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Listeria monocytogenes is an important pathogen responsible for listeriosis, a serious foodborne illness associated with high mortality rates. Therefore, L. monocytogenes is considered a challenge for the food industry due to the ability of some strains to persist in food-associated environments. Biofilm production is presumed to contribute to increased L. monocytogenes resistance and persistence. The aims of this study were to (1) assess the biofilm formation of L. monocytogenes isolates from a meat processing facility and sheep farm previously characterized and subjected to whole-genome sequencing and (2) perform a comparative genomic analysis to compare the biofilm formation and the presence of a known set of biofilm-associated genes and related resistance or persistence markers. Among the 37 L. monocytogenes isolates of 15 sequence types and four serogroups involved in this study, 14%, 62%, and 24% resulted in the formation of weak, moderate, and strong biofilm, respectively. Increased biofilm-forming ability was associated with the presence of the stress survival islet 1 (SSI-1), inlL, and the truncated inlA genes. Combining the phenotypic and genotypic data may contribute to understanding the relationships between biofilm-associated genes and L. monocytogenes biofilm-forming ability, enabling improvement in the control of this foodborne pathogen. [ABSTRACT FROM AUTHOR]

The COVID-19 era brought about new medical challenges, which, together with nosocomial bacterial infections, resulted in an enormous burden for the healthcare system. One of the most alarming nosocomial threats was carbapenem-resistant Klebsiella pneumoniae (CRKP). Monitoring CRKP incidence and antimicrobial resistance globally and locally is vitally important. In a retrospective study, the incidence of CRKP in the pre-COVID-19 period (2017–2019) and the COVID-19 pandemic (2020–2022) was investigated in the Central Military Hospital in Ružomberok, Slovak Republic. The relative incidence of CRKP significantly increased during the COVID-19 period—by 4.8 times, from 0.18 to 0.76%. At the same time, 47% of CRKP-positive patients also had COVID-19. Twenty-six KPC and sixty-nine NDM-producing isolates were identified. CRKPs isolated in the year 2022 were submitted to whole genome sequencing, and their susceptibility was tested to cefiderocol, ceftazidime–avibactam, imipenem–relebactam and meropenem–vaborbactam, with excellent results. KPC-producing isolates were also highly susceptible to colistin (92%). The NDM isolates revealed lower susceptibility rates, including only 57% colistin susceptibility. ST-307 prevailed in KPC and ST-11 in NDM isolates. Despite the excellent activity of new antimicrobials, rational antibiotic policy must be thoroughly followed, supported by complementary treatments and strict anti-epidemic precautions. [ABSTRACT FROM AUTHOR]

The terminal restriction fragment length polymorphism munities from different collecting places was evaluated was used to determine the bacterial diversity in rhizo- by principal component analysis. Results showed that sphere of maize (Zea mays L.) collected from four sites the most different bacterial community originated from of experimental field plot in two dates of the vegetation marginal part of the experimental field plot collected in season (July and September). The 16S rRNA gene was September was caused probably by combination of the amplified from metagenomic DNA using universal eubac- marginal effect and drought before sampling date in Sep- terial primers and PCR products were digested separately tember. Other rhizosphere samples showed from moderate with three restriction enzymes. Significant differences in to small differences in the structure of the bacterial com- the number of terminal restriction fragments among rhi- munity. Nevertheless, significant differences among all zosphere samples and between sampling dates were not collected bacterial communities were not observed. detected (P < 0.05). Variation within the bacterial communities from different collecting places was evaluated by principal component analysis. Results showed that the most different bacterial community originated from marginal part of the experimental field plot collected in September was caused probably by combination of the marginal effect and drought before sampling date in September. Other rhizosphere samples showed from moderate to small differences in the structure of the bacterial community. Nevertheless, significant differences among all collected bacterial communities were not observed.

