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September 22, 2019

Genome sequencing and comparative analysis of Stenotrophomonas acidaminiphila reveal evolutionary insights into sulfamethoxazole resistance.

Stenotrophomonas acidaminiphila is an aerobic, glucose non-fermentative, Gram-negative bacterium that been isolated from various environmental sources, particularly aquatic ecosystems. Although resistance to multiple antimicrobial agents has been reported in S. acidaminiphila, the mechanisms are largely unknown. Here, for the first time, we report the complete genome and antimicrobial resistome analysis of a clinical isolate S. acidaminiphila SUNEO which is resistant to sulfamethoxazole. Comparative analysis among closely related strains identified common and strain-specific genes. In particular, comparison with a sulfamethoxazole-sensitive strain identified a mutation within the sulfonamide-binding site of folP in SUNEO, which may reduce the binding affinity of sulfamethoxazole. Selection pressure analysis indicated folP in SUNEO is under purifying selection, which may be owing to long-term administration of sulfonamide against Stenotrophomonas.

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September 22, 2019

The African Bullfrog (Pyxicephalus adspersus) genome unites the two ancestral ingredients for making vertebrate sex chromosomes

Heteromorphic sex chromosomes have evolved repeatedly among vertebrate lineages despite largely deleterious reductions in gene dose. Understanding how this gene dose problem is overcome is hampered by the lack of genomic information at the base of tetrapods and comparisons across the evolutionary history of vertebrates. To address this problem, we produced a chromosome-level genome assembly for the African Bullfrog (Pyxicephalus adspersus)–an amphibian with heteromorphic ZW sex chromosomes–and discovered that the Bullfrog Z is surprisingly homologous to substantial portions of the human X. Using this new reference genome, we identified ancestral synteny among the sex chromosomes of major vertebrate lineages, showing that non-mammalian sex chromosomes are strongly associated with a single vertebrate ancestral chromosome, while mammals are associated with another that displays increased haploinsufficiency. The sex chromosomes of the African Bullfrog however, share genomic blocks with both humans and non-mammalian vertebrates, connecting the two ancestral chromosome sequences that repeatedly characterize vertebrate sex chromosomes. Our results highlight the consistency of sex-linked sequences despite sex determination system lability and reveal the repeated use of two major genomic sequence blocks during vertebrate sex chromosome evolution.

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September 22, 2019

Modeling trophic dependencies and exchanges among insects’ bacterial symbionts in a host-simulated environment.

Individual organisms are linked to their communities and ecosystems via metabolic activities. Metabolic exchanges and co-dependencies have long been suggested to have a pivotal role in determining community structure. In phloem-feeding insects such metabolic interactions with bacteria enable complementation of their deprived nutrition. The phloem-feeding whitefly Bemisia tabaci (Hemiptera: Aleyrodidae) harbors an obligatory symbiotic bacterium, as well as varying combinations of facultative symbionts. This well-defined bacterial community in B. tabaci serves here as a case study for a comprehensive and systematic survey of metabolic interactions within the bacterial community and their associations with documented occurrences of bacterial combinations. We first reconstructed the metabolic networks of five common B. tabaci symbionts genera (Portiera, Rickettsia, Hamiltonella, Cardinium and Wolbachia), and then used network analysis approaches to predict: (1) species-specific metabolic capacities in a simulated bacteriocyte-like environment; (2) metabolic capacities of the corresponding species’ combinations, and (3) dependencies of each species on different media components.The predictions for metabolic capacities of the symbionts in the host environment were in general agreement with previously reported genome analyses, each focused on the single-species level. The analysis suggests several previously un-reported routes for complementary interactions and estimated the dependency of each symbiont in specific host metabolites. No clear association was detected between metabolic co-dependencies and co-occurrence patterns.The analysis generated predictions for testable hypotheses of metabolic exchanges and co-dependencies in bacterial communities and by crossing them with co-occurrence profiles, contextualized interaction patterns into a wider ecological perspective.

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September 22, 2019

Pseudomonas aeruginosa L10: A hydrocarbon-degrading, biosurfactant-producing, and plant-growth-promoting endophytic bacterium isolated from a reed (Phragmites australis).

