A novel flavobacterial phage abundant during green tide, representing a new viral family, Zblingviridae.

Flavobacteriia are the dominant and active bacteria during algal blooms and play an important role in polysaccharide degradation. However, little is known about phages infecting Flavobacteriia, especially during green tide. In this study, a novel virus, vB_TgeS_JQ, infecting Flavobacteriia was isolated from the surface water of the Golden Beach of Qingdao, China. Transmission electron microscopy demonstrated that vB_TgeS_JQ had the morphology of siphovirus. The experiments showed that it was stable from -20°C to 45°C and pH 5 to pH 8, with latent and burst periods both lasting for 20 min. Genomic analysis showed that the phage vB_TgeS_JQ contained a 40,712-bp dsDNA genome with a GC content of 30.70%, encoding 74 open-reading frames. Four putative auxiliary metabolic genes were identified, encoding electron transfer-flavoprotein dehydrogenase, calcineurin-like phosphoesterase, phosphoribosyl-ATP pyrophosphohydrolase, and TOPRIM nucleotidyl hydrolase. The abundance of phage vB_TgeS_JQ was higher during Ulva prolifera (U. prolifera) blooms compared with other marine environments. The phylogenetic and comparative genomic analyses revealed that vB_TgeS_JQ exhibited significant differences from all other phage isolates in the databases and therefore was classified as an undiscovered viral family, named Zblingviridae. In summary, this study expands the knowledge about the genomic, phylogenetic diversity and distribution of flavobacterial phages (flavophages), especially their roles during U. prolifera blooms. IMPORTANCE: The phage vB_TgeS_JQ was the first flavobacterial phage isolated during green tide, representing a new family in Caudoviricetes and named Zblingviridae. The abundance of phage vB_TgeS_JQ was higher during the Ulva prolifera blooms. This study provides insights into the genomic, phylogenetic diversity, and distribution of flavophages, especially their roles during U. prolifera blooms.

Isolation, characterization, and potential application of Acinetobacter baumannii phages against extensively drug-resistant strains.

One of the significant issues in treating bacterial infections is the increasing prevalence of extensively drug-resistant (XDR) strains of Acinetobacter baumannii. In the face of limited or no viable treatment options for extensively drug-resistant (XDR) bacteria, there is a renewed interest in utilizing bacteriophages as a treatment option. Three Acinetobacter phages (vB_AbaS_Ftm, vB_AbaS_Eva, and vB_AbaS_Gln) were identified from hospital sewage and analyzed for their morphology, host ranges, and their genome sequences were determined and annotated. These phages and vB_AbaS_SA1 were combined to form a phage cocktail. The antibacterial effects of this cocktail and its combinations with selected antimicrobial agents were evaluated against the XDR A. baumannii strains. The phages exhibited siphovirus morphology. Out of a total of 30 XDR A. baumannii isolates, 33% were sensitive to vB_AbaS_Ftm, 30% to vB_AbaS_Gln, and 16.66% to vB_AbaS_Eva. When these phages were combined with antibiotics, they demonstrated a synergistic effect. The genome sizes of vB_AbaS_Ftm, vB_AbaS_Eva, and vB_AbaS_Gln were 48487, 50174, and 50043 base pairs (bp), respectively, and showed high similarity. Phage cocktail, when combined with antibiotics, showed synergistic effects on extensively drug-resistant (XDR) strains of A. baumannii. However, the need for further study to fully understand the mechanisms of action and potential limitations of using these phages is highlighted.

Characterization and genome analysis of a broad host range lytic phage vB_SenS_TUMS_E19 against Salmonella enterica and its efficiency evaluation in the liquid egg.

