Characterization of Six Leuconostoc fallax Bacteriophages Isolated from an Industrial Sauerkraut Fermentation

Six bacteriophages active against Leuconostoc fallax strains were isolated from industrial sauerkraut fermentation brines. These phages were characterized as to host range, morphology, structural proteins, and genome fingerprint. They were exclusively lytic against the species L. fallax and had different host ranges among the strains of this species tested. Morphologically, three of the phages were assigned to the family Siphoviridae, and the three others were assigned to the family Myoviridae. Major capsid proteins detected by electrophoresis were distinct for each of the two morphotypes. Restriction fragment length polymorphism analysis and randomly amplified polymorphic DNA fingerprinting showed that all six phages were genetically distinct. These results revealed for the first time the existence of bacteriophages that are active against L. fallax and confirmed the presence and diversity of bacteriophages in a sauerkraut fermentation. Since a variety of L. fallax strains have been shown to be present in sauerkraut fermentation, bacteriophages active against L. fallax are likely to contribute to the microbial ecology of sauerkraut fermentation and could be responsible for some of the variability observed in this type of fermentation.     

Isolation and Characterization of Campylobacter Bacteriophages from Retail Poultry

The ability of phages to survive processing is an important aspect of their potential use in the biocontrol of Campylobacter in poultry production. To this end, we have developed a procedure to recover Campylobacter bacteriophages from chilled and frozen retail poultry and have validated the sensitivity of the method by using a characterized Campylobacter phage (i.e., NCTC 12674). By using this method, we have shown that Campylobacter phages can survive on retail chicken under commercial storage conditions. Retail chicken portions purchased in the United Kingdom were screened for the presence of endogenous Campylobacter phages. Thirty-four Campylobacter bacteriophages were isolated from 300 chilled retail chicken portions, but none could be recovered from 150 frozen chicken portions. The phage isolates were characterized according to their lytic profiles, morphology, and genome size. The free-range products were significantly more likely to harbor phages (P < 0.001 by single-factor analysis of variance) than were standard or economy products. This study demonstrates that Campylobacter bacteriophages, along with their hosts, can survive commercial poultry processing procedures and that the phages exhibited a wide range of recovery rates from chicken skin stored at 4°C.     

Isolation and Characterization of Bacteriophages Infecting Staphylococcus epidermidis

Bacteriophages infecting Staphylococcus epidermidis were isolated by mitomycin C induction. Three distinct phages (vB_SepiS-phiIPLA5, vB_SepiS-phiIPLA6, and vB_SepiS-phiIPLA7)—defined by plaque morphology, structure, virion proteins pattern, DNA restriction bands, and host range—were obtained. One-step growth curves of bacteriophages under optimal growth conditions for S. epidermidis F12 revealed eclipse and latent periods of 5–10 and 10–15 min, respectively, with burst sizes of about 5 to 30 PFU per infected cell. Transmission electron microscopy revealed that the phages were of similar size and belonged to the Siphoviridae family. Phage phi-IPLA7 had the broadest host range infecting 21 out of 65 S. epidermidis isolates. Phage phi-IPLA5 seemed to be a virulent phage probably derived from phi-IPLA6. Phages phi-IPLA5 and phi-IPLA7 exhibited increasing plaques surrounded by a halo that could be indicative of a polysaccharide depolymerase activity. Viable counts, determined during the infection of S. epidermidis F12, confirmed that phi-IPLA5 had a potent lytic capability and reduced S. epidermidis population by 5.67 log units in 8 h of incubation; in the presence of the mixture of phi-IPLA6 and phi-IPLA7, however, a reduction of 2.27 log units was detected     

Isolation and Characterization of Two Basic Internal Proteins from the T-Even Bacteriophages

Two species of basic internal proteins were found in osmotic shock supernatant solutions of bacteriophages T4B, T4D, T2H, T2L, and T6. The major species of protein isolated had a molecular weight of approximately 21,000 daltons, whereas the minor protein molecular weight was near 9,500 daltons. The two protein species exhibited unique isoelectric points and amino acid compositions. The 21,000-dalton protein of T2L showed major electrophoretic and compositional differences from the other 21,000-dalton proteins isolated. Similarities between the 21,000-dalton proteins and phage lysozyme are discussed.     

Isolation and Characterization of Temperate Bacteriophages of Clostridium difficile

The lack of information on bacteriophages of Clostridium difficile prompted this study. Three of 56 clinical C. difficile isolates yielded double-stranded DNA phages C2, C5, C6, and C8 upon induction. Superinfection and DNA analyses revealed relatedness between the phages, while partial sequencing of C2 showed nucleotide homology to the sequenced C. difficile strain CD630.     

