Subgroups in Group III of RNA Phages

In our previous studies, RNA phage strains were separated into 3 major groups on the basis of filtration efficiency through Millipore filters. In the present study, the strains of group III were shown to be further divided into 3 subgroups: (a) Qβ, NH, SG; (b) VK, SO; (c) ST. This subgrouping was found to be compatible with the serological grouping and pronase sensitivity with the exception of strain NM. Strain NM was classified in subgroup (a) by the Millipore filtration and in subgroup (b) by the other two methods.     

Comparative Persistence of Subgroups of F-Specific RNA Phages in River Water

F-specific (F+) RNA phages are widely used as indicators for the presence of fecal contamination and/or enteric viruses in water, and identifying subgroups of F+ RNA phages provides an approach for microbial source tracking. Different survival characteristics of the F+ RNA phage subgroups result in a misinterpretation of their original proportion in water, thus giving misleading information when they are used for microbial source tracking. This study investigated the comparative persistence of subgroups of F+ RNA phages in river water under different conditions. Results suggested that temperature and pH are the major factors affecting the persistence of F+ RNA phages in river water, and organic substances promote phage survival. The comparative persistence patterns of subgroups of F+ RNA phages varied and may bias extrapolation of their initial proportions in surface water. Thus, the characteristics of water should be taken into consideration and the results should be carefully interpreted when F+ RNA phages are used for microbial source tracking.     

Comparison of DNA sizes in a group of giant Pseudomonas aeruginosa phages by the PFGE method

Genome sizes of Pseudomonas aeruginosa phages phiKZ and EL earlier determined by sequence analysis were shown to correspond to sizes of their DNAs assessed by pulse-electrophoresis (PFGE). Putative "redundant" genes in phiKZ phage genome are supposed to control functions promoting vigorous growth of the phage belonging to this species, compared to phages of EL species.     

An oligonucleotide hybridization assay for the identification and enumeration of F-specific RNA phages in surface water

F-specific RNA phages can be used as model organisms for enteric viruses to monitor the effectiveness of sewage treatment, and to assess the potential contamination of surface water with these viruses. In this paper a method is described which identifies RNA phages quantitatively by a plaque hybridization assay. Oligonucleotide probes were developed that can assign phages to their phylogenetic subgroups. Such a distinction is important, since some subgroups preferentially occur in sewage of human origin, while others tend to be associated with animal wastewater. The method has been tested on a large number of isolates and represents an improvement in time and reliability over the previously used serological classification.     

Occurrence, survival, and persistence of human adenoviruses and F-specific RNA phages in raw groundwater

Detection of specific genetic markers can rapidly identify the presence of enteric viruses in groundwater. However, comparison of stability characteristics between genetic and infectivity markers is necessary to better interpret molecular data. Human adenovirus serotype 2 (HAdV2), in conjunction with MS2 phages or GA phages, was spiked into raw groundwater microcosms. Viral stability was periodically assessed by both infectivity and real-time PCR methods. The results of this yearlong study suggest that adenoviruses have the most stable persistence profile and an ability to survive for a long time in groundwater. According to a linear regression model, infectivity reductions of HAdV2 ranged from 0.0076 log(10)/day (4°C) to 0.0279 log(10)/day (20°C) and were significantly lower than those observed for phages. No adenoviral genome degradation was observed at 4°C, and the reduction was estimated at 0.0036 log(10)/day at 20°C. Occurrence study showed that DNA of human adenoviruses could be observed in groundwater from a confined aquifer (7 of the 60 samples were positive by real-time PCR), while no fecal indicators were detected. In agreement with the persistence of genetic markers, the presence of adenoviral DNA in groundwater may be misleading in term of health risk, especially in the absence of information on the infective status.     

Isolation and partial characterization of phages infectingCitrobacter intermedius C3

Several new bacteriophages infecting strain C3 ofCitrobacter intermedius have been isolated from fresh waters. The physicochemical properties, plaque morphology, growth characteristics, virion structure, and immunology of eight bacteriophage isolates are reported. The phages are classified into three groups according to their characteristics.     

Rearrangements in the 5′ Nontranslated Region and Phylogenetic Analyses of Cucumber Mosaic Virus RNA 3 Indicate Radial Evolution of Three Subgroups

Cucumber mosaic virus (CMV) has been divided into two subgroups based on serological data, peptide mapping of the coat protein, nucleic acid hybridization, and nucleotide sequence similarity. Analyses of a number of recently isolated strains suggest a further division of the subgroup I strains. Alignment of the 5' nontranslated regions of RNA 3 for 26 strains of CMV suggests the division of CMV into subgroups IA, IB, and II and suggests that rearrangements, deletions, and insertions in this region may have been the precursors of the subsequent radiation of each subgroup. Phylogeny analyses of CMV using the coat protein open reading frame of 53 strains strongly support the further division of subgroup I into IA and IB. In addition, strains within each subgroup radiate from a single point of origin, indicating that they have evolved from a single common ancestor for each subgroup.     

Enzymatic synthesis of 3-(3-amino-3-carboxypropyl)uridine in Escherichia coli phenylalanine transfer RNA: transfer of the 3-amino-acid-3-carboxypropyl group from S-adenosylmethionine

A novel enzyme which catalyzes the transfer of the 3-amino-3-carboxypropyl group into tRNA to form 3-(3-amino-3-carboxypropyl)uridine was isolated from $\text{Escherichia coli}$. The enzyme required S-adenosylmethionine as donor molecule.     

A closely related group of RNA-dependent RNA polymerases from double-stranded RNA viruses.

Probably one of the first proteinaceous enzymes was an RNA-dependent RNA polymerase (RDRP). Although there are several conserved motifs present in the RDRPs of most positive and double-stranded RNA (dsRNA) viruses, the RDRPs of the dsRNA viruses show no detectable sequence similarity outside the conserved motifs. There is now, however, a group of dsRNA viruses of lower eucaryotes whose RDRPs are detectably similar. The origin of this sequence similarity appears to be common descent from one or more noninfectious viruses of a progenitor cell, an origin that predates the differentiation of protozoans and fungi. The cause of this preservation of sequence appears to be constraints placed on the RDRP by the life-style of these viruses--the maintenance of a stable, persistent, noninfectious state.     

3' Terminal labelling of RNA of RNA with beta-32P-pyrophosphate group and its application to the sequence analysis of 5S RNA from Streptomyces griseus.

Nucleotide pyrophosphate transferase isolated from Streptomyces griseus is used to transfer pyrophosphate group from gamma-32P-ATP to the 3'-OH of tRNA, generating a strictly terminal label at its 3' end. Using yeast tRNAPhe as model compound, it is demonstrated that the labelled molecule is suitable for rapid gel sequencing by both enzymatic and chemical methods. RNA molecules terminated by pyrimidine nucleoside are poor pyrophosphate acceptors. To label RNAs of this kind, first guanosine 5'-phosphate 3'-(beta-32P)-pyrophosphate (pGpp) is prepared from gamma-32P-ATP and GMP by nucleotide pyrophosphate transferase. pGpp is then ligated to the 3' end of RNA by T4 RNA ligase. The complete nucleotide sequence of 5S RNA from Streptomyces griseus is established by rapid gel sequencing methods performed on 3'-(beta-32P)-pyrophosphate labelled molecule.