Agnieszka Necel - Academia.edu (original) (raw)

Papers by Agnieszka Necel

Frontiers in Cellular and Infection Microbiology

Phage therapy is a promising alternative treatment of bacterial infections in human and animals. ... more Phage therapy is a promising alternative treatment of bacterial infections in human and animals. Nevertheless, despite the appearance of many bacterial strains resistant to antibiotics, these drugs still remain important therapeutics used in human and veterinary medicine. Although experimental phage therapy of infections caused by Salmonella enterica was described previously by many groups, those studies focused solely on effects caused by bacteriophages. Here, we compared the use of phage therapy (employing a cocktail composed of two previously isolated and characterized bacteriophages, vB_SenM-2 and vB_Sen-TO17) and antibiotics (enrofloxacin and colistin) in chickens infected experimentally with S. enterica serovar Typhimurium. We found that the efficacies of both types of therapies (i.e. the use of antibiotics and phage cocktail) were high and very similar to one another when the treatment was applied shortly (one day) after the infection. Under these conditions, S. Typhimurium w...

1“GERMS”The University of Gdansk Student Chapter of the American Society for Microbiology, affili... more 1“GERMS”The University of Gdansk Student Chapter of the American Society for Microbiology, affiliated with the Department of Molecular Biology, University of Gdansk, Wita Stwosza 59, 80-308 Gdańsk, Poland; 2Department of Molecular Biology, University of Gdansk, Wita Stwosza 59, 80-308 Gdańsk, Poland; 3Laboratory of Molecular Biology affiliated with University of Gdansk, Institute of Biochemistry and Biophysics Polish Academy of Sciences, Wita Stwosza 59, 80-308 Gdańsk, Poland

Phage Therapy: A Practical Approach, 2019

Bacteriophage therapy is usually considered as a procedure to treat human diseases. However, the ... more Bacteriophage therapy is usually considered as a procedure to treat human diseases. However, the use of bacteriophages to treat bacterial infections in animals is perhaps more advanced than human phage therapy. This is due to the fact that formal regulations on the use of various methods in treatment of animals are considerably less stringent than those for human therapies. This applies also to phage therapy. On the other hand, from studies on animals and development of phage therapy for animals, we can learn a lot about mechanisms and efficacy of application of phages to treat various infections. In this chapter, current stage of development of phage therapy for animals is presented and discussed. Applications of phages in animal breeding, aviculture, and aquaculture are described. The use of phages in food protection is also mentioned. Methods for phage isolation, propagation, purification, and administration in animal phage therapy are also briefly described and discussed.

Acta Biochimica Polonica, 2019

Shiga toxin-producing Escherichia coli (STEC) is a group of pathogenic strains responsible for hu... more Shiga toxin-producing Escherichia coli (STEC) is a group of pathogenic strains responsible for human infections that result in bloody diarrhea and hemorrhagic colitis, often with severe complications. The main virulence factors of STEC are Shiga toxins encoded by stx genes located in genomes of Shiga toxin-converting bacteriophages (Stx phages). These bacterial viruses are clustered in the lambdoid bacteriophages family represented by phage λ. Here, we report that expression of orf73 from the exo-xis region of the phage genome promotes the lysogenic pathway of development of λ and Φ24B phages. We demonstrated that the mutant phages with deletions of orf73 revealed higher burst size during the lytic cycle. Moreover, survival rates of E. coli infected with mutant bacteriophages were lower relative to wild-type viruses. Additionally, orf73 deletion negatively influenced the lysogenization process of E. coli host cells. We conclude that orf73 plays an important biological role in the de...

