家蚕内网虫属微孢虫核糖体小亚单位RNA(SSUrRNA)核心序列的克隆和序列分析

Endoreticulatus bombycis is a new pathogenic microspordia isolated from the silkworm larvae.With polymerase chain reaction(PCR) method,we amplified a fragment of the core sequence of the small subunit ribosomal RNA( SSUr-RNA) of Endoreticulatus bombycis.The SSUrRNA fragment was inserted into pMD18-T Vector and then cloned.It had a length of 1230 nucleotides and a percentage GC content of 51.3%.The accession number in Genbank is gill 181769|AY009115.The secondary structure model of the SSUrRNA of Endoreticulatus bombycis was constructed both on the bases of the sequence alignment and RNAFOLD program of the PC-GENE package.The secondary structure model revealed that Endoreticulatus bombycis lacked many cukaryotic hclices.such as heli10,helix 11.helix 18,helix 43 and helix 46,but it has V4 region and has more of prokaryotic character.Analyzed by Blast,Endoreticulatus bombycis.Endoreticulatus schubergi and pleistophora sp.(Sd-Nu-IW8201) have a high similarity,over 98%,Phylogeny tree of Endoreticulatus and Pleistophora species showed that Endoreticulatus and Pleistophora sp.(Sd0Nu-IW8201) was in a group and the other Pleistophora species were in another group[Acta Zoologica Sinica 49(3):414-420,2001].     

Vertical transmission and overwintering of microsporidia in the gypsy moth, Lymantria dispar

Vertical transmission and the overwintering success of three different microsporidia infecting Lymantria dispar (Lepidoptera: Lymantriidae) larvae were investigated. Endoreticulatus schubergi, a midgut pathogen, was transmitted to offspring via female and male via the egg chorion (transovum transmission). Between 8% and 29% of the emerging larvae became infected. No spores of E. schubergi were found in surface-washed eggs. Nosema lymantriae, a microsporidium that causes systemic infections, was transovarially transmitted. Between 35% and 72% of the progeny were infected. Vairimorpha disparis, a fat body pathogen, was not vertically transmitted. The infectivity of spores that overwintered in cadavers of infected L. dispar varied by species, placement in the environment, and weather conditions. Spores of E. schubergi were still infective after an eight month exposure period of cadavers on the ground. Spores of N. lymantriae and V. disparis remained highly infective only when cadavers overwintered under a more or less continuous snow cover for four months.     

Occurrence of Cystosporogenes sp. (Protozoa, Microsporidia) in a multi-species insect production facility and its elimination from a colony of the eastern spruce budworm, Choristoneura fumiferana (Clem.) (Lepidoptera: Tortricidae)

We have isolated a microsporidium from a laboratory colony of the eastern spruce budworm, Choristoneura fumiferana (Clem.) (Lepidoptera: Tortricidae). Light and electron microscopic investigations showed that gross pathology and ultrastructure of our isolate are similar to those described for Cystosporogenes legeri from the European grape vine moth, Lobesia botrana. Comparative phylogenetic analysis of the small subunit rDNA using maximum likelihood, maximum parsimony, and neighbour joining distance methods revealed perfect homology with the C. legeri sequence. The microsporidian was infectious to other Choristoneura species, as well as Malacosoma disstria, Lymantria dispar, and Lambdina fiscellaria. Incubation of infected egg masses at 41 degrees C for 20 min followed by 30 min in 33% formaldehyde did not reduce disease incidence in larval offspring. Exposure of one or two generations to fumagillin at 6000 ppm or higher eliminated infection in adult moths, but also reduced colony fitness. A clean colony was established by conducting individual matings and selecting disease-free offspring.     

Microsporidian infections in Lymantria dispar larvae: interactions and effects of multiple species infections on pathogen horizontal transmission

The interactions in multiple species infections and effects on the horizontal transmission of three microsporidian species, Vairimorpha disparis, Nosema lymantriae and Endoreticulatus schubergi, infecting Lymantria dispar were evaluated in the laboratory. Simultaneous and sequential inoculations of host larvae were performed and the resulting infections were evaluated. Test larvae were exposed to the inoculated larvae to measure horizontal transmission. Dual species infections demonstrated interspecific competition between Nosema and Vairimorpha in the host larvae, but no observable competition occurred between Endoreticulatus and either of the other microsporidian species. Timing of inoculation was an important factor determining the outcome of competition between Nosema and Vairimorpha. The species inoculated first showed a higher rate of successful establishment; a time lag of 7 days between inoculations allowed the first species to essentially exclude the second. The microsporidia differed in efficiency of horizontal transmission. Nosema and Endoreticulatus were transmitted at very high rates, close to 100%. Horizontal transmission of Vairimorpha was less efficient, ranging from 25% to a maximum of 75%. The patterns of infection observed in inoculated larvae were reflected in the test larvae that acquired infections in the horizontal transmission experiments. Competition with Vairimorpha suppressed horizontal transmission of Nosema after simultaneous and sequential inoculation. In simultaneous inoculation experiments Endoreticulatus had no effect on transmission of Nosema and Vairimorpha.     

