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The Penaeus vannamei aquaculture industry is facing a significant challenge in the form of hepatopancreatic microsporidiosis (HPM) caused by Enterocytozoon hepatopenaei (EHP), resulting in substantial economic losses. However, the extent of knowledge regarding the mechanisms by which shrimp resist EHP is limited. We screened resistant and susceptible shrimp and found that resistant shrimp had lower EHP load and less tissue damage. To gain insight into the molecular mechanisms underlying the EHP resistance of shrimp, a comparison was conducted at the transcriptional level between the resistant and susceptible families. Transcriptomic analysis of shrimp hepatopancreas revealed significant differences between the resistant and susceptible families. Compared to the susceptible family, the immune system of the resistant family was activated. The resistant family showed up-regulation in the expression of cathepsin L, C-type lectin, penaeidin, chitinase genes, and metabolism of xenobiotics by cytochrome P450-related genes. Additionally, the resistant shrimp exhibited a higher capacity for amino acid uptake. The observed differences in the resistant and susceptible family transcriptome may contribute to the shrimp's resistance to EHP.

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Hepatopancreatic microsporidiosis (HPM), caused by the microsporidium Ecytonucleospora hepatopenaei (EHP) leads to retarded growth and enhanced susceptibility to other diseases in shrimp resulting in a major loss for the shrimp industry worldwide. It is little understood how EHP infects its host and hijacks its cellular machinery to replicate and exert clinical manifestations in infected shrimp. Since the initial record of HPM, histopathology and polymerase chain reaction (PCR)-based assays were developed for the detection of EHP to prevent spread of the disease. Availability of an antibody-based detection method would complement these existing diagnostic tools and be useful in studying EHP pathogenesis. We describe here an immunofluorescence assay (IFA) for detecting EHP using monoclonal antibodies (mAbs) that were originally developed against Cryptosporidium parvum, a coccidian parasite that infects calves (Bos taurus), other agriculturally important animals, and humans. Forty-one mAbs were screened and two mAbs, 3E2 and 3A12, were found to detect EHP successfully. The utility of these mAbs in detecting EHP was further assessed by testing 36 experimentally challenged EHP-infected shrimp (Penaeus vannamei). EHP-detection data from infected shrimp were compared by Hematoxylin and Eosin (H&E) histology, real-time PCR, and immunofluorescence. The data show IFA using mAbs 3E2 and 3A12 could successfully detect EHP and that the sensitivity of detection is comparable to H&E histology and quantitative PCR. Availability of mAbs that can detect EHP is expected to be immensely beneficial in HPM diagnosis. Since the pathobiology of C. parvum has been so widely studied, these cross-reactive mAbs may also aid in gaining some insight into EHP pathogenesis and disease.

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AIM: To investigate the quality of life of women with endometriosis before treatment and 3 months after the start of surgical and/or conservative treatment. SAMPLE AND METHODOLOGY: The sample comprised of 38 patients, of whom 26 underwent surgical treatment, 6 had pharmacological treatment, and 6 had both surgical and pharmacological treatment. The Endometriosis Health Profile (EHP-30) questionnaire in the Czech version and the Numeric Rating Scale (NRS) were used to assess quality of life. The questionnaires were completed before treatment and 3 months into the treatment. RESULTS AND DISCUSSION: When comparing quality of life with the EHP-30 questionnaire, 3 months after the start of treatment, significantly better quality of life scores were found in all domains except the domain "Infertility." Statistically significant improvement was observed in the domains of "Control and powerlessness," "Emotional well-being," and "Pain" (P < 0.0001). Pain assessment using NRS showed subjective improvement in pain during menstruation, outside menstruation, during intercourse, micturition, and defecation. Statistically significant improvement was reported in pain during menstruation and outside menstruation (P < 0.0001). CONCLUSION: Treatment of endometriosis improves the quality of life and also leads to a subjective reduction of pain intensity as one of the main symptoms of the disease.

