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September 22, 2019

Cross-species comparison of the gut: Differential gene expression sheds light on biological differences in closely related tenebrionids.

The gut is one of the primary interfaces between an insect and its environment. Understanding gene expression profiles in the insect gut can provide insight into interactions with the environment as well as identify potential control methods for pests. We compared the expression profiles of transcripts from the gut of larval stages of two coleopteran insects, Tenebrio molitor and Tribolium castaneum. These tenebrionids have different life cycles, varying in the duration and number of larval instars. T. castaneum has a sequenced genome and has been a model for coleopterans, and we recently obtained a draft genome for T. molitor. We assembled gut transcriptome reads from each insect to their respective genomes and filtered mapped reads to RPKM>1, yielding 11,521 and 17,871 genes in the T. castaneum and T. molitor datasets, respectively. There were identical GO terms in each dataset, and enrichment analyses also identified shared GO terms. From these datasets, we compiled an ortholog list of 6907 genes; 45% of the total assembled reads from T. castaneum were found in the top 25 orthologs, but only 27% of assembled reads were found in the top 25 T. molitor orthologs. There were 2281 genes unique to T. castaneum, and 2088 predicted genes unique to T. molitor, although improvements to the T. molitor genome will likely reduce these numbers as more orthologs are identified. We highlight a few unique genes in T. castaneum or T. molitor that may relate to distinct biological functions. A large number of putative genes expressed in the larval gut with uncharacterized functions (36 and 68% from T. castaneum and T. molitor, respectively) support the need for further research. These data are the first step in building a comprehensive understanding of the physiology of the gut in tenebrionid insects, illustrating commonalities and differences that may be related to speciation and environmental adaptation. Published by Elsevier Ltd.

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September 22, 2019

Microsatellite polymorphism in the endangered snail kite reveals a panmictic, low diversity population

Genetic structure and genetic diversity are key population characteristics that can inform conservation decisions, such as delineating management units or assessing potential risks for inbreeding depression. Evidence of genetic structuring or low genetic diversity in the critically endangered snail kite (Rostrhamus sociabilis plumbeus) would have implications for monitoring and planning decisions. Recent work on understanding connectivity across the snail kite range indicated that there is less dispersal between northern and southern parts of the current range, and that dispersal is shaped by individual habitat preference. We examine whether there is neutral genetic structure and the amount of genetic variation in the population by non-lethally sampling 235 nestlings from unique nests across the entire breeding range between 2013 and 2014. Data on 15 microsatellite revealed low diversity (e.g., Na?=?2.54, He?=?0.37) and range-wide panmixia based on AMOVA, Bayesian clustering, spatial autocorrelation, isolation by distance, and spatially explicit ordination analyses. Our results emphasize that long-term recovery goals and management strategies should be based on viewing snail kites as a single genetic population, despite evidence for non-random dispersal between wetlands over ecological time scales. These results also highlight the need to understand potential effects of low genetic diversity on population dynamics and viability of snail kites. More broadly, these results add to the growing evidence for potential discrepancies between dispersal and genetic patterns, emphasizing that care should be taken if using one to interpret the other, particularly for widely-ranging species.

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September 22, 2019

Challenges of Francisella classification exemplified by an atypical clinical isolate.

The accumulation of sequenced Francisella strains has made it increasingly apparent that the 16S rRNA gene alone is not enough to stratify the Francisella genus into precise and clinically useful classifications. Continued whole-genome sequencing of isolates will provide a larger base of knowledge for targeted approaches with broad applicability. Additionally, examination of genomic information on a case-by-case basis will help resolve outstanding questions regarding strain stratification. We report the complete genome sequence of a clinical isolate, designated here as F. novicida-like strain TCH2015, acquired from the lymph node of a 6-year-old male. Two features were atypical for F. novicida: exhibition of functional oxidase activity and additional gene content, including proposed virulence determinants. These differences, which could potentially impact virulence and clinical diagnosis, emphasize the need for more comprehensive methods to profile Francisella isolates. This study highlights the value of whole-genome sequencing, which will lead to a more robust database of environmental and clinical genomes and inform strategies to improve detection and classification of Francisella strains. Copyright © 2017 Elsevier Inc. All rights reserved.

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September 22, 2019

Capnocytophaga endodontalis sp. nov., isolated from a human refractory periapical abscess.

