Occurrence and potentially zoonotic genotypes of Enterocytozoon bieneusi in wild rhesus macaques (Macaca mulatta) living in Nanwan Monkey Island, Hainan, China: a public health concern.

BACKGROUND: Enterocytozoon bieneusi, a microsporidian species, is a zoonotic pathogen found in both humans and animals. Here, we determined the prevalence, explored the different genotypes of E. bieneusi in wild rhesus macaques (Macaca mulatta) (Hainan Island of China), and assessed their zoonotic potential. METHODS: We collected 173 fecal specimens from wild rhesus macaques living in Nanwan Monkey Island, Hainan, China. Subsequently, we identified and genotyped E. bieneusi using nested PCR analysis amplification of the internal transcribed spacer region (ITS) of the rRNA gene. Lastly, a neighbor-joining tree was built based on gene sequences from the ITS region of E. bieneusi. RESULTS: Of the 173 specimens from wild rhesus macaques, 26 (15%) were infected with E. bieneusi. We identified six genotypes of E. bieneusi, of which five were known: PigEBITS7 (n = 20), D (n = 2), Type IV (n = 1), Peru6 (n = 1), Henan-III (n = 1), and a novel genotype: HNM-IX (n = 1). From the phylogenetic analysis, the six genotypes identified here were all clustered into zoonotic group 1. CONCLUSION: This study is the first report to detect E. bieneusi infection in wild rhesus macaques from Hainan, China. Human-pathogenic genotypes D, Henan-III, Peru6, PigEbITS7, and Type IV in the wild rhesus macaques support these animals infected with E. bieneusi have a public health significance.

Enterocytozoon bieneusi in Bovine Viral Diarrhea Virus (BVDV) infected and noninfected cattle herds.

Enterocytozoon bieneusi known as a causative agent of opportunistic infections instigating diarrhoea in AIDS patients was identified also in a number of immunocompetent patients and in a wide range of animals, including cattle. In the present study we tested if the Bovine Viral Diarrhea Virus (BVDV), the most common pathogen underlying immunosuppressive Bovine Viral Diarrhoea (BVD), can enhance the occurrence of opportunistic infections with E. bieneusi in cattle. Six dairy farms were investigated using ELISA to detect antibodies against or antigens arising from BVDV in collected sera. A total of 240 individual faecal samples from four age groups were examined for the presence of E. bieneusi by nested PCR. Sequence analysis of six E. bieneusi positive samples revealed the presence of the genotype I of E. bieneusi, previously described in cattle. The hypothesis expecting higher prevalence of E. bieneusi in BVDV positive cattle herds was not confirmed in this study; however this is the first description about E. bieneusi in cattle in the Czech Republic.

The growing prevalence of Nosema ceranae in honey bees in Spain, an emerging problem for the last decade.

Microsporidiosis caused by infection with Nosema apis or Nosema ceranae has become one of the most widespread diseases of honey bees and can cause important economic losses for beekeepers. Honey can be contaminated by spores of both species and it has been reported as a suitable matrix to study the field prevalence of other honey bee sporulated pathogens. Historical honey sample collections from the CAR laboratory (Centro Apícola Regional) were analyzed by PCR to identify the earliest instance of emergence, and to determine whether the presence of Nosema spp. in honey was linked to the spread of these microsporidia in honey bee apiaries. A total of 240 frozen honey samples were analyzed by PCR and the results compared with rates of Nosema spp. infection in worker bee samples from different years and geographical areas. The presence of Nosema spp. in hive-stored honey from naturally infected honey bee colonies (from an experimental apiary) was also monitored, and although collected honey bees resulted in a more suitable sample to study the presence of microsporidian parasites in the colonies, a high probability of finding Nosema spp. in their hive-stored honey was observed. The first honey sample in which N. ceranae was detected dates back to the year 2000. In subsequent years, the number of samples containing N. ceranae tended to increase, as did the detection of Nosema spp. in adult worker bees. The presence of N. ceranae as early as 2000, long before generalized bee depopulation and colony losses in 2004 may be consistent with a long incubation period for nosemosis type C or related with other unknown factors. The current prevalence of nosemosis, primarily due to N. ceranae, has reached epidemic levels in Spain as confirmed by the analysis of worker honey bees and commercial honey.

