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PubMed1981

Dispersal and fate of coliphages in the River Saar.

The dispersal of coliphages was analysed quantitatively in longitudinal profiles of the partially canalized River Saar. The numbers of coliphages ranged between 0 and 2380 PFU (plaque-forming units)/ml in the water and between 0 and 2550 PFU/g in the sediments, depending on the degree of fecal pollution. In canalized parts the phages were found mainly in the sediments, whereas in non-canalized parts they prevailed in the water phase (Fig. 3). Determinations performed at municipal sewage water outfalls revealed that in the canalized part most of the phages precipitated to the sediment on a water course of only about 250 m (Fig. 5). In the non-canalized part of the River Saar, sediments were not contaminated to this extent, since phages rather remained in the water, due to high current velocities (Fig. 5). According to these results coliphages are, just like some fecal bacteria, closely associated with particles. They are rapidly inactivated in the sediment and water (inactivation rates: 0.12 to 0.66 d-1).

Clostridium perfringens↗

PubMed1998

Yeast positive-stranded virus-like RNA replicons. 20 S and 23 S RNA terminal nucleotide sequences and 3' end secondary structures resemble those of RNA coliphages.

Saccharomyces cerevisiae strains carry single-stranded RNAs called 20 S RNA and 23 S RNA. These RNAs and their double-stranded counterparts, W and T dsRNAs, have been cloned and sequenced. A few nucleotides at both ends, however, remained unknown. These RNAs do not encode coat proteins but their own RNA-dependent RNA polymerases that share a high degree of conservation to each other. The polymerases are also similar to the replicases of RNA coliphages, such as Qbeta. Here we have determined the nucleotide sequences of W and T dsRNAs at both ends using reverse transcriptase polymerase chain reaction-generated cDNA clones. We confirmed the terminal sequences by primer-extension and RNase protection experiments. Furthermore, these analyses demonstrated that W and T dsRNAs and their single-stranded RNA counterparts (i) are linear molecules, (ii) have identical nucleotide sequences at their ends, and (iii) have no poly(A) tails at their 3' ends. Both 20 S and 23 S RNAs have GGGGC at the 5' ends and the complementary 5-nucleotides sequence, GCCCC-OH, at their 3' ends. S1 and V1 secondary structure-mapping of the 3' ends of 20 S and 23 S RNAs shows the presence of a stem-loop structure that partially overlaps with the conserved 3' end sequence. Nucleotide sequences and stem-loop structures similar to those described here have been found at the 3' ends of RNA coliphages. These data, together with the similarity of the RNA-dependent RNA polymerases encoded among these RNAs and RNA coliphages, suggest that 20 S and 23 S RNAs are plus-strand single-stranded virus-like RNA replicons in yeast.

Cloning, Molecular↗

PubMed1982

Adsorption of coliphages T1 and T7 to clay minerals.

Coliphages T1 and T7 of Escherichia coli were absorbed by kaolinite (K) and montmorillonite (M). Maximum adsorption of T7 (96%) to M was greater than that of T1 (84%), but the adsorption of both coliphages to K was the same (99%). Positively charged sites (i.e., anion exchange sites) on the clays appeared to be primarily responsible for the adsorption of T1 to K but only partially responsible for the adsorption of T1 to M; equilibrium adsorption isotherms of T1 to K and M did not show a correlation between adsorption and the cation exchange capacity of the clays, and the reduction in adsorption caused by sodium metaphosphate (a polyanion that interacts with positively charged sites on clay) was more pronounced with K than with M. The equilibrium adsorption isotherms of T7 to K and M suggested a correlation between adsorption and the cation exchange capacity of the clays. However, studies with sodium metaphosphate indicated that T7 also adsorbed to positively charged sites on the clays, especially on K. Adsorption of the coliphages to positively charged sites was greater with K than with M, probably because the ratio of positively charged sites to negatively charged sites was greater on K than on M.

Adsorption↗

PubMed2003

The genome sequence of Yersinia pestis bacteriophage phiA1122 reveals an intimate history with the coliphage T3 and T7 genomes.

