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Submitted in fulfillment of the academic requirement for a degree in Doctor of Philosophy (Ph.D.) in Food Science and Technology, Durban University of Technology, Durban, South Africa, 2017. / There has been a global increase in fresh produce consumption, due to its attendant nutritional a nd health benefits. On the other hand, increase in the outbreak of diseases, accompanied with health and economic implications, have been traced to this deve lopment. A good number of pathogenic contaminants along the food chain have been identified as causative agent s with Listeria monocytogenes identified as one of such. Among other control strategies, the use of bacteriophage, was recommended as a palliative measure. Furthermore, the a ppli cation of artificial neural networks (ANN) in food safety remains an emerging concept in risk assessment study. Therefore, the aim of this research is to investigate the effect of bacteriophage or phage control and artificial neural network prediction in the inactivation of L. monocytogenes ATCC 7644 on fresh produce. Fresh-cut tomato and carrot were artificially inoculated with L. monocytogenes (108 CFU/ml) and subjected to antimicrobial treatment of Listex P100 bacteriophage (108 PFU/ml), sucrose monolaurate (SML at 100, 250 and 400 ppm), with chlorine (sodium hypochlorite at 200 ppm) used as control. Also, application of ANN to predict the risk effect of antimicrobial treatments of bacteriophage, sucrose monolaurate and chlorine was evaluated on the fresh-cut produce. Mathematical models were developed using a linear regression and sigmoid (hyperbolic and logistic) activation function-(120). Data sets were trained using Back propagation ANN, containing one hidden layer with four hidden neurons. Furthermore, carbon utilization profile of phage-treated L. monocytogenes using phenotypic micro array method was evaluated. In the first phase, susceptibility of L. monocytogenes subjected to certain stress-adapted conditions (acid,-adapted AA, chlorine-adapted CA, heat-adapted HA) and non-adapted-NA to phage treatment inoculated on the fresh-cut produce stored for 10 days at 4, 10 and 25oC was evaluated. The second phase investigated the combination of bacteriophage and sucrose monolaurate (using chlorine at 200 ppm as control) to inhibit the L. monocytogenes growth on the fresh-cut produce stored for 6 days at 4, 10 and 25oC. Physicochemical properties (pH, titratable acid-TTA, total soluble solids-TSS, and colour values-CIE L* a* b*) of the fresh produce after treatment were evaluated. In the third phase, ANN as a predictive tool was used to evaluate the risk involved in the relationship among the initial bacterial load, fresh-produce type, antimicrobial concentration and residual bacteria. In the final phase, 100 µL of phage-treated L. monocytogenes was introduced into a 96-micro well plate impregnated with a tetrazolium dye. The Carbon utilization profile was evaluated at intervals of 4 hours for 48 hours using a biolog micro station. Generally, L. monocytogenes grew on both fresh-cut produce and the storage temperature did not adversely affect the lytic ability of the phage treatment. Antimicrobial treatment of phage and sucrose monolaurate had minimal variations on the physicochemical properties of both fresh-cut samples. All stress-adapted and non-adapted L. monocytogenes were (p ≤ 0.05) susceptible to bacteriophage control. Phage treatment reduced non-adapted, acid adapted, chlorine-adapted, and heat-adapted L. monocytogenes population by 0.57, 0.81, 0.86 and 0.95 log CFU/ml in fresh-cut tomato, and 2.26, 2.41, 2.49 and 2.54 log CFU/ml in fresh cut carrot respectively. Furthermore, the additive effect of SML at 100 and 250 ppm had no significant effect on phage lysis. However, combination of phage with SML at 400 ppm significantly (p ≤ 0.05) resulted in 1 and 3 fold reductions in tomato and carrot respectively. Control treatment with chlorine resulted in 1-2 log reductions on both fresh produce. Algorithm data set trained using ANN gave 100% accuracy. Prediction with logistic activation function showed the highest positive correlation relationship between predicted and observed values with ~ 0.99 R2-value and MSE of 0.0831. Carbon utilization profile showed hexose and pentose sugars-ribose, glucose, fructose and sugars were maximally utilized while oligosaccharide sugars of sucrose, cellobiose and gentiobiose were similarly observed to be utilized. Notably, utilization of glucose-6-phosphate which determines L. monocytogenes pathogenicity was not very pronounced in the carbon profile. Bacteriophage application in the inactivation of L. monocytogenes contamination of fresh produce provides a safe means of control. Its perceived limitation however, can be overcome by combining with other antimicrobials. Similarly, the use of artificial neural networks prediction, remains an improved approach to harness the potential risk that could occur through this method. / D

