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PubMed2003

RNA-binding protein-mediated translational repression of transgene expression in plants.

We have demonstrated that RNA-binding proteins from coliphages and yeast can function as translational repressors in plants. RNA sequences called translational operators were inserted at a cap-proximal position in the 5'-UTR of mRNAs of two reporter genes, gus or aroA:CP4. Translation of the reporter mRNAs was efficiently repressed when the RNA binding protein that specifically binds to its cognate operator was co-expressed. The efficiency of translational repression by RNA-binding protein positively correlated with the amount of binding protein in transformed plant cells. Detailed studies on coliphage MS2 coat protein-mediated translational repression also suggested that the efficiency of translational repression was position-dependent. A translational operator situated at the cap-proximal position was more efficient in conferring repression than one that was placed cap-distal. Translational repression can be an efficient means for regulation of transgene expression, thereby broadening current approaches for transgene regulation in plants.

3-Phosphoshikimate 1-Carboxyvinyltransferase↗
PubMed2000

Determination of enteroviruses, hepatitis A virus, bacteriophages and Escherichia coli in Adriatic Sea mussels.

The aim of the present study was to evaluate the incidence of enteric viruses in mussels and to verify the possibility of using phages as indirect indicators of mussel viral contamination. Mussels (36 samples) collected from three different areas of the Adriatic Sea were analysed to determine the following parameters: Escherichia coli, somatic coliphage (T6 phage), F-Plus (MS2 phage), B40-8 (phage of Bacteroides fragilis), enteroviruses and hepatitis A virus. Most of the results of the bacteriological analysis (most probable number (MPN) ml-1) were in accordance with the bacteriological limits established by European law, with the exception of seven samples. The bacteriophage analyses were always negative for F-Plus and B40-8, with the exception of a few samples, whereas the somatic coliphages were generally between 0 and 20 MPN g-1, with the exception of two samples (110 MPN g-1). The virological analysis showed five samples positive for the presence of enteroviruses and 13 for the presence of hepatitis A virus (in three samples both viruses were present). Most of these samples presented acceptable bacteriological parameters and the bacteriophages were absent or their value was generally very low. The results show that the detection of E. coli and phages does not seem to be a good indicator of viral contamination.

Animals↗
PubMed2002

Removal and inactivation of indicator bacteriophages in fresh waters.

AIMS: The removal and inactivation of faecal coliform (FC) bacteria, enterococci (ENT), sulphite-reducing clostridia (SRC), somatic coliphages, F-specific RNA bacteriophages and bacteriophages infecting Bacteroides fragilis in fresh waters. METHODS AND RESULTS: Removal was studied in two areas of a river. The results showed different removal of each group of microbes. Faecal coliform bacteria were removed faster than any other, whereas SRC and bacteriophages infecting Bact. fragilis were the most persistent. Inactivation was measured by 'in situ' experiments, which showed significant differences in survival of the different groups of bacterial and bacteriophage indicators. The SRC and bacteriophages were more resistant than faecal coliforms and enterococci, with the exception of F-specific RNA bacteriophages in the summer. Inactivation experiments with pure cultures of bacteriophages confirmed that phage B40-8 of Bact. fragilis was the most resistant. CONCLUSIONS: Bacteria and bacteriophages show different resistance to natural inactivation. The use of phages allows information to be obtained in addition to that provided by bacterial indicators. Somatic coliphages and phages infecting Bact. fragilis might supply that indicator function. SIGNIFICANCE AND IMPACT OF THE STUDY: Confirmation was obtained that bacteriophages provided additional information to that provided by bacterial indicators to monitor the natural inactivation of viruses and/or pathogens.

Bacteria↗
PubMed2001

Bacteriological and virological quality of seawater bathing areas along the Tyrrhenian coast.

Monitoring was carried out during summer 1997 along a selected area of the Tyrrhenian coast near the Tiber river mouth. Fifty-eight seawater samples, collected from 19 stations, were examined for coliforms, streptococci, Enteroviruses, Salmonellae, coliphages, Bacteroides fragilis phages, Pseudomonas, alophilic Vibrios, Aeromonas and yeasts. Salmonellae and coliphages were isolated in 3 and 12 out of 58 samples, respectively. Enteroviruses and Bacteroides fragilis phages were not isolated. Reoviruses were isolated only from 2 out of 58 samples. A limited number of samples of the northern stations located near the Tiber and other river mouths exceeded the guide values for bathing water by the EU Directive. All the southern stations, located near canals, were of very good microbiological quality. Pseudomonas, Vibrio, Aeromonas and yeasts were isolated from all stations and their values in 100 ml of seawater were 10-10(6), 10-10(6), 0-10(6) and 1-10(3), respectively. An extensive disinfection practice carried out on domestic wastes, which are discharged in rivers and canals, probably brought pollution levels of most stations to values within the bacterial standards. The spread of Pseudomonas, Aeromonas, etc. showed that all the coastal area studied was characterized by the presence of organic matter coming from land that can support the presence of opportunistic pathogens and other microbial flora.

