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Biomedical subjects

D O Cliver

Publications and source records attributed to D O Cliver.

At least 19 recordsLinked to original sources
PubMed1995

Effects of temperature and host cell growth phase on replication of F-specific RNA coliphage Q beta.

Human enteric viruses have been found in groundwater in the absence of fecal coliforms. Because detection of human enteric viruses is costly, time-consuming, and lacking in sensitivity, F-specific RNA (FRNA) coliphages, which infect Escherichia coli by attachment to F pili, are being examined for suitability as indicators of human enteric viruses in groundwater. Temperatures and host cell growth conditions that constrain F-pilus expression will limit FRNA coliphage replication in groundwater and wastewater, as is desirable in an indicator. Below 25 degrees C F-pilus synthesis ceases; FRNA coliphage Qbeta did not replicate below this temperature in batch cultures. One-step replication studies indicated that the replicative cycle is prolonged and that fewer progeny are released as the temperature decreases. The decreases in phage replication observed in the one-step replication studies were a consequence of fewer cells infected as the temperature was lowered or as host cells entered stationary phase. The numbers of phage particles released from infected cells did not change. The minimum temperature for replication of Qbeta, 25 degrees C, is not maintained in wastewater and does not occur in Wisconsin groundwater. On the basis of temperature and host cell growth phase, we have concluded that extensive replication of FRNA coliphages does not occur in wastewater and groundwater in Wisconsin and areas with similar cool climates.

Anaerobiosis↗
PubMed1992

Inactivation of poliovirus type 1 in mixed human and swine wastes and by bacteria from swine manure.

The persistence of poliovirus type 1 (PO1) in mixed septic tank effluent and swine manure slurry was determined, and the antiviral effects of several bacterial cultures isolated from swine manure slurry were demonstrated. In two field experiments, PO1 was consistently inactivated more rapidly in the mixed waste than in the control Dulbecco's phosphate-buffered saline (D-PBS). D values (time [in days] for a 90% reduction of virus titer) were 18.7 and 29.9 for the mixed waste and 56.5 and 51.8 for the D-PBS control, respectively. The virus inactivation in the mixed waste was temperature dependent. A comparison of PO1 inactivation in raw mixed waste, autoclaved mixed waste, and bacterium-free filtrate of raw mixed waste at the same pH and temperatures provided an initial demonstration that the virus inactivation in the mixed waste is related, at least in part, to microbial activity. At 25 degrees C, the D value was 6.8 for the mixed waste, 11.2 for the autoclaved mixed waste, and 10.5 for the bacterium-free filtrate of raw mixed waste. At 37 degrees C, D values were 1.3, 3.9, and 3.1 for these three suspending media, respectively. Three bacterial isolates which had shown antiviral effects in a screening test each caused virus inactivation in autoclaved mixed waste, in which the effect of other microorganisms was excluded. Inhibition of PO1 inactivation by protease inhibitors suggests that the virus inactivation in the mixed waste was due in part to proteolytic enzymes produced by bacteria in the waste.

Animals↗
PubMed1992

Degradation of Giardia lamblia cysts in mixed human and swine wastes.

This study was conducted to determine the persistence of Giardia lamblia cysts in mixed septic tank effluent and swine manure slurry and to correlate fluorescein diacetate-propidium iodide staining of G. lamblia cysts with their morphology under low-voltage scanning electron microscopy. Under field conditions, G. lamblia cysts were degraded more rapidly in the mixed waste than in the control Dulbecco's phosphate-buffered saline (PBS). For total and viable cysts, the mixed waste had D values (time for a 90% reduction in number of cysts) of 18.3 and 15.5 days, and the Dulbecco's PBS control had D values of 41.6 and 26.8 days. The rates of cyst degradation in septic tank effluent and in Dulbecco's PBS were similar. Increasing the proportion of swine manure slurry in the mixed waste favored degradation of the parasite. These results indicate that the mixed waste treatment was the predominant factor affecting the cyst persistence and that it was swine manure slurry that played the role of degrading the parasite. Visualization of viable and nonviable Giardia cysts with low-voltage scanning electron microscopy revealed an excellent correlation between the viability of the cysts determined by fluorescein diacetate-propidium iodide staining and their electron microscopic morphology.

Animals↗
PubMed1988

Coxsackievirus-cell interactions that initiate infection in porcine ileal explants.

Coxsackievirus B5 (CB5) labeled with tritiated uridine was used to trace the interaction of the virus with explant cultures of porcine ileum. Similarly labeled human poliovirus 1 (PO 1), which is not specifically retained by porcine tissue, was used as a control. The explant procedure employed could maintain ileal tissue in a differentiated state for up to 48 hours. Porcine ileum was acquired from both young (4-6 week-old) and adult (9-11 month-old) animals. Inoculated explants of either absorptive or lymphoid tissue were incubated at temperatures selected to permit either viral adsorption or penetration and elution to occur. Retention of radioactive virus was quantitated by liquid scintillation counting and localized by autoradiography. Only in absorptive tissue explants from young animals did adsorption of CB5 at 6 degrees C exceed penetration at 37 degrees C. This suggested that incubation at 6 degrees C may not be an appropriate condition for studying enterovirus adsorption in explants. CB5 penetrated most efficiently into lymphoid tissue explants from young animals, indicating that these tissues could discriminate between CB5 and PO 1. In explants from adults, CB5 penetrated equally well into lymphoid and absorptive tissues. Virus penetrated into the absorptive epithelial cells and, possibly, the lamina propria near the villous tips. Low efficiency of penetration, and the non-critical function of these target cells, may help account for the characteristic lack of gastrointestinal symptoms in enterovirus infections.