A novel Gram-negative, capnophilic, fusiform bacterium, designated strain ChDC OS43T, was isolated from a human refractory periapical abscess in the left mandibular second molar and was characterized by polyphasic taxonomic analysis. The 16S rRNA gene sequence revealed that the strain belongs to the genus Capnocytophaga, as it showed sequence similarities to Capnocytophaga ochracea ATCC 27872T(96.30%) and C. sputigena ATCC 33612T(96.16%). The prevalent fatty acids of strain ChDC OS43Twere isoC15:0(57.54%), C16:0(5.93%), C16:03OH (5.72%), and C18:1cis 9 (4.41%). The complete genome of strain ChDC OS43Twas 3,412,686 bp, and the G+C content was 38.2 mol%. The average nucleotide identity (ANI) value between strain ChDC OS43Tand C. ochracea ATCC 27872Tor C. sputigena ATCC 33612Twas >92.01%. The genome-to-genome distance (GGD) value between strain ChDC OS43Tand C. ochracea ATCC 27872Tor C. sputigena ATCC 33612Twas 32.0 and 45.7%, respectively. Based on the results of phenotypic, chemotaxonomic, and phylogenetic analysis, strain ChDC OS43T(=?KCOM 1579T?=?KCTC 5562T?=?KCCM 42841T?=?JCM 32133T) should be classified as the type strain of a novel species of genus Capnocytophaga, for which the name Capnocytophaga endodontalis sp. nov. is proposed.

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September 22, 2019

The sequence of the salamander.

The genome of the aquatic axolotl salamander, a native of Mexico’s lakes, has yielded some surprises, and the technique used could point the way to analysis of other organisms that have complex genomes with large numbers of sequence repeats, such as the lungfish and many species of plants.

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September 22, 2019

Genomics of habitat choice and adaptive evolution in a deep-sea fish.

Intraspecific diversity promotes evolutionary change, and when partitioned among geographic regions or habitats can form the basis for speciation. Marine species live in an environment that can provide as much scope for diversification in the vertical as in the horizontal dimension. Understanding the relevant mechanisms will contribute significantly to our understanding of eco-evolutionary processes and effective biodiversity conservation. Here, we provide an annotated genome assembly for the deep-sea fish Coryphaenoides rupestris and re-sequencing data to show that differentiation at non-synonymous sites in functional loci distinguishes individuals living at different depths, independent of horizontal spatial distance. Our data indicate disruptive selection at these loci; however, we find no clear evidence for differentiation at neutral loci that may indicate assortative mating. We propose that individuals with distinct genotypes at relevant loci segregate by depth as they mature (supported by survey data), which may be associated with ecotype differentiation linked to distinct phenotypic requirements at different depths.

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September 22, 2019

Genome analysis of Fimbriiglobus ruber SP5T, a planctomycete with confirmed chitinolytic capability.

Members of the bacterial order Planctomycetales have often been observed in associations with Crustacea. The ability to degrade chitin, however, has never been reported for any of the cultured planctomycetes although utilization of N-acetylglucosamine (GlcNAc) as a sole carbon and nitrogen source is well recognized for these bacteria. Here, we demonstrate the chitinolytic capability of a member of the family Gemmataceae, Fimbriiglobus ruber SP5T, which was isolated from a peat bog. As revealed by metatranscriptomic analysis of chitin-amended peat, the pool of 16S rRNA reads from F. ruber increased in response to chitin availability. Strain SP5T displayed only weak growth on amorphous chitin as a sole source of carbon but grew well with chitin as a source of nitrogen. The genome of F. ruber SP5T is 12.364 Mb in size and is the largest among all currently determined planctomycete genomes. It encodes several enzymes putatively involved in chitin degradation, including two chitinases affiliated with the glycoside hydrolase (GH) family GH18, GH20 family ß-N-acetylglucosaminidase, and the complete set of enzymes required for utilization of GlcNAc. The gene encoding one of the predicted chitinases was expressed in Escherichia coli, and the endochitinase activity of the recombinant enzyme was confirmed. The genome also contains genes required for the assembly of type IV pili, which may be used to adhere to chitin and possibly other biopolymers. The ability to use chitin as a source of nitrogen is of special importance for planctomycetes that inhabit N-depleted ombrotrophic wetlands. IMPORTANCE Planctomycetes represent an important part of the microbial community in Sphagnum-dominated peatlands, but their potential functions in these ecosystems remain poorly understood. This study reports the presence of chitinolytic potential in one of the recently described peat-inhabiting members of the family Gemmataceae, Fimbriiglobus ruber SP5T This planctomycete uses chitin, a major constituent of fungal cell walls and exoskeletons of peat-inhabiting arthropods, as a source of nitrogen in N-depleted ombrotrophic Sphagnum-dominated peatlands. This study reports the chitin-degrading capability of representatives of the order Planctomycetales. Copyright © 2018 American Society for Microbiology.

