Anammox enrichment culture has unexpected capabilities to biotransform azole contaminants of emerging concern.

Azoles are contaminants of emerging concern. They have a ubiquitous presence in the environment due to their wide variety of uses. This study investigated the fate of two commonly occurring azole compounds in an anammox enrichment culture. The results showed that 1H-pyrazole (PA) and 1H-1,2,4-triazole (TA) were biotransformed yielding major biotransformation products, 3-amino-1H-pyrazole and 3-amino-1H-1,2,4-triazole, respectively. Nitrate and glucose greatly stimulated the biotransformation. Under optimized conditions, 80.7% of PA and 16.4% of TA were biotransformed in an incubation period of 6 days. High molar product yield of 84.5% and 83.6% was observed per mole of PA and TA biotransformed, respectively. This novel and selective biotransformation constitutes the first report on the microbial biotransformation of PA and is amongst the very few reports on the biotransformation of TA. This study also provides evidence that anammox enrichments have unexpected capabilities to biotransform organic contaminants of emerging concern.

Molecular Dating of the Emergence of Anaerobic Rumen Fungi and the Impact of Laterally Acquired Genes.

The anaerobic gut fungi (AGF), or Neocallimastigomycota, inhabit the rumen and alimentary tract of herbivorous mammals, where they play important roles in the degradation of plant fiber. Comparative genomic and phylogenomic analyses of the AGF have long been hampered by their fastidious growth condition, as well as their large (up to 200 Mb) and AT-biased (78 to 84%) genomes. We sequenced 21 AGF transcriptomes and combined them with 5 available AGF genome sequences to explore their evolutionary relationships, time their divergence, and characterize gene gain/loss patterns associated with their evolution. We estimate that the most recent common ancestor of the AGF diverged 66 (±10) million years ago, a time frame that coincides with the evolution of grasses (Poaceae), as well as the mammalian transition from insectivory to herbivory. The concordance of independent estimations suggests that AGF have been important in shaping the success of mammalian herbivory transition by improving the efficiency of energy acquisition from recalcitrant plant materials. Comparative genomics identified multiple lineage-specific genes in the AGF, two of which were acquired from rumen gut bacteria and animal hosts via horizontal gene transfer (HGT). A third AGF domain, plant-like polysaccharide lyase, represents a novel gene in fungi that potentially aids AGF to degrade pectin. Analysis of genomic and transcriptomic sequences confirmed both the presence and expression of these lineage-specific genes in nearly all AGF clades. These genetic elements may contribute to the exceptional abilities of AGF to degrade plant biomass and enable metabolism of the rumen microbes and animal hosts.IMPORTANCE Anaerobic fungi living in the rumen of herbivorous mammals possess an extraordinary ability to degrade plant biomass. We examined the origin and genomic composition of these poorly characterized anaerobic gut fungi using both transcriptome and genomic data. Phylogenomics and molecular dating analyses found remarkable concurrence of the divergence times of the rumen fungi, the forage grasses, and the dietary shift of ancestral mammals from primarily insectivory to herbivory. Comparative genomics identified unique machinery in these fungi to utilize plant polysaccharides. The rumen fungi were also identified with the ability to code for three protein domains with putative functions in plant pectin degradation and microbial defense, which were absent from all other fungal organisms (examined over 1,000 fungal genomes). Two of these domains were likely acquired from rumen gut bacteria and animal hosts separately via horizontal gene transfer. The third one is a plant-like polysaccharide lyase, representing a unique fungal enzyme with potential pectin breakdown abilities.

Genome Sequence of a Bovine Isolate of Pasteurella multocida Strain 232.

We present here the genome sequence of Pasteurella multocida 232, a bacterium that is associated with pneumonia in humans as well as in many animal species. The genome of Pasteurella multocida 232 has an N 50 value of 187.32 kb and a total size of 2.34 Mb.

Nanopore ultra-long read sequencing technology for antimicrobial resistance detection in Mannheimia haemolytica.

Disruptive innovations in long-range, cost-effective direct template nucleic acid sequencing are transforming clinical and diagnostic medicine. A multidrug resistant strain and a pan-susceptible strain of Mannheimia haemolytica, isolated from pneumonic bovine lung samples, were sequenced at 146× and 111× coverage, respectively with Oxford Nanopore Technologies MinION. De novo assembly produced a complete genome for the non-resistant strain and a nearly complete assembly for the drug resistant strain. Functional annotation using RAST (Rapid Annotations using Subsystems Technology), CARD (Comprehensive Antibiotic Resistance Database) and ResFinder databases identified genes conferring resistance to different classes of antibiotics including ß-lactams, tetracyclines, lincosamides, phenicols, aminoglycosides, sulfonamides and macrolides. Resistance phenotypes of the M. haemolytica strains were determined by minimum inhibitory concentration (MIC) of the antibiotics. Sequencing with a highly portable MinION device corresponded to MIC assays with most of the antimicrobial resistant determinants being identified with as few as 5437 reads, except for the genes responsible for resistance to Fluoroquinolones. The resulting quality assemblies and AMR gene annotation highlight the efficiency of ultra-long read, whole-genome sequencing (WGS) as a valuable tool in diagnostic veterinary medicine.

