Order Caudovirales Research Articles

Insights into the global freshwater virome.

Viruses are by far the most abundant life forms on this planet. Yet, the full viral diversity remains mostly unknown, especially in environments like freshwater. Therefore, we aimed to study freshwater viruses in a global context. To this end, we downloaded 380 publicly available viral metagenomes (>1 TB). More than 60% of these metagenomes were discarded based on their levels of cellular contamination assessed by ribosomal DNA content. For the remaining metagenomes, assembled contigs were decontaminated using two consecutive steps, eventually yielding 273,365 viral contigs longer than 1,000 bp. Long enough contigs (≥ 10 kb) were clustered to identify novel genomes/genome fragments. We could recover 549 complete circular and high-quality draft genomes, out of which 10 were recognized as being novel. Functional annotation of these genomes showed that most of the annotated coding sequences are DNA metabolic genes or phage structural genes. On the other hand, taxonomic analysis of viral contigs showed that most of the assigned contigs belonged to the order Caudovirales, particularly the families of Siphoviridae, Myoviridae, and Podoviridae. The recovered viral contigs contained several auxiliary metabolic genes belonging to several metabolic pathways, especially carbohydrate and amino acid metabolism in addition to photosynthesis as well as hydrocarbon degradation and antibiotic resistance. Overall, we present here a set of prudently chosen viral contigs, which should not only help better understanding of freshwater viruses but also be a valuable resource for future virome studies.

Characterization of the Urinary Metagenome and Virome in Healthy Children.

Recent advances in next-generation sequencing and metagenomic studies have provided insights into the microbial profile of different body sites. However, research on the microbial composition of urine is limited, particularly in children. The goal of this study was to optimize and develop reproducible metagenome and virome protocols using a small volume of urine samples collected from healthy children. We collected midstream urine specimens from 40 healthy children. Using the metagenomics shotgun approach, we tested various protocols. Different microbial roots such as Archaea, Bacteria, Eukaryota, and Viruses were successfully identified using our optimized urine protocol. Our data reflected much variation in the microbial fingerprints of children. Girls had significantly higher levels of Firmicutes, whereas boys had significantly higher levels of Actinobacteria. The genus Anaerococcus dominated the urinary bacteriome of healthy girls, with a significant increase in Anaerococcus prevotii, Anaerococcus vaginalis, and Veillonella parvula (p-value < 0.001) when compared with that of boys. An increased relative abundance of Xylanimonas and Arthrobacter, with a significantly high abundance of Arthrobacter sp. FB24 (p-value 0.0028) and Arthrobacter aurescences (p-value 0.015), was observed in boys. The urinary mycobiome showed a significant rise in the genus Malassezia and Malassezia globose fungus (p-value 0.009) in girls, whereas genus Saccharomyces (p-value 0.009) was significantly high in boys. The beta diversity of the urinary mycobiome was found to differ between different age groups. Boys had significantly more Mastadenovirus and Human mastadenovirus-A in their urinary virome than girls. With increasing age, we noticed an increase in the relative abundance of the order Caudovirales. Our optimized protocols allowed us to identify the unique microbes for each sex by using an adequate volume of urine (3–10 mL) to screen for the bacteriome, mycobiome, and virome profiles in the urine of healthy children. To the best of our knowledge, this is the first study to characterize the metagenomics profiles of urine in a healthy pediatric population.

Characterisation of broad-spectrum phiKZ like jumbo phage and its utilisation in controlling multidrug-resistant Pseudomonas aeruginosa isolates