Bathing Beaches↗
PubMed2001

Chemical and microbial characterization of household graywater.

In arid areas, the search for efficient methods to conserve water is of paramount importance. One of the methods of water conservation available today is graywater recycling--the reuse of water from the sinks, showers, washing machine, and dishwasher in a home. The purpose of this project was to characterize the chemical and microbial quality of graywater from a single-family home with two adults. Water samples from a graywater holding tank were analyzed over a seven-month period for total coliforms, fecal coliforms, fecal streptococci, Staphylococcus aureus (S. aureus), Pseudomonas aeruginosa (P. aeruginosa), and coliphages. The pH, turbidity, biological oxygen demand (BOD), suspended solids (SS), electrical conductivity (EC), sulfates (SO4), and chlorides (Cl) were also measured. The mean numbers of total coliforms, fecal coliforms, fecal streptococci, and P. aeruginosa were 8.03 x 10(7), 5.63 x 10(5), 2.38 x 10(2), and 1.99 x 10(4) CFU/100 mL, respectively. S. aureus and coliphages were not detected. In the chemical analysis, mean values of 7.47 for pH, 43 nephelometric turbidity units (NTU) for turbidity, 64.85 mg/L for BOD, 35.09 mg/L for SS, 0.43 mS/cm for EC, 59.59 mg/L for SO4, and 20.54 mg/L for Cl were measured. These data were compared to data taken in 1986 and 1987, when two adults and one child lived in the household. Analysis showed no statistically significant difference in levels of total coliforms and suspended solids between the two data sets. There were statistically significant differences in levels of fecal coliforms, pH, turbidity, chlorides, sulfates, and BOD between the two households. Fecal coliforms, turbidity, and BOD were higher in the household with two adults and one child. Levels of Cl, SO4, and pH were higher in the household with two adults.

Adult↗
PubMed1985

Virus persistence in groundwater.

More than 50% of the outbreaks of waterborne disease in the United States are due to the consumption of contaminated groundwater. An estimated 65% of the cases in these outbreaks are caused by enteric viruses. Little, however, is known about the persistence of viruses in groundwater. The purpose of this study was to determine whether measurable chemical and physical factors correlate with virus survival in groundwater. Groundwater samples were obtained from 11 sites throughout the United States. Water temperature was measured at the time of collection. Several physical and chemical characteristics, including pH, nitrates, turbidity, and hardness, were determined for each sample. Separate water samples were inoculated with each of three viruses (poliovirus 1, echovirus 1, and MS-2 coliphage) and incubated at the in situ groundwater temperature; selected samples were also incubated at other temperatures. Assays were performed at predetermined intervals over a 30-day period to determine the number of infective viruses remaining. Multiple regression analysis revealed that temperature was the only variable significantly correlated with the decay rates of all three viruses. No significant differences were found among the decay rates of the three viruses, an indication that MS-2 coliphage might be used as a model of animal virus survival in groundwater.

Bacteriophages↗
PubMed1985

Effect of an activated sludge wastewater treatment plant on ambient air densities of aerosols containing bacteria and viruses.

Bacteria- and virus-containing aerosols were studied during the late summer and fall seasons in a midwestern suburb of the United States before and during the start-up and operation of an unenclosed activated sludge wastewater treatment plant. The study showed that the air in this suburban area contained low-level densities of indicator microorganisms. After the plant began operating, the densities of total aerobic bacteria-containing particles, standard plate count bacteria, total coliforms, fecal coliforms, fecal streptococci, and coliphages increased significantly in the air within the perimeter of the plant. Before plant operations, bacteria were detected from five genera, Klebsiella, Enterobacter, Serratia, Salmonella, and Aeromonas. During plant operations, the number of genera identified increased to 11. In addition to those genera found before plant operations, Escherichia, Providencia, Citrobacter, Acinetobacter, Pasteurella, and Proteus, were also identified. Enteric viruses were detected in low densities from the air emissions of this plant. Only standard plate count bacteria remained at significantly higher than base-line densities beyond 250 m downwind from the center of the aeration tanks. Fecal streptococci and coliphages appeared to be more stable in aerosols than the other indicator microorganisms studied. In general, the densities of microorganism-containing aerosols were higher at night than during the day. The techniques used in this study may be employed to establish microorganism-containing aerosol exposure during epidemiological investigations.

