Molecular prevalence and genotypes of Enterocytozoon bieneusi in cancer patients under chemotherapy in Aegean region of Türkiye.

BACKGROUND: Enterocytozoon bieneusi is the most common species found in humans. Although E. bieneusi has been investigated in humans, genotype profile of E. bieneusi is not known in Türkiye. METHODS: In this study, we screened E. bieneusi in patients (n = 94) with different types of malignant solid tumors by Real Time PCR and then sequenced E. bieneusi positive samples. All cancer patients were undergoing chemotherapy and had diarrhea. Moreover, as control groups, we also screened E. bieneusi in patients with diarrhea (n = 50) and without diarrhea (n = 50). RESULTS: Among all patients analyzed, 33 (17%) were found to be E. bieneusi-positive. As the patients were categorized, the molecular prevalence of E. bieneusi increased to 25.5% among cancer patients with diarrhea. However, the molecular prevalence of E. bieneusi was found to be lower in patients with presenting only diarrhea (8%) and patients without diarrhea (10%). The high molecular prevalence value detected among cancer patients with diarrhea was also statistically significant compared to other patient groups (P = 0.00112 and P = 0.0269). Among the 33 Real Time PCR positive samples, 10 of them were amplified by nested PCR and among these 10 samples, 6 of them were successfully genotyped. The phylogenetic tree showed the presence of D and Type IV which were also identified in stray cats living in Izmir in our previous study. CONCLUSIONS: High molecular prevalence value indicates the importance of screening stool samples of cancer patients with diarrhea for E. bieneusi and genotyping results indicate that D and Type IV are circulating between humans and cats.

Enterocytozoon bieneusi Infection after Hematopoietic Stem Cell Transplant in Child, Argentina.

We report a case of Enterocytozoon bieneusi infection in a pediatric hematopoietic stem cell transplant recipient in Argentina. Spores were visualized in feces using Calcofluor White and modified trichrome stainings. PCR and sequencing identified E. bieneusi genotype D in fecal samples and liver samples, confirming extraintestinal dissemination of the parasite.

Widespread distribution of human-infective Enterocytozoon bieneusi genotypes in small rodents in northeast China and phylogeny and zoonotic implications revisited.

Enterocytozoon bieneusi features high genetic diversity among host species and environmental sources and over 500 genotypes in 11 phylogenetic groups have been defined. Here we investigated 291 small rodents in Heilongjiang province, northeast China, for the presence of E. bieneusi by PCR of the ribosomal internal transcribed spacer (ITS). Nine of 60 (15.0 %) gray squirrels from a park in Harbin, 120 of 201 (59.7 %) guinea pigs from a pet shop in Harbin, and two of 30 (6.7 %) peridomestic rats from a pasture in Qiqihar were positive for the parasite. Six known genotypes (EbpB, SCC-1, SCC-2, D, S7 and HLJ-CP1) and two novel genotypes (NESQ1 and NEGP1) were identified by sequence analysis of the ITS, with EbpB, SCC-1, SCC-2 and NESQ1 found in squirrels, D, S7 and NEGP1 in guinea pigs, and EbpB and HLJ-CP1 in rats. Widespread distribution of human-infective Group 10 genotype S7 and Group 1 genotype D in guinea pigs raised our concerns about the importance of pet animals as zoonotic reservoirs of microsporidiosis. Co-occurrence of Group 1 genotypes D and HLJ-CP1 in cancer patients and rodents in Heilongjiang indicated a possibility of zoonotic transmission. The host range of Group 1 genotype EbpB previously considered pig-adapted was extended. A potential variant of genotype S7, namely NESQ1, went into the existing Group 10 in phylogenetic analysis. The other new genotype, NEGP1, was clustered in an undefined clade we proposed as Group 15. With the emerging epidemiologic evidence, the host specificity of existing E. bieneusi genotypes is now being challenged.

Microscopic and Molecular Investigation of Intestinal Microsporidia in HIV + /AIDS and Cancer Patients Undergoing Chemotherapy in Mazandaran Province, North of Iran.