The emergence of highly virulent multidrug-resistant P. aeruginosa has become increasingly evident among hospital-acquired infections and has raised the need for alternative therapies. Phage therapy can be one such alternative to antibiotic therapy to combat multidrug-resistant pathogenic bacteria, but this requires the availability of phages with a broad host range. In this study, isolation and molecular characterisation of P. aeruginosa specific phages were carried out. A total of 17 phages isolated showed different spectra of activity and efficiency of lysis against 82 isolates of P. aeruginosa obtained from clinical samples (n = 13), hospital effluent (n = 46) and fish processing plant effluent (n = 23). Antibiotic susceptibility test results revealed multi-drug resistance in 61 of the total 82 isolates. Three new jumbo lytic P. aeruginosa specific broad host range phages were isolated and characterised in this present study belonged to the family Myoviridae (order Caudovirales). The genetic analysis of ɸU5 revealed that phage has a genome size of 282.6 kbp with 373 putative open reading frames (ORFs), and its genetic architecture is similar to phiKZ like jumbo phages infecting P. aeruginosa. The bacteriophages isolated in this study had lytic ability against biofilm-forming and multidrug-resistant P. aeruginosa and could be candidates for further studies towards phage therapy.

Genomic characterization of a novel bacteriophage STP55 revealed its prominent capacity in disrupting the dual-species biofilm formed by Salmonella Typhimurium and Escherichia coli O157: H7 strains.

Salmonella and Escherichia coli are important foodborne pathogens, forming bacterial biofilms that contribute to their virulence, antimicrobial resistance, and survival on surfaces. Broad lytic phages are promising alternatives to conventional technologies for pathogen biocontrol and reducing biofilms. Herein, we isolated and characterized a novel polyvalent phage STP55 that not only lyse some serotypes of Salmonella, but also some E. coli strains. It had a wide range of pH (4-12) and thermal (30-60°C) tolerances. The latent time was determined to be 10min in the one-step growth experiment. Morphological observations by transmission electron microscopy and phylogenetic analysis using terminase gene classified STP55 to family Ackermannviridae in the order Caudovirales, with a complex tail structure. The genome was found to comprise 157,708bp double-stranded DNA, with 44.57% GC content, 207 predicted ORFs and with no genes associated with antibiotic resistance, toxins, lysogeny, and virulence factors. Particularly, phage STP55 was able to inhibit single- and dual-species biofilms formation by S. Typhimurium ATCC 14028 and E. coli O157: H7, with a reduction percentage of 51.0%, 47.8% and 52.8%, respectively. Moreover, more than 65.0%, 72.9% and 46.2% of an established, single- and dual-species biofilms by S. Typhimurium ATCC 14028 and E. coli O157: H7 were removed after 8h exposure to the phage STP55, respectively. The elimination effect of STP55 on dual-species biofilm formed on lettuce was further observed by SEM. Overall, our results demonstrated that STP55 is a promising antimicrobial against Salmonella and E. coli.

Enterobacteria Phage Ac3's Genome Annotation and Host Range Analysis Against the ECOR Reference Library.

Host range analyses and genome sequencing/annotation of bacteriophage isolates allow more effective development of tools for applications in medicine, agriculture, and the environment and expand our understanding of phage biology. Here we present the complete sequence of phage Ac3's assembled and annotated genome (accession OK040907). Originally referred to simply as "3," Ac3 has previously been described as a T4-like bacteriophage belonging to the Myoviridae family in the Caudovirales order of tailed bacteriophages. Using a combination of spot tests and full plate plaque assays, Ac3's permissive and adsorptive host range were evaluated against the ECOR Reference Library; a panel of 72 Escherichia coli isolates meant to represent the diversity of E. coli. Spot assays revealed that Ac3 could adsorb to 43 of the 72 strains (59.7%), whereas plaque assays demonstrated Ac3's ability to complete replication within 27 of the 72 strains (37.5%). By overlaying spot test and plaque assay results, 16 of the 45 nonpermissive ECOR strains (35.5%) were highlighted as being able to support Ac3's adsorption and tail contraction, but not its replication. Further characterization of Ac3 is still needed, however, the study presented here provides a solid starting point for future research.