Clostridium difficile is the most common bacterial cause of infectious diarrhoea in healthcare environments and in 2014 was responsible for 13,785 infections in the UK. C. difficile infection (CDI) is spread via the faecal-oral route and by contact with contaminated surfaces. However, despite the healthcare concerns no tests are available to validate if sufficient cleaning has been conducted. In addition, Polymerase Chain Reaction (PCR) and Enzyme Immunoassays (EIAs)-based tests used to diagnose CDI lack sensitivity and specificity and hence false negative results are commonly obtained. To overcome these concerns the aim of the PhD research has been to develop the first diagnostic test that exploits the specific interactions of C. difficile bacteriophages (phages), viruses that specifically infect and kill C. difficile. In order to develop a C. difficile phage-based test, first suitable phages that can be used for the test were identified and this was conducted by screening 35 different C. difficile phages against 160 clinically relevant C. difficile isolates. Five phages were found to infect the most number of isolates and were investigated further to identify whether a phage-based diagnostic could be developed based on phages binding (adsorption) to different C. difficile subgroups. However, for all five phages, adsorption rates were not consistently high for C. difficile subgroups in comparison to other common bacteria found in similar locations to C. difficile. Therefore, to increase specificity of the phage-based diagnostic test a new approach was taken by tagging two phages with luminescence luxAB genes (reporter phages), which would be expressed once C. difficile cells were infected with the phages. To design the C. difficile reporter phages, non-essential phage genes were replaced with the luxAB genes, but this study revealed mutagenesis of C. difficile was troublesome and extensive optimisation was required. In addition, once the reporter phages had successfully been constructed the luxAB genes were unstable within the phage genome and were lost during phage replication. Despite extensive optimisation and due to time constrains the luxAB genes were not stabilised within the phages but future work will focus on stabilising the genes.

Masters of Science / Thermophilic bacteriophages provide simple model systems for understanding biochemical and biological adaptation mechanisms at elevated temperatures. The essential objectives of this study were to characterise the physicochemical properties of select Geobacillus bacteriophages and to sequence their complete genomes. The later objective is believed to be an essential prerequisite to the engineering of a sitespecific integration vector for the stable cloning of exogenous genes into host bacteria. Bacteriophages were assayed at 55oC by the agar overlay technique using dry Karoo soils as source material. A pure strain of bacteriophage called GV1 (for Geobacillus stearothermophilus virus 1) was isolated with the strain Geobacillus stearothermophilus TAU3A1. Plaques were medium sized (2 to 4 mm diameters), with regular contour, clear, and without resistant cells. Host range specificity study showed that GV1 was lytic on thirteen thermophilic Bacillus-like strains tested, including strains of Geobacillus stearothermophilus, G. thermoglucosidasius, B. licheniformis, Anoxybacillus idirlerensis, and A. kuwalawohkensis. However, GV1 failed to infect a mesophilic strain of Bacillus megaterium. TEM analysis of semipurified particles revealed that the phage belongs to the family of Siphoviridae. Morphological characteristics included a long tail of approximately 100 nm and a hexagonal head of approximately 50 nm diameter. Viability and stability studies showed that the phage was best maintained at -80oC in PMN buffer supplemented with 20% glycerol. It was stable at a pH range of 5.5 to 7.5 and MgCl2 and CaCl2 concentration of 0.001 M. hermostability experiments, conducted over short periods of time, showed that GV1 was stable over the temperature range 50 to 75oC, with optimum at 55oC. The study of phage-host interactions showed that phage articles inhibited the initial growth of infected cultures in the first six hours post-infection, presumably while mature phages were released. This was followed by a steady recovery of the growth rate. Atempts to obtain pure particles and to extract and sequence phage DNA were unsuccessful due to the low titer nature of the phage.

