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Vitamin B12 is a critical nutrient in vegan and vegetarian lifestyles as plant-based vitamin sources are rare. Traditional fermented foods could be enriched by adding vitamin B12-producing bacteria to offer non-animal vitamin sources. The aim was to isolate a vitamin B12 producer that is capable of producing the human-active vitamin even at low pH values so that it can be used in fruit juice fortification. Therefore, fermented foods (homemade and industrial) and probiotics were screened for vitamin B12 production strains. A modified microbiological vitamin B12 assay based on Lactobacillus delbrueckii subsp. lactis DSM 20355 was used to identify vitamin B12-containing samples and the presence of vitamin B12-producing strains. The screening resulted in isolating several positive strains for vitamin B12 formation derived from sourdough. Mass spectrometry confirmed the biosynthesis of solely the human physiologically active form. Species identification carried out by the German Strain Collection of Microorganisms and Cell Cultures resulted in two species: Acetobacter orientalis and Acetobacter malorum, of which two isolates were further characterised. The potential for cobalamin biosynthesises in food matrixes was demonstrated for A. malorum HFD 3141 and A. orientalis HFD 3031 in apple juice at different pH values (2.85–3.80). The isolates synthesised up to 18.89 µg/L and 7.97 µg/L vitamin B12 at pH 3.80. The results of this study suggest that acetic acid bacteria (AAB) and fermented acetic acid foods are promising resources for vitamin B12 and its producers, which might have been overlooked in the past.

This study explores the development of whole-grain sourdough bread with reduced FODMAP (fermentable oligosaccharides, disaccharides, monosaccharides, and polyols) content to offer dietary solutions for individuals with irritable bowel syndrome (IBS). Three sourdough breads were prepared using different lactic acid bacteria (LAB) strains including Lactiplantibacillus plantarum FST1.7 (SD-FST1.7), Lacticaseibacillus paracasei R3 (SD-R3), and Pediococcus pentosaceus RYE106 (SD-RYE106). A control sourdough bread was prepared using baker’s yeast (SD-control). In vitro digestion and in vitro colonic fermentation were employed on bread samples with cellulose (negative control) and inulin (positive control), followed by 16S rRNA sequencing and short-chain fatty acid (SCFA) analysis to evaluate the impact on gut microbiota and SCFA levels. Alpha and beta diversity did not reveal any significant differences within the groups following in vitro colonic fermentation (FDR > 0.05). Taxonomic analysis displayed Firmicutes as the predominant phylum across all fecal samples at the end of colonic fermentation. Actinobacteriota was significantly lower in cellulose fermented fecal samples compared to samples fermented with SD-Control (ANCOMBC, FDR = 0.02) and inulin (ANCOMBC, FDR = 0.0001). Fecal samples fermented with inulin had significantly higher Bacteroidota levels compared to those fermented with cellulose (ANCOMBC, FDR =0.002). Acetate levels were higher in fecal samples fermented with SD-FST1.7 compared to those fermented with SD-R3 and SD-RYE106 (p = 0.03 for both). Positive correlations between butyrate and Lachnospira, Agathobacter, and Bifidobacterium were observed, demonstrating the potential of sourdough fermentation to influence gut health and support IBS management.

Recently, case studies have been published regarding the application of mycobacteriophage (MP) therapy (MPT) in patients with multi-antibiotic-resistant infections. A major limitation in the development of MPT is the paucity of therapeutically useful MP. As there are approximately 10,000 MP that have yet to be sequenced, it is possible that characterization of this cohort would increase the repertoire of useful MP. This study aims to contribute to such a strategy, by characterizing a cohort of 7 mycobacteriophages. Sequencing analyses revealed that the MP have unique sequences, and subsequent gene annotation revealed differences in gene organization. Notably, MP LOCARD has the largest genome and operons encoding for glycosyltransferases. Taxonomic analysis executed with VIRIDIC, Gegenees and VICTOR revealed that LOCARD belongs to a different genus than the other phages and is the foundational member of one of three novel species identified in this study. LOCARD, LOCV2, and LOCV5 were selected as representative members of their species and subjected to phenotypic analyses to compare their stability under biologically and industrially relevant conditions. Again LOCARD stood out, as it was unaffected by the typical temperatures (37 °C) and salinity (0.9%) experienced in mammals, while the viability of LOCV2 and LOCV5 was significantly reduced. LOCARD was also tolerant to pH 10, low levels of antiviral detergent and was the least impacted by a single freeze-thaw cycle. When all these results are considered, it indicates that LOCARD in particular, has potential therapeutic and/or diagnostics applications, given its resilience towards physiological and storage conditions.

Staphylococcus aureus is frequently highlighted as a priority for novel drug research due to its pathogenicity and ability to develop antibiotic resistance. Coagulase-negative staphylococci (CoNS) are resident flora of the skin and nares. Previous studies have confirmed their ability to kill and prevent colonization by S. aureus through the production of bioactive substances. This study screened a bank of 37 CoNS for their ability to inhibit the growth of methicillin-resistant S. aureus (MRSA). Deferred antagonism assays, growth curves, and antibiofilm testing performed with the cell-free supernatant derived from overnight CoNS cultures indicated antimicrobial and antibiofilm effects against MRSA indicators. Whole genome sequencing and BAGEL4 analysis of 11 CoNS isolates shortlisted for the inhibitory effects they displayed against MRSA led to the identification of two strains possessing complete putative bacteriocin operons. The operons were predicted to encode a nukacin variant and a novel epilancin variant. From this point, strains Staphylococcus hominis C14 and Staphylococcus epidermidis C33 became the focus of the investigation. Through HPLC, a peptide identical to previously characterized nukacin KQU-131 and a novel epilancin variant were isolated from cultures of C14 and C33, respectively. Mass spectrometry confirmed the presence of each peptide in the active fractions. Spot-on-lawn assays demonstrated both bacteriocins could inhibit the growth of an MRSA indicator. The identification of natural products with clinically relevant activity is important in today’s climate of escalating antimicrobial resistance and a depleting antibiotic pipeline. These findings also highlight the prospective role CoNS may play as a source of bioactive substances with activity against critical pathogens.

Abstract Bacterial vaginosis (BV) is a recurrent dysbiosis that is frequently associated with preterm birth, increased risk for acquisition of human immunodeficiency virus (HIV) and other sexually transmitted infections (STIs). The overgrowth of a key pathobiont, Gardnerella vaginalis, as a recalcitrant biofilm is central to the development of this dysbiosis. Overgrowth of vaginal biofilms, seeded by initial G. vaginalis colonization, leads to recurrent symptomatic BV which is poorly resolved by classically used antibiotics. In this light, the use of bacteriophages and/or their proteins, represents a promising alternative. Here we identify 84 diverse anti-Gardnerella endolysins across 7 protein families. A subset of 36 endolysin candidates were refactored and overexpressed in an E. coli BL21 (DE3) system and 5 biochemically and structurally diverse endolysins were fully characterized. Each candidate endolysin showed good lytic activity against planktonic G. vaginalis ATCC14018, as well as G. vaginalis clinical isolates. These endolysin candidates were assayed in biofilm prevention and disruption assays, with biofilm disruption at low microgram concentrations (5 μg/ml) observed. In addition to clonal G. vaginalis biofilms, endolysin candidates could also successfully disrupt polyspecies biofilms. Importantly, none of our candidates showed lytic activity against commensal lactobacilli present in the vaginal microbiota such as L. crispatus, L. jensenii, L. gasseri, and L. iners or against Atopobium vaginae (currently classified as Fannyhessa vaginae). The potency and selectivity of these novel endolysins constitute a promising alternative treatment to combat BV, avoiding problems associated with antibiotic resistance, while retaining beneficial commensal bacteria in the vaginal flora. The diverse library of candidates reported here represents a strong repository of endolysins for further preclinical development.

