MATERIALS AND METHODS
spores of B. subtilis were most resistant to freezing. E.
coli succumbed more rapidly than the other organisms
General information
during storage.
Kraft (1992) reported that although considerable
Snow made from secondary wastewater at a treat-
strain variation exists, vegetative cells of cocci are re-
ment facility in Carrabassett Valley, Maine, was used
sistant to freezing and frozen storage, and gram-nega-
in these studies. To determine the fate of bacteria as a
tive bacteria are less resistant in general than gram-posi-
result of spraying, wastewater was collected after it
tive bacteria. Spores are very resistant to freezing. With
entered the spray system, and freshly fallen manufac-
the exception of the gram-positive Cl. perfringens,
tured snow was collected. Because this treatment facil-
Mackey's (1984) data also appear to support the claim
ity operates only when it is able to make snow, we were
that gram-positive cells are more resistant to freezing.
unable to conduct any tests that would allow us to de-
Organisms in logarithmic growth phase are not as re-
termine whether cell losses that are associated with
sistant as those in stationary phase.
spraying were due to freezing or to the rapid change in
pressure at the spray nozzle.
Effects on other microbial pathogens
Snow columns, which were placed in a temperature-
Although there is a great deal of information on the
controlled environment or outdoors, were used to de-
effect of chilling, freezing, and rewarming (thawing)
termine the impact of overwintering and spring melt
on bacterial survival, we have not been able to find much
on bacteria. For these columns, the freshly fallen, manu-
information on the effect of these processes on other
factured snow was collected and placed in plastic stor-
types of pathogenic, sewage microorganisms, such as
age bags, which were then placed in insulated coolers
helminth eggs, protozoa, or viruses. A study by Sanin
for transport back to our laboratory. The harvested snow
was stored at 10C (15F) until it was used to build
et al. (1994) did compare survival of several microor-
ganisms in frozen sludge (Table 1). They found that
the snow columns.
fecal streptococci and Ascaris eggs (parasitic worms)
The snow columns were placed either outdoors or
were the most resistant to the effects of freezing, that
in a low-temperature incubator that was cycled from
temperatures from 8C to +14C. The snow columns
bacteriophage and polio virus were less susceptible than
fecal coliforms, and that the protozoan oocysts were
were cycled so that the snow column would undergo
completely destroyed by freezing (>8-log reduction).
numerous freezethaw cycles. There was no set pat-
With respect to other water treatment processes,
tern to the cycling except that daytime temperatures
Ridgway (1984) reported that in water, the degree of
tended to be warmer than nighttime temperatures. The
resistance to inactivation of various types of microor-
pattern of warming and cooling was deliberately erratic
ganisms was vegetative bacteria < viruses < bacterial
to simulate a typical New England winter. Meltwater
spores and protozoa.
was sampled to determine the fate of bacteria during
Table 1. Reduction of pathogenic and indicator microorganisms in
two different sludges by freezethaw conditioning. (From Sanin et
al. 1994.)
Overall log reduction
Aerobically
Anaerobically
digested sludge
digested sludge
Fecal coliforms
1.90
1.10
Fecal streptococci
0.21
0.20
Salmonella
0.54
0.74
Viral Plaque Forming Units
0.80
0.85
Poliovirus
1.08
1.47
Helminth ova1
0.06
0.03
Protozoa oocysts2
>8.00
>8.00
1Ascaris
2Cryptosporidium
parvum
4
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