The increasing incidence of infections due to antibiotic resistant bacteria has led to renewed interest in bacteriophages (= phages) and phage therapy. Although phage therapy has been applied to control bacterial diseases in plants, poultry, livestock and humans, its application in aquaculture is still relatively limited. The emergence of phage-resistant bacterial mutants has been considered to be one of the major limitations of phage therapy. This study aimed to (i) isolate and characterise phages; (ii) select phages and their bacterial hosts to set up in vivo phage therapy models with aquaculture animals, and estimate the efficiency of phage therapy; (iii) investigate the generation and characteristics of phage-resistant mutants, and thus estimate the consequence of applying phage therapy when phage-resistant mutants emerge; and (iv) discuss the prospects for application of phages in aquaculture. Two Vibrio isolates and their phages were isolated from a Scottish marine fish farm. Based on the results of conventional phenotype testing and 16S rRNA gene sequencing analysis, the two vibrios, V9 and V13, were identified as Vibrio splendidus and Vibrio cyclitrophicus, respectively. The bacterial characteristics including morphology, temperature and salinity range of growth, production of extracellular enzymes, and the possession of virulence genes were examined. According to the morphological characteristics observed using transmission electron microscopy by negative staining, phage PVS9 of V. splendidus V9 was identified as a myophage, while phage PVC13 of V. cyclitrophicus V13 was identified as a siphophage. The phages could only lyse one bacterial host strain and their genomic DNA was double stranded with a size of ~46 kb. The two Vibrio isolates were found to be non- or of low virulence to rainbow trout, goldsinny wrasse and Artemia in pathogenicity experiments. Thus an in vivo phage therapy model could not be set up using these Vibrio isolates and their phages. Two phages pAS-3 and pAS-6 were isolated using the Aeromonas salmonicida subsp. salmonicida Hooke strain as the host. Phages pAS-3 and pAS-6 had a similar genome size of ~50 kb, and the same relatively narrow host range within A. salmonicida subsp. salmonicida strains. The siphophage pAS-3 formed clear plaques and inhibited A. salmonicida Hooke growth in vitro completely for at least 18 hours when using MOI = 1,000, whereas the podophage pAS-6 formed turbid plaques and weakly inhibited Hooke growth. Rainbow trout exposed by intraperitoneal injection with 0.1 mL of the raw phage preparations at a concentration of 108 PUF mL-1 showed no adverse effects over 14 days. In the phage therapy trial, fish were firstly injected with 1 x 102 CFU fish-1 of A. salmonicida Hooke, then immediately injected with phage preparations of pAS-3 and pAS-6, respectively, using MOI = 10,000. Compared with the control group (which did not receive phage treatment), phage treated groups showed a delay in the time to death, and lower mortalities. However, the mortalities and time to death between phage treated and non-treated groups were not significantly different. Phage-resistant mutants of pathogenic A. salmonicida strain Hooke were induced by repeatedly challenging with phage pAS-3. One of the mutants, termed HM, was chosen to compare the characteristics with the parental wild-type strain Hooke. Test results including the formation of ‘smooth’ colonies on TSA, autoagglutination negative, the formation of creamy colonies on Coomassie Brilliant Blue agar, and the degradation of a thick/furry layered structure on the cell surface indicated a deficiency of the A-layer in the phage-resistant mutant HM. Therefore, it was deduced that the A-layer either directly acted as the receptor of A. salmonicida phage pAS-3, or was affected indirectly by the change of an unknown phage receptor. The greater wax moth larvae model was used to compare the virulence of the phage-resistant mutant HM and the parental wild-type strain Hooke, as it is an ethically acceptable animal model, which has the advantages of being low cost and convenient for injection, and is also a recognised alternative model for bacterial pathogens of fish. The results showed that virulence of the phage-resistant mutant HM did not decline in the greater wax moth larvae model compared with that of the parental wild-type strain Hooke. In conclusion, different approaches were used to isolate and characterise phages from different aquaculture environments for potential use in phage therapy. A rainbow trout model was set up using pathogenic A. salmonicida strain Hooke and two A. salmonicida phages pAS-3 and pAS-6. The use of phage treatment led to lower cumulative mortalities and delay to the time of death, although the differences between the groups were not significant, futher work is required to determine if these phages have potential in phage therapy. The consequence of applying phage therapy when phage-resistant mutants emerge was estimated based on their characteristics and virulence, and no decline in virulence of the phage-resistant mutant from this study indicates the importance of fully testing the virulence of phage-resistant mutants before carrying out large scale field trials of phage therapy. It appears feasible to use phage therapy as an alternative approach to control bacterial infections in aquaculture, but further studies are required to focus on improving effectiveness, and also to overcome the concrete limitations and hurdles in application and commercialisation. Moreover, a broader range of applications of phages in aquaculture should be explored.