BACKGROUND: The aim of this study was to identify Enterocytozoon bieneusi and Encephalitozoon spp. in fecal samples of HIV + /AIDS and cancer patients undergoing chemotherapy, and comparing the results to healthy individuals in Mazandaran province, north of Iran. METHODS: Stool samples were collected from 50 HIV + /AIDS patients, 50 cancer patients, and 50 healthy samples referred to medical centers in north of Iran. Stool samples were kept in 2.5% potassium dichromate at 4 °C, and stained by modified trichrome for light microscopy examination. The multiplex/nested-PCR targeted the small subunit ribosomal RNA (SSU rRNA) gene. To characterize genotypes, the nested PCR products sequenced by Bioneer Company and was subjected to phylogenetic analyses. RESULTS: Ten of 50 samples (20%) of HIV + /AIDS patients, 5 of 50 samples (10%) of cancer patients, and 1 of healthy individuals (2%) were microscopically positive. From 50 HIV + / AIDS patients, E. bieneusi and Encephalitozoon spp. were detected in 10 (20%) and 6 (12%) cases, respectively. Furthermore, among cancer patients, 7 (14%) and 2 (4%) cases were E. bieneusi and Encephalitozoon spp., respectively. Out of 50 samples of healthy individuals, only 3 (6%) cases of E. bieneusi were observed. The genotypes D and M were detected among positive samples of E. bieneusi. CONCLUSIONS: E. bieneusi and then Encephalitozoon spp. are common intestinal microsporidia in HIV + /AIDS patients and cancer patients undergoing chemotherapy in Mazandaran province. E. bieneusi genotype D seems to be the predominant genotype in Mazandaran province. Due to the considerable prevalence of intestinal microsporidia, physicians are advised to pay more attention to this opportunistic infection in high-risk groups.

Detection of a novel microsporidium with intranuclear localization in farmed Penaeus vannamei from Latin America.

Microsporidia are emerging intracellular parasites of most known animal phyla in all ecological niches. In shrimp aquaculture, the microsporidium Enterocytozoon hepatopenaei (EHP) is a major cause of concern inflicting tremendous losses to shrimp producers in southeast Asia. During a histopathological examination of Penaeus vannamei samples originating in a country from Latin America presenting slow growth, we observed abnormal nuclei in the epithelial cells of the hepatopancreas. A PCR screening of the samples using DNA isolated from paraffin embedded tissues for the SSU rRNA gene of EHP provided a 149 bp amplicon. In situ hybridization using the SSU rRNA gene probe provided a positive signal in the nuclei instead of the cytoplasm. Sequence analysis of the SSU rRNA gene product revealed a 91.3 %, 89.2 % and 85.4 % sequence identity to Enterocytozoon bieneusi, E. hepatopenaei and Enterospora canceri respectively. Furthermore, phylogenetic analysis revealed the newly discovered microsporidium clustered with E. bieneusi. Considering the intranuclear location of the novel microsporidium and the differences in the sequence of the SSU rRNA, we tentatively consider this parasite a new member of the genus Enterospora sp. The pathogenicity and distribution of the shrimp Enterospora sp. are currently unknown. Our future efforts are focused on the characterization and development of diagnostic tools for this parasite to understand if it acts as an emergent pathogen that might require surveillance to prevent its spread.

Enterocytozoon bieneusi and Encephalitozoon intestinalis (microsporidia) in HIV-positive patients in central Spain.