Salmonella enterica serovars are zoonotic bacterial that cause foodborne enteritis. Due to bacteria's antibiotic resistance, using bacteriophages for biocontrol and treatment is a new therapeutic approach. In this study, we isolated, characterized, and analyzed the genome of vB_SenS_TUMS_E19 (E19), a broad host range Salmonella bacteriophage, and evaluated the influence of E19 on liquid eggs infected with Salmonella enterica serovar Enteritidis. Transmission electron microscopy showed that the isolated bacteriophage had a siphovirus morphotype. E19 showed rapid adsorption (92% in 5 min), a short latent period (18 min), a large burst size (156 PFU per cell), and a broad host range against different Salmonella enterica serovars. Whole-genome sequencing analysis indicated that the isolated phage had a 42 813 bp long genome with 49.8% G + C content. Neither tRNA genes nor those associated with antibiotic resistance, virulence factors, or lysogenic formation were detected in the genome. The efficacy of E19 was evaluated in liquid eggs inoculated with S. Enteritidis at 4 and 25 °C, and results showed that it could effectively eradicate S. Enteritidis in just 30 min and prevented its growth up to 72 h. Our findings indicate that E19 can be an alternative to a preservative to control Salmonella in food samples and help prevent and treat salmonellosis.

Morphological, biological, and genomic characterization of Klebsiella pneumoniae phage vB_Kpn_ZC2.

BACKGROUND: Bacteriophages (phages) are one of the most promising alternatives to traditional antibiotic therapies, especially against multidrug-resistant bacteria. Klebsiella pneumoniae is considered to be an opportunistic pathogen that can cause life-threatening infections. Thus, this study aims at the characterization of a novel isolated phage vB_Kpn_ZC2 (ZCKP2, for short). METHODS: The phage ZCKP2 was isolated from sewage water by using the clinical isolate KP/08 as a host strain. The isolated bacteriophage was purified and amplified, followed by testing of its molecular weight using Pulse-Field Gel Electrophoresis (PFGE), transmission electron microscopy, antibacterial activity against a panel of other Klebsiella pneumoniae hosts, stability studies, and whole genome sequencing. RESULTS: Phage ZCKP2 belongs morphologically to siphoviruses as indicated from the Transmission Electron Microscopy microgram. The Pulsed Field Gel Electrophoresis and the phage sequencing estimated the phage genome size of 48.2 kbp. Moreover, the absence of lysogeny-related genes, antibiotic resistance genes, and virulence genes in the annotated genome suggests that phage ZCKP2 is safe for therapeutic use. Genome-based taxonomic analysis indicates that phage ZCKP2 represents a new family that has not been formally rated yet. In addition, phage ZCKP2 preserved high stability at different temperatures and pH values (-20 - 70 °C and pH 4 - 9). For the antibacterial activity, phage ZCKP2 maintained consistent clear zones on KP/08 bacteria along with other hosts, in addition to effective bacterial killing over time at different MOIs (0.1, 1, and 10). Also, the genome annotation predicted antibacterial lytic enzymes. Furthermore, the topology of class II holins was predicted in some putative proteins with dual transmembrane domains that contribute significantly to antibacterial activity. Phage ZCKP2 characterization demonstrates safety and efficiency against multidrug-resistant K. pneumoniae, hence ZCKP2 is a good candidate for further in vivo and phage therapy clinical applications.

Jojan: a novel virus that lyses Stenotrophomonas maltophilia from dog.

Stenotrophomonas maltophilia is a Gram-negative bacterium widely distributed in the environment and associated with nosocomial infections, pneumonia, and bacteremia in humans and other mammals. We have isolated and sequenced a new virus that lyses the S. maltophilia strain from a dog skin. The virus has a siphovirus-like morphology and a linear dsDNA genome 60,804 pb in length with terminal repeats, four tRNA genes, and 111 putative proteins. The annotated genes resemble the corresponding genes of some siphoviruses, but the unique genome arrangement and limited similarity of the encoded proteins suggest that this virus does not belong to any known genus. The virus uses zinc metallopeptidase for lysis of its host. This enzyme is active in the presence of Zn2+ or Mg2+ ions and maintains its bactericidal activity up to 50 °C. Both the virus itself and the endolysin specifically degrade only the host bacterial strain.