The expansion of sequence type 131 (ST131) extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (E. coli) represents major worldwide challenges. E. coli strains originating from healthcare facilities (labeled No. 1 and No. 2) of the University Hospital Bratislava (UHB) were analyzed for ST131 emergence, including its (sub)lineages and clonal relatedness. Antimicrobial resistance was determined in most strains. Of a total of 354 E. coli strains, 263 (74.3%) belonged to ST131; of these, 177 (67.3%) were from No. 1. Generally, among 260 ST131 E. coli, clades A/B were confirmed in 20 (7.7%), while clade C was noted in 240 (92.3%) strains; within them, subclades were detected as follows: C0 (17; 7.1%), C1 (3; 1.2%), and C2 (220; 91.7%). Among fifteen randomly selected E. coli strains that were investigated for ST and clonal relatedness, seven STs were identified: eight (53.3%) ST131, two (13.3%) ST73, and one each (6.7%) of ST10, ST12, ST14, ST1193, and ST1196. From No. 1, two ST131 in the first internal clinic and one ST131 from No. 2 in the aftercare department were highly clonally related, suggesting possible epidemiological association. Antimicrobial resistance was as follows: ciprofloxacin 93.8%, ceftazidime 78.4%, meropenem 0%, fosfomycin 2.9% and nitrofurantoin 1.4%. Prevention of ESBL-producing E. coli dissemination, especially for ST131 clade C2, is inevitably necessary for reducing drug resistance and decreasing healthcare-associated infections. [ABSTRACT FROM AUTHOR]

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Carbapenem-resistant (CR) Klebsiella pneumoniae represents an urgent worldwide threat. We focused on CR K. pneumoniae in selected facilities of the University Hospital Bratislava (UHB) to investigate sequence types (STs), clonal relatedness, and antimicrobial resistance. Suspected carbapenem-nonsusceptible K. pneumoniae strains were obtained from hospitalized patients. Further examination included carbapenemase confirmation, cgMLST, and quantitative susceptibility testing. Of the total 41 CR K. pneumoniae strains, 26 (63.4%) were determined as ST11 in hospital No. 1; of these ST11, 22 (84.6%) were found in the first internal clinic. Six (14.6%) ST258 and three (7.3%) ST584 occurred in hospital No. 2; the most, i.e., four (66.7%), ST258 were detected in the geriatric clinic. In hospital No. 3, we found two (4.8%) ST584 and one (2.4%) ST258. Of the ST11 set, 24 (92.3%) produced NDM-1. Furthermore, seven (87.5) ST258 and five (100%) ST584 strains generated KPC-2. Antimicrobial resistance was as follows: ertapenem 97.6%, meropenem 63.4%, tigecycline 7.3%, eravacycline 7.3%, and colistin 2.5%. We revealed a presumably epidemiological association of ST11 with transmission, particularly in the first internal clinic of hospital No. 1, while ST258 and ST584 were related to interhospital dissemination between medical facilities No. 2 and No. 3. It is essential to prevent the circulation of these pathogens within and between healthcare facilities. [ABSTRACT FROM AUTHOR]

We present a biological profile of 16 Aspergillus niger environmental isolates from different types of soils and solid substrates across a pH range, from an ultra-acidic (<3.5) to a very strongly alkaline (>9.0) environment. The soils and solid substrates also differ in varying degrees of anthropic pollution, which in most cases is caused by several centuries of mining activity at old mining sites, sludge beds, ore deposits, stream sediments, and coal dust. The values of toxic elements (As, Sb, Zn, Cu, Pb) very often exceed the limit values. The isolates possess different macro- and micromorphological features. All the identifications of Aspergillus niger isolates were confirmed by molecular PCR analysis and their similarity was expressed by RAMP analysis. The biochemical profile of isolates based on FF-MicroPlate tests from the Biolog system showed identical biochemical reactions in 50 tests, while in 46 tests the utilisation reactions differed. The highest similarity of strains isolated from substrates with the same pH, as well as the most suitable biochemical tests for analysis of the phenotypic similarity of isolated strains, were confirmed when evaluating the biochemical profile using multicriterial analysis in the Canoco program. The isolates were screened for mycotoxin production by thin-layer chromatography (TLC), as well. Two of them were able to synthesise ochratoxin A, while none produced fumonisins under experimental conditions. Presence of toxic compounds in contaminated sites may affect environmental microscopic fungi and cause the genome alteration, which may result in changes of their physiology, including the production of different (secondary) metabolites, such as mycotoxins. [ABSTRACT FROM AUTHOR]

