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1.
Viruses ; 12(6)2020 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-32486497

RESUMO

Acinetobacter baumannii is an opportunistic pathogen that presents a serious clinical challenge due to its increasing resistance to all available antibiotics. Phage therapy has been introduced recently to treat antibiotic-incurable A. baumannii infections. In search for new A. baumannii specific bacteriophages, 20 clinical A. baumannii strains were used in two pools in an attempt to enrich phages from sewage. The enrichment resulted in induction of resident prophage(s) and three temperate bacteriophages, named vB_AbaS_fEg-Aba01, vB_AbaS_fLi-Aba02 and vB_AbaS_fLi-Aba03, all able to infect only one strain (#6597) of the 20 clinical strains, were isolated. Morphological characteristics obtained by transmission electron microscopy together with the genomic information revealed that the phages belong to the family Siphoviridae. The ca. 35 kb genomic sequences of the phages were >99% identical to each other. The linear ds DNA genomes of the phages contained 10 nt cohesive end termini, 52-54 predicted genes, an attP site and one tRNA gene each. A database search revealed an >99% identical prophage in the genome of A. baumannii strain AbPK1 (acc. no. CP024576.1). Over 99% identical prophages were also identified from two of the original 20 clinical strains (#5707 and #5920) and both were shown to be spontaneously inducible, thus very likely being the origins of the isolated phages. The phage vB_AbaS_fEg-Aba01 was also able to lysogenize the susceptible strain #6597 demonstrating that it was fully functional. The phages showed a very narrow host range infecting only two A. baumannii strains. In conclusion, we have isolated and characterized three novel temperate Siphoviridae phages that infect A. baumannii.

2.
Viruses ; 12(6)2020 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-32517038

RESUMO

YerA41 is a Myoviridae bacteriophage that was originally isolated due its ability to infect Yersinia ruckeri bacteria, the causative agent of enteric redmouth disease of salmonid fish. Several attempts to determine its genomic DNA sequence using traditional and next generation sequencing technologies failed, indicating that the phage genome is modified in such a way that it is an unsuitable template for PCR amplification and for conventional sequencing. To determine the YerA41 genome sequence, we performed RNA-sequencing from phage-infected Y. ruckeri cells at different time points post-infection. The host-genome specific reads were subtracted and de novo assembly was performed on the remaining unaligned reads. This resulted in nine phage-specific scaffolds with a total length of 143 kb that shared only low level and scattered identity to known sequences deposited in DNA databases. Annotation of the sequences revealed 201 predicted genes, most of which found no homologs in the databases. Proteome studies identified altogether 63 phage particle-associated proteins. The RNA-sequencing data were used to characterize the transcriptional control of YerA41 and to investigate its impact on the bacterial gene expression. Overall, our results indicate that RNA-sequencing can be successfully used to obtain the genomic sequence of non-sequencable phages, providing simultaneous information about the phage-host interactions during the process of infection.

3.
Viruses ; 12(5)2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32429141

RESUMO

The lytic phage, fHe-Kpn01 was isolated from sewage water using an extended-spectrum beta-lactamase-producing strain of Klebsiella pneumoniae as a host. The genome is 43,329 bp in size and contains direct terminal repeats of 222 bp. The genome contains 56 predicted genes, of which proteomics analysis detected 29 different proteins in purified phage particles. Comparison of fHe-Kpn01 to other phages, both morphologically and genetically, indicated that the phage belongs to the family Podoviridae and genus Drulisvirus. Because fHe-Kpn01 is strictly lytic and does not carry any known resistance or virulence genes, it is suitable for phage therapy. It has, however, a narrow host range since it infected only three of the 72 tested K. pneumoniae strains, two of which were of capsule type KL62. After annotation of the predicted genes based on the similarity to genes of known function and proteomics results on the virion-associated proteins, 22 gene products remained annotated as hypothetical proteins of unknown function (HPUF). These fHe-Kpn01 HPUFs were screened for their toxicity in Escherichia coli. Three of the HPUFs, encoded by the genes g10, g22, and g38, were confirmed to be toxic.

