Distribution and genotypic analysis of Enterocytozoon bieneusi from wild boars in Korea.

Enterocytozoon bieneusi, an important microsporidian fungus, causes chronic diarrhea in humans and animals worldwide. Out of the 502 fecal samples from wild boars, 13 were positive for the E. bieneusi internal transcribed spacer region, with a prevalence of 2.6%. Six E. bieneusi genotypes, D, EbpC, and four novel KWB1-KWB4, were identified with zoonotic potential. Genotypes D (subgroup 1a) and EbpC (subgroup 1d) were first reported in Korean swine and Korea, respectively; KWB1-KWB4 (subgroup 1e) were most prevalent in this study. Because zoonotic genotypes have been identified, E. bieneusi transmission through wild boars must be closely monitored for proper prevention and treatment, despite their low prevalence. LAY SUMMARY: Enterocytozoon bieneusi is an important microsporidian fungus. Its sequences from wild boars were identified with zoonotic potential. Genotypes D and EbpC were first reported in Korean swine and Korea, respectively. E. bieneusi should be closely monitored to properly prevent and treat animals.

Transcriptional response of honey bee (Apis mellifera) to differential nutritional status and Nosema infection.

BACKGROUND: Bees are confronting several environmental challenges, including the intermingled effects of malnutrition and disease. Intuitively, pollen is the healthiest nutritional choice, however, commercial substitutes, such as Bee-Pro and MegaBee, are widely used. Herein we examined how feeding natural and artificial diets shapes transcription in the abdomen of the honey bee, and how transcription shifts in combination with Nosema parasitism. RESULTS: Gene ontology enrichment revealed that, compared with poor diet (carbohydrates [C]), bees fed pollen (P > C), Bee-Pro (B > C), and MegaBee (M > C) showed a broad upregulation of metabolic processes, especially lipids; however, pollen feeding promoted more functions, and superior proteolysis. The superiority of the pollen diet was also evident through the remarkable overexpression of vitellogenin in bees fed pollen instead of MegaBee or Bee-Pro. Upregulation of bioprocesses under carbohydrates feeding compared to pollen (C > P) provided a clear poor nutritional status, uncovering stark expression changes that were slight or absent relatively to Bee-Pro (C > B) or MegaBee (C > M). Poor diet feeding (C > P) induced starvation response genes and hippo signaling pathway, while it repressed growth through different mechanisms. Carbohydrate feeding (C > P) also elicited 'adult behavior', and developmental processes suggesting transition to foraging. Finally, it altered the 'circadian rhythm', reflecting the role of this mechanism in the adaptation to nutritional stress in mammals. Nosema-infected bees fed pollen compared to carbohydrates (PN > CN) upheld certain bioprocesses of uninfected bees (P > C). Poor nutritional status was more apparent against pollen (CN > PN) than Bee-Pro (CN > BN) or MegaBee (CN > MN). Nosema accentuated the effects of malnutrition since more starvation-response genes and stress response mechanisms were upregulated in CN > PN compared to C > P. The bioprocess 'Macromolecular complex assembly' was also enriched in CN > PN, and involved genes associated with human HIV and/or influenza, thus providing potential candidates for bee-Nosema interactions. Finally, the enzyme Duox emerged as essential for guts defense in bees, similarly to Drosophila. CONCLUSIONS: These results provide evidence of the superior nutritional status of bees fed pollen instead of artificial substitutes in terms of overall health, even in the presence of a pathogen.

A Large Collection of Novel Nematode-Infecting Microsporidia and Their Diverse Interactions with Caenorhabditis elegans and Other Related Nematodes.