This work was aimed at determining the occurrence and diversity of Listeria monocytogenes in a traditional meat-processing facility to reveal persistent contamination. A total of 268 samples, including 196 environmental samples and 72 meat samples were collected during a four-year period, and 70 were found to be L. monocytogenes positive. Molecular serotyping of 77 isolates classified these strains into four serogroups, with the majority of 34 strains in serogroup IIa. To reveal the persistent contamination, subtyping by AscI/ApaI-PFGE was applied. Cluster analysis resulted in strain discrimination into 18 profiles. Strains with PFGE/MLST-ST classification as 1/ST9, 2/ST14 and 9/ST2 were predominant and considered persistent. Based on the results of PFGE, a spatial and temporal map of persistent L. monocytogenes contamination was drawn. Subtyping by PFGE was used as a reliable tool to reveal persistent contamination and, complemented with virulence characteristics, can contribute to the improvement of L. monocytogenes control programs in food processing. Practical application Listeria monocytogenes is a food-borne pathogen with significant public health and economic impact. This study, focused on prevalence and tracing of L. monocytogenes strains in a traditional meat-processing facility, revealed persistent contamination by the pathogen. Data of PFGE profiling from repeated samplings over a four-year period facilitated to draw a spatial and temporal map of persistent L. monocytogenes strains spreading in a meat-processing facility and, subsequently, will allow optimisation of cleaning and disinfection measures to ensure hygiene. Based on the detection of virulence factors, persistent ECIII and comK prophage-positive strains were observed, indicating a higher risk of contamination by more virulent L. monocytogenes strains. The obtained results clearly indicate, that most attention should be paid to control strategy improvement in the meat processing chain focused preferably on prevention, not only to final products. [ABSTRACT FROM AUTHOR]

Overall cell surface hydrophobicity (CSH) is predicted to play an important role during biofilm formation in Candida albicans but is the result of many expressed proteins. This study compares the CSH status and CSH1 gene expression in C. albicans planktonic cells, sessile biofilm, and dispersal cells. Greater percentages of hydrophobic cells were found in non-adhered (1.5 h) and dispersal forms (24 or 48 h) (41.34±4.17% and 39.52±7.45%, respectively), compared with overnight planktonic cultures (21.69±3.60%). Results from quantitative real-time PCR confirmed greater up-regulation of the CSH1 gene in sessile biofilm compared with both planktonic culture and dispersal cells. Up-regulation was also greater in dispersal cells compared with planktonic culture. The markedly increased CSH found both in C. albicans biofilm, and in cells released during biofilm formation could provide an advantage to dispersing cells building new biofilm. [ABSTRACT FROM AUTHOR]

Hemp seeds are increasingly produced on small farms in Slovakia and used for production of food. We detected contamination of 1.4 x 107 CFU·g-1 coliform coliform bacteria in hemp seeds from a small farm and isolated three strains of Escherichia coli. These were characterized by GTG-typing and by whole genome sequencing. The three strains were indistinguishable from each other by GTG-typing but different from a reference strain E. coli CCM 2024. They were not distinguished either by multi-locus sequence type (MLST) deduced from whole genome sequences, as all strains were of sequence type 58, or by core genome-based MLST (cgMLST), as all strains belonged to type 148521. All strains were found to be fumC4/fimH32 type and serotype H21, O-antigen was not identified. No plasmid origin was detected. No gene encoding for shiga toxin was detected. Several virulence-associated genes were detected in all three strains at high identities with sequences in the database, namely, fimH, lpfA, terC, csgA, hlyE, nlpI, yehB, yehC and yehD. Further virulence-associated genes were detected at shorter lengths, namely, fdeC, gad and yehA. Several antibiotic resistance genes were also identified. The strains were found to be virtually identical. With a probability of being a human pathogen, they need to be addressed for on-farm food production safety. [ABSTRACT FROM AUTHOR]

