bats in this study are known human pathogens. Thus, straw-colored fruit bats in Nigeria are mainly infected with host-adapted Cryptosporidium and E. bieneusi genotypes.

Prevalence and genetic diversity of Enterocytozoon bieneusi in sheep in China

BackgroundEnterocytozoon bieneusi is a common species of microsporidia that not only influences human health but also threatens animal productive performance and value. However, there have been no systematic studies of the prevalence of E. bieneusi in sheep in China.ResultsA total of 953 fecal specimens were collected from sheep from 11 provinces across five regions of China and analyzed for E. bieneusi by nested PCR targeting the ribosomal internal transcribed spacer (ITS). Enterocytozoon bieneusi infections were detected in four regions, with an overall infection rate of 20.4% (194/953). The highest infection rate was detected in pre-weaned lambs (25.0%), followed by post-weaned lambs (22.2%) and adult sheep (14.6%). Enterocytozoon bieneusi was found in nine of the 11 tested provinces, with infection rates between 2.9–51.7%. Eleven genotypes were identified based on ITS analysis, including seven known genotypes (BEB6, CHG1, CHG3, CHS7, CHS8, COS-I and NESH5) and four novel genotypes (CHHLJS1, CHHLJS2, CHNXS1 and CHXJS1). All 11 genotypes were clustered into group 2, and the zoonotic genotype BEB6 was the dominant genotype (n = 129, 66.5%) in sheep.ConclusionThe prevalence of E. bieneusi was studied in five regions representing most areas where sheep are bred in China. This is the first report of E. bieneusi infection in sheep for seven Chinese provinces. Geographical differences were detected in the distribution of E. bieneusi genotypes, but no differences were found among sheep in different age groups. The zoonotic genotype BEB6 was the dominant genotype, indicating that sheep are a potential source of zoonotic microsporidiosis in China. These results improve our knowledge of the epidemiology of E. bieneusi in sheep in China.

Prevalence and genotypes of Enterocytozoon bieneusi in pigs in southern China

Enterocytozoon bieneusi is one of the most important causative agents of microsporidiosis, causing diarrhoea the symptoms of enteric disease in humans and animals. Although there is some information on the prevalence and genotypes of E. bieneusi in China, there is still a lack of data in pigs in southern China. In the present study, a total of 396 faecal specimens were collected from pigs in Zhejiang, Guangdong and Yunnan provinces in southern China, and were examined by nested PCR amplification of the ribosomal internal transcribed spacer (ITS) for the prevalence and genotypes of E. bieneusi. The overall prevalence of E. bieneusi in pigs was 31.57% (125/396), forming 15 genotypes, including 9 known genotypes (EbpC, EbpA, D, G, H, PigEBITS5, Henan-IV, KIN-1, CHS5) and 6 novel genotypes (GD1, ZJ1, ZJ2, YN1, YN2 and YN3), which were all clustered into Group 1. Moreover, multilocus sequence typing (MLST) showed that 6, 3, 4 and 5 types were identified in MS1, MS3, MS7 and MS4 loci, respectively, representing four multilocus genotypes (MLGs), designated as MLGs novel-1 to novel-4 in the present study. This is the first detailed study of E. bieneusi using MLST in pigs in southern China, which extended information about the distribution of E. bieneusi genotypes in China.

Occurrence and molecular characterization of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi from Tibetan sheep in Gansu, China

Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are globally ubiquitous pathogens. However, little is known about the occurrence and distribution of Cryptosporidium spp., G. duodenalis, and E. bieneusi in Tibetan sheep. In the present study, fecal specimens of 177 Tibetan sheep were examined by nested PCR. 4.5% (n = 8), 1.7% (n = 3) and 34.5% (n = 61) of the Tibetan sheep were positive for Cryptosporidium spp., G. duodenalis, and E. bieneusi, respectively. Cryptosporidium ubiquitum was the only species found by small subunit (SSU) rRNA-based PCR, and subtyping of C. ubiquitum (n = 6) by 60-kDa glycoprotein (gp60) revealed that all positive isolates belonged to zoonotic XIIa subtype 2. Multilocus genotyping at the SSU rRNA and β-giardin (bg) genes suggested that three G. duodenalis positive specimens belonged to assemblage E. Sequences analysis of internal transcribed spacer (ITS) gene characterized four E. bieneusi genotypes which all belonged to Group 2, one novel CGS1 (n = 3), and three known: CM7 (n = 34), BEB6 (n = 22), and CHS3 (n = 2). We employed multilocus sequence typing (MLST) using three microsatellites (MS1, MS3 and MS7), one minisatellite (MS4), and sequence analysis of MLST yielded 3, 2, 2 and 2 genotypes at the MS1, MS3, MS4, and MS7 loci, respectively, forming 4 MLGs. Our findings elucidate the occurrence and distribution of three zoonotic pathogens in Tibetan sheep in China. More subsequent and detailed data are required to better understand the transmission of Cryptosporidium spp., G. duodenalis, and E. bieneusi in sheep.