Microsporidia are fungi-related intracellular pathogens that may infect virtually all animals, but are poorly understood. The nematode Caenorhabditis elegans has recently become a model host for studying microsporidia through the identification of its natural microsporidian pathogen Nematocida parisii. However, it was unclear how widespread and diverse microsporidia infections are in C. elegans or other related nematodes in the wild. Here we describe the isolation and culture of 47 nematodes with microsporidian infections. N. parisii is found to be the most common microsporidia infecting C. elegans in the wild. In addition, we further describe and name six new species in the Nematocida genus. Our sampling and phylogenetic analysis further identify two subclades that are genetically distinct from Nematocida, and we name them Enteropsectra and Pancytospora. Interestingly, unlike Nematocida, these two genera belong to the main clade of microsporidia that includes human pathogens. All of these microsporidia are horizontally transmitted and most specifically infect intestinal cells, except Pancytospora epiphaga that replicates mostly in the epidermis of its Caenorhabditis host. At the subcellular level in the infected host cell, spores of the novel genus Enteropsectra show a characteristic apical distribution and exit via budding off of the plasma membrane, instead of exiting via exocytosis as spores of Nematocida. Host specificity is broad for some microsporidia, narrow for others: indeed, some microsporidia can infect Oscheius tipulae but not its sister species Oscheius sp. 3, and conversely some microsporidia found infecting Oscheius sp. 3 do not infect O. tipulae. We also show that N. ausubeli fails to strongly induce in C. elegans the transcription of genes that are induced by other Nematocida species, suggesting it has evolved mechanisms to prevent induction of this host response. Altogether, these newly isolated species illustrate the diversity and ubiquity of microsporidian infections in nematodes, and provide a rich resource to investigate host-parasite coevolution in tractable nematode hosts.

A new microsporidium, Vairimorpha subcoccinellae n. sp. (Microsporidia: Burenellidae), isolated from Subcoccinella vigintiquatuorpunctata L. (Coleoptera: Coccinellidae).

A new microsporidium was isolated from Subcoccinella vigintiquatuorpunctata L. (Coleoptera: Coccinellidae), a pest of Galega officinalis L. in Turkey. Infection in larval and adult stages was systemic with mature spores produced in the midgut, gonads, Malpighian tubules and, most extensively, fat body tissues. The microsporidium was polymorphic with two sporulation sequences producing two types of spores, binucleate spores with 13-15 coils of the polar tube, and uninucleate spores with 7 coils of the polar tube that developed within a sporophorous vesicle (SPV) to form meiospores. The 16S small subunit rRNA (SSU rRNA) gene of the microsporidium was sequenced and compared with twenty-seven microsporidian sequences from GenBank. Based on the phylogenetic analysis of the SSU rRNA sequence, this microsporidium is unique within the Vairimorpha group. Morphological and genetic characters indicate that the described microsporidium is dissimilar to all known Vairimorpha species, and so is named here as Vairimorpha subcoccinellae n. sp.

A wild C. elegans strain has enhanced epithelial immunity to a natural microsporidian parasite.

Microbial pathogens impose selective pressures on their hosts, and combatting these pathogens is fundamental to the propagation of a species. Innate immunity is an ancient system that provides the foundation for pathogen resistance, with epithelial cells in humans increasingly appreciated to play key roles in innate defense. Here, we show that the nematode C. elegans displays genetic variation in epithelial immunity against intestinal infection by its natural pathogen, Nematocida parisii. This pathogen belongs to the microsporidia phylum, which comprises a large phylum of over 1400 species of fungal-related parasites that can infect all animals, including humans, but are poorly understood. Strikingly, we find that a wild C. elegans strain from Hawaii is able to clear intracellular infection by N. parisii, with this ability restricted to young larval animals. Notably, infection of older larvae does not impair progeny production, while infection of younger larvae does. The early-life immunity of Hawaiian larvae enables them to produce more progeny later in life, providing a selective advantage in a laboratory setting--in the presence of parasite it is able to out-compete a susceptible strain in just a few generations. We show that enhanced immunity is dominant to susceptibility, and we use quantitative trait locus mapping to identify four genomic loci associated with resistance. Furthermore, we generate near-isogenic strains to directly demonstrate that two of these loci influence resistance. Thus, our findings show that early-life immunity of C. elegans against microsporidia is a complex trait that enables the host to produce more progeny later in life, likely improving its evolutionary success.