Seven variable-number tandem repeat (VNTR) markers, which had previously proven to be suitable for typing of Salmonella enterica serovar Typhimurium or S. enterica serovar Enteritidis, were evaluated for their discrimination power in multilocus VNTR analysis (MLVA) on a panel of 83 S. enterica strains belonging to 23 different serovars, which had been isolated from food and animal samples. Strains varied not only in numbers of tandem repeats, but also in the presence or absence of VNTR alleles. The greatest polymorphism was observed in STTR5, STTR3 and STTR7 loci. Based on combined allelic profiles of all seven VNTR loci, it was possible to distinguish as many as 50 different MLVA profiles. The MLVA clustering of strains parallelled serotyping results to a reasonable extent. In addition, a high degree of intra-serovar variability was observed in strains of serovars Bareilly, Bredeney, Dublin, Enteritidis, Havana, Saintpaul, Senftenberg, Schwarzengrund and Typhimurium. MLVA proved to be more discriminative than dnaN housekeeping gene sequencing. The optimized MLVA method can be advantageously used in tracing of Salmonella contamination of food products. [ABSTRACT FROM AUTHOR]

Lysogenic bacteriophages are a significant source of variability in closely related Salmonella strains. In this study, screening for diversity of 152 Salmonella Typhimurium strains was performed using PCR detection of selected prophage regions derived from phages P22, Gifsy-1, Gifsy-2, Fels-1, ST104 and SopEΦ. A high degree of variability was observed in the presence of specific genes. Based on the presence of particular prophage genes, we divided strains into 37 different PCR-prophage profiles; 20 of them were represented by only a single strain. Using multilocus variable number tandem repeats analysis (MLVA), 152 Salmonella strains were separated into 82 MLVA strings. Similar grouping of Salmonella strains was observed in the case of PCR-prophage detection and MLVA and the results corresponded well with the phage type of strains. However, several Salmonella strains were detected, which were closely related according to MLVA; yet, they differed in PCR phage profiles. The observations support a view that integration/excision of bacteriophages in Salmonella strains are frequent events shaping the bacterial genome. [ABSTRACT FROM AUTHOR]

Microbial contamination of oil seeds can cause their deterioration leading to reduction of the quality of the produced oil. Knowledge of the levels and composition of microbial communities may be therefore helpful to set up the effective control measures. Seeds of hemp and poppy (blue and white cultivars) from two farms in Slovakia were analysed by culture-based microbiological methods and by metagenomic high-throughput sequencing of 16S rDNA and 28S rDNA to provide information on surface bacterial and fungal communities. The results of culture-based analysis showed a comparatively high level of contamination by bacteria and by fungi of hemp seeds. High-throughput sequencing revealed that the bacterial community of hemp seeds consisted mainly of families Enterobacteriaceae and Pseudomonadaceae. The fungal community was composed mainly of Pleosporaceae, Davidiellaceae and Tremellaceae. In poppy seeds, the bacterial community was found to be composed mainly of Enterobacteriaceae (genus Klebsiella), Pseudomonadaceae and Xanthomonadaceae or Streptomycetaceae. The fungal community was composed mainly of Pleosporaceae and Davidiellaceae (genus Cladosporium), while some samples were characterized by increased levels of Trichocomaceae (genus Aspergillus). The determined presence of pseudomonads and aspergilli in certain seeds may indicate that more effective cleaning and sanitation will be required as they could affect the produced oil. [ABSTRACT FROM AUTHOR]

Urinary tract infections (UTIs) are among the events that most frequently need medical intervention. Uropathogenic Escherichia coli are frequently their causative agents and the infections are sometimes complicated by the presence of polyresistant nosocomial strains. Phage therapy is a tool that has good prospects for the treatment of these infections. In the present study, we isolated and characterized two bacteriophages with broad host specificity against a panel of local uropathogenic E. coli strains and combined them into a phage cocktail. According to genome sequencing, these phages were closely related and belonged to the Tequatrovirus genus. The newly isolated phages showed very good activity on a panel of local clinical E. coli strains from urinary tract infections. In the form of a two-phage cocktail, they were active on E. coli strains belonging to phylogroups B2 and D, with relatively lower activity in B1 and no response in phylogroup A. Our study is a preliminary step toward the establishment of a national phage bank containing local, well-characterized phages with therapeutic potential for patients in Slovakia. [ABSTRACT FROM AUTHOR]

