THE SURVIVAL OF ESCHERICHIA COLI IN GROUNDWATER

JFP ENGELBRECHT

CSIR, Tel: 021 888 2659, Fax: 021 888 2686, E-mail pengelbr@csir.co.za

ABSTRACT

Microorganisms will be the dominant forms of life and, in most cases; they will be the only
forms of life present in aquifers. However, with very few exceptions the only waterborne
microbial pathogens of man are essentially human bacteria, viruses and protozoa. Thus, in
considering the safety of drinking water from the point of view of infectious diseases one
can almost completely ignore any source of infectious agents except human and animal
excreta. Therefore, it is important to understand the survival of faecal and pathogenic
bacteria, introduced to groundwater by anthropogenic activities.

The supply of clean water to all the people of South Africa is a necessity. The
contamination of groundwater is a fact. The die-off rate of Escherichia coli in groundwater
50 90
is related to many factors, some known and some unknown. The T and T are studied
by many researchers all over the world, but the results vary from aquifer environment to
aquifer environment and from researcher to researcher.

The survival of Escherichia coli in groundwater was studied. A literature study, a

laboratory study and a field investigation were done. From the literature it was found that
every study, conducted on the survival of bacteria in groundwater, has its own limitations,
resulting in the variations of the prediction of the survival of microorganisms in
groundwater. However, there is enough evidence that survival of Escherichia coli in
groundwater does occur. From the laboratory and field investigation it is concluded that
there are a decline in numbers over time. That Escherichia coli survived for more than 40
days in the laboratory. Escherichia coli does survive in the environment. Escherichia
coli numbers in the soil are higher than in the subsurface water. That Escherichia coli
can move from surface water to groundwater.

INTRODUCTION

The survival of non-soil bacteria, introduced to groundwater by anthropogenic activities, is

of great importance to drinking water quality. Although groundwater scientists know that
indicator bacteria and pathogens does survive to a certain extent in groundwater, the
overwhelming majority of water managers, engineers and the general public believes
otherwise. Groundwater has long been considered to be of excellent quality because of
the soil barrier providing effective isolation of groundwater from surface pollutants.
However, if microorganisms survive the passage through the vadose zone, they will
contaminate groundwater and may emerge in springs and boreholes used as drinking
water supplies (1). The number and variety of the microorganisms in natural waters vary
greatly in different places and under different conditions. Bacteria are washed into the
water from the air, the soil and from almost every conceivable object. The faeces of
animals contain vast numbers of bacteria and many enter natural water systems.

The cysts and vegetative forms of protozoa, bacteria and viruses are the three groups of
microorganisms that may travel through porous geologic media. These microorganisms
differ greatly in size and survival potential in the environment. Each group has members
that are capable of causing disease in humans and animals. Water-born diseases
transmitted by pathogenic organisms include infectious hepatitis, cholera, typhoid,
bacterial dysentery, amoebic dysentery, cryptosporidiosis and giardiasis (2).

The sizes of laboratory grown bacterial cells are in the order of 1 micrometer, but the
smallest survival forms in soil may be only a few tenths of a micrometer in diameter (3).
The sizes of openings in subsurface material can be assumed to be variable and are
generally not measured, but porosity and permeability measurements on aquifer
sediments indicate that adequate spaces for bacteria exist in many sediment types, even
in some rather dense porous rocks (4). The interstices of the shallow aquifer sediments
can easily accommodate bacteria and probably protozoa and fungi as well. Larger
organisms will be excluded from most subsurface formations, except for gravelly and
cavernous aquifers. Therefore, microorganisms will be the dominant forms of life and in
most cases, they will be the only forms of life present in aquifers. However, with very few
exceptions the only waterborne microbial pathogens of man are essentially human
bacteria, viruses and protozoa, and in considering the safety of drinking water from the
point of view of infectious diseases one can almost completely ignore any source of
infectious agents except human and animal excreta.

LITERATURE STUDY

Movement
The factors that control the transport of bacteria through porous media are not well
understood while the study of microbial movement in the field under unsaturated flow
conditions have received only limited study to date. However, advection, dispersion,
deposition (clogging) and entrainment (declogging) are all processes that affect transport
in noticeable ways.

In two case studies (5 & 6) where water with a bacterial load was injected into a 177 feet
deep borehole, the bacteria were detected after 9 days in an observation borehole, 60 feet
away. However, under winter conditions, no bacteria were detected in the observation
borehole.

Percolation beds (7) were set-up in parallel across a shallow stratum of sand and gravel
confined in an old river bed and secondary sewage effluent, given tertiary treatment in
oxidation ponds was pumped into it. The infiltrated water moved down this confined
channel a total of 1500 feet to an interceptor trench dug across the channel. Boreholes
were located 200 feet and 400 feet downstream from the percolation beds. The median
value of faecal streptococci in the oxidation pond fluent was 4500/100mL, while median
values in the 200 feet and 400 feet boreholes and the interceptor trench were 20, 48 and
6.8, respectively.