Nano-microbial based technology employing polyvalent phage conjugate: A next generation weapon for antimicrobial resistance lurking behind wastewater

Worldwide, due to a dearth of innovative interventions, new forms of antimicrobial resistance (AMR) are being discovered every day in clinical and environmental settings. Therefore, it is necessary to remove these contaminants directly or indirectly from the environment. Nanomicrobial-based technology employing nanomaterials with microbes is a new paradigm that finds a place in the antimicrobial crisis. Microbial entities such as phages can be used to treat antimicrobial resistance, but phage resistance is challenging and limits its applicability. Similarly, nanotechnology will not be able to selectively remove resistant strains from the environment individually. Therefore, we employ nanomicrobial-based technology that aims to fill these gaps. In the present study, polyvalent phages were isolated from wastewater with an easy-to-use modified multi-host sequential approach, characterized and conjugated with magnetite (Fe3O4) nanoparticles with the modified formulation to form nanomicrobial conjugates (NMCs). These NMCs were subjected to characterization and in vitro antibacterial studies. The results indicated a significant polyvalency of phages in the order of Caudovirales. Transmission electron microscopy (TEM) analysis of Fe3O4 nanoparticles formed by the co-precipitation method showed a particle size of 30 ± 5 nm and the selected area electron diffraction (SAED) pattern indicates a single-phase crystalline structure. To form NMCs, isolated phages (105 PFU/mL) were immobilized onto Fe3O4 nanoparticles. Further, surface modification of Fe3O4 nanoparticles enables the covalent association of phages. Biosurfactant-functionalized Fe3O4 nanoparticles (FNMCs) were found to have higher phage loading capacity, with a significant value of p < 0.0127 and a zeta potential of −22.2 mV. TEM studies and in vitro biofilm assay showed that NMCs exhibit promising antibacterial activity against various resistant bacterial strains. Pilot studies showed that NMCs can selectively eliminate up to 98.3% of AMR in wastewater. Thus, these findings indicate a synergistic effect of both phage and nanomaterial and this technology is expected to be a new lead in wastewater management.

Analysis of the anaerobic digestion metagenome under environmental stresses stimulating prophage induction

BackgroundThe viral community has the potential to influence the structure of the microbiome and thus the yield of the anaerobic digestion process. However, the virome composition in anaerobic digestion is still under-investigated. A viral induction experiment was conducted on separate batches undergoing a series of DNA-damaging stresses, in order to coerce temperate viruses to enter the lytic cycle.ResultsThe sequencing of the metagenome revealed a viral community almost entirely composed of tailed bacteriophages of the order Caudovirales. Following a binning procedure 1,092 viral and 120 prokaryotic genomes were reconstructed, 64 of which included an integrated prophage in their sequence.Clustering of coverage profiles revealed the presence of species, both viral and microbial, sharing similar reactions to shocks. A group of viral genomes, which increase under organic overload and decrease under basic pH, uniquely encode the yopX gene, which is involved in the induction of temperate prophages. Moreover, the in-silico functional analysis revealed an enrichment of sialidases in viral genomes. These genes are associated with tail proteins and, as such, are hypothesised to be involved in the interaction with the host. Archaea registered the most pronounced changes in relation to shocks and featured behaviours not shared with other species. Subsequently, data from 123 different samples of the global anaerobic digestion database was used to determine coverage profiles of host and viral genomes on a broader scale.ConclusionsViruses are key components in anaerobic digestion environments, shaping the microbial guilds which drive the methanogenesis process. In turn, environmental conditions are pivotal in shaping the viral community and the rate of induction of temperate viruses. This study provides an initial insight into the complexity of the anaerobic digestion virome and its relation with the microbial community and the diverse environmental parameters.5iPig9MR3ASALE62Tsjw9nVideo