Intestinal microsporidiosis in Strasbourg from 2014 to 2016: emergence of an Enterocytozoon bieneusi genotype of Asian origin

Microsporidia cause opportunistic infections in highly immunodeficient individuals. Few studies on the epidemiology of these infections have been conducted in France. Between 2014 and 2016, we undertook a study to estimate the prevalence and circulating genotypes of this fungus-related micro-organism among the population of Strasbourg University Hospital. Samples were collected from hospitalized patients and analyzed using microscopy and molecular assays. Strains from positive subjects were sequenced for genotyping. Only 7/661 patients (1.1%) were positive for microsporidia, and the only species identified was Enterocytozoon bieneusi. Two patients presented immunodeficiency linked to AIDS, and five transplant recipients presented immunodeficiency linked to immunosuppressive therapies. Only five patients received specific antimicrosporidial treatment, but clinical outcomes were good in all cases. We identified four genotypes: A and D in patients with AIDS, and C and S9 in transplant recipients. To date, genotype S9 has been described only once. This genotype is similar to those found in farm animals in China. Because some of these animals have been introduced to Central Europe, we postulate that this genotype might be of Asian origin. Thus, genotyping microsporidial strains may be of epidemiological and clinical interest to identify potential outbreak sources depending on the infecting strains.

Molecular characterizations of Cryptosporidium spp. and Enterocytozoon bieneusi in brown rats (Rattus norvegicus) from Heilongjiang Province, China

BackgroundCryptosporidium spp. and Enterocytozoon bieneusi are prevalent zoonotic pathogens responsible for the high burden of diarrheal diseases worldwide. Rodents are globally overpopulated and are known as reservoirs or carriers of a variety of zoonotic pathogens including Cryptosporidium spp. and E. bieneusi. However, few data are available on genetic characterizations of both pathogens in rodents in China. The aim of the present work was to determine the prevalence and genetic characterizations of Cryptosporidium spp. and E. bieneusi in brown rats (Rattus norvegicus) from Heilongjiang, China.MethodsA total of 242 wild brown rats were captured in Heilongjiang Province of China. A fresh fecal specimen was collected directly from the intestinal and rectal content of each brown rat. All the fecal specimens were examined for the presence of Cryptosporidium spp. and E. bieneusi by PCR and sequencing of the partial small subunit (SSU) rRNA gene and the internal transcribed spacer (ITS) region of the rRNA gene of the two pathogens, respectively.ResultsThe infection rate was 9.1% (22/242) for Cryptosporidium spp. and 7.9% (19/242) for E. bieneusi. Sequence analysis confirmed the presence of C. ubiquitum (1/22, 4.5%) and three genotypes of Cryptosporidium, including Cryptosporidium rat genotype I (14/22, 63.6%), Cryptosporidium rat genotype IV (6/22, 27.3%) and Cryptosporidium suis-like genotype (1/22, 4.5%). Meanwhile, two E. bieneusi genotypes were identified, including D (17/19, 89.5%) and Peru6 (2/19, 10.5%).ConclusionsTo the best of our knowledge, Enterocytozoon bieneusi genotype Peru6 was identified in rodents for the first time globally and Cryptosporidium rat genotype I and Cryptosporidium rat genotype IV were found in rats in China for the first time. The finding of zoonotic C. ubiquitum and C. suis-like genotype, as well as E. bieneusi genotypes, suggests that brown rats pose a threat to human health. It is necessary to control brown rat population in the investigated areas and improve local people’s awareness of the transmission risk of the two pathogens from brown rats to humans.

Molecular and phylogenetic evidences of dispersion of human-infecting microsporidia to vegetable farms via irrigation with treated wastewater: One-year follow up.