The movement of coliform bacteria in primary settled sewage through porous sand was
investigated (8). Primary settled sewage was infiltrated into four inch diameter columns of
comparatively permeable agricultural soil. The percolate was sampled at various depths
below the soil surface. From the results it was evident that not only was the surface
stratum the limiting horizon for movement of sewage particulate, but also that it greatly
restricted the entrance of bacteria into the soil. The problem of bacterial removal was
resolved into two distinct areas: (i) effect of surface stratum and (ii) the behaviour of
bacteria in the subsurface in the absence of particulate matter. Near the surface of the soil
an abrupt drop in coliform bacteria in the percolant occurred, but farther down in the
column, below the first few centimetres of clogged surface stratum, the decrease appeared
to be a logarithmic function of the distance traversed by the contaminant percolate. The
study concluded that mechanical straining and sedimentation are the two removal
mechanisms. For a short period following the application of bacteria-laden water to a
porous medium both mechanisms are operative. The capacity for removal by straining in
the first layer of grains is soon reached, and is thereafter inoperative, or saturated, leaving
only sedimentation operative in that region.

In Ottawa, Canada, a 7.93 m long septic tile was installed in sandy clay (9). The
groundwater level fluctuated between near ground level during spring snow melt period to
a depth of 2.44 to 3.05 m during summer and early autumn periods. Vertical and
horizontal permeability tests of the soil, taken at 0.61 m depth, showed mean values of 1.8
-5 -1 -3 -1
x 10 cm s , and 1.1 x 10 cm s respectively. Part of the effluent from a septic tank from
an individual household was diverted to the septic tile. Groundwater samples were
collected down slope of the septic tile at distances of 0, 3.05, 6.1, 9.15, 12.2 and 15.25 m
at 2 m depth from the end of the septic tile. The groundwater table was 0.15 m below the
bottom of the septic tile trench. According to this study, most of the provinces in Canada
specify, at that time, a minimum distance of 15.25 to 30.5 m between a drinking water
supply well (shallow) and a septic tank. The level of high groundwater table should be at a
minimum distance of 1.22 m from the trench bottom. The study concluded that although a
decline of microorganism levels occur over distance, relative high levels of microorganisms
were found in groundwater at a horizontal distance of 15.25 m from a septic tile. The high
level of microorganisms at 15.25 m were due to a fluctuating groundwater table which
limited the travel distance in the unsaturated soil to 0 to 0.15 m, thereby limiting organism
removal through filtration / adsorption. However, the data presented in the paper,
indicates that the travelling distance may be double that of 15.25 m with the decline in
colony forming units of only one log.

Goyal, Keswick, B.H. and Gerba (10) monitored three slow rate operational land treatment
sites. Two sites received secondary treated effluent while the third site received primary
sewage that were aerated and then stored in lagoons. A total of thirteen soil samples and
twenty-three groundwater samples were collected from all three sites. The soil samples
were collected to a depth of 18 cm. The groundwater samples were collected to a depth of
27,5 m. Viruses were detected in 6 of the 13 soil samples at two of the sites, while 11 of
the 26 groundwater samples showed viruses at all three sites. Total coliforms, faecal
coliforms and faecal streptococci analysis were done on the groundwater samples only.
Total coliforms were detected in all the groundwater samples while faecal coliforms and
faecal streptococci were detected in 60% and 44% of the groundwater samples
respectively. They concluded that their study proved that viruses can occur at considerable
depths beneath slow rate sewage land application sites where crop irrigation of
wastewater is practised.

Gerba and co-workers (11) reported the isolation of viruses from as deep as 30 m and
from as far as 100 m from sewage treatment basins in Texas, Arizona, Michigan, and
Israel.

Viral and bacterial contamination of groundwater by on-site wastewater treatment systems

was studied in an experimental field on the North Carolina coast by Moe, Cogger &
Sobsey (12). They used a low pressure dosing system that applied septic tank effluent
(STE) at three hydraulic loading rates to two, adjacent drain fields with the same sandy soil
but different water table depths (high watertable = 0.3 to 0.45 m and low watertable = 0.6
to 0.9 m). Known amounts of bovine enterovirus type 1 (BE-1) were added to the STE.
Groundwater samples were collected twice weekly from PVC monitoring wells (1.5 to 1.9
m deep) placed 0.3, 1.5 and 3 m down gradient from each site. STE samples were taken
from the distribution lines of each field during dosing cycles. The groundwater samples
were analysed for BE-1 twice weekly before, during and after virus dosing periods for three
weeks. Faecal coliforms, faecal streptococci and coliphages were also measured
periodically. Concentrations of BE-1, faecal coliforms, faecal streptococci and coliphages
were reduced from their levels in the STE in both fields. The reductions were significantly
greater in the field with the deep groundwater table. BE-1 was detected in groundwater by
the second day of virus dosing and persisted as long as 30 days post-dosing.