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OBJECTIVE: To evaluate the effect of relugolix combination therapy (relugolix CT; 40 mg relugolix, 1 mg estradiol, and 0.5 mg norethisterone acetate) for up to 2 years in the SPIRIT long-term extension study on functioning and health-related quality of life (QoL), using the Endometriosis Health Profile (EHP)-30 questionnaire, and assess how changes in QoL domains correlated with improvements in dysmenorrhea as well as nonmenstrual pelvic pain (NMPP). DESIGN: Long-term extension study of the SPIRIT phase 3 trials. SETTING: Clinics and University Hospitals. PATIENT(S): Premenopausal women with moderate-to-severe endometriosis pain who previously completed the randomized SPIRIT trials were eligible to enroll in an 80-week long-term extension where all women received relugolix CT. INTERVENTION(S): Relugolix CT (relugolix 40 mg, estradiol 1 mg, and norethindrone acetate 0.5 mg). MAIN OUTCOME MEASURE(S): Least squares (LS) mean changes in the EHP-30 domain and total scores from baseline (pivotal) were analyzed using a mixed-effects model. Results up to 104 weeks are reported by a pivotal trial treatment group with a focus on the relugolix CT group (i.e., relugolix CT or placebo for 24 weeks, or delayed relugolix CT [relugolix 40 mg monotherapy for 12 weeks, followed by relugolix CT for 12 weeks]). In addition, the relationships between changes in dysmenorrhea and NMPP as well as changes in EHP-30 scores were assessed. RESULT(S): In the 277 women treated with relugolix CT, LS mean EHP-30 pain domain scores improved by 57.8% (LS mean change: -32.8; 95% CI: -35.5, -30.1), 66.4% (LS mean change: -37.7; 95% CI: -40.3, -35.0), and 72.2% (LS mean change: -41.3; 95% CI: -43.9, -38.7) at weeks 24, 52, and 104, respectively. The proportions of women with clinically meaningful improvement in the EHP-30 pain domain were 75.9%, 83.6%, and 88.6% at weeks 24, 52, and 104, respectively. Non-pain EHP-30 domain and total scores likewise improved. A positive correlation between changes in dysmenorrhea/NMPP and all EHP-30 domain scores was observed. Results were similar for the delayed relugolix CT and placebo → relugolix CT groups. CONCLUSION(S): Sustained reduction of endometriosis-associated pain with relugolix CT observed up to 104 weeks was accompanied by improvements in functioning and health-related QoL. These findings complement the results of the pivotal SPIRIT trials, which showed that relugolix combination therapy significantly reduced dysmenorrhea, NMPP, and dyspareunia vs. placebo in premenopausal women with endometriosis-associated pain. CLINICAL TRIAL REGISTRATION NUMBER: Registration/clinicaltrials.gov identifier: SPIRIT Extension Study (NCT03654274).

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The microsporidian Enterocytozoon hepatopenaei (EHP) is a major threat to shrimp health worldwide. Severe EHP infections in shrimp cause growth retardation and increase susceptibility to opportunistic infections. EHP produces spores with a chitin wall that enables them to survive prolonged environmental exposure. Previous studies showed that polar tube extrusion is a prerequisite for EHP infection, such that inhibiting extrusion should prevent infection. Using a proteomic approach, polar tube protein 2 of EHP (EhPTP2) was found abundantly in protein extracts obtained from extruded spores. Using an immunofluorescent antibody against EhPTP2 for immunohistochemistry, extruded spores were found in the shrimp hepatopancreas (HP) and intestine, but not in the stomach. We hypothesized that presence of EhPTP2 might be required for successful EHP spore extrusion. To test this hypothesis, we injected EhPTP2-specific double-stranded RNA (dsRNA) and found that it significantly diminished EHP copy numbers in infected shrimp. This indicated reduced amplification of EHP-infected cells in the HP by spores released from previously infected cells. In addition, injection of the dsRNA into EHP-infected shrimp prior to their use in cohabitation with naïve shrimp significantly (p < 0.05) reduced the rate of EHP transmission to naïve shrimp. The results revealed that EhPTP2 plays a crucial role in the life cycle of EHP and that dsRNA targeting EHP mRNA can effectively reach the parasite developing in host cells. This approach is a model for future investigations to identify critical genes for EHP survival and spread as potential targets for preventative and therapeutic measures in shrimp.

