Recovery of viruses from field samples of raw, digested, and lagoon-dried sludges.

In a 22-month study, viruses were detected in 84% (62/74) of raw, 53% (19/36) of anaerobically digested, and 39% (11/28) of lagoon-dried sludge samples. Lagoon sludge contained detectable viruses (reovirus and enterovirus groups) even after 8 months of retention. Because of such prolonged virus survival in sludge, care must be taken in its disposal or utilization.

Viral pollution of surface waters due to chlorinated primary effluents.

The role of chlorinated primary effluents in viral pollution of the Ottawa River (Ontario) was assessed by examining 282 field samples of wastewaters from two different sewage treatment plants over a 2-year period. The talc-Celite technique was used for sample concentration, and BS-C-1 cells were employed for virus detection. Viruses were detected in 80% (75/94) of raw sewage, 72% (68/94) of primary effluent, and 56% (53/94) of chlorinated effluent samples. Both raw sewage and primary effluent samples contained about 100 viral infective units (VIU) per 100 ml. Chlorination produced a 10- to 50-fold reduction in VIU and gave nearly 2.7 VIU/100 ml of chlorinated primary effluent. With a combined daily chlorinated primary effluent output of approximately 3.7 x 10(8) liters, these two plants were discharging 1.0 x 10(10) VIU per day. Because the river has a mean annual flow of 8.0 x 10(10) liters per day, these two sources alone produced a virus loading of 1.0 VIU/8 liters of the river water. This river also receives at least 9.0 x 10(7) liters of raw sewage per day and undetermined but substantial amounts of storm waters and agricultural wastes. It is used for recreation and acts as a source of potable water for some 6.0 x 10(5) people. In view of the potential of water for disease transmission, discharge of such wastes into the water environment needs to be minimized.

Exogenous bacterial contamination of donor blood.

The Canadian Red Cross blood transfusion service has followed a set protocol for phlebotomy and collection of a unit of blood. Recent requirements for automated testing have necessitated that a second tube of blood be obtained from the blood line following collection of the unit. Evaluation of the techniques used, however, has indicated the possibility of bacterial contamination from the skin of donors, from insertion of the needle through an unsterile rubber stopper, and through backflow from a nonsterile vacuum tube. To test these possibilities swabs were taken from skin and stoppers of vacuum tubes. Further, vacuum tubes were deliberately contaminated with Escherichia coli. The normal sampling procedure, which involves stripping the donor line to refill and mix the blood, was then followed. This resulted in contamination of the segments and even the blood bag. These findings led to modification of the standard bleeding technique, whereby stripping was eliminated and sterile vacuum tubes were to be used at all times.

Cancer-associated serum galactosyltransferase activity. Demonstration in an animal model system.

Two different lines of solid tumors were produced in outbred hamsters by subcutaneous injection of polyoma transformed BHK cells. Growth of the tumors correlated with the appearance in serum of an electrophoretically distinct peak of galactosyltransferase: NeuAc-, Gal-free fetuin acceptor activity on polyacrylamide gels. This slow moving peak of enzyme activity (GT-HH) was detected before solid tumors could be grossly observed and the amount of activity in this peak was also found to be linearly related with growth of the tumor. GT-IIH was not detectable in control animals and separated from a faster migrating major area of serum galactosyltransferase activity (GT-IH) found in sera of both control and tumor-bearing hamsters. These two activities were shown to maintain their respective mobilities on re-electrophoresis. Solubilized enzyme derived from excised tumors demonstrated an electrophoretic mobility on polyacrylamide gels identical to that for GT-IIH present in serum from tumor-bearing animals. In contrast, enzyme activity solubilized from livers of both control or tumor-bearing hamsters showed a mobility similar to that of the faster moving serum galactosyltransferase enzyme activity, i.e. GT-IH. In addition, medium derived from nonconfluent BHKpy cells in tissue culture contained galactosyltransferase activity which co-electrophoresed with the slower migrating characteristics of galactosyltransferase activities derived from serum (control and tumor-bearing), solid tumors, liver and BHKpy cells in tissue culture were compared. All kinetic properties were similar with the exception that the Km UDP-galactose of GT-IIH (1.0 X 10(-5) M) was half that of GT-IH (2.0 X 10(-5) M).

Isolation of apparently wild strains of poliovirus type 1 from sewage in the Ottawa area.

In the first 4 months of 1974, 140 gauze pad samples of sewage collected in the Ottawa area were analysed by the BS-C-1 cell system for the presence of viruses pathogenic for humans. Viruses were isolated from 111 (79%) of the samples. Of the 72 (65%) isolates identified by serology and electron microscopic examination, 56 (78%) were reoviruses and 16 (22%), enteroviruses. The enterovirus isolates included one coxsackievirus B4, one vaccine strain of poliovirus type 3, nine vaccine strains of poliovirus type 1 and five strains of poliovirus type 1 that proved by serodifferentiation and temperature marker tests to be different from vaccine strains. The fact that these strains were present in the community sewage in readily detectable concentrations at a time when immunity against polioviruses is declining in such communities is a cause for concern.

Comparison of talc-Celite and polyelectrolyte 60 in virus recovery from sewage: development of technique and experiments with poliovirus (type 1, Sabin)-contaminated multilitre samples.