Aging↗
PubMed1987

Enterovirus replication in porcine ileal explants.

Organ explants of porcine ileum were cultured in different media for up to 48 h. Tissue preservation was evaluated by light microscopy and by transmission and scanning electron microscopy. Cellular structure was well maintained after incubation for 48 h in CMRL-1066 supplemented with insulin and cortisone. Explants of absorptive or lymphoid tissue from young or adult pigs were incubated with either coxsackievirus B5 (which is infectious for swine) or human poliovirus type 1 (which served as a control) for 24 h at 37 degrees C. Progeny virus was detected by plaque assay. Replication was most evident in the absorptive tissue explants from young pigs. In tissues from adults, replication occurred equally well in absorptive and lymphoid tissues. Infection in explants was inefficient, and the yield of progeny virus was low.

Animals↗
PubMed1986

Enumeration of enterovirus particles by scanning electron microscopy.

Enumeration of virus particles requires relatively concentrated and uniformly dispersed virus preparations, which is difficult to achieve by the usual methods of negative staining and transmission electron microscopy. We have developed an electrophoretic method that concentrates enteroviruses onto a polycarbonate membrane for examination by high-resolution scanning electron microscopy. The electrophoretic apparatus comprises three chambers in electrical series, each containing 3.5 ml of dilute buffer. The center chamber is inoculated with virus. A 15-nm porosity membrane, which does not pass virus, separates the center from the side chambers. A constant current is applied, and chilled buffer is pumped past the electrodes for 2 h. The virus suspension is recovered, and changes in titer (or radioactivity if labeled virus is used) due to electrophoresis are measured. Buffer pH, relative to the viral isoelectric points, determines the direction of virus migration. Particle counts are calculated from the mean of 25 randomly chosen fields photographed at 35-60,000 X magnification and related to titers measured by plaque assay.

Electrophoresis↗
PubMed1984

A broad-spectrum enzyme-linked immunosorbent assay for the detection of human enteric viruses.

An enzyme-linked immunosorbent assay (ELISA) test has been devised for detection of a broad spectrum of human enteric viruses, based on the use of poly-L-lysine as a nonspecific adhesive to hold the virus particles in the test wells and of pooled human immune serum globulin to mark the virus for detection with commercial goat anti-human IgG antibody conjugated with horseradish peroxidase. Detection of five human enteroviruses and a reovirus at levels of 10 most probable number of cytopathogenic units (MPNCU) to 10 PFU per well was achieved, whereas no reaction was seen with two porcine enteroviruses. When two virus types are present in a sample, the ELISA reactions augment each other.

Antigens, Viral↗
PubMed1984

Septage treatments to reduce the numbers of bacteria and polioviruses.

Disposal of the pumped contents of septic tanks (septage) represents a possible means of dissemination of enteric pathogens including viruses, since persistence of enteroviruses in septic tank sludge for greater than 100 days has been demonstrated. The risk of exposure to potentially infectious agents can be reduced by disinfecting septages before their disposal. Of the septage disinfectants examined (technical and analytical grade glutaraldehyde, hydrogen peroxide, heat treatments, and a combination of heat and hydrogen peroxide), the treatment including hydrogen peroxide (5 mg, plus 0.33 mg of trichloroacetic acid, per ml of septage) and 55 degrees C killed virtually all the bacteria in septage within 1 h, whereas 55 degrees C alone inactivated inoculated polioviruses within 30 min. Virus was the most sensitive to heat, whereas fecal coliforms appeared to be the most sensitive to all chemical treatments. The responses of fecal streptococci and virus to both grades of glutaraldehyde (each at 1 mg/ml) were similar. Virus was more resistant than either fecal streptococci or total bacteria to low concentrations of hydrogen peroxide (1 to 5 mg/ml); however, virus and fecal streptococci were more labile than total bacteria to the highest peroxide concentration (10 mg/ml) examined. It is possible that the treatment combining heat and hydrogen peroxide was the most effective in reducing the concentrations of all bacteria, because catalase and peroxidases as well as other enzymes were heat inactivated, although catalase seems the most likely cause of damage. However, this most effective treatment does not appear to be practical for on-site use as performed, so further work on septage disinfection is recommended.

Bacteria↗
PubMed1983

Membrane filter evaluations using poliovirus.

Poliovirus 1, with and without radioactive label, has been used to evaluate membrane filters for several purposes in virology. Filters of several polymers will pass virus and retain larger particles. Cellulose nitrate can adsorb virus for concentration purposes. Polycarbonate does not adsorb virus and is well suited to particle sizing.