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Infectious hypodermal and hematopoietic necrosis virus (IHHNV) is one of the linearly single-stranded DNA viruses. Ecytonucleospora hepatopenaei (EHP) is an intracellular parasitic microsporidian. IHHNV and EHP are pathogens that have been widely prevalent in shrimp farming. Both of them are associated with growth retardation of the penaeid shrimp, which causes serious economic losses to shrimp farming. Shrimp can be co-infected with IHHNV and EHP. In this study, a rapid duplex polymerase chain reaction (PCR) was developed and optimized for the simultaneous detection of EHP and IHHNV. The detection limit of the duplex PCR could reach 1.5 × 102 copies for EHP and IHHNV. A total of 578 Litopenaeus vannamei samples were detected by the established duplex PCR detection method. The results suggested that 398 samples were infected with EHP, 362 samples were infected with IHHNV, and 265 samples were co-infected with EHP and IHHNV. The case-control analysis of the detected shrimp samples showed a certain synergistic effect between EHP and IHHNV.

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The SARS-CoV-2 papain-like protease (PLpro) and main protease (Mpro) are nucleophilic cysteine enzymes that catalyze hydrolysis of the viral polyproteins pp1a/1ab. By contrast with Mpro, PLpro is also a deubiquitinase (DUB) that accepts post-translationally modified human proteins as substrates. Here we report studies on the DUB activity of PLpro using synthetic Nε-lysine-branched oligopeptides as substrates that mimic post-translational protein modifications by ubiquitin (Ub) or Ub-like modifiers (UBLs), such as interferon stimulated gene 15 (ISG15). Mass spectrometry (MS)-based assays confirm the DUB activity of isolated recombinant PLpro. They reveal that the sequence of both the peptide fragment derived from the post-translationally modified protein and that derived from the UBL affects PLpro catalysis; the nature of substrate binding in the S sites appears to be more important for catalytic efficiency than binding in the S' sites. Importantly, the results reflect the reported cellular substrate selectivity of PLpro, i.e. human proteins conjugated to ISG15 are better substrates than those conjugated to Ub or other UBLs. The combined experimental and modelling results imply that PLpro catalysis is affected not only by the identity of the substrate residues binding in the S and S' sites, but also by the substrate fold and the conformational dynamics of the blocking loop 2 of the PLpro:substrate complex. Nε-Lysine-branched oligopeptides thus have potential to help the identification of PLpro substrates. More generally, the results imply that MS-based assays with Nε-lysine-branched oligopeptides have potential to monitor catalysis by human DUBs and hence to inform on their substrate preferences.

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BACKGROUND: Endometriosis often leads to a decrease in Quality of Life (QoL), due to its impact on various aspects of women's lives, such as social life, mental health, sex life, and working capacity. Although previous studies have assessed QoL in women with endometriosis, few studies have explored the impact of different clinical variables on QoL. The aim of this study was to investigate how women with endometriosis perceive their QoL, and to analyze which clinical factors are associated with QoL. METHODS: The Endometriosis Health Profile-30 and the ENDOCARE Questionnaire were distributed to 1000 women diagnosed with endometriosis from 10 different clinics across Sweden. The responses from 476 women were included in univariate and multivariable regression analyses, where the clinical factors were correlated with overall QoL and QoL dimensions. RESULTS: The women participating in this study reported a low QoL. The clinical factors that showed a significant correlation with overall QoL were age at first onset of endometriosis symptoms (ß= -0.64, p < 0.001), having more than 10 visits to general practitioners before referral to a gynecologist (ß = 5.58, p = 0.036), current or previous mental health issues (ß = 7.98, p < 0.001) patient-centeredness (ß= -2.59, p < 0.001) and use of opioids (ß = 7.14, p = 0.002). CONCLUSIONS: This study shows that opioid use and mental health issues were associated with a worse QoL, whereas a higher degree of patient-centeredness was associated with a better QoL. The association between opioid use and a worse QoL might not entirely be caused by the opioid use itself but also by symptom severity and mental health issues. An improved patient-centeredness and more focus on taking care of mental health issues would reasonably result in a better QoL for women with endometriosis.