Microsporidia are fungi-related eukaryotic intracellular parasites that opportunistically infect immunocompromised individuals such as those infected by the human immunodeficiency virus (HIV). Among them, Enterocytozoon bieneusi and Encephalitozoon spp. are the most clinically relevant species. We investigated the occurrence and genetic diversity of microsporidial and protist infections in mostly immunocompetent HIV-positive patients in Madrid, Spain. A structured questionnaire was used to retrieve data on factors potentially associated with an increased risk of infection, including sexual attitudes and sex-risk behaviour. Faecal samples (n = 96) from 81 HIV-positive patients were collected and analysed by molecular (PCR and Sanger sequencing) methods. Two microsporidial pathogens were detected: Ent. bieneusi (2.5%, 95% CI: 0.3-8.6) and Enc.intestinalis (4.9%, 95% CI: 1.4-12.2). The two Ent. bieneusi isolates were identified as zoonotic genotype A. Among protists, Entamoeba dispar was the species most prevalently found (33.3%, 95% CI: 23.2-44.7), followed by Blastocystis spp. (19.8%, 95% CI: 11.7-30.1), Giardia duodenalis (13.6%, 95% CI: 7.0-23.0), and Cryptosporidium spp. and Entamoeba histolytica (2.5%, 95% CI: 0.3-8.6 each). Cyclospora cayetanensis and Cystoisospora belli were not detected. Subtypes ST1 (70.6%, 12/17) and ST3 (29.4%, 5/17) were identified within Blastocystis sp., sub-assemblages AII and BIII (50%, 1/2 each) within G. duodenalis, and Cry. parvum and canine-adapted Cry. canis (50%, 1/2 each) within Cryptosporidium spp. Microsporidial and protist parasites were frequent in well-controlled, mostly immunocompetent HIV-positive patients and should be included in diagnostic algorithms when diarrhoea is present.

Opportunistic microsporidial and protist intestinal infections were relatively common in well-controlled HIV-positive patients in Madrid, Spain. These agents should be suspected and appropriately diagnosed in HIV-positive patients presenting with diarrhoea regardless of their immunological status.

Molecular identification and genetic characteristics of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in human immunodeficiency virus/acquired immunodeficiency syndrome patients in Shanghai, China.

BACKGROUND: Opportunistic infections are a ubiquitous complication in human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) patients. Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are common opportunistic intestinal pathogens in humans. In China, despite the number of HIV/AIDS patients being extremely large, only a few studies have investigated opportunistic infections caused by intestinal pathogens in this patient population. The aims of this study were to elucidate the occurrence and genetic characteristics of Cryptosporidium spp., G. duodenalis, and E. bieneusi in HIV/AIDS patients. METHODS: We collected fecal specimens from 155 HIV/AIDS patients (one from each patient). All of the specimens were examined for the presence of the pathogens by genotyping using polymerase chain reaction and sequencing of the small subunit ribosomal RNA gene for Cryptosporidium spp.; the triosephosphate isomerase, ß-giardin and glutamate dehydrogenase genes for G. duodenalis; and the internal transcribed spacer region of the rRNA gene for E. bieneusi. The Cryptosporidium-positive specimens were further subtyped by polymerase chain reacion and sequencing of the 60-kDa glycoprotein gene. RESULTS: Six (3.9%), three (1.9%), and eight (5.2%) HIV/AIDS patients were positive for Cryptosporidium spp., G. duodenalis, and E. bieneusi, respectively. No statistical differences were observed in occurrence rate between the groups by gender, clinical symptom (diarrhea), and CD4+ cell count. Four Cryptosporidium species were identified: Cryptosporidium hominis (n = 2), Cryptosporidium parvum (n = 1), Cryptosporidium meleagridis (n = 1), and Cryptosporidium andersoni (n = 2). Furthermore, two C. hominis subtypes (IeA12G3T3 and IaA28R4) were detected. Three G. duodenalis-positive specimens were successfully amplified and sequenced at the triosephosphate isomerase and ß-giardin loci, which led to the identification of assemblages C and B, respectively. Seven genotypes (D, Type IV, EbpC, Peru11, EbpD, A, and I) were identified in E. bieneusi-positive specimens. CONCLUSIONS: Our findings should increase awareness of AIDS-related opportunistic intestinal pathogens, and indicate the need for routine examination in clinical practice for the detection of Cryptosporidium spp., G. duodenalis, and E. bieneusi. Homology analyses of the three intestinal pathogens at the nucleotide and/or amino acid levels indicated their zoonotic potential.

An isothermal enzymatic recombinase amplification (ERA) assay for rapid and accurate detection of Enterocytozoon hepatopenaei infection in shrimp.