4.
Viruses ; 12(2)2020 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-31979276

RESUMO

Emergence of antibiotic-resistant bacteria is a serious threat to the public health. This is also true for Staphylococcus aureus and other staphylococci. Staphylococcus phages Stab20, Stab21, Stab22, and Stab23, were isolated in Albania. Based on genomic and phylogenetic analysis, they were classified to genus Kayvirus of the subfamily Twortvirinae. In this work, we describe the in-depth characterization of the phages that electron microscopy confirmed to be myoviruses. These phages showed tolerance to pH range of 5.4 to 9.4, to maximum UV radiation energy of 25 µJ/cm2, to temperatures up to 45 °C, and to ethanol concentrations up to 25%, and complete resistance to chloroform. The adsorption rate constants of the phages ranged between 1.0 × 10-9 mL/min and 4.7 × 10-9 mL/min, and the burst size was from 42 to 130 plaque-forming units. The phages Stab20, 21, 22, and 23, originally isolated using Staphylococcus xylosus as a host, demonstrated varied host ranges among different Staphylococcus strains suggesting that they could be included in cocktail formulations for therapeutic or bio-control purpose. Phage particle proteomes, consisting on average of ca 60-70 gene products, revealed, in addition to straight-forward structural proteins, also the presence of enzymes such DNA polymerase, helicases, recombinases, exonucleases, and RNA ligase polymer. They are likely to be injected into the bacteria along with the genomic DNA to take over the host metabolism as soon as possible after infection.

5.
Microb Pathog ; 141: 103993, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31988008

RESUMO

Yersinia pestis, a Gram-negative bacterium, is the etiologic agent of plague. A hallmark of Y. pestis infection is the organism's ability to rapidly disseminate through an animal host. Y. pestis expresses the outer membrane protein, Ail (Attachment invasion locus), which is associated with host invasion and serum resistance. However, whether Ail plays a role in host dissemination remains unclear. In this study, C57BL/6J mice were challenged with a defined Y. pestis strain, KimD27, or an isogenic ail-deleted mutant derived from KimD27 via metacarpal paw pad inoculation, nasal drops, orogastric infection, or tail vein injection to mimic bubonic, pneumonic, oral, or septicemic plague, respectively. Our results showed that ail-deleted Y. pestis KimD27 lost the ability to invade host cells, leading to failed host dissemination in the pneumonic and oral plague models but not in the bubonic or septicemic plague models, which do not require invasiveness. Therefore, this study demonstrated that whether Ail plays a role in Y. pestis pathogenesis depends on the infection route.

6.
Viruses ; 11(12)2019 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-31795231

RESUMO

One of the human- and animal-pathogenic species in genus Yersinia is Yersinia enterocolitica, a food-borne zoonotic pathogen that causes enteric infections, mesenteric lymphadenitis, and sometimes sequelae such as reactive arthritis and erythema nodosum. Y. enterocolitica is able to proliferate at 4 C, making it dangerous if contaminated food products are stored under refrigeration. The most common source of Y. enterocolitica is raw pork meat. Microbiological detection of the bacteria from food products is hampered by its slow growth rate as other bacteria overgrow it. Bacteriophages can be exploited in several ways to increase food safety with regards to contamination by Y. enterocolitica. For example, Yersinia phages could be useful in keeping the contamination of food products under control, or, alternatively, the specificity of the phages could be exploited in developing rapid and sensitive diagnostic tools for the identification of the bacteria in food products. In this review, we will discuss the present state of the research on these topics.

7.
Viruses ; 11(11)2019 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-31739448

RESUMO

The rapid emergence of antibiotic resistance among many pathogenic bacteria has created a profound need to discover new alternatives to antibiotics. Bacteriophages, the viruses of microbes, express special proteins to overtake the metabolism of the bacterial host they infect, the best known of which are involved in bacterial lysis. However, the functions of majority of bacteriophage encoded gene products are not known, i.e., they represent the hypothetical proteins of unknown function (HPUFs). In the current study we present a phage genomics-based screening approach to identify phage HPUFs with antibacterial activity with a long-term goal to use them as leads to find unknown targets to develop novel antibacterial compounds. The screening assay is based on the inhibition of bacterial growth when a toxic gene is expression-cloned into a plasmid vector. It utilizes an optimized plating assay producing a significant difference in the number of transformants after ligation of the toxic and non-toxic genes into a cloning vector. The screening assay was first tested and optimized using several known toxic and non-toxic genes. Then, it was applied to screen 94 HPUFs of bacteriophage φR1-RT, and identified four HPUFs that were toxic to Escherichia coli. This optimized assay is in principle useful in the search for bactericidal proteins of any phage, and also opens new possibilities to understanding the strategies bacteriophages use to overtake bacterial hosts.