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OBJECTIVES: To validate and determine the psychometric properties of the Malay version of the endometriosis health profile-30 (EHP-30) by confirmatory factor analysis. METHODS: A cross-sectional study was carried out in the main city of Malaysia at a tertiary teaching hospital between January to April 2021. A total of 218 women diagnosed with endometriosis symptoms were recruited using the universal sampling method to answer the questionnaire. RESULTS: The revised Malay version of the EHP-30 with 28 items demonstrated that the factor loading of the 28 items had an acceptable value range between 0.60-0.90. The model fit was acceptable after the inclusion of 28 items correlated errors of the root mean square of error approximation: 0.072, 90% confidence interval: [0.065-0.080], comparative fit index (0.939), Tucker-Lewis index (0.932), and Chi-square/degrees of Freedom (2.135). The Cronbach's alpha ranged from 0.89-0.97. Concurrent validity for the composite reliability was between 0.88-0.96, while the average variance extracted was between 0.65-0.74. CONCLUSION: This revised Malay version of the EHP-30 is a reliable and valid tool that can be used for the next study.

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Viruses use microRNAs (miRNAs) to impair the host antiviral response and facilitate viral infection by expressing their own miRNAs or co-opting cellular miRNAs. miRNAs inhibit translation initiation of their target mRNAs by recruiting the GIGYF2-4EHP (or EIF4E2) translation repressor complex to the mRNA 5'-cap structure. We recently reported that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-encoded non-structural protein 2 (NSP2) interacts with GIGYF2. This interaction is critical for blocking translation of the Ifnb1 mRNA that encodes the cytokine interferon ß, and thereby impairs the host antiviral response. However, it is not known whether NSP2 also affects miRNA-mediated silencing. Here, we demonstrate the pervasive augmentation of miRNA-mediated translational repression of cellular mRNAs by NSP2. We show that NSP2 interacts with argonaute 2 (AGO2), the core component of the miRNA-induced silencing complex (miRISC), via GIGYF2 and enhances the translational repression mediated by natural miRNA-binding sites in the 3' untranslated region of cellular mRNAs. Our data reveal an additional layer of the complex mechanism by which SARS-CoV-2 and likely other coronaviruses manipulate the host gene expression program by co-opting the host miRNA-mediated silencing machinery.

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Segmental rectum resections for indications other than endometriosis were reported to result in up to 40% sexual dysfunctions. We, therefore, evaluated sexual function after low bowel resection (n = 33) for deep endometriosis in comparison with conservative excision (n = 23). Sexual function was evaluated with the FSFI-19 (Female Sexuality Functioning Index) and EHP 30 (Endometriosis Health Profile). The pain was evaluated with visual analogue scales. Linear excision and bowel resections improved FSFI, EHP 30, and postoperative pain comparably. By univariate analysis, a decreased sexual function was strongly associated with pain both before (p < 0.0001) and after surgery (p = 0.0012), age (p = 0.05), and duration of surgery (p = 0.023). By multivariate analysis (proc logistic), the FSFI after surgery was predicted only by FSFI before or EHP after surgery. No differences were found between low bowel segmental resection and a more conservative excision. In conclusion, improving pain after surgery can explain the improvement in sexual function. A deleterious effect of a bowel resection on sexual function was not observed for endometriosis. Sexual function in women with endometriosis can be evaluated using a simplified questionnaire such as FSFI-6.