Encapsulated medication is a common method of administering therapeutic treatments. As researchers explore alternative therapies, it is likely that encapsulation will remain a feature of these novel treatments, particularly when routes of delivery are considered. For instance, alginate-encapsulation is often favoured where gastric digestion poses an obstacle. When exposed to cations (namely Ca2+), alginate readily forms gels that are resilient to acidic conditions and readily dissociate in response to mid-range pH. This action can be extremely valuable for the encapsulation of phages. The efficient delivery of phages to the intestine is important when considering mycobacteriophage (MP) therapy (or MP prophylaxis) for disseminated mycobacterial infections and chronic gastroenteritis conditions. This study presents the design and in vitro validation of an alginate-encapsulated MP capable of releasing phages in a pH-dependent manner. Ultimately, it is shown that encapsulated phages pretreated with simulated gastric fluid (SGF) are capable of releasing viable phages into simulated intestinal fluid (SIF) and thereby reducing the mycobacterial numbers in spiked SIF by 90%. These findings suggest that alginate encapsulation may be a viable option for therapeutic and prophylactic approaches to the management of intestinal mycobacterial disease, such as Johne’s disease.

Fighting food waste caused by spoilage yeasts and developing more natural forms of preservatives are critical points for the food industry. Plant antimicrobial peptides have biological activity against a broad spectrum of microorganisms, which could represent a valid alternative to the current preservative agents. In this work, the Pharabitis nil antimicrobial peptides Pn-AMP1 and Pn-AMP2 were chemically synthesised, and the potency of their antiyeast activity against common spoilage yeasts was measured. Pn-AMP1 showed a better inhibitory effect than Pn-AMP2, therefore it was further characterised. Pn-AMP1 displayed fast inhibitory activity against yeast strains Kluyveromyces lactis and Zygosaccharomyces bailii, and the antiyeast mechanism of membrane permeabilisation was detected. Its safety in terms of human consumption was studied and no adverse effects were found. Lastly, the stability of the peptide in different environments and conditions, such as heat, high salt, and a range of pH were studied, in addition to their antiyeast activity in different food matrices such as a soft drink and dairy product, further supporting Pn-AMP1’s potential for use in food preservation.

16S rRNA amplicon sequencing was used to investigate changes in the broiler gastrointestinal tract (GIT) microbiota throughout the rearing period and in combination with antibiotic treatment. Thirty birds (from a commercial flock) were removed at multiple points throughout the rearing period on days 13, 27, and 33, euthanised, and their GIT aseptically removed and divided into upper (the crop, proventriculus, and the gizzard), middle (the duodenum, jejunum, and ileum) and lower (the large intestine, the caeca, and the cloaca) sections. In a separate commercial flock, on the same farm with similar husbandry practices and feed, doxycycline (100 mg/ml per kg body weight) was administered in drinking water between day 8 and 12 (inclusive) of the production cycle. Birds were removed on days, 13, 27, and 33 and GIT samples prepared as above. The contents of three merged samples from each GIT section were pooled (n = 60), the DNA extracted and analysed by 16S rRNA amplicon metagenomic sequencing and analysed. Major changes in the broiler microbiota were observed as the birds aged particularly with the Firmicutes/Bacteroidetes ratio (F:B) of the lower GIT. Moreover, Chao1, ACE, and Shannon indices showed the antibiotic treatment significantly altered the microbiota, and this change persisted throughout the rearing period. Further research is required to investigate the effect of these changes on bird performance, susceptibility to infections and Campylobacter carriage.

FODMAPs are fermentable oligo-, di-, monosaccharides, and polyols. The application of homofermentative lactic acid bacteria (LAB) has been investigated as a promising approach for producing low-FODMAP whole-wheat bread. The low-FODMAP diet is recommended to treat irritable bowel syndrome (IBS). Wheat flour is staple to many diets and is a significant source of fructans, which are considered FODMAPs. The reduction of fructans via sourdough fermentation, generally associated with heterofermentative lactic acid bacteria (LAB), often leads to the accumulation of other FODMAPs. A collection of 244 wild-type LAB strains was isolated from different environments and their specific FODMAP utilisation profiles established. Three homofermentative strains were selected for production of whole-wheat sourdough bread. These were Lactiplantibacillus plantarum FST1.7 (FST1.7), Lacticaseibacillus paracasei R3 (R3), and Pediococcus pentosaceus RYE106 (RYE106). Carbohydrate levels in flour, sourdoughs (before and after 48 h fermentation), and resulting breads were analysed via HPAEC-PAD and compared with whole-wheat bread leavened with baker’s yeast. While strain R3 was the most efficient in FODMAP reduction, breads produced with all three test strains had FODMAP content below cut-off levels that would trigger IBS symptoms. Results of this study highlighted the potential of homofermentative LAB in producing low-FODMAP whole-wheat bread.

Proteins and peptides belonging to the plant immune system can possess natural antibacterial, antifungal and antiviral properties. Due to their broad range of activity and stability, they represent promising novel alternatives to commonly used antifungal agents to fight the emergence of resistant strains. An isolation protocol was optimised to target proteins found in plants’ defence system, and it was applied to white mustard (Brassica hirta) seeds. Firstly, a ∼14 kDa protein with activity against S. cerevisiae was extracted and purified; secondly, the protein was identified as the mustard Napin protein named Allergen Sin a 1. Napin is the name given to seed storage (2S) albumin proteins belonging to the Brassicaceae family. While several Napins have been described for their antimicrobial potential, Sin a 1 has been mainly studied for its allergenic properties. The antimicrobial activity of Sin a 1 is described and characterised for the first time in this study; it possesses antifungal and antiyeast in vitro activity, but no antibacterial activity was recorded. The yeasts Zygosaccharomyces bailii Sa 1403 and Saccharomyces cerevisiae DSM 70449 along with the filamentous fungi Fusarium culmorum FST 4.05 were amongst the most senstitive strains to Sin a 1 (MICs range 3–6 μM). The antimicrobial mechanism of membrane permeabilisation was detected, and in general, the antifungal activity of Sin a 1 seemed to be expressed in a dose-dependent manner. Data collected confirmed Sin a 1 to be a stable and compact protein, as it displayed resistance to α-chymotrypsin digestion, heat denaturation and insensitivity to pH variations and the presence of salts. In addition, the protein did not show cytotoxicity towards mammalian cells.