Molecular Characterization of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in Captive Wildlife at Zhengzhou Zoo, China.

Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are common gastrointestinal protists in humans and animals. Two hundred and three fecal specimens from 80 wildlife species were collected in Zhengzhou Zoo and their genomic DNA extracted. Three intestinal pathogens were characterized with a DNA sequence analysis of different loci. Cryptosporidium felis, C. baileyi, and avian genotype III were identified in three specimens (1.5%), the manul, red-crowned crane, and cockatiel, respectively. Giardia duodenalis was also found in five specimens (2.5%) firstly: assemblage B in a white-cheeked gibbon and beaver, and assemblage F in a Chinese leopard and two Siberian tigers, respectively. Thirteen genotypes of E. bieneusi (seven previously reported genotypes and six new genotypes) were detected in 32 specimens (15.8%), of which most were reported for the first time. A phylogenetic analysis of E. bieneusi showed that five genotypes (three known and two new) clustered in group 1; three known genotypes clustered in group 2; one known genotype clustered in group 4; and the remaining four genotypes clustered in a new group. In conclusion, zoonotic Cryptosporidium spp., G. duodenalis, and E. bieneusi are maintained in wildlife and transmitted between them. Zoonotic disease outbreaks of these infectious agents possibly originate in wildlife reservoirs.

Proteomic analysis of BmN cells (Bombyx mori) in response to infection with Nosema bombycis.

Nosema bombycis (N. bombycis, Nb) is an obligate intracellular parasite, which can cause pebrine disease in the silkworm. To investigate the effects of N. bombycis infection on the host cells, proteomes from BmN cells that had or had not been infected with N. bombycis at different infection stages were characterized with two-dimensional gel electrophoresis and MALDI-TOF/TOF mass spectrometry, which identified 24 differentially expressed host proteins with significant intensity differences (P < 0.05) at least at one time point in mock- and N. bombycis infected cells. Notably, gene ontology analyses showed that these proteins are involved in many important biological reactions. During the infection phase, proteins involved in energy metabolism and oxidative stress had up-regulated expression. Two proteins participated in ubiquitin-dependent protein catabolic process had down-regulated expression. Quantitative real-time polymerase chain reaction was used to analyze the transcriptional profiles of these identified proteins. Taken together, the abundance changes, putative functions, and participation in biological reactions for the identified proteins produce a host-responsive protein model in N. bombycis-infected BmN cells. These findings further our knowledge about the effect of energy defect parasites on the host cells.

Molecular detection of Enterocytozoon bieneusi in farmed Asiatic brush-tailed porcupines (Atherurus macrourus) and bamboo rats (Rhizomys pruinosus) from Hainan Province, China: Common occurrence, wide genetic variation and high zoonotic potential.