Aerosols↗
PubMed1994

Cloning of the early promoters of Pseudomonas aeruginosa bacteriophage D3: sequence of the immunity region of D3.

The early promoters of bacteriophage D3 of Pseudomonas aeruginosa were cloned and physically mapped to the right 25% of the phage genome. The promoters were cloned into promoter selection vector pQF26, and their relative strengths, the direction of transcription, and whether they were directly regulated by repressor were determined. A 3.3-kb fragment of the genome containing the immunity region was sequenced and analyzed (GenBank accession number: L22692). The promoter activity associated with this region was determined to be bidirectional and repressible, indicating that this region contains operator-promoter complexes. Sequence and functional analyses suggest that this region is analogous to the immunity region of coliphage lambda. Two strong promoters, one of which was repressible, were found to be located adjacent to the immunity region. Clear-plaque mutant phage D3c contains insertion element IS222, which causes it to behave as a repressor-negative (c1) variant. The site of insertion of IS222 was sequenced and determined to lie within the c1 gene open reading frame. This phage shows remarkable similarity in genomic organization to coliphage lambda and its relatives.

Amino Acid Sequence↗
PubMed1999

Cloning and analysis of the capsid morphogenesis genes of Pseudomonas aeruginosa bacteriophage D3: another example of protein chain mail?

The terminal DNA restriction fragments (PstI-D and -B) of Pseudomonas aeruginosa bacteriophage D3 were ligated, cloned, and sequenced. Of the nine open reading frames in this 8.3-kb fragment, four were identified as encoding large-subunit terminase, portal, ClpP protease, and major head proteins. The portal and capsid proteins showed significant homology with proteins of the lambdoid coliphage HK97. Phage D3 was purified by CsCl equilibrium gradient centrifugation (rho = 1.533 g/ml), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed six proteins with molecular masses of 186, 91, 79, 70, 45, and 32 kDa. The pattern was unusual, since a major band corresponding to the expected head protein (43 kDa) was missing and a significant amount of the protein was retained in the stacking gel. The amino terminus of the 186-kDa protein was sequenced, revealing that the D3 head is composed of cross-linked 31-kDa protein subunits, resulting from the proteolysis of the 43-kDa precursor. This is identical to the situation observed with coliphage HK97.

Amino Acid Sequence↗
PubMed2003

Genome sequences of two closely related Vibrio parahaemolyticus phages, VP16T and VP16C.

Two bacteriophages of an environmental isolate of Vibrio parahaemolyticus were isolated and sequenced. The VP16T and VP16C phages were separated from a mixed lysate based on plaque morphology and exhibit 73 to 88% sequence identity over about 80% of their genomes. Only about 25% of their predicted open reading frames are similar to genes with known functions in the GenBank database. Both phages have cos sites and open reading frames encoding proteins closely related to coliphage lambda's terminase protein (the large subunit). Like in coliphage lambda and other siphophages, a large operon in each phage appears to encode proteins involved in DNA packaging and capsid assembly and presumably in host lysis; we refer to this as the structural operon. In addition, both phages have open reading frames closely related to genes encoding DNA polymerase and helicase proteins. Both phages also encode several putative transcription regulators, an apparent polypeptide deformylase, and a protein related to a virulence-associated protein, VapE, of Dichelobacter nodosus. Despite the similarity of the proteins and genome organization, each of the phages also encodes a few proteins not encoded by the other. We did not identify genes closely related to genes encoding integrase proteins belonging to either the tyrosine or serine recombinase family, and we have no evidence so far that these phages can lysogenize the V. parahaemolyticus strain 16 host. Surprisingly for active lytic viruses, the two phages have a codon usage that is very different than that of the host, suggesting the possibility that they may be relative newcomers to growth in V. parahaemolyticus. The DNA sequences should allow us to characterize the lifestyles of VP16T and VP16C and the interactions between these phages and their host at the molecular level, as well as their relationships to other marine and nonmarine phages.

Bacteriophages↗
PubMed1975

Molecular weight of DNA from actinophage MSP2.

Actinophage MSP2 is infectious for Streptomyces venezuelae S13. Based upon electron microscopy of coliphage T4 mixed with MSP2, MSP2 had a head about 48 +/- 2 nm wide and 87 +/- 5 nm long. DNA from polyoma virus and from coliphages T4 and T7 served as reference markers in estimating the molecular weight of MSP2 DNA from sedimentation in sucrose gradients. Denatured MSP2 DNA was estimated to be about 17 x 10(6) and double-stranded MSP2 DNA was about (36 +/- 1.6) x 10(6) in molecular weight.