Background: A rapid in-house TM4 mycobacteriophage-based assay, to identify multidrug resistance against various anti-tuberculosis drugs, using the fast-growing Mycobacterium smegmatis mc2 155 in a microtiter plate format was evaluated, based on phage viability assays. Methods: A variety of parameters were optimized before the study including the minimum incubation time for the drugs, phage and M. smegmatis mc2 155 to be in contact. An increase in phage numbers over 2 h was indicative that M. smegmatis mc2 155 is resistant to the drugs under investigation, however when phage numbers remained static, M. smegmatis mc2 155 found to be sensitive to the drug. Results: The study confirmed that the data are statistically significant and that M. smegmatis mc2 155 is, in fact, sensitive to isonazid, iifampicin, pyranzaimide, and ethambutol as phage numbers doubled over 2 h (P = 0.015, 0.018, 0.014, and 0.020). The study also confirmed that M. smegmatis mc2 155 is resistant to the drugs ampicillin, erythromycin, amoxicillin streptomycin as phage numbers remain static over the same 2 h period (P = 0.028, 0.052, 0.049, and 0.04). This drug-susceptibility profiling of eight different drugs against M. smegmatis mc2 155 was detected in as little as 1½ days with a cost of ~ one euro and fifteen cent to test four drugs. Conclusion: This test is rapid to perform and will have widespread applications in drug-susceptibility testing of other members of the mycobacterial genus. In addition, the platform could also be used as a tool for high-throughput screening of novel antimycobacterial drugs. The main assets of this assay include its relatively cheap cost, versatility, and quick turnaround time.

Campylobacter is the most common foodborne pathogen in developed countries and most cases are associated with poultry. This study investigated the effect of three anti-Campylobacter water additives on broiler growth and on the caecal microbiota at harvest using 16S rRNA amplicon sequencing. Mixtures of organic acids (OA) and essential oils (EO) were administered to broilers for the entirety of the production cycle (35 d) and medium-chain fatty acids (MCFA) for 5 d immediately before harvest, under commercial conditions. Bird weight gain was significantly (p < 0.001) reduced in broilers receiving the OA and EO treatments. While this was most likely due to reduced water intake and corresponding lower feed consumption, changes to the caecal microbiota may also have contributed. Firmicutes made up over 75% of the bacteria regardless of sample type, while the minor phyla included Bacteroidetes, Actinobacteria, Melainabacteria, and Proteobacteria. There were no significant (p > 0.05) differences in the alpha diversity as measured using ACE, Chao1, and Shannon indices, except for control (water) versus MCFA and OA versus MCFA, using the Wilcox test. In contrast, there was a significant (p < 0.05) difference in beta diversity when the treated were compared to the untreated control and main flock samples, while linear discriminant analysis effect size (LeFSe) identified three OTUs that were present in the control but absent in the treated birds. It was concluded that the water additives tested adversely affected broiler performance, which may, at least in part, be due to changes in the caecal microbiota, assuming that the altered microbiota at day 35 is indicative of a change throughout the production cycle.

A milk-alternative produced from lentil protein isolate was fermented with three multifunctional strains of lactic acid bacteria, Leuconostoc citreum TR116, Leuconostoc pseudomesenteroides MP070, and Lacticaseibacillus paracasei FST 6.1. As a control, a commercial starter culture containing Streptococcus thermophilus was used. The metabolic performance of these strains and the techno-functional properties of the resulting yogurt alternatives (YA) were studied. Microbial growth was evaluated by cell counts, acidification, and carbohydrate metabolization. The structure of the YA was investigated by textural and rheological analyses and confocal laser scanning microscopy (CLSM). Production of antifungal compounds, the influence of fermentation on the content of FODMAPs, and typical metabolites were analyzed, and a sensory analysis was performed. The results revealed an exponential microbial growth in the lentil base substrate supported by typical acidification, which indicates a suitable environment for the selected strains. The resulting YA showed a gel-like texture typical for non-stirred yogurts, and high water holding capacity. The tested strains produced much higher levels of antifungal phenolic compounds than the commercial control and are therefore promising candidates as adjunct cultures for shelf-life extension. The Leuconostoc strains produced mannitol from fructose and could thus be applied in sugar-reduced YA. Preliminary sensory analysis showed high acceptance for YA produced with Lacticaseibacillus paracasei FST 6.1, and a yogurt-like flavor not statistically different to that produced by the control. Overall, each tested strain possessed promising functionalities with great potential for application in fermented plant-based dairy-alternatives.

Background: Most publicly available genomes of Salmonella enterica are from human disease in the US and the UK, or from domesticated animals in the US. Methods: Here we describe a historical collection of 10,000 strains isolated between 1891-2010 in 73 different countries. They encompass a broad range of sources, ranging from rivers through reptiles to the diversity of all S. enterica isolated on the island of Ireland between 2000 and 2005. Genomic DNA was isolated, and sequenced by Illumina short read sequencing. Results: The short reads are publicly available in the Short Reads Archive. They were also uploaded to EnteroBase, which assembled and annotated draft genomes. 9769 draft genomes which passed quality control were genotyped with multiple levels of multilocus sequence typing, and used to predict serovars. Genomes were assigned to hierarchical clusters on the basis of numbers of pair-wise allelic differences in core genes, which were mapped to genetic Lineages within phylogenetic trees. Conclusions: The University of Warwick/University College Cork (UoWUCC) project greatly extends the geographic sources, dates and core genomic diversity of publicly available S. enterica genomes. We illustrate these features by an overview of core genomic Lineages within 33,000 publicly available Salmonella genomes whose strains were isolated before 2011. We also present detailed examinations of HC400, HC900 and HC2000 hierarchical clusters within exemplar Lineages, including serovars Typhimurium, Enteritidis and Mbandaka. These analyses confirm the polyphyletic nature of multiple serovars while showing that discrete clusters with geographical specificity can be reliably recognized by hierarchical clustering approaches. The results also demonstrate that the genomes sequenced here provide an important counterbalance to the sampling bias which is so dominant in current genomic sequencing.

Bacteriophages (phages) of the genus Kayvirus of Staphylococcus aureus are promising agents for therapeutic applications. In this study, we isolated Kayvirus phages, SAM1 and SAM2, from the Fersisi commercial phage cocktail (George Eliava Institute, Tbilisi, Georgia), which exhibits high sequence homology with phage K (≥94%, BLASTn). We found that phages SAM1 and SAM2 infected 95% and 86% of 21 MRSA of differing sequence types (MLST, SCCmec type) obtained from the Irish National MRSA collection, respectively. We conducted differential transcriptomic analysis by RNA-Seq on phage SAM1 during host infection, showing differential expression of its genes at different points during host infection. This analysis also allowed the identification of potentially adverse outcomes in the application of these phages to target MRSA as therapy. The interaction of phage SAM1 on the host caused the upregulation of prophage genes. Additionally, phage infection was found to cause the slight upregulation of host genes implicated in virulence factors relating to hemolysins, immune evasion, and adhesion, but also the downregulation of genes associated with enterotoxins. The findings of this study give further insights into the biology of kayviruses and their use as therapeutics.