We investigated the occurrence and genotypic diversity of E. bieneusi in farmed Asiatic brush-tailed porcupines and bamboo rats from Hainan Province, China. Four hundred and sixty-seven fresh feces were collected from 164 Asiatic brush-tailed porcupines and 303 bamboo rats. DNA extraction from the feces and genotyping of E. bieneusi were performed by the amplification of the internal transcribed spacer (ITS) region of rDNA of E. bieneusi using PCR. A neighbor-joining tree was constructed based on the sequences obtained here and other sequences of E. bieneusi genotypes stored in Genbank. The total rate of infection with E. bieneusi was 32.5% (152/467), with 14.6% (24/164) in Asiatic brush-tailed porcupines and 42.2% (128/303) in bamboo rats infected. Seventeen genotypes of E. bieneusi were identified including 12 known genotypes, i.e., D (n = 78), Henan-III (n = 21), SHW7 (n = 19), KIN-1 (n = 11), ETMK5 (n = 7), TypeIV (n = 4), EbpD (n = 2), EbpA (n = 1), EbpC (n = 1), S7 (n = 1), HNPL-III (n = 1), HNR-VII (n = 1), and five novel genotypes named as HNZS-I (n = 1) and HNHZ-I to HNHZ-IV (n = 1 per genotype). Phylogenetic analysis revealed that all the genotypes found here except genotype S7 fell into Group 1. The present study demonstrated a relatively high prevalence of E. bieneusi infection (32.5%) and a large genetic variation of E. bieneusi (seventeen genotypes) in farmed Asiatic brush-tailed porcupines and bamboo rats in Hainan, China. The high proportion (78.3%) of zoonotic genotypes identified in the animals investigated here suggests that there is the potential for zoonotic or cross-species transmission which may pose a serious public health threat in the area. Public education on the management of Asiatic brush-tailed porcupines and bamboo rats should be implemented in the investigated areas.

Survival and immune response of drones of a Nosemosis tolerant honey bee strain towards N. ceranae infections.

Honey bee colonies (Apis mellifera) have been selected for low level of Nosema in Denmark over decades and Nosema is now rarely found in bee colonies from these breeding lines. We compared the immune response of a selected and an unselected honey bee lineage, taking advantage of the haploid males to study its potential impact on the tolerance toward Nosema ceranae, a novel introduced microsporidian pathogen. After artificial infections of the N. ceranae spores, the lineage selected for Nosema tolerance showed a higher N. ceranae spore load, a lower mortality and an up-regulated immune response. The differences in the response of the innate immune system between the selected and unselected lineage were strongest at day six post infection. In particular genes of the Toll pathway were up-regulated in the selected strain, probably is the main immune pathway involved in N. ceranae infection response. After decades of selective breeding for Nosema tolerance in the Danish strain, it appears these bees are tolerant to N. ceranae infections.

[Contribution of PCR for detection and identification of intestinal microsporidia in HIV-infected patients]. / Apport de la PCR dans la recherche et l'identification des microsporidies intestinales chez les sujets infectés par le VIH.

AIM: Intestinal microsporidiosis are among the most frequent opportunistic diseases in immunocompromised subjects. This study aimed to evaluate the contribution of PCR for a better detection and species identification of microsporidia in stool specimens of HIV-infected patients. PATIENTS AND METHODS: Stool samples obtained from 119 HIV-infected Tunisian subjects were screened for intestinal microsporidiosis by light microscopy using Weber's modified Trichrome stain and by a PCR method using universal primers V1/PMP2 which amplified a common fragment of the small subunit rRNA gene of microsporidia. The obtained PCR products were then sequenced using an ABI PRISM 377 DNA sequencer. RESULTS: The results showed a better sensitivity of PCR in the detection of microsporidia with an infection rate of 14.3% significantly higher than that of 6.7% obtained by light microscopy (p=0.03). As previously described, intestinal microsporidiosis was associated with low CD4 cell counts; 23.9% infection rate in patients having CD4 cell count under 200/mm(3) against 5.6% in patients with higher CD4 cell count (p=0.008). The sequencing of 15 out of the 17 positive PCR products has confirmed in all cases the species identified based on the PCR fragment size i.e., 250pb for Enterocytozoon bieneusi (seven cases) and about 270pb for Encephalitozoon intestinalis (nine cases); one case revealed a double infection. CONCLUSION: PCR proved to be more effective than classical Trichrome stain for the diagnosis of intestinal microsporidiosis. Moreover, the ability of PCR to identify the species involved could also be useful for cases management.

Conservation of Nematocida microsporidia gene expression and host response in Caenorhabditis nematodes.

