Spiroplasma Bacteria Enhance Survival of Drosophila hydei Attacked by the Parasitic Wasp Leptopilina heterotoma

Maternally-transmitted associations between endosymbiotic bacteria and insects are ubiquitous. While many of these associations are obligate and mutually beneficial, many are facultative, and the mechanism(s) by which these microbes persist in their host lineages remain elusive. Inherited microbes with imperfect transmission are expected to be lost from their host lineages if no other mechanisms increase their persistence (i.e., host reproductive manipulation and/or fitness benefits to host). Indeed numerous facultative heritable endosymbionts are reproductive manipulators. Nevertheless, many do not manipulate reproduction, so they are expected to confer fitness benefits to their hosts, as has been shown in several studies that report defense against natural enemies, tolerance to environmental stress, and increased fecundity.We examined whether larval to adult survival of Drosophila hydei against attack by a common parasitoid wasp (Leptopilina heterotoma), differed between uninfected flies and flies that were artificially infected with Spiroplasma, a heritable endosymbiont of Drosophila hydei that does not appear to manipulate host reproduction. Survival was significantly greater for Spiroplasma-infected flies, and the effect of Spiroplasma infection was most evident during the host's pupal stage. We examined whether or not increased survival of Spiroplasma-infected flies was due to reduced oviposition by the wasp (i.e., pre-oviposition mechanism). The number of wasp eggs per fly larva did not differ significantly between Spiroplasma-free and Spiroplasma-infected fly larvae, suggesting that differential fly survival is due to a post-oviposition mechanism.Our results suggest that Spiroplasma confers protection to D. hydei against wasp parasitism. This is to our knowledge the first report of a potential defensive mutualism in the genus Spiroplasma. Whether it explains the persistence and high abundance of this strain in natural populations of D. hydei, as well as the widespread distribution of heritable Spiroplasma in Drosophila and other arthropods, remains to be investigated

A One-Step Real-Time Multiplex PCR for Screening Y-Chromosomal Microdeletions without Downstream Amplicon Size Analysis

BACKGROUND: Y-chromosomal microdeletions (YCMD) are one of the major genetic causes for non-obstructive azoospermia. Genetic testing for YCMD by multiplex polymerase chain reaction (PCR) is an established method for quick and robust screening of deletions in the AZF regions of the Y-chromosome. Multiplex PCRs have the advantage of including a control gene in every reaction and significantly reducing the number of reactions needed to screen the relevant genomic markers. PRINCIPAL FINDINGS: The widely established "EAA/EMQN best practice guidelines for molecular diagnosis of Y-chromosomal microdeletions (2004)" were used as a basis for designing a real-time multiplex PCR system, in which the YCMD can simply be identified by their melting points. For this reason, some AZF primers were substituted by primers for regions in their genomic proximity, and the ZFX/ZFY control primer was exchanged by the AMELX/AMELY control primer. Furthermore, we substituted the classical SybrGreen I dye by the novel and high-performing DNA-binding dye EvaGreen™ and put substantial effort in titrating the primer combinations in respect to optimal melting peak separation and peak size. SIGNIFICANCE: With these changes, we were able to develop a platform-independent and robust real-time based multiplex PCR, which makes the need for amplicon identification by electrophoretic sizing expendable. By using an open-source system for real-time PCR analysis, we further demonstrate the applicability of automated melting point and YCMD detection

A transcriptomic analysis of Echinococcus granulosus larval stages:implications for parasite biology and host adaptation

The cestode Echinococcus granulosus--the agent of cystic echinococcosis, a zoonosis affecting humans and domestic animals worldwide--is an excellent model for the study of host-parasite cross-talk that interfaces with two mammalian hosts. To develop the molecular analysis of these interactions, we carried out an EST survey of E. granulosus larval stages. We report the salient features of this study with a focus on genes reflecting physiological adaptations of different parasite stages.We generated ~10,000 ESTs from two sets of full-length enriched libraries (derived from oligo-capped and trans-spliced cDNAs) prepared with three parasite materials: hydatid cyst wall, larval worms (protoscoleces), and pepsin/H(+)-activated protoscoleces. The ESTs were clustered into 2700 distinct gene products. In the context of the biology of E. granulosus, our analyses reveal: (i) a diverse group of abundant long non-protein coding transcripts showing homology to a middle repetitive element (EgBRep) that could either be active molecular species or represent precursors of small RNAs (like piRNAs); (ii) an up-regulation of fermentative pathways in the tissue of the cyst wall; (iii) highly expressed thiol- and selenol-dependent antioxidant enzyme targets of thioredoxin glutathione reductase, the functional hub of redox metabolism in parasitic flatworms; (iv) candidate apomucins for the external layer of the tissue-dwelling hydatid cyst, a mucin-rich structure that is critical for survival in the intermediate host; (v) a set of tetraspanins, a protein family that appears to have expanded in the cestode lineage; and (vi) a set of platyhelminth-specific gene products that may offer targets for novel pan-platyhelminth drug development.This survey has greatly increased the quality and the quantity of the molecular information on E. granulosus and constitutes a valuable resource for gene prediction on the parasite genome and for further genomic and proteomic analyses focused on cestodes and platyhelminths