8.
Microbiol Resour Announc ; 8(38)2019 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-31537666

RESUMO

We report the genomic sequences of phages KpCHEMY26 and KpGranit, isolated in Israel during a worldwide effort against a multidrug- and phage-resistant strain of Klebsiella pneumoniae from a patient in Finland. These results demonstrate the importance of an efficient worldwide network for collaborating in personalized therapy for infectious diseases.

9.
Front Microbiol ; 10: 1674, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31396188

RESUMO

The production of phages for therapeutic purposes demands fast, efficient and scalable purification procedures. Phage lysates have a wide range of impurities, of which endotoxins of gram-negative bacteria and protein toxins produced by many pathogenic bacterial species are harmful to humans. The highest allowed endotoxin concentration for parenterally applied medicines is 5 EU/kg/h. The aim of this study was to evaluate the feasibility of different purification methods in endotoxin and protein toxin removal in the production of phage preparations for clinical use. In the purification assays, we utilized three phages: Escherichia phage vB_EcoM_fHoEco02, Acinetobacter phage vB_ApiM_fHyAci03, and Staphylococcus phage vB_SauM_fRuSau02. The purification methods tested in the study were precipitation with polyethylene glycol, ultracentrifugation, ultrafiltration, anion exchange chromatography, octanol extraction, two different endotoxin removal columns, and different combinations thereof. The efficiency of the applied purification protocols was evaluated by measuring phage titer and either endotoxins or staphylococcal enterotoxins A and C (SEA and SEC, respectively) from samples taken from different purification steps. The most efficient procedure in endotoxin removal was the combination of ultrafiltration and EndoTrap HD affinity column, which was able to reduce the endotoxin-to-phage ratio of vB_EcoM_fHoEco02 lysate from 3.5 × 104 Endotoxin Units (EU)/109 plaque forming units (PFU) to 0.09 EU/109 PFU. The combination of ultrafiltration and anion exchange chromatography resulted in ratio 96 EU/109 PFU, and the addition of octanol extraction step into this procedure still reduced this ratio threefold. The other methods tested either resulted to less efficient endotoxin removal or required the use of harmful chemicals that should be avoided when producing phage preparations for medical use. Ultrafiltration with 100,000 MWCO efficiently removed enterotoxins from vB_SauM_fRuSau02 lysate (from 1.3 to 0.06 ng SEA/109 PFU), and anion exchange chromatography reduced the enterotoxin concentration below 0.25 ng/ml, the detection limit of the assay.

10.
PLoS One ; 14(7): e0219422, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31287844

RESUMO

Yersinia pseudotuberculosis is an important foodborne pathogen threatening modern food safety due to its ability to survive and grow at low temperatures. DEAD-box RNA helicase CsdA has been shown to play an important role in the low-temperature growth of psychrotrophic Y. pseudotuberculosis. A total of five DEAD-box RNA helicase genes (rhlB, csdA, rhlE, dbpA, srmB) have been identified in Y. pseudotuberculosis IP32953. However, their role in various stress conditions used in food production is unclear. We studied the involvement of the DEAD-box RNA helicase-encoding genes in the cold tolerance of Y. pseudotuberculosis IP32953 using quantitative real-time reverse transcription (RT-qPCR) and mutational analysis. Quantitative RT-PCR revealed that mRNA transcriptional levels of csdA, rhlE, dbpA and srmB were significantly higher after cold shock at 3°C compared to non-shocked culture at 28°C, suggesting the involvement of these four genes in cold shock response at the transcriptional level. The deletion of csdA ceased growth, while the deletion of dbpA or srmB significantly impaired growth at 3°C, suggesting the requirement of these three genes in Y. pseudotuberculosis at low temperatures. Growth of each DEAD-box RNA helicase mutant was also investigated under pH, osmotic, ethanol and oxidative stress conditions. The five helicase-encoding genes did not play major roles in the growth of Y. pseudotuberculosis IP32953 under pH, osmotic, ethanol or oxidative stress.