Lytic bacteriophage vB_KmiS-Kmi2C disrupts biofilms formed by members of the Klebsiella oxytoca complex, and represents a novel virus family and genus.

AIMS: This study aimed to characterize the lytic phage vB_KmiS-Kmi2C, isolated from sewage water on a GES-positive strain of Klebsiella michiganensis. METHODS AND RESULTS: Comparative phylogenetic and network-based analyses were used to characterize the genome of phage vB_KmiS-Kmi2C (circular genome of 42 234 bp predicted to encode 55 genes), demonstrating it shared little similarity with other known phages. The phage was lytic on clinical strains of K. oxytoca (n = 2) and K. michiganensis (n = 4), and was found to both prevent biofilm formation and disrupt established biofilms produced by these strains. CONCLUSIONS: We have identified a phage capable of killing clinically relevant members of the K. oxytoca complex (KoC). The phage represents a novel virus family (proposed name Dilsviridae) and genus (proposed name Dilsvirus).

In planta interactions of a novel bacteriophage against Pseudomonas syringae pv. tomato.

The biology and biotechnology of bacteriophages have been extensively studied in recent years to explore new and environmentally friendly methods of controlling phytopathogenic bacteria. Pseudomonas syringae pv. tomato (Pst) is responsible for bacterial speck disease in tomato plants, leading to decreased yield. Disease management strategies rely on the use of copper-based pesticides. The biological control of Pst with the use of bacteriophages could be an alternative environmentally friendly approach to diminish the detrimental effects of Pst in tomato cultivations. The lytic efficacy of bacteriophages can be used in biocontrol-based disease management strategies. Here, we report the isolation and complete characterization of a bacteriophage, named Medea1, which was also tested in planta against Pst, under greenhouse conditions. The application of Medea1 as a root drenching inoculum or foliar spraying reduced 2.5- and fourfold on average, respectively, Pst symptoms in tomato plants, compared to a control group. In addition, it was observed that defense-related genes PR1b and Pin2 were upregulated in the phage-treated plants. Our research explores a new genus of Pseudomonas phages and explores its biocontrol potential against Pst, by utilizing its lytic nature and ability to trigger the immune response of plants. KEY POINTS: • Medea1 is a newly reported bacteriophage against Pseudomonas syringae pv. tomato having genomic similarities with the phiPSA1 bacteriophage • Two application strategies were reported, one by root drenching the plants with a phage-based solution and one by foliar spraying, showing up to 60- and 6-fold reduction of Pst population and disease severity in some cases, respectively, compared to control • Bacteriophage Medea1 induced the expression of the plant defense-related genes Pin2 and PR1b.

Characterization of a Vibriophage Infecting Pathogenic Vibrio harveyi.

Bacterial diseases caused by Vibrio spp. are prevalent in aquaculture and can lead to high mortality rates among aquatic species and significant economic losses. With the increasing emergence of multidrug-resistant Vibrio strains, phage therapy is being explored as a potential alternative to antibiotics for biocontrol of infectious diseases. Here, a new lytic phage named vB_VhaS_R21Y (R21Y) was isolated against Vibrio harveyi BVH1 obtained from seawater from a scallop-farming area in Rongcheng, China. Its morphology, infection cycle, lytic profile, phage stability, and genetic features were characterized. Transmission electronic microscopy indicated that R21Y is siphovirus-like, comprising an icosahedral head (diameter 73.31 ± 2.09 nm) and long noncontractile tail (205.55 ± 0.75 nm). In a one-step growth experiment, R21Y had a 40-min latent period and a burst size of 35 phage particles per infected cell. R21Y was highly species-specific in the host range test and was relatively stable at pH 4-10 and 4-55 °C. Genomic analysis showed that R21Y is a double-stranded DNA virus with a genome size of 82,795 bp and GC content of 47.48%. Its high tolerance and lytic activity indicated that R21Y may be a candidate for phage therapy in controlling vibriosis in aquacultural systems.