For virus recovery from sewage, a mixture of talc and Celite was tested as a possible inexpensive substitute for polyelectrolyte 60 (PE 60). After adjustment of pH to 6 and the addition of 45-60 plaque forming units (PFU)/ml of poliovirus type I (Sabin) to the sewage sample under test, 100 ml of it was passed through either a PE 60 (400 mg) or a talc (300 mg)-Celite (100 mg) layer; the layer-adsorbed virus was eluted with 10 ml of 10% fetal calf serum (FCS) in saline (pH 7.2). In these experiments, PE 60 layers recovered 73-80% (mean 76%) of the input virus. In comparison, virus recoveries with the talc-Celite layers were 65-70% (mean 68%). Passage of 5 litres of raw sewage (containing 50 to 1.26 X 10(5) PFU/100 ml of the poliovirus) through the talc (15 g)-Celite (5 g) layers and virus elution with 50 ml of 10% FCS in saline gave virus recoveries of 33-63% (mean 49%). Except for pH adjustment and prefiltration through two layers of gauze to remove large solids, no other sample pretreatment was found to be necessary. Application of this technique to recovery of indigenous viruses from field samples of raw sewage and effluents has been highly satisfactory.

Comparison of four eluents in the recovery of indigenous viruses from raw sludge.

The efficiency of 3% casein hydrolysate (CH), 3% lactalbumin hydrolysate (LH), 3% beef extract (BE), and 10% fetal calf serum (FCS) was compared for the recovery of viruses from raw sludge. CH and LH proved to be inefficient and were eliminated from the study after initial testing. In tests with 20 different samples of raw sludge, beef extract eluted virus in 15 (75%) and FCS revealed virus in 19 (95%) of the samples using BS-C-1 cells. That different eluents were not eluting different viruses from the same sample was shown by the serologic and electron-microscopic examination of 43% (18/42) of the isolates. The identified viruses included members of the entero- (coxsackie B, and polio) and reo-virus groups.

An immunoprecipitin study of the incidence of influenza A antibodies in animal sera in the Ottawa area.

A survey of over 600 'normal' sera from 14 animal species by immunoprecipitin tests in cellulose acetate using viron antigens revealed a high incidence of precipitating activity against a broad range of influenza A virus strains, particularly A2hHong Kong/1/68 and /PR8. However, serum treatments trypsin-heat-periodate, NaIO4, V. cholerae receptor-destroying enzyme (RDE), or kaolin eliminated most precipitating activity, which suggests that it was due to "non-specific" inhibitors of influenze viruses. A resistant minority could not be identified as inhibitor or antibody on this basis. Precipitation of the influenza A major type-specific antigen in virus-soluble antigens by human 7S gamma globulin antibody (IgG), demonstrated to be specific for influenza virus, was established as a reference reaction to identify similar immunoprecipitin reactions occurring between virus-soluble antigens and normal or immune sera. Complement fixation tests provided supplementary evidence for the presence of influenza A antibodies in these sera. Influenza A antibodies were found in only a few sera of six animal species: cat, dog, rabbit, goat, chipmunk, and sheep. Thus the animal species examined in the Ottawa area have not revealed an unequivocal reservoir for human influenza A viruses.

Hospital-acquired infection: present and future impact and need for positive action.

Surveillance of infections at the Ottawa General Hospital between September 1, 1971 and August 31, 1972 showed that the overall infection rate was 13.5% of which 5.6% was community-acquired while 7.9% was of nosocomial origin. These figures are comparable to those for equivalent hospitals in the United States and lower than those reported from the Boston City Hospital, but they nevertheless indicate that over half the infected patients in the hospital were infected after admission. Urinary tract infections accounted for 44.8% of all nosocomial infections and clearly dominated the picture. The postoperative wound infection rate was 3.9% and accounted for only 18% of nosocomial infections. It is probable that these findings are representative of general hospitals throughout Canada and indicate conditions which will not long be tolerated. The knowledge and techniques exist for the prevention of all hospital cross-infection and much autogenous infection. Specific measures are suggested for working towards this goal. These are (1) the replacement of archaic hospitals and hospital facilities, (2) the establishment in every hospital of an efficient surveillance program, (3) the institution of good catheterization and catheter care techniques, and (4) the establishment by hospitals of a quality control program whereby a specific explanation is required for every infection occurring within the hospital.

Use of polyelectrolytes and electron microscopy for detection of viruses from stool.

Insoluble polyelectrolytes (PE60) were used for the concentration of viruses from stool specimens, confirming the results of Wallis et al. (1969). Ten percent suspensions inoculated with poliovirus type 3 were used in these experiments. A small number of stool specimens from patients naturally infected with enteroviruses were also tested. Preferential adsorption of viruses to PE60 was maximum at a pH range of 4.5 to 6.0. The elution of the adsorbed viruses was optimal at pH 8.5. Other parameters were also investigated. Electron microscopy was used successfully to detect the eluted viruses.

Hazard inherent in microbial tracers: reduction of risk by the use of Bacillus stearothermophilus spores in aerobiology.

The use of a "biological tracer" forms an essential part of many aerobiological experiments. Where release of such tracers is likely to result in deliberate or inadvertent human exposure, safety becomes a primary consideration in the selection of the tracer organism. Of the three most commonly used organisms, namely Bacillus subtilis, Escherichia coli, and Serratia marcescens, only the first comes near to satisfying the need for nonpathogenicity and even it has been incriminated as a cause of human infection, sometimes with a fatal outcome. The relevant characteristics of B. stearothermophilus were, therefore, investigated. Because it can grow only at elevated temperatures (minimum 41 C; optimum 56 C), it should not pose a threat to human health and this view is supported by experimental evidence to be presented. It is extremely easy to grow and maintain in the laboratory, and spore suspensions are easily prepared and stored. It withstands the stresses of aerosolization and sampling and its stability in the aerosol state compares favorably with that of B. subtilis var. niger.