Background: Most publicly available genomes of Salmonella enterica are from human disease in the US and the UK, or from domesticated animals in the US. Methods: Here we describe a historical collection of 10,000 strains isolated between 1891-2010 in 73 different countries. They encompass a broad range of sources, ranging from rivers through reptiles to the diversity of all S. enterica isolated on the island of Ireland between 2000 and 2005. Genomic DNA was isolated, and sequenced by Illumina short read sequencing. Results: The short reads are publicly available in the Short Reads Archive. They were also uploaded to EnteroBase, which assembled and annotated draft genomes. 9769 draft genomes which passed quality control were genotyped with multiple levels of multilocus sequence typing, and used to predict serovars. Genomes were assigned to hierarchical clusters on the basis of numbers of pair-wise allelic differences in core genes, which were mapped to genetic Lineages within phylogenetic trees. Conclusions: The University of Warwick/University College Cork (UoWUCC) project greatly extends the geographic sources, dates and core genomic diversity of publicly available S. enterica genomes. We illustrate these features by an overview of core genomic Lineages within 33,000 publicly available Salmonella genomes whose strains were isolated before 2011. We also present detailed examinations of HC400, HC900 and HC2000 hierarchical clusters within exemplar Lineages, including serovars Typhimurium, Enteritidis and Mbandaka. These analyses confirm the polyphyletic nature of multiple serovars while showing that discrete clusters with geographical specificity can be reliably recognized by hierarchical clustering approaches. The results also demonstrate that the genomes sequenced here provide an important counterbalance to the sampling bias which is so dominant in current genomic sequencing.

Abstract The true prevalence of tet(A), which codes for a tetracycline efflux pump, in thermophilic Camplyobacter spp. requires clarification after reports emerged in Iran (2014) and Kenya (2016) of the novel detection of tet(A) in Campylobacter. During our investigation of antibiotic resistance mechanisms in a sample of Irish thermophilic Campylobacter broiler isolates, it was determined that 100% of tetracycline-resistant isolates (n = 119) harboured tet(O). Accessory tetracycline-resistance mechanisms were considered as tetracycline minimum inhibitory concentrations ranged from 4 to ≥ 64 mg/L. Primers previously reported for the detection of tet(A) in Campylobacter failed to produce an amplicon using a positive control strain (Escherichia coli K12 SK1592 containing the pBR322 plasmid) and a selection of Campylobacter isolates. Accordingly, we designed new tet(A)-targeting primers on SnapGene2.3.2 that successfully generated a 407 bp product from the positive control strain only. Further in silico analysis using BLASTn and SnapGene2.3.2 revealed that previously reported Campylobacter tet(A) sequences deposited on GenBank shared 100% homology with Campylobacter tet(O). We postulate that this gave rise to the erroneous report of a high tet(A) prevalence among a pool of Kenyan broiler Campylobacter isolates that were tested using primers designed based on these apparent tet(A) sequences. In conclusion, further work would be required to determine whether the homology between tet(A) potentially present in Campylobacter and known tet(A) genes would be sufficient to allow amplification using the primers designed in our study. Finally, the existence of tet(A) in thermophilic Campylobacter spp. remains to be demonstrated.

Bacteriophage endolysins and their derivatives have strong potential as antibacterial agents considering the increasing prevalence of antibiotic resistance in common bacterial pathogens. The peptidoglycan degrading peptidase CHAPk, a truncated derivate of staphylococcal phage K endolysin (LysK), has proven efficacy in preventing and disrupting staphylococcal biofilms. Nevertheless, the concentration of CHAPk required to eliminate populations of stationary-phase cells was previously found to be four-fold higher than that for log-phase cells. Moreover, CHAPk-mediated lysis of stationary-phase cells was observed to be slower than for log-phase cultures. In the present study, we report the fusion of a 165 amino acid fragment containing CHAPk with a 136 amino acid fragment containing the cell-binding domain of the bacteriocin lysostaphin to create a chimeric enzyme designated CHAPk-SH3blys in the vector pET28a. The chimeric protein was employed in concentrations as low as 5 μg/mL, producing a reduction in turbidity in 7-day-old cultures, whereas the original CHAPk required at least 20 μg/mL to achieve this. Where 7-day old liquid cultures were used, the chimeric enzyme exhibited a 16-fold lower MIC than CHAPk. In terms of biofilm prevention, a concentration of 1 μg/mL of the chimeric enzyme was sufficient, whereas for CHAPk, 125 μg/mL was needed. Moreover, the chimeric enzyme exhibited total biofilm disruption when 5 μg/mL was employed in 4-h assays, whereas CHAPk could only partially disrupt the biofilms at this concentration. This study demonstrates that the cell-binding domain from lysostaphin can make the phage endolysin CHAPk more effective against sessile staphylococcal cells.

Bacteriophages that lyse Salmonella enterica are potential tools to target and control Salmonella infections. Investigating the host range of Salmonella phages is a key to understand their impact on bacterial ecology, coevolution and inform their use in intervention strategies. Virus–host infection networks have been used to characterize the “predator–prey” interactions between phages and bacteria and provide insights into host range and specificity. Here, we characterize the target-range and infection profiles of 13 Salmonella phage clones against a diverse set of 141 Salmonella strains. The environmental source and taxonomy contributed to the observed infection profiles, and genetically proximal phages shared similar infection profiles. Using in vitro infection data, we analyzed the structure of the Salmonella phage–bacteria infection network. The network has a non-random nested organization and weak modularity suggesting a gradient of target-range from generalist to specialist species with nested subsets, which are also observed within and across the different phage infection profile groups. Our results have implications for our understanding of the coevolutionary mechanisms shaping the ecological interactions between Salmonella phages and their bacterial hosts and can inform strategies for targeting Salmonella enterica with specific phage preparations.

This study investigates the effect of stocking density and population dynamics on broiler growth rates and productivity, while further validating the ability of the biosecurity cubes (BC) to protect birds from Campylobacter. In our methodology, six BC were constructed in a commercial broiler house containing approximately 28,500 birds. During three trials, the BC were stocked at densities of 12, 14, 16, 18, 20 and 22 birds/m2, with the main flock (20 birds/m2) considered the control. Periodically, 10 birds per density were weighed and examined. The Campylobacter status of the birds was monitored via faecal samples using the ISO 10272: 2017. The stocking density for maximum calculated yield was 20 (trials 1 and 2) or 22 birds/m2 (trial 3), followed by 18, 16, 14 and 12. At the stocking rate of 20 birds/m2, the birds in the pen grew faster than those at the same density in the main flock achieving 2 Kg 3–6 days faster. Birds in the BC were observed to be generally healthier, and in some cases, remained Campylobacter negative, even after the main flock was infected. Our results conclude that dividing the flock into sub-flocks of approximately 20 birds/m2 using BC could increase productivity up to 20%, while preventing Campylobacter.

Grain legumes, such as faba beans, have been investigated as promising ingredients to enhance the nutritional value of wheat bread. However, a detrimental effect on technological bread quality was often reported. Furthermore, considerable amounts of antinutritional compounds present in faba beans are a subject of concern. Sourdough-like fermentation can positively affect baking performance and nutritional attributes of faba bean flours. The multifunctional lactic acid bacteria strain Leuconostoc citreum TR116 was employed to ferment two faba bean flours with different protein contents (dehulled flour (DF); high-protein flour (PR)). The strain’s fermentation profile (growth, acidification, carbohydrate metabolism and antifungal phenolic acids) was monitored in both substrates. The fermentates were applied in regular wheat bread by replacing 15% of wheat flour. Water absorption, gluten aggregation behaviour, bread quality characteristics and in vitro starch digestibility were compared to formulations containing unfermented DF and PR and to a control wheat bread. Similar microbial growth, carbohydrate consumption as well as production of lactic and acetic acid were observed in both faba bean ingredients. A less pronounced pH drop as well as a slightly higher amount of antifungal phenolic acids were measured in the PR fermentate. Fermentation caused a striking improvement of the ingredients’ baking performance. GlutoPeak measurements allowed for an association of this observation with an improved gluten aggregation. Given its higher potential to improve protein quality in cereal products, the PR fermentate seemed generally more promising as functional ingredient due to its positive impact on bread quality and only moderately increased starch digestibility in bread.

