Identification of Mycobacterium pseudoshottsii in the Eastern Mediterranean.

Among the numerous pathogenic nontuberculous mycobacteria (NTM), which may cause disease in both poikilothermic and homoeothermic organisms, members of the unique clade Mycobacterium ulcerans/Mycobacterium marinum (MuMC) may cause disease in both fish and humans. Here, we describe the emergence of Mycobacterium pseudoshottsii, one of the four MuMC members, in Israel. For many years, M. marinum was the dominant NTM that was diagnosed in Israel as a fish pathogen. To the best of our knowledge, this is the first isolation and genomic characterization of M. pseudoshottsii infecting edible fish from two different fish species farmed in offshore sea cages in the eastern Mediterranean as well as in a recirculating aquaculture system in Israel. We compared the M. pseudoshottsii whole-genome sequences to all available genomic sequences of MuMC in free, publicly accessible databases. IMPORTANCE Mycobacterium pseudoshottsii was first detected in 1997 in the USA, infecting wild striped bass (Morone saxatilis). Since then, several reports from different countries worldwide have shown its capacity to become established in new regions as well as its pathogenicity to saltwater and euryhaline finfish of different genera. Our phylogenetic analysis revealed that the Mycobacterium ulcerans/Mycobacterium marinum clade (MuMC) is divided into two main branches: one that includes M. marinum and M. pseudoshottsii, and the second, which includes other M. marinum isolates as well as two isolates of M. shottsii. Our results reinforce the proposition that the geographical distribution of M. pseudoshottsii is much more extensive than is commonly believed. The emergence of M. pseudoshottsii in different parts of the world and its pathogenic traits that affect finfish of different genera may be a cause for concern among fish farmers, researchers, and environmental organizations.

Infection by the Parasite Myxobolus bejeranoi (Cnidaria: Myxozoa) Suppresses the Immune System of Hybrid Tilapia.

Myxozoa (Cnidaria) is a large group of microscopic obligate endoparasites that can cause emerging diseases, affecting wild fish populations and fisheries. Recently, the myxozoan Myxobolus bejeranoi was found to infect the gills of hybrid tilapia (Nile tilapia (Oreochromis niloticus) × Jordan/blue tilapia (O. aureus)), causing high morbidity and mortality. Here, we used comparative transcriptomics to elucidate the molecular processes occurring in the fish host following infection by M. bejeranoi. Fish were exposed to pond water containing actinospores for 24 h and the effects of minor, intermediate, and severe infections on the sporulation site, the gills, and on the hematopoietic organs, head kidney and spleen, were compared. Enrichment analysis for GO and KEGG pathways indicated immune system activation in gills at severe infection, whereas in the head kidney a broad immune suppression included deactivation of cytokines and GATA3 transcription factor responsible for T helper cell differentiation. In the spleen, the cytotoxic effector proteins perforin and granzyme B were downregulated and insulin, which may function as an immunomodulatory hormone inducing systemic immune suppression, was upregulated. These findings suggest that M. bejeranoi is a highly efficient parasite that disables the defense mechanisms of its fish host hybrid tilapia.

Comparative study of population genomic approaches for mapping colony-level traits.

Social insect colonies exhibit colony-level phenotypes such as social immunity and task coordination, which are produced by the individual phenotypes. Mapping the genetic basis of such phenotypes requires associating the colony-level phenotype with the genotypes in the colony. In this paper, we examine alternative approaches to DNA extraction, library construction, and sequencing for genome wide association studies (GWAS) of colony-level traits using a population sample of Cataglyphis niger ants. We evaluate the accuracy of allele frequency estimation from sequencing a pool of individuals (pool-seq) from each colony using either whole-genome sequencing or reduced representation genomic sequencing. Based on empirical measurement of the experimental noise in sequenced DNA pools, we show that reduced representation pool-seq is drastically less accurate than whole-genome pool-seq. Surprisingly, normalized pooling of samples did not result in greater accuracy than un-normalized pooling. Subsequently, we evaluate the power of the alternative approaches for detecting quantitative trait loci (QTL) of colony-level traits by using simulations that account for an environmental effect on the phenotype. Our results can inform experimental designs and enable optimizing the power of GWAS depending on budget, availability of samples and research goals. We conclude that for a given budget, sequencing un-normalized pools of individuals from each colony provides optimal QTL detection power.