Initiation of transcription is a major step in the regulation of gene expression. A dominant theme in regulation of gene expression lies in understanding the mechanism involved in selective expression of the genes in response to external or internal stimuli. Gene regulatory proteins bind DNA at specific sites either cognate to the promoters they act upon or at a distance, thereby exerting their effect by turning on (activation) or turning off (repression) the genes. Response of these factors to the environmental signals is further achieved by the DNA binding affinity of the transcription factors that can be modulated by small ligands, concentrations of which may fluctuate in response to nutrient availability and stress. Bacteriophages achieve a high degree of efficiency in gene expression by evolving elegant strategies of transcriptional control. mom gene of enterobacteriophage Mu serves as an excellent model to understand this elaborate regulation of gene expression. The gene encodes a unique DNA modification function that confers an anti-restriction phenotype to the phage genome. Though dispensable for phage growth, it is fascinating in two respects (i) a novel modification; (ii) regulation follows a complex scheme without precedence in prokaryotes. mom is the last gene to be expressed during the phage lytic life cycle. Premature expression of the gene is deleterious to both host and phage and hence it is under a complex regulatory network. Dam methylase, a host encoded protein acts as a positive regulator of gene expression, an example where methylation has been shown to play a positive role in regulating tranascription. OxyR, another host encoded protein negatively regulates mom gene expression. Dam methylation prevents the binding OxyR to its site located in the mom regulatory region. The regulatory interplay also involves two phage encoded proteins. C, a middle gene product is essential for transcriptional switch from middle to late genes and Com, a late gene product, for enhancing translation of mom mRNA. Thus, C and Com serve as transcriptional and translational activators of mom gene expression. Pmom is a weak promoter with both -10 and -35 elements away from consensus and a sub-optimal 19 bp spacer element encompassing a stretch of 6T residues that act as negative elements. ‘T stretch’ is known to induce a kink in the DNA. The sub-optimal spacer region makes the promoter elements out of phase and RNAP by itself cannot bind at mom promoter. C protein exerts its effect in activation in a multistep mechanism. The protein binds DNA as a dimer overlapping the promoter and unwinds the DNA, realigning the promoter elements, thus recruiting the RNAP. In the next step, it enhances the promoter clearance by the enzyme, thus enhancing the rate of productive transcription. With this prevailing knowledge on C mediated mom gene expression, the present thesis work describes the experiments carried out to further understand the molecular mechanism of second step activation at Pmom. Genetic and biochemical analysis were carried out to identify the interacting surface of C protein on RNAP. Subsequently, studies have been extended to understand the C mediated transactivation at other late promoters- lys, I, P, which encode for the lysis and morphogenetic functions of the phage. Finally, Mg2+ coordinating residues in C protein were identified to decipher the ligand induced conformational changes in the activator protein required for its transactivator function. Chapter I, a general introduction to the thesis, deals with the detailed discussion on gene expression and its regulatory mechanisms. RNA polymerase (RNAP) being the central molecule of gene expression (transcription) its organization and assembly are discussed. With the availability of the high resolution crystal structures of bacterial RNAP, an in-depth review on RNAP structure in terms of its potential regulatory targets, conformational changes associated with the formation of a functional holoenzyme, and during its transition from initiation to elongation processes have been described. Regulation of transcription with an emphasis on activation mechanism, ligand mediated allosteric transitions in regulatory proteins and the polymerase-activator interactions are discussed citing a few examples. The chapter concludes by introducing bacteriophage Mu and mom gene and its regulation by C. The objectives of the thesis form the concluding section of the chapter. Activators are capable of resurrecting defective promoters in response to cellular demands. The unusual, multistep activation of mom promoter (Pmom) by C protein involves activator mediated promoter unwinding to recruit RNA Polymerase (RNAP) and subsequent enhanced promoter clearance of the enzyme. The first step of transactivation is an interaction independent step, while the later might involve a transient interaction between C and one of the subunits of RNAP. Previous studies pointed out β′ subunit to be the most probable interaction partner. Chapter II comprises the genetic and biochemical studies carried out to confirm this observation. Employing a genetic screen mutations in rpoC gene (encoding the β′ subunit of RNAP), were isolated which result in the defective RNAP. The mutant RNAPs were assayed for their C specific activity by in vivo transactivation assays. Such mutants have been purified and characterized to understand their effect at different steps of C mediated mom gene expression during transcription initiation. The mutant RNAP had normal transcription activity with typical σ70 promoters but exhibited reduced productive transcription and enhanced abortive initiation on