Evaluation Studies as Topic↗
PubMed1983

Attachment of Salmonella to mammalian cells in vitro.

The attachment of Salmonella typhimurium strain PHL67342 to several mammalian tissue culture cell lines was investigated. Strain PHL67342 failed to attach in significant numbers to the Buffalo green monkey (BGM), swine testicular (ST), and HeLa cell lines. Significant attachment was observed with the Henle intestinal cell line. Log-phase cells of strain PHL67342 attached in greatest numbers to the Henle cells after 45 min of incubation at 37 degrees C. Attachment to the Henle cells was not affected by D-mannose or D-galactose, but was markedly inhibited by high concentrations of alpha-methyl-D-mannoside. Also, Salmonella lipopolysaccharide had no effect on the attachment of strain PHL67342 to the Henle cells. Fimbriae were not detected on the bacterial cells used in the adherence experiments. These results suggest that some bacterial factor(s) other than fimbriae and lipopolysaccharide mediate the attachment of strain PHL67342 to the Henle cells.

Adhesiveness↗
PubMed1982

Emetic action of staphylococcal enterotoxin A on weanling pigs.

Peroral and intraduodenal administration of staphylococcal enterotoxin A (SEA) to weanling pigs elicited an emetic response. Peroral administration of an emetic dose of SEA resulted in a single emetic episode occurring 90 to 180 min after dosing. Intraduodenal administration via a surgically implanted catheter of 100 or 150 micrograms of SEA resulted in multiple emetic episodes occurring 150 to 210 min after dosing, suggesting an intestinal site of action for SEA. The 50% emetic dose for perorally administered SEA was between 40 and 50 micrograms for 4.1- to 9.1-kg weanling pigs and 20 micrograms in 0.9- to 2.3-kg weanling pigs. Neurobehavioral responses, including alternating periods of drowsiness and restlessness, staggering, temporary loss of the righting reflex, and refusal to feed were also observed in pigs given an oral dose of SEA. Based on the demonstrated responsiveness of pigs to SEA, pigs should be considered suitable animal models for studies on the sites and mode of action of the toxin.

Administration, Oral↗
PubMed1978

Inactivation of enteroviruses by ascorbic acid and sodium bisulfite.

Poliovirus type 1, coxsackievirus type A9, and echovirus type 7 were inactivated by sodium bisulfite and ascorbic acid. Inactivation rates depended upon concentration, temperature, and pH. RNA infectivity was lost during inactivation; the capsid was also altered by these inactivating agents, as determined by enzyme sensitivity assays and by tests of adsorption to cells. Structural modifications of the virus particles were not identical, suggesting that the mechanism of inactivation by ascorbic acid differs from that of sodium bisulfite.

Ascorbic Acid↗
PubMed1976

Effect of acid pH, salts, and temperature on the infectivity and physical integrity of enteroviruses.

At 2 degrees and 30 degrees C, enteroviruses are more stable on the acid than on the alkaline side of neutrality. In the range from pH 3 to 9, temperature is so influential that the fastest inactivation rate at 2 degrees C is slower than the slowest inactivation rate at 30 degrees C. Specific ions or salts also affect the rate of inactivation of enteroviruses. NaCl and other chloride salts enhance the inactivation of poliovirus at pH 3. NaCl is considerably less effective against poliovirus in the range of pH 4.5 to 7.0 than at pH less than 4.5. Loss of RNA infectivity of the virus particle proceeds as rapidly as the loss of infectivity of the particle itself, except at pH 3 in the presence of MgCl2. Inactivation results in alterations to the physical integrity of enteroviruses. At pH 5 and 7, RNA hydrolysis of poliovirus particles occurs; and at pH3, 5,6, and 7 the nucleic acid becomes susceptible to ribonuclease. Only virus particles inactivated at pH 3 show a sensitivity to chymotrypsin. The hemagglutinins of echovirus type 7 are destroyed during inactivation at pH 3,4,5, and 6; but at pH 6 this alteration precedes the loss of infectivity. The pH of the suspension is a primary determinant of the mechanism of virus destruction and possibly of the loss of infectivity at these temperatures.

Cell Line↗
PubMed1975

Virus association with wastewater solids.

The solids produced at an urban wastewater treatment plant, tested semiquantitatively, almost all contained human intestinal viruses. Reoviruses and five or more types of enteroviruses were present. Sludge, digested anaerobically at 30 degrees-32 degrees C, and grit contained measurable levels of viruses. Until reliable means of inactivating the viruses have been developed and implemented, great care should be taken in disposing of these solids.

Enterovirus↗
PubMed1973

Filtration methods for recovering enteroviruses from foods.

A set of simple, rapid filtration methods has been developed for recovering enteroviruses from foods. The methods employ quite inexpensive equipment, and the resulting extract might be concentrated in any of several ways. Three model enteroviruses were apparently recovered with at least 80% efficiency when inoculated, at levels ranging from thousands to one plaque-forming unit per 20-g sample, into eleven foods representing six major food groups.

Animals↗