Genome Sequence of a Spontaneous Nonhemolytic Mutant of Mannheimia haemolytica 16041065 GH.

We report here the draft genome sequence of a spontaneous nonhemolytic mutant of Mannheimia haemolytica 16041065 GH. This mutant arose during routine passage and was devoid of hemolytic activity on standard blood agars. This genome sequence had a total size of 2.7 Mb with an N50 of 117 kb.

Genome Sequence of Mannheimia haemolytica Serotype 1 Strain 16041065 BH.

Here, we report the genome sequence of Mannheimia haemolytica serotype 1 strain 16041065 BH, which was recently isolated from a Midwestern calf that died due to Mannheimia haemolytica-induced pneumonia. This genome comprised a total of 2.7 Mb, with an N50 of 122 kb, and maintained hemolytic activity when grown on blood heart infusion agar supplemented with 5% sheep's blood.

A Tissue-Mapped Axolotl De Novo Transcriptome Enables Identification of Limb Regeneration Factors.

Mammals have extremely limited regenerative capabilities; however, axolotls are profoundly regenerative and can replace entire limbs. The mechanisms underlying limb regeneration remain poorly understood, partly because the enormous and incompletely sequenced genomes of axolotls have hindered the study of genes facilitating regeneration. We assembled and annotated a de novo transcriptome using RNA-sequencing profiles for a broad spectrum of tissues that is estimated to have near-complete sequence information for 88% of axolotl genes. We devised expression analyses that identified the axolotl orthologs of cirbp and kazald1 as highly expressed and enriched in blastemas. Using morpholino anti-sense oligonucleotides, we find evidence that cirbp plays a cytoprotective role during limb regeneration whereas manipulation of kazald1 expression disrupts regeneration. Our transcriptome and annotation resources greatly complement previous transcriptomic studies and will be a valuable resource for future research in regenerative biology.

Penicillium echinulatum secretome analysis reveals the fungi potential for degradation of lignocellulosic biomass.

BACKGROUND: The enzymatic degradation of lignocellulosic materials by fungal enzyme systems has been extensively studied due to its effectiveness in the liberation of fermentable sugars for bioethanol production. Recently, variants of the fungus Penicillium echinulatum have been described as a great producer of cellulases and considered a promising strain for the bioethanol industry. RESULTS: Penicillium echinulatum, wild-type 2HH and its mutant strain S1M29, were grown on four different carbon sources: cellulose, sugar cane bagasse pretreated by steam explosion (SCB), glucose, and glycerol for 120 h. Samples collected at 24, 96, and 120 h were used for enzymatic measurement, and the 96-h one was also used for secretome analysis by 1D-PAGE LC-MS/MS. A total of 165 proteins were identified, and more than one-third of these proteins belong to CAZy families. Glycosyl hydrolases (GH) are the most abundant group, being represented in larger quantities by GH3, 5, 17, 43, and 72. Cellobiohydrolases, endoglucanases, ß-glycosidases, xylanases, ß-xylosidases, and mannanases were found, and in minor quantities, pectinases, ligninases, and amylases were also found. Swollenin and esterases were also identified. CONCLUSIONS: Our study revealed differences in the two strains of P. echinulatum in several aspects in which the mutation improved the production of enzymes related to lignocellulosic biomass deconstruction. Considering the spectral counting analysis, the mutant strain S1M29 was more efficient in the production of enzymes involved in cellulose and hemicellulose degradation, despite having a nearly identical CAZy enzymatic repertoire. Moreover, S1M29 secretes more quantities of protein on SCB than on cellulose, relevant information when considering the production of cellulases using raw materials at low cost. Glucose, and especially glycerol, were used mainly for the production of amylases and ligninases.

Draft Genome Sequence of the Thermophile Thermus filiformis ATCC 43280, Producer of Carotenoid-(Di)glucoside-Branched Fatty Acid (Di)esters and Source of Hyperthermostable Enzymes of Biotechnological Interest.

Here, we present the draft genome sequence of Thermus filiformis strain ATCC 43280, a thermophile bacterium capable of producing glycosylated carotenoids acylated with branched fatty acids and enzymes of biotechnological potential.

De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis.

De novo assembly of RNA-seq data enables researchers to study transcriptomes without the need for a genome sequence; this approach can be usefully applied, for instance, in research on 'non-model organisms' of ecological and evolutionary importance, cancer samples or the microbiome. In this protocol we describe the use of the Trinity platform for de novo transcriptome assembly from RNA-seq data in non-model organisms. We also present Trinity-supported companion utilities for downstream applications, including RSEM for transcript abundance estimation, R/Bioconductor packages for identifying differentially expressed transcripts across samples and approaches to identify protein-coding genes. In the procedure, we provide a workflow for genome-independent transcriptome analysis leveraging the Trinity platform. The software, documentation and demonstrations are freely available from http://trinityrnaseq.sourceforge.net. The run time of this protocol is highly dependent on the size and complexity of data to be analyzed. The example data set analyzed in the procedure detailed herein can be processed in less than 5 h.