Euglena gracilis is a fresh-water flagellate possessing secondary chloroplasts of green algal origin. In contrast with organisms possessing primary plastids, mRNA levels of nucleus-encoded genes for chloroplast proteins in E. gracilis depend on neither light nor plastid function. However, it remains unknown, if all these mRNAs are trans-spliced and possess spliced leader sequence at the 5′-end and if trans-splicing depends on light or functional plastids. This study revealed that polyadenylated mRNAs encoding the chloroplast proteins glyceraldehyde-3-phosphate dehydrogenase ( GapA), cytochrome f ( PetA), and subunit O of photosystem II ( PsbO) are trans-spliced irrespective of light or plastid function. [ABSTRACT FROM AUTHOR]

Genomes of 21 strains of lactic acid bacteria isolated from Slovakian traditional cheeses were sequenced on an Illumina MiSeq platform. Subsequently, they were analysed regarding taxonomic classification, presence of genes encoding defence systems, antibiotic resistance and production of biogenic amines. Thirteen strains were found to carry genes encoding at least one bacteriocin, 18 carried genes encoding at least one restriction–modification system, all strains carried 1–6 prophages and 9 strains had CRISPR-Cas systems. CRISPR-Cas type II-A was the most common, containing 0–24 spacers. Only 10% spacers were found to be homological to known bacteriophage or plasmid sequences in databases. Two Enterococcus faecium strains and a Lactococcus lactis strain carried antibiotic resistance genes. Genes encoding for ornithine decarboxylase were detected in four strains and genes encoding for agmatine deiminase were detected in four strains. Lactobacillus paraplantarum 251 L appeared to be the most interesting strain, as it contained genes encoding for two bacteriocins, a restriction–modification system, two CRISPR-Cas systems, four prophages and no genes connected with antibiotic resistance or production of biogenic amines. [ABSTRACT FROM AUTHOR]

Lactobacillaceae are candidates for starter or adjunct cultures for the production of bryndza cheese with traditional organoleptic properties, in particular well adapted strains isolated from bryndza cheese or from ewes' whey. Whole genome sequencing of 34 such strains was carried out, their taxonomic classification was done and the genomes were analysed for the presence of genes responsible for antibiotic resistance or production of biogenic amines. Taxonomic classification of the strains was achieved using several bioinformatic tools, with most weight assigned to Type Genome Server (TYGS). Based on this, qualified presumption of safety (QPS) status as defined by European Food Safety Authority could be assigned to 25 strains, which belonged to species Lacticaseibacillus paracasei/casei, Limosilactobacillus fermentum, Lactobacillus helveticus, Lactiplantibacillus plantarum and Leuconostoc lactis. None of the strains was found to harbour in the genome any acquired antimicrobial resistance genes to clinically relevant antimicrobials. None of the strains contained genes conferring production of the two most hazardous biogenic amines, histamine and tyramine. All strains were predicted to be non-pathogenic for humans by PathogenFinder (Technical University of Denmark, Copenhagen, Denmark). Based on the results, 25 out of 34 tested strains fulfilled safety requirements for the development of starter or adjunct cultures for the production of bryndza cheese with traditional organoleptic properties. [ABSTRACT FROM AUTHOR]

Listeria monocytogenes is an important foodborne pathogen ubiquitous in environment with a potential to colonize food production facilities for long periods. In the present study, diversity of L. monocytogenes isolates from two facilities processing food of animal origin in Slovakia was evaluated using several molecular typing methods. Fifty-five isolates clustered by pulsed-field gel electrophoresis (PFGE) to 15 pulsotypes in our previous studies were subjected to multiple locus variable-number tandem repeat analysis (MLVA) resulting in strain discrimination into 16 MLVA profiles. For these L. monocytogenes isolates, MLVA showed a slightly higher discriminatory power compared to PFGE when one PFGE cluster was divided into two MLVA clusters. Multi-locus sequence typing (MLST) based on whole genome sequencing (WGS) separated 35 L. monocytogenes isolates into 13 sequence types (ST) and 16 groups covering isolates with less than 10 allelic differences in core genome (cg) MLST analysis, five of them containing more than one isolate. PFGE and MLVA allowed reliable primary discrimination of L. monocytogenes isolates. WGS data allowed the comparison of genome diversity required for confirmation of persistent L. monocytogenes. [ABSTRACT FROM AUTHOR]

