Division of the genus Chryseobacterium: Observation of discontinuities in amino acid identity values, a possible consequence of major extinction events, guides transfer of nine species to the genus Epilithonimonas, eleven species to the genus Kaistella, and three species to the genus Halpernia gen. nov., with description of Kaistella daneshvariae sp. nov. and Epilithonimonas vandammei sp. nov. derived from clinical specimens.

The genus Chryseobacterium in the family Weeksellaceae is known to be polyphyletic. Amino acid identity (AAI) values were calculated from whole-genome sequences of species of the genus Chryseobacterium, and their distribution was found to be multi-modal. These naturally-occurring non-continuities were leveraged to standardise genus assignment of these species. We speculate that this multi-modal distribution is a consequence of loss of biodiversity during major extinction events, leading to the concept that a bacterial genus corresponds to a set of species that diversified since the Permian extinction. Transfer of nine species (Chryseobacterium arachidiradicis, Chryseobacterium bovis, Chryseobacterium caeni, Chryseobacterium hispanicum, Chryseobacterium hominis, Chryseobacterium hungaricum,, Chryseobacterium pallidum and Chryseobacterium zeae) to the genus Epilithonimonas and eleven (Chryseobacterium anthropi, Chryseobacterium antarcticum, Chryseobacterium carnis, Chryseobacterium chaponense, Chryseobacterium haifense, Chryseobacterium jeonii, Chryseobacterium montanum, Chryseobacterium palustre, Chryseobacterium solincola, Chryseobacterium treverense and Chryseobacterium yonginense) to the genus Kaistella is proposed. Two novel species are described: Kaistella daneshvariae sp. nov. and Epilithonimonas vandammei sp. nov. Evidence is presented to support the assignment of Planobacterium taklimakanense to a genus apart from Chryseobacterium, to which Planobacterium salipaludis comb nov. also belongs. The novel genus Halpernia is proposed, to contain the type species Halpernia frigidisoli comb. nov., along with Halpernia humi comb. nov., and Halpernia marina comb. nov.

A Real-Time Multiplex PCR Assay for Detection of Elizabethkingia Species and Differentiation between Elizabethkingia anophelis and E. meningoseptica.

Nosocomial infections of Elizabethkingia species can have fatal outcomes if not identified and treated properly. The current diagnostic tools available require culture and isolation, which can extend the reporting time and delay treatment. Using comparative genomics, we developed an efficient multiplex real-time PCR for the simultaneous detection of all known species of Elizabethkingia, as well as differentiating the two most commonly reported species, Elizabethkingia anophelis and Elizabethkingia meningoseptica.

Revisiting the taxonomy of the genus Elizabethkingia using whole-genome sequencing, optical mapping, and MALDI-TOF, along with proposal of three novel Elizabethkingia species: Elizabethkingia bruuniana sp. nov., Elizabethkingia ursingii sp. nov., and Elizabethkingia occulta sp. nov.

The genus Elizabethkingia is genetically heterogeneous, and the phenotypic similarities between recognized species pose challenges in correct identification of clinically derived isolates. In addition to the type species Elizabethkingia meningoseptica, and more recently proposed Elizabethkingia miricola, Elizabethkingia anophelis and Elizabethkingia endophytica, four genomospecies have long been recognized. By comparing historic DNA-DNA hybridization results with whole genome sequences, optical maps, and MALDI-TOF mass spectra on a large and diverse set of strains, we propose a comprehensive taxonomic revision of this genus. Genomospecies 1 and 2 contain the type strains E. anophelis and E. miricola, respectively. Genomospecies 3 and 4 are herein proposed as novel species named as Elizabethkingia bruuniana sp. nov. (type strain, G0146T = DSM 2975T = CCUG 69503T = CIP 111191T) and Elizabethkingia ursingii sp. nov. (type strain, G4122T = DSM 2974T = CCUG 69496T = CIP 111192T), respectively. Finally, the new species Elizabethkingia occulta sp. nov. (type strain G4070T = DSM 2976T = CCUG 69505T = CIP 111193T), is proposed.

A Genetic Locus in Elizabethkingia anophelis Associated with Elevated Vancomycin Resistance and Multiple Antibiotic Reduced Susceptibility.