Assuntos
Temperatura Baixa , RNA Helicases DEAD-box/genética , Etanol/metabolismo , Concentração de Íons de Hidrogênio , Pressão Osmótica , Estresse Oxidativo , Yersinia pseudotuberculosis/genética , Yersinia pseudotuberculosis/metabolismo , Mutação
11.
Arch Virol ; 164(8): 2197-2199, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31123962

RESUMO

We present here the isolation and characterization of Acinetobacter pittii phage vB_ApiM_fHyAci03 (fHyAci03), which belongs to the family Myoviridae. The fHyAci03 genome was found to be 165,975 bp in length and predicted to contain 255 genes. While the whole genome was 92.4% identical to Acinetobacter baumannii phage KARL-1, phylogenetic analysis based on phage long distal tail fiber amino acid sequences assigned fHyAci03 and KARL-1 to different subclusters, reflecting their different host species. Together with phylogenetic analysis, genome comparisons indicated that fHyAci03 is a novel member of the subfamily Tevenvirinae. Host range experiments revealed that fHyAci03 could infect two clinical strains of Acinetobacter nosocomialis and six clinical strains of A. pittii. Thus, fHyAci03 is a novel lytic phage that infects clinical Acinetobacter strains and represents a potential new candidate to be used in phage therapy.


Assuntos
Acinetobacter/virologia , Bacteriófagos/genética , DNA Viral/genética , Genoma Viral/genética , Especificidade de Hospedeiro/genética , Myoviridae/genética , Filogenia
12.
Arch Virol ; 164(8): 2171-2173, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31093759

RESUMO

We report here the annotation of the complete genomes of four novel lytic Staphylococcus phages; Stab20, Stab21, Stab22 and Stab23. These phages have double-stranded DNA genomes ranging between 153,338 and 155,962 bp in size with terminal repeats of 10,814-12,304 bp. The genome analysis suggests that they represent new phage species within the genus Kayvirus in the subfamily Twortvirinae of the family Herelleviridae.


Assuntos
Myoviridae/genética , Staphylococcus/genética , DNA Viral/genética , Genoma Viral/genética , Genômica/métodos , Filogenia , Análise de Sequência de DNA/métodos , Fagos de Staphylococcus/genética
13.
Infect Immun ; 87(8)2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31085704

RESUMO

Salmonella enterica serovar Typhimurium, a Gram-negative bacterium, can cause infectious diseases ranging from gastroenteritis to systemic dissemination and infection. However, the molecular mechanisms underlying this bacterial dissemination have yet to be elucidated. A study indicated that using the lipopolysaccharide (LPS) core as a ligand, S Typhimurium was able to bind human dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (hCD209a), an HIV receptor that promotes viral dissemination by hijacking antigen-presenting cells (APCs). In this study, we showed that S Typhimurium interacted with CD209s, leading to the invasion of APCs and potentially the dissemination to regional lymph nodes, spleen, and liver in mice. Shielding of the exposed LPS core through the expression of O-antigen reduces dissemination and infection. Thus, we propose that similar to HIV, S Typhimurium may also utilize APCs via interactions with CD209s as a way to disseminate to the lymph nodes, spleen, and liver to initiate host infection.


Assuntos
Moléculas de Adesão Celular/fisiologia , Lectinas Tipo C/fisiologia , Receptores de Superfície Celular/fisiologia , Salmonella typhimurium/patogenicidade , Animais , Células Apresentadoras de Antígenos/microbiologia , Feminino , Interações Hospedeiro-Patógeno , Humanos , Lipopolissacarídeos/fisiologia , Mananas/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Antígenos O/fisiologia , Nódulos Linfáticos Agregados/fisiologia , Fagocitose , Células RAW 264.7
14.
Front Immunol ; 10: 96, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30915064

