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July 7, 2019

First report of cfr-encoding plasmids in the pandemic sequence type (ST) 22 methicillin-resistant Staphylococcus aureus Staphylococcal cassette chromosome mec type-IV clone.

Linezolid is often the drug of last resort for serious methicillin-resistant Staphylococcus aureus (MRSA) infections. Linezolid resistance is mediated by mutations in 23S rRNA and genes for ribosomal proteins, cfr encoding phenicol, lincosamide, oxazolidinone, pleuromutilin and streptogramin A (PhLOPSA) resistance, its homolgue cfr(B) or optrA conferring oxazolidinone and phenicol resistance. Linezolid resistance is rare in S. aureus, and cfr even rarer. This study investigated the clonality and linezolid resistance mechanisms of two MRSA isolates from patients in separate Irish hospitals. Isolates were subjected to cfr PCR, PhLOPSA susceptibility testing, 23S rRNA PCR and sequencing, DNA microarray profiling, spa typing, pulsed-field gel electrophoresis (PFGE), plasmid curing and conjugative transfer. Whole-genome sequencing was used for single nucleotide variant (SNV) analysis, multilocus-sequence typing, L-protein mutation identification, cfr-plasmid sequence analysis and optrA and cfr(B) detection. Isolates M12/0145 and M13/0401 exhibited linezolid MICs of 64 and 16 mg/liter, respectively, and harbored identical 23S rRNA and L22 mutations, but M12/0145 exhibited the mutation in 2/6 23S rRNA alleles compared to 1/5 in M13/0401. Both isolates were ST22-MRSA-IV/t032, harbored cfr, exhibited the PhLOPSA phenotype and lacked optrA and cfr(B). They differed by five PFGE bands and 603 SNVs. Isolate M12/0145 harbored cfr and fexA on a 41-kb conjugative pSCFS3-type plasmid, whereas M13/0401 harbored cfr and lsa(B) on a novel 27-kb plasmid. This is the first report of cfr in the pandemic ST22-MRSA-IV clone. Different cfr plasmids and mutations associated with linezolid resistance in genotypically distinct ST22-MRSA-IV isolates highlights that prudent management of linezolid use is essential. Copyright © 2016 Shore et al.

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July 7, 2019

Finished genome sequence and methylome of the cyanide-degrading Pseudomonas pseudoalcaligenes strain CECT5344 as resolved by single-molecule real-time sequencing.

Pseudomonas pseudoalcaligenes CECT5344 tolerates cyanide and is also able to utilize cyanide and cyano-derivatives as a nitrogen source under alkaline conditions. The strain is considered as candidate for bioremediation of habitats contaminated with cyanide-containing liquid wastes. Information on the genome sequence of the strain CECT5344 became available previously. The P. pseudoalcaligenes CECT5344 genome was now resequenced by applying the single molecule, real-time (SMRT(®)) sequencing technique developed by Pacific Biosciences. The complete and finished genome sequence of the strain consists of a 4,696,984 bp chromosome featuring a GC-content of 62.34%. Comparative analyses between the new and previous versions of the P. pseudoalcaligenes CECT5344 genome sequence revealed additional regions in the new sequence that were missed in the older version. These additional regions mostly represent mobile genetic elements. Moreover, five additional genes predicted to play a role in sulfoxide reduction are present in the newly established genome sequence. The P. pseudoalcaligenes CECT5344 genome sequence is highly related to the genome sequences of different Pseudomonas mendocina strains. Approximately, 70% of all genes are shared between P. pseudoalcaligenes and P. mendocina. In contrast to P. mendocina, putative pathogenicity genes were not identified in the P. pseudoalcaligenes CECT5344 genome. P. pseudoalcaligenes CECT5344 possesses unique genes for nitrilases and mercury resistance proteins that are of importance for survival in habitats contaminated with cyano- and mercury compounds. As an additional feature of the SMRT sequencing technology, the methylome of P. pseudoalcaligenes was established. Six sequence motifs featuring methylated adenine residues (m6A) were identified in the genome. The genome encodes several methyltransferases, some of which may be considered for methylation of the m6A motifs identified. The complete genome sequence of the strain CECT5344 now provides the basis for exploitation of genetic features for biotechnological purposes. Copyright © 2016 Elsevier B.V. All rights reserved.

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July 7, 2019

Extensive mobilome-driven genome diversification in mouse gut-associated Bacteroides vulgatus mpk.