To date, Certrevirus is one of two genera of bacteriophage (phage), with phages infecting Pectobacterium atrosepticum, an economically important phytopathogen that causes potato blackleg and soft rot disease. This study provides a detailed description of Pectobacterium phage CB7 (vB_PatM_CB7), which specifically infects P. atrosepticum. Host range, morphology, latent period, burst size and stability at different conditions of temperature and pH were examined. Analysis of its genome (142.8 kbp) shows that the phage forms a new species of Certrevirus, sharing sequence similarity with other members, highlighting conservation within the genus. Conserved elements include a putative early promoter like that of the Escherichia coli sigma70 promoter, which was found to be shared with other genus members. A number of dissimilarities were observed, relating to DNA methylation and nucleotide metabolism. Some members do not have homologues of a cytosine methylase and anaerobic nucleotide reductase subunits NrdD and NrdG, respectively. Furthermore, the genome of CB7 contains one of the largest numbers of homing endonucleases described in a single phage genome in the literature to date, with a total of 23 belonging to the HNH and LAGLIDADG families. Analysis by RT-PCR of the HNH homing endonuclease residing within introns of genes for the large terminase, DNA polymerase, ribonucleotide reductase subunits NrdA and NrdB show that they are splicing competent. Electrospray ionization-tandem mass spectrometry (ESI-MS/MS) was also performed on the virion of CB7, allowing the identification of 26 structural proteins—20 of which were found to be shared with the type phages of the genera of Vequintavirus and Seunavirus. The results of this study provide greater insights into the phages of the Certrevirus genus as well as the subfamily Vequintavirinae.

Campylobacteriosis is the leading cause of human bacterial gastroenteritis, very often associated with poultry consumption. Thermophilic Campylobacter (Campylobacter jejuni and Campylobacter coli) isolates (n = 158) recovered from broiler neck skin and caecal contents in Ireland over a one-year period, resistant to at least one of three clinically relevant antimicrobial classes, were screened for resistance determinants. All ciprofloxacin-resistant isolates (n = 99) harboured the C257T nucleotide mutation (conferring the Thr-86-Ile substitution) in conjunction with other synonymous and nonsynonymous mutations, which may have epidemiological value. The A2075G nucleotide mutation and amino acid substitutions in L4 and L22 were detected in all erythromycin-resistant isolates (n = 5). The tetO gene was detected in 100% (n = 119) of tetracycline-resistant isolates and three of which were found to harbour the mosaic tetracycline resistance gene tetO/32/O. Two streptomycin-resistant C. jejuni isolates (isolated from the same flock) harboured ant(6)-Ib, located in a multidrug resistance genomic island, containing aminoglycoside, streptothricin (satA) and tetracycline resistance genes (truncated tetO and mosaic tetO/32/O). The ant(6)-Ie gene was identified in two streptomycin-resistant C. coli isolates. This study highlights the widespread acquisition of antimicrobial resistance determinants among chicken-associated Campylobacter isolates, through horizontal gene transfer or clonal expansion of resistant lineages. The stability of such resistance determinants is compounded by the fluidity of mobile genetic element.

A series of fifteen new N-alkoxyphenylanilides of 3-hydroxynaphthalene-2-carboxylic acid was prepared and characterized. Primary in vitro screening of the synthesized compounds was performed against Staphylococcus aureus, three methicillin-resistant S. aureus strains, Mycobacterium tuberculosis H37Ra and M. avium subsp. paratuberculosis. Some of the tested compounds showed antibacterial and antimycobacterial activity against the tested strains comparable with or higher than that of the standards ampicillin or rifampicin. 3-Hydroxy-N-(2-propoxyphenyl)naphthalene-2-carboxamide and N-[2-(but-2-yloxy)-phenyl]-3-hydroxynaphthalene-2-carboxamide had MIC = 12 µM against all methicillin-resistant S. aureus strains; thus their activity is 4-fold higher than that of ampicillin. The second mentioned compound as well as 3-hydroxy-N-[3-(prop-2-yloxy)phenyl]-naphthalene-2-carboxamide had MICs = 23 µM and 24 µM against M. tuberculosis respectively. N-[2-(But-2-yloxy)phenyl]-3-hydroxynaphthalene-2-carboxamide demonstrated higher activity against M. avium subsp. paratuberculosis than rifampicin. Screening of the cytotoxicity of the most effective antimycobacterial compounds was performed using THP-1 cells, and no significant lethal effect was observed for the most potent compounds. The compounds were additionally tested for their activity related to inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. N-(3-Ethoxyphenyl)-3-hydroxynaphthalene-2-carboxamide (IC50 = 4.5 µM) was the most active PET inhibitor. The structure-activity relationships are discussed.

In this study, a series of twenty-two ring-substituted naphthalene-1-carboxanilides were prepared and characterized. Primary in vitro screening of the synthesized carboxanilides was performed against Mycobacterium avium subsp. paratuberculosis. N-(2-Methoxyphenyl)naphthalene-1-carboxamide, N-(3-methoxy-phenyl)naphthalene-1-carboxamide, N-(3-methylphenyl)naphthalene-1-carboxamide, N-(4-methylphenyl)naphthalene-1-carboxamide and N-(3-fluorophenyl)naphthalene-1-carboxamide showed against M. avium subsp. paratuberculosis two-fold higher activity than rifampicin and three-fold higher activity than ciprofloxacin. The most effective antimycobacterial compounds demonstrated insignificant toxicity against the human monocytic leukemia THP-1 cell line. The testing of biological activity of the compounds was completed with the study of photosynthetic electron transport (PET) inhibition in isolated spinach (Spinacia oleracea L.) chloroplasts. The PET-inhibiting activity expressed by IC50 value of the most active compound N-[4-(trifluoromethyl)phenyl]naphthalene-1-carboxamide was 59 μmol/L. The structure-activity relationships are discussed.

In this work a series of 15 N-benzylamine substituted 5-amino-6-methyl-pyrazine-2,3-dicarbonitriles was prepared by the aminodehalogenation reactions using microwave assisted synthesis with experimentally set and proven conditions. This approach for the aminodehalogenation reaction was chosen due to its higher yields and shorter reaction times. The products of this reaction were characterized by IR, NMR and other analytical data. The compounds were evaluated for their antibacterial, antifungal and herbicidal activity. Compounds 3 (R = 3,4-Cl), 9 (R = 2-Cl) and 11 (R = 4-CF3) showed good antimycobacterial activity against Mycobacterium tuberculosis (MIC = 6.25 µg/mL). It was found that the lipophilicity is important for antimycobacterial activity and the best substitution on the benzyl moiety of the compounds is a halogen or trifluoromethyl group according to Craig’s plot. The activities against bacteria or fungi were insignificant. The presented compounds also inhibited photosynthetic electron transport in spinach chloroplasts and the IC50 values of the active compounds varied in the range from 16.4 to 487.0 µmol/L. The most active substances were 2 (R = 3-CF3), 3 (R = 3,4-Cl) and 11 (R = 4-CF3). A linear dependence between lipophilicity and herbicidal activity was observed.

