LysM Receptor-Like Kinase LYK9 of Pisum Sativum L. May Regulate Plant Responses to Chitooligosaccharides Differing in Structure.

This study focused on the interactions of pea (Pisum sativum L.) plants with phytopathogenic and beneficial fungi. Here, we examined whether the lysin-motif (LysM) receptor-like kinase PsLYK9 is directly involved in the perception of long- and short-chain chitooligosaccharides (COs) released after hydrolysis of the cell walls of phytopathogenic fungi and identified in arbuscular mycorrhizal (AM) fungal exudates. The identification and analysis of pea mutants impaired in the lyk9 gene confirmed the involvement of PsLYK9 in symbiosis development with AM fungi. Additionally, PsLYK9 regulated the immune response and resistance to phytopathogenic fungi, suggesting its bifunctional role. The existence of co-receptors may provide explanations for the potential dual role of PsLYK9 in the regulation of interactions with pathogenic and AM fungi. Co-immunoprecipitation assay revealed that PsLYK9 and two proposed co-receptors, PsLYR4 and PsLYR3, can form complexes. Analysis of binding capacity showed that PsLYK9 and PsLYR4, synthesized as extracellular domains in insect cells, were able to bind the deacetylated (DA) oligomers CO5-DA-CO8-DA. Our results suggest that the receptor complex consisting of PsLYK9 and PsLYR4 can trigger a signal pathway that stimulates the immune response in peas. However, PsLYR3 seems not to be involved in the perception of CO4-5, as a possible co-receptor of PsLYK9.

Hexokinase as a versatile molecular genetic marker for Microsporidia.

Hexokinase (HK) is a core glycolytic enzyme of Microsporidia which regulates host cell metabolic processes. The goal of the present study was to test for the utility of HK for molecular phylogenetics, species identification and molecular detection of microsporidia in infected insects. HK sequence-based reconstructions were essentially similar to those based upon largest subunit RNA polymerase (RPB1) gene sequences, as well as previously published rRNA gene and genome-based trees. Comparing HK sequences allowed clear differentiation of closely related taxa, such as Nosema bombycis and Nosema pyrausta. In Nosema ceranae, unique SNPs were found for an isolate from wild colonies of the Burzyan dark honey bee as compared with the isolates from domesticated European honey bee. Similarly, in Encephalitozoon cuniculi, HK was as effective as RPB1 for discrimination of isolates belonging to different ITS genotypes. Amplification using species-specific primers flanking short fragments at the 3'-end of HK gene showed the presence of infection in insect tissues infected with N. pyrausta, Nosema ceranae and Paranosema (Antonospora) locustae. For the latter parasite species, HK expression was also demonstrated at early stages of infection using total mRNA extracts of locust larvae. These results indicate the suitability of HK as a novel tool for molecular genetic studies of Microsporidia.

Ultrastructure and molecular phylogeny of Anisofilariata chironomi g.n. sp.n. (Microsporidia: Terresporidia) from Chironomus plumosus L. (Diptera: Chironomidae).

Larvae of Chironomus plumosus, collected in North-Western Russia in September 2008, were infected with a microsporidium possessing broadly oval uninucleate spores in sporophorous vesicles. Sporogony and spore ultrastructure of this microsporidium differed from that of known microsporidian species, suggesting establishment of a new species, Anisofilariata chironomi, being a type species of a new genus. Sporogony di-, tetra-, octo-, and 16-sporoblastic. Fixed and stained spores are 4.7-6.8 x 3.4-5.4 microm in size, the spore measurements varying depending upon the number of spores in the sporophorous vesicle. The polaroplast is bipartite, with anterior and posterior parts composed of very thin and thick lamellae, respectively, and occupies the major volume of the spore. The polar filament is anisofilar, with two broad proximal and 10-13 narrow distal coils arranged in 2-4 layers. The sporophorous vesicle is bounded by a thin membrane and contains multiple tubular structures. Small subunit ribosomal DNA phylogeny showed basal position of the new microsporidium to a cluster uniting microsporidia infecting ciliates (Euplotespora binucleata), microcrustaceans (Glugoides intestinalis, Mrazekia macrocyclopis), lepidopteran insects (Cystosporogenes spp., Endoreticulatus spp.) and human (Vittaforma corneae), nested within Clade IV sensu Vossbrinck and Debrunner-Vossbrinck (2005 Folia Parasitol 52:131-142). No close phylogenetic relationships were found between A. chironomi and microsporidia from other dipteran hosts.