Cronobacter spp. are responsible for rare but fatal cases of infection in neonates and immunocompromised persons. These bacteria are ubiquitous in the environment and were also isolated from various foods, the powdered infant formula being the main vehicle for its transmission in neonatal infections. In the present study, effects of four bacteriophages against Cronobacter in liquid media and in biofilm were assessed. Ten selected Cronobacter isolates belonging to four species showed variable inhibition by phages in minimal medium with maltose and in powdered infant formula. Bacteriophages significantly decreased growth and biofilm production of the bacterial cultures in maltose minimal medium, the highest effect being observed in the phage cocktail samples. However, the efficiency of bacteriophages for biofilm removal in powdered infant formula was limited only to three strains. Our results will facilitate development of new strategies for Cronobacter control in food and food processing factories. [ABSTRACT FROM AUTHOR]

Polymerase chain reaction (PCR)-based systems were developed to detect the genes prtP, pepN, pepX and bcaT, which are involved in production of aroma-active compounds, and their presence was investigated in 17 strains of lactic acid bacteria isolated from ewes' raw milk-based cheeses. Using the developed PCR systems, along with the previously published PCR systems specific for Lactococcus-derived genes, five strains were found to contain all targeted genes (three Lactobacillus paracasei strains, one Lactobacillus casei and one Lactococcus lactis). In 14 Lactobacillus strains, prtP was detected in 5 strains, pepN in 14 strains, pepX in 12 strains and bcaT in 10 strains. Effectivity of the performance of the PCR systems was checked with whole-genome sequences of the strains, which showed that all strains containing sequences homological (at ≥ 90 %) to primers were correctly amplified. Whole-genome sequences of the strains were used also to determine the representativeness of the PCR systems, which showed that individual genes were correctly detected in all Lactobacillus and Lactococcus strains by at least one PCR system. Knowledge on the presence of protease genes in bacterial strains is the basis for studying their expression and involvement in formation of volatile aroma-active compounds during cheese ripening. [ABSTRACT FROM AUTHOR]

The aim of the study was to investigate the occurence and diversity of Listeria monocytogenes in three ewes' milk processing factories in Slovakia, and to reveal its sources. A total of 639 samples from the cheese production chain were collected during the years 2011-2014, out of which 20 (3.1%) were found to be positive for L. monocytogenes. L. ivanovii was detected in two samples and other Listeria species in 309 (48.4%) samples. Sporadic L. monocytogenes contamination was observed in all three factories with 10% positive samples being the products, while 90% of positive samples were associated with the production environment. By molecular serotyping using multiplex polymerase chain reaction, the L. monocytogenes isolates were classified in three serogroups - 80% in serogroup IIa, 10% in serogroup IIc and 10% in serogroup IVb. Testing for markers specific for three epidemic clones (ECI-ECIII) produced negative results. Molecular diversity evaluation of isolated strains by AscI-restriction and pulsed-field gel electrophoresis (PFGE) resulted in strain discrimination into 14 clusters at 100% similarity. The observed occasional incidence of L. monocytogenes and high strain diversity indicated the origin of contamination from external sources. [ABSTRACT FROM AUTHOR]

Cronobacter spp. is responsible for rare but fatal cases of infections in neonates and immunocompromised infants. The reconstituded powdered infant formula (PIF) is one of main sources of newborn infections. The aim of our study was to assess application of recombinant human cathelicidin LL-37, an antimicrobial peptide, alone and in combination with bacteriophages against Cronobacter in PIF. The minimal inhibitory concentration (MIC) of LL-37 determined by radial diffusion method for ten representative Cronobacter strains ranged from 2.56 µg⋅ml-1 to 8.83 µg⋅ml-1, and the values were below MIC of E. coli standard strain. By testing the antimicrobial activity in liquid growth medium, substantial inhibition of strains was observed at 30 µg⋅ml-1 LL-37, whereas 20 µg⋅ml-1 showed only moderate effect. However, higher peptide concentrations were necessary to inhibit Cronobacter in reconstituted PIF. By using 50 µg⋅ml-1 and 100 µg⋅ml-1 LL-37, numbers of cells decreased after 4-hour cultivation by approx. 70% and 96%, respectively. Finally, simultaneous application of LL-37 and bacteriophage Dev-CS-701 resulted in synergistic effect, as a 1 300-fold reduction of cell numbers in PIF was observed. Application of the two antimicrobial agents thus facilitated their reduced dosing and decreased the probability of generation of phage-resistant cells. [ABSTRACT FROM AUTHOR]

