Andreas Linde

Vairimorpha disparis n. comb. (Microsporidia: Burenellidae): a redescription and taxonomic revision of Thelohania disparis Timofejeva 1956, a microsporidian parasite of the gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae).

ABSTRACT. Investigation of pathogens of populations of the gypsy moth, Lymantria dispar (L.) in Central and Eastern Europe revealed the existence of a microsporidium (Fungi: Microsporidia) of the genus Vairimorpha. The parasite produced three spore morphotypes. Internally infective spores are formed in the gut and adjacent muscle and connective tissue; single diplokaryotic spores and monokaryotic spores grouped by eight in sporophorous vesicles develop in the fat body tissues. The small subunit rDNA gene sequences of various isolates of the Vairimorpha microsporidia, obtained from L. dispar in various habitats in the investigated region, revealed their mutual identity. In phylogenetic analyses, the organism clustered with other L. dispar microsporidia that form only diplokaryotic spores in the sporogony cycle. The octospores of certain microsporidia infecting Lepidoptera that were previously described as Thelohania spp., have recently been shown to be one of the several spore morphotypes produced by species in the genus Vairimorpha. Because the description and drawings of a parasite described as Thelohania disparis by Timofejeva fit the characteristics of Vairimorpha, and all octospore‐producing microsporidia collected from L. dispar since 1985 are genetically identical Vairimorpha species, it is believed that the parasite characterized here is identical to T. disparis Timofejeva 1956, and is herein redescribed, characterized, and transferred to the genus Vairimorpha as the new combination Vairimorpha disparis n. comb.

Monitoring the establishment and prevalence of the fungal entomopathogen Entomophaga maimaiga in two Lymantria dispar L. populations in Bulgaria

The establishment and prevalence of the entomopathogenic fungus Entomophaga maimaiga, (Zygomycetes, Entomophthorales), introduced into two gypsy moth populations in Bulgaria, was monitored during 2000–2004. In the Karlovo Region population, where E. maimaiga was introduced in 1999, the fungus was recovered every year and the prevalence of infection varied from 6.1% to 15.9%. A microsporidian infection caused by an Endoreticulatus sp. (Protista, Microsporidia) was also recorded every year and the prevalence varied from 2.1% and 5.0%. In the Svoge Region population, where E. maimaiga was introduced in late 2000 and first found in 2002, the prevalence of infection varied from 8.8% to 13.8%. Larval parasitism caused by Cotesia melanoscela, Protapanteles liparidis (Hymenoptera, Braconidae) and species of tachinids (Diptera, Tachinidae) was also recorded in the larvae of both populations. We provide a rationale as to why other countries should consider introducing E. maimaiga for biological control of Lymantria dispar populations.

Pathogens of bark beetles (Curculionidae: Scolytinae) and other beetles in Bulgaria

The presence and diversity of beetle pathogens associated with different tree species in Bulgaria was investigated. In total, 818 specimens belonging to 22 beetle species were examined. Pathogens occurred in 9 host species. The infections were found in the gut (virus, nematodes, protozoans, microsporidia) and haemolymph (nematodes) of the infected insects. The following pathogen species: ItEPV, Beauveria bassiana, B. brongniartii, Isaria farinosa, Gregarina typographi, Gregarina spp., Chytridiopsis typographi, Chytridiopsis sp., and nematodes were revealed. Insects with mycoses were mummified and filled up with fungal structures, formed pseudosclerotium. Most pathogens were established in bark beetles (Scolytinae). For the first time, the fungus B. bassiana was reported in Tomicus piniperda, Orthotomicus longicollis, O. erosus, X. spinole, Taphrorychus villifrons and Phylobius sp. in Bulgaria. We also present the first records of gregarines in O. longicollis, Acanthocinus aedilis, Rhagium inquisitor, Pyrochroa coccinea, and of the microsporidium Chytridiopsis sp. in O. longicollis.

