An ecophysiological explanation for manganese enrichment in rock varnish.

Desert varnish is a dark rock coating that forms in arid environments worldwide. It is highly and selectively enriched in manganese, the mechanism for which has been a long-standing geological mystery. We collected varnish samples from diverse sites across the western United States, examined them in petrographic thin section using microscale chemical imaging techniques, and investigated the associated microbial communities using 16S amplicon and shotgun metagenomic DNA sequencing. Our analyses described a material governed by sunlight, water, and manganese redox cycling that hosts an unusually aerobic microbial ecosystem characterized by a remarkable abundance of photosynthetic Cyanobacteria in the genus Chroococcidiopsis as the major autotrophic constituent. We then showed that diverse Cyanobacteria, including the relevant Chroococcidiopsis taxon, accumulate extraordinary amounts of intracellular manganese-over two orders of magnitude higher manganese content than other cells. The speciation of this manganese determined by advanced paramagnetic resonance techniques suggested that the Cyanobacteria use it as a catalytic antioxidant-a valuable adaptation for coping with the substantial oxidative stress present in this environment. Taken together, these results indicated that the manganese enrichment in varnish is related to its specific uptake and use by likely founding members of varnish microbial communities.

Hidden Markov Model: a shortest unique representative approach to detect the protein toxins, virulence factors and antibiotic resistance genes.

OBJECTIVE: Currently, next generation sequencing (NGS) is widely used to decode potential novel or variant pathogens both in emergent outbreaks and in routine clinical practice. However, the efficient identification of novel or diverged pathogenomic compositions remains a big challenge. It is especially true for short DNA sequence fragments from NGS, since sequence similarity searching is vulnerable to false negatives or false positives, as is mismatching or matching with unrelated proteins. Therefore, this study aimed to establish a bioinformatics approach that can generate unique motif sequences for profiling searching, resulting in high specificity and sensitivity. RESULTS: In this study, we introduced a Shortest Unique Representative Hidden Markov Model (HMM) approach to identify bacterial toxin, virulence factor (VF), and antimicrobial resistance (AR) in short sequence reads. We first construct unique representative domain sequences of toxin genes, VFs, and ARs to avoid potential false positives, and then to use HMM models to accurately identify potential toxin, VF, and AR fragments. The benchmark shows this approach can achieve relatively high specificity and sensitivity if the appropriate cutoff value is applied. Our approach can be used to recognize the protein sequences of known toxins and pathogens, identifies their common characteristics and then searches for similar sequences in other organisms.

The Distinctive Evolution of orfX Clostridium parabotulinum Strains and Their Botulinum Neurotoxin Type A and F Gene Clusters Is Influenced by Environmental Factors and Gene Interactions via Mobile Genetic Elements.

Of the seven currently known botulinum neurotoxin-producing species of Clostridium, C. parabotulinum, or C. botulinum Group I, is the species associated with the majority of human botulism cases worldwide. Phylogenetic analysis of these bacteria reveals a diverse species with multiple genomic clades. The neurotoxins they produce are also diverse, with over 20 subtypes currently represented. The existence of different bont genes within very similar genomes and of the same bont genes/gene clusters within different bacterial variants/species indicates that they have evolved independently. The neurotoxin genes are associated with one of two toxin gene cluster types containing either hemagglutinin (ha) genes or orfX genes. These genes may be located within the chromosome or extrachromosomal elements such as large plasmids. Although BoNT-producing C parabotulinum bacteria are distributed globally, they are more ubiquitous in certain specific geographic regions. Notably, northern hemisphere strains primarily contain ha gene clusters while southern hemisphere strains have a preponderance of orfX gene clusters. OrfX C. parabotulinum strains constitute a subset of this species that contain highly conserved bont gene clusters having a diverse range of bont genes. While much has been written about strains with ha gene clusters, less attention has been devoted to those with orfX gene clusters. The recent sequencing of 28 orfX C. parabotulinum strains and the availability of an additional 91 strains for analysis provides an opportunity to compare genomic relationships and identify unique toxin gene cluster characteristics and locations within this species subset in depth. The mechanisms behind the independent processes of bacteria evolution and generation of toxin diversity are explored through the examination of bacterial relationships relating to source locations and evidence of horizontal transfer of genetic material among different bacterial variants, particularly concerning bont gene clusters. Analysis of the content and locations of the bont gene clusters offers insights into common mechanisms of genetic transfer, chromosomal integration, and development of diversity among these genes.