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September 22, 2019

Characterization of Lactobacillus amylolyticus L6 as potential probiotics based on genome sequence and corresponding phenotypes

The potential of newly isolated Lactobacillus amylolyticus L6 as probiotics was investigated based on the whole genome sequence and corresponding phenotypes. With Lactobacillus acidophilus NCFM as positive control, several established methods of evaluating potential probiotics were performed on L. amylolyticus L6. The results indicated that L. amylolyticus L6 retained higher viability in human gastrointestinal (GI) tract and it also had strong inhibitory effect on pathogenic bacteria. Meanwhile, the candidate probiotics exhibited similar adhesion level as that of L. acidophilus NCFM in vitro test. As for carbohydrate utilization profile, L. amylolyticus L6 had high ability of utilizing raffinose and stachyose which were known as flatulence factors in soybean products. And this strain could also utilize starch. Besides, the mechanisms of probiotic and metabolic properties for L. amylolyticus L6 were further illustrated with the identification of related genes through the analysis of genome sequence. Therefore, we proposed that L. amylolyticus L6 have the potential to be used as probiotics from phenotypes to genotypes. And it is the first time that the complete genome sequence of L. amylolyticus L6 and the potential of this strain to be used as probiotics were reported in this study.

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September 22, 2019

Comparative genome analysis reveals a complex population structure of Legionella pneumophila subspecies.

The majority of Legionnaires’ disease (LD) cases are caused by Legionella pneumophila, a genetically heterogeneous species composed of at least 17 serogroups. Previously, it was demonstrated that L. pneumophila consists of three subspecies: pneumophila, fraseri and pascullei. During an LD outbreak investigation in 2012, we detected that representatives of both subspecies fraseri and pascullei colonized the same water system and that the outbreak-causing strain was a new member of the least represented subspecies pascullei. We used partial sequence based typing consensus patterns to mine an international database for additional representatives of fraseri and pascullei subspecies. As a result, we identified 46 sequence types (STs) belonging to subspecies fraseri and two STs belonging to subspecies pascullei. Moreover, a recent retrospective whole genome sequencing analysis of isolates from New York State LD clusters revealed the presence of a fourth L. pneumophila subspecies that we have termed raphaeli. This subspecies consists of 15 STs. Comparative analysis was conducted using the genomes of multiple members of all four L. pneumophila subspecies. Whereas each subspecies forms a distinct phylogenetic clade within the L. pneumophila species, they share more average nucleotide identity with each other than with other Legionella species. Unique genes for each subspecies were identified and could be used for rapid subspecies detection. Improved taxonomic classification of L. pneumophila strains may help identify environmental niches and virulence attributes associated with these genetically distinct subspecies. Published by Elsevier B.V.

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September 22, 2019

Genetic basis of chromosomally-encoded mcr-1 gene.

Compared with plasmid-borne mcr-1, the occurrence of chromosomally-encoded mcr-1 is rare although it has been reported in several cases. This study aimed to investigate the genetic features of chromosomally-encoded mcr-1 among Escherichia coli strains as well as the potential genetic basis governing mobilisation of mcr-1 in bacterial chromosomes. The genome sequences of 16 E. coli strains containing a chromosomal mcr-1 gene were obtained and analysed. Phylogenetic and whole-genome sequencing (WGS) analysis demonstrated that mcr-1 was associated with four major types of genetic arrangements, namely ISApl1-mcr1-orf, Tn6330, complex Tn6330 and ?Tn6330 in chromosomes of genetically unrelated E. coli strains. The mcr-1-carrying mobile elements were shown to insert into the AT-rich region, which was also the case for ISApl1. Analysis of complete E. coli genome sequences showed that there were multiple copies of ISApl1 present in E. coli chromosomes that also carried mcr-1, whilst all mcr-1-negative chromosomes were absent of any copy of ISApl1, suggesting the strong association of ISApl1 and mcr-1. Insertion of ISApl1 into E. coli chromosomes may be a prerequisite for the insertion of mcr-1-carrying mobile elements. Insertion of mcr-1 into E. coli chromosomes would enable it to become intrinsically resistant, which is expected to become more prevalent. Policy on the prudent use of colistin both in veterinary and clinical settings should be imposed globally to further prevent dissemination of mcr-1 in E. coli and other bacterial pathogens. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

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September 22, 2019

Transcriptional profiling, molecular cloning, and functional analysis of C1 inhibitor, the main regulator of the complement system in black rockfish, Sebastes schlegelii.