Application of a novel lytic phage vB_EcoM_SQ17 for the biocontrol of Enterohemorrhagic Escherichia coli O157:H7 and Enterotoxigenic E. coli in food matrices.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 and Enterotoxigenic E. coli (ETEC) are important foodborne pathogens, causing serious food poisoning outbreaks worldwide. Bacteriophages, as novel antibacterial agents, have been increasingly exploited to control foodborne pathogens. In this study, a novel broad-host range lytic phage vB_EcoM_SQ17 (SQ17), was isolated, characterized, and evaluated for its potential to control bacterial counts in vitro and in three different food matrices (milk, raw beef, and fresh lettuce). Phage SQ17 was capable of infecting EHEC O157:H7, ETEC, and other E. coli strains. Morphology, one-step growth, and stability assay showed that phage SQ17 belongs to the Caudovirales order, Myoviridae family, and Mosigvirus genus. It has a short latent period of 10 min, a burst size of 71 PFU/infected cell, high stability between pH 4 to 12 as well as thermostability between 30°C and 60°C for 60 min. Genome sequencing analysis revealed that the genome of SQ17 does not contain any genes associated with antibiotic resistance, toxins, lysogeny, or virulence factors, indicating the potential safe application of phage SQ17 in the food industry. In Luria-Bertani (LB) medium, phage SQ17 significantly decreased the viable counts of EHEC O157:H7 by more than 2.40 log CFU/ml (p < 0.05) after 6 h of incubation at 37°C. Phage SQ17 showed great potential to be applied for biocontrol of EHEC O157:H7 in milk and raw beef. In fresh lettuce, treatment with SQ17 also resulted in significant reduction of viable cell counts of EHEC O157:H7 and ETEC at both 4°C and 25°C. Our results demonstrate that SQ17 is a good candidate for application as an EHEC O157:H7 and ETEC biocontrol agent in the processing stages of food production and food preservation.

Comprehensive deciphering prophages in genus Acetobacter on the ecology, genomic features, toxin-antitoxin system, and linkage with CRISPR-Cas system.

Acetobacter is the predominant microbe in vinegar production, particularly in those natural fermentations that are achieved by complex microbial communities. Co-evolution of prophages with Acetobacter, including integration, release, and dissemination, heavily affects the genome stability and production performance of industrial strains. However, little has been discussed yet about prophages in Acetobacter. Here, prophage prediction analysis using 148 available genomes from 34 Acetobacter species was carried out. In addition, the type II toxin–antitoxin systems (TAs) and CRISPR-Cas systems encoded by prophages or the chromosome were analyzed. Totally, 12,000 prophage fragments were found, of which 350 putatively active prophages were identified in 86.5% of the selected genomes. Most of the active prophages (83.4%) belonged to the order Caudovirales dominated by the families Siphoviridae and Myroviridae prophages (71.4%). Notably, Acetobacter strains survived in complex environments that frequently carried multiple prophages compared with that in restricted habits. Acetobacter prophages showed high genome diversity and horizontal gene transfer across different bacterial species by genomic feature characterization, average nucleotide identity (ANI), and gene structure visualization analyses. About 31.14% of prophages carry type II TAS, suggesting its important role in addiction, bacterial defense, and growth-associated bioprocesses to prophages and hosts. Intriguingly, the genes coding for Cse1, Cse2, Cse3, Cse4, and Cas5e involved in type I-E and Csy4 involved in type I-F CRISPR arrays were firstly found in two prophages. Type II-C CRISPR-Cas system existed only in Acetobacter aceti, while the other Acetobacter species harbored the intact or eroded type I CRISPR-Cas systems. Totally, the results of this study provide fundamental clues for future studies on the role of prophages in the cell physiology and environmental behavior of Acetobacter.

Characterization and anti-biofilm activity of bacteriophages against urinary tract Enterococcus faecalis isolates

