Complete genome sequence of the newly discovered temperate Clostridioides difficile bacteriophage phiCDKH01 of the family Siphoviridae

A temperate siphovirus, phiCDKH01, was obtained from a clinical isolate of Clostridioides difficile. The phage genome is a 45,089-bp linear double-stranded DNA molecule with an average G+C content of 28.7%. It shows low similarity to known phage genomes, except for phiCD24-1. Genomic and phylogenetic analysis revealed that phiCDKH01 is a newly discovered phage. Sixty-six putative ORFs were predicted in the genome, 37 of which code for proteins with predicted functions. The phiCDKH01 prophage was localized in the host genome. The results of this study increase our knowledge about the genetic diversity of tailed phages.

Complete genome sequence of a clinical isolate of Clostridioides difficile bacteriophage phiCDKH01 of the family Siphoviridae

A new temperate phiCDKH01 siphophage was obtained from clinical isolate of Clostridioides difficile. The phage genome is a 45,089 bp linear double-stranded DNA molecule with an average G + C content of 28.7%. It shows low similarity to known phage genomes except for phiCD24-1. Genomic and phylogenetic analysis revealed that phiCDKH01 is a novel phage. 66 putative ORFs were predicted in the genome, 37 of which code for proteins with predicted functions. The phiCDKH01 prophage has been localized in the host genome. Results of this study increases genetic diversity of known tailed phages.

Ubiquity, Diversity, and Genomic Complexity of Cyanophages in Freshwater Environments

Cyanophages are viruses that infect cyanobacteria (also known as blue-green algae) and are ubiquitious in marine and freshwater environments. In recent years, freshwater cyanophages have attracted more attention because they can affect global freshwater ecosystems. The spatial distribution and morphological diversity of cyanophage populations were examined in Lake Donghu with three trophic regions: hypertrophic, eutrophic, and mesotrophic regions. The surprisingly high viral abundance (ranging from 108 to 109 phage mL−1) and morphological diversity were detected. Most of them have tails and belong to the families Siphoviridae, Myoviridae, and Podoviridae. Various morphotypes were observed, such as prolate-headed virus-like particles and lemon-shaped virus-like particles. In addition, some cyanophages were studied by virological experiments and whole-genome analyses, combined with morphological observation. For example, three cyanophages were isolated and their whole genomes were sequenced. Contractile tail myonophage MaMV-DC infects bloom-forming cyanobacterium Microcystis aeruginosa. Tailless cyanophage Planktothrix agardhii virus isolated from Lake Donghu (PaV-LD) infects filamentous cyanobacterium. Short-tail podovirus A-4L can infect the model cyanobacterium Anabaena sp. strain PCC 7120. The MaMV-DC genome contains 169,223 bp encoding 170 putative open reading frames (ORFs). The PaV-LD genome posseses 95,299 bp encoding 142 putative ORFs. The genome of short-tail podovirus A-4L has 41,750 bp encoding 38 putative ORFs. There are significant differences in their genomic size and encoded tail proteins, but all three cyanophages contain genes that are not commonly found in phages. By studying the vast biodiversity of viruses in freshwater environments, these novel findings of cyanophages broaden our insights, and allow us to gain more useful knowledge about the global impact of these viruses in freshwater ecosystems.

Characterization of a Novel Thermobifida fusca Bacteriophage P318

Thermobifida fusca is of biotechnological interest due to its ability to produce an array of plant cell wall hydrolytic enzymes. Nonetheless, only one T. fusca bacteriophage with genome information has been reported to date. This study was aimed at discovering more relevant bacteriophages to expand the existing knowledge of phage diversity for this host species. With this end in view, a thermostable T. fusca bacteriophage P318, which belongs to the Siphoviridae family, was isolated and characterized. P318 has a double-stranded DNA genome of 48,045 base pairs with 3′-extended COS ends, on which 52 putative ORFs are organized into clusters responsible for the order of genome replication, virion morphogenesis, and the regulation of the lytic/lysogenic cycle. In comparison with T. fusca and the previously discovered bacteriophage P1312, P318 has a much lower G+C content in its genome except at the region encompassing ORF42, which produced a protein with unknown function. P1312 and P318 share very few similarities in their genomes except for the regions encompassing ORF42 of P318 and ORF51 of P1312 that are homologous. Thus, acquisition of ORF42 by lateral gene transfer might be an important step in the evolution of P318.

