Is the Insect Cuticle the only Entry Gate for Fungal Infection? Insights into Alternative Modes of Action of Entomopathogenic Fungi.

Entomopathogenic fungi are the only insect pathogens able to infect their host by adhesion to the surface and penetration through the cuticle. Although the possibility of fungal infection per os was described almost a century ago, there is an information gap of several decades regarding this topic, which was poorly explored due to the continuous elucidation of cuticular infection processes that lead to insect death by mycosis. Recently, with the advent of next-generation sequencing technologies, the genomes of the main entomopathogenic fungi became available, and many fungal genes potentially useful for oral infection were described. Among the entomopathogenic Hypocreales that have been sequenced, Beauveria bassiana (Balsamo-Crivelli) Vuillemin (Cordycipitaceae) is the main candidate to explore this pathway since it has a major number of shared genes with other non-fungal pathogens that infect orally, such as Bacillus thuringiensis Berliner (Bacillales: Bacillaceae). This finding gives B. bassiana a potential advantage over other entomopathogenic fungi: the possibility to infect through both routes, oral and cuticular. In this review, we explore all known entry gates for entomopathogenic fungi, with emphasis on the infection per os. We also set out the fungal infection process in a more integral approach, as a need to exploit its full potential for insect control, considering all of its virulence factors and the conditions needed to improve its virulence against insect that might offer some resistance to the common infection through the cuticle.

The limpet transcription factors of Triatoma infestans regulate the response to fungal infection and modulate the expression pattern of defensin genes.

As part of the innate humoral response to microbial attack, insects activate the expression of antimicrobial peptides (AMP). Understanding the regulatory mechanisms of this response in the Chagas disease vector Triatoma infestans is important since biological control strategies against pyrethroid-resistant insect populations were recently addressed by using the entomopathogenic fungus Beauveria bassiana. By bioinformatics, gene expression, and silencing techniques in T. infestans nymphs, we achieved sequence and functional characterization of two variants of the limpet transcription factor (Tilimpet) and studied their role as regulators of the AMP expression, particularly defensins, in fungus-infected insects. We found that Tilimpet variants may act differentially since they have divergent sequences and different relative expression ratios, suggesting that Tilimpet-2 could be the main regulator of the higher expressed defensins and Tilimpet-1 might play a complementary or more general role. Also, the six defensins (Tidef-1 to Tidef-6) exhibited different expression levels in fungus-infected nymphs, consistent with their phylogenetic clustering. This study aims to contribute to a better understanding of T. infestans immune response in which limpet is involved, after challenge by B. bassiana infection.

Tracing the coevolution between Triatoma infestans and its fungal pathogen Beauveria bassiana.

The chemical control of Triatoma infestans, the major Chagas disease vector in southern South America, has been threatened in the last years by the emergence of pyrethroid-resistant bug populations. As an alternative approach, the efficacy of the entomopathogenic fungus Beauveria bassiana to control T. infestans populations (regardless their pyrethroid susceptibility) has been demonstrated. Growing research efforts on the interaction between T. infestans and B. bassiana by molecular, ecological, biochemical and behavioral traits has allowed framing such interaction as an evolutionary arms race. This review will focus on the relationships established in this particular host-pathogen system, compiling available data on the relevance of fungal pathogenesis, insect behavior, population dynamics and human intervention to favor fungal dissemination in bug populations. The current snapshot shows the fungus ahead in the evolutionary arms race and predicts a promissory landscape for the biological control of Chagas disease vectors.

Transcriptome profiling of Diachasmimorpha longicaudata towards useful molecular tools for population management.

BACKGROUND: Diachasmimorpha longicaudata (Hymenoptera: Braconidae) is a solitary parasitoid of Tephritidae (Diptera) fruit flies of economic importance currently being mass-reared in bio-factories and successfully used worldwide. A peculiar biological aspect of Hymenoptera is its haplo-diploid life cycle, where females (diploid) develop from fertilized eggs and males (haploid) from unfertilized eggs. Diploid males were described in many species and recently evidenced in D. longicaudata by mean of inbreeding studies. Sex determination in this parasitoid is based on the Complementary Sex Determination (CSD) system, with alleles from at least one locus involved in early steps of this pathway. Since limited information is available about genetics of this parasitoid species, a deeper analysis on D. longicaudata's genomics is required to provide molecular tools for achieving a more cost effective production under artificial rearing conditions. RESULTS: We report here the first transcriptome analysis of male-larvae, adult females and adult males of D. longicaudata using 454-pyrosequencing. A total of 469766 reads were analyzed and 8483 high-quality isotigs were assembled. After functional annotation, a total of 51686 unigenes were produced, from which, 7021 isotigs and 20227 singletons had at least one BLAST hit against the NCBI non-redundant protein database. A preliminary comparison of adult female and male evidenced that 98 transcripts showed differential expression profiles, with at least a 10-fold difference. Among the functionally annotated transcripts we detected four sequences potentially involved in sex determination and three homologues to two known genes involved in the sex determination cascade. Finally, a total of 4674SimpleSequence Repeats (SSRs) were in silico identified and characterized. CONCLUSION: The information obtained here will significantly contribute to the development of D. longicaudata functional genomics, genetics and population-based genome studies. Thousands of new microsatellite markers were identified as toolkits for population genetics analysis. The transcriptome characterized here is the starting point to elucidate the molecular bases of the sex determination mechanism in this species.

Genetic diversity of Burkholderia contaminans isolates from cystic fibrosis patients in Argentina.

A total of 120 Burkholderia cepacia complex isolates collected during 2004-2010 from 66 patients in two cystic fibrosis reference centers in Argentina were analyzed. Burkholderia contaminans was the species most frequently recovered (57.6%), followed by Burkholderia cenocepacia (15%), a species distribution not reported so far. The recA-PCR-based techniques applied to the B. contaminans isolates revealed that 85% of the population carried the recA-ST-71 allele. Our results showed the utility of BOX-PCR genotyping in analyzing B. contaminans diversity. This approach allowed us to address clonal transmission during an outbreak and the genetic changes occurring in infecting bacteria over the course of chronic infection.