山西高平市农家酸菜汁中乳酸菌的分离及鉴定

对山西高平市农家酸菜汁中乳酸菌含量和菌种进行分析研究,经分离筛选得到3株产酸能力较强的菌株,对其进行鉴定并对其生理特性进行研究。通过对其形态特征、菌落特征及生化特征进行分析可知,其中2株为植物乳杆菌(Lactobacillus plantarum),1株为短乳杆菌(Lactobacillus breris)。对筛选到的乳酸菌进行酸菜发酵性能研究发现,其中菌株L2在发酵24 h后,发酵液pH值达到3.47、酸度达到1.67 g乳酸/100 mL、菌落数达到1.37×107cfu/mL的高密度富集状态,且发酵无异味,口味自然,效果较好。     

Isolation and Partial Characterization of Two Aeromonas hydrophila Bacteriophages

Two Aeromonas hydrophila bacteriophages, Aeh1 and Aeh2, were isolated from sewage. Both phages showed binal symmetry. The dimensions of A. hydrophila phages Aeh1 and Aeh2 differed from those of the other Aeromonas phages. Also, phage Aeh2 was the largest Aeromonas phage studied to date. Phage Aeh1 formed small, clear plaques, and phage Aeh2 formed turbid plaques with clear centers. Both phages were sensitive to chloroform treatment, being totally inactivated after treatment for 1 h at 60°C at pH 3 and 11. However, the infectivity of Aeh1 phage stocks increased by approximately fivefold after they were treated at pH 10 for 1 h at 22°C. Phages Aeh1 and Aeh2 were serologically unrelated and had latent periods of 39 and 52 min, respectively. The average burst sizes of phages Aeh1 and Aeh2 were 17 and 92 PFU per cell, respectively. Phage Aeh1 infected 13 of 22 A. hydrophila strains tested, whereas phage Aeh2 infected only its original host. Phage Aeh1 infected some A. hydrophila strains only at or below 37°C. Neither phage infected the two A. (Plesiomonas) shigelloides strains used in this study.     

Isolation and Preliminary Characterization of Bacteriophages for Bdellovibrio bacteriovorus

Ten bacteriophages that attack and lyse saprophytic strains of Bdellovibrio bacteriovorus were isolated. Morphological, serological, and host-range studies revealed that there were four different bdellovibrio phages present among the isolates. One of the phages lysed a strain of B. bacteriovorus that requires the presence of a suitable bacterial host for growth. The phage attached to the bdellovibrio cells in the absence of the bacterial host cells; lysis occurred only in the presence of host cells. The 19 saprophytic bdellovibrio strains employed in the phage host-range studies were grouped on the basis of their susceptibility to phage lysis.     

Escherichia coli K Bacteriophages I. Isolation and Introductory Characterization of Five Escherichia coli K Bacteriophages

A set of five Escherichia coli K phages has been isolated. These phages are adsorbed to and lyse the capsular forms of the host bacteria, whereas their spontaneous, acapsular mutants are not affected. All host strains are heavily encapsulated test strains for E. coli K antigens of the thermostable A type and they readily segregate acapsular mutants. In four of the phage-host systems, all secondary growth obtained was found to be acapsular. When tested for host-range mutants on 38 strains of E. coli and Klebsiella, less than one mutant per 10(5) plaque-forming units was found. No cross-reacting neutralizing antibodies were obtained when rabbits were immunized with the K phages. The latent periods (between 16 and 30 min) and average burst sizes (between 145 and 580) were determined by one-step growth experiments.     

Isolation and Partial Characterization of Bacteriophages of the Phytopathogen Pseudomonas syringae

Bacteriophages isolated from culture supernatants of Pseudomonas syringae pv. syringae and from sewage were identified. The DNA from each phage was isolated and digested with the restriction endonuclease EcoRI. Eight isolates were determined to be different, with two phage isolates from sewage having restriction patterns identical to two phages from culture supernatants. The sizes of the phage DNA ranged from 24 to49 kilobases for isolates from sewage and from 39 to 52.5 kilobases for the isolates from culture supernatants. Buoyant densities of phage particles in CsCl varied from 1.498 to 1.507 g/cm³ for isolates from sewage and from 1.506 to 1.516 g/cm³ for isolates from culture supernatants. Electron microscopy revealed four morphological types. Based on plaque-forming ability of culture supernatants, 31 out of 47 strains of P. syringae are probably lysogenic.     