Toxins, 2021

Shiga toxin-producing Escherichia coli (STEC) can cause severe infections in humans, leading to s... more Shiga toxin-producing Escherichia coli (STEC) can cause severe infections in humans, leading to serious diseases and dangerous complications, such as hemolytic-uremic syndrome. Although cattle are a major reservoir of STEC, the most commonly occurring source of human infections are food products (e.g., vegetables) contaminated with cow feces (often due to the use of natural fertilizers in agriculture). Since the use of antibiotics against STEC is controversial, other methods for protection of food against contaminations by these bacteria are required. Here, we propose a validation system for selection of bacteriophages against STEC contamination. As a model system, we have employed a STEC-specific bacteriophage vB_Eco4M-7 and the E. coli O157:H7 strain no. 86-24, bearing Shiga toxin-converting prophage ST2-8624 (Δstx2::cat gfp). When these bacteria were administered on the surface of sliced cucumber (as a model vegetable), significant decrease in number viable E. coli cells was obse...

Viruses, 2021

Phage therapy is one of main alternative option for antibiotic treatment of bacterial infections,... more Phage therapy is one of main alternative option for antibiotic treatment of bacterial infections, particularly in the era of appearance of pathogenic strains revealing resistance to most or even all known antibiotics. Enterococcus faecalis is one of such pathogens causing serious human infections. In the light of high level of biodiversity of bacteriophages and specificity of phages to bacterial species or even strains, development of effective phage therapy depend, between others, on identification and characterization of a large collection of these viruses, including understanding of their interactions with host bacterial cells. Recently, isolation of molecular characterization of bacteriophage vB_EfaS-271, infecting E. faecalis strains have been reported. In this report, phage–host interactions are reported, including ability of vB_EfaS-271 to infect bacteria forming biofilms, efficiency of eliminating bacterial cells from cultures depending on multiplicity of infection (m.o.i.),...

Antibiotics, 2021

In addition to specific antibiotic resistance, the formation of bacterial biofilm causes another ... more In addition to specific antibiotic resistance, the formation of bacterial biofilm causes another level of complications in attempts to eradicate pathogenic or harmful bacteria, including difficult penetration of drugs through biofilm structures to bacterial cells, impairment of immunological response of the host, and accumulation of various bioactive compounds (enzymes and others) affecting host physiology and changing local pH values, which further influence various biological functions. In this review article, we provide an overview on the formation of bacterial biofilm and its properties, and then we focus on the possible use of phage-derived depolymerases to combat bacterial cells included in this complex structure. On the basis of the literature review, we conclude that, although these bacteriophage-encoded enzymes may be effective in destroying specific compounds involved in the formation of biofilm, they are rarely sufficient to eradicate all bacterial cells. Nevertheless, a ...

Virus Genes, 2020

The exo-xis region of lambdoid phages contains open reading frames and genes that appear to be ev... more The exo-xis region of lambdoid phages contains open reading frames and genes that appear to be evolutionarily important. However, this region has received little attention up to now. In this study, we provided evidence that ea22, the largest gene of this region, favors the lysogenic pathway over the lytic pathway in contrast to other characterized exo-xis region genes including ea8.5, orf61, orf60a, and orf63. Our assays also suggest some functional analogies between Ea22 and the phage integrase protein (Int). While it is unsurprising that Ea22 operates similarly in both λ and Stx phages, we have observed some distinctions that may arise from considerable sequence dissimilarity at the carboxy termini of each protein.

Viruses, 2018

The exo-xis region of lambdoid bacteriophage genomes contains several established and potential g... more The exo-xis region of lambdoid bacteriophage genomes contains several established and potential genes that are evolutionarily conserved, but not essential for phage propagation under laboratory conditions. Nevertheless, deletion or overexpression of either the whole exo-xis region and important regulatory elements can significantly influence the regulation of phage development. This report defines specific roles for orf60a and orf61 in bacteriophage λ and Φ24B, a specific Shiga toxin-converting phage with clinical relevance. We observed that mutant phages bearing deletions of orf60a and orf61 impaired two central aspects of phage development: the lysis-versus-lysogenization decision and prophage induction. These effects were more pronounced for phage Φ24B than for λ. Surprisingly, adsorption of phage Φ24B on Escherichia coli host cells was less efficient in the absence of either orf60a or orf61. We conclude that these open reading frames (ORFs) play important, but not essential, rol...