Bacterial endophytes with the capacity to degrade petroleum hydrocarbons and promote plant growth may facilitate phytoremediation for the removal of petroleum hydrocarbons from contaminated soils. A hydrocarbon-degrading, biosurfactant-producing, and plant-growth-promoting endophytic bacterium, Pseudomonas aeruginosa L10, was isolated from the roots of a reed, Phragmites australis, in the Yellow River Delta, Shandong, China. P. aeruginosa L10 efficiently degraded C10-C26n-alkanes from diesel oil, as well as common polycyclic aromatic hydrocarbons (PAHs) such as naphthalene, phenanthrene, and pyrene. In addition, P. aeruginosa L10 could produce biosurfactant, which was confirmed by the oil spreading method, and surface tension determination of inocula. Moreover, P. aeruginosa L10 had plant growth-stimulating attributes, including siderophore and indole-3-acetic acid (IAA) release, along with 1-aminocyclopropane-1-carboxylic (ACC) deaminase activity. To explore the mechanisms underlying the phenotypic traits of endophytic P. aeruginosa L10, we sequenced its complete genome. From the genome, we identified genes related to petroleum hydrocarbon degradation, such as putative genes encoding monooxygenase, dioxygenase, alcohol dehydrogenase, and aldehyde dehydrogenase. Genome annotation revealed that P. aeruginosa L10 contained a gene cluster involved in the biosynthesis of rhamnolipids, rhlABRI, which should be responsible for the observed biosurfactant activity. We also identified two clusters of genes involved in the biosynthesis of siderophore (pvcABCD and pchABCDREFG). The genome also harbored tryptophan biosynthetic genes (trpAB, trpDC, trpE, trpF, and trpG) that are responsible for IAA synthesis. Moreover, the genome contained the ACC deaminase gene essential for ACC deaminase activity. This study will facilitate applications of endophytic P. aeruginosa L10 to phytoremediation by advancing the understanding of hydrocarbon degradation, biosurfactant synthesis, and mutualistic interactions between endophytes and host plants.

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September 22, 2019

Acquisition of resistance to carbapenem and macrolide-mediated quorum sensing inhibition by Pseudomonas aeruginosa via ICE Tn4371 6385

Pseudomonas aeruginosa can cause life-threatening infections in immunocompromised patients. The first-line agents to treat P. aeruginosa infections are carbapenems. However, the emergence of carbapenem-resistant P. aeruginosa strains greatly compromised the effec- tiveness of carbapenem treatment, which makes the surveillance on their spreading and transmission important. Here we characterized the full-length genomes of two carbapenem- resistant P. aeruginosa clinical isolates that are capable of producing New Delhi metallo-ß- lactamase-1 (NDM-1). We show that blaNDM-1 is carried by a novel integrative and conjugative element (ICE) ICETn43716385, which also carries the macrolide resistance gene msr(E) and the florfenicol resistance gene floR. By exogenously expressing msr(E) in P. aeruginosa laboratory strains, we show that Msr(E) can abolish azithromycin-mediated quorum sensing inhibition in vitro and anti-Pseudomonas effect in vivo. We conclude that ICEs are important in transmitting carbapenem resistance, and that anti-virulence treatment of P. aeruginosa infections using sub-inhibitory concentrations of macrolides can be challenged by horizontal gene transfer.

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September 22, 2019

Structure and biosynthesis of mayamycin B, a new polyketide with antibacterial activity from Streptomyces sp. 120454.

Mayamycin B, a new antibacterial type II polyketide, together with its known congener mayamycin A, were isolated from Streptomyces sp. 120454. The structure of new compound was elucidated by extensive spectroscopic analysis and comparison with literature data. Sequencing and bioinformatics analysis revealed the biosynthetic gene cluster for mayamycins A and B.

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September 22, 2019

A genome comparison of T7-like Podoviruses that infect Caulobacter crescentus.

Bacteriophages remain an understudied component of bacterial communities. Therefore, our laboratory has initiated an effort to isolate large numbers of bacteriophages that infect Caulobacter crescentus to provide an estimate of the diversity of bacteriophages that infect this common environmental bacterium. The majority of the new isolates are phicbkviruses, a genus of giant viruses that appear to be Caulobacter specific. However, we have also isolated several Podoviruses with icosahedral heads and small tails. One of these Podoviruses, designated Lullwater, is similar to two previously isolated Caulobacter phages, Cd1 and Percy. All three have genomes that are approximately 45 kb and contain approximately 30 genes. The gene order is conserved among the three genomes with one of the genes coding for a DNA polymerase that has homology to the family of T7 DNA polymerases. Phylogenetic trees based on either the DNA polymerase or the RNA polymerase amino acid sequences suggests that the three phages represent a new branch of the T7virus tree. Based on these similarities, we concluded that Cd1, Lullwater, and Percy comprise a new group in the T7virus genus.