Characterization of Parageobacillus Bacteriophage vB_PtoS_NIIg3.2-A Representative of a New Genus within Thermophilic Siphoviruses.

A high temperature-adapted bacteriophage, vB_PtoS_NIIg3.2 (NIIg3.2), was isolated in Lithuania from compost heaps using Parageobacillus toebii strain NIIg-3 as a host for phage propagation. Furthermore, NIIg3.2 was active against four strains of Geobacillus thermodenitrificans, and it infected the host cells from 50 to 80 °C. Transmission electron microscopy analysis revealed siphovirus morphology characterized by an isometric head (~59 nm in diameter) and a noncontractile tail (~226 nm in length). The double-stranded DNA genome of NIIg3.2 (38,970 bp) contained 71 probable protein-encoding genes and no genes for tRNA. In total, 29 NIIg3.2 ORFs were given a putative functional annotation, including those coding for the proteins responsible for DNA packaging, virion structure/morphogenesis, phage-host interactions, lysis/lysogeny, replication/regulation, and nucleotide metabolism. Based on comparative phylogenetic and bioinformatic analysis, NIIg3.2 cannot be assigned to any genus currently recognized by ICTV and potentially represents a new one within siphoviruses. The results of this study not only extend our knowledge about poorly explored thermophilic bacteriophages but also provide new insights for further investigation and understanding the evolution of Bacilllus-group bacteria-infecting viruses.

A Genomic Analysis of the Bacillus Bacteriophage Kirovirus kirovense Kirov and Its Ability to Preserve Milk.

Bacteriophages are widely recognized as alternatives to traditional antibiotics commonly used in the treatment of bacterial infection diseases and in the food industry, as phages offer a potential solution in combating multidrug-resistant bacterial pathogens. In this study, we describe a novel bacteriophage, Kirovirus kirovense Kirov, which infects members of the Bacillus cereus group. Kirovirus kirovense Kirov is a broad-host-range phage belonging to the Caudoviricetes class. Its chromosome is a linear 165,667 bp double-stranded DNA molecule that contains two short, direct terminal repeats, each 284 bp long. According to bioinformatics predictions, the genomic DNA contains 275 protein-coding genes and 5 tRNA genes. A comparative genomic analysis suggests that Kirovirus kirovense Kirov is a novel species within the Kirovirus genus, belonging to the Andregratiavirinae subfamily. Kirovirus kirovense Kirov demonstrates the ability to preserve and decontaminate B. cereus from cow milk when present in milk at a concentration of 104 PFU/mL. After 4 h of incubation with the phage, the bacterial titer drops from 105 to less than 102 CFU/mL.

Characterization and Genomic Analysis of ɸSHP3, a New Transposable Bacteriophage Infecting Stenotrophomonas maltophilia.

This study describes a novel transposable bacteriophage, ɸSHP3, continuously released by Stenotrophomonas maltophilia strain c31. Morphological observation and genomic analysis revealed that ɸSHP3 is a siphovirus with a 37,611-bp genome that encodes 51 putative proteins. Genomic comparisons indicated that ɸSHP3 is a B3-like transposable phage. Its genome configuration is similar to that of Pseudomonas phage B3, except for the DNA modification module. Similar to B3-like phages, the putative transposase B of ɸSHP3 is a homolog of the type two secretion component ExeA, which is proposed to serve as a potential virulence factor. Moreover, most proteins of ɸSHP3 have homologs in transposable phages, but only ɸSHP3 carries an RdgC-like protein encoded by gene 3, which exhibits exonuclease activity in vitro Two genes and their promoters coding for ɸSHP3 regulatory proteins were identified and appear to control the lytic-lysogenic switch. One of the proteins represses one promoter activity and confers immunity to ɸSHP3 superinfection in vivo The short regulatory region, in addition to the canonical bacterial promoter sequences, displays one LexA and two CpxR recognition sequences. This suggests that LexA and the CpxR/CpxA two-component system might be involved in the control of the ɸSHP3 genetic switch.IMPORTANCES. maltophilia is an emerging global pathogenic bacterium that displays genetic diversity in both environmental and clinical strains. Transposable phages have long been known to improve the genetic diversity of bacterial strains by transposition. More than a dozen phages of S. maltophilia have been characterized. However, no transposable phage infecting S. maltophilia has been reported to date. Characterization of the first transposable phage, ɸSHP3, from S. maltophilia will contribute to our understanding of host-phage interactions and genetic diversity, especially the interchange of genetic materials among S. maltophilia.