A comparative analysis of methods for de novo assembly of hymenopteran genomes using either haploid or diploid samples.

Diverse invertebrate taxa including all 200,000 species of Hymenoptera (ants, bees, wasps and sawflies) have a haplodiploid sex determination system, where females are diploid and males are haploid. Thus, hymenopteran genome projects can make use of DNA from a single haploid male sample, which is assumed advantageous for genome assembly. For the purpose of gene annotation, transcriptome sequencing is usually conducted using RNA from a pool of individuals. We conducted a comparative analysis of genome and transcriptome assembly and annotation methods, using genetic sources of different ploidy: (1) DNA from a haploid male or a diploid female (2) RNA from the same haploid male or a pool of individuals. We predicted that the use of a haploid male as opposed to a diploid female will simplify the genome assembly and gene annotation thanks to the lack of heterozygosity. Using DNA and RNA from the same haploid individual is expected to provide better confidence in transcript-to-genome alignment, and improve the annotation of gene structure in terms of the exon/intron boundaries. The haploid genome assemblies proved to be more contiguous, with both contig and scaffold N50 size at least threefold greater than their diploid counterparts. Completeness evaluation showed mixed results. The SOAPdenovo2 diploid assembly was missing more genes than the haploid assembly. The SPAdes diploid assembly had more complete genes, but a higher level of duplicates, and a greatly overestimated genome size. When aligning the two transcriptomes against the male genome, the male transcriptome gave 2-3% more complete transcripts than the pool transcriptome for genes with comparable expression levels in both transcriptomes. However, this advantage disappears in the final results of the gene annotation pipeline that incorporates evidence from homologous proteins. The RNA pool is still required to obtain the full transcriptome with genes that are expressed in other life stages and castes. In conclusion, the use of a haploid source material for a de novo genome project provides a substantial advantage to the quality of the genome draft and the use of RNA from the same haploid individual for transcriptome to genome alignment provides a minor advantage for genes that are expressed in the adult male.

Gene duplication in trypanosomatids - two DED1 paralogs are functionally redundant and differentially expressed during the life cycle.

DED1/VAS belong to the DEAD-box family of RNA helicases that are associated with translation initiation in higher eukaryotes. Here we report on two DED1/VAS homologs that were identified in the genome of Leishmania. The two paralogs include all the domains that are typical of DEAD-box proteins and a phylogenetic analysis suggests that their duplication predates the branching of DED1 and VAS, which took place along with the appearance of early metazoans. The two Leishmania DED1 paralogs complement a yeast strain that fails to express the endogenous DED1, suggesting that they are responsible for a similar function. This is also supported by RNAi-mediated silencing experiments performed in Trypanosoma brucei. The two proteins are functionally redundant, since defects in protein synthesis and cell growth arrest were observed only when both paralogs were eliminated. A partial stage-specific specialization is observed, as LeishDED1-2 is more abundant in promastigotes, whereas expression of LeishDED1-1 increases in amastigotes. Duplication of an essential gene usually offers a safety net against mutations but in this case it also generated two proteins with stage specific expression.

Cryo-electron tomography: gaining insight into cellular processes by structural approaches.

Visualization of cellular processes at a resolution of the individual protein should involve integrative and complementary approaches that can eventually draw realistic functional and cellular landscapes. Electron tomography of vitrified but otherwise unaltered cells emerges as a central method for three-dimensional reconstruction of cellular architecture at a resolution of 2-6 nm. While a combination of correlative light-based microscopy with cryo-electron tomography (cryo-ET) provides medium-resolution insight into pivotal cellular processes, fitting high-resolution structural approaches, for example, X-ray crystallography, into reconstructed macromolecular assemblies provides unprecedented information on native protein assemblies. Thus, cryo-ET bridges the resolution gap between cellular and structural biology. In this article, we focus on the study of eukaryotic cells and macromolecular complexes in a close-to-life-state. We discuss recent developments and structural findings enabling major strides to be made in understanding complex physiological functions.