RESUMO

Yersinia pestis, a Gram-negative bacterium and the etiologic agent of plague, has evolved from Yersinia pseudotuberculosis, a cause of a mild enteric disease. However, the molecular and biological mechanisms of how Y. pseudotuberculosis evolved to such a remarkably virulent pathogen, Y. pestis, are not clear. The ability to initiate a rapid bacterial dissemination is a characteristic hallmark of Y. pestis infection. A distinguishing characteristic between the two Yersinia species is that Y. pseudotuberculosis strains possess an O-antigen of lipopolysaccharide (LPS) while Y. pestis has lost the O-antigen during evolution and therefore exposes its core LPS. In this study, we showed that Y. pestis utilizes its core LPS to interact with SIGNR1 (CD209b), a C-type lectin receptor on antigen presenting cells (APCs), leading to bacterial dissemination to lymph nodes, spleen and liver, and the initiation of a systemic infection. We therefore propose that the loss of O-antigen represents a critical step in the evolution of Y. pseudotuberculosis into Y. pestis in terms of hijacking APCs, promoting bacterial dissemination and causing the plague.


Assuntos
Moléculas de Adesão Celular/imunologia , Interações Hospedeiro-Patógeno/imunologia , Lectinas Tipo C/imunologia , Lipopolissacarídeos/imunologia , Peste/imunologia , Receptores de Superfície Celular/imunologia , Yersinia pestis/fisiologia , Animais , Células Apresentadoras de Antígenos/imunologia , Moléculas de Adesão Celular/genética , Linhagem Celular , Feminino , Células HeLa , Humanos , Lectinas Tipo C/genética , Macrófagos/imunologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Superfície Celular/genética , Yersinia pseudotuberculosis/fisiologia , Infecções por Yersinia pseudotuberculosis/imunologia
15.
Front Microbiol ; 10: 150, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30787919

RESUMO

Background: Alpha-mangostin (α-MG) is a natural xanthone reported to exhibit rapid bactericidal activity against Gram-positive bacteria, and may therefore have potential clinical application in healthcare sectors. This study sought to identify the impact of α-MG on Staphylococcus epidermidis RP62A through integrated advanced omic technologies. Methods: S. epidermidis was challenged with sub-MIC (0.875 µg/ml) of α-MG at various time points and the differential expression pattern of genes/proteins were analyzed in the absence and presence of α-MG using RNA sequencing and LC-MS/MS experiments. Bioinformatic tools were used to categorize the biological processes, molecular functions and KEGG pathways of differentially expressed genes/proteins. qRT-PCR was employed to validate the results obtained from these analyses. Results: Transcriptomic and proteomic profiling of α-MG treated cells indicated that genes/proteins affected by α-MG treatment were associated with diverse cellular functions. The greatest reduction in expression was observed in transcription of genes conferring cytoplasmic membrane integrity (yidC2, secA and mscL), cell division (ftsY and divlB), teichoic acid biosynthesis (tagG and dltA), fatty-acid biosynthesis (accB, accC, fabD, fabH, fabI, and fabZ), biofilm formation (icaA) and DNA replication and repair machinery (polA, polC, dnaE, and uvrA). Those with increased expression were involved in oxidative (katA and sodA) and cellular stress response (clpB, clpC, groEL, and asp23). The qRT-PCR analysis substantiated the results obtained from transcriptomic and proteomic profiling studies. Conclusion: Combining transcriptomic and proteomic methods provided comprehensive information about the antibacterial mode of action of α-MG. The obtained results suggest that α-MG targets S. epidermidis through multifarious mechanisms, and especially prompts that loss of cytoplasmic membrane integrity leads to rapid onset of bactericidal activity.

16.
Infect Immun ; 87(1)2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30348825

RESUMO

Yersinia pseudotuberculosis is a Gram-negative enteropathogen and causes gastrointestinal infections. It disseminates from gut to mesenteric lymph nodes (MLNs), spleen, and liver of infected humans and animals. Although the molecular mechanisms for dissemination and infection are unclear, many Gram-negative enteropathogens presumably invade the small intestine via Peyer's patches to initiate dissemination. In this study, we demonstrate that Y. pseudotuberculosis utilizes its lipopolysaccharide (LPS) core to interact with CD209 receptors, leading to invasion of human dendritic cells (DCs) and murine macrophages. These Y. pseudotuberculosis-CD209 interactions result in bacterial dissemination to MLNs, spleens, and livers of both wild-type and Peyer's patch-deficient mice. The blocking of the Y. pseudotuberculosis-CD209 interactions by expression of O-antigen and with oligosaccharides reduces infectivity. Based on the well-documented studies in which HIV-CD209 interaction leads to viral dissemination, we therefore propose an infection route for Y. pseudotuberculosis where this pathogen, after penetrating the intestinal mucosal membrane, hijacks the Y. pseudotuberculosis-CD209 interaction antigen-presenting cells to reach their target destinations, MLNs, spleens, and livers.