Pantoea Bacteriophage vB_PagS_MED16-A Siphovirus Containing a 2'-Deoxy-7-amido-7-deazaguanosine-Modified DNA.

A novel siphovirus, vB_PagS_MED16 (MED16) was isolated in Lithuania using Pantoea agglomerans strain BSL for the phage propagation. The double-stranded DNA genome of MED16 (46,103 bp) contains 73 predicted open reading frames (ORFs) encoding proteins, but no tRNA. Our comparative sequence analysis revealed that 26 of these ORFs code for unique proteins that have no reliable identity when compared to database entries. Based on phylogenetic analysis, MED16 represents a new genus with siphovirus morphology. In total, 35 MED16 ORFs were given a putative functional annotation, including those coding for the proteins responsible for virion morphogenesis, phage-host interactions, and DNA metabolism. In addition, a gene encoding a preQ0 DNA deoxyribosyltransferase (DpdA) is present in the genome of MED16 and the LC-MS/MS analysis indicates 2'-deoxy-7-amido-7-deazaguanosine (dADG)-modified phage DNA, which, to our knowledge, has never been experimentally validated in genomes of Pantoea phages. Thus, the data presented in this study provide new information on Pantoea-infecting viruses and offer novel insights into the diversity of DNA modifications in bacteriophages.

A novel phage from periodontal pockets associated with chronic periodontitis.

Bacteriophages often constitute the majority of periodontal viral communities, but phages that infect oral bacteria remain uncharacterized. Here, we present the genetic analysis of the genome of a novel siphovirus, named Siphoviridae_29632, which was isolated from a patient with periodontitis using a viral metagenomics-based approach. Among 43 predicted open reading frames (ORFs) in the genome, the viral genes encoding structural proteins were distinct from the counterparts of other viruses, although a distant homology is shared among viral morphogenesis proteins. A total of 28 predicted coding sequences had significant homology to other known phage ORF sequences. In addition, the prevalence of Siphoviridae_29632 in a cohort of patients with chronic periodontitis was 41.67%, which was significantly higher than that in the healthy group (4.55%, P < 0.001), suggesting that this virus as well as its hosts may contribute to the ecological environment favored for chronic periodontitis.

Isolation and characterization of the first phage infecting ecologically important marine bacteria Erythrobacter.

BACKGROUND: Erythrobacter comprises a widespread and ecologically significant genus of marine bacteria. However, no phage infecting Erythrobacter spp. has been reported to date. This study describes the isolation and characterization of phage vB_EliS-R6L from Erythrobacter. METHODS: Standard virus enrichment and double-layer agar methods were used to isolate and characterize the phage. Morphology was observed by transmission electron microscopy, and a one-step growth curve assay was performed. The phage genome was sequenced using the Illumina Miseq platform and annotated using standard bioinformatics tools. Phylogenetic analyses were performed based on the deduced amino acid sequences of terminase, endolysin, portal protein, and major capsid protein, and genome recruitment analysis was conducted using Jiulong River Estuary Virome, Pacific Ocean Virome and Global Ocean Survey databases. RESULTS: A novel phage, vB_EliS-R6L, from coastal waters of Xiamen, China, was isolated and found to infect the marine bacterium Erythrobacter litoralis DSM 8509. Morphological observation and genome analysis revealed that phage vB_EliS-R6L is a siphovirus with a 65.7-kb genome that encodes 108 putative gene products. The phage exhibits growth at a wide range of temperature and pH conditions. Genes encoding five methylase-related proteins were found in the genome, and recognition site predictions suggested its resistance to restriction-modification host systems. Genomic comparisons and phylogenetic analyses indicate that phage vB_EliS-R6L is distinct from other known phages. Metagenomic recruitment analysis revealed that vB_EliS-R6L-like phages are widespread in marine environments, with likely distribution in coastal waters. CONCLUSIONS: Isolation of the first Erythrobacter phage (vB_EliS-R6L) will contribute to our understanding of host-phage interactions, the ecology of marine Erythrobacter and viral metagenome annotation efforts.

