Explore the vast range of titles available.

Using a pyrosequencing-based custom-made sequencer BIGIS-4, we sequenced a Gram-negative bacterium Glaciecola mesophila sp. nov. ( Gmn ) isolated from marine invertebrate specimens. We generated 152043 sequencing reads with a mean high-quality length of 406 bp, and assembled them using the BIGIS-4 post-processing module. No systematic low-quality data was detected beyond expected homopolymer-derived errors. The assembled Gmn genome is 5144318 bp in length and harbors 4303 annotated genes. A large number of metabolic genes correspond to various nutrients from surface marine invertebrates. Its abundant cold-tolerant and cellular signaling and related genes reveal a fundamental adaptation to low-temperature marine environment.

A number of basic and applied questions in ecology and environmental management require the characterization of soil and leaf litter faunal diversity. Recent advances in high-throughput sequencing of barcode-gene amplicons （＇metabarcoding＇） have made it possible to survey biodiversity in a robust and efficient way. However, one obstacle to the widespread adoption of this technique is the need to choose amongst many candidates for bioinformatic processing of the raw sequencing data. We com- pare three candidate pipelines for the processing of 18S small subunit rDNA metabarcode data from solid substrates： （i） USEARCH/CROP, （ii） Denoiser/UCLUST, and （iii） OCTUPUS. The three pipelines produced reassuringly similar and highly correlated assessments of community composition that are dominated by taxa known to characterize the sampled environments However, OCTUPUS appears to inflate phylogenetic diversity, because of higher sequence noise. We therefore recommend either the USEARCH/CROP or Denoiser/UCLUST pipelines, both of which can be run within the QIIME （Quantitative In- sights Into Microbial Ecology） environment.

The application of the aero‐derivative gas turbine to warship propulsion is described in detail. Comparisons of the different roles of the warship and aircraft are made to better understand the marine propulsion system solutions and methods of operation and control. The demanding marine environment is discussed including the effects and mitigation of salt spray, green water exposure, inlet and exhaust location relative the engine installation and the need for infrared suppression. Accommodation of the wide range of operational requirements has lead to complex machinery arrangements with boost and cruise engines operating separately or in combination together with a Controllable Reversible Propeller (CRP). Ancillary systems are described including blow‐down starting and engine washing systems, fuel supply and purification systems. The importance of modeling of the ship and its propulsion machinery as an essential tool for understanding and optimizing the various control modes and machinery combinations is emphasized. A typical control strategy is presented wherein powerplant selection to match ship requirements is automated as well as the scheduling of throttle and propeller pitch to meet immediate ship speed demands during various maneuvers.

Background Granulomatous amoebic encephalitis (GAE) is an infrequent and fatal infectious disease worldwide. Antemortem diagnosis in this condition is very difficult because clinical manifestations and neuroimaging are nonspecific. Case presentation A 60-year-old Japanese woman was admitted with a chief complaint of left homonymous hemianopsia. Brain-MRI showed extensive necrotizing lesions enhanced by gadolinium, in the right frontal lobe, right occipital lobe, and left parietal lobe. Epithelioid granulomas of unknown etiology were found in the biopsied brain specimens. Shotgun metagenomic sequencing using a next-generation sequencer detected DNA fragments of Balamuthia mandrillaris in the tissue specimens. The diagnosis of granulomatous amoebic encephalitis was confirmed using an amoeba-specific polymerase chain reaction and immunostaining on the biopsied tissues. Conclusions Shotgun metagenomics is useful for the diagnosis of central nervous system infections such as GAE wherein the pathogens are difficult to identify.

This study aims to demonstrate the effectiveness of using biological oxidation for hydrogen sulfide (H2S) control. A long-term experiment was conducted using a rod-shaped electrode made of highly conductive concrete, which provided an electron pathway for H2S mitigation. Bacterial flora analysis was conducted using PCR-DGGE and metagenomic analysis by next-generation sequencing to identify electricity-producing bacteria. Results showed that H2S was effectively mitigated, and electricity-producing bacteria, including Geobacter sp. and Pelobacter sp., were found around the inner surface of the anode. The study found that highly conductive concrete can create an electron pathway for biological oxidation of H2S. Oxygen from the air layer near the surface of the water can act as an electron acceptor, even under anaerobic conditions, enabling effective H2S control in sewer systems.