Enterocytozoon hepatopenaei (EHP) is a kind of microsporidian parasite belonging to fungi, and poses a serious threat to prawn farmers. Due to the lack of effective treatments for EHP, the establishment of a rapid and sensitive detection method would be beneficial to the control and prevention of this prawn parasitic disease. In this study, an isothermal enzymatic recombinase amplification (EHP-ERA) assay that could diagnose EHP within 20 min at 42 °C was developed and evaluated. The determined final concentrations of primers and probe in the reaction system were 400 nM and 120 nM, respectively. EHP-ERA was carried out within 13 min (24.31 ± 0.37 Ct) with a detection limit of 10 copies/µL. The results of specificity test showed that EHP-ERA had no cross-reactivity with white spot syndrome virus (WSSV), Vibrio parahaemolyticus strain causing acute hepatopancreatic necrosis disease (VpAHPND), and infectious hypodermal and hematopoietic necrosis virus (IHHNV) and specific pathogen free (SPF) shrimp. Using 32 clinical samples, the practical diagnostic results of EHP-ERA was consistent with nested PCR and real-time PCR (qPCR) under the premise of less time-consuming and simpler operation. In summary, we established a simple, rapid, and effective ERA assay for the detection of EHP, which had great potential to be widely used in both lab and practical usage.

Concurrent infection of a novel genotype of hepatopancreatic parvovirus and Enterocytozoon hepatopenaei in Penaeus vannamei in Taiwan.

Hepatopancreatic parvovirus (HPV) and Enterocytozoon hepatopenaei (EHP) are emerging and reemerging pathogens in shrimps. In the present study, a novel genotype of HPV concurrently infected with EHP in Penaeus vannamei in Taiwan leading to severe atrophy and damage of hepatopancreas were confirmed by histopathology, in situ hybridization, and PCR. The novel genotype of HPV exhibited 66%-69.5% sequence identities with all known HPVs and carried unique amino acid deletions and insertions in the VP gene. According to phylogenetic analysis, the Taiwan HPV isolates were classified as the genotype IV. The present study not only provided the histopathological and molecular proof of HPV and EHP co-infection in Taiwan, but also revealed the importance of investigating the geographical expansion of novel HPV genotypes.

Establishment and application of a TaqMan probe-based qPCR for the detection of Enterocytozoon hepatopenaei in shrimp Litopenaeus vannamei.

Enterocytozoon hepatopenaei (EHP) is a common parasite that invades the epithelial cytoplasm in the hepatopancreas of shrimp Litopenaeus vannamei and results in slow growth of the host shrimps to cause significant economic loss in shrimp aquaculture. In this study, a TaqMan probe-based qPCR for quantitative detection of EHP was established. A pair of specific primers and a TaqMan probe were designed based on the sequence of cysteine desulfurase gene (NFS1) of EHP. The standard curve between cycle threshold (Ct) and the logarithmic starting quantity (SQ) of the template was determined as Ct = - 3.231 lg (SQ) + 40.638, with a correlation coefficient (R2) of 0.998 and an amplification efficiency of 103.9%. The lower limit of quantification was 1.67 × 101 copies/µL for this TaqMan probe-based qPCR and 1.67 × 103 copies/µL for the conventional PCR. The TaqMan probe-based qPCR established in the research was 100 times more sensitive than the conventional PCR method. In addition, the results of clinical sample detection indicated that the present technique was efficient in detecting EHP in the hepatopancreas, feces, water, and pond bottom mud samples. Therefore, the established TaqMan probe-based qPCR is a suitable technique for detecting EHP in both shrimp and aquatic environment samples.

The successful treatment of Enterocytozoon bieneusi Microsporidiosis with nitazoxanide in a patient with B-ALL: A Case Report.

Introduction: Enterocytozoon bieneusi (E. bieneusi) Microsporidia can cause opportunistic infections in immunocompromised patients and is also an emerging disease in these individuals. Its clinical manifestations are chronic diarrhea and severe wasting syndrome, these can be extremely debilitating and carry a significant risk of death for immunocompromised patients. Often, microsporidia cannot be confirmed immediately by routine examination and culture. Effective and available treatment options are limited for infections caused by E. bieneusi in humans. Such cases are very rare in Chinese Mainland. Case presentation: A 47-year-old male had recurrent, profuse watery diarrhea and abdominal discomfort for more than 7 months, with a fever for 5 days. Two years earlier, he received treatment with a modified BFM-90 protocol for acute B cell lymphoblastic leukemia and is currently in the final stages of maintenance therapy with oral methotrexate and mercaptopurine. The leukemia was assessed as still in remission two months ago. PET/CT showed massive peritoneal fluid accumulation and a high uptake area in the diffused peritoneum (SUVmax 12.57), suggesting tumor invasion or microbial infections. However, broad-spectrum antibacterial therapies were ineffective. Metagenomic sequencing of plasma and peritoneal fluid showed no suggestion of the existence of a tumor but instead showed a high sequence number of DNA and RNA of the Microsporidia. His albendazole treatment failed and subsequent treatment with nitazoxanide successfully resolved the infection. Conclusion: This case shows that we should consider the possibility of atypical pathogen infection in patients with hematologic malignancy who repeatedly develop unexplained diarrhea with wasting. mNGS can help rule out malignant neoplasms and diagnose infections. Our results suggest that nitazoxanide effectively treats E. bieneusi microsporidia infections.