A novel Halomonas ventosae-specific virulent halovirus isolated from the Qiaohou salt mine in Yunnan, Southwest China.

Although Halomonas phages belonging to the families Myoviridae and Siphoviridae have been reported, no virulent Halomonas siphoviruses are known. In this study, a virulent bacteriophage, QHHSV-1, of the family Siphoviridae that specifically infects H. ventosae QH52-2 was isolated from the Qiaohou salt mine. Restriction analysis indicated that QHHSV-1 is a dsDNA virus with a genome size of 33.5-39.5 kb. Transmission electron microscopy showed that QHHSV-1 is a typical representative of the Siphoviridae, with an icosahedral head (47 nm in diameter) and a non-contractile tail (75 nm in length). We also assessed the adsorption rate of QHHSV-1 for the host bacterium and found significant inhibition after the addition of 10 mM CaCl2. Based on a one-step growth curve, we determined a latent period of 30 min and a burst size of 73 PFU/infected cell. At the optimal pH of 8.0, 25.9 and 15.2 % of the phages survived after a 60-min incubation at 50 and 60 °C, respectively. Phage replication was possible at a wide range of salt concentrations, from 2.0 to 20 % (w/v), with an optimum concentration of 5 %. The survival of QHHSV-1 at different salt concentrations decreased with time and 25 % survival after 25 days at 30 % salt concentration.

Genome sequence of Soos: a siphovirus of the CP cluster infecting Gordonia rubripertincta  .

Novel actinobacteriophage Soos was isolated and purified from Southern Indiana soil using host Gordonia rubripertincta NRRL B-16540. Sequencing revealed a 57,509 bp circularly permuted genome encoding 87 predicted protein-coding genes. Soos is only the third phage in cluster CP, along with phages Clawz and Sting.

Complete genome and characteristics of cluster BC bacteriophage SoJo, isolated using Streptomyces mirabilis NRRL B-2400 in Columbia, MD.

Here, we present bacteriophage SoJo, a siphovirus infecting Streptomyces mirabilis, with a circularly permuted genome of 39 kbp and GC content of 71.5%. Its genome length and content are similar to that of other phages in the Actinobacteriophage Database BC cluster. SoJo was isolated from soil in Columbia, MD, USA.

A Genome of Temperate Enterococcus Bacteriophage Placed in a Space of Pooled Viral Dark Matter Sequences.

In the human gut, temperate bacteriophages interact with bacteria through predation and horizontal gene transfer. Relying on taxonomic data, metagenomic studies have associated shifts in phage abundance with a number of human diseases. The temperate bacteriophage VEsP-1 with siphovirus morphology was isolated from a sample of river water using Enterococcus faecalis as a host. Starting from the whole genome sequence of VEsP-1, we retrieved related phage genomes in blastp searches of the tail protein and large terminase sequences, and blastn searches of the whole genome sequences, with matches compiled from several different databases, and visualized a part of viral dark matter sequence space. The genome network and phylogenomic analyses resulted in the proposal of a novel genus "Vespunovirus", consisting of temperate, mainly metagenomic phages infecting Enterococcus spp.