Strong biofilm-forming Enterococcus feacalis urinary tract pathogens (n = 35) were used to determine the lytic spectrum of six bacteriophages isolated from sewage samples. Only 17 Enterococcus feacalis isolates gave lytic zones with the tested bacteriophages from which five isolates were susceptible to all of them. The isolated enterococcal phages are characterized by wide range of thermal (30–90 °C) and pH (3–10) stability. They belong to order Caudovirales, from which four bacteriophages (EPA, EPB, EPD, EPF) belong to family Myoviridae and two (EPC, EPE) belong to family Siphoviridae. In addition, they have promising antibiofilm activity against the tested strong-forming biofilm E. faecalis isolates. The enterococcal phages reduced the formed and preformed biofilms to a range of 38.02–45.7% and 71.0–80.0%, respectively, as compared to the control. The same promising activities were obtained on studying the anti-adherent effect of the tested bacteriophages on the adherence of bacterial cells to the surface of urinary catheter segments. They reduced the number of adherent cells to a range of 30.8–43.8% and eradicated the pre-adherent cells to a range of 48.2–71.1%, as compared to the control. Overall, the obtained promising antibiofilm activity makes these phages good candidates for application in preventing and treating biofilm associated Enterococcus faecalis infections.

Dynamics of nasopharyngeal tract phageome and association with disease severity and age of patients during three waves of COVID-19.

In December 2019, several patients were hospitalized and diagnosed with severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection, which subsequently led to a global pandemic. To date, there are no studies evaluating the relationship between the respiratory phageome and the SARS‐CoV‐2 infection. The current study investigated the phageome profiles in the nasopharyngeal swabs collected from 55 patients during the three different waves of coronavirus disease 2019 (COVID‐19) in the Campania Region (Southern Italy). Data obtained from the taxonomic profiling show that phage families belonging to the order Caudovirales have a high abundance in the patient samples. Moreover, the severity of the COVID‐19 infection seems to be correlated with the phage abundance.

Isolation and Characterization of Novel Lytic Bacteriophage vB_RsoP_BMB50 infecting Ralstonia solanacearum.

Ralstonia solanacearum is a soil-borne phytopathogen, and it can cause bacterial wilt disease in a variety of key crops around the world, thus resulting in enormous financial losses. However, there is a lack of effective, green, and safe prevention and control measures against increasingly devastating bacterial wilt disease. Bacteriophages (phages) are considered as potential biocontrol agents against bacterial wilt disease. Although many phages infecting R. solanacearum have been isolated, so far, these Ralstonia phages are still insufficient to deal with the diversity of the bacteria of R. solanacearum. In this study, a novel lytic bacteriophage vB_RsoP_BMB50 infecting multiple R. solanacearum was isolated from tomato fields in Dalian, China. Transmission electron microscopy and genomics analysis indicated that vB_RsoP_BMB50 belonged to the subfamily Okabevirinae, Autographiviridae family, and order Caudovirales, and it comprised a double-stranded DNA with a full length of 43,665bp and a mean G+C content of 61.79%, containing 53 open reading frames (ORFs). This novel phage exhibited a large burst size, high temperature stability (4-50 °C), and strong pH tolerance (pH 5-10). Comparative analyses and phylogenetic analyses revealed that vB_RsoP_BMB50 represented a novel Ralstonia phage genus since it exhibited a low sequence similarity to other phages in the GenBank database. Due to its broad lytic spectrum, high thermal stability, and strong pH tolerance, vB_RsoP_BMB50 is considered as an effective candidate biocontrol agent against bacterial wilt disease caused by R. solanacearum.

Effect of encapsulation and natural polyphenolic compounds on bacteriophage stability and activity on Escherichia coli in Lactuca sativa L. var. longifolia

AbstractThe goals of the present study were to characterize the phages isolated by magnetic nanocomplex, to determine the effect of natural polyphenols on phage activity against E. coli and the stability of free and encapsulated bacteriophages under UV irradiation, and their efficacy against E. coli on romaine lettuce (Lactuca sativa L. var. longifolia) as a fresh food model. The morphology of the two magnetically isolated phages were characterized using the transmission electron microscope (TEM). All isolated phages were classified in a morphological family of tailed phages of the order Caudovirales, Myoviridae family. The PCR results showed that the stx1 and cI genes were not on any phage. Assays were carried out to determine the effect of four polyphenolic natural extracts on phages under UV radiation. The highest activity and stability of phages expressed in phage titer (PFU/ml) were observed in the presence of polyphenols from tarbush leaves (Flourensia cernua DC). Coliphages were encapsulated in alginate/chitosan and alginate/PVA carriers with and without polyphenols from tarbush leaves. Encapsulation led to a higher phage stability under UV radiation. All encapsulated formulations and free coliphages were applied on the surfaces of romaine lettuce containing E. coli cells. An effective reduction of 2 log of E. coli titers from lettuce was observed. Encapsulated phage formulations with and without polyphenols from tarbush leaves could be considered for future application in fresh food preservation.