Identification of two potential aetiological agents of chronic diarrhoea in an immunocompromised patient in Cuba using conventional and molecular diagnostic techniques.

The aetiology of diarrhoea in a patient in Cuba with HIV was investigated. Although molecular diagnostics are still not used in many under-resourced settings, here traditional methods were supported by use of PCR. This approach enabled detection of a dual infection (Cystoisospora belli and Enterocytozoon bieneusi), the latter of which was not identified by microscopy with Didier's trichromic staining.

Development of a PCR assay for the effective detection of Enterocytozoon hepatopenaei (EHP) and investigation of EHP prevalence in Shandong Province, China.

Enterocytozoon hepatopenaei (EHP), a recently reported pathogen in the penaeid shrimp, is spreading widely and seriously threatening Penaeus (Litopenaeus) vannamei aquaculture. This study aimed to develop a new and more sensitive polymerase chain reaction (PCR) method for the effective detection of EHP. An EHP PCR assay with a pair of primers specifically amplifying a 358 bp EHP DNA fragment was developed, which was demonstrated to be capable of detecting as low as 2 × 101 copies of EHP and is specific for EHP without cross reaction with DNA samples prepared from five common shrimp pathogens, including white spot syndrome virus (WSSV), infectious hypodermal and haematopoietic virus (IHHNV), hepatopancreatic parvovirus (HPV), infectious myonecrosis virus (IMNV), and yellow head virus (YHV). This new assay is more specific and more sensitive than the previously published EHP PCR methods. With the PCR assay developed in this study, we investigated the prevalence of EHP in four areas of Shandong, China by testing a total of 639 shrimp samples collected from Yantai, Binzhou, Dongying, and Weifang. The results showed that the EHP positive rate reached 51.2%, indicating that EHP is prevalent in shrimp culture in China.

Microscopio and PCR-based detection of microsporidia spores in human stool samples

Abstract Microsporidia are obligate intracellular fungi with a remarkable ability to infect a wide range of invertebrate and vertebrate hosts. Namely, Enterocytozoon bieneusi is the most frequently microsporidia reported worldwide, and mainly associated with chronic diarrea and wasting syndrome in AIDS patients. Microscopy and PCR-based detection techniques are effective for diagnosis and identification of species and genotypes; however, these methods should be standardized in each laboratory. In this study, we performed microscopy and nested PCR techniques with PCR product sequencing to detect E. bieneusi in human stool samples. These techniques, if applied together, might prove useful for diagnosis and future epidemiological studies of intestinal microsporidiosis in Argentina.

Resumen Los microsporidios son hongos intracelulares obligados con una notable capacidad para infectar una amplia gama de hospedadores invertebrados y vertebrados. Enterocytozoon bieneusi es el microsporidio más frecuentemente reportado en todo el mundo, principalmente tricrómicaasociado con diarrea crónica y síndrome debilitante en pacientes con sida. Las técnicas dedetección basadas en microscopía y PCR son útiles para el diagnóstico y la identificación deespecies y genotipos, pero estos métodos deben estar estandarizados en cada laboratorio.En este estudio evaluamos técnicas de microscopía y PCR anidada, con secuenciación de losproductos, para detectar E. bieneusi en muestras de heces humanas. Estas técnicas, usadas con-juntamente, podrían ser útiles para su aplicación en el diagnóstico de microsporidiosis intestinaly para realizar estudios epidemiológicos de esta afección en Argentina.