Mining cytochrome p450 genes through next generation sequencing and metagenomic analysis from Binh Chau hot spring

Cytochrome P450s (CYPs) are one of the largest distributed enzymes, which catalyze more than 20 different reactions. At present, there has been an increasing realization of the power of P450 biocatalysts for the industrial synthesis of pharmaceuticals, agrochemicals, bulk chemicals, food ingredients, etc. On the other hand, the conditions of industrial processes at high temperature, high-pressure or in chemical solvent require the enzymes, which catalyze the bioconversion, have a specific properties such as thermostability, chemical tolerance or barophilicity. Up to date, the number of thermostable P450s is limited. Nowadays, DNA-metagenome technique gives us a chance to catch novel genes and unique interesting enzymes from microbial community in certain ecology. In this paper, metagenomic DNA extracted from water samples from Binh Chau hot spring was sequenced using Illumila’s HiSeq platform and was analysed to mining putative genes encoding cytochrome P450. The sequencing generated 9.4 Gb of reads containing 156,093 putative ORFs, of these, 106,903 genes were annotated in NCBI non-redundant protein sequence database. Among all the ORFs were annotated, 68 putative ORFs encoding cytochrome P450 were found belong to 36 specific groups of cytochrome P450 protein family. Of these, the melting temperature (Tm) from thirty-six completed ORFs was predicted for a better understanding of thermodynamic stability.

Mining cytochrome p450 genes through next generation sequencing and metagenomic analysis from Binh Chau hot spring

Cytochrome P450s (CYPs) are one of the largest distributed enzymes, which catalyze more than 20 different reactions. At present, there has been an increasing realization of the power of P450 biocatalysts for the industrial synthesis of pharmaceuticals, agrochemicals, bulk chemicals, food ingredients, etc. On the other hand, the conditions of industrial processes at high temperature, high-pressure or in chemical solvent require the enzymes, which catalyze the bioconversion, have a specific properties such as thermostability, chemical tolerance or barophilicity. Up to date, the number of thermostable P450s is limited. Nowadays, DNA-metagenome technique gives us a chance to catch novel genes and unique interesting enzymes from microbial community in certain ecology. In this paper, metagenomic DNA extracted from water samples from Binh Chau hot spring was sequenced using Illumila’s HiSeq platform and was analysed to mining putative genes encoding cytochrome P450. The sequencing generated 9.4 Gb of reads containing 156,093 putative ORFs, of these, 106,903 genes were annotated in NCBI non-redundant protein sequence database. Among all the ORFs were annotated, 68 putative ORFs encoding cytochrome P450 were found belong to 36 specific groups of cytochrome P450 protein family. Of these, the melting temperature (Tm) from thirty-six completed ORFs was predicted for a better understanding of thermodynamic stability.

Complete genome sequence of new virulent actinophages belonging to Streptomyces flavovirens

Complete genome sequence of new virulent actinophages belonging to Streptomyces flavovirens We report biological and physiochemical features and use NGS to provide the complete annotated genomes for two new strains (Sf1 and Sf3) of the virulent phage Streptomyces flavovirens, isolated from Egyptian soil samples. The S. flavovirens phages (Sf1 and Sf3) show high adsorption rates (82 and 85 %, respectively), indicating a strong specificity to their host, and their burst sizes were 1.95 and 2.49 virions per mL. The phage genomes are parts of a singleton cluster with sizes of 43,150 bp and 60,934 bp, respectively. the assignment of possible functions to 19 and 28 putative ORFs were identified, which included phage structural proteins, lysis components and metabolic proteins. Comparative genomic analysis revealed significant homology between the two phages and the closest Streptomyces phage (VWB phages). However, the phylogenetic analysis highlighted that the isolated phages belong to the BG Streptomyces phage group but are clearly separated, representing a novel sub-cluster.

Complete genome sequence of new virulent actinophages belonging to Streptomyces flavovirens

Complete genome sequence of new virulent actinophages belonging to Streptomyces flavovirens We report biological and physiochemical features and use NGS to provide the complete annotated genomes for two new strains (Sf1 and Sf3) of the virulent phage Streptomyces flavovirens, isolated from Egyptian soil samples. The S. flavovirens phages (Sf1 and Sf3) show high adsorption rates (82 and 85 %, respectively), indicating a strong specificity to their host, and their burst sizes were 1.95 and 2.49 virions per mL. The phage genomes are parts of a singleton cluster with sizes of 43,150 bp and 60,934 bp, respectively. the assignment of possible functions to 19 and 28 putative ORFs were identified, which included phage structural proteins, lysis components and metabolic proteins. Comparative genomic analysis revealed significant homology between the two phages and the closest Streptomyces phage (VWB phages). However, the phylogenetic analysis highlighted that the isolated phages belong to the BG Streptomyces phage group but are clearly separated, representing a novel sub-cluster.