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White feces syndrome (WFS) is a commercially important disease in Penaeus vannamei (whiteleg shrimp) farming. The aetiology beyond the white or golden white midgut with mediocre growth performance producing a floating mass of white fecal strings in WFS-affected shrimp farms remains uncharted. To give WFS a perception of pathobiome, healthy P. vannamei shrimps were subjected to an enteric microsporidian Enterocytozoon hepatopenaei (EHP) infection along with Vibrio harveyi and V. alginolyticus in different combinations. Immune responses in haemolymph (total haemocyte count (THC), prophenoloxidase activity (proPO), respiratory burst activity (RBA), superoxide dismutase activity (SOD) and catalase activity (CAT)), plasma biochemical changes (aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP)) and digestive enzymes activity (alpha-amylase (AMY), lipase (LIP) and protease (PRO)) were assessed in the challenged shrimps at 5, 10 and 15 days post-infection (dpi). The microbial interactions between the EHP and Vibrio spp. have led to the formation of WFS in the challenged shrimps. The histological sections of the hepatopancreas revealed the presence of EHP along with colonized bacterial masses, leading to the formation of aggregated transformed microvilli (ATM) structures and increased sloughing of lipid vacuoles into the tubule lumen. A significantly decreased THC and increased proPO levels, dysregulated antioxidant system, prominent hepatic damage, reduced energy metabolism and higher lipid production were the key records supporting that EHP-associated WFS in P. vannamei is due to the pathobiome.

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Infection by the microsporidian parasite Enterocytozoon hepatopenaei (EHP) has become a significant problem in the shrimp cultivation industry in Asian countries like Thailand, China, India, Vietnam, Indonesia, and Malaysia. The outbreak of this microsporidian parasite is predominantly related to the existence of macrofauna-carriers of EHP. However, information about potential macrofauna-carriers of EHP in rearing ponds is still limited. In this study, the screening of EHP in potential macrofauna-carriers was conducted in farming ponds of Penaeus vannamei in three states in Malaysia, namely Penang, Kedah, and Johor. A total of 82 macrofauna specimens (phyla: Arthropoda, Mollusca, and Chordata) were amplified through a polymerase chain reaction (PCR) assay targeting genes encoding spore wall proteins (SWP) of EHP. The PCR results showed an average prevalence of EHP (82.93%) from three phyla (Arthropoda, Mollusca and Chordata). The phylogenetic tree generated from the macrofauna sequences was revealed to be identical to the EHP-infected shrimp specimens from Malaysia (MW000458, MW000459, and MW000460), as well as those from India (KY674537), Thailand (MG015710), Vietnam (KY593132), and Indonesia (KY593133). These findings suggest that certain macrofauna species in shrimp ponds of P. vannamei are carriers of EHP spores and could be potential transmission vectors. This study provides preliminary information for the prevention of EHP infections that can be initiated at the pond stage by eradicating macrofauna species identified as potential vectors.

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The consolidation of learned information into long-lasting memories requires the strengthening of synaptic connections through de novo protein synthesis. Translation initiation factors play a cardinal role in gating the production of new proteins thereby regulating memory formation. Both positive and negative regulators of translation play a critical role in learning and memory consolidation. The eukaryotic initiation factor 4E (eIF4E) homologous protein (4EHP, encoded by the gene Eif4e2) is a pivotal negative regulator of translation but its role in learning and memory is unknown. To address this gap in knowledge, we generated excitatory (glutamatergic: CaMKIIα-positive) and inhibitory (GABAergic: GAD65-positive) conditional knockout mice for 4EHP, which were analyzed in various behavioral memory tasks. Knockout of 4EHP in Camk2a-expressing neurons (4EHP-cKOexc) did not impact long-term memory in either contextual fear conditioning or Morris water maze tasks. Similarly, long-term contextual fear memory was not altered in Gad2-directed 4EHP knockout mice (4EHP-cKOinh). However, when subjected to a short-term T-maze working memory task, both mouse models exhibited impaired cognition. We therefore tested the hypothesis that de novo protein synthesis plays a direct role in working memory. We discovered that phosphorylation of ribosomal protein S6, a measure of mTORC1 activity, is dramatically reduced in the CA1 hippocampus of 4EHP-cKOexc mice. Consistently, genetic reduction of mTORC1 activity in either excitatory or inhibitory neurons was sufficient to impair working memory. Taken together, these findings indicate that translational control by 4EHP and mTORC1 in both excitatory and inhibitory neurons are necessary for working memory.