Prevalence and Genetic Characterization of Cryptosporidium and Microsporidia Infecting Hematological Malignancy Patients.

PURPOSE: To evaluate the prevalence of Cryptosporidium and Microsporidia, associated risk factors and species identification in patients with haematological malignancies (HM). METHODS: A total of 148 consecutive patients with HM and 101 healthy subjects were evaluated for Cryptosporidium and Microsporidia using modified Kinyoun and modified Trichrome staining. Clinical, demographic and laboratory parameters were studied. The species of Cryptosporidium and Microsporidia were studied using PCR-RFLP. RESULTS: Of 148 HM patients initially screened, 47 were excluded from the final analysis due to inadequate clinical records. Patients with HM [n = 101, 63 (62.4%) male] more often had Cryptosporidium than healthy subjects [n = 101, 65 (74.4%) male] [3/101 (3%) vs. 0/101 (0%), p = 0.02]. Two of 101 (2%) HM patients and none of the healthy subjects had Microsporidia (p = 0.155). Diarrhea was more prevalent in HM patients with Cryptosporidium than those without [3, 100% vs. 39/96, 40.62%; p = 0.04). Both patients infected with Microsporidia presented with persistent diarrhea and fever. Cryptosporidium hominis was identified in all the three HM patients. Enterocytozoon bieneusi was identified in one HM patient infected with Microsporidia, which was classified as genotype Ind2. CONCLUSION: Cryptosporidium and Microsporidia may infect HM patients leading to overwhelming diarrhea. The commonest species of Cryptosporidium and Microsporidia found to infect HM patients are C. hominis and E. bieneusi.

Detection of human intestinal protozoan parasites in vegetables and fruits: a review.

Diarrheal diseases caused by intestinal protozoan parasites are a major food-borne public health problem across the world. Vegetables and fruits provide important nutrients and minerals, but are also common sources of some food-borne human pathogenic microorganisms. The contamination of raw vegetables and fruits with human pathogenic parasites are now a global public health threat, despite the health benefits of these foods in non-pharmacological prophylaxes against diseases. A large number of reports have documented the contamination of vegetables or fruits with human pathogenic microorganisms. In this paper, we reviewed the contamination and detection methods of human pathogenic intestinal protozoans that are frequently recovered from raw vegetables and fruits. The protozoan parasites include Cryptosporidium spp., Giardia duodenalis, Cyclospora cayetanensis, Entamoeba spp., Toxoplasma gondii, Balantioides coli, Blastocystis sp., Cystoisospora belli and Enterocytozoon bieneusi. The risk factors involved in the contamination of vegetables and fruits with parasites are also assessed.

Molecular epidemiology of Enterocytozoon bieneusi and Encephalitozoon sp., among immunocompromised and immunocompetent subjects in Iran.

Intestinal microsporidiosis is known as an opportunistic infection in immunocompromised patients. The current study aimed to investigate intestinal microsporidia infection in human subjects with/without immunodeficiency. Totally, 600 stool samples were collected from immunocompromised (254) and immunocompetent (346) subjects. DNA extraction was performed and the SSU rRNA and the ITS genes were amplified to detect and characterize microsporidia and the relevant genotypes. Phylogenetic trees were drawn using MEGA7 software to illustrate the correlation between isolates. From 600 enrolled subjects, 283 and 317 were male and female, respectively. The average age ± SD of all tested subjects was 28.85 ± 26.92. The results of PCR demonstrated the presence of E. bieneusi and Encephalitozoon sp., among 10/600 (1.67%) and 26/600 (4.33%) of samples, respectively. Accordingly, E. bieneusi was seen among 4/346 (1.15%), 1/53 (1.88%), 3/124 (2.42%), and 2/63 (3.17%), and Encephalitozoon sp., was detected from 17/346 (4.91%), 3/53 (5.36%), 4/124 (3.22%) and 2/63 (3.17%) of healthy subjects, RA patients, cancer patients, and transplantation recipients, respectively. Statistical significant correlation was not seen between the presence of microsporidia and age, gender, stool appearance, and geographical region. Molecular analysis showed that all E. bieneusi were the genotype D. Phylogenetic tree demonstrated no classification according to the presence/absence of immunodeficiency, geographical locations and presence of diarrhea. The high prevalence of Encephalitozoon sp., in comparison to E. bieneusi in this study suggested the importance of this genus alongside with E. bieneusi in Iran. In addition, predominance of the genotype D highlighted the wide distribution of this genotype in Iran.