Comparative genomic and phenotypic characterization of invasive non-typhoidal Salmonella isolates from Siaya, Kenya.

Non-typhoidal Salmonella (NTS) is a major global health concern that often causes bloodstream infections in areas of the world affected by malnutrition and comorbidities such as HIV and malaria. Developing a strategy to control the emergence and spread of highly invasive and antimicrobial resistant NTS isolates requires a comprehensive analysis of epidemiological factors and molecular pathogenesis. Here, we characterize 11 NTS isolates that caused bloodstream infections in pediatric patients in Siaya, Kenya from 2003-2010. Nine isolates were identified as S. Typhimurium sequence type 313 while the other two were S. Enteritidis. Comprehensive genotypic and phenotypic analyses were performed to compare these isolates to those previously identified in sub-Saharan Africa. We identified a S. Typhimurium isolate referred to as UGA14 that displayed novel plasmid, pseudogene and resistance features as compared to other isolates reported from Africa. Notably, UGA14 is able to ferment both lactose and sucrose due to the acquisition of insertion elements on the pKST313 plasmid. These findings show for the first time the co-evolution of plasmid-mediated lactose and sucrose metabolism along with cephalosporin resistance in NTS further elucidating the evolutionary mechanisms of invasive NTS phenotypes. These results further support the use of combined genomic and phenotypic approaches to detect and characterize atypical NTS isolates in order to advance biosurveillance efforts that inform countermeasures aimed at controlling invasive and antimicrobial resistant NTS.

Genomic Characterization of Newly Completed Genomes of Botulinum Neurotoxin-Producing Species from Argentina, Australia, and Africa.

Botulinum neurotoxin-producing clostridia are diverse in the types of toxins they produce as well as in their overall genomic composition. They are globally distributed, with prevalent species and toxin types found within distinct geographic regions, but related strains containing the same toxin types may also be located on distinct continents. The mechanisms behind the spread of these bacteria and the independent movements of their bont genes may be understood through examination of their genetic backgrounds. The generation of 15 complete genomic sequences from bacteria isolated in Argentina, Australia, and Africa allows for a thorough examination of genome features, including overall relationships, bont gene cluster locations and arrangements, and plasmid comparisons, in bacteria isolated from various areas in the southern hemisphere. Insights gained from these examinations provide an understanding of the mechanisms behind the independent movements of these elements among distinct species.

Genome Sequence of Staphylococcus pettenkoferi Strain SMA0010-04 (UGA20), a Clinical Isolate from Siaya County Referral Hospital in Siaya, Kenya.

Here, we report the sequence of a Staphylococcus pettenkoferi clinical isolate, strain SMA0010-04 (UGA20), which contains the PC1 beta-lactamase (blaZ) gene.

Genome Sequences of a Staphylococcus aureus Clinical Isolate, Strain SMA0034-04 (UGA22), from Siaya County Referral Hospital in Siaya, Kenya.

Here, we report the genome sequences of a Staphylococcus aureus clinical isolate, strain SMA0034-04 (UGA22), which contains one chromosome and one plasmid. We also reveal that isolate SMA0034-04 (UGA22) contains loci in the genome that encode multiple exotoxins.

Defending Our Public Biological Databases as a Global Critical Infrastructure.

Progress in modern biology is being driven, in part, by the large amounts of freely available data in public resources such as the International Nucleotide Sequence Database Collaboration (INSDC), the world's primary database of biological sequence (and related) information. INSDC and similar databases have dramatically increased the pace of fundamental biological discovery and enabled a host of innovative therapeutic, diagnostic, and forensic applications. However, as high-value, openly shared resources with a high degree of assumed trust, these repositories share compelling similarities to the early days of the Internet. Consequently, as public biological databases continue to increase in size and importance, we expect that they will face the same threats as undefended cyberspace. There is a unique opportunity, before a significant breach and loss of trust occurs, to ensure they evolve with quality and security as a design philosophy rather than costly "retrofitted" mitigations. This Perspective surveys some potential quality assurance and security weaknesses in existing open genomic and proteomic repositories, describes methods to mitigate the likelihood of both intentional and unintentional errors, and offers recommendations for risk mitigation based on lessons learned from cybersecurity.

Genome Sequence of a Staphylococcus xylosus Clinical Isolate, Strain SMA0341-04 (UGA5), from Siaya County Referral Hospital in Siaya, Kenya.