Listeria monocytogenes is a ubiquitous Gram-positive bacterium that is a major concern for food business operators because of its pathogenicity and ability to form biofilms in food production environments. Bacteriophages (phages) have been evaluated as biocontrol agents for L. monocytogenes in a number of studies and, indeed, certain phages have been approved for use as anti-listerial agents in food processing environments (ListShield and PhageGuard Listex). Endolysins are proteins produced by phages in the host cell. They cleave the peptidoglycan cell wall, thus allowing release of progeny phage into the environment. In this study, the amidase domain of the phage vB_LmoS_293 endolysin (293-amidase) was cloned and expressed in Escherichia. coli (E. coli). Muralytic activity at different concentrations, pH and temperature values, lytic spectrum and activity against biofilms was determined for the purified 293-amidase protein. The results showed activity on autoclaved cells at three different temperatures (20 °C, 37 °C and 50 °C), with a wider specificity (L. monocytogenes 473 and 3099, a serotype 4b and serogroup 1/2b-3b-7, respectively) compared to the phage itself, which targets only L. monocytogenes serotypes 4b and 4e. The protein also inhibits biofilm formation on abiotic surfaces. These results show the potential of using recombinant antimicrobial proteins against pathogens in the food production environment.

In this study, a series of twenty-five ring-substituted 4-arylamino-7-chloroquinolinium chlorides were prepared and characterized. The compounds were tested for their activity related to inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts and also primary in vitro screening of the synthesized compounds was performed against mycobacterial species. 4-[(2-Bromophenyl)amino]-7-chloroquinolinium chloride showed high biological activity against M. marinum, M. kansasii, M. smegmatis and 7-chloro-4-[(2-methylphenyl)amino]quinolinium chloride demonstrated noteworthy biological activity against M. smegmatis and M. avium subsp. paratuberculosis. The most effective compounds demonstrated quite low toxicity (LD50 > 20 μmol/L) against the human monocytic leukemia THP-1 cell line within preliminary in vitro cytotoxicity screening. The tested compounds were found to inhibit PET in photosystem II. The PET-inhibiting activity expressed by IC50 value of the most active compound 7-chloro-4-[(3-trifluoromethylphenyl)amino]quinolinium chloride was 27 μmol/L and PET-inhibiting activity of ortho-substituted compounds was significantly lower than this of meta- and para-substituted ones. The structure-activity relationships are discussed for all compounds.

In this study, a series of twenty-two ring-substituted 2-hydroxynaphthalene-1‑carboxanilides were prepared and characterized. Primary in vitro screening of the synthesized compounds was performed against Staphylococcus aureus, three methicillin-resistant S. aureus strains, Mycobacterium marinum, M. kasasii, M. smegmatis. and M. avium paratuberculosis. The compounds were also tested for their activity related to inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. 2-Hydroxy-N-phenylnaphthalene-1-carboxanilide and 2-hydroxy-N-(3-trifluoromethylphenyl)naphthalene-1-carboxamide (IC50 = 29 µmol/L) were the most active PET inhibitors. Some of tested compounds showed the antibacterial and antimycobacterial activity against the tested strains comparable or higher than the standards ampicillin or isoniazid. Thus, for example, 2-hydroxy-N-(3-nitrophenyl)naphthalene-1-carboxamide showed MIC = 26.0 µmol/L against methicillin-resistant S. aureus and MIC = 51.9 µmol/L against M. marinum, or 2-hydroxy-N-phenylnaphthalene-1-carboxamide demonstrated MIC = 15.2 µmol/L against M. kansasii. The structure-activity relationships for all compounds are discussed.

Bacteriophages (phages) are viruses that infect bacterial hosts, and since their discovery over a century ago they have been primarily exploited to control bacterial populations and to serve as tools in molecular biology. In this commentary, we highlight recent diverse advances in the field of phage research, going beyond bacterial control using whole phage, to areas including biocontrol using phage-derived enzybiotics, diagnostics, drug discovery, novel drug delivery systems and bionanotechnology.

In this study, a series of thirty-five substituted quinoline-2-carboxamides and thirty-three substituted naphthalene-2-carboxamides were prepared and characterized. They were tested for their activity related to the inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. Primary in vitro screening of the synthesized compounds was also performed against four mycobacterial species. N-Cycloheptylquinoline-2-carboxamide, N-cyclohexylquinoline-2-carboxamide and N-(2-phenylethyl)quinoline-2-carboxamide showed higher activity against M. tuberculosis than the standards isoniazid or pyrazinamide and 2-(pyrrolidin-1-ylcarbonyl)quinoline and 1-(2-naphthoyl)pyrrolidine expressed higher activity against M. kansasii and M. avium paratuberculosis than the standards isoniazid or pyrazinamide. The most effective antimycobacterial compounds demonstrated insignificant toxicity against the human monocytic leukemia THP-1 cell line. The PET-inhibiting activity expressed by IC50 value of the most active compound N-benzyl-2-naphthamide was 7.5 μmol/L. For all compounds, the structure-activity relationships are discussed.

In this study, a series of twelve ring-substituted salicylanilides and carbamoylphenylcarbamates were prepared and characterized. The compounds were analyzed using RP-HPLC to determine lipophilicity. They were tested for their activity related to the inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. Moreover, their site of action in the photosynthetic apparatus was determined. Primary in vitro screening of the synthesized compounds was also performed against mycobacterial, bacterial and fungal strains. Several compounds showed biological activity comparable with or higher than the standards 3-(3,4-dichlorophenyl)-1,1-dimethylurea, isoniazid, penicillin G, ciprofloxacin or fluconazole. The most active compounds showed minimal anti-proliferative activity against human cells in culture, indicating they would have low cytotoxicity. For all compounds, the relationships between lipophilicity and the chemical structure are discussed.

Pectobacterium atrosepticum is a phytopathogen of economic importance as it is the causative agent of potato blackleg and soft rot. Here we describe the Pectobacterium phage vB_PatP_CB5 (abbreviated as CB5), which specifically infects the bacterium. The bacteriophage is characterized in detail and TEM micrographs indicate that it belongs to the Podoviridae family. CB5 shares significant pairwise nucleotide identity (≥80%) with P. atrosepticum phages φM1, Peat1, and PP90 and also shares common genome organization. Phylograms constructed using conserved proteins and whole-genome comparison-based amino acid sequences show that these phages form a distinct clade within the Autographivirinae. They also possess conserved RNA polymerase recognition and specificity loop sequences. Their lysis cassette resembles that of KP34virus, containing in sequential order a U-spanin, a holin, and a signal–arrest–release (SAR) endolysin. However, they share low pairwise nucleotide identity with the type phage of the KP34virus genus, Klebsiella phage KP34. In addition, phage KP34 does not possess several conserved proteins associated with these P. atrosepticum phages. As such, we propose the allocation of phages CB5, Peat1, φM1, and PP90 to a separate new genus designated Phimunavirus.