A Cocktail of Three Virulent Phages Controls Multidrug-Resistant Salmonella Enteritidis Infection in Poultry.

Salmonella enterica is not only the most common pathogen of poultry and poultry-derived products but is also a significant foodborne pathogen. In recent years, many S. enterica isolates have exhibited multi-drug resistance, which places huge pressure on global economy and health. Since phages are an attractive alternative to biocontrol pathogens, we isolated a total of 15 Salmonella phages from sewage effluent, sediment, and chicken manure. The GRNsp1, GRNsp3, GRNsp6, GRNsp21, GRNsp27, GRNsp30, GRNsp50, and GRNsp51 phages exhibited a wide host range against S. enterica serovars Enteritidis and Typhimurium in vitro. In particular, GRNsp51 exerted highly efficient lytic effects against a large proportion of S. Enteritidis and S. Typhimurium strains isolated from different regions of China. Meanwhile, GRNsp8 expanded the host range of GRNsp6 and GRNsp51. Based on their host ranges and lytic capacities, GRNsp6, GRNssp8, and GRNsp51 were selected for further investigation. Morphology, one-step growth curves, and stability assays revealed that GRNsp6, GRNsp8, and GRNsp51 all belong to the Caudovirales order and display relatively short latency periods with broad pH and thermal stability. Genomic analysis indicated that the genomes of these three phages contained no genes related to virulence, antibiotic resistance, or lysogeny. In addition, we tested the effectiveness of a cocktail composed of these three phages against S. Enteritidis in a chicken model. Treatment with the oral phage cocktail 24 h before or alongside Salmonella challenge significantly reduced colonization of the intestinal tract and decreased the mRNA expression of IL-6, IFN-γ, and IL-1β in the duodenum. Together, these findings indicate that a cocktail of the GRNsp6, GRNsp8, and GRNsp51 phages could serve as an effective antimicrobial therapeutic agent against multidrug-resistant Salmonella in animal production to mitigate infections by multiple zoonotic Salmonella species.

Isolation and Characterization of vB_kpnM_17-11, a Novel Phage Efficient Against Carbapenem-Resistant Klebsiella pneumoniae.

Phages and phage-encoded proteins exhibit promising prospects in the treatment of Carbapenem-Resistant Klebsiella pneumoniae (CRKP) infections. In this study, a novel Klebsiella pneumoniae phage vB_kpnM_17-11 was isolated and identified by using a CRKP host. vB_kpnM_17-11 has an icosahedral head and a retractable tail. The latent and exponential phases were 30 and 60 minutes, respectively; the burst size was 31.7 PFU/cell and the optimal MOI was 0.001. vB_kpnM_17-11 remained stable in a wide range of pH (4-8) and temperature (4-40°C). The genome of vB_kpnM_17-11 is 165,894 bp, double-stranded DNA (dsDNA), containing 275 Open Reading Frames (ORFs). It belongs to the family of Myoviridae, order Caudovirales, and has a close evolutionary relationship with Klebsiella phage PKO111. Sequence analysis showed that the 4530 bp orf022 of vB_kpnM_17-11 encodes a putative depolymerase. In vitro testing demonstrated that vB_kpnM_17-11 can decrease the number of K. pneumoniae by 105-fold. In a mouse model of infection, phage administration improved survival and reduced the number of K. pneumoniae in the abdominal cavity by 104-fold. In conclusion, vB_kpnM_17-11 showed excellent in vitro and in vivo performance against K. pneumoniae infection and constitutes a promising candidate for the development of phage therapy against CRKP.