Assuntos
Moléculas de Adesão Celular/metabolismo , Células Dendríticas/microbiologia , Endocitose , Interações Hospedeiro-Patógeno , Lectinas Tipo C/metabolismo , Lipopolissacarídeos/metabolismo , Macrófagos/microbiologia , Receptores de Superfície Celular/metabolismo , Yersinia pseudotuberculosis/patogenicidade , Animais , Aderência Bacteriana , Células Cultivadas , Modelos Animais de Doenças , Humanos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Ligação Proteica , Yersiniose/microbiologia , Yersiniose/patologia , Yersiniose/fisiopatologia
17.
PeerJ ; 6: e5470, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30155366

RESUMO

Background: Extraintestinal pathogenic Escherichia coli (ExPEC) is an important bacterium and responsible for many bloodstream infections, including urinary tract infections and even fatal bacteremia. The aim of this research was to investigate whether ExPEC strains isolated from Turkish blood cultures have a relationship between 16S rRNA based phylogenetic clusters and antibiotic resistance profiles, virulence factors or clonal lineages. Methods: Phenotypically identified ExPEC blood culture isolates (n = 104) were included in this study. The 16S rRNA partial sequence analysis was performed for genotypic identification of ExPEC isolates. Antibiotic susceptibility and Extended-Spectrum ß-Lactamase testing of isolates were performed. Phylogenetic classification (A, B1, B2 and D), Multi Locus Sequence Typing analysis and virulence-associated genes were investigated. Results: Based on 16S rRNA partial sequence analysis, 97 out of 104 (93.26%) ExPEC isolates were confirmed as E. coli. Ampicillin (74.22%) and cefuroxime axetil (65.97%) resistances had the highest frequencies among the ExPEC isolates. In terms of phylogenetic classification of ExPEC, D (38.14%, 37/97) was the most prevalent group after A (29.89%, 29/97), B2 (20.61%, 20/97), and B1 (11.34%, 11/97). The sequence types of the 20 ExPEC isolates belonging to the B2 phylogenetic group were analyzed by Multi Locus Sequence Typing. Ten isolates out of 20 (50.0%) were identified as ST131. The other STs were ST95 (n = 1), ST14 (n = 1), ST10 (n = 1), ST69 (n = 1), ST1722 (n = 2), ST141 (n = 1), ST88 (n = 1), ST80 (n = 1), and ST998 (n = 1). Of the ST131 strains, six (60%, 6/10) represented serogroup O25. The most common virulence factor genes were serum resistance factor gene, traT (55.7%) aerobactin siderophore receptor and yersiniabactin encoding genes iutA (45.3%) and fyuA (50.5%), respectively. In addition, PAI (41.2%), iroN (23.7%), hlyA (15.4%), kpsII (13.4%), ompT (13.4%), papG (12.4%), iss (9.3%), cnf1 (7.2%), ibeA (2.06%), and sfaS (2.06%) genes were present in the ExPEC isolates. Conclusion: The 16S rRNA-based phylogenetic relationship tree analysis showed that a large cluster was present among 97 ExPEC isolates along with related reference strains. There were 21 main clusters with 32 closely related subclusters. Based on our findings, different clonal lineages of ExPEC can display different antibiotic susceptibilities and virulence properties. We also concluded that virulence factors were not distributed depending on phylogenetic groups (A, B1, B2, and D). The ExPEC isolates belonging to the same phylogenetic group and sequence type could display different resistance and virulence characteristics.

18.
Genome Announc ; 6(22)2018 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-29853497

RESUMO

Escherichia phages vB_EcoM-fFiEco06 and vB_EcoM-fHoEco02 were found to have 167,076-bp and 167,064-bp genomes, respectively. They are members of genus T4virus, and they are 99.96% identical to each other. The host ranges of the phages are different, probably due to a few differences in their tail protein amino acid sequences.