Negative correlation between Nosema ceranae spore loads and deformed wing virus infection levels in adult honey bee workers.

Interactions between pathogens might contribute to honey bee colony losses. Here we investigated if there is an association between the microsporidian Nosema ceranae and the deformed wing virus (DWV) in different body sections of individual honey bee workers (Apis mellifera ligustica) under exclusion of the vector Varroa destructor. Our data provide correlational evidence for antagonistic interactions between the two pathogens in the midgut of the bees.

Intestinal microsporidiosis in diarrheal patients infected with human immunodeficiency virus-1 in Addis Ababa, Ethiopia.

Intestinal microsporidiosis is most commonly associated with persistent diarrhea in advanced AIDS cases. To determine the prevalence and clinical manifestations of this infection in HIV/AIDS patients, a single fresh stool sample and blood were collected from 243 (214 HIV-positive and 29 HIV-negative) diarrheal patients. The presence of intestinal microsporidiosis in the stool was determined by Uvitex-2B staining and a PCR-based detection method. HIV screening was done by using ELISA, and reactive samples were confirmed by Western blotting. The CD4+ cell count was analyzed using FACScan. Out of 243 diarrheal patients, 39 (16.0%) cases were positive for intestinal microsporidial infection by either of the methods used. Of the 39, only 18 cases positive by microscopy were also positive by PCR. Based on PCR and microscopic analyses the microsporidial parasites were identified as Enterocytozoon bieneusi (30), Ecephalitozoon intestinalis (6), and double infections (3). All microsporidia-positive cases were HIV-positive, and 92.3% had diarrhea for over 4 weeks. The diarrhea was watery in 79.5% of the patients. Weight loss >10% was recorded in 37 (94.9%) cases. The CD4+ cell count was <100 cells/mm(3) in 84.4% of subjects, and 59.4% of the patients had a CD4+ cell count of < or =50 cells/mm(3), with a mean of 22.8 cells/mm(3). This study revealed that intestinal microsporidiosis is a common cause of chronic diarrhea and severe weight loss in advanced AIDS patients in Ethiopia. This condition is attributable mainly to E. bieneusi. Thus, early diagnosis of intestinal microsporidiosis in HIV/AIDS patients would certainly be helpful in the understanding and management of diarrheal illness.

Microsporidiosis in Venezuela: prevalence of intestinal microsporidiosis and its contribution to diarrhea in a group of human immunodeficiency virus-infected patients from Zulia State.

Microsporidia are recognized as a cause of morbidity among patients infected with the human immunodeficiency virus (HIV). Infection rates for intestinal microsporidiosis in HIV-infected patients from Venezuela are unknown. To determine the prevalence and pathogenic role of microsporidia in these patients in northwestern Venezuela, a case control study was conducted in 103 outpatients (mean +/- SD age = 37.3 +/- 5.6 years). Microsporidia were detected using unconcentrated formalin-fixed stools examined by Weber's chromotrope-based staining method. For identification of coccidia, modified Ziehl-Neelsen carbolfuchsin staining of formalin-ether concentrates were used, and for other pathogenic parasites, iron hematoxylin-stained smears and formalin-ether concentrates were examined. Microsporidial infections were detected in 14 (13.6%) of 103 patients and 39 (37.9%) had other parasitic pathogens. No significant difference was noted in the occurrence of the infection in patients with diarrhea (13 of 74, 17.6%) and controls (1 of 29, 3.4%) (P = 0.118). Nevertheless, this result may be due to the small sample size (n = 14) of infected individuals. The proportions of other pathogens in patients with or without diarrhea were not significantly different (P = 0.828). Microsporidiosis is common among the HIV-infected population in northwestern Venezuela. However, its pathogenic role in these patients is uncertain and warrants further investigation.