International Journal of Molecular Sciences, 2020

A newly isolated bacteriophage infecting Enterococcus faecalis strains has been characterized, in... more A newly isolated bacteriophage infecting Enterococcus faecalis strains has been characterized, including determination of its molecular features. This phage, named vB_EfaS-271, has been classified as a Siphoviridae member, according to electron microscopy characterization of the virions, composed of a 50 nm-diameter head and a long, flexible, noncontractable tail (219 × 12.5 nm). Analysis of the whole dsDNA genome of this phage showed that it consists of 40,197 bp and functional modules containing genes coding for proteins that are involved in DNA replication (including DNA polymerase/primase), morphogenesis, packaging and cell lysis. Mass spectrometry analysis allowed us to identify several phage-encoded proteins. vB_EfaS-271 reveals a relatively narrow host range, as it is able to infect only a few E. faecalis strains. On the other hand, it is a virulent phage (unable to lysogenize host cells), effectively and quickly destroying cultures of sensitive host bacteria, with a latent p...

Scientific Reports, 2020

The characterization of a recently isolated bacteriophage, vB_Eco4M-7, which effectively infects ... more The characterization of a recently isolated bacteriophage, vB_Eco4M-7, which effectively infects many, though not all, Escherichia coli O157 strains, is presented. The genome of this phage comprises double-stranded DNA, 68,084 bp in length, with a GC content of 46.2%. It contains 96 putative open reading frames (ORFs). Among them, the putative functions of only 35 ORFs were predicted (36.5%), whereas 61 ORFs (63.5%) were classified as hypothetical proteins. The genome of phage vB_Eco4M-7 does not contain genes coding for integrase, recombinase, repressors or excisionase, which are the main markers of temperate viruses. Therefore, we conclude that phage vB_Eco4M-7 should be considered a lytic virus. This was confirmed by monitoring phage lytic development by a one-step growth experiment. Moreover, the phage forms relatively small uniform plaques (1 mm diameter) with no properties of lysogenization. Electron microscopic analyses indicated that vB_Eco4M-7 belongs to the Myoviridae fami...

Frontiers in Microbiology, 2019

Morphological, biological, and genetic characteristics of a virulent Siphoviridae phage, named vB... more Morphological, biological, and genetic characteristics of a virulent Siphoviridae phage, named vB-EcoS-95, is reported. This phage was isolated from urban sewage. It was found to infect some Escherichia coli strains giving clear plaques. The genome of this phage is composed of 50,910 bp and contains 89 ORFs. Importantly, none of the predicted ORFs shows any similarity with known pathogenic factors that would prevent its use in medicine. Genome sequence analysis of vB-EcoS-95 revealed 74% similarity to genomic sequence of Shigella phage pSf-1. Compared to pSf-1, phage vb-EcoS-95 does not infect Shigella strains and has an efficient bacteriolytic activity against some E. coli strains. One-step growth analysis revealed that this phage has a very short latent period (4 min), and average burst size of 115 plaque forming units per cell, which points to its high infectivity of host cells and strong lytic activity. The bacteriolytic effect of vB-EcoS-95 was tested also on biofilm-producing strains. These results indicate that vB-EcoS-95 is a newly discovered E. coli phage that may be potentially used to control the formation of biofilms.