Recently, a Salmonella Typhi isolate producing CTX-M-15 extended spectrum β-lactamase (ESBL) and with decreased ciprofloxacin susceptibility was isolated in the Democratic Republic of the Congo. We have selected bacteriophages that show strong lytic activity against this isolate and have potential for phage-based treatment of S. Typhi, and Salmonella in general.

This perspective paper follows up on earlier communications on bacteriophage therapy that we wrote as a multidisciplinary and intercontinental expert-panel when we first met at a bacteriophage conference hosted by the Eliava Institute in Tbilisi, Georgia in 2015. In the context of a society that is confronted with an ever-increasing number of antibiotic-resistant bacteria, we build on the previously made recommendations and specifically address how the Nagoya Protocol might impact the further development of bacteriophage therapy. By reviewing a number of recently conducted case studies with bacteriophages involving patients with bacterial infections that could no longer be successfully treated by regular antibiotic therapy, we again stress the urgency and significance of the development of international guidelines and frameworks that might facilitate the legal and effective application of bacteriophage therapy by physicians and the receiving patients. Additionally, we list and comment on several recently started and ongoing clinical studies, including highly desired double-blind placebo-controlled randomized clinical trials. We conclude with an outlook on how recently developed DNA editing technologies are expected to further control and enhance the efficient application of bacteriophages.

The increase in antibiotic resistance in pathogenic bacteria is a public health danger requiring alternative treatment options, and this has led to renewed interest in phage therapy. In this respect, we describe the distinct host ranges of Staphylococcus phage K, and two other K-like phages against 23 isolates, including 21 methicillin-resistant S. aureus (MRSA) representative sequence types representing the Irish National MRSA Reference Laboratory collection. The two K-like phages were isolated from the Fersisi therapeutic phage mix from the Tbilisi Eliava Institute, and were designated B1 (vB_SauM_B1) and JA1 (vB_SauM_JA1). The sequence relatedness of B1 and JA1 to phage K was observed to be 95% and 94% respectively. In terms of host range on the 23 Staphylococcus isolates, B1 and JA1 infected 73.9% and 78.2% respectively, whereas K infected only 43.5%. Eleven open reading frames (ORFs) present in both phages B1 and JA1 but absent in phage K were identified by comparative genomic analysis. These ORFs were also found to be present in the genomes of phages (Team 1, vB_SauM-fRuSau02, Sb_1 and ISP) that are components of several commercial phage mixtures with reported wide host ranges. This is the first comparative study of therapeutic staphylococcal phages within the recently described genus Kayvirus.

In this study, a series of fourteen ring-substituted 8-hydroxyquinoline derivatives were prepared. The synthesis procedures are presented. The compounds were analyzed using RP-HPLC to determine lipophilicity. They were tested for their activity related to inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. Primary in vitro screening of the synthesized compounds was also performed against four mycobacterial strains and against eight fungal strains. Several compounds showed biological activity comparable with or higher than the standards isoniazid or fluconazole. For all the compounds, the relationships between the lipophilicity and the chemical structure of the studied compounds are discussed.

In this study, series of ring-substituted 2-styrylquinazolin-4(3H)-one and 4-chloro-2-styrylquinazoline derivatives were prepared. The syntheses of the discussed compounds are presented. The compounds were analyzed by RP-HPLC to determine lipophilicity. They were tested for their inhibitory activity on photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. Primary in vitro screening of the synthesized compounds was also performed against four mycobacterial strains and against eight fungal strains. Several compounds showed biological activity comparable with or higher than that of the standard isoniazid. It was found that the electronic properties of the R substituent, and not the total lipophilicity of the compound, were decisive for the photosynthesis-inhibiting activity of tested compounds.

In the study, a series of twelve ring-substituted 4-hydroxy-1H-quinolin-2-one derivatives were prepared. The procedures for synthesis of the compounds are presented. The compounds were analyzed using RP-HPLC to determine lipophilicity and tested for their photosynthesis-inhibiting activity using spinach (Spinacia oleracea L.) chloroplasts. All the synthesized compounds were also evaluated for antifungal activity using in vitro screening with eight fungal strains. For all the compounds, the relationships between the lipophilicity and the chemical structure of the studied compounds are discussed, as well as their structure-activity relationships (SAR).

Novel 1-(2-{3-/4-[(alkoxycarbonyl)amino]phenyl}-2-hydroxyethyl)-4-(2-fluorophenyl)-piperazin-1-ium chlorides (alkoxy = methoxy to butoxy; 8a–h) have been designed and synthesized through multistep reactions as a part of on-going research programme focused on finding new antimycobacterials. Lipophilic properties of these compounds were estimated by RP-HPLC using methanol/water mobile phases with a various volume fraction of the organic modifier. The log kw values, which were extrapolated from intercepts of a linear relationship between the logarithm of a retention factor k (log k) and volume fraction of a mobile phase modifier (ϕM), varied from 2.113 (compound 8e) to 2.930 (8h) and indicated relatively high lipophilicity of these salts. Electronic properties of the molecules 8a–h were investigated by evaluation of their UV/Vis spectra. In a next phase of the research, the compounds 8a–h were in vitro screened against M. tuberculosis CNCTC My 331/88 (identical with H37Rv and ATCC 2794), M. kansasii CNCTC My 235/80 (identical with ATCC 12478), a M. kansasii 6 509/96 clinical isolate, M. avium CNCTC My 330/80 (identical with ATCC 25291) and M. avium intracellulare ATCC 13950, respectively, as well as against M. kansasii CIT11/06, M. avium subsp. paratuberculosis CIT03 and M. avium hominissuis CIT10/08 clinical isolates using isoniazid, ethambutol, ofloxacin, ciprofloxacin or pyrazinamide as reference drugs. The tested compounds 8a–h were found to be the most promising against M. tuberculosis; a MIC = 8 μM was observed for the most effective 1-(2-{4-[(butoxycarbonyl)amino]phenyl}-2-hydroxyethyl)-4-(2-fluorophenyl)piperazin-1-ium chloride (8h). In addition, all of them showed low (insignificant) in vitro toxicity against a human monocytic leukemia THP-1 cell line, as observed LD50 values > 30 μM indicated. The structure–antimycobacterial activity relationships of the analyzed 8a–h series are also discussed.

New antibacterial agents are urgently needed for the elimination of biofilm-forming bacteria that are highly resistant to traditional antimicrobial agents. Proliferation of such bacteria can lead to significant economic losses in the agri-food sector. This study demonstrates the potential of the bacteriophage-derived peptidase, CHAPK, as a biocidal agent for the rapid disruption of biofilm-forming staphylococci, commonly associated with bovine mastitis. Purified CHAPK applied to biofilms of Staphylococcus aureus DPC5246 completely eliminated the staphylococcal biofilms within 4 h. In addition, CHAPK was able to prevent biofilm formation by this strain. The CHAPK lysin also reduced S. aureus in a skin decolonization model. Our data demonstrates the potential of CHAPK as a biocidal agent for prevention and treatment of biofilm-associated staphylococcal infections or as a decontaminating agent in the food and healthcare sectors.