Isolation, characterization, and genome analysis of a broad host range Salmonella phage vB_SenS_TUMS_E4: a candidate bacteriophage for biocontrol.

Salmonella enteritidis is one of the most important foodborne pathogens that cause numerous outbreaks worldwide. Some strains of Salmonella have become progressively resistant to antibiotics, so they could represent a critical threat to public health and have led to the use of alternative therapeutic approaches like phage therapy. In this study, a lytic phage, vB_SenS_TUMS_E4 (E4), was isolated from poultry effluent and characterized to evaluate its potential and efficacy for bio-controlling S. enteritidis in foods. Transmission electron microscopy revealed that E4 has a siphovirus morphotype, with an isometric head and non-contractile tail. Determining the host range showed that this phage can effectively infect different motile as well as non-motile Salmonella enterica serovars. The biological characteristics of E4 showed that it has a short latent period of about 15 min and a large burst size of 287 PFU/cell, and is also significantly stable in a broad range of pHs and temperatures. The E4 whole genome contains 43,018 bp and encodes 60 coding sequences (CDSs) but no tRNA genes. Bioinformatics analysis revealed that the genome of E4 lacks any genes related to lysogeny behavior, antibiotic resistance, toxins, or virulence factors. The efficacy of phage E4 as a bio-control agent was assessed in various foodstuffs inoculated with S. enteritidis at 4°C and 25°C, and the resulting data indicated that it could eradicate S. enteritidis after a very short time of 15 min. The findings of the present study showed that E4 is a hopeful candidate as a bio-control agent against S. enteritidis and has the potential to be used in various foodstuffs.

Characterization and genomic analysis of phage vB_ValR_NF, representing a new viral family prevalent in the Ulva prolifera blooms.

Introduction: Vibrio is an important bacterial genus containing many pathogenic species. Although more and more Vibrio phages were isolated, the genome, ecology and evolution of Vibrio phages and their roles in bacteriophage therapy, have not been fully revealed. Methods: Novel Vibrio phage vB_ValR_NF infecting Vibrio alginolyticus was isolated from the coastal waters of Qingdao during the Ulva prolifera blooms, Characterization and genomic feature of phage vB_ValR_NF has been analysed using phage isolation, sequencing and metagenome method. Results and Discussion: Phage vB_ValR_NF has a siphoviral morphology (icosahedral head 114±1 nm in diameter; a tail length of 231±1 nm), a short latent period (30 minutes) and a large burst size (113 virions per cell), and the thermal/pH stability study showed that phage vB_ValR_NF was highly tolerant to a range of pHs (4-12) and temperatures (-20 - 45 °C), respectively. Host range analysis suggests that phage vB_ValR_NF not only has a high inhibitory ability against the host strain V. alginolyticus, but also can infect 7 other Vibrio strains. In addition, the phage vB_ValR_NF has a double-stranded 44, 507 bp DNA genome, with 43.10 % GC content and 75 open reading frames. Three auxiliary metabolic genes associated with aldehyde dehydrogenase, serine/threonine protein phosphatase and calcineurin-like phosphoesterase were predicted, might help the host V. alginolyticus occupy the survival advantage, thus improving the survival chance of phage vB_ValR_NF under harsh conditions. This point can be supported by the higher abundance of phage vB_ValR_NF during the U. prolifera blooms than in other marine environments. Further phylogenetic and genomic analysis shows that the viral group represented by Vibrio phage vB_ValR_NF is different from other well-defined reference viruses, and can be classified into a new family, named Ruirongviridae. In general, as a new marine phage infecting V. alginolyticus, phage vB_ValR_NF provides basic information for further molecular research on phage-host interactions and evolution, and may unravel a novel insight into changes in the community structure of organisms during the U. prolifera blooms. At the same time, its high tolerance to extreme conditions and excellent bactericidal ability will become important reference factors when evaluating the potential of phage vB_ValR_NF in bacteriophage therapy in the future.