The Gram-negative Elizabethkingia express multiple antibiotic resistance and cause severe opportunistic infections. Vancomycin is commonly used to treat Gram-positive infections and has also been used to treat Elizabethkingia infections, even though Gram-negative organisms possess a vancomycin permeability barrier. Elizabethkingia anophelis appeared relatively vancomycin-susceptible and challenge with this drug led to morphological changes indicating cell lysis. In stark contrast, vancomycin growth challenge revealed that E. anophelis populations refractory to vancomycin emerged. In addition, E. anophelis vancomycin-selected mutants arose at high frequencies and demonstrated elevated vancomycin resistance and reduced susceptibility to other antimicrobials. All mutants possessed a SNP in a gene (vsr1 = vancomycin-susceptibility regulator 1) encoding a PadR family transcriptional regulator located in the putative operon vsr1-ORF551, which is conserved in other Elizabethkingia spp as well. This is the first report linking a padR homologue (vsr1) to antimicrobial resistance in a Gram-negative organism. We provide evidence to support that vsr1 acts as a negative regulator of vsr1-ORF551 and that vsr1-ORF551 upregulation is observed in vancomycin-selected mutants. Vancomycin-selected mutants also demonstrated reduced cell length indicating that cell wall synthesis is affected. ORF551 is a membrane-spanning protein with a small phage shock protein conserved domain. We hypothesize that since vancomycin-resistance is a function of membrane permeability in Gram-negative organisms, it is likely that the antimicrobial resistance mechanism in the vancomycin-selected mutants involves altered drug permeability.

Neural network based integration of assays to assess pathogenic potential.

Limited data significantly hinders our capability of biothreat assessment of novel bacterial strains. Integration of data from additional sources that can provide context about the strain can address this challenge. Datasets from different sources, however, are generated with a specific objective and which makes integration challenging. Here, we developed a deep learning-based approach called the neural network embedding model (NNEM) that integrates data from conventional assays designed to classify species with new assays that interrogate hallmarks of pathogenicity for biothreat assessment. We used a dataset of metabolic characteristics from a de-identified set of known bacterial strains that the Special Bacteriology Reference Laboratory (SBRL) of the Centers for Disease Control and Prevention (CDC) has curated for use in species identification. The NNEM transformed results from SBRL assays into vectors to supplement unrelated pathogenicity assays from de-identified microbes. The enrichment resulted in a significant improvement in accuracy of 9% for biothreat. Importantly, the dataset used in our analysis is large, but noisy. Therefore, the performance of our system is expected to improve as additional types of pathogenicity assays are developed and deployed. The proposed NNEM strategy thus provides a generalizable framework for enrichment of datasets with previously collected assays indicative of species.

Reevaluation of the taxonomic status of recently described species of Enterococcus: evidence that E. thailandicus is a senior subjective synonym of "E. sanguinicola" and confirmation of E. caccae as a species distinct from E. silesiacus.

Several of the more recently proposed new species of Enterococcus are nearly identical based on 16S rRNA gene sequence analysis and phenotypic traits. In the present study, DNA-DNA reassociation experiments, in conjunction with sequencing of the 16S rRNA and rpoB genes, provided evidence that "Enterococcus sanguinicola" and Enterococcus thailandicus actually represent the same species. In contrast, Enterococcus caccae and Enterococcus silesiacus, two other species with nearly identical 16S rRNA gene sequences, were confirmed to be separate species.

Draft Genome Sequence of Kroppenstedtia sanguinis X0209T, a Clinical Isolate Recovered from Human Blood.

Kroppenstedtia sanguinis X0209T, a thermoactinomycete, was isolated from the blood of a patient in Sweden. We report on the draft genome sequence obtained with an Illumina MiSeq instrument. The assembled genome totaled 3.73 Mb and encoded 3,583 proteins. Putative genes for virulence, transposons, and biosynthetic gene clusters have been identified.

The draft genomes of Elizabethkingia anophelis of equine origin are genetically similar to three isolates from human clinical specimens.

We report the isolation and characterization of two Elizabethkingia anophelis strains (OSUVM-1 and OSUVM-2) isolated from sources associated with horses in Oklahoma. Both strains appeared susceptible to fluoroquinolones and demonstrated high MICs to all cell wall active antimicrobials including vancomycin, along with aminoglycosides, fusidic acid, chloramphenicol, and tetracycline. Typical of the Elizabethkingia, both draft genomes contained multiple copies of ß-lactamase genes as well as genes predicted to function in antimicrobial efflux. Phylogenetic analysis of the draft genomes revealed that OSUVM-1 and OSUVM-2 differ by only 6 SNPs and are in a clade with 3 strains of Elizabethkingia anophelis that were responsible for human infections. These findings therefore raise the possibility that Elizabethkingia might have the potential to move between humans and animals in a manner similar to known zoonotic pathogens.