Human-infecting microsporidia are a group of spore-forming eukaryotic microorganisms that can infect both animals and humans. Recent evidences indicate waterborne transmission of microsporidia spores to human via either drinking water or irrigation of vegetable farms with contaminated water resources. The current study aimed to evaluate the presence of human-infecting microsporidia in treated wastewater (TW) and vegetable farms irrigated with treated wastewater during a year. Totally, twelve samples of each treated wastewater and vegetables were collected. In order to recover microsporidia spores, filtration using cellulose nitrate membrane (pore size 0.4 μm) and sedimentation were employed. DNA extraction was performed for all samples and genus/species were characterized using specific primers. In order to characterize genotypes, ITS fragment of E. bieneusi was amplified, sequenced and compared in GenBank database. Phylogenetic tree was employed to analysis the probable correlation between obtained genotypes with those E. bieneusi genotypes, which were previously obtained from human and animals from same region. After nested PCR, expected fragments of E. bieneusi and Encephalitozoon spp were observed among 5/12 (41.7%) and 1/12 (8.33%) of vegetable samples, respectively. From total of 12 TW samples, expected fragments of E. bieneusi and Encephalitozoon spp were amplified among 7/12 (53.8%) and 1/12 (8.33%) of TW samples, respectively. Genotypes D and E were characterized from both TW and vegetables samples. Phylogenetic analysis showed close-relationship between E. bieneusi from TW and vegetable samples with E. bieneusi from animals and humans obtained from the same region. Our findings suggested the key role of animals in epidemiology of zoonotic transmission of E. bieneusi. Moreover, our findings revealed the occurrence of human-infecting microsporidia in treated wastewater because of either insufficiency of treatment process or distribution of microsporidia spores in wastewater treatment plant via animals.

Genetic diversity within dominant Enterocytozoon bieneusi genotypes in pre-weaned calves

BackgroundCattle are commonly infected with the microsporidian parasite Enterocytozoon bieneusi. Sequence characterization of E. bieneusi in these animals at the ribosomal internal transcribed spacer (ITS) locus had identified I, J and BEB4 as the dominant genotypes. However, current studies on E. bieneusi in dairy cattle are mostly on infection rates and genotype distribution. This study aims to examine the intragenotypic diversity within dominant E. bieneusi genotypes in pre-weaned dairy calves in Shanghai, China.MethodsEnterocytozoon bieneusi genotypes and subtypes were identified by PCR sequence analysis of ITS and multilocus sequence typing (MLST), based on material from farms. Chi-square test was used to examine differences in E. bieneusi infection rates between farms or age groups.ResultsThe overall infection rate of E. bieneusi was 26.5% (214/809), ranging from 12.6% (Farm 5) to 38.5% (Farm 4). Infection rates increased with age during early life, with the peak infection rate (43.0%; 43/100) occurring at six weeks. Four genotypes were present, including J (n = 145, 67.8%), BEB4 (n = 59, 27.6%), CHN4 (n = 4, 1.9%) and CHN15 (n = 1, 0.5%), with the former two belonging to Group 2 and the latter two belonging to Group 1. Differences were detected in the distribution of the dominant genotypes J and BEB4 among five study farms. Altogether, 10 multilocus genotypes (MLGs) were identified in the two dominant ITS genotypes, including MLG-J1 to MLG-J8 of genotype J and MLG-B1 to MLG-B2 of genotype BEB4. MLG-B1 and MLG-B2 were recovered in Farms 1, 2 and 5, whereas MLG-J1 to MLG-J5 and MLG-J6 to MLG-J8 were found in Farms 3 and 4, respectively.ConclusionsThere is extensive genetic heterogeneity within the dominant E. bieneusi genotypes J and BEB4 in dairy calves in Shanghai, China, and MLST should be used in molecular epidemiological studies of E. bieneusi in cattle.

Enterocytozoon bieneusi genotypes in Tibetan sheep and yaks.

Few studies have been conducted on the distribution of Enterocytozoon bieneusi genotypes in Tibetan sheep and yaks, which live outdoors in extreme climate with high altitude. In this study, fecal specimens from 312 Tibetan sheep and 554 yaks in Qinghai, China, were collected and examined for E. bieneusi by PCR-sequence analysis of the ribosomal internal transcribed spacer. Among them, 73 (23.4%) specimens from Tibetan sheep and 40 (7.2%) from yaks were positive for E. bieneusi. There were eight E. bieneusi genotypes in Tibetan sheep, including three known ones (BEB6, COS-I, and NESH5) and five novel ones (named as CHS13-CHS17). Similarly, seven E. bieneusi genotypes were found in yaks, including five known ones (J, BEB4, BEB6, COS-I, and NESH5) and two novel ones (named as CHN13 and CHN14). Most of the E. bieneusi genotypes and all frequent ones identified in the study belonged to group 2. One new subgroup of genotypes was identified within group 1. The distribution of E. bieneusi genotypes was different between Tibetan sheep and yaks, with BEB6 as the dominant one (42.5%) in Tibetan sheep and J as the dominant one (47.5%) in yaks. These data support the occurrence of host adaptation among E. bieneusi genotypes within group 2.