深圳赤潮中霍乱弧菌噬菌体的分离筛选及生物学特性分析

本文以从鲍鱼肠道中分离出的霍乱弧菌SWBC-a为宿主菌,采用双层平板法从2004年夏季深圳大梅沙海域赤潮海水样品中分离得到10株噬菌体。通过对分离出的噬菌体效价、RTD值的测定、形态特征的观察及对属内外细菌的交叉裂解谱的研究,筛选出3株裂解谱较宽的强效噬菌体。选取其中2株高效裂解噬菌体PsaA和PsaH做生物学特性分析,证明除镁离子有利于噬菌体裂解外,温度、pH值、紫外照射、柠檬酸钠对噬菌体裂解都有不同程度的抑制。本研究结果为进一步利用噬菌体研发用于消除霍乱弧菌的微生态制剂和诊断食源性霍乱弧菌提供了理论基础。     

Isolation of Bacteriophages from Group H Streptococci

Temperate phages were isolated from 5 of the 17 strains of group H streptococci tested.     

Bacteriophages of Rhodopseudomonas spheroides: Isolation and Characterization of a Rhodopseudomonas spheroides Bacteriophage

A DNA-containing bacteriophage, designated RS1, infecting Rhodopseudomonas spheroides 2.4.1, has been isolated from sewage. The buoyant density of RS1 in CsCl equilibrium centrifugation is 1.50 g/cm(3), and the buoyant density of RS1 DNA is 1.706. The phage possesses a polyhedral head, approximately 65 nm in diameter, and a tail 60 nm long. When grown on aerobic cells, RS1 has a latent period of 120 min and an average burst size of 20. When grown on anaerobic cells, RS1 has a latent period of 150 min, and a burst size similar to that observed during aerobic infection. The adsorption rate constant of RS1 to aerobic cells is 1.2 x 10(-9) ml/min, and 0.58 x 10(-9) ml/min to anaerobic cells. Adsorption of RS1 to R. spheroides requires the presence of divalent cations.     

Isolation and Characterization of Bacteriophages from Inland Saline Aquaculture Environments to Control Vibrio parahaemolyticus Contamination in Shrimp

Among the various bacterial pathogens associated with the aquaculture environment, Vibrio parahaemolyticus the important one from shrimp and human health aspects. Though having been around for several decades, phage-based control of bacterial pathogens (phage therapy) has recently re-emerged as an attractive alternative due to the availability of modern phage characterization tools and the global emergence of antibiotic-resistant bacteria. In the present study, a total of 12 V. parahaemolyticus specific phages were isolated from 264 water samples collected from inland saline shrimp culture farms. During the host range analysis against standard/field isolates of V. parahaemolyticus and other bacterial species, lytic activity was observed against 2.3–45.5% of tested V. parahaemolyticus isolates. No lytic activity was observed against other bacterial species. For genomic characterization, high-quality phage nucleic acid with concentrations ranging from 7.66 to 220 ng/µl was isolated from 9 phages. After digestion treatments with DNase, RNase and S1 nuclease, the nature of phage nucleic acid was determined as ssDNA and dsDNA for 7 and 2 phages respectively. During transmission electron microscopy analysis of phage V5, it was found to have a filamentous shape making it a member of the family Inoviridae. During efficacy study of phage against V. parahaemolyticus in shrimp, 78.1% reduction in bacterial counts was observed within 1 h of phage application. These results indicate the potential of phage therapy for the control of V. parahaemoyticus in shrimp.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12088-021-00934-6.     

Isolation of Yersinia ruckeri Bacteriophages

Eight bacteriophages effective against Yersinia ruckeri, the enteric redmouth disease bacterium, were isolated. Phage YerA41, a tailed icosahedral virus isolated from sewage enrichments, lysed 34 of 35 strains of Y. ruckeri serovar I, but was inactive against 15 strains belonging to three other serological groups. Six other phages lysed strains of serovars II, V, and I′, a subgroup of serovar I. YerL62, a phage obtained by mitomycin C induction, was specific for one of three serovar V strains. These bacteriophages, particularly YerA41, have potential value for fish disease diagnostic work.     

Isolation and Characterization of Lytic Properties of Bacteriophages Specific for M. haemolytica Strains

ResultsEight bacteriophages were obtained, like typical of the families Myoviridae, Siphoviridae and Podoviridae. Most of the bacteriophages exhibited lytic properties against the M. haemolytica strains. Restriction analysis revealed similarities to the P2-like phage obtained from the strain M. haemolytica BAA-410. The most similar profiles were observed in the case of bacteriophages φA1 and φA5. All of the bacteriophages obtained were characterized by the presence of additional fragments in the restriction profiles with respect to the P2-like reference phage. In the analysis of PCR products for the P2-like reference phage phi-MhaA1-PHL101 ({"type":"entrez-nucleotide","attrs":{"text":"DQ426904","term_id":"90110542","term_text":"DQ426904"}}DQ426904) and the phages of the M. haemolytica serotypes, a 734-bp phage PCR product was obtained. The primers were programmed in Primer-Blast software using the structure of the sequence {"type":"entrez-nucleotide","attrs":{"text":"DQ426904","term_id":"90110542","term_text":"DQ426904"}}DQ426904 of reference phage PHL101.ConclusionsThe results obtained indicate the need for further research aimed at isolating and characterizing bacteriophages, including sequence analysis of selected fragments. Moreover, standardization of methods for obtaining them in order to eliminate M. haemolytica bacteria involved in the etiopathogenesis of BRDC is essential.     