Identification of a novel small-molecule inhibitor of the hypoxia-inducible factor 1 pathway.

Hypoxia-inducible factor 1 (HIF-1) is the central mediator of cellular responses to low oxygen and has recently become an important therapeutic target for solid tumor therapy. Inhibition of HIF-1 is expected to result in the attenuation of hypoxia-inducible genes, which are vital to many aspects of tumor biology, including adaptative responses for survival under anaerobic conditions. To identify small molecules inhibiting the HIF-1 pathway, we did a biological screen on a 10,000-membered natural product-like combinatorial library. The compounds of the library, which share a 2,2-dimethylbenzopyran structural motif, were tested for their ability to inhibit the hypoxic activation of an alkaline phosphatase reporter gene under the control of hypoxia-responsive elements in human glioma cells. This effort led to the discovery of 103D5R, a novel small-molecule inhibitor of HIF-1alpha. 103D5R markedly decreased HIF-1alpha protein levels induced by hypoxia or cobaltous ions in a dose- and time-dependent manner, whereas minimally affecting global cellular protein expression levels, including that of control proteins such as HIF-1beta, IkappaBalpha, and beta-actin. The inhibitory activity of 103D5R against HIF-1alpha was clearly shown under normoxia and hypoxia in cells derived from different cancer types, including glioma, prostate, and breast cancers. This inhibition prevented the activation of HIF-1 target genes under hypoxia such as vascular endothelial growth factor (VEGF) and glucose transporter-1 (Glut-1). Investigations into the molecular mechanism showed that 103D5R strongly reduced HIF-1alpha protein synthesis, whereas HIF-1alpha mRNA levels and HIF-1alpha degradation were not affected. 103D5R inhibited the phosphorylation of Akt, Erk1/2, and stress-activated protein kinase/c-jun-NH(2)-kinase, without changing the total levels of these proteins. Further studies on the mechanism of action of 103D5R will likely provide new insights into its validity/applicability for the pharmacologic targeting of HIF-1alpha for therapeutic purposes.

Twelve Complete Reference Genomes of Clinical Isolates in the Capnocytophaga Genus.

We report here 1 near-complete genome sequence and 12 complete genome sequences for clinical Capnocytophaga isolates. Total read coverages ranged from 211× to 737×, and genome sizes ranged from 2.41 Mb to 3.10 Mb. These genomes will enable a more comprehensive taxonomic evaluation of the Capnocytophaga genus.

Complete Circularized Genome Sequences of Four Strains of Elizabethkingia anophelis, Including Two Novel Strains Isolated from Wild-Caught Anopheles sinensis.

We provide complete circularized genome sequences of two mosquito-derived Elizabethkingia anophelis strains with draft sequences currently in the public domain (R26 and Ag1), and two novel E. anophelis strains derived from a different mosquito species, Anopheles sinensis (AR4-6 and AR6-8). The genetic similarity of all four mosquito-derived strains is remarkable.

Evolutionary dynamics and genomic features of the Elizabethkingia anophelis 2015 to 2016 Wisconsin outbreak strain.

An atypically large outbreak of Elizabethkingia anophelis infections occurred in Wisconsin. Here we show that it was caused by a single strain with thirteen characteristic genomic regions. Strikingly, the outbreak isolates show an accelerated evolutionary rate and an atypical mutational spectrum. Six phylogenetic sub-clusters with distinctive temporal and geographic dynamics are revealed, and their last common ancestor existed approximately one year before the first recognized human infection. Unlike other E. anophelis, the outbreak strain had a disrupted DNA repair mutY gene caused by insertion of an integrative and conjugative element. This genomic change probably contributed to the high evolutionary rate of the outbreak strain and may have increased its adaptability, as many mutations in protein-coding genes occurred during the outbreak. This unique discovery of an outbreak caused by a naturally occurring mutator bacterial pathogen provides a dramatic example of the potential impact of pathogen evolutionary dynamics on infectious disease epidemiology.

Draft Genome Sequences of Strains Representing Each of the Elizabethkingia Genomospecies Previously Determined by DNA-DNA Hybridization.