Like many other Bacteroides species, Bacteroides vulgatus strain mpk, a mouse fecal isolate which was shown to promote intestinal homeostasis, utilizes a variety of mobile elements for genome evolution. Based on sequences collected by Pacific Biosciences SMRT sequencing technology, we discuss the challenges of assembling and studying a bacterial genome of high plasticity. Additionally, we conducted comparative genomics comparing this commensal strain with the B. vulgatus type strain ATCC 8482 as well as multiple other Bacteroides and Parabacteroides strains to reveal the most important differences and identify the unique features of B. vulgatus mpk. The genome of B. vulgatus mpk harbors a large and diverse set of mobile element proteins compared with other sequenced Bacteroides strains. We found evidence of a number of different horizontal gene transfer events and a genome landscape that has been extensively altered by different mobilization events. A CRISPR/Cas system could be identified that provides a possible mechanism for preventing the integration of invading external DNA. We propose that the high genome plasticity and the introduced genome instabilities of B. vulgatus mpk arising from the various mobilization events might play an important role not only in its adaptation to the challenging intestinal environment in general, but also in its ability to interact with the gut microbiota.

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July 7, 2019

Complete genome sequence of antibiotic and anticancer agent violacein producing Massilia sp. strain NR 4-1.

Massilia sp. NR 4-1 was a violacein producing strain newly isolated from topsoil under nutmeg tree, Torreya nucifera in Korean national monument Bijarim Forest. Violacein is a novel class of drug exhibiting anticancer and antibiotic activities originated from l-tryptophan. Here, we present the complete genome of Massilia sp. strain NR 4-1 of 6,361,416bp and total 5285 coding sequences (CDSs) including a complete violacein biosynthesis pathway, vioABCDE. The genome sequence of Massilia sp. NR 4-1 will provide stable and efficient biotechnological applications of violacein production. Copyright © 2016 Elsevier B.V. All rights reserved.

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July 7, 2019

Complete genome sequence of Enterococcus faecalis LD33, a bacteriocin-producing strain.

Enterococcus faecalis LD33 strain was originally isolated from traditional naturally fermented cream in Inner Mongolia of China. Its complete genome sequence was carried out using the Illumina Hiseq and the PacBio RSII platform. The genome only has a circular chromosome and a GC content of 37.58%. Other core information shown in the genome sequencing results further insight on this bacterium’s genetic elements for bacteriocin production and the genes related to respiratory chain. Copyright © 2016 Elsevier B.V. All rights reserved.

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July 7, 2019

Detection of translocatable units in a blaCTX-M-15 extended-spectrum ß-lactamase-producing ST131 Escherichia coli isolate using a hybrid sequencing approach.

Sir,Escherichia coli sequence type 131 (ST131) producing CTX- M-type [3-lactamases are the most common extended-spectrum [3-lactamase (ESBL)-producing strains and are of high virulence potential. In particular, the blal-;X.M.[5 gene is often encoded on a conjugative plasmid and less frequently on the chromo- some. The presence of identical bluCTX.M.[5 alleles on both the chromosome and on a plasmid in the same strain has been reported [1], suggesting transfer ofthese genes between these two locations.

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July 7, 2019

Dissemination of the mcr-1 colistin resistance gene.

Since our first report on plasmid- mediated colistin resistance gene mcr-1,1 strains previously collected in seven countries (Denmark, the Netherlands, Laos, Nigeria, Thailand, France, and the UK) have been found to carry mcr-1.2–6 Furthermore, the sequences in GenBank show that mcr-1 might also be circulating in Portugal and Malaysia. The earliest mcr-1- positive strain was collected from cattle in France in 2008 (GenBank accession number LMBK01000308). These findings confirm our initial concern that mcr-1 could have already disseminated worldwide.

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July 7, 2019

Genome sequencing and analysis of the first complete genome of Lactobacillus kunkeei strain MP2, an Apis mellifera gut isolate