C-dependent Pmom. Experiments carried out to probe the interaction between C and mutant RNAP revealed that the physical interaction per se is not disrupted between the two proteins. Post C-mediated recruitment of RNAP to the promoter, transient interactions between the two proteins appears to induce subtle conformational changes in RNAP leading to an enhanced promoter clearance. Transactiavtor protein C is essential for the expression of other late genes lys, I, P apart from mom during the phage life cycle. Although the mechanism of multistep activation at Pmom has been elucidated, little is known on the transactivation from lys, I and P promoters. Chapter III includes studies carried out to understand the process of activation at these promoters. Owing to the differences in their C-binding site and promoter architecture it was important to investigate the differential effect of C, if any at lys, I , P promoters compared to that at Pmom. Activators in prokaryotes are shown to stimulate different steps of transcription initiation pathway ranging from the polymerase binding to the promoters to the post recruitment steps of isomerization and promoter clearance. Effect of C at different steps of transcription initiation pathway was analysed. The results indicate that C is absolutely essential for transcription from lys, I and P promoters similar to mom. However, at these promoters C exerts its effect at the step of Isomerisation from closed complex to open complex formation. Thus, C acts at a single step here and the mode of activation is different from that observed at Pmom. C dimer binds DNA with high affinity and sequence specificity, to an interrupted palindromic sequence overlapping the -35 element of mom promoter. Mg2+ mediated conformational transitions in C protein are essential for its DNA binding and transactivation functions. Chapter IV deals with the identification of the Mg2+ coordinating residues in C protein. Primary sequence analyses lead to the identification of a putative metal coordinating motif (EXDXD) towards the N-terminus of the protein. These residues were subjected to site directed mutagenesis to infer their role in Mg2+ coordination, its associated allosteric transition required for specific interaction with DNA. Mutants showed an altered Mg2+ induced conformation, compromised DNA binding and reduced levels of transcription activation when compared to C protein. Though Mg2+ is widely used in various DNA transaction reactions, this study provides the first insights on the importance of metal-ion induced allosteric transitions in regulating transcription factor function.

La diversité des génomes de virus infectant les bactéries, les bactériophages (ou phage en abrégé), est telle qu’il est difficile de les classer de manière satisfaisante, la notion d’espèce elle-même ne faisant pas accord dans la communauté scientifique. A la racine de cette diversité, un des facteurs clé est la recombinaison de l’ADN, qui est élevée chez les bactériophages, et permet des échanges de gènes entre entités parfois fort différentes. Mes travaux se sont centrés sur la recombinaison homologue chez les bactériophages, et en particulier sur la protéine centrale de ce processus, la recombinase. J’ai montré pour deux grands types de recombinases phagiques, de type Rad52 et Sak4, que celles-ci étaient beaucoup moins fidèles dans le processus de recombinaison, comparées à la recombinase bactérienne RecA. De plus, pour Sak4, j’ai observé que cette recombinaison se produisait par appariement simple brin, et qu’elle dépendait entièrement in vivo d’une SSB phagique, dont le gène est situé à proximité du gène sak4 sur le chromosome du phage. Les échanges génétiques sont donc grandement facilités pour les phages contenant ce type de recombinases, mais ils ne sont pas non plus anarchiques : la recombinaison s’observe jusque 22% de divergence, mais deux séquences à 50% de divergence ne peuvent recombiner. Tout se passe donc comme si la notion d’espèce devait être élargie chez les phages par rapport aux bactéries, pour inclure dans un même groupe des génomes portant des traces d’échanges récents de matériel génétique par recombinaison homologue (ce que l’on appelle le mosaïcisme). / The diversity of the viruses infecting bacteria (bacteriophages, or phages for short) is so important that it is difficult to classify them in a pertinent way, and the species notion itself is a matter of debate among specialists. At the root of this diversity, one of the key factors is DNA recombination, which occurs at high levels among phages, and permits gene exchanges among entities that are sometimes very distant. My research has focused on homologous recombination in phages, and in particular on the protein that is key to the process, the recombinase. I have shown, for two different types of recombinases, Rad52-like and Sak4-like, that their fidelity was relaxed, compared to the bacterial recombinase, RecA. Moreover, for Sak4, a protein that had not been studied before, I showed that recombination occurs by single strand annealing, and that it is strictly dependent in vivo on the co-expression of its cognate SSB protein, whose gene is often encoded nearby in phage genomes encoding sak4. Genetic exchanges are therefore greatly facilitated for phages encoding these types of recombinases. Nevertheless, exchanges are not anarchical: recombination is seen up to 22% diverged substrates, but 50% diverged DNA sequences will not recombine. It may be that the species notion should be enlarged for phages, so as to include into a same group all phages exhibiting traces of recent exchanges of genetic material (the so-called mosaicism).