Interspecies transmission of Enterozytozoon bieneusi supported by observations in laboratory animals and phylogeny.

Enterocytozoon bieneusi is emerging as an important cause of chronic diarrhoea in AIDS patients. Its reservoirs and transmission patterns are unknown. In this study, we have examined E. bieneusi sequences from four Rhesus macaques of different origin, which were kept at one animal facility. The sequences were identical in all animals, which suggested that infection had occurred within the facility. Full sequence agreement of E. bieneusi from macaques was found with an E. bieneusi genotype that occurs frequently in humans. To clarify, the relevance of possible inter-species transmission from man to macaque, a phylogenetic analysis was conducted including all sequences of E. bieneusi deposited in GenBank. The hitherto used system of diverse nomenclatures could be reduced to an outlier group and three main lineages, one of which could be further sub-divided into five subgroups. Based in this phylogeny, an association of parasites and host species could be observed for main lineages 2 and 3, as well as for most of the subgroups of main lineage 1. For confirmation, the phylogeny of main lineage 1 was reconstructed with an alternative method of distance estimation, yielding essentially the same parasite-host associations. Zoonotic potential of E. bieneusi is thus supported on a phylogenetic basis.

Intestinal microsporidiosis in HIV infected patients with chronic diarrhea in Thailand.

Microsporidia have been recognized as emerging opportunistic agents affecting multiple organs. Intestinal microsporidiosis caused by Enterocytozoon bieneusi and Encephalitozoon intestinalis is a common disease which is associated with gastrointestinal symptoms, particularly in AIDS patients. So far, information on the frequency of this enteric disease in Thailand is not available. Therefore, the present study was undertaken to investigate the prevalence of intestinal microsporidiosis in HIV infected persons with chronic diarrhea. From 1995 to 1996, multiple diarrheal stool specimens were received and examined for the presence of the organism using Weber's modified trichrome staining method and transmission electron microscopy for confirmation. Twenty-two of 66 patients (33.3%) were positive for microsporidia which appeared as pink-red spores of 0.8-1.2 x 0.7-0.9 microm with the characteristic transverse or oblique band representing the coiled polar filament. Clinical features of these patients included chronic diarrhea (100%), weight loss (100%), abdominal pain (77%), fever (36%), vomiting (36%) and anorexia (18%). Transmission electron microscopic examination of fecal specimens from the 22 patients with positive staining results revealed E. bieneusi in 18 cases.

Identification of an Enterocytozoon bieneusi-like microsporidian parasite in simian-immunodeficiency-virus-inoculated macaques with hepatobiliary disease.

Enterocytozoon bieneusi is a common opportunistic pathogen of human patients with acquired immune deficiency syndrome (AIDS) causing significant morbidity and mortality. In a retrospective analysis utilizing conventional histochemical techniques, in situ hybridization, polymerase chain reaction, and ultrastructural examination, we identified 18 simian-immunodeficiency-virus-infected macaques (16 Macaca mulatta, 1 M. nemestrina, and 1 M. cyclopis) with Enterocytozoon infection of the hepatobiliary system and small intestine. The organisms were readily identified in the bile ducts and gall bladder by special stains and by in situ hybridization using a probe directed against the small subunit ribosomal RNA of human origin E. bieneusi. Infection of the biliary system was associated with a nonsuppurative and proliferative cholecystitis and choledochitis. Hepatic involvement was characterized by bridging portal fibrosis and nodular hepatocellular regeneration accompanied by marked bile ductular and septal duct hyperplasia. Ultrastructurally, all developmental stages of the organism were found in direct contact with the host cell cytoplasm; spores and sporoblasts contained a double layer of polar tubes. Sequencing of a 607-bp segment of the small subunit ribosomal RNA revealed 97 and 100% identity to two clones of small subunit ribosomal RNA derived from E. bieneusi of human origin. Extensive morphological and genetic similarities between the simian and human enterocytozoons suggest that experimentally infected macaques may serve as a useful model of microsporidial infection in AIDS.