Splenic gene expression profiling in White Leghorn layer inoculated with the Salmonella enterica serovar Enteritidis

Salmonella enterica serovar Enteritidis (SE) is a foodborne pathogen that can threaten human health through contaminated poultry products. Live poultry, chicken eggs and meat are primary sources of human salmonellosis. To understand the genetic resistance of egg‐type chickens in response to SE inoculation, global gene expression in the spleen of 20‐week‐old White Leghorn was measured using the Agilent 4 × 44 K chicken microarray at 7 and 14 days following SE inoculation (dpi). Results showed that there were 1363 genes significantly differentially expressed between inoculated and non‐inoculated groups at 7 dpi (I7/N7), of which 682 were up‐regulated and 681 were down‐regulated genes. By contrast, 688 differentially expressed genes were observed at 14 dpi (I14/N14), of which 371 were up‐regulated genes and 317 were down‐regulated genes. There were 33 and 28 immune‐related genes significantly differentially expressed in the comparisons of I7/N7 and I14/N14 respectively. Functional annotation revealed that several Gene Ontology (GO) terms related to immunity were significantly enriched between the inoculated and non‐inoculated groups at 14 dpi but not at 7 dpi, despite a similar number of immune‐related genes identified between I7/N7 and I14/N14. The immune response to SE inoculation changes with different time points following SE inoculation. The complicated interaction between the immune system and metabolism contributes to the immune responses to SE inoculation of egg‐type chickens at 14 dpi at the onset of lay. GC, TNFSF8, CD86, CD274, BLB1 and BLB2 play important roles in response to SE inoculation. The results from this study will deepen the current understanding of the genetic response of the egg‐type chicken to SE inoculation at the onset of egg laying.     

Distribution and function of prophage phiRv1 and phiRv2 among Mycobacterium tuberculosis complex

Mycobacterium tuberculosis complex (MTBC) is notorious for causing diseases, such as tuberculosis. Tuberculosis caused by M. tuberculosis remains a global public health concern. Two prophages, phiRv1 and phiRv2, can be found among most MTBC genomes. However, no precise functions have been assigned for the two prophages. In this paper, to find out the function of these two prophages, the distribution and function of phiRv1 and phiRv2 in MTBC genomes were analyzed from multiple omics data. We found that complex insertion, deletion, and reorganization appeared on the locus of two prophages in MTBC genomes; some genes of the two prophages can be translated and are functional from proteomic data; the expression of other prophage genes, such as Rv1577c, Rv2650c, Rv2652c, Rv2659c, and Rv2658c, can vary with environmental stresses and might enhance the fitness of MTBC. These data will facilitate our in-depth understanding of their function.     

The tRNA A76 Hydroxyl Groups Control Partitioning of the tRNA-dependent Pre- and Post-transfer Editing Pathways in Class I tRNA Synthetase

Aminoacyl-tRNA synthetases catalyze ATP-dependent covalent coupling of cognate amino acids and tRNAs for ribosomal protein synthesis. Escherichia coli isoleucyl-tRNA synthetase (IleRS) exploits both the tRNA-dependent pre- and post-transfer editing pathways to minimize errors in translation. However, the molecular mechanisms by which tRNA^Ile organizes the synthetic site to enhance pre-transfer editing, an idiosyncratic feature of IleRS, remains elusive. Here we show that tRNA^Ile affects both the synthetic and editing reactions localized within the IleRS synthetic site. In a complex with cognate tRNA, IleRS exhibits a 10-fold faster aminoacyl-AMP hydrolysis and a 10-fold drop in amino acid affinity relative to the free enzyme. Remarkably, the specificity against non-cognate valine was not improved by the presence of tRNA in either of these processes. Instead, amino acid specificity is determined by the protein component per se, whereas the tRNA promotes catalytic performance of the synthetic site, bringing about less error-prone and kinetically optimized isoleucyl-tRNA^Ile synthesis under cellular conditions. Finally, the extent to which tRNA^Ile modulates activation and pre-transfer editing is independent of the intactness of its 3′-end. This finding decouples aminoacylation and pre-transfer editing within the IleRS synthetic site and further demonstrates that the A76 hydroxyl groups participate in post-transfer editing only. The data are consistent with a model whereby the 3′-end of the tRNA remains free to sample different positions within the IleRS·tRNA complex, whereas the fine-tuning of the synthetic site is attained via conformational rearrangement of the enzyme through the interactions with the remaining parts of the tRNA body.     