We report here the genome sequence of a Staphylococcus xylosus clinical isolate, strain SMA0341-04 (UGA5), which contains one chromosome and at least one plasmid. Notably, strain SMA0341-04 (UGA5) contains the tetracycline efflux major facilitator superfamily (MFS) transporter (tetK) gene.

Draft Genome Sequences of Two Staphylococcus warneri Clinical Isolates, Strains SMA0023-04 (UGA3) and SMA0670-05 (UGA28), from Siaya County Referral Hospital, Siaya, Kenya.

We report the complete draft genome sequences of two Staphylococcus warneri clinical isolates, strains SMA0023-04 (UGA3) and SMA0670-05 (UGA28), each of which contains one chromosome and at least one plasmid. Isolate SMA0023-04 (UGA3) contains tetracycline efflux major facilitator superfamily (MFS) transporter (tetK), macrolide resistance (msrC and mphC), and beta-lactamase (blaZ) genes on its plasmids.

Exception to the Rule: Genomic Characterization of Naturally Occurring Unusual Vibrio cholerae Strains with a Single Chromosome.

The genetic make-up of most bacteria is encoded in a single chromosome while about 10% have more than one chromosome. Among these, Vibrio cholerae, with two chromosomes, has served as a model system to study various aspects of chromosome maintenance, mainly replication, and faithful partitioning of multipartite genomes. Here, we describe the genomic characterization of strains that are an exception to the two chromosome rules: naturally occurring single-chromosome V. cholerae. Whole genome sequence analyses of NSCV1 and NSCV2 (natural single-chromosome vibrio) revealed that the Chr1 and Chr2 fusion junctions contain prophages, IS elements, and direct repeats, in addition to large-scale chromosomal rearrangements such as inversions, insertions, and long tandem repeats elsewhere in the chromosome compared to prototypical two chromosome V. cholerae genomes. Many of the known cholera virulence factors are absent. The two origins of replication and associated genes are generally intact with synonymous mutations in some genes, as are recA and mismatch repair (MMR) genes dam, mutH, and mutL; MutS function is probably impaired in NSCV2. These strains are ideal tools for studying mechanistic aspects of maintenance of chromosomes with multiple origins and other rearrangements and the biological, functional, and evolutionary significance of multipartite genome architecture in general.

Complete Genome Sequence of Veillonella atypica OK5, the First Transformable Strain in the Species.

The Veillonella atypica strain OK5 was isolated from a human saliva sample and was the first strain shown to be genetically transformable in the Veillonella genus. Genetic studies using this strain have helped us gain much insight into the ecology of human oral biofilms. Here, we report the complete genome sequence of V. atypica OK5.

Genome Sequence of Porphyromonas gingivalis Strain A7A1-28.

Porphyromonas gingivalis is an oral opportunistic pathogen. Sequenced P. gingivalis laboratory strains display limited diversity in antigens that modulate host responses. Here, we present the genome sequence of A7A1-28, a strain possessing atypical fimbrillin and capsule types, with a single contig of 2,249,024 bp and a G+C content of 48.58%.

Genome Sequence of Porphyromonas gingivalis Strain 381.

Porphyromonas gingivalis is associated with both oral and systemic diseases. Strain-specific P. gingivalis invasion phenotypes do not reliably predict disease presentation during in vivo studies. Here, we present the genome sequence of 381, a common laboratory strain, with a single contig of 2,378,872 bp and a G+C content of 48.36%.

Comparative genomic analyses reveal broad diversity in botulinum-toxin-producing Clostridia.