Background The human leukocyte antigen (HLA) region, the 3.8-Mb segment of the human genome at 6p21, has been associated with more than 100 different diseases, mostly autoimmune diseases. Due to the complex nature of HLA genes, there are difficulties in elucidating complete HLA gene sequences especially HLA gene haplotype structures by the conventional sequencing method. We propose a novel, accurate, and cost-effective method for generating phase-defined complete sequencing of HLA genes by using indexed multiplex next generation sequencing. Results A total of 33 HLA homozygous samples, 11 HLA heterozygous samples, and 3 parents-child families were subjected to phase-defined HLA gene sequencing. We applied long-range PCR to amplify six HLA genes ( HLA-A, -C, -B, DRB1, -DQB1 , and –DPB1 ) followed by transposase-based library construction and multiplex sequencing with the MiSeq sequencer. Paired-end reads (2 × 250 bp) derived from the sequencer were aligned to the six HLA gene segments of UCSC hg19 allowing at most 80 bases mismatch. For HLA homozygous samples, the six amplicons of an individual were pooled and simultaneously sequenced and mapped as an individual-tagging method. The paired-end reads were aligned to corresponding genes of UCSC hg19 and unambiguous, continuous sequences were obtained. For HLA heterozygous samples, each amplicon was separately sequenced and mapped as a gene-tagging method. After alignments, we detected informative paired-end reads harboring SNVs on both forward and reverse reads that are used to separate two chromosomes and to generate two phase-defined sequences in an individual. Consequently, we were able to determine the phase-defined HLA gene sequences from promoter to 3′-UTR and assign up to 8-digit HLA allele numbers, regardless of whether the alleles are rare or novel. Parent–child trio-based sequencing validated our sequencing and phasing methods. Conclusions Our protocol generated phased-defined sequences of the entire HLA genes, resulting in high resolution HLA typing and new allele detection.

Oceanic island ecosystems are vulnerable to the introduction of alien species, and they provide a habitat for many endangered species. Knowing the diet of an endangered animal is important for appropriate nature restoration efforts on oceanic islands because introduced species may be a major component of the diets of some endangered species. DNA barcoding techniques together with next‐generation sequencing may provide more detailed information on animal diets than other traditional methods. We performed a diet analysis using 48 fecal samples from the critically endangered red‐headed wood pigeon that is endemic to the Ogasawara Islands based on chloroplast trnL P6 loop sequences. The frequency of each detected plant taxa was compared with a microhistological analysis of the same sample set. The DNA barcoding approach detected a much larger number of plants than the microhistological analysis. Plants that were difficult to identify by microhistological analysis after being digested in the pigeon stomachs were frequently identified only by DNA barcoding. The results of the barcoding analysis indicated the frequent consumption of introduced species, in addition to several native species, by the red‐headed wood pigeon. The rapid eradication of specific introduced species may reduce the food resources available to this endangered bird; thus, balancing eradication efforts with the restoration of native food plants should be considered. Although some technical problems still exist, the trnL approach to next‐generation sequencing may contribute to a better understanding of oceanic island ecosystems and their conservation. We performed diet analysis of critically endangered red‐headed wood pigeon using next‐generation sequencer. The results indicated frequent use of introduced species, not only native species, by the pigeon.

The coronavirus disease 2019 (COVID-19) pandemic has crudely demonstrated the need for massive and rapid diagnostics. By the first week of July, more than 10,000,000 positive cases of COVID-19 have been reported worldwide, although this number could be greatly underestimated. In the case of an epidemic emergency, the first line of response should be based on commercially available and validated resources. Here, we demonstrate the use of the miniPCR, a commercial compact and portable PCR device recently available on the market, in combination with a commercial well-plate reader as a diagnostic system for detecting genetic material of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of COVID-19. We used the miniPCR to detect and amplify SARS-CoV-2 DNA sequences using the sets of initiators recommended by the World Health Organization (WHO) for targeting three different regions that encode for the N protein. Prior to amplification, samples were combined with a DNA intercalating reagent (i.e., EvaGreen Dye). Sample fluorescence after amplification was then read using a commercial 96-well plate reader. This straightforward method allows the detection and amplification of SARS-CoV-2 nucleic acids in the range of ~625 to 2×105 DNA copies. The accuracy and simplicity of this diagnostics strategy may provide a cost-efficient and reliable alternative for COVID-19 pandemic testing, particularly in underdeveloped regions where RT-QPCR instrument availability may be limited. The portability, ease of use, and reproducibility of the miniPCR makes it a reliable alternative for deployment in point-of-care SARS-CoV-2 detection efforts during pandemics.

Background A simple and accurate molecular diagnostic method for malaria is urgently needed due to the limitations of conventional microscopic examination. In this study, we demonstrate a new diagnostic procedure for human malaria using loop mediated isothermal amplification (LAMP) and the MinION™ nanopore sequencer. Methods We generated specific LAMP primers targeting the 18S–rRNA gene of all five human Plasmodium species including two P. ovale subspecies ( P. falciparum, P. vivax, P. ovale wallikeri, P. ovale curtisi , P. knowlesi and P. malariae ) and examined human blood samples collected from 63 malaria patients in Indonesia. Additionally, we performed amplicon sequencing of our LAMP products using MinION™ nanopore sequencer to identify each Plasmodium species. Results Our LAMP method allowed amplification of all targeted 18S–rRNA genes of the reference plasmids with detection limits of 10–100 copies per reaction. Among the 63 clinical samples, 54 and 55 samples were positive by nested PCR and our LAMP method, respectively. Identification of the Plasmodium species by LAMP amplicon sequencing analysis using the MinION™ was consistent with the reference plasmid sequences and the results of nested PCR. Conclusions Our diagnostic method combined with LAMP and MinION™ could become a simple and accurate tool for the identification of human Plasmodium species, even in resource-limited situations.