Os elementos genéticos móveis encontrados no genoma da bactéria Xylella fastidiosa (Xf) são representados principalmente por bacteriófagos (na forma de profagos inseridos no genoma) e ilhas genômicas. As integrases são responsáveis pelo processo de mobilização (integração e/ou excisão) destes elementos, através do mecanismo de recombinação sítio-específica. Bacteriófagos e ilhas genômicas estão associados a eventos de rearranjos genômicos e à aquisição e ou interrupção de genes importantes para bactéria, tendo implicação direta na diversidade e organização genômica e, por conseqüência, na diferenciação entre linhagens. A extensão e o impacto desses eventos é o foco deste trabalho, através da análise in-silico das integrases e sua associação com regiões de profagos e ilhas genômicas, no genoma de quatro linhagens de Xf. Os dados aqui apresentados corroboram o papel das integrases e seus elementos genéticos móveis associados como agentes chaves no processo de diversidade e evolução da organização genômica entre as quatro linhagens de Xf. / The mobile genetic elements found in the genome of the bacterium Xylella fastidiosa (Xf) are mainly represented by bacteriophages (as prophages inserted into the genome) and genomic islands. The integrases are responsible for the process of mobilization (integration and / or excision) of these elements through the mechanism of site-specific recombination. Bacteriophages and genomic islands are associated with events of genomic rearrangements and the acquisition and or interruption of important genes for bacteria, with direct involvement in diversity and genomic organization and, consequently, the differentiation between strains. The extent and impact of these events are the focus of this work, through in-silico analysis of integrase and association with regions of prophages and genomic islands in the genome of four strains of Xf. The data presented here support the role of integrases and their mobile genetic elements associated as key players in the process of evolution and diversity of genomic organization between the four strains of Xf.

<p>U ovom radu vr&scaron;ena je izolacija sojeva<em>&nbsp; P. shigelloides</em>&nbsp; iz uzoraka povr&scaron;inskih voda<br />Panonske nizije i formirana je kolekcija kultura, nakon čega je obavljena karakterizacija<br />faktora virulencije i antibiotske rezistencije&nbsp; izolovanih sojeva. Pored toga vr&scaron;ena je<br />izolacija bakteriofaga specifičnih za vrstu <em>P. shigelloides</em> iz uzoraka povr&scaron;inskih i otpadnih voda kao i njihova karakterizacija. Karakterizacija je podrazumevala sledeće testove: ispitivanje efikasnosti lize, litičkog spektra, uticaja različitih ekolo&scaron;kih faktora,<br />određivanje proteinskog profila i karakterizaciju genoma. Pored toga, vr&scaron;ena je detekcija<br />izolovanih bakteriofaga kori&scaron;ćenjem protočnog citometra. Rezultati ovog rada su potvrdili<br />prisustvo bakterije <em>P. shigelloides</em>&nbsp; i njenih specifičnih bakteriofaga u povr&scaron;inskim&nbsp; vodama Panonske nizije. Budući da su bakteriofagi specifični za vrstu&nbsp;<em> P. shigelloides&nbsp;</em> po prvi put izolovani, rezultati ovog rada mogu predstavljajati prvi&nbsp; korak u ispitivanjima vezanim za regulaciju brojnosti ove bakterije putem primene bakteriofaga.</p> / <p>We analyzed surface waters collected in Panonian plane, for the presence of bacterium&nbsp; <em>P. shigelloides</em>. The bacterial strains were preserved and characterized for virulence factors and antibiotic resistance. In addition, isolation of&nbsp; <em>P. shigelloides</em>&nbsp; specific bacteriophages was performed. Characterization of isolated phages included the following tests: examination of lytic efficacy, lytic spectrum, the influence of various ecological factors, the determination of the protein profile and the characterization of the genome. In addition, isolated bacteriophages were detected using a flow cytometer. The results of this paper demonstrate that the surface waters of Panonian plane contain bacterium&nbsp; <em>P. shigelloides&nbsp;</em> and its specific bacteriophages. Since&nbsp; <em>P. shigelloides</em>&nbsp; &ndash;&nbsp; specific bacteriophages are for the&nbsp;first time isolated, the results of this paper can represent the first step in the studies related to the regulation of the number of this bacterium by the application of bacteriophages.</p>

No description available.