Divergent Cryptosporidium parvum subtype and Enterocytozoon bieneusi genotypes in dromedary camels in Algeria.

Little information is available on the occurrence of the zoonotic protists Cryptosporidium spp. and none on Enterocytozoon bieneusi in camels. This preliminary study was conducted to examine the identity of Cryptosporidium subtypes and E. bieneusi genotypes in dromedary camels in Algeria. A total of 39 fecal specimens were collected from young camels. PCR-sequence analysis of the small subunit rRNA was used to detect and genotype Cryptosporidium spp. Cryptosporidium parvum present was further subtyped by sequence analysis of the 60kDa glycoprotein gene. PCR-sequence analysis of the ribosomal internal transcribed spacer gene was used to detect and genotype E. bieneusi. Altogether, two and eight of the specimens analyzed were positive for C. parvum and E. bieneusi, respectively. The former was identified as a new subtype that is genetically related to the C. hominis If subtype family, whereas the latter was identified as two related genotypes (Macaque1 and a novel genotype) in the newly assigned E. bieneusi genotype group 8. Although they are not known hosts for C. parvum and E. bieneusi, camels are apparently infected with genetically distinct variants of these pathogens.

Stray cats are more frequently infected with zoonotic protists than pet cats.

Faecal samples were collected from cats kept as pets (n = 120) and stray cats (n = 135) in Central Europe (Czech Republic, Poland and Slovakia) and screened for the presence of Cryptosporidium spp., Giardia intestinalis (Kunstler, 1882), Encephalitozoon spp. and Enterocytozoon bieneusi Desportes, Le Charpentier, Galian, Bernard, Cochand-Priollet, Lavergne, Ravisse et Modigliani, 1985 by PCR analysis of the small-subunit of rRNA (Cryptosporidium spp. and G. intestinalis) and ITS (microsporidia) genes. Sequence analysis of targeted genes revealed the presence of C. felis Iseki, 1979, G. intestinalis assemblage F, E. cuniculi Levaditi, Nicolau et Schoen, 1923 genotype II, and E. bieneusi genotype D. There was no correlation between the occurrence of detected parasites and sex, presence of diarrhoea or drug treatment (drug containing pyrantel and praziquantel). Compared to pet cats (7%), stray cats (30%) were statistically more frequently infected with protist parasites and overall may present a greater risk to human health.

Zoonotic and host-adapted genotypes of Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi in dairy cattle in Hebei and Tianjin, China

A total of 1040 fecal samples, collected from 12 dairy cattle farms in Hebei and Tianjin, near the Bohai area of China, were screened for Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi by polymerase chain reaction. The overall prevalence for Cryptosporidium, G. duodenalis and E. bieneusi was 1.0% (n=10), 4.7% (n=49) and 19.4% (n=202), respectively. Ten Cryptosporidium-positive samples were identified as C. parvum by DNA sequence analysis of the small subunit rRNA (SSU rRNA) gene. DNA sequencing of the 60-kDa glycoprotein gene revealed that the C. parvum samples were all subtype IIdA19G1. Forty-nine G. duodenalis-positive samples belonged to assemblage E (n=47) and assemblage E mixed with A (n=2), based on the sequenced SSU rRNA, triosephosphate isomerase, and glutamate dehydrogenase genes. Sequence analysis of the internal transcribed spacer (ITS) gene identified six known E. bieneusi genotypes, I (n=87), J (n=83), BEB4 (n=18), BEB6 (n=3), N (n=1) and Ebpc (n=1), along with three new genotypes, CHC6 (n=1), CHC7 (n=1) and CHC8 (n=7). Phylogenetic analysis showed that Ebpc belonged to zoonotic group 1, whereas the other E. bieneusi genotypes clustered within group 2. More studies are needed to better understand the species distributions and public health significance of these pathogens in the study areas.