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September 22, 2019

Co-occurrence of mcr-1 in the chromosome and on an IncHI2 plasmid: persistence of colistin resistance in Escherichia coli.

Two colistin-resistant Escherichia coli strains (FS13Z2S and FS3Z6C) possessing chromosomally encoded mcr-1 isolated from swine were characterised. Whole-genome sequencing revealed that in strain FS13Z2S mcr-1 occurred in triplicate in the chromosome with another copy encoded on a pHNSHP45-2-like IncHI2 plasmid, whereas in strain FS3Z6C only one copy mcr-1 was inserted in the chromosome. It seems likely that the triplication of chromosomal copies of mcr-1 in FS13Z2S is due to intramolecular transposition events via a composite transposon containing an mcr-1 cassette bracketed by two copies of insertion sequence ISApl1, and the pap2 gene at the insertion site was truncated by an IS1294-like element. In plasmid pFS13Z2S and the chromosome of strain FS3Z6C, only a single copy of ISApl1 was present upstream of the mcr-1 cassette. The two strains exhibited similar colistin minimum inhibitory concentrations (MICs) and featured phosphoethanolamine addition to lipid A, without regard to the copy number of mcr-1. The mcr-1-harbouring plasmid was unstable in wild-type strain FS13Z2S and was quickly lost after 7 days of passage on colistin-free Luria-Bertani broth containing 0.5% SDS, but the mcr-1 copies on the chromosome persisted. These results reveal that the single copy of mcr-1 could result in modification of lipopolysaccharide (LPS) and cause colistin resistance in E. coli. Acquisition of multiple copies of mcr-1, especially on the chromosome, would facilitate stable persistence of colistin resistance in the host strain. Copyright © 2018 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

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September 22, 2019

Coexistence of mcr-1, blaKPC-2 and two copies of fosA3 in a clinical Escherichia coli strain isolated from urine.

Here we report the first clinical Escherichia coli isolate co-harboring mcr-1, blaKPC-2 and two copies of fosA3 from China. The five plasmids of the isolate were completely sequenced and analyzed. Gene mcr-1 and blaKPC-2 were located on IncI2 and IncR plasmid, respectively. A variety of other resistance determinants such as fosA3 (two copies), blaCTX-M-123, blaOXA-1 and blaCTX-M-65 were also identified from the rest plasmids. Copyright © 2018 Elsevier B.V. All rights reserved.

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September 22, 2019

Genome-based evolutionary history of Pseudomonas spp.

Pseudomonas is a large and diverse genus of Gammaproteobacteria. To provide a framework for discovery of evolutionary and taxonomic relationships of these bacteria, we compared the genomes of type strains of 163 species and 3 additional subspecies of Pseudomonas, including 118 genomes sequenced herein. A maximum likelihood phylogeny of the 166 type strains based on protein sequences of 100 single-copy orthologous genes revealed thirteen groups of Pseudomonas, composed of two to sixty three species each. Pairwise average nucleotide identities and alignment fractions were calculated for the data set of the 166 type strains and 1224 genomes of Pseudomonas available in public databases. Results revealed that 394 of the 1224 genomes were distinct from any type strain, suggesting that the type strains represent only a fraction of the genomic diversity of the genus. The core genome of Pseudomonas was determined to contain 794 genes conferring primarily housekeeping functions. The results of this study provide a phylogenetic framework for future studies aiming to resolve the classification and phylogenetic relationships, identify new gene functions and phenotypes, and explore the ecological and metabolic potential of the Pseudomonas spp.© 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

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September 22, 2019

Insect symbionts as valuable grist for the biotechnological mill: an alkaliphilic silkworm gut bacterium for efficient lactic acid production.