A series of twenty substituted 2-hydroxy-3-[(2-aryloxyethyl)amino]propyl 4-[(alkoxycarbonyl)amino]benzoates were prepared and characterized. As similar compounds have been described as potential antimycobacterials, primary in vitro screening of the synthesized carbamates was also performed against two mycobacterial species. 2-Hydroxy-3-[2-(2,6-dimethoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride, 2-hydroxy-3-[2-(4-methoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride, and 2-hydroxy-3-[2-(2-methoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride showed higher activity against M. avium subsp. paratuberculosis and M. intracellulare than the standards ciprofloxacin, isoniazid, or pyrazinamide. Cytotoxicity assay of effective compounds was performed using the human monocytic leukaemia THP-1 cell line. Compounds with predicted amphiphilic properties were also tested for their effects on the rate of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. All butyl derivatives significantly stimulated the rate of PET, indicating that the compounds can induce conformational changes in thylakoid membranes resulting in an increase of their permeability and so causing uncoupling of phosphorylation from electron transport.

The Eighth Central European Conference “Chemistry towards Biology” was held in Brno, Czech Republic, on August 28–September 1, 2016 to bring together experts in biology, chemistry and design of bioactive compounds; promote the exchange of scientific results, methods and ideas; and encourage cooperation between researchers from all over the world. The topics of the conference covered “Chemistry towards Biology”, meaning that the event welcomed chemists working on biology-related problems, biologists using chemical methods, and students and other researchers of the respective areas that fall within the common scope of chemistry and biology. The authors of this manuscript are plenary speakers and other participants of the symposium and members of their research teams. The following summary highlights the major points/topics of the meeting.

This research was focused on in silico characterization and in vitro biological testing of the series of the compounds carrying a N-arylpiperazine moiety. The in silico investigation was based on the prediction of electronic, steric and lipohydrophilic features. The molecules were screened against Mycobacterium avium subsp. paratuberculosis CIT03, M. smegmatis ATCC 700084, M. kansasii DSM 44162, M. marinum CAMP 5644, Staphylococcus aureus ATCC 29213, methicillin-resistant S. aureus 63718, Escherichia coli ATCC 25922, Enterococcus faecalis ATCC 29212, Candida albicans CCM 8261, C. parapsilosis CCM 8260 and C. krusei CCM 8271, respectively, by standardized microdilution methods. The eventual antiproliferative (cytotoxic) impact of those compounds was examined on a human monocytic leukemia THP-1 cell line, as a part of the biological study. Promising potential against M. kansasii was found for 1-[3-(3-ethoxyphenylcarbamoyl)oxy-2-hydroxypropyl]-4-(3-trifluoromethylphenyl)piperazin-1-ium chloride (MIC = 31.75 μM), which was comparable to the activity of isoniazid (INH; MIC = 29.17 μM). Moreover, 1-{2-hydroxy-3-(3-methoxyphenylcarbamoyl)oxy)propyl}-4-(4-fluorophenyl)piperazin-1-ium chloride was even more effective (MIC = 17.62 μM) against given mycobacterium. Among the tested N-arylpiperazines, 1-{2-hydroxy-3-(4-methoxyphenylcarbamoyl)oxy)propyl}-4-(3-trifluorometh-ylphenyl)piperazin-1-ium chloride was the most efficient against M. marinum (MIC = 65.32 μM). One of the common features of all investigated substances was their insignificant antiproliferative (i.e., non-cytotoxic) effect. The study discussed structure–antimicrobial activity relationships considering electronic, steric and lipophilic properties.

A series of nineteen N-(alkoxyphenyl)-2-hydroxynaphthalene-1-carboxamides and a series of their nineteen positional isomers N-(alkoxyphenyl)-1-hydroxynaphthalene-2-carboxamides were prepared and characterized. Primary in vitro screening of all the synthesized compounds was performed against Mycobacterium tuberculosis H37Ra, M. kansasii and M. smegmatis. Screening of the cytotoxicity of the compounds was performed using human monocytic leukemia THP-1 cells. Some of the tested compounds showed antimycobacterial activity comparable with or higher than that of rifampicin. For example, 2-hydroxy-N-(4-propoxyphenyl)-naphthalene-1-carboxamide showed the highest activity (MIC = 12 µM) against M. tuberculosis with insignificant cytotoxicity. N-[3-(But-2-yloxy)phenyl]- and N-[4-(but-2-yloxy)phenyl]-2-hydroxy-naphthalene-1-carboxamide demonstrated high activity against all tested mycobacterial strains and insignificant cytotoxicity. N-(Alkoxyphenyl)-1-hydroxynaphthalene-2-carboxamides demonstrated rather high effect against M. smegmatis and M. kansasii and strong antiproliferative effect against the human THP-1 cell line. Lipophilicity was found as the main physicochemical parameter influencing the activity. A significant decrease of mycobacterial cell metabolism (viability of M. tuberculosis H37Ra) was observed using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide) assay. Structure-activity relationships are discussed.

ABSTRACT Pseudomonas aeruginosa is a common cause of infection in the lungs of patients with cystic fibrosis (CF). In addition, biofilm formation and antibiotic resistance of Pseudomonas are major problems that can complicate antibiotic therapy. We evaluated the efficacy of using bacteriophages to kill the pathogen in both biofilms and in the murine lung. We isolated and characterized two phages from a local wastewater treatment plant, a myovirus (ϕNH-4) and a podovirus (ϕMR299-2). Both phages were active against clinical isolates of P. aeruginosa. Together, the two phages killed all 9 clinical isolate strains tested, including both mucoid and nonmucoid strains. An equal mixture of the two phages was effective in killing P. aeruginosa NH57388A (mucoid) and P. aeruginosa MR299 (nonmucoid) strains when growing as a biofilm on a cystic fibrosis bronchial epithelial CFBE41o- cell line. Phage titers increased almost 100-fold over a 24-h period, confirming replication of the phage. Furthermore, the phage mix was also effective in killing the pathogen in murine lungs containing 1 × 107 to 2 × 107 P. aeruginosa. Pseudomonas was effectively cleared (reduced by a magnitude of at least 3 to 4 log units) from murine lungs in 6 h. Our study demonstrates the efficacy of these two phages in killing clinical Pseudomonas isolates in the murine lung or as a biofilm on a pulmonary cell line and supports the growing interest in using phage therapy for the control and treatment of multidrug-resistant Pseudomonas lung infections in CF patients. IMPORTANCE Given the rise in antibiotic resistance, nonantibiotic therapies are required for the treatment of infection. This is particularly true for the treatment of Pseudomonas infection in patients with cystic fibrosis. We have identified two bacterial viruses (bacteriophages) that can kill Pseudomonas growing on human lung cells and in an animal model of lung infection. The use of bacteriophages is particularly appropriate because the killing agent can replicate on the target cell, generating fresh copies of the bacteriophage. Thus, in the presence of a target, the killing agent multiplies. By using two bacteriophages we can reduce the risk of resistant colonies developing at the site of infection. Bacteriophage therapy is an exciting field, and this study represents an important demonstration of efficacy in validated infection models.