C1-inhibitor (C1inh) plays a crucial role in assuring homeostasis and is the central regulator of the complement activation involved in immunity and inflammation. A C1-inhibitor gene from Sebastes schlegelii was identified and designated as SsC1inh. The identified genomic DNA and cDNA sequences were 6837 bp and 2161 bp, respectively. The genomic DNA possessed 11 exons, interrupted by 10 introns. The amino acid sequence possessed two immunoglobulin-like domains and a serpin domain. Multiple sequence alignment revealed that the serpin domain of SsC1inh was highly conserved among analyzed species where the two immunoglobulin-like domains showed divergence. The distinctiveness of teleost C1inh from other homologs was indicated by the phylogenetic analysis, genomic DNA organization, and their extended N-terminal amino acid sequences. Under normal physiological conditions, SsC1inh mRNA was most expressed in the liver, followed by the gills. The involvement of SsC1inh in homeostasis was demonstrated by modulated transcription profiles in the liver and spleen upon pathogenic stress by different immune stimulants. The protease inhibitory potential of recombinant SsC1inh (rSsC1inh) and the potentiation effect of heparin on rSsC1inh was demonstrated against C1esterase and thrombin. For the first time, the anti-protease activity of the teleost C1inh against its natural substrates C1r and C1s was proved in this study. The protease assay conducted with recombinant black rockfish C1r and C1s proteins in the presence or absence of rSsC1inh showed that the activities of both proteases were significantly diminished by rSsC1inh. Taken together, results from the present study indicate that SsC1inh actively plays a significant role in maintaining homeostasis in the immune system of black rock fish. Copyright © 2018. Published by Elsevier Ltd.

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September 22, 2019

Benefit from decline: the primary transcriptome of Alteromonas macleodii str. Te101 during Trichodesmium demise.

Interactions between co-existing microorganisms deeply affect the physiology of the involved organisms and, ultimately, the function of the ecosystem as a whole. Copiotrophic Alteromonas are marine gammaproteobacteria that thrive during the late stages of phytoplankton blooms in the marine environment and in laboratory co-cultures with cyanobacteria such as Trichodesmium. The response of this heterotroph to the sometimes rapid and transient changes in nutrient supply when the phototroph crashes is not well understood. Here, we isolated and sequenced the strain Alteromonas macleodii str. Te101 from a laboratory culture of Trichodesmium erythraeum IMS101, yielding a chromosome of 4.63?Mb and a single plasmid of 237?kb. Increasing salinities to =43 ppt inhibited the growth of Trichodesmium but stimulated growth of the associated Alteromonas. We characterized the transcriptomic responses of both microorganisms and identified the complement of active transcriptional start sites in Alteromonas at single-nucleotide resolution. In replicate cultures, a similar set of genes became activated in Alteromonas when growth rates of Trichodesmium declined and mortality was high. The parallel activation of fliA, rpoS and of flagellar assembly and growth-related genes indicated that Alteromonas might have increased cell motility, growth, and multiple biosynthetic activities. Genes with the highest expression in the data set were three small RNAs (Aln1a-c) that were identified as analogs of the small RNAs CsrB-C in E. coli or RsmX-Z in pathogenic bacteria. Together with the carbon storage protein A (CsrA) homolog Te101_05290, these RNAs likely control the expression of numerous genes in responding to changes in the environment.

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September 22, 2019

Pathogen-specific binding soluble Down syndrome cell adhesion molecule (Dscam) regulates phagocytosis via membrane-bound Dscam in crab.

The Down syndrome cell adhesion molecule (Dscam) gene is an extraordinary example of diversity that can produce thousands of isoforms and has so far been found only in insects and crustaceans. Cumulative evidence indicates that Dscam may contribute to the mechanistic foundations of specific immune responses in insects. However, the mechanism and functions of Dscam in relation to pathogens and immunity remain largely unknown. In this study, we identified the genome organization and alternative Dscam exons from Chinese mitten crab, Eriocheir sinensis. These variants, designated EsDscam, potentially produce 30,600 isoforms due to three alternatively spliced immunoglobulin (Ig) domains and a transmembrane domain. EsDscam was significantly upregulated after bacterial challenge at both mRNA and protein levels. Moreover, bacterial specific EsDscam isoforms were found to bind specifically with the original bacteria to facilitate efficient clearance. Furthermore, bacteria-specific binding of soluble EsDscam via the complete Ig1-Ig4 domain significantly enhanced elimination of the original bacteria via phagocytosis by hemocytes; this function was abolished by partial Ig1-Ig4 domain truncation. Further studies showed that knockdown of membrane-bound EsDscam inhibited the ability of EsDscam with the same extracellular region to promote bacterial phagocytosis. Immunocytochemistry indicated colocalization of the soluble and membrane-bound forms of EsDscam at the hemocyte surface. Far-Western and coimmunoprecipitation assays demonstrated homotypic interactions between EsDscam isoforms. This study provides insights into a mechanism by which soluble Dscam regulates hemocyte phagocytosis via bacteria-specific binding and specific interactions with membrane-bound Dscam as a phagocytic receptor.