Insects constitute the most abundant and diverse animal class and act as hosts to an extraordinary variety of symbiotic microorganisms. These microbes living inside the insects play critical roles in host biology and are also valuable bioresources. Enterococcus mundtii EMB156, isolated from the larval gut (gut pH >10) of the model organism Bombyx mori (Lepidoptera: Bombycidae), efficiently produces lactic acid, an important metabolite for industrial production of bioplastic materials. E. mundtii EMB156 grows well under alkaline conditions and stably converts various carbon sources into lactic acid, offering advantages in downstream fermentative processes. High-yield lactic acid production can be achieved by the strain EMB156 from renewable biomass substrates under alkaline pretreatments. Single-molecule real-time (SMRT) sequencing technology revealed its 3.01 Mbp whole genome sequence. A total of 2956 protein-coding sequences, 65 tRNA genes, and 6 rRNA operons were predicted in the EMB156 chromosome. Remarkable genomic features responsible for lactic acid fermentation included key enzymes involved in the pentose phosphate (PP)/glycolytic pathway, and an alpha amylase and xylose isomerase were characterized in EMB156. This genomic information coincides with the phenotype of E. mundtii EMB156, reflecting its metabolic flexibility in efficient lactate fermentation, and established a foundation for future biotechnological application. Interestingly, enzyme activities of amylase were quite stable in high-pH broths, indicating a possible mechanism for strong EMB156 growth in an alkaline environment, thereby facilitating lactic acid production. Together, these findings implied that valuable lactic acid-producing bacteria can be discovered efficiently by screening under the extremely alkaline conditions, as exemplified by gut microbial symbionts of Lepidoptera insects.

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September 22, 2019

The genome of Artemisia annua provides insight into the evolution of Asteraceae family and artemisinin biosynthesis.

Artemisia annua, commonly known as sweet wormwood or Qinghao, is a shrub native to China and has long been used for medicinal purposes. A. annua is now cultivated globally as the only natural source of a potent anti-malarial compound, artemisinin. Here, we report a high-quality draft assembly of the 1.74-gigabase genome of A. annua, which is highly heterozygous, rich in repetitive sequences, and contains 63 226 protein-coding genes, one of the largest numbers among the sequenced plant species. We found that, as one of a few sequenced genomes in the Asteraceae, the A. annua genome contains a large number of genes specific to this large angiosperm clade. Notably, the expansion and functional diversification of genes encoding enzymes involved in terpene biosynthesis are consistent with the evolution of the artemisinin biosynthetic pathway. We further revealed by transcriptome profiling that A. annua has evolved the sophisticated transcriptional regulatory networks underlying artemisinin biosynthesis. Based on comprehensive genomic and transcriptomic analyses we generated transgenic A. annua lines producing high levels of artemisinin, which are now ready for large-scale production and thereby will help meet the challenge of increasing global demand of artemisinin. Copyright © 2018 The Author. Published by Elsevier Inc. All rights reserved.

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September 22, 2019

Sequence analysis of IncA/C and IncI1 plasmids isolated from multidrug-resistant Salmonella Newport using Single-Molecule Real-Time Sequencing.

Multidrug-resistant (MDR) plasmids play an important role in disseminating antimicrobial resistance genes. To elucidate the antimicrobial resistance gene compositions in A/C incompatibility complex (IncA/C) plasmids carried by animal-derived MDR Salmonella Newport, and to investigate the spread mechanism of IncA/C plasmids, this study characterizes the complete nucleotide sequences of IncA/C plasmids by comparative analysis. Complete nucleotide sequencing of plasmids and chromosomes of six MDR Salmonella Newport strains was performed using PacBio RSII. Open reading frames were assigned using prokaryotic genome annotation pipeline (PGAP). To understand genomic diversity and evolutionary relationships among Salmonella Newport IncA/C plasmids, we included three complete IncA/C plasmid sequences with similar backbones from Salmonella Newport and Escherichia coli: pSN254, pAM04528, and peH4H, and additional 200 draft chromosomes. With the exception of canine isolate CVM22462, which contained an additional IncI1 plasmid, each of the six MDR Salmonella Newport strains contained only the IncA/C plasmid. These IncA/C plasmids (including references) ranged in size from 80.1 (pCVM21538) to 176.5?kb (pSN254) and carried various resistance genes. Resistance genes floR, tetA, tetR, strA, strB, sul, and mer were identified in all IncA/C plasmids. Additionally, blaCMY-2 and sugE were present in all IncA/C plasmids, excepting pCVM21538. Plasmid pCVM22462 was capable of being transferred by conjugation. The IncI1 plasmid pCVM22462b in CVM22462 carried blaCMY-2 and sugE. Our data showed that MDR Salmonella Newport strains carrying similar IncA/C plasmids clustered together in the phylogenetic tree using chromosome sequences and the IncA/C plasmids from animal-derived Salmonella Newport contained diverse resistance genes. In the current study, we analyzed genomic diversities and phylogenetic relationships among MDR Salmonella Newport using complete plasmids and chromosome sequences and provided possible spread mechanism of IncA/C plasmids in Salmonella Newport Lineage II.