Comparison of infectious agents detected from hatchery and wild juvenile Coho salmon in British Columbia, 2008-2018.

Infectious diseases are potential contributors to decline in Coho salmon (Oncorhynchus kisutch) populations. Although pathogens are theoretically considered to pose higher risk in high-density rearing environments like hatcheries, there is no direct evidence that hatchery-origin Coho salmon increase the transmission of infectious agents to sympatric wild populations. This study was undertaken to compare prevalence, burden, and diversity of infectious agents between hatchery-reared and wild juvenile Coho salmon in British Columbia (BC), Canada. In total, 2,655 juvenile Coho salmon were collected between 2008 and 2018 from four regions of freshwater and saltwater in BC. High-throughput microfluidics qPCR was employed for simultaneous detection of 36 infectious agents from mixed-tissue samples (gill, brain, heart, liver, and kidney). Thirty-one agents were detected at least once, including ten with prevalence >5%. Candidatus Brachiomonas cysticola, Paraneuclospora theridion, and Parvicapsula pseudobranchiocola were the most prevalent agents. Diversity and burden of infectious agents were substantially higher in marine environment than in freshwater. In Mainland BC, infectious burden and diversity were significantly lower in hatchery smolts than in wild counterparts, whereas in other regions, there were no significant differences. Observed differences in freshwater were predominantly driven by three parasites, Loma salmonae, Myxobolus arcticus, and Parvicapsula kabatai. In saltwater, there were no consistent differences in agent prevalence between hatchery and wild fish shared among the west and east coasts of Vancouver Island. Although some agents showed differential infectious patterns between regions, annual variations likely contributed to this signal. Our findings do not support the hypothesis that hatchery smolts carry higher burdens of infectious agents than conspecific wild fish, reducing the potential risk of transfer to wild smolts at this life stage. Moreover, we provide a baseline of infectious agents in juvenile Coho salmon that will be used in future research and modeling potential correlations between infectious profiles and marine survival.

Genotyping Approach for Potential Common Source of Enterocytozoon bieneusi Infection in Hematology Unit.

Microsporidiosis is a fungal infection that generally causes digestive disorders, especially in immunocompromised hosts. Over a 4-day period in January 2018, 3 patients with hematologic malignancies who were admitted to the hematology unit of a hospital in France received diagnoses of Enterocytozoon bieneusi microsporidiosis. This unusually high incidence was investigated by sequence analysis at the internal transcribed spacer rDNA locus and then by 3 microsatellites and 1 minisatellite for multilocus genotyping. The 3 isolates had many sequence similarities and belonged to a new genotype closely related to genotype C. In addition, multilocus genotyping showed high genetic distances with all the other strains collected from epidemiologically unrelated persons; none of these strains belonged to the new genotype. These data confirm the epidemiologic link among the 3 patients and support a common source of infection.

Detection of Enterocytozoon hepatopenaei using an invasive but non-lethal sampling method in shrimp (Penaeus vannamei).

The detection of enteric pathogens that cause diseases in shrimp involves the sacrifice of the host to obtain tissue samples for diagnosis. In this study, we describe an invasive but non-lethal sampling methodology using a syringe to collect biopsy samples from the hepatopancreas (HP) of Penaeus vannamei to detect the microsporidian pathogen, Enterocytozoon hepatopenaei (EHP), by qPCR and transmission electron microscopy. EHP was detected in all the infected shrimp by qPCR. The shrimp infected by the microsporidian showed 65% survival at 7 days post-sampling. Transmission electron microscopic examination of the biopsy samples revealed numerous spores of the pathogen. The presence of EHP was further confirmed by histology and in situ hybridization from HP tissue samples. The data shows that a hepatopancreas biopsy could be a viable means of detecting enteric pathogens in shrimp, and the method could be valuable in sampling broodstock and natural populations without the need to sacrifice the animals.