Frontiers in Microbiology, 2017

Lambdoid bacteriophages form a group of viruses that shares a common schema of genome organizatio... more Lambdoid bacteriophages form a group of viruses that shares a common schema of genome organization and lifecycle. Some of them can play crucial roles in creating the pathogenic profiles of Escherichia coli strains. For example, Shiga toxin-producing E. coli (STEC) acquired stx genes, encoding Shiga toxins, via lambdoid prophages (Stx phages). The results obtained so far present the evidence for the relation between the exo-xis region of the phage genome and lambdoid phage development, however molecular mechanisms of activities of the exo-xis genes' products are still unknown. In view of this, we decided to determine the influence of the uncharacterized open reading frame orf63 of the exo-xis region on lambdoid phages development using recombinant prophages, λ and Stx phage 24 B. We have demonstrated that orf63 codes for a folded protein, thus, it is a functional gene. NMR spectroscopy and analytical gel filtration were used to extend this observation further. From backbone chemical shifts, Orf63 is oligomeric in solution, likely a trimer and consistent with its small size (63 aa.), is comprised of two helices, likely intertwined to form the oligomer. We observed that the deletion of phage orf63 does not impair the intracellular lambdoid phage lytic development, however delays the time and decreases the efficiency of prophage induction and in consequence results in increased survival of E. coli during phage lytic development. Additionally, the deletion of phage orf63 negatively influences expression of the major phage genes and open reading frames from the exo-xis region during prophage induction with hydrogen peroxide. We conclude, that lambdoid phage orf63 may have specific functions in the regulation of lambdoid phages development, especially at the stage of the lysis vs. lysogenization decision. Besides, orf63 probably participates in the regulation of the level of expression of essential phage genes and open reading frames from the exo-xis region during prophage induction.

Scientific Reports, 2016

A large scale analysis presented in this article focuses on biological and physiological variety ... more A large scale analysis presented in this article focuses on biological and physiological variety of bacteriophages. A collection of 83 bacteriophages, isolated from urban sewage and able to propagate in cells of different bacterial hosts, has been obtained (60 infecting Escherichia coli, 10 infecting Pseudomonas aeruginosa, 4 infecting Salmonella enterica, 3 infecting Staphylococcus sciuri, and 6 infecting Enterococcus faecalis). High biological diversity of the collection is indicated by its characteristics, both morphological (electron microscopic analyses) and biological (host range, plaque size and morphology, growth at various temperatures, thermal inactivation, sensitivity to low and high pH, sensitivity to osmotic stress, survivability upon treatment with organic solvents and detergents), and further supported by hierarchical cluster analysis. By the end of the research no larger collection of phages from a single environmental source investigated by these means had been foun...

Toxins, 2015

Shiga toxin-converting bacteriophages (Stx phages) are present as prophages in Shiga toxin-produc... more Shiga toxin-converting bacteriophages (Stx phages) are present as prophages in Shiga toxin-producing Escherichia coli (STEC) strains. Theses phages can be transmitted to previously non-pathogenic E. coli cells making them potential producers of Shiga toxins, as they bear genes for these toxins in their genomes. Therefore, sensitivity of Stx phage virions to various conditions is important in both natural processes of spreading of these viruses and potential prophylactic control of appearance of novel pathogenic E. coli strains. In this report we provide evidence that virions of Stx phages are significantly more sensitive to UV irradiation than bacteriophage λ. Following UV irradiation of Stx virions at the dose of 50 J/m 2 , their infectivity dropped by 1-3 log10, depending on the kind of phage. Under these conditions, a considerable release of phage DNA from virions was observed, and electron

Antibiotics

Apart from antibiotic resistance of pathogenic bacteria, the formation of biofilms is a feature t... more Apart from antibiotic resistance of pathogenic bacteria, the formation of biofilms is a feature that makes bacterial infections especially difficulty to treat. Shiga toxin-producing Escherichia coli (STEC) strains are dangerous pathogens, causing severe infections in humans, and capable of biofilm production. We have reported previously the identification and characterization of the vB_Eco4-M7 bacteriophage, infecting various STEC strains. It was suggested that this phage might be potentially used in phage therapy against these bacteria. Here, we tested the effects of vB_Eco4-M7 alone or in a phage cocktail with another STEC-infecting phage, and/or in a combination with different antibiotics (ciprofloxacin and rifampicin) on biofilm formed by a model STEC strain, named E. coli O157:H7 (ST2-8624). The vB_Eco4-M7 phage appeared effective in anti-biofilm action in all these experimental conditions (2–3-fold reduction of the biofilm density, and 2–3 orders of magnitude reduction of the ...