Bacteriophages Isolated From Turkeys Infecting Diverse Salmonella Serovars.

Salmonella is one of the leading causes of foodborne illnesses worldwide. The rapid emergence of multidrug-resistant Salmonella strains has increased global concern for salmonellosis. Recent studies have shown that bacteriophages (phages) are novel and the most promising antibacterial agents for biocontrol in foods because phages specifically kill target bacteria without affecting other bacteria, do not alter organoleptic properties or nutritional quality of foods, and are safe and environmentally friendly. Due to the vast variation in Salmonella serotypes, large numbers of different and highly virulent Salmonella phages with broad host ranges are needed. This study isolated 14 Salmonella phages from turkey fecal and cecal samples. Six phages (Φ205, Φ206, Φ207, ΦEnt, ΦMont, and Φ13314) were selected for characterization. These phages were from all three families in the Caudovirales order. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed that each phage had a unique structural protein profile. Each phage had a distinct host range. Φ207 and ΦEnt are both siphophages. They shared eight hosts, including seven different Salmonella serovars and one Shigella sonnei strain. These two phages showed different restriction banding patterns generated through EcoRI or HindIII digestion, but shared three bands from EcoRI digestion. ΦEnt displayed the broadest and very unusual host range infecting 11 Salmonella strains from nine serovars and three Shigella strains from two species, and thus was further characterized. The one-step growth curve revealed that ΦEnt had a short latent period (10 min) and relatively large burst size (100 PFU/infected cell). ΦEnt and its host showed better thermal stabilities in tryptic soy broth than in saline at 63 or 72°C. In the model food system (cucumber juice or beef broth), ΦEnt infection [regardless of the multiplicity of infections (MOIs) of 1, 10, and 100] resulted in more than 5-log10 reduction in Salmonella concentration within 4 or 5 h. Such high lytic activity combined with its remarkably broad and unusual host range and good thermal stability suggested that ΦEnt is a novel Salmonella phage with great potential to be used as an effective biocontrol agent against diverse Salmonella serovars in foods.

Features of the gut prokaryotic virome of Japanese patients with Crohn's disease.

The gut virome is mainly composed of bacteriophages and influences gut homeostasis and pathogenic conditions. In this study, we analyzed the gut prokaryotic virome in Japanese patients with Crohn's disease (CD). We collected 19 fecal samples from CD patients and 16 samples from healthy controls. The gut bacteriome was analyzed by 16S rRNA gene sequencing and the virome was profiled by shotgun metagenomic sequencing. Despite no differences in richness and evenness, there was a significant difference in the overall structure of the gut virome between CD patients and controls (P = 0.013). CrAssphage and Staphylococcus virus, belonging to the order Caudovirales, were dominant in the gut virome of controls and CD patients. The abundance of crAssphage was significantly greater in CD patients than controls (P = 0.021). Lactococcus, Enterococcus and Lactobacillus phages were present only in CD patients, while Xanthomonas and Escherichia phages were unique to the controls. In the gut bacteriome of CD patients, richness and evenness were significantly lower, and a significant difference in the overall structure was observed between groups (P = 0.014). The gut bacteriome of CD patients was characterized by a decrease of the genera Faecalibacterium, Roseburia, and Ruminococcus and an increase of the family Enterobacteriaceae. There were more significant correlations between viruses and bacteria in CD patients than controls. The gut virome of CD patients was distinct from that of healthy controls in a Japanese population. An altered gut virome may be one of the factors associated with the bacterial dysbiosis of CD.

Metagenomic analysis of wastewater phageome from a University Hospital in Turkey.