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September 22, 2019

Extensive gene amplification as a mechanism for piperacillin-tazobactam resistance in Escherichia coli.

Although the TEM-1 ß-lactamase (BlaTEM-1) hydrolyzes penicillins and narrow-spectrum cephalosporins, organisms expressing this enzyme are typically susceptible to ß-lactam/ß-lactamase inhibitor combinations such as piperacillin-tazobactam (TZP). However, our previous work led to the discovery of 28 clinical isolates of Escherichia coli resistant to TZP that contained only blaTEM-1 One of these isolates, E. coli 907355, was investigated further in this study. E. coli 907355 exhibited significantly higher ß-lactamase activity and BlaTEM-1 protein levels when grown in the presence of subinhibitory concentrations of TZP. A corresponding TZP-dependent increase in blaTEM-1 copy number was also observed, with as many as 113 copies of the gene detected per cell. These results suggest that TZP treatment promotes an increase in blaTEM-1 gene dosage, allowing BlaTEM-1 to reach high enough levels to overcome inactivation by the available tazobactam in the culture. To better understand the nature of the blaTEM-1 copy number proliferation, whole-genome sequence (WGS) analysis was performed on E. coli 907355 in the absence and presence of TZP. The WGS data revealed that the blaTEM-1 gene is located in a 10-kb genomic resistance module (GRM) that contains multiple resistance genes and mobile genetic elements. The GRM was found to be tandemly repeated at least 5 times within a p1ESCUM/p1ECUMN-like plasmid when bacteria were grown in the presence of TZP.IMPORTANCE Understanding how bacteria acquire resistance to antibiotics is essential for treating infected patients effectively, as well as preventing the spread of resistant organisms. In this study, a clinical isolate of E. coli was identified that dedicated more than 15% of its genome toward tandem amplification of a ~10-kb resistance module, allowing it to escape antibiotic-mediated killing. Our research is significant in that it provides one possible explanation for clinical isolates that exhibit discordant behavior when tested for antibiotic resistance by different phenotypic methods. Our research also shows that GRM amplification is difficult to detect by short-read WGS technologies. Analysis of raw long-read sequence data was required to confirm GRM amplification as a mechanism of antibiotic resistance. Copyright © 2018 Schechter et al.

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September 22, 2019

A large-scale comparative metagenomic study reveals the functional interactions in six bloom-forming Microcystis-epibiont communities.

Cyanobacterial blooms are worldwide issues of societal concern and scientific interest. Lake Taihu and Lake Dianchi, two of the largest lakes in China, have been suffering from annual Microcystis-based blooms over the past two decades. These two eutrophic lakes differ in both nutrient load and environmental parameters, where Microcystis microbiota consisting of different Microcystis morphospecies and associated bacteria (epibionts) have dominated. We conducted a comprehensive metagenomic study that analyzed species diversity, community structure, functional components, metabolic pathways and networks to investigate functional interactions among the members of six Microcystis-epibiont communities in these two lakes. Our integrated metagenomic pipeline consisted of efficient assembly, binning, annotation, and quality assurance methods that ensured high-quality genome reconstruction. This study provides a total of 68 reconstructed genomes including six complete Microcystis genomes and 28 high quality bacterial genomes of epibionts belonging to 14 distinct taxa. This metagenomic dataset constitutes the largest reference genome catalog available for genome-centric studies of the Microcystis microbiome. Epibiont community composition appears to be dynamic rather than fixed, and the functional profiles of communities were related to the environment of origin. This study demonstrates mutualistic interactions between Microcystis and epibionts at genetic and metabolic levels. Metabolic pathway reconstruction provided evidence for functional complementation in nitrogen and sulfur cycles, fatty acid catabolism, vitamin synthesis, and aromatic compound degradation among community members. Thus, bacterial social interactions within Microcystis-epibiont communities not only shape species composition, but also stabilize the communities functional profiles. These interactions appear to play an important role in environmental adaptation of Microcystis colonies.

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