Isolation from Sewage Influent and Characterization of Novel Staphylococcus aureus Bacteriophages with Wide Host Ranges and Potent Lytic Capabilities

Staphylococcus aureus is a gram-positive pathogen that causes a variety of diseases, including bovine mastitis, which has severe economic consequences. Standard antibiotic treatment results in selection of resistant strains, leading to a need for alternative treatments, such as bacteriophage therapy. Forty-nine S. aureus isolates were obtained from the milk of mastitic cows for use in screening of staphylococcal phages. Fifteen isolates which were positive for both coagulase and hemolysin were assayed by PCR for variation in the X region and the immunoglobulin G-binding region of the protein A gene (spa) and in the carboxy terminus of the coagulase gene (coa) and for the presence of enterotoxin C, G, H, and I genes. The host ranges of 52 phages isolated from sewage influent were determined by performing spot tests with the 15 S. aureus isolates, and two phages were subsequently chosen for further analysis. ΦSA039 had the widest host range, producing clear plaques on 13 of the 15 isolates (87%), while ΦSA012 produced clear plaques on 8 isolates (53%) and was the only phage that produced a clear plaque on a nonmastitic S. aureus strain. Transmission electron microscopy revealed that the phages were similar sizes and belonged to the Myoviridae family. Measurement of optical densities during coculture with S. aureus isolates confirmed the breadth of the ΦSA039 host range and showed that ΦSA012 had potent lytic capability. ΦSA012-resistant bacteria did not appear for three of seven isolates tested (43%) after 65 h of incubation. These two phages are proposed as candidates for phage therapy of bovine mastitis.In the dairy industry, mastitis is a widespread problem responsible for important decreases in milk production. Economic losses of $100 million per year have been estimated for farms in Hokkaido, one of the largest milk-producing areas in Japan (28). Mastitis can be caused by over 150 different microorganisms, and one of the most important of these organisms is Staphylococcus aureus (22). After diagnosis of mastitis, the standard treatment regimen consists of isolating the diseased cow and treating it with antibiotics. However, this approach has drawbacks, such as its high cost and the eradication of harmless or beneficial organisms due to the lack of specificity of antibiotics. Additionally, the incidence of antibiotic-resistant bacteria has increased in recent years (4). As a result, there has been renewed interest in the use of other natural or engineered antimicrobial agents as an alternative or supplementary treatment for staphylococcal diseases such as mastitis (11, 21, 26). One group of alternatives with great potential involves bacteriophages (phages) and their derivatives, and a number of promising candidates have been described (2, 5, 7, 13, 17, 18, 27), notably bacteriophage K.One of the main obstacles to successful treatment of mastitis using phages is the fact that most phages are able to infect only a very narrow range of hosts. Given the plural etiology of many mastitis cases, it is desirable to find a phage-based treatment that targets a wide range of pathogens. One proposed method is a combination of two or more phages with different and broad host ranges in a cocktail (25, 29). This method has been shown to delay considerably the appearance of phage-resistant cells in Escherichia coli cultures, providing hope that similar success can be achieved using S. aureus phages.In this paper we describe the isolation of two novel phages, designated ΦSA012 and ΦSA039, that were found to have a lytic effect on a broad range of S. aureus isolates obtained from mastitic milk. ΦSA012 and ΦSA039 were selected after host range analysis from a total of 52 phages screened from sewage influent. These two phages were characterized morphologically by using electron microscopy, and their effects on representative strains of S. aureus were examined by coculture in milk.     

Characterization of Salmonella Bacteriophages Isolated from Swine Lagoon Effluent

Four Salmonella bacteriophages that had been originally isolated from swine manure lagoons were characterized and compared to each other and to well-known Salmonella phages P22 and Felix 01. Host ranges of the lagoon phages were similar to each other in spot tests on reference strains of Salmonella, but differed slightly from each other on a panel of Salmonella lagoon strains. In single-step growth at 35°C the lagoon phages had latent periods of 15 to 20 min and burst sizes from 100 to 230. The lagoon phages and P22 were purified by cesium chloride (CsCl) gradient centrifugation and used to produce specific antisera and DNA. The lagoon phages were indistinguishable from each other but distinct from P22 and Felix 01 in immunodiffusion and infectivity neutralization tests and in restriction digest analysis. Mention of a trade name, proprietary product, or specific equipment does not constitute a guarantee, warranty, or endorsement by the USDA and does not imply its approval to the exclusion of other products that may be suitable.