Development of genetically modified (GM) food has generated worldwide public controversy over food safety issues. To assess the food safety aspects and potential risk of horizontal gene transfer (HGT) from GM plants and food derived thereof, we have chosen aroA CP4 coding EPSPS (5-enolpyruvylshikimate-3-phosphate synthase) from GM maize. During the study of possible HGT of aroA CP4 gene from GM food, we have discovered and characterized truncated form of aroA CP4 within the cloning experiments. Comparison of properties of recombinant E. coli strains with both forms of CP4 EPSPS gene were performed on DNA and protein levels. We report on cloning, expression, purification and enzyme activity studies of both genes. The specific activities of both enzymes were indistinguishable indicating that the truncated CP4 EPSPS enzyme retained the activity of a full-length CP4 EPSPS. The in vitro growth test showed that both forms of aroA CP4 genes grew well at concentrations of 10-40 mmol⋅l-1 glyphosate and the in vivo complementation test revealed uniform growth. The results indicate that both full-length and truncated aroA CP4 have similar characteristics (full length CP4 EPSPS specific activity 0.063 ± 0.01 µmol⋅min-1⋅mg-1; truncated CP4 EPSPS specific activity 0.055 ± 0.01 µmol⋅min-1⋅mg-1). These results should also be taken into account in risk analysis of possible unintended effects from GM plants. [ABSTRACT FROM AUTHOR]

Numerous factors associated with sample preparation, DNA extraction, primer choice, sequencing platform and data analysis can affect the accuracy of 16S rRNA sequencing results. The DNA extraction method is considered critical for the success of sequencing as it can be the source of considerable variations in the analysis of the microbiome. In this study, the impact of various DNA extraction methods on the results of analysis of bacterial communities in cheese was evaluated. DNA was isolated from Mozzarella as a model cheese using optimized bead-based homogenization followed by different extraction procedures. Five commercial kits and two open-formula DNA extraction protocols were evaluated for amplicon sequencing of a 16S rRNA fragment of ~1460 bp. In addition, model cheese samples artificially contaminated by defined concentrations of Listeria monocytogenes and Escherichia coli , as representatives of Gram positive and Gram negative bacteria, were analysed. Six out of seven DNA extraction procedures were found to be able to provide amplifiable bacterial DNA suitable for 16S rRNA sequence analysis, but individual extraction procedures led to variable results. In particular, lysis supported with bead-beating led to a higher proportion of G+ bacteria in relative abundance profiles, probably because of the more efficient cell wall disruption. Artificially added bacterial species were reliably detected with a quantitative response. The results demonstrated a risk in comparing the data on bacterial communities in cheese when different DNA extraction protocols are used and highlighted the need to choose a standardized approach when comparison across multiple sequencing runs is required. • DNA extraction method is crucial for the success of 16S rRNA sequencing analysis. • Individual extraction procedures led to variable results of microbiome profiling. • Mechanically assisted lysis promoted higher representation of G+ bacteria. • Artificially added bacteria were detected with quantitative response. [ABSTRACT FROM AUTHOR]