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The cap-binding protein 4EHP/eIF4E2 has been a recent object of interest in the field of post-transcriptional gene regulation and translational control. From ribosome-associated quality control, to RNA decay and microRNA-mediated gene silencing, this member of the eIF4E protein family regulates gene expression through numerous pathways. Low in abundance but ubiquitously expressed, 4EHP interacts with different binding partners to form multiple protein complexes that regulate translation in a variety of biological contexts. Documented functions of 4EHP primarily relate to its role as a translational repressor, but recent findings indicate that it might also participate in the activation of translation in specific settings. In this review, we discuss the known functions, properties and mechanisms that involve 4EHP in the control of gene expression. We also discuss our current understanding of how 4EHP processes are regulated in eukaryotic cells, and the diseases implicated with dysregulation of 4EHP-mediated translational control.

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Here we report for the first time a laboratory challenge model for Enterocytozoon hepatopenaei (EHP) to determine the difference of two Specific Pathogen Free (SPF) lines of Penaeus vannamei shrimp. These lines were experimentally challenged using EHP-infected fecal strings as inoculum. Real-time PCR and histopathology assays were performed to confirm EHP infection and evaluate differences in EHP susceptibility in the two genetic lines screened. Although the histopathology of the hepatopancreas tissue showed EHP lesions in both challenged groups, the histological lesions were more pronounced in one of the SPF lines. Quantitative PCR results revealed that animals displaying less hepatocellular damage have lower EHP load compared to animals displaying more pronounced pathological changes. There was no significant difference in final survival at 36 days post-infection in these lines with survival ranging between 80 and 100%. The data showed that mortality as an endpoint metric is not a suitable parameter to determine genetic susceptibility to EHP. Instead, histopathological changes in hepatopancreas, EHP load of the same tissue, and growth retardation would be better metrics to screen EHP susceptibility in P. vannamei. The results show the feasibility of screening genetic lines of P. vannamei for EHP resistance/tolerance using fecal string as an inoculum and, assessing histopathological changes, EHP load, and weight as indicators of resistance.

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PURPOSE: To translate, adapt and validate the Endometriosis Health Profile-5 (EHP-5) in Croatian population. METHODS: This validation study is a part of a prospective, observational study (EHP-5 CRO) with aim of implementation of EHP-5 and to provide better insight in quality of life consideration of women with endometriosis in Croatian clinical practice. A 150 consecutive patients with surgically proven endometriosis were enrolled. The translation to Croatian followed standardized procedure. Cronbach's Alpha was calculated to calculate internal consistency reliability of EHP-5. The test-retest reliability was calculated using intraclass correlation coefficient (ICC). The t test for independent samples was used to assess known-groups validity. RESULTS: Both EHP-5 core and EHP-5 modular parts of the questionnaire had good internal consistency, assessed by the Cronbach's Alpha coefficient (α = 0.793 and α = 0.842, respectively). Obtained results indicate very good reliability for core as well as for modular part of EHP-5 questionnaire (ICC = 0.896 and 0.936, respectively). The independent t test showed that women who reported their pain with VAS scale 7 or more had significantly higher results (p < 0.001) on EHP-5 (M = 50.63) compared with women who reported their pain 6 or less (M = 26.91). Furthermore, we found statistically significant difference between women who are infertile with women who are fertile (p < 0.001), whereby infertile women had higher average result on EHP-5 (M = 49.55) compared with fertile women (M = 34.36). CONCLUSIONS: The Croatian version of the EHP-5 have very good psychometric characteristics and can be used as a reliable tool for assessing patients with endometriosis in everyday clinical practice.