The genome sequence of bacteriophage phiA1122 has been determined. phiA1122 grows on almost all isolates of Yersinia pestis and is used by the Centers for Disease Control and Prevention as a diagnostic agent for the causative agent of plague. phiA1122 is very closely related to coliphage T7; the two genomes are colinear, and the genome-wide level of nucleotide identity is about 89%. However, a quarter of the phiA1122 genome, one that includes about half of the morphogenetic and maturation functions, is significantly more closely related to coliphage T3 than to T7. It is proposed that the yersiniophage phiA1122 recombined with a close relative of the Y. enterocolitica phage phiYeO3-12 to yield progeny phages, one of which became the classic T3 coliphage of Demerec and Fano (M. Demerec and U. Fano, Genetics 30:119-136, 1945).

Amino Acid Sequence↗

PubMed1993

A two-dimensional model at the nucleotide level for the central hairpin of coliphage Q beta RNA.

We describe the construction and testing of a structural model at the nucleotide level for conformation CH of the central hairpin of genomic RNA from coliphage Q beta. The model was developed with the computer program MFOLD using both optimal and suboptimal predictions. Structural information obtained by electron microscopic analysis of Kleinschmidt spreadings of Q beta RNA was used to guide the modeling. The model was tested in solution with three enzymatic probes: RNase T1, RNase T2, and RNase V1, as well as four chemical probes: dimethylsulfate, diethylpyrocarbonate, kethoxal and 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide metho-p-toluene sulfonate (CMCT). The structural analyses in solution are consistent with the predicted structural model. The model is also supported by comparative structural analysis with the related coliphage SP. The model provides a structural basis for published biochemical and genetic studies implicating large, long-range structural features in the co-regulation of viral coat and replicase expression. In addition, we show that the read-through region of the viral protein A1 forms a separate structural domain, and we suggest that it functions as a nucleation site that participates in the folding and refolding of the molecule during replication and translation. In addition to the central hairpin, we have analyzed the structure of the viral coat initiation region. Our studies show that the entire region consists of small local hairpins and that 26 nucleotides immediately surrounding the coat initiation codon are single-stranded.

Base Sequence↗

PubMed1998

Removal of waterborne human enteric viruses and coliphages with oxidized coal.

Human enteric viruses and coliphages have been detected in water that has undergone what is generally considered adequate treatment, including chlorination. Because small numbers of virus particles are needed for the initiation of a productive virus infection, the presence of any number of virus particles in water resources will always be of concern. In this investigation the ability of oxidized coal to remove viruses from water was investigated. The oxicoal product was found to be able to remove not only coliphages, but also various pathogenic human viruses from seeded water sources. Removal was dependent upon the type of virus, the period of exposure, and the concentration of oxidized coal.

Coal↗

PubMed1976

Inactivation of the T7 coliphage by monofunctional alkylating agents. Action of phage adsorption and injection of its DNA.

Alkylation by ethyl or methyl methanesulfonate to an extent that inactivates more than 99.5% of T7 coliphages has no effect on phage adsorption on Escherichia coli B cells, but decreases the amount of phage DNA injected into the host cells. Depurination interferes with the injection of the phage DNA. Failure to inject the whole phage genome thus appears to be a cause of the immediate as well as of the delayed inactivation of the T7 coliphage treated by monofunctional alkylating agents; the hypothesis that it is the only cause of inactivation, although not very likely, cannot be excluded at the present time.

Binding Sites↗

PubMed2005

Response of four types of coliphages to high hydrostatic pressure.

Pressure inactivation of four types of coliphages, varphiX 174 (ssDNA virus), MS2 (ssRNA virus), lambda imm434 (dsDNA virus) and T4 (dsDNA virus), was studied to evaluate their potential as human enteric viral surrogates for use in validation of commercial pressure processing treatments. Phage varphiX 174 demonstrated an unexpected high resistance to pressure with no more than 1-log(10) reduction observed following exposures to 350-600 MPa. There was no greater than 1-log(10) reduction below 500 MPa for MS2 in modified phosphate-buffered saline, but a 3.3-log(10) reduction was observed for MS2 pressurized at 600 MPa. Coliphages lambda imm434 and T4 were relatively sensitive to pressure in demonstrating inactivation at 350 MPa. At 21 degrees C, lambda imm434 was inactivated in modified phosphate-buffered saline or Dulbecco's Modified Eagle's Medium plus 5% fetal bovine sera by at least 7.5-log(10) when exposed to 400 MPa for 5 min. Treatment at 450 MPa for 5 min was necessary to obtain a log(10) reduction of 6-7 for T4.