Draft genome sequences of Elizabethkingia meningoseptica and representatives of each of its four historically described genomospecies were sequenced here. Preliminary analysis suggests that Elizabethkingia miricola belongs to genomospecies 2, and both Elizabethkingia anophelis and Elizabethkingia endophytica are most similar to genomospecies 1.

Complete Genome Sequences of Four Strains from the 2015-2016 Elizabethkingia anophelis Outbreak.

The complete circularized genome sequences of selected specimens from the largest known Elizabethkingia anophelis outbreak to date are described here. Genomic rearrangements observed among the outbreak strains are discussed.

Functional evolution of ADAMTS genes: evidence from analyses of phylogeny and gene organization.

BACKGROUND: The ADAMTS (A Disintegrin-like and Metalloprotease with Thrombospondin motifs) proteins are a family of metalloproteases with sequence similarity to the ADAM proteases, that contain the thrombospondin type 1 sequence repeat motifs (TSRs) common to extracellular matrix proteins. ADAMTS proteins have recently gained attention with the discovery of their role in a variety of diseases, including tissue and blood disorders, cancer, osteoarthritis, Alzheimer's and the genetic syndromes Weill-Marchesani syndrome (ADAMTS10), thrombotic thrombocytopenic purpura (ADAMTS13), and Ehlers-Danlos syndrome type VIIC (ADAMTS2) in humans and belted white-spotting mutation in mice (ADAMTS20). RESULTS: Phylogenetic analysis and comparison of the exon/intron organization of vertebrate (Homo, Mus, Fugu), chordate (Ciona) and invertebrate (Drosophila and Caenorhabditis) ADAMTS homologs has elucidated the evolutionary relationships of this important gene family, which comprises 19 members in humans. CONCLUSIONS: The evolutionary history of ADAMTS genes in vertebrate genomes has been marked by rampant gene duplication, including a retrotransposition that gave rise to a distinct ADAMTS subfamily (ADAMTS1, -4, -5, -8, -15) that may have distinct aggrecanase and angiogenesis functions.

Complete Genome Sequences for Two Strains of a Novel Fastidious, Partially Acid-Fast, Gram-Positive Corynebacterineae Bacterium, Derived from Human Clinical Samples.

Here we report the complete genome sequences of two strains of the novel fastidious, partially acid-fast, Gram-positive bacillus "Lawsonella clevelandensis" (proposed). Their clinical relevance and unusual growth characteristics make them intriguing candidates for whole-genome sequencing.

Complete Genome Sequence of Strain H5989 of a Novel Devosia Species.

The CDC Special Bacteriology Reference Laboratory (SBRL) collection of human clinical pathogens contains several strains from the genus Devosia, usually found environmentally. We provide here the complete genome of strain H5989, which was isolated from a human cerebrospinal fluid (CSF) specimen and represents a putative novel species in the genus Devosia.

Genome Sequences of Oblitimonas alkaliphila gen. nov. sp. nov. (Proposed), a Novel Bacterium of the Pseudomonadaceae Family.

Results obtained through 16S rRNA gene sequencing and phenotypic testing of eight related, but unidentified, isolates located in a historical collection at the Centers for Disease Control and Prevention suggested that these isolates belong to a novel genera of bacteria. The genomes of the bacteria, to be named Oblitimonas alkaphilia gen. nov. sp. nov., were sequenced using Illumina technology. Closed genomes were produced for all eight isolates.

Exploration of neuroendocrine and immune gene expression in peripheral blood mononuclear cells.

As pathways of communication between the nervous, endocrine, and immune systems are identified, the importance of the interplay of these systems for health and well-being is increasingly recognized. In this study, we created a comprehensive database of 1622 genes likely to be involved in synthetic, biochemical, and regulatory psycho-neuroendocrine-immune (PNI) pathways. Expression of 1058 of these genes was detected in the peripheral blood by querying both a peripheral blood-specific expressed sequence tag (EST) database and a peripheral blood database generated from microarray evaluation of 30,000 genes. Several neural and endocrine genes were expressed in the peripheral blood including hormone receptors, a hormone-responsive transcription factor, and neurotransmitter receptors. These findings document the expression of nervous and endocrine genes in the peripheral blood that have previously only been characterized in the respective system tissues, and indicate that the blood is a rich source of information that should help in deciphering the communication between the mind and the body.