BACKGROUND: Clostridium botulinum is a diverse group of bacteria characterized by the production of botulinum neurotoxin. Botulinum neurotoxins are classified into serotypes (BoNT/A-G), which are produced by six species/Groups of Clostridia, but the genetic background of the bacteria remains poorly understood. The purpose of this study was to use comparative genomics to provide insights into the genetic diversity and evolutionary history of bacteria that produce the potent botulinum neurotoxin. RESULTS: Comparative genomic analyses of over 170 Clostridia genomes, including our draft genome assemblies for 59 newly sequenced Clostridia strains from six continents and publicly available genomic data, provided in-depth insights into the diversity and distribution of BoNT-producing bacteria. These newly sequenced strains included Group I and II strains that express BoNT/A,/B,/E, or/F as well as bivalent strains. BoNT-producing Clostridia and closely related Clostridia species were delineated with a variety of methods including 16S rRNA gene, concatenated marker genes, core genome and concatenated multi-locus sequencing typing (MLST) gene phylogenies that related whole genome sequenced strains to publicly available strains and sequence types. These analyses illustrated the phylogenetic diversity in each Group and the diversity of genomic backgrounds that express the same toxin type or subtype. Comparisons of the botulinum neurotoxin genes did not identify novel toxin types or variants. CONCLUSIONS: This study represents one of the most comprehensive analyses of whole genome sequence data for Group I and II BoNT-producing strains. Read data and draft genome assemblies generated for 59 isolates will be a resource to the research community. Core genome phylogenies proved to be a powerful tool for differentiating BoNT-producing strains and can provide a framework for the study of these bacteria. Comparative genomic analyses of Clostridia species illustrate the diversity of botulinum-neurotoxin-producing strains and the plasticity of the genomic backgrounds in which bont genes are found.

Genome Sequence of Porphyromonas gingivalis Strain AJW4.

Porphyromonas gingivalis is associated with oral and systemic diseases. Strain-specific P. gingivalis invasion phenotypes have been correlated with disease presentation in infected laboratory animals. Here, we present the genome sequence of AJW4, a minimally invasive strain, with a single contig of 2,372,492 bp and a G+C content of 48.27%.

Genome Sequence of Porphyromonas gingivalis Strain A7436.

Porphyromonas gingivalis is strongly associated with periodontitis. P. gingivalis strain trafficking and tissue homing differ widely, even among presumptive closely related strains, such as W83 and A7436. Here, we present the genome sequence of A7436 with a single contig of 2,367,029 bp and a G+C content of 48.33%.

Genomic sequences of six botulinum neurotoxin-producing strains representing three clostridial species illustrate the mobility and diversity of botulinum neurotoxin genes.

The whole genomes for six botulinum neurotoxin-producing clostridial strains were sequenced to provide references for under-represented toxin types, bivalent strains or unusual toxin complexes associated with a bont gene. The strains include three Clostridium botulinum Group I strains (CDC 297, CDC 1436, and Prevot 594), a Group II C. botulinum strain (Eklund 202F), a Group IV Clostridium argentinense strain (CDC 2741), and a Group V Clostridium baratii strain (Sullivan). Comparisons of the Group I genomic sequences revealed close relationships and conservation of toxin gene locations with previously published Group I C. botulinum genomes. The bont/F6 gene of strain Eklund 202F was determined to be a chimeric toxin gene composed of bont/F1 and bont/F2. The serotype G strain CDC 2741 remained unfinished in 20 contigs with the bont/G located within a 1.15Mb contig, indicating a possible chromosomal location for this toxin gene. Within the genome of C. baratii Sullivan strain, direct repeats of IS1182 insertion sequence (IS) elements were identified flanking the bont/F7 toxin complex that may be the mechanism of bont insertion into C. baratii. Highlights of the six strains are described and release of their genomic sequences will allow further study of unusual neurotoxin-producing clostridial strains.

Recruiting human microbiome shotgun data to site-specific reference genomes.

The human body consists of innumerable multifaceted environments that predispose colonization by a number of distinct microbial communities, which play fundamental roles in human health and disease. In addition to community surveys and shotgun metagenomes that seek to explore the composition and diversity of these microbiomes, there are significant efforts to sequence reference microbial genomes from many body sites of healthy adults. To illustrate the utility of reference genomes when studying more complex metagenomes, we present a reference-based analysis of sequence reads generated from 55 shotgun metagenomes, selected from 5 major body sites, including 16 sub-sites. Interestingly, between 13% and 92% (62.3% average) of these shotgun reads were aligned to a then-complete list of 2780 reference genomes, including 1583 references for the human microbiome. However, no reference genome was universally found in all body sites. For any given metagenome, the body site-specific reference genomes, derived from the same body site as the sample, accounted for an average of 58.8% of the mapped reads. While different body sites did differ in abundant genera, proximal or symmetrical body sites were found to be most similar to one another. The extent of variation observed, both between individuals sampled within the same microenvironment, or at the same site within the same individual over time, calls into question comparative studies across individuals even if sampled at the same body site. This study illustrates the high utility of reference genomes and the need for further site-specific reference microbial genome sequencing, even within the already well-sampled human microbiome.