Bacteriophages↗
PubMed1984

Morphology of Pasteurella multocida bacteriophages.

Twenty-one tailed phages with icosahedral heads belong to the Myoviridae, Siphoviridae, and Podoviridae families and to four morphological types. Type AU, with 10 phages, has a contractile tail and is morphologically identical with coliphage P2. Lysates contain contracted tail sheaths assembled end-to-end and abnormal structures with long tails and multiple tail sheaths. Types C-2 and 32, with one and three phages, respectively, have long, noncontractile tails. Type 22 includes seven phages, has a short tail, and resembles coliphage T7. Our results agree with previous biological data and suggest that types AU, C-2, 32, and 22 correspond to four different phage species.

Bacteriophages↗
PubMedDate not supplied

Microbiological indicators of water quality in the Xochimilco canals, Mexico City.

OBJECTIVE: To quantify microbiology indicators of fecal contamination in the effluents of two waste water treatment plants and in samples collected in several canals in Xochimilco. MATERIAL AND METHODS: A cross sectional study was performed. Ten sites, 5 from plant effluents and 5 from canals, were selected for sampling during November and December 2001. Fecal coliforms and enterococci were quantified by membrane filtration, male specific (F+) and somatic coliphages by double agar layer technique, and Cryptosporidium oocysts and Giardia cysts by concentration with Envirocheck filter followed by immunofluorescence microscopy quantification. The average of organisms counts from effluents and canal water were compared with t Student test. RESULTS: Treated water discharge in canals showed a low count of Fecal Coliforms (average 40.4/100 ml), enterococci (average 58.8/100 ml) and Cryptosporidium oocysts (average 13.2/100 l), while coliphages and Giardia cyst rendered higher counts (average 1467.5/100 ml and 1199.8/100 l, respectively) suggesting the water treatment methods could fail to remove these agents. A significant lower count of Giardia cysts (average 45/100 l) and no Cryptosporidium oocysts were found in irrigation canals, which suggests a natural clearance of these pathogens. Strains of Escherichia coli isolated in one of the canals contaminated with sewage had antimicrobial multi-resistance that was transferred by conjugation suggesting that resistance is encoded in a plasmid potentially transferable to other pathogenic bacteria. CONCLUSIONS: Cost effective and culturally acceptable waste treatment methods will require careful planning and consultation if they are to be adopted and mantained by local populations.

Mexico↗
PubMed2006

UV disinfection of wastewater effluents for unrestricted irrigation.

Wastewater reuse in arid regions is important for the production of a water resource to be utilised for non-potable purposes and to prevent the environmental transmission of disease-causing agents. This study was conducted to evaluate the effect of water quality on the comparative disinfection efficiency of viruses, bacteria and spores by UV irradiation. Furthermore, the microbial quality of effluent produced by coagulation, high rate filtration (HRF) and either UV irradiation or chlorination was determined. Using low pressure collimated beam, a UV dose of 80 mWs/cm2 was needed to achieve a 3-log10 inactivation of either rotavirus SA-11 or coliphage MS2, whereas over 5-log10 inactivation of E. coli was reached with a dose of only 20 mWs/cm2. B. subtilis inactivation was found to be linear up to a dose of 40 mWs/cm2 and then a tailing up to a UV dose of 120 mWs/cm2 was observed. It is worth noting that effluent turbidity of < 5 NTU did not influence the inactivation efficiency of UV irradiation. Operation of a pilot plant to treat secondary effluent by coagulation, HRF and UV disinfection at a UV dose of 80 mWs/cm2 resulted in the production of high quality effluent in compliance with the Israel standards for unrestricted irrigation (< 10 CFU/100 mL faecal coliform and turbidity of < 5 NTU). Sulphite reducing clostridia (SRC) were found to be more resistant than coliphages and F coliform for UV irradiation. The results of this study indicated that UV disinfection is suitable for the production of effluents for unrestricted irrigation of food crops.

Bacteria↗
PubMedDate not supplied

Reduction of enteric microorganisms at the Upper Occoquan Sewage Authority Water Reclamation Plant.