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September 22, 2019

Catabolism of 2-hydroxypyridine by Burkholderia sp. MAK1: a five-gene cluster encoded 2-hydroxypyridine 5-monooxygenase HpdABCDE catalyses the first step of biodegradation.

Microbial degradation of 2-hydroxypyridine usually results in the formation of a blue pigment (nicotine blue). In contrast, the Burkholderia sp. strain MAK1 bacterium utilizes 2-hydroxypyridine without the accumulation of nicotine blue. This scarcely investigated degradation pathway presumably employs 2-hydroxypyridine 5-monooxygenase, an elusive enzyme that has been hypothesized but has yet to be identified or characterized. The isolation of the mutant strain Burkholderia sp. MAK1 ?P5 that is unable to utilize 2-hydroxypyridine has led to the identification of a gene cluster (designated hpd) which is responsible for the degradation of 2-hydroxypyridine. The activity of 2-hydroxypyridine 5-monooxygenase has been assigned to a soluble diiron monooxygenase (SDIMO) encoded by a five-gene cluster (hpdA, hpdB, hpdC, hpdD, and hpdE). A 4.5-kb DNA fragment containing all five genes has been successfully expressed in Burkholderia sp. MAK1 ?P5 cells. We have proved that the recombinant HpdABCDE protein catalyzes the enzymatic turnover of 2-hydroxypyridine to 2,5-dihydroxypyridine. Moreover, we have confirmed that emerging 2,5-dihydroxypyridine is a substrate for HpdF, an enzyme similar to 2,5-dihydroxypyridine 5,6-dioxygenases that are involved in the catabolic pathways of nicotine and nicotinic acid. The proteins and genes identified in this study have allowed the identification of a novel degradation pathway of 2-hydroxypyridine. Our results provide a better understanding of the biodegradation of pyridine derivatives in nature. Also, the discovered 2-hydroxypyridine 5-monooxygenase may be an attractive catalyst for the regioselective synthesis of various N-heterocyclic compounds.IMPORTANCE The degradation pathway of 2-hydroxypyridine without the accumulation of a blue pigment is relatively unexplored, as, to our knowledge, no genetic data related to this process have ever been presented. In this paper, we describe genes and enzymes involved in this little-studied catabolic pathway. This work provides new insights into the metabolism of 2-hydroxypyridine in nature. A broad-range substrate specificity of 2-hydroxypyridine 5-monooxygenase, a key enzyme in the degradation, makes this biocatalyst attractive for the regioselective hydroxylation of pyridine derivatives. Copyright © 2018 American Society for Microbiology.

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September 22, 2019

The presence of colistin resistance gene mcr-1 and -3 in ESBL producing Escherichia coli isolated from food in Ho Chi Minh City, Vietnam.

Colistin is indicated for the treatment of multidrug-resistant gram-negative bacterial infections. However, the spread of colistin-resistant bacteria harbouring an mcr gene has become a serious concern. This study investigated local foods in Vietnam for contamination with colistin-resistant bacteria. A total of 261 extended-spectrum ß-lactamase (ESBL)- and AmpC-producing Escherichia coli isolates from 330 meat and seafood products were analysed for colistin susceptibility and the presence of mcr genes. Approximately, 24% (62/261) of ESBL- or AmpC-producing E. coli isolates showed colistin resistance; 97% (60/62) of colistin-resistant isolates harboured mcr-1, whereas 3% (2/62) harboured mcr-3. As the result of plasmid analysis of two strains, both plasmids harbouring mcr-3 revealed that plasmid replicon type was IncFII. Sequencing analysis indicated that an insertion sequence was present near mcr-3, suggesting that IncFII plasmids harbouring mcr-3 could be transferred to other bacterial species by horizontal transfer of the plasmid or transfer with some insertion sequence. In conclusion, ESBL-producing E. coli and AmpC-producing E. coli have acquired colistin resistance because 24% of such isolates show colistin resistance and 3% of the colistin-resistant strains harbour mcr-3. We reported the present of the mcr-3-carrying ESBL-producing E. coli isolated from pork in Vietnam.

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