Analysis of Variance↗

PubMed2001

Rapid coliphage detection assay.

A rapid coliphage detection assay was developed, based on the phage-induced release of beta-galactosidase from cells of Escherichia coli. The assay could detect as few as five coliphage per sample without an overnight incubation period. The range of acceptable assay parameters was identified.

Coliphages↗

PubMed1998

Fecal coliform-related bacterial and coliphage populations in five lakes of southeastern Spain.

Aerobic heterotrophic bacteria, fecal and total coliforms, fecal streptococci and coliphages were isolated from five protected lakes in the Antequera area of Spain over the time from January to March (1994-96). The water samples contained large number of heterotrophic bacteria (mean counts 0.2 to 5.0 x 10(7) cfu per 100 ml). Most of the lakes contained fecal streptococci and a relationship between streptococci and salinity of the water samples was established. Coliphages were isolated from lakes containing fecal coliform and these bacteria were taxonomically identified as E. coli. Coliform bacilli do not seem to be an adequate indicator of fecal pollution for these ephemeral small lakes.

Animals↗

PubMed1978

The behaviour of f2 coliphage in activated sludge treatment.

A model activated sludge treatment plant was used which was functionally very similar to a full-scale plant. It was inoculated with f2 coliphage and the titres of virus in the influent, the mixed liquor and the effluent were monitored regularly. The distribution of the virus in the solids and liquid fractions of the mixed liquor was in the ratio of 18:82 and 20.4% of the influent virus was recovered in the effluent. After inoculation was stopped the titre of virus in the solids fraction of the mixed liquor remained high and unaltered for up to 70 h, whereas the value for effluent reverted to the low background titre originally present. These results are discussed in relation to those reported for poliovirus and it is concluded that f2 coliphage is not a suitable model for studies of the behaviour of human enteroviruses.

Coliphages↗

PubMed2003

Bacterial host strains that support replication of somatic coliphages.

Somatic coliphages detected by Escherichia coli strain WG5 have been proposed as potential indicators of water quality. Their potential replication in the water environment is considered a drawback for their use as indicators. However, the contribution of replication outside the gut to the total numbers has never been quantified. It has not been determined either the fraction of bacterial strains that might support replication of phages detected by strain WG5 in the water environment. We examined the sensitivity of 291 host strains to 25 phages by streaking slants of the presumptive host strain onto an agar layer that contains bacteriophages, which gives a total of 7275 combinations (sensitivity tests). Only a 3.02% of the tests showed sensitivity. Additionally, six environmental strains were used as hosts to count phages in sewage and seawater. Phages isolated on these strains were used to infect strain WG5. The environmental strains detected 1 log10 fewer phages than strain WG5 in sewage and seawater. The fraction of phages that were detected by the six strains and that also infected strain WG5 ranged from < 0.07% to < 2.0% of the total amount of bacteriophages detected by strain WG5 in the same samples. Our results confirm that less than 3% of naturally occurring hosts support replication of phages infecting E. coli. We conclude that the contribution of replication to the number of somatic coliphages detected in the aquatic environment is negligible.

Coliphages↗

PubMed1973

Nucleotide sequences of similar size from the coliphage R17 genome.

A sequence of 33 nucleotides from the coliphage R17 RNA genome was determined. It constitutes the main component of a mixture of fragments that migrate together on electrophoresis in a separation according to molecular weight. Fragments of comparable chain length from 3' end of RNA from coliphage R17, from a region preceding and overlapping the coat-protein cistron ribosome binding site and from the beginning of the A-protein cistron, were also found and characterized. ;Hairpin'-like secondary structures are proposed for the longer fragments, one of which appears to have a tetranucleotide excised in the loop region.

Autoradiography↗

PubMed1999

Study of the potential relationship between the morphology of infectious somatic coliphages and their persistence in the environment.