The Upper Occoquan Sewage Authority (UOSA) Water Reclamation Plant, Centreville, Virginia, is a state-of-the-art wastewater treatment plant that was created to treat area wastewater and provide protection for the Occoquan Reservoir. This study investigated UOSA's unit processes as barriers to pathogenic as well as altemative and traditional-indicator microorganisms. Samples were collected once a month for 1 year from eight sites within UOSA's advanced wastewater reclamation plant. The eight sites were monitored for indicator bacteria total and fecal coliforms, enterococci, Clostridium, coliphage (the virus that infects Escherichia coli), human enteroviruses, and enteric protozoa. Overall, the plant was able to achieve a 5- to 7-log10 reduction of bacteria, 5-log10 reduction of enteroviruses, 4-log10 reduction for Clostridium, and 4.6-log10 reduction of protozoa. Total coliforms, enterococci, Clostridium, coliphage, Cryptosporidium, and Giardia were all detected in four or fewer samples of the final effluent. No enteroviruses or fecal coliforms were detected in the final effluent. The microbiological quality of reclaimed water and the reservoir water were compared. In every case, the treated wastewater was of a better quality than the ambient water in the reservoir, thus indicating that the reclaimed water will not adversely affect the water quality for downstream users.

Animals↗
PubMed2001

Removal of bacterial indicators and pathogens from dairy wastewater by a multi-component treatment system.

Microbial removal by a multi-component treatment system for dairy and municipal wastewater is being studied in Arizona, USA. The system consists of paired solids separators, anaerobic lagoons, aerobic ponds and constructed wetlands cells. The organisms under study include: total coliform, fecal coliform, enterovirus, Listeria monocytogenes, Clostridium perfringens, coliphage, Giardia lamblia and Cryptosporidium parvum. Organism removal rates from dairy wastewater varied from 13.2 per cent for fecal coliform to 94.9 per cent for coliphage. It appears that the much higher turbidity of the dairy wastewater, nearly 1,300 NTU, decreased the treatment systems' ability to remove some microbial indicators and pathogens. Information from this study can be used to determine the adequacy of multi-component treatment systems for the control of wastewater-borne pathogens, both in municipal treatment systems as well as in confined animal feeding operations (CAFO). This information also can assist municipalities and the CAFO industry in the implementation of rational and efficient treatment strategies for appropriate reuse of wastewaters.

Agriculture↗
PubMed2001

Occurrence and die-off of indicator organisms in the sediment in two constructed wetlands.

The interest in constructed wetlands for municipal wastewater and stormwater treatment has recently increased but data for the reduction efficiency of indicator organisms are often restricted to the water phase. In a full-scale wastewater wetland in Sweden fecal coliforms and enterococci were reduced by 97-99.9% and coliphages by approximately 70%. The factors affecting the reduction are however less well understood. In two full-scale wetlands, for stormwater and wastewater treatment, an assessment has been done of the particle associated fraction of indicator organisms. No significant differences in the particle-associated numbers were seen between the inlet and the outlet of the wetlands, but the amounts of sedimenting particles varied between the two sites. In the stormwater wetland the amount of sedimenting particles at the outlet was 3% of the amount at the inlet, while the wastewater wetland had much lower particle removal efficiency. The reduction of suspended particles seems to be the main factor for bacterial elimination from the water phase, governed by vegetation and design. In the sediment, survival of presumptive E. coli, fecal enterococci, Clostridium and coliphages were long with T50-values of 27, 27, 252 and 370 days, respectively. The organisms can however be reintroduced by resuspension. Viruses in the water phase may be of main concern for a risk assessment of receiving waters.

Ecosystem↗
PubMed2001

River water quality improvement by natural and constructed wetland systems in the tropical semi-arid region of northeastern Brazil.

The efficiencies of a natural Typha spp wetland (Wn) formed on a river bed and its effluent treatment in a constructed wetland (Wc, subsurface horizontal flow) were investigated in northeastern Brazil (Paraiba State). The Wc system (12 tanks with stone gravel, 4.13 m2, 0.22 m3, 20 Typha spp rhizomes, m(-2) each, with 38, 29, and 19 mm x d(-1) hydraulic loadings, and 5, 7, and 10 days HRT) was fed daily with effluent from a Wn. Wn removal presented the highest values after Typha spp were cut during the 5th week. Removal values were (1st and 2nd periods or before and after cutting): 75% and 81% BOD5; 10-53% total phosphorus; 13%-55% ammonia; 89%-91% FC; 90-96% coliphages and bacteriophages. Wc removals increased with time with best results on 10 d HRT. Removals were also higher in the 2nd period: 74%-78% BOD5; 58%-82% ammonia; 90% FC; 94-98% FS; and 92%-96% coliphages and bacteriophages. Despite the high remaining values of FC (1.4 x 10(4) CFU/100 ml) and FX (4 x 10(3) CFU/100 ml), the removals were satisfactory and HRT dependent, suggesting a gradual optimization of the system with time. The Wc exhibited good efficiency for improving water quality from polluted river.

Brazil↗