Genomic diversity of burkholderia pseudomalleiIsolates, Colombia

We report an analysis of the genomic diversity of isolates of Burkholderia pseudomallei, the cause of melioidosis, recovered in Colombia from routine surveillance during 2016–2017. B. pseudomallei appears genetically diverse, suggesting it is well established and has spread across the region

Streptacidiphilus bronchialis sp. nov., a ciprofloxacin-resistant bacterium from a human clinical specimen; reclassification of Streptomyces griseoplanus as Streptacidiphilus griseoplanus comb. nov. and emended description of the genus Streptacidiphil

17 páginasThe taxonomic position of strain 15-057AT, an acidophilic actinobacterium isolated from the bronchial lavage of an 80-year-old male, was determined using a polyphasic approach incorporating morphological, phenotypic, chemotaxonomic and genomic analyses. Pairwise 16S rRNA gene sequence similarities calculated using the GGDC web server between strain 15-057AT and its closest phylogenetic neighbours, Streptomyces griseoplanus NBRC 12779T and Streptacidiphilus oryzae TH49T, were 99.7 and 97.6 %, respectively. The G+C content of isolate 15-057AT was determined to be 72.6 mol%. DNA–DNA relatedness and average nucleotide identity between isolate 15-057AT and Streptomyces griseoplanus DSM 40009T were 29.2±2.5 % and 85.97 %, respectively. Chemotaxonomic features of isolate 15-057AT were consistent with its assignment within the genus Streptacidiphilus: the whole-cell hydrolysate contained ll-diaminopimelic acid as the diagnostic diamino acid and glucose, mannose and ribose as cell-wall sugars; the major menaquinone was MK9(H8); the polar lipid profile consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, glycophospholipid, aminoglycophospholipid and an unknown lipid; the major fatty acids were anteiso-C15 : 0 and iso-C16 : 0. Phenotypic and morphological traits distinguished isolate 15-057AT from its closest phylogenetic neighbours. The results of our taxonomic analyses showed that strain 15-057AT represents a novel species within the evolutionary radiation of the genus Streptacidiphilus, for which the name Streptacidiphilus bronchialis sp. nov. is proposed. The type strain is 15-057AT (=DSM 106435T=ATCC BAA-2934T).Peer reviewe

De-MetaST-BLAST: A Tool for the Validation of Degenerate Primer Sets and Data Mining of Publicly Available Metagenomes

<div><p>Development and use of primer sets to amplify nucleic acid sequences of interest is fundamental to studies spanning many life science disciplines. As such, the validation of primer sets is essential. Several computer programs have been created to aid in the initial selection of primer sequences that may or may not require multiple nucleotide combinations (<em>i.e.,</em> degeneracies). Conversely, validation of primer specificity has remained largely unchanged for several decades, and there are currently few available programs that allows for an evaluation of primers containing degenerate nucleotide bases. To alleviate this gap, we developed the program De-MetaST that performs an <em>in silico</em> amplification using user defined nucleotide sequence dataset(s) and primer sequences that may contain degenerate bases. The program returns an output file that contains the <em>in silico</em> amplicons. When De-MetaST is paired with NCBI’s BLAST (De-MetaST-BLAST), the program also returns the top 10 nr NCBI database hits for each recovered <em>in silico</em> amplicon. While the original motivation for development of this search tool was degenerate primer validation using the wealth of nucleotide sequences available in environmental metagenome and metatranscriptome databases, this search tool has potential utility in many data mining applications.</p> </div

Flowchart outlining De-MetaST-BLAST user actions and corresponding computational processes.

<p>Fwd, Forward; Rev, Reverse; NCBI, National Center for Biotechnology Information.</p

Runtime duration of De-MetaST.

a<p>The datasets used for benchmarking were manipulations of the Waseca Farm Soil metagenome (AAFX01000000); the average sequence read length in these datasets is 1117 bp.</p

De-MetaST transformation of nucleotide sequences into a binary representation.

<p>The binary representation for each of the 16 possible nucleotide character inputs is shown in the upper box. The lower box provides an example of the transformation using a mock primer sequence. Spaced gaps are shown for instructional purposes and do not occur in the De-MetaST search routine.</p

<i>boxB</i> and 16S rRNA gene <i>in silico</i> amplicons identified in representative metagenomes using De-MetaST-BLAST.

a<p>The primers <i>boxB</i>171F (^5′ CARGGNGAYACNGARCC ^3′) and <i>boxB</i>265R (^5′ YTTNCCRTCNCKRTCNGT ^3′) were used to target an approximately 300 bp region of <i>boxB</i>.</p>b<p>Unique reads were identified using MOTHUR (v.1.27.0) <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050362#pone.0050362-Schloss1" target="_blank">[46]</a>.</p>c<p>The primers 358f (^5′CCTACGGGAGGCAGCAG^3′) and 517r (^5′ATTACCGCGGCTGCTGG^3′) <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050362#pone.0050362-Muyzer1" target="_blank">[47]</a> were used to target an approximately 190 bp amplicon in the 16S rRNA gene.</p>d<p>Average read length was estimated by dividing the database size by number of reads. The AntarcticaAquatic database is dominated by pyrosequencing derived reads (98% of all reads), while the GOS dataset is dominated by Sanger derived reads; the exact distribution for GOS reads is not available.</p

Phylogeography of Burkholderia pseudomallei Isolates, Western Hemisphere

The bacterium Burkholderia pseudomallei causes melioidosis, which is mainly associated with tropical areas. We analyzed single-nucleotide polymorphisms (SNPs) among genome sequences from isolates of B. pseudomallei that originated in the Western Hemisphere by comparing them with genome sequences of isolates that originated in the Eastern Hemisphere. Analysis indicated that isolates from the Western Hemisphere form a distinct clade, which supports the hypothesis that these isolates were derived from a constricted seeding event from Africa. Subclades have been resolved that are associated with specific regions within the Western Hemisphere and suggest that isolates might be correlated geographically with cases of melioidosis. One isolate associated with a former World War II prisoner of war was believed to represent illness 62 years after exposure in Southeast Asia. However, analysis suggested the isolate originated in Central or South America

Example of De-MetaST-BLAST output.

<p>Text within the box denotes the spreadsheet output for a <i>boxB</i> primer set search against the WASECA Farm Soil Metagenome (AAFX01000000) <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050362#pone.0050362-Tringe1" target="_blank">[41]</a> that recovers two <i>in silico</i> amplicons. Column descriptors are shown in color; select columns have been truncated due to space constraints. For the “excision info” column, the first alphanumeric character reports the “hit” number within a read (i.e. “1” indicates it is the first <i>in silico</i> amplicon found within a single read). The subsequent alphanumeric characters denote the primer orientation yielding the amplicon (F = forward, R = reverse). Whether a unique read identifier is returned is contingent upon the database itself.</p

Burkholderia thailandensis Isolated from Infected Wound, Arkansas, USA

The bacterium Burkholderia thailandensis, a member of the Burkholderia pseudomallei complex, is generally considered nonpathogenic; however, on rare occasions, B. thailandensis infections have been reported. We describe a clinical isolate of B. thailandensis, BtAR2017, recovered from a patient with an infected wound in Arkansas, USA, in 2017