Pathogens of bark beetles (Coleoptera: Curculionidae) in Bulgarian forests

The occurrence and prevalence of bark beetle pathogens in forest stands in Bulgaria were investigated in 944 specimens belonging to 21 bark beetle species. Protozoa, microsporidia, fungi and nematodes occurred in 19 of all investigated species. The infections were found in the gut (nematodes, gregarines, microsporidia), gonads (microsporidia) and hemolymph (nematodes) of the infected insects. Protozoan species (Gregarina typographi, Gregarina spp.) were detected in eight bark beetle species. Morphometric data about G. typographi and Gregarina spp. are presented. The prevalence of the gregarines varied between 1.4% and 64.2%. Microsporidia of the genera Nosema and Chytridiopsis were revealed in three bark beetle species. The prevalence of microsporidia ranged between 1.5% and 11.8%. This is the first report of a microsporidium in Taphrorychus villifrons and of gregarines in T. villifrons, Pityogenes bistridentatus, P. conjunctus, and Orthotomicus erosus. The fungus Beauveria bassiana was found in 3.4% of Hylurgops palliatus specimens. Nematodes (in gut and haemolymph) were revealed in 19 bark beetle species and their prevalence varied between 10% and 98.5%.

First record of Entomophaga maimaiga (Entomophthorales: Entomophthoraceae) in Georgia

Abstract In 2005, high levels of mortality occurred in an outbreak of the gypsy moth population in Georgia. Resting spores typical of entomophthoralean fungi were found within larval cadavers and molecular analyses confirmed that the pathogen was Entomophaga maimaiga. This is the first record of this entomopathogen in Georgia and in this part of Europe.

Review of the genus Endoreticulatus (Microsporidia, Encephalitozoonidae) with description of a new species isolated from the grasshopper Poecilimon thoracicus (Orthoptera: Tettigoniidae) and transfer of Microsporidium itiiti Malone to the genus

The historic genus Pleistophora (Plistophora) is a highly polyphyletic clade with invertebrate Microsporidia reassigned to several new genera since the 1980s. Two genera, Endoreticulatus and Cystosporogenes, clearly separate into distinct but closely related clades based on small subunit ribosomal RNA analysis but are included in different families that are each polyphyletic. A microsporidium with morphology resembling the Endoreticulatus/Cystosporogenes clade was isolated from the grasshopper Poecilimon thoracicus from a site in Northwest Bulgaria. It produced intense infections in the digestive tract of the host but no behavioral changes were noted in infected individuals. Prevalence of the microsporidium increased over the active feeding season yearly. Mature spores were oval and measured 2.58±0.21 μm×1.34±0.24 μm, with 16 to approximately 32 spores in a parasitophorous vacuole. The spores were uninucleate and polar filament coils numbered 8-9 situated in a single row. The spore polaroplast consisted of an anterior lamellar section and a posterior vesicular section, and the posterior vacuole was reduced. Analyses of a 1221 bp partial SSU-rRNA sequence indicated that the isolate is more closely related to the Endoreticulatus clade than to Cystosporogenes, but shows earlier phylogenetic separation from species infecting Lepidoptera and represents a new species, Endoreticulatus poecilimonae. To compare sequences of Endoreticulatus spp. from Lepidoptera to those infecting other insect orders, an isolate, Microsporidium itiitiMalone (1985), described from the Argentine stem weevil, Listronotus bonariensis, was sequenced. Like the grasshopper isolate, the weevil isolate is closely related but basal to the lepidopteran Endoreticulatus clade. The original description combined with the new sequence data confirms species status and permits transfer of the isolate from Microsporidium, a genus erected for microsporidian species of uncertain taxonomic status, to Endoreticulatus.

Ultrastructure, characteristic features and occurrence of Nosema leptinotarsae Lipa, 1968, a microsporidian pathogen of Leptinotarsa decemlineata (Coleoptera, Chrysomelidae)

This paper presents the ultrastructure and characteristic features of Nosema leptinotarsae infecting L. decemlineata collected in Trabzon (Turkey). Fresh spores are oval, 4.69 ± 0.19 μm in length and 2.43 ± 0.12 μm in width (n = 50). The spore wall is thick and measures 180 to 250 nm and consists of a clear endospore (130 to 170 nm) and an electron-dense, uniform exospore (50 to 80 nm). The polar filament is isofilar and has 15–16 coils. The well-developed polaroplast has a lamellated structure with thin lamellae. The ultrastructural features of Nosema leptinotarsae differ from those of other microsporidia infecting chrysomelids.