Fitness Is Strongly Influenced by Rare Mutations of Large Effect in a Microbial Mutation Accumulation Experiment

Our understanding of the evolutionary consequences of mutation relies heavily on estimates of the rate and fitness effect of spontaneous mutations generated by mutation accumulation (MA) experiments. We performed a classic MA experiment in which frequent sampling of MA lines was combined with whole genome resequencing to develop a high-resolution picture of the effect of spontaneous mutations in a hypermutator (ΔmutS) strain of the bacterium Pseudomonas aeruginosa. After ∼644 generations of mutation accumulation, MA lines had accumulated an average of 118 mutations, and we found that average fitness across all lines decayed linearly over time. Detailed analyses of the dynamics of fitness change in individual lines revealed that a large fraction of the total decay in fitness (42.3%) was attributable to the fixation of rare, highly deleterious mutations (comprising only 0.5% of fixed mutations). Furthermore, we found that at least 0.64% of mutations were beneficial and probably fixed due to positive selection. The majority of mutations that fixed (82.4%) were base substitutions and we failed to find any signatures of selection on nonsynonymous or intergenic mutations. Short indels made up a much smaller fraction of the mutations that were fixed (17.4%), but we found evidence of strong selection against indels that caused frameshift mutations in coding regions. These results help to quantify the amount of natural selection present in microbial MA experiments and demonstrate that changes in fitness are strongly influenced by rare mutations of large effect.     

R-Loop Mediated trans Action of the APOLO Long Noncoding RNA

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Tropical terrestrial invertebrates—Where to from here?

There are over one million described invertebrate species on Earth, the majority of which are likely to inhabit the highly biodiverse rain forests around the equator. These are some of the most vulnerable ecosystems on Earth due to the pressures of deforestation and climate change with many of their inhabitants at risk of extinction. Invertebrates play a major role in ecosystem functioning from decomposition and nutrient cycling to herbivory and pollination; however, while our understanding of these roles is improving, we are far from being able to predict the consequences of further deforestation, climate change, and biodiversity loss due to the lack of comparative data and the high proportion of species which remain to be discovered. As we move into an era of increased pressure on old-growth habitats and biodiversity, it is imperative that we understand how changes to invertebrate communities, and the extinction of species, affect ecosystems. Innovative and comprehensive methods that approach these issues are needed. Here, we highlight priorities for future tropical terrestrial invertebrate research such as the efficiency of sustainable land management, exploration of innovative methods for better understanding of invertebrate ecology and behavior, and quantifying the role of invertebrates in ecosystem functioning.     

The mitochondrial genome of Gatzara jezoensis (Neuroptera: Myrmeleontidae) and phylogenetic analysis of Neuroptera

Phylogenetic relationship within Neuroptera is controversial, particularly for the various hypotheses based on both morphological and molecular evidence. In the present study, we determined the complete mitochondrial genome (mitogenome) of Gatzara jezoensis, which is the second representative of the tribe Dendroleontini. The G. jezoensis mitogenome contained the conserved set of 37 mitochondrial genes and a putative control region, with a conserved gene arrangement which was similar to that of most sequenced neuropteran mitogenomes. All transfer RNAs exhibited the canonical cloverleaf secondary structure, except for trnS^(AGN). The control region contained two conserved elements (ploy-T stretch and ATGGTTCAAYAAAATAAYYCYCTC motif) and abundant microsatellite-like elements. The phylogenetic analysis of sequenced neuropteran mitogenomes using the concatenated protein-coding genes (PCGs) and ribosomal genes recovered the monophyly of Myrmeleontidae, which revealed this dataset could generate the more robust phylogeny of Neuroptera than that of 13 PCGs dataset.     

Enhancing plasmid transformation efficiency and enabling CRISPR-Cas9/Cpf1-based genome editing in Clostridium tyrobutyricum

Clostridium tyrobutyricum ATCC 25755 is known as a natural hyper-butyrate producer with great potentials as an excellent platform to be engineered for valuable biochemical production from renewable resources. However, limited transformation efficiency and the lack of genetic manipulation tools have hampered the broader applications of this micro-organism. In this study, the effects of Type I restriction-modification system and native plasmid on conjugation efficiency of C. tyrobutyricum were investigated through gene deletion. The deletion of Type I restriction endonuclease resulted in a 3.7-fold increase in conjugation efficiency, while the additional elimination of the native plasmid further enhanced conjugation efficiency to 6.05 ± 0.75 × 10³ CFU/ml-donor, which was 15.3-fold higher than the wild-type strain. Fermentation results indicated that the deletion of those two genetic elements did not significantly influence the end-products production in the resultant mutant ΔRMIΔNP. Thanks to the increased conjugation efficiency, the CRISPR-Cas9/Cpf1 systems, which previously could not be implemented in C. tyrobutyricum, were successfully employed for genome editing in ΔRMIΔNP with an efficiency of 12.5–25%. Altogether, approaches we developed herein offer valuable guidance for establishing efficient DNA transformation methods in nonmodel micro-organisms. The ΔRMIΔNP mutant can serve as a great chassis to be engineered for diverse valuable biofuel and biochemical production.