Microsporidia are obligate intracellular parasites that are known to infect most types of animals. Many species of microsporidia can infect multiple related hosts, but it is not known if microsporidia express different genes depending upon which host species is infected or if the host response to infection is specific to each microsporidia species. To address these questions, we took advantage of two species of Nematocida microsporidia, N. parisii and N. ausubeli, that infect two species of Caenorhabditis nematodes, C. elegans and C. briggsae. We performed RNA-seq at several time points for each host infected with either microsporidia species. We observed that Nematocida transcription was largely independent of its host. We also observed that the host transcriptional response was similar when infected with either microsporidia species. Finally, we analyzed if the host response to microsporidia infection was conserved across host species. We observed that although many of the genes upregulated in response to infection are not direct orthologs, the same expanded gene families are upregulated in both Caenorhabditis hosts. Together our results describe the transcriptional interactions of Nematocida infection in Caenorhabditis hosts and demonstrate that these responses are evolutionarily conserved.

Factors That Determine Microsporidia Infection and Host Specificity.

Microsporidia are a large phylum of obligate intracellular parasites that infect an extremely diverse range of animals and protists. In this chapter, we review what is currently known about microsporidia host specificity and what factors influence microsporidia infection. Extensive sampling in nature from related hosts has provided insight into the host range of many microsporidia species. These field studies have been supported by experiments conducted in controlled laboratory environments which have helped to demonstrate host specificity. Together, these approaches have revealed that, while examples of generalist species exist, microsporidia specificity is often narrow, and species typically infect one or several closely related hosts. For microsporidia to successfully infect and complete their life cycle within a compatible host, several steps must occur, including spore germination, host cell invasion, and proliferation of the parasite within the host tissue. Many factors influence infection, including temperature, seasonality, nutrient availability, and the presence or absence of microbes, as well as the developmental stage, sex, and genetics of the host. Several studies have identified host genomic regions that influence resistance to microsporidia, and future work is likely to uncover molecular mechanisms of microsporidia host specificity in more detail.

Recent Advances with Fish Microsporidia.

There have been several significant new findings regarding Microsporidia of fishes over the last decade. Here we provide an update on new taxa, new hosts and new diseases in captive and wild fishes since 2013. The importance of microsporidiosis continues to increase with the rapid growth of finfish aquaculture and the dramatic increase in the use of zebrafish as a model in biomedical research. In addition to reviewing new taxa and microsporidian diseases, we include discussions on advances with diagnostic methods, impacts of microsporidia on fish beyond morbidity and mortality, novel findings with transmission and invertebrate hosts, and a summary of the phylogenetics of fish microsporidia.

Immune Response to Microsporidia.

Microsporidia are a group of pathogens, which can pose severe risks to the immunocompromised population, such as HIV-infected individuals or organ transplant recipients. Adaptive immunity has been reported to be critical for protection, and mice depleted of T cells are unable to control these infections. In a mouse model of infection, CD8 T cells have been found to be the primary effector cells and are responsible for protecting the infected host. Also, as infection is acquired via a peroral route, CD8 T cells in the gut compartment act as a first line of defense against these pathogens. Thus, generation of a robust CD8 T-cell response exhibiting polyfunctional ability is critical for host survival. In this chapter, we describe the effector CD8 T cells generated during microsporidia infection and the factors that may be essential for generating protective immunity against these understudied but significant pathogens. Overall, this chapter will highlight the necessity for a better understanding of the development of CD8 T-cell responses in gut-associated lymphoid tissue (GALT) and provide some insights into therapies that may be used to restore defective CD8 T-cell functionality in an immunocompromised situation.

Identification and subcellular localization analysis of membrane protein Ycf 1 in the microsporidian Nosema bombycis.