The First Record of Nucleopolyhedrovirus Isolated from the Gypsy Moth, Lymantria dispar (Lepidoptera, Lymantriidae) in Turkey

OBJECTIVE The gypsy moth, Lymantria dispar L. (Lepidoptera, Lymantriidae) is a common pest of forests and fruit trees throughout the world. This insect is also a major serious pest in Turkey. Nowadays L. dispar can be managed by biological control methods especially, using entomopathogenic viruses. The aim of this study is to characterize entomopathogenic viruses and is the first record of nucleopolyhedrovirus isolated from the gypsy moth in Turkey. METHODS PIBs obtained from infected larvae were measured and photographed using an Olympus BX51 microscope with a DP-25 digital camera and a DP2-BSW Soft Imaging System and examined with a Philips 208 electron microscope (TEM). RESULTS The virus had the typical characteristics of nucleopolyhedroviruses. The dimension of the polyhedral inclusion bodies (PIBs) was 2.03±0.25 µm. PIBs varied in size from 1.65 to 2.21 µm and were usually polygonal in shape. Virions in PIBs contained 1 to 8 nucleocapsids per virion. The size of the viral particles was 366.67±54.72 (312-500) x 42.95±6.12 (30-47) nm. CONCLUSION The isolation and characterization of a pure isolate of Lymantria dispar multinucleopolyhedrovirus (LdMNPV-TR) from Turkey is presented for the first time.

Two different and sublethal isolates of Nosema lymantriae (Microsporidia) reduce the reproductive success of their host, Lymantria dispar

Abstract We examined the effects of two microsporidian isolates of Nosema lymantriae (Germany isolate; Schweinfurt isolate) on the reproductive success of Lymantria dispar L. All possible mating combinations were tested. Both isolates affected the fecundity of infected females and the hatch of neonates. The infection of female L. dispar with either isolate resulted in a higher proportion of non-viable eggs; the survival of neonates during egg stage was not affected. When L. dispar larvae were infected with N. lymantriae [Germany] the number of eggs per egg mass decreased between 24 and 61%. When both adults were infected, the hatch rate decreased to 26%. While the infection of the male or the female host with the Germany isolate affected the number of eggs per egg mass and the hatch of progeny, we did not find a significant effect when male hosts were infected with the Schweinfurt isolate; only infection of the female L. dispar resulted in a reduction of the number of eggs per egg mass between 26 and 37%.

Nosema pieriae sp. n. (Microsporida, Nosematidae): A New Microsporidian Pathogen of the Cabbage Butterfly Pieris brassicae L. (Lepidoptera: Pieridae)

A new microsporidian pathogen of the cabbage butterfly, Pieris brassicae is described based on light microscopy, ultrastructural characteristics and comparative small subunit rDNA analysis. The pathogen infects the gut of P. brassicae. All development stages are in direct contact with the host cell cytoplasm. Meronts are spherical or ovoid. Spherical meronts measure 3.68 ± 0.73 × 3.32 ± 1.09 µm and ovoid meronts 4.04 ± 0.74 × 2.63 ± 0.49 µm. Sporonts are spherical to elongate (4.52 ± 0.48 × 2.16 ± 0.27 µm). Sporoblasts are elongated and measure 4.67 ± 0.60 × 2.30 ± 0.30 µm in length. Fresh spores with nuclei arranged in a diplokaryon are oval and measure 5.29 ± 0.55 µm in length and 2.31 ± 0.29 µm in width. Spores stained with Giemsas stain measure 4.21 ± 0.50 µm in length and 1.91 ± 0.24 µm in width. Spores have an isofilar polar filament with six coils. All morphological, ultrastructural and molecular features indicate that the described microsporidium belongs to the genus Nosema and confirm that it has different taxonomic characters than other microsporidia infecting Pieris spp.