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BACKROUND: Endometriosis is one of the most common gynecological illnesses causing extensive psychological, physical and social impact on patient's life and exerts negative effects on health-related quality of Life (HRQoL). However, the effects of surgery on the postoperative HRQoL in the different endometriosis subgroups have not been fully evaluated. METHODS: We performed a comparative retrospective study between 2014 and 2018 at the Medical University of Vienna, including all patients with surgically confirmed endometriosis who had completed the standardized Endometriosis Health Profile-30 (EHP-30) questionnaire 1 day after surgery (the questions refer to the 4 weeks preoperatively) and 6-10 weeks postoperatively. RESULTS: Compared to preoperative values, we found significant benefits, regarding postoperative conditions, in our study group (n = 115) in all five categories, "pain" (HR 0.78, p < 0.001); "self-determination" (HR 0.92, p < 0.001); "emotional health" (HR 0.83, p < 0.001);" social environment" (HR 0.67, p < 0.001); and "self-image" (HR 0.47, p < 0.001). Patients with only peritoneal endometriosis had the lowest preoperative clinical symptoms and there were no significant changes in any of the categories. In the subgroups deep infiltrating endometriosis (DIE) and DIE + ovarian endometrioma, surgical intervention results in a significantly greater improvement in all categories of EHP 30 compared to ovarian endometrioma without DIE or peritoneal endometriosis. CONCLUSION: Our study shows, that especially women with DIE-with or without ovarian endometrioma-demonstrate a more pronounced benefit from surgical therapy compared to patients with peritoneal endometriosis or endometrioma without DIE.

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In the Republic of Korea, Enterocytozoon hepatopenaei (EHP) was first isolated from Pacific whiteleg shrimp in April 2020; however, there are no existing reports of EHP infection in other shrimp or prawns. Here, we aimed to investigate EHP infection and its prevalence in giant freshwater prawn farms in the Republic of Korea. We tested prawns from 22 farms for EHP infection, and samples from eight farms showed positive EHP infection results in 2021. In EHP-infected prawn farms, the prevalence ranged from 4.9% to 18.2%. The prevalence of EHP infection in the Republic of Korea, derived from the prevalence in prawn farms, was estimated to be 0.8% in 2021. The proliferation of EHP was observed within the hepatopancreatic epithelial cells of prawns using H&E and Giemsa staining. Mature EHP was observed in the sinus between epithelial cells of the digestive tubules. Phylogenetic analysis revealed a clade distinct from the previously reported EHP in Pacific whiteleg shrimps. This is the first report of EHP infection in a giant freshwater prawn in the Republic of Korea, where the prevalence of EHP infection is not high, but it is recognized as an emerging disease that requires periodic monitoring and quarantine management in giant freshwater prawns.

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Enterocytozoon hepatopenaei (EHP), is an emerging microsporidian pathogen responsible for hepatopancreatic microsporidiasis (HPM) in shrimps and is associated with severe growth retardation. The disease causes economic losses in shrimp aquaculture. In this study, EHP spore germination was induced and demonstrated with a scanning electron microscope (SEM). The ions (cations and anions) generated by high-energy electrons during frozen water radiolysis in the SEM specimen chamber induce EHP spore germination. This study is the first to demonstrate the induction of a microsporidian spore germination by ions generated under SEM. This study will enhance our understanding of EHP biology, life cycle and lead to the development of prophylactics and therapeutics for EHP control. Also, this method will help standardize the study of germination in other microsporidians.

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