Genes located within the ФAH14b genome and proteins homologous to the ФAH14b-encoded proteins found in selected <i>Pseudomonas</i> spp. genomes and Enterobacteria phage P4.

<p>Searches were performed with the following cut offs <i>E</i> value < 1e-10 and < 1e-40. The hits obtained with e-value 1e^-40 are bolded.</p

Genes located within the ФAH14a genome and proteins homologous to the ФAH14a-encoded proteins found in selected <i>Pseudomonas</i> spp. genomes and <i>Pseudomonas</i> phages.

<p>Searches were performed with the following cut offs <i>E</i> value < 1e-10 and < 1e-40. The hits obtained with e-value 1e^-40 are bolded.</p

Genome organization of the prophages ΦAH14a and ΦAH14b of <i>Pseudomonas</i> sp. ANT_H14.

<p>Arrows indicate the transcriptional orientation of the genes. The gray shaded blocks represent genetic modules identified within the prophages. The genes encoding tail assembly protein I (TAPI) are indicated.</p

Two Inducible Prophages of an Antarctic <i>Pseudomonas</i> sp. ANT_H14 Use the Same Capsid for Packaging Their Genomes – Characterization of a Novel Phage Helper-Satellite System - Fig 5

<p><b>Restriction patterns of DNAs</b>: isolated from viral particles (a mixture of ФAH14a and ФAH14b genomic DNA) cleaved with selected REases: HpaII (CCGG), MspI (CCGG) and SmaI (CCCGGG) [panel A] and <i>Pseudomonas</i> sp. ANT_H14 genomic DNA cleaved with SmaI (CCCGGG) [panel B]. Digest mixtures were electrophoresed on 0.8% agarose gels and stained with ethidium bromide. ND–undigested DNA isolated from viral particles; λ/S–DNA of λ <i>dam</i>^<i>-</i> <i>dcm</i>^<i>-</i> digested with SmaI; Ps–undigested <i>Pseudomonas</i> sp. ANT_H14 genomic DNA; Ps/S–ANT_H14 genomic DNA digested with SmaI; M–GeneRuler 100–10,000 bp size marker.</p

Separation of ΦAH14a and ΦAH14b virion proteins by SDS-polyacrylamide gel electrophoresis (12%).

<p>M–Page Ruler prestained protein ladder SM0671 (Thermo Scientific).</p

Restriction patterns of DNA extracted from purified virions cleaved with the selected REases.

<p>Panel A: HindIII (H), Eco32I (E32), SalI (S) and EcoRI (EI). ND, undigested DNA. M, GeneRuler 100- to 10,000-bp size marker. λ/H, λ DNA cleaved with HindIII. ~14 kb restriction fragment of EcoR32I digested virion DNA is marked with a triangle. Panel B: Arrows indicate restriction fragments assigned to ФAH14a (left) and ФAH14b (right) genomic DNA, obtained by SalI digestion.</p

Presentation1_Diversity of antibiotic resistance gene variants at subsequent stages of the wastewater treatment process revealed by a metagenomic analysis of PCR amplicons.pdf

Wastewater treatment plants have been recognised as point sources of various antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARG) which are considered recently emerging biological contaminants. So far, culture-based and molecular-based methods have been successfully applied to monitor antimicrobial resistance (AMR) in WWTPs. However, the methods applied do not permit the comprehensive identification of the true diversity of ARGs. In this study we applied next-generation sequencing for a metagenomic analysis of PCR amplicons of ARGs from the subsequent stages of the analysed WWTP. The presence of 14 genes conferring resistance to different antibiotic families was screened by PCR. In the next step, three genes were selected for detailed analysis of changes of the profile of ARG variants along the process. A relative abundance of 79 variants was analysed. The highest diversity was revealed in the ermF gene, with 52 variants. The relative abundance of some variants changed along the purification process, and some ARG variants might be present in novel hosts for which they were currently unassigned. Additionally, we identified a pool of novel ARG variants present in the studied WWTP. Overall, the results obtained indicated that the applied method is sufficient for analysing ARG variant diversity.</p

Plasmids of Carotenoid-Producing <i>Paracoccus</i> spp. (<i>Alphaproteobacteria</i>) - Structure, Diversity and Evolution

<div><p>Plasmids are components of many bacterial genomes. They enable the spread of a large pool of genetic information via lateral gene transfer. Many bacterial strains contain mega-sized replicons and these are particularly common in <i>Alphaproteobacteria</i>. Considerably less is known about smaller alphaproteobacterial plasmids. We analyzed the genomes of 14 such plasmids residing in 4 multireplicon carotenoid-producing strains of the genus <i>Paracoccus</i> (<i>Alphaproteobacteria</i>): <i>P. aestuarii</i> DSM 19484, <i>P. haeundaensis</i> LG P-21903, <i>P. marcusii</i> DSM 11574 and <i>P. marcusii</i> OS22. Comparative analyses revealed mosaic structures of the plasmids and recombinational shuffling of diverse genetic modules involved in (i) plasmid replication, (ii) stabilization (including toxin-antitoxin systems of the <i>relBE</i>/<i>parDE</i>, <i>tad</i>-<i>ata</i>, <i>higBA</i>, <i>mazEF</i> and <i>toxBA</i> families) and (iii) mobilization for conjugal transfer (encoding relaxases of the Mob_Q, Mob_P or Mob_V families). A common feature of the majority of the plasmids is the presence of AT-rich sequence islets (located downstream of <i>exc1</i>-like genes) containing genes, whose homologs are conserved in the chromosomes of many bacteria (encoding e.g. RelA/SpoT, SMC-like proteins and a retron-type reverse transcriptase). The results of this study have provided insight into the diversity and plasticity of plasmids of <i>Paracoccus</i> spp., and of the entire <i>Alphaproteobacteria</i>. Some of the identified plasmids contain replication systems not described previously in this class of bacteria. The composition of the plasmid genomes revealed frequent transfer of chromosomal genes into plasmids, which significantly enriches the pool of mobile DNA that can participate in lateral transfer. Many strains of <i>Paracoccus</i> spp. have great biotechnological potential, and the plasmid vectors constructed in this study will facilitate genetic studies of these bacteria. </p> </div

Schematic structure of the REP modules analyzed in this study.

<p>The color-coded keys show the species and strain of origin of each plasmid (circles) and identified direct repeats (DRs), inverted repeats (IRs) as well as predicted DnaA and IHF binding sites (mixed shapes). The sequences of the iteron-like DRs are presented next to the relevant diagrams with a consensus sequence shown for DRs of plasmids with related REP modules. Blue arrows indicate the <i>rep</i> genes and their transcriptional orientation. Specific motifs identified within the aa sequences of the Rep proteins are indicated by colored rounded bars. A+T and G+C indicate DNA regions of lower or higher than average G+C content, respectively. The components of the REP modules are not shown to scale.</p

Distribution of the REP modules analyzed in this study in the <i>Paracoccus</i> spp. genomes.

<p>A specific DNA probe (fragment of a <i>rep</i> gene amplified by PCR and DIG-labeled) was prepared for each analyzed REP module and used in dot blot hybridization analysis with total DNA isolated from 20 strains of <i>Paracoccus</i> spp. The results are presented as a matrix. The relatedness of the tested <i>Paracoccus</i> strains is shown beneath by a phylogenetic tree based on their 16S rDNA sequences. The tree was constructed by the neighbor-joining algorithm with Kimura corrected distances. The statistical support for the internal nodes was determined by 1000 bootstrap replicates and values of >50% are shown. The <i>Paracoccus</i> strains from which the plasmids were isolated are denoted by red text. </p