Microsporidia are obligate intracellular parasites that can infect a wide range of vertebrates and invertebrates including humans and insects, such as silkworm and bees. The microsporidium Nosema bombycis can cause pebrine in Bombyx mori, which is the most destructive disease in the sericulture industry. Although membrane proteins are involved in a wide range of cellular functions and part of many important metabolic pathways, there are rare reports about the membrane proteins of microsporidia up to now. We screened a putative membrane protein Ycf 1 from the midgut transcriptome of the N. bombycis-infected silkworm. Gene cloning and bioinformatics analysis showed that the Ycf 1 gene contains a complete open reading frame (ORF) of 969 bp in length encoding a 322 amino acid polypeptide that has one signal peptide and one transmembrane domain. Indirect immunofluorescence results showed that Ycf 1 protein is distributed on the plasma membrane. Expression pattern analysis showed that the Ycf 1 gene expressed in all developmental stages of N. bombycis. Knockdown of the Ycf 1 gene by RNAi effectively inhibited the proliferation of N. bombycis. These results indicated that Ycf 1 is a membrane protein and plays an important role in the life cycle of N. bombycis.

Molecular diversity of the microsporidium Kneallhazia solenopsae reveals an expanded host range among fire ants in North America.

Kneallhazia solenopsae is a pathogenic microsporidium that infects the fire ants Solenopsis invicta and Solenopsis richteri in South America and the USA. In this study, we analyzed the prevalence and molecular diversity of K. solenopsae in fire ants from North and South America. We report the first empirical evidence of K. solenopsae infections in the tropical fire ant, Solenopsis geminata, and S. geminata×Solenopsis xyloni hybrids, revealing an expanded host range for this microsporidium. We also analyzed the molecular diversity at the 16S ribosomal RNA gene in K. solenopsae from the ant hosts S.invicta, S. richteri, S. geminata and S. geminata×S. xyloni hybrids from North America, Argentina and Brazil. We found 22 16S haplotypes. One of these haplotypes (WD_1) appears to be widely distributed, and is found in S. invicta from the USA and S. geminata from southern Mexico. Phylogenetic analyses of 16S sequences revealed that K. solenopsae haplotypes fall into one of two major clades that are differentiated by 2-3%. In some cases, multiple K. solenopsae haplotypes per colony were found, suggesting either an incomplete homogenization among gene copies within the 16S gene cluster or multiple K. solenopsae variants simultaneously infecting host colonies.

Genotyping of Enterocytozoon bieneusi among captive long-tailed macaques (Macaca fascicularis) in Hainan Province: High genetic diversity and zoonotic potential.

Enterocytozoon bieneusi is a potentially important zoonotic pathogen. However, there is no information on E. bieneusi infection of captive long-tailed macaques (Macaca fascicularis) in Hainan Province, China. Here 193 fecal specimens of M. fascicularis were collected from a breeding base in Hainan Province, China, housing non-human primates for experimental use. E. bieneusi was identified and genotyped by nested PCR analysis of the internal transcribed spacer (ITS) region of the rRNA gene. A total of 59 (30.6%) specimens were PCR-positive for E. bieneusi and 16 ITS genotypes were identified including nine known genotypes: Type IV (n = 19), D (n = 11), CM1 (n = 8), PigEBITS7 (n = 4), Pongo2 (n = 4), Peru8 (n = 3), Peru11 (n = 1), WL21 (n = 1) and CM2 (n = 1) and seven novel genotypes HNM-I to HNM-VII (one each). Importantly, genotypes D, Type IV, Peru8, PigEBITS7, and Peru11, which were the predominant (38/59, 64.4%) genotypes identified among captive M. fascicularis in this study, are also well-known human-pathogenic genotypes. All the genotypes of E. bieneusi identified here, including the seven novel ones, belonged to zoonotic Group 1. This is the first report of the identification of E. bieneusi in M. fascicularis in Hainan Province, China. The finding that the numerous known human-pathogenic types and seven novel genotypes of E. bieneusi all belong to zoonotic Group 1 indicates the possibility of transmission of this important pathogenic parasite between M. fascicularis and humans.

Enterocytozoon hepatopenaei sp. nov. (Microsporida: Enterocytozoonidae), a parasite of the black tiger shrimp Penaeus monodon (Decapoda: Penaeidae): Fine structure and phylogenetic relationships.

