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Next-generation sequencing (NGS)-library preparation for whole-genome sequencing (WGS) starts with DNA fragmentation, and sonication is a physical approach used most often due to its simplicity and reproducibility. However, the commercially available Covaris instrument has a high price for both the device and consumables. Here, we describe our in-house method of DNA shearing by sonication with small (100–600 µm) glass beads and an ultrasonic bath. The fragmentation conditions were optimized for the bacterial WGS with ∼550-bp fragment size (the ultrasonic bath water temperature 5–10°C, glass beads 0.06 g, the fragmentation time 50 s) and for human DNA with ∼250 bp (fragmentation with the same parameters for 4 min). Fragmentation results were compared with the Covaris instrument for preparing several bacterial NGS libraries for Illumina NGS platforms by several characteristics. We obtained close mean fragment lengths (523–623 versus 480–646), similar mono- and dinucleotide specificity of shearing, and comparable indicators of read alignment and de novo assembly for both methods. Thus, the described method is a new fast, and effective DNA fragmentation approach that can be used in different WGS applications.

Extended-spectrum beta-lactamases (ESBL) and AmpC producing-Escherichia coli have spread worldwide, but data about ESBL-producing-E. coli in the Northern and Eastern regions of Europe is scant. The aim of this study has been to describe the phenotypical and molecular epidemiology of different ESBL/AmpC/Carbapenemases genes in E. coli strains isolated from the Baltic States (Estonia, Latvia, and Lithuania), Norway and St. Petersburg (Russia), and to determine the predominant multilocus sequence type and single nucleotide polymorphisms diversity of E. coli isolates deduced by whole genome sequencing (WGS). A total of 10,780 clinical E. coli strains were screened for reduced sensitivity to third-generation cephalosporins. They were collected from 21 hospitals located in Estonia, Latvia, Lithuania, Norway and St. Petersburg during a 5 month period in 2012. The overall prevalence of ESBL/AmpC strains was 4.7% by phenotypical test and 3.9% by sequencing. We found more strains with the ESBL/AmpC phenotype and genotype in St. Petersburg and Latvia than other countries. Of phenotypic E. coli strains, 85% contained confirmed ESBL genes (including blaCTX–M, blaTEM–29, blaTEM–71), AmpC genes (blaCMY–59, blaACT–12/–15/–20, blaESC–6, blaFEC–1, blaDHA–1), or carbapenemase genes (blaNDM–1). blaCTX–M–1, blaCTX–M–14 and blaCTX–M–15 were found in all countries, but blaCTX–M–15 prevalence was higher in Latvia than in St. Petersburg (Russia), Estonia, Norway and Lithuania. The dominating AmpC genes were blaCMY–59 in the Baltic States and Norway, and blaDHA–1 in St. Petersburg. E. coli strains belonged to 83 different sequence types, of which the most prevalent was ST131 (40%). In conclusion, we generally found low ESBL/AmpC/Carbapenemase prevalence in E. coli strains isolated in Northern/Eastern Europe. However, several inter-country differences in distribution of particular genes and multilocus sequence types were found.

Foodborne disease is one of the major public health problems worldwide. The conventional methods used to detect foodborne pathogens require multiple incubation steps and take a long time to results. Thus, there is an urgent need for biosensors that can determine pathogens without special sample preparation. This paper outlines a method for discriminating E. coli contaminated meat since this bacterium has been implicated as one of the main causative agents of food illness. The method is based on the laser-induced fluorescence response of the Flavin group that is presented among E. coli bacteria’s metabolites. The fluorescence activity of metabolites produced by five E. coli strains was investigated. The list of analyzed strains contains ATCC 25922 E. coli, Enteropathogenic E. coli, Enteroinvasive E. coli, Enterotoxigenic E. coli, and Enteroaggregative E. coli. The preliminary results have allowed developing the statistical model using a part of the fluorescence spectra in the range of 520-560 nm when excited at 450 nm. The proposed model is aimed to differentiate contaminated and uncontaminated meat samples. It has been confirmed that the examined technique provides detection of a bacterial concentration of 106 CFU∙cm-2 in five hours after initial contamination at room temperature. The further improvement of the method by using fluorescent probes is discussed.

Extended-spectrum beta-lactamases (ESBL) and AmpC producing-Escherichia coli have spread worldwide, but data about ESBL-producing-E. coli in the Northern and Eastern regions of Europe is scant. The aim of this study has been to describe the phenotypical and molecular epidemiology of different ESBL/AmpC/Carbapenemases genes in E. coli strains isolated from the Baltic States (Estonia, Latvia, and Lithuania), Norway and St. Petersburg (Russia), and to determine the predominant multilocus sequence type and single nucleotide polymorphisms diversity of E. coli isolates deduced by whole genome sequencing (WGS). A total of 10,780 clinical E. coli strains were screened for reduced sensitivity to third-generation cephalosporins. They were collected from 21 hospitals located in Estonia, Latvia, Lithuania, Norway and St. Petersburg during a 5 month period in 2012. The overall prevalence of ESBL/AmpC strains was 4.7% by phenotypical test and 3.9% by sequencing. We found more strains with the ESBL/AmpC phenotype and genotype in St. Petersburg and Latvia than other countries. Of phenotypic E. coli strains, 85% contained confirmed ESBL genes (including blaCTX–M, blaTEM–29, blaTEM–71), AmpC genes (blaCMY–59, blaACT–12/–15/–20, blaESC–6, blaFEC–1, blaDHA–1), or carbapenemase genes (blaNDM–1). blaCTX–M–1, blaCTX–M–14 and blaCTX–M–15 were found in all countries, but blaCTX–M–15 prevalence was higher in Latvia than in St. Petersburg (Russia), Estonia, Norway and Lithuania. The dominating AmpC genes were blaCMY–59 in the Baltic States and Norway, and blaDHA–1 in St. Petersburg. E. coli strains belonged to 83 different sequence types, of which the most prevalent was ST131 (40%). In conclusion, we generally found low ESBL/AmpC/Carbapenemase prevalence in E. coli strains isolated in Northern/Eastern Europe. However, several inter-country differences in distribution of particular genes and multilocus sequence types were found.

The aim of this study was to compare the prevalence of different virulence factor (VF) genes in extended-spectrum beta-lactamase (ESBL) producingEscherichia colistrains isolated from the Baltic Sea region. A total of 432 strains of phenotypically ESBL positiveE. coliwere collected from 20 institutions located in Estonia, Latvia, Lithuania, and the region of St. Petersburg in Russia from January to May 2012 and analyzed for phylogenetic group and prevalence of 23 VF genes. The strains were collected from clinical material (urine, blood, wound, and respiratory tract). Bacterial isolates were compared according to phylogenetic group, clinical material, and geographical origin. Most of the VF genes were concentrated within phylogenetic group B2 and/or D. When comparing strains isolated from different countries, it was found that strains originating from Estonia and Latvia belonged mainly to group B2 and strains from Lithuania and Russia mainly to groups B2 and D. The P-fimbrial adhesin genepapEFwas more prevalent in Russian strains, colicin genecvaCin Lithuanian strains, and capsular genekpsMTIIin Latvian strains; serum resistant genetraTwas less prevalent in Estonian strains. The regional differences of VF genes remained statistically significant after taking into account the phylogenetic distribution in the countries.