Three microsporidian pathogens infecting Lymantria dispar larvae do not differ in their success in horizontal transmission

We quantified horizontal transmission of three microsporidian pathogens, Endoreticulatus schubergi, Nosema lymantriae and Vairimorpha disparis that infect Lymantria dispar larvae in an experiment using caged, potted oak plants. Despite marked differences in the modes of spore release from infectious hosts, no significant differences in the transmission success to uninfected, susceptible test hosts were ascertained between the tested microsporidian species. The density of initially inoculated larvae and the exposure period, on the other hand, did influence the number of infected test larvae. Depending on the density of inoculated larvae (10%, 30% or 50%), between 0% and 26% of the test larvae became infected with one of the three tested microsporidian pathogens after an exposure period of 6 days. When the exposure period was 12 days, between 11% and 76% of the test larvae became infected.     

Ultrastructural study on a novel microsporidian, Endoreticulatus eriocheir sp. nov. (Microsporidia, Encephalitozoonidae), parasite of Chinese mitten crab, Eriocheir sinensis (Crustacea, Decapoda)

A microsporidian pathogen, infecting the epithelial cells of the hepatopancreas of Chinese mitten crab, Eriocheir sinensis, was studied by electron microscopy. The detailed ultrastructure of life cycle of the pathogen including proliferative and sporogonic developmental stages are provided. All stages of the parasite are haplokaryotic and develop in a vacuole bounded by a single membrane in contact with host cell cytoplasm. Sporogenesis is synchronous with the same developmental stage in one vacuole. Sporogony shows a characteristic of multinucleate sporogonial plasmodia divided by rosette-like division, producing 4 or 8 sporoblasts. The mature spore is ellipsoidal, length (mean) 1.7 microm, width 1.0 microm, with a uninucleate in the center of the sporoplasm, 7 turns of the polar filament, a bell-like polaroplast of compact membranes and obliquely positioned posterior vacuole. The morphological characteristics of this novel microsporidian pathogen have led us to assign the parasite to a new species of Endoreticulatus, E. eriocheir sp. nov., that has not been reported previously from crab.     

Small Subunit Ribosomal DNA Phylogeny of Various Microsporidia with Emphasis on AIDS Related Forms

ABSTRACT. Phylogenetic analysis of the small subunit ribosomal DNA of a broad range of representative microsporidia including five species from humans ( Enterocytozoon bieneusi, Nosema corneum, Septata intestinalis, Encephalitozoon hellem and Encephalitozoon cuniculi ), reveals that human microsporidia are polyphyletic in origin. Septata intestinalis and E. hellem are very similar to the mammalian parasite E. cuniculi . Based on the results of our phylogenetic analysis, we suggest that S. intestinalis be designated Encephalitozoon intestinalis . Furthermore, analysis of our data indicates that N. corneum is much more closely related to the insect parasite Endoreticulatus schubergi than it is to other Nosema species. This finding is supported by recent studies which have shown a similarity between E. schubergi and N. corneum based on the origin and development of the parasitophorous vacuole. Thus these opportunistic microsporidian parasites can originate from hosts closely or distantly related to humans. Finally, the phylogeny based on small subunit ribosomal DNA sequences is highly inconsistent with traditional classifications based on morphological characters. Many of the important morphological characters (diplokaryon, sporophorous vesicle, and meiosis) appear to have multiple origins.     

Morphological and molecular investigations of a microsporidium infecting the European grape vine moth,Lobesia botrana Den. et Schiff., and its taxonomic determination as Cystosporogenes legeri nov. comb

We have isolated a microsporidium from a laboratory stock of the European grape vine moth, Lobesia botrana Den. et Schiff. (Lepidoptera, Tortricidae). Screening of this stock showed an infection rate of more than 90%, whereas field collected larvae from three different locations in Rhineland-Palatinate (Germany) did not demonstrate any signs of infection. Light and electron microscopic investigations of infected insects showed that gross pathology, morphology, and ultrastructure of the microsporidium are similar to those described earlier for Pleistophora legeri. Comparative phylogenetic analysis of the small subunit rDNA using maximum likelihood, maximum parsimony, and neighbour joining distance methods showed that our isolate was closely related to Cystosporogenes operophterae. Based on our morphological and molecular investigations we propose to rename this species Cystosporogenes legeri nov. comb.