19.
Viruses ; 10(4)2018 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-29614052

RESUMO

Yersinia enterocolitica causes enteric infections in humans and animals. Human infections are often caused by contaminated pork meat. Y. enterocolitica colonizes pig tonsils and pigs secrete both the human pathogen and its specific bacteriophages into the stools. In this work, sixteen Y. enterocolitica-infecting lytic bacteriophages isolated from pig stools originating from several pig farms were characterized. All phages belong to the Podoviridae family and their genomes range between 38,391-40,451 bp in size. The overall genome organization of all the phages resembled that of T7-like phages, having 3-6 host RNA polymerase (RNAP)-specific promoters at the beginning of the genomes and 11-13 phage RNAP-specific promoters as well as 3-5 rho-independent terminators, scattered throughout the genomes. Using a ligation-based approach, the physical termini of the genomes containing direct terminal repeats of 190-224 bp were established. No genes associated with lysogeny nor any toxin, virulence factor or antibiotic resistance genes were present in the genomes. Even though the phages had been isolated from different pig farms the nucleotide sequences of their genomes were 90-97% identical suggesting that the phages were undergoing microevolution within and between the farms. Lipopolysaccharide was found to be the surface receptor of all but one of the phages. The phages are classified as new species within the T7virus genus of Autographivirinae subfamily.


Assuntos
Genoma Viral , Genômica , Podoviridae/fisiologia , Doenças dos Suínos/microbiologia , Yersiniose/veterinária , Yersinia enterocolitica/virologia , Animais , Bacteriólise , Biologia Computacional/métodos , Evolução Molecular , Genômica/métodos , Humanos , Anotação de Sequência Molecular , Filogenia , Podoviridae/classificação , Podoviridae/isolamento & purificação , Podoviridae/ultraestrutura , Suínos , Sequenciamento Completo do Exoma
20.
Int J Food Microbiol ; 271: 33-47, 2018 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-29477807

RESUMO

Yersinia enterocolitica, the primary cause of yersiniosis, is one of the most important foodborne pathogens globally and is associated with the consumption of raw contaminated pork. In the current study, four virulent bacteriophages (phages), one of Podoviridae (fHe-Yen3-01) and three of Myoviridae (fHe-Yen9-01, fHe-Yen9-02, and fHe-Yen9-03), capable of infecting Y. enterocolitica were isolated and characterized. fHe-Yen9-01 had the broadest host range (61.3% of strains, 65/106). It demonstrated a latent period of 35 min and a burst size of 33 plaque-forming units/cell, and was found to have a genome of 167,773 bp with 34.79% GC content. To evaluate the effectiveness of phage fHe-Yen9-01 against Y. enterocolitica O:9 strain Ruokola/71, we designed an experimental model of the food market environment. Phage treatment after bacterial inoculation of food samples, including raw pork (4 °C, 72 h), ready-to-eat pork (26 °C, 12 h), and milk (4 °C, 72 h), prevented bacterial growth throughout the experiments, with counts decreasing by 1-3 logs from the original levels of 2-4 × 103 CFU/g or ml. Similarly, when artificially contaminated kitchen utensils, such as wooden and plastic cutting boards and knives, and artificial hands, were treated with phages for 2 h, bacterial growth was effectively inhibited, with counts decreasing by 1-2 logs from the original levels of ca 104 CFU/cm2 or ml. To the best of our knowledge, this is the first report of the successful application of phages for the control of Y. enterocolitica growth in food and on kitchen utensils.


Assuntos
Bacteriófagos/patogenicidade , Contaminação de Alimentos/prevenção & controle , Carne Vermelha/microbiologia , Yersiniose/prevenção & controle , Yersinia enterocolitica/crescimento & desenvolvimento , Yersinia enterocolitica/virologia , Animais , Bacteriófagos/crescimento & desenvolvimento , Bacteriófagos/isolamento & purificação , Feminino , Microbiologia de Alimentos , Especificidade de Hospedeiro , Camundongos , Camundongos Endogâmicos BALB C , Leite/microbiologia , Suínos , Yersiniose/microbiologia
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