Phage DNA analysis gives opportunity to understand living ecosystem of the environment where the samples are taken. In the present study, we analyzed phage DNA obtained from wastewater sample of university hospital sewage. After filtration, long high-speed centrifugation was done to collect phages. DNA was extracted from pellet by phenol chloroform extraction and used for NGS sequencing. The host profile, taxonomic and functional analyses were performed using MG-RAST, and ResFinder program was used for resistance gene detection. High amounts of reads belong to bacteriophage groups (~ 95%) from our DNA sample were obtained and all bacteriophage reads were found belonging to Caudovirales order and Myoviridae (56%), Siphoviridae (43%), and Podoviridae (0.02%) families. The most common host genera were Escherichia (88.20%), Salmonella (5.49%) and Staphylococcus (5.19%). SEED subsystems hits were mostly structural parts and KEGG Orthology hits were nucleotide- and carbohydrate metabolism-related genes. No anti-microbial resistance genes were detected. Our bacteriophage DNA purification method is favorable for phage metagenomic studies. Dominance of coliphages may explain infrequent Podoviridae. Dominancy of structural genes and auxiliary genes is probably due to abundance of lytic phages in our sample. Absence of antibiotic resistance genes even in hospital environment phages indicates that phages are not important carrier of resistance genes.

An Anti-MRSA Phage From Raw Fish Rinse: Stability Evaluation and Production Optimization.

Accumulating evidence has denoted the danger of resistance in tenacious organisms like methicillin-resistant Staphylococcus aureus (MRSA). MRSA, a supple bacterium that adopts a variety of antibiotic resistance mechanisms, is the cause of multiple life-threatening conditions. Approaching a post-antibiotic era, bacteria-specific natural predators, bacteriophages, are now given the chance to prove eligible for joining the antibacterial weaponry. Considering the foregoing, this study aimed at isolating bacteriophages with promising anti-MRSA lytic activity, followed by characterization and optimization of the production of the bacteriophage with the broadest host range. Five phages were isolated from different environmental sources including the rinse of raw chicken egg, raw milk, and, remarkably, the raw meat rinses of chicken and fish. Examined for lytic activity against a set of 23 MRSA isolates collected from various clinical specimens, all five phages showed relatively broad host ranges with the bacteriophage originally isolated from raw fish rinse showing lytic activity against all the isolates tested. This phage is suggested to be a member of Siphoviridae family, order Caudovirales, as revealed by electron microscopy. It also exhibited good thermal stability and viability at different pH grades. Moreover, it showed reasonable stability against UV light and all viricidal organic solvents tested. Optimization using D-optimal design by response surface methodology was carried out to enhance the phage yield. The optimum conditions suggested by the generated model were a pH value of 7, a carbon source of 0.5% w/v sucrose, and a nitrogen source of 0.1% w/v peptone, at a temperature of 28°C and a bacterial inoculum size of 107 CFU/ml, resulting in a 2 log-fold increase in the produced bacteriophage titer. Overall, the above findings indicate the lytic ability inflicted by this virus on MRSA. Apparently, its stability under some of the extreme conditions tested implies its potential to be a candidate for pharmaceutical formulation as an anti-MRSA therapeutic tool. We hope that bacteriophages could tip the balance in favor of the human front in their battle against multidrug-resistant pathogens.

PHAGES OF BACTERIА OF THE GENUS BACILLUS ISOLATED FROM THE AQUATIC ENVIRONMENT

SummaryDespite the fact that bacteria of the genus Bacillus isolated from the soil have been studied for centuries, there are still topics that are not sufficiently covered or need further research. Most often, members of the genus Bacillus are isolated from soil or food. In recent years, these bacteria have begun to isolate from various aquatic biocenoses of ecosystems of oceans, seas, estuaries, lakes, rivers. Studies of such isolates indicate that bacteria of almost all species of the genus Bacillus are infected with bacteriophages of the order Caudovirales, which have caudal processes, integrase and excision systems necessary for the lysogenic development cycle. Although most of the bacteriophages found belong to the order Caudovirales, they have a wide range of differences, such as the relationship to temperature and pH, the impact on metabolism and sporulation of the host. The review presents data from the modern literature on bacteriophages that infect bacteria of the genus Bacillus isolated from aquatic biocenoses, features of their structure, chemical composition, genome structure and interaction with the host cell.