Background. The honey bee (Apis mellifera) is the most important pollinator in agriculture worldwide. However, the number of honey bees has fallen significantly since 2006, becoming a huge ecological problem nowadays. The principal cause is CCD, or Colony Collapse Disorder, characterized by the seemingly spontaneous abandonment of hives by their workers. One of the characteristics of CCD in honey bees is the alteration of the bacterial communities in their gastrointestinal tract, mainly due to the decrease of Firmicutes populations, such as the Lactobacilli. At this time, the causes of these alterations remain unknown. We recently isolated a strain of Lactobacillus kunkeei (L. kunkeei strain MP2) from the gut of Chilean honey bees. L. kunkeei, is one of the most commonly isolated bacterium from the honey bee gut and is highly versatile in different ecological niches. In this study, we aimed to elucidate in detail, the L. kunkeei genetic background and perform a comparative genome analysis with other Lactobacillus species. Methods. L. kunkeei MP2 was originally isolated from the guts of Chilean A. mellifera individuals. Genome sequencing was done using Pacific Biosciences single-molecule real-time sequencing technology. De novo assembly was performed using Celera assembler. The genome was annotated using Prokka, and functional information was added using the EggNOG 3.1 database. In addition, genomic islands were predicted using IslandViewer, and pro-phage sequences using PHAST. Comparisons between L. kunkeei MP2 with other L. kunkeei, and Lactobacillus strains were done using Roary. Results. The complete genome of L. kunkeei MP2 comprises one circular chromosome of 1,614,522 nt. with a GC content of 36,9%. Pangenome analysis with 16 L. kunkeei strains, identified 113 unique genes, most of them related to phage insertions. A large and unique region of L. kunkeei MP2 genome contains several genes that encode for phage structural protein and replication components. Comparative analysis of MP2 with other Lactobacillus species, identified several unique genes of L. kunkeei MP2 related with metabolism, biofilm generation, survival under stress conditions, and mobile genetic elements (MGEs). Discussion. The presence of multiple mobile genetic elements, including phage sequences, suggest a high degree of genetic variability in L. kunkeei. Its versatility and ability to survive in different ecological niches (bee guts, flowers, fruits among others) could be given by its genetic capacity to change and adapt to different environments. L. kunkeei could be a new source of Lactobacillus with beneficial properties. Indeed, L. kunkeei MP2 could play an important role in honey bee nutrition through the synthesis of components as isoprenoids.

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July 7, 2019

Complete genome sequence of Salmonella enterica serovar Typhimurium strain SO3 (sequence type 302) isolated from a baby with meningitis in Mexico.

The complete genome of Salmonella entericaserovar Typhimurium strain SO3 (sequence type 302), isolated from a fatal meningitis infection in Mexico, was determined using PacBio technology. The chromosome hosts six complete prophages and is predicted to harbor 51 genomic islands, including 13 pathogenicity islands (SPIs). It carries the Salmonella virulence plasmid (pSTV). Copyright © 2016 Vinuesa et al.

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July 7, 2019

Genome sequence of Aeromicrobium erythreum NRRL B-3381, an erythromycin-producing bacterium of the Nocardioidaceae.

Aeromicrobium erythreumNRRL B-3381 has a 3,629,239-bp circular genome that has 72% G+C content. There are at least 3,121 coding sequences (CDSs), two rRNA gene operons, and 47 tRNAs. The genome and erythromycin (ery) biosynthetic gene sequences provide resources for metabolic and combinatorial engineering of polyketides. Copyright © 2016 Harrell and Miller.

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July 7, 2019

Mechanisms involved in acquisition of blaNDM genes by IncA/C2 and IncFIIY plasmids.

blaNDM genes confer carbapenem resistance and have been identified on transferable plasmids belonging to different incompatibility (Inc) groups. Here we present the complete sequences of four plasmids carrying a blaNDM gene, pKP1-NDM-1, pEC2-NDM-3, pECL3-NDM-1 and pEC4-NDM-6, from four clinical samples originating from four different patients. Different plasmids carry segments that align to different parts of the blaNDM region found on Acinetobacter plasmids. pKP1-NDM-1 and pEC2-NDM-3, from Klebsiella pneumoniae and Escherichia coli, respectively, were identified as type 1 IncA/C2 plasmids with almost identical backbones. Different regions carrying blaNDM are inserted in different locations in the antibiotic resistance island known as ARI-A and ISCR1 may have been involved in acquisition of blaNDM-3 by pEC2-NDM-3. pECL3-NDM-1 and pEC4-NDM-6, from Enterobacter cloacae and E. coli, respectively, have similar IncFIIY backbones but different regions carrying blaNDM are found in different locations. Tn3-derived Inverted-repeat Transposable Elements (TIME) appear to have been involved in acquisition of blaNDM-6 by pEC4-NDM-6 and the rmtC 16S rRNA methylase gene by IncFIIY plasmids. Characterisation of these plasmids further demonstrates that even very closely related plasmids may have acquired blaNDM genes by different mechanisms. These findings also illustrate the complex relationships between antimicrobial resistance genes, transposable elements and plasmids and provide insights into the possible routes for transmission of blaNDM genes amongst species of the Enterobacteriaceae family. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019

Prediction of putative resistance islands in a carbapenem-resistant Acinetobacter baumannii global clone 2 clinical isolate.