Comparative analysis of protein-coding and long non-coding transcripts based on RNA sequence features

RNA plays an important role in the intracellular cell life and in the organism in general. Besides the well-established protein coding RNAs (messenger RNAs, mRNAs), long non-coding RNAs (lncRNAs) have gained the attention of recent researchers. Although lncRNAs have been classified as non-coding, some authors reported the presence of corresponding sequences in ribosome profiling data (Ribo-seq). Ribo-seq technology is a powerful experimental tool utilized to characterize RNA translation in cell with focus on initiation (harringtonine, lactimidomycin) and elongation (cycloheximide). By exploiting translation starts obtained from the Ribo-seq experiment, we developed a novel position weight matrix model for the prediction of translation starts. This model allowed us to achieve 96% accuracy of discrimination between human mRNAs and lncRNAs. When the same model was used for the prediction of putative ORFs in RNAs, we discovered that the majority of lncRNAs contained only small ORFs ([Formula: see text][Formula: see text]nt) in contrast to mRNAs.

First Report of Tomato yellow leaf curl Kanchanaburi virus Infecting Eggplant in Laos

Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) is a bipartite begomovirus (genus Begomovirus, family Geminiviridae) reported to infect tomato and eggplant in Thailand and Vietnam (1,2). In April 2013, eggplant (Solanum melongena L.) plants exhibiting yellow mosaic symptoms were found in a suburb of Vientiane, Laos. Three symptomatic samples were collected. Total DNA was extracted from leaves by the CTAB method, and used as template for PCR using the degenerate primer pair AV494/CoPR (3). The PCR results suggested that the plants were infected by a begomovirus. The begomoviral genome was amplified by rolling circle amplification (RCA) with TempliPhi kit (GE Healthcare) following the manufacturer's protocol. RCA product was digested with the endonucleases BamH I, EcoR I, Hind III, Kpn I, Pst I, and Xba I, respectively. The fragments about 2.1 kbp (with Pst I digestion) and 1.5 kbp (with Xba I digestion) in size were cloned and sequenced. The sequence of the 2.1-kbp fragment showed similarity with begomovirus DNA-A component. A pair of primers for amplification of the full-length DNA-A, AF (5′-CTTCATCGTTTCTCAGCATCAT-3′) and AR (5′-CACTTGCACACGATCTCTAAGA-3′) were designed from the 2.1-kbp sequence. The full-length DNA-A was 2,752 nucleotides and encoded six putative ORFs (GenBank Accession No. KF218820). The sequence of the 1.5-kbp fragment shared similarity with begomoviruses DNA-B. The begomoviral circular DNA-B was amplified using the pair of primers BF (5′-GTAACAGCCGAAGTGCACG-3′) and BR (5′-AATGGAGAGACACCAGTCTGCC-3′) designed from the 1.5-kbp sequence. PCR yielded a product of expected size (~1.4 kbp). The full-length DNA-B sequence was obtained by assembling the two sequences. The DNA-B was 2,734 nucleotides and encoded two putative ORFs (GenBank Accession No. KF218821). The sequences of DNA-A and DNA-B of isolate Laos shared the highest nucleotide sequences identities at 99.0% and 98.0% with those of TYLCKaV-[TH:Kan 1:01] (AF511529), and [TH:Kan 2:Egg:01] (AF511527), respectively. The results indicated that the virus associated with eggplant yellow mosaic disease was an isolate of TYLCKaV. To our knowledge, this is the first report of this begomovirus in Laos. Our results indicate that this virus may be spreading in Southeast Asia and scientists there should be aware of this virus when developing begomovirus-resistant varieties of tomato or eggplant. References: (1) S. K. Green et al. Plant Dis. 87:446, 2003. (2) C. Ha et al. J. Gen. Virol. 89:312, 2008.(3) Z. F. He et al. Arch. Virol. 154:1199, 2009.