Infection by and genotype characteristics of Enterocytozoon bieneusi in HIV/AIDS patients from Guangxi Zhuang autonomous region, China

BackgroundEnterocytozoon bieneusi has been increasingly reported to infect humans and various mammals. Microsporidia cause diarrhea in HIV-infected patients worldwide. PCR amplification and sequencing based on the internal transcribed spacer region have been used to describe the genotypes of E. bieneusi and transmission of microsporidiosis.MethodsIn this study, we examined E. bieneusi infection and genotypes in HIV-positive patients in Guangxi, China. Stool specimens were collected from 285 HIV-positive patients and 303 HIV-negative individuals. E. bieneusi genotypes were characterized using nested PCR and sequencing.ResultsThirty-three (11.58%) HIV-positive patients were infected with microsporidia, and no infection was found in the 303 healthy controls. Three new genotypes were identified and named as GX25, GX456, and GX458; four known genotypes, PigEBITS7, Type IV/K, D, and Ebpc, were also identified. Our data showed that the positive rate for microsporidia was significantly higher in the rural patients than in the other occupation groups. In addition, the positive rate for microsporidia was significantly higher in the patients who drink unboiled water than in those with other drinking water sources.ConclusionsOur results will provide baseline data for preventing and controlling E. bieneusi infection in HIV/AIDS patients. Further studies are required to clarify the epidemiology and potential sources of microsporidia. Our study showed that microsporidium infection occurs in the HIV/AIDS patients in Guangxi, China.

Genotyping of Enterocytozoon bieneusi and Subtyping of Blastocystis in Cancer Patients: Relationship to Diarrhea and Assessment of Zoonotic Transmission

Enterocytozoon bieneusi (E. bieneusi) and Blastocystis are common pathogens responsible for diarrhea in humans, especially in immunocompromised individuals. The number of cancer patients has been increasing and diarrhea is a common clinical symptom in the treatment of cancers. To understand the prevalences and genotypes/subtypes of E. bieneusi and Blastocystis in cancer patients in China, to track the infection sources, and to explore the relationships between E. bieneusi and Blastocystis infections and diarrhea, 381 fecal specimens were collected from cancer patients. Each of them was analyzed for the presence of E. bieneusi and Blastocystis by PCR amplifying and sequencing the ITS region of the rRNA gene and the barcode region of the SSU rRNA gene, respectively. 1.3 and 7.1% of cancer patients were positive for E. bieneusi and Blastocystis, respectively. No statistical differences were observed in the infection rates between the groups by age, gender, and residence. E. bieneusi and Blastocystis were both significantly more common in cancer patients with diarrhea, and significant relationship of Blastocystis to diarrhea was found in chemotherapy group. Two E. bieneusi genotypes (D and a novel one named as HLJ-CP1) and two Blastocystis subtypes (ST1 and ST3) were identified with three novel ST1 sequences. This is the first report of occurrence and molecular characterizations of E. bieneusi and Blastocystis in cancer patients in China. E. bieneusi genotype D and Blastocystis ST1 and ST3 have been identified in humans and animals while one novel E. bieneusi genotype falling into zoonotic group 1, implying a potential of zoonotic transmission.

Molecular characterization of Enterocytozoon bieneusi isolates in laboratory macaques in north China: zoonotic concerns.

The significance of wild and zoo nonhuman primates (NHPs) as potential sources of human Enterocytozoon bieneusi infections has been increasingly appreciated, while the role of laboratory NHPs in zoonotic transmission of microsporidiosis remains elusive. In this study, the infection rate, genetic characteristic, and zoonotic potential of E. bieneusi were investigated for 205 laboratory macaques in Beijing, north China. The parasite was identified in 37 (18.0%) animals by nested PCR and sequencing of the ribosomal internal transcribed spacer (ITS), with an infection rate of 25.6% in Macaca fascicularis (34/133) and 4.2% in Macaca mulatta (3/72). The differences in infection rate between the two species of macaques and between young macaques aged ≤5 years (29.6%, 32/108) and adults aged >5 years (5.2%, 5/97) were significant (p<0.01). Analysis of the ITS sequence polymorphisms recognized eight known genotypes (CC4, CM1, CM2, D, Peru8, Peru11, Type IV, and WL21) and two new genotypes (named as CMB1 and CMB2), with well-known human-pathogenic genotypes (D, Peru8, Peru11, and Type IV) most frequently detected. The rest genotypes (CC4, CM1, CM2, WL21, CMB1, and CMB2) were clustered into zoonotic group 1 in phylogenetic analysis. The high diversity and widespread presence of the human-pathogenic or group 1 E. bieneusi genotypes in laboratory NHPs, notably M. fascicularis and the young animals, suggest potential of zoonotic transmission. These findings imply that laboratory rhesus macaques could be significant reservoirs for human microsporidiosis.