We investigated the whole genome sequence (WGS) of a carbapenem-resistant Acinetobacter baumannii isolate belonging to the global clone 2 (GC2) and predicted resistance islands using a software tool.A. baumannii strain YU-R612 was isolated from the sputum of a 61-yr-old man with sepsis. The WGS of the YU-R612 strain was obtained by using the PacBio RS II Sequencing System (Pacific Biosciences Inc., USA). Antimicrobial resistance genes and resistance islands were analyzed by using ResFinder and Genomic Island Prediction software (GIPSy), respectively.The YU-R612 genome consisted of a circular chromosome (ca. 4,075 kb) and two plasmids (ca. 74 kb and 5 kb). Its sequence type (ST) under the Oxford scheme was ST191, consistent with assignment to GC2. ResFinder analysis showed that YU-R612 possessed the following resistance genes: four ß-lactamase genes bla(ADC-30), bla(OXA-66), bla(OXA-23), and bla(TEM-1); armA, aadA1, and aacA4 as aminoglycoside resistance-encoding genes; aac(6′)Ib-cr for fluoroquinolone resistance; msr(E) for macrolide, lincosamide, and streptogramin B resistance; catB8 for phenicol resistance; and sul1 for sulfonamide resistance. By GIPSy analysis, six putative resistant islands (PRIs) were determined on the YU-R612 chromosome. Among them, PRI1 possessed two copies of Tn2009 carrying bla(OXA-23), and PRI5 carried two copies of a class I integron carrying sul1 and armA genes.By prediction of resistance islands in the carbapenem-resistant A. baumannii YU-R612 GC2 strain isolated in Korea, PRIs were detected on the chromosome that possessed Tn2009 and class I integrons. The prediction of resistance islands using software tools was useful for analysis of the WGS.

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July 7, 2019

Complete nucleotide sequence of pH11, an IncHI2 plasmid conferring multi-antibiotic resistance and multi-heavy metal resistance genes in a clinical Klebsiella pneumoniae isolate.

The complete 284,628bp sequence of pH11, an IncHI2 plasmid, was determined through single-molecule, real-time (SMRT) sequencing. Harbored by a clinical Klebsiella pneumoniae strain H11, and isolated in Beijing, this plasmid contains multiple antibiotic resistance genes, including catA2, aac(6′)-Ib, strB, strA, dfrA19, blaTEM-1, blaSHV-12, sul1, qacE delta 1, ereA, arr2, and aac3. The aac(6′)-Ib is carried by a class I integron. Plasmid pH11 also carries several genes associated with resistance to heavy metals, such as tellurium, mercury, cobalt, zinc, nickel, copper, lead and cadmium. This plasmid exhibits numerous characteristics, including HipBA and RelBE toxin-antitoxin systems, two major transfer (Tra) regions closely related to those of Salmonella enterica serovar plasmid pRH-R27, a type II restriction modification system (EcoRII R-M system), several methyltransferases and methylases and genes encoding Hha and StpA. These characteristics suggest that pH11 may adapt to various hosts and environments. Multiple insertion sequence elements, transposases, recombinases, resolvases and integrases are scattered throughout pH11. The presence of these genes may indicate that horizontal gene transfer occurs frequently in pH11 and thus may facilitate the dissemination of antimicrobial resistance determinants. Our data suggest that pH11 is a chimera gradually assembled through the integration of different horizontally acquired DNA segments via transposition or homologous recombination. Copyright © 2016 Elsevier Inc. All rights reserved.

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July 7, 2019

Complete genome sequence of Streptomyces venezuelae ATCC 15439, producer of the methymycin/pikromycin family of macrolide antibiotics, using PacBio technology.

Here, we report the complete genome sequence of Streptomyces venezuelae ATCC 15439, a producer of the methymycin/pikromycin family of macrolide antibiotics and a model host for natural product studies, obtained exclusively using PacBio sequencing technology. The 9.03-Mbp genome harbors 8,775 genes and 11 polyketide and nonribosomal peptide natural product gene clusters. Copyright © 2016 He et al.

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