This paper investigates the differences in some metabolites using Biolog FF Microplate™ system and the production of organic acids such as β-hydroxybutyric, p-hydroxyphenylacetic, and others. Another group of organic acids such as gluconic, oxalic and citric acid were studied during cultivation in a liquid medium. Four different Aspergillus niger (An) wild type strains were used as a model organism. Three strains, from the Banská Štiavnica – Šobov (An – Š), Pezinok (An – P) and Slovinky (An – Sl) localities were isolated from contaminated old mining areas with soil with ultra acidic to strong alkaline reactions. The fourth strain isolated from the Gabčíkovovo (An – G) locality was used for comparative purposes. According to the RAMP analysis, the strains are clustered into two groups, An – Š and An – P (similarity 82%), An – G and An – Sl (similarity 64%) which correlates with the pH values of the original environment. However, significant differences were found in metabolic processes in the reaction with a wide range of organic acids. In general, the reactions with D-lactic acid and D-malic acid correlate with the results of the RAMP analysis of fungal genotype similarities, the An – Š and An – P strains had an identical negative reaction, and an identical positive reaction was found in the An – Sl and An – G strains. During incubation the wild-type strains produced substantial amounts of gluconic acid, oxalic acid and small amounts of citric acid. The appearance and accumulation of organic acids was found to be highly pH dependent with the most active strain isolated from an ultra-acidic environment. The comparative strain differs entirely in the production of oxalic acid.This paper investigates the differences in some metabolites using Biolog FF Microplate™ system and the production of organic acids such as β-hydroxybutyric, p-hydroxyphenylacetic, and others. Another group of organic acids such as gluconic, oxalic and citric acid were studied during cultivation in a liquid medium. Four different Aspergillus niger (An) wild type strains were used as a model organism. Three strains, from the Banská Štiavnica – Šobov (An – Š), Pezinok (An – P) and Slovinky (An – Sl) localities were isolated from contaminated old mining areas with soil with ultra acidic to strong alkaline reactions. The fourth strain isolated from the Gabčíkovovo (An – G) locality was used for comparative purposes. According to the RAMP analysis, the strains are clustered into two groups, An – Š and An – P (similarity 82%), An – G and An – Sl (similarity 64%) which correlates with the pH values of the original environment. However, significant differences were found in metabolic processes in the reaction with a wide range of organic acids. In general, the reactions with D-lactic acid and D-malic acid correlate with the results of the RAMP analysis of fungal genotype similarities, the An – Š and An – P strains had an identical negative reaction, and an identical positive reaction was found in the An – Sl and An – G strains. During incubation the wild-type strains produced substantial amounts of gluconic acid, oxalic acid and small amounts of citric acid. The appearance and accumulation of organic acids was found to be highly pH dependent with the most active strain isolated from an ultra-acidic environment. The comparative strain differs entirely in the production of oxalic acid.This paper investigates the differences in some metabolites using Biolog FF Microplate™ system and the production of organic acids such as β-hydroxybutyric, p-hydroxyphenylacetic, and others. Another group of organic acids such as gluconic, oxalic and citric acid were studied during cultivation in a liquid medium. Four different Aspergillus niger (An) wild type strains were used as a model organism. Three strains, from the Banská Štiavnica – Šobov (An – Š), Pezinok (An – P) and Slovinky (An – Sl) localities were isolated from contaminated old mining areas with soil with ultra acidic to strong alkaline reactions. The fourth strain isolated from the Gabčíkovovo (An – G) locality was used for comparative purposes. According to the RAMP analysis, the strains are clustered into two groups, An – Š and An – P (similarity 82%), An – G and An – Sl (similarity 64%) which correlates with the pH values of the original environment. However, significant differences were found in metabolic processes in the reaction with a wide range of organic acids. In general, the reactions with D-lactic acid and D-malic acid correlate with the results of the RAMP analysis of fungal genotype similarities, the An – Š and An – P strains had an identical negative reaction, and an identical positive reaction was found in the An – Sl and An – G strains. During incubation the wild-type strains produced substantial amounts of gluconic acid, oxalic acid and small amounts of citric acid. The appearance and accumulation of organic acids was found to be highly pH dependent with the most active strain isolated from an ultra-acidic environment. The comparative strain differs entirely in the production of oxalic acid. [ABSTRACT FROM AUTHOR]

Abstract Salmonella 4,[5],12:i:- has become the third most common serotype in Europe, including the Czech Republic. In this study, phenotypic and genotypic methods for a more detailed description of this serotype were used. Analysis of a limited number of isolates revealed that 76% of them belonged to phage type DT193. Also, rare phage types DT208 and U311 were identified. In total, 88.6% of the isolates were resistant to at least four antimicrobial agents. In this study, 24 multilocus variable-number tandem-repeat analysis profiles were detected, and some of them matched with the profiles recently described in Europe.