A new microsporidian species, Enterocytozoon hepatopenaei sp. nov., is described from the hepatopancreas of the black tiger shrimp Penaeus monodon (Crustacea: Decapoda). Different stages of the parasite are described, from early sporogonal plasmodia to mature spores in the cytoplasm of host-cells. The multinucleate sporogonal plasmodia existed in direct contact with the host-cell cytoplasm and contained numerous small blebs at the surface. Binary fission of the plasmodial nuclei occurred during early plasmodial development and numerous pre-sporoblasts were formed within the plasmodium. Electron-dense disks and precursors of the polar tubule developed in the cytoplasm of the plasmodium prior to budding of early sporoblasts from the plasmodial surface. Mature spores were oval, measuring 0.7x1.1microm and contained a single nucleus, 5-6 coils of the polar filament, a posterior vacuole, an anchoring disk attached to the polar filament, and a thick electron-dense wall. The wall was composed of a plasmalemma, an electron-lucent endospore (10nm) and an electron-dense exospore (2nm). DNA primers designed from microsporidian SSU rRNA were used to amplify an 848bp product from the parasite genome (GenBank FJ496356). The sequenced product had 84% identity to the matching region of SSU rRNA from Enterocytozoon bieneusi. Based upon ultrastructural features unique to the family Enterocytozoonidae, cytoplasmic location of the plasmodia and SSU rRNA sequence identity 16% different from E. bieneusi, the parasite was considered to be a new species, E. hepatopenaei, within the genus Enterocytozoon.

Molecular evidence of Enterocytozoon bieneusi in Japan.

Enterocytozoon bieneusi is an emerging and clinically significant enteric pathogen in humans associated mainly with chronic diarrhea. It has been found in a variety of wild, domestic and companion mammals and birds. To date, epidemiological surveys of E. bieneusi infection in humans, other mammals and birds have been performed in more than 21 countries in Africa, the Americas, Australasia and Europe. In Asia E. bieneusi has been found in India, Thailand, Vietnam and Korea, but it has been quite unclear whether this pathogen is present in Japan. In the present study, we examined 149 DNAs extracted from 45 human (9 of them HIV-positive) and 104 animal fecal samples by PCR. Two dogs and a cat were positive and their genotypes were found to be dog specific and zoonotic (genotype K) types, respectively. Present study is the first record of E. bieneusi in Japan.

Natural variation in the roles of C. elegans autophagy components during microsporidia infection.

Natural genetic variation can determine the outcome of an infection, and often reflects the co-evolutionary battle between hosts and pathogens. We previously found that a natural variant of the nematode Caenorhabditis elegans from Hawaii (HW) has increased resistance against natural microsporidian pathogens in the Nematocida genus, when compared to the standard laboratory strain of N2. In particular, HW animals can clear infection, while N2 animals cannot. In addition, HW animals have lower levels of initial colonization of Nematocida inside intestinal cells, compared to N2. Here we investigate how this natural variation in resistance relates to autophagy. We found that there is much better targeting of autophagy-related machinery to parasites under conditions where they are cleared. In particular, ubiquitin targeting to Nematocida cells correlates very well with their subsequent clearance in terms of timing, host strain and age, as well as species of Nematocida. Furthermore, clearance correlates with targeting of the LGG-2/LC3 autophagy protein to parasite cells, with HW animals having much more efficient targeting of LGG-2 to parasite cells than N2 animals. Surprisingly, however, we found that LGG-2 is not required to clear infection. Instead, we found that LGG-2/LC3 regulates Nematocida colonization inside intestinal cells. Interestingly, LGG-2/LC3 regulates intracellular colonization only in the HW strain, and not in N2. Altogether these results demonstrate that there is natural genetic variation in an LGG-2-dependent process that regulates microsporidia colonization inside intestinal cells, although not microsporidia clearance.