The proportions of different morphological types of infectious somatic coliphages were determined in faecally polluted freshwaters. Myoviridae, followed by Siphoviridae, were the most frequently isolated morphological types in raw sewage, treated sewage and river water collected a few metres downstream from a sewage outfall. However, in river water collected further downstream from the pollution point, in river water after 'in situ' inactivation experiments and in chlorinated raw and treated sewage significant changes in the proportions of the different somatic coliphage morphological types occurred. In all cases, Siphoviridae, especially those with flexible and curled tails, became more abundant to the detriment of Myoviridae.

Caudovirales↗

PubMed1997

The dual role of Apl in prophage induction of coliphage 186.

In the present study we show that the Apl protein of the temperate coliphage 186 combines, in one protein, the activities of the coliphage lambda proteins Cro and Xis. We have shown previously that Apl represses both the lysogenic promoter, pL, and the major lytic promoter, pR, and is required for excision of the prophage. Apl binds at two locations on the phage chromosome, i.e. between pR and pL and at the phage-attachment site. Using an in vivo recombination assay, we now show that the role of Apl in excision is in the process itself and is not simply a consequence of repression of pR or pL. To study the repressive role of Apl at the switch promoters we isolated Apl-resistant operator mutants and used them to demonstrate a requirement for Apl in the efficient derepression of the lysogenic promoter during prophage induction. We conclude that Apl is both an excisionase and transcriptional repressor.

Attachment Sites, Microbiological↗

PubMed1979

Interaction of Escherichia coli RNA polymerase with promoters of several coliphage and plasmid DNAs.

The interaction of Escherichia coli RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) with restriction fragments obtained from various E. coli related DNAs was studied in vitro. The DNAs investigated included several coliphage genomes (T5, lambda, T7, fd) and plasmid DNAs (pML 21, pSC101). By using the nitrocellulose filter binding of the enzyme-DNA complexes, fragment-specific relative rates of complex formation as well as complex stabilities were determined. Promoter-specific relative rates of polymerase binding were derived from fragment-specific rates by taking into account the number of major binding sites for RNA polymerase within several DNAs. Estimates of the stability of complexes formed between some major binding sites and the enzyme were obtained by studying the rate of complex decay. Both characteristics--rate of complex formation and rate of decay--varied widely and independently of each other. The promoters reacting most efficiently with E. coli RNA polymerase were found in the early region of coliphage T5 whereas some promoters in pML 21, or for example, the lambda promoter PI, belong to signals binding the enzyme most slowly. Based on the second-order rate constant determined for the interaction of E. coli RNA polymerase with promoters of phage fd, the fastest promoters characterized so far reacted with rates in the order of 10(8) M-1s-1. The hierarchy of promoters established here is of interest from the viewpoint that promoter strength correlates with the rate of polymerase binding. Among the promoters studied here this rate spans a range of 2 orders of magnitude.

Base Sequence↗

PubMed1989

UV induction of coliphage 186: prophage induction as an SOS function.

Our results show that UV induction of the 186 prophage depends upon the phage function Tum, with the mutant phenotype of turbid plaques on mitomycin plates and the expression of which is controlled by the host LexA protein. Tum function, encoded near the right-hand end of the coliphage 186 chromosome, is under the control of promoter p95. This promoter is overlapped by a sequence closely related to the consensus sequence of the LexA-binding site. It is proposed that inactivation of LexA after UV irradiation (or by genetic means) leads to prophage induction by permitting expression of Tum which, by unknown means, induces prophage. This mechanism is basically different from that seen with the UV-inducible lambdoid coliphages, which are not regulated by LexA.

Base Sequence↗

PubMed2003

Removal of Encephalitozoon intestinalis, calicivirus, and coliphages by conventional drinking water treatment.

The removal of the Microsporidia, Encephalitozoon intestinalis, feline calicivirus and coliphages MS-2, PRD-1, and Fr were evaluated during conventional drinking water treatment in a pilot plant. The treatment consisted of coagulation, sedimentation, and mixed media filtration. Fr coliphage was removed the most (3.21 log), followed by feline calicivirus (3.05 log), E. coli (2.67 log), E. intestinalis (2.47 log), MS-2 (2.51 log). and PRD-1 (1.85 log). With the exception of PRD-1 the greatest removal of the viruses occurred during the flocculation step of the water treatment process.

Animals↗