They concluded that there was no consistent pattern of increased microbial concentrations
in groundwater at greater hydraulic loading rates of the STE. They further stated that their
results indicated that the use of septic tank soil adsorption systems in permeable soils with
shallow water tables can contaminate groundwater supplies. It also demonstrates the
importance of unsaturated flow for efficient removal of enteric viruses and bacteria.

Fontes and co-workers (13) added bacteria as a pulse to the tops of columns of clean
quartz sand with an artificial groundwater solution pumped through the columns. Their
experimental variables tested were ionic strength of the effluent, grain size of the porous
medium, bacterial cell size and heterogeneities within the medium. They results indicate
that, even in cases of efficient filtration, significant numbers of bacteria can be transported
through porous media.

Survival
The outbreaks of typhoid fewer at the turn of the century from eating raw vegetables grown
on soil fertilised with raw sewage resulted in extensive studies of the survival of enteric
bacteria in soil (11).

Hagedorn and co-workers (14) studied the movement and survival time of Escherichia coli
and Streptococcus faecalis in septic tank effluent. They reported survival time of 32+ days
with moving distances of 30m in soil with a surface gradient of only 2%.

Kibbey and co-workers (15) inoculated five different soils (16 of each) with faecal
streptococci. The soils were incubated under different temperature and moisture
conditions. The soils were sampled and analysed over a period of 120 days. Their results
showed that Streptococcus faecalis, regardless of soil type, died faster under drier
conditions at warmer temperature and survived the longest under moist conditions at
cooler temperatures. The average 95% reduction time for the bacteria under saturated
conditions was 53 days except for a silty clay loam where the 95% reduction had not
occurred by the termination of the study at 120 days.

After a large waterborne disease outbreak of Escherichia coli O157 : H7, Rice and co-
workers (16) studied the survival of the epidemic strain (C4195) together with another
strain (932) and a typical Escherichia coli (strain R1), a fresh isolate from the Ohio river.
The bacteria was inoculated in the laboratory into 250 mL bottles filled with groundwater
collected from the town supply boreholes. The bottles were incubated in the dark at 5 C
and 20 C. Samples were collected at various timed intervals and analysed. No significant
reduction for all three organisms was observed at both temperatures until the 7th day.
After the 7th day, die-off at 20 C was more rapid with a 5 log10 reduction by day 35. The
strains incubated at 5 C showed only a 3 log10 reduction by day 70.
Filip and co-workers (17) carried out model experiments in the laboratory on the survival of
Escherichia coli, Salmonella typhimurium, Pseudomonas aeruginosa, Yersinia
enterocolitica, Staphylococcus aureus, Streptococcus faecalis, Bacillus cereus, Bacillus
megaterium and Clostridium perfringens in groundwater and groundwater with sand.
Groundwater was placed in portions of 500 mL in Erlenmeyer flasks. In some of the
Erlenmeyer flasks, 250 g portions of sand was added. All were inoculated with the
individual bacteria and kept at 10 C in the dark. Samples were taken at 0, 2, 5, 10, 20,
...100 day intervals. Total die-off for Bacillus megaterium ocurred after 12 days and for
Staphylococcus aureus after 30 days. All the rest survived past the 100 days. Several
prolonged tests indicated that the same bacterial concentrations remained almost stable
up to 300 days.

The movement and survival of bacteria in porous media was investigated by Kott (18)
using bacterial filtration experiments. Bacterial suspension was pumped onto a tube filled
with sand at a rate of 1 mL/min. The survival of bacteria was determined after timed
periods and at various depths. The survival of viruses was determined in dry sand in the
tubes kept at room temperature. Poliovirus 1 survived for over 77 days while f2
bacteriophage survived for more than 217 days. Escherichia coli and Streptococcus
faecalis survived for more than 42 days. Shigella flexneri did not survive longer than 13
days while Salmonella typhimurium survived for 28 days and Salmonella typhi
disappeared within 4 days.

Entry and co-workers (19) studied the movement and survival of total and faecal coliform
bacteria in waste water applied to land with different vegetation types. The water samples
were collected in surface runoff collectors, suction lysimeters and wells. Bacterial counts
after 90 to 120 days, 30 m from application point were down to background levels.

In a natural ecosystem (i.e. the subsurface) populations of the many different organisms
interact in many ways. These interactions may be negative (decreased growth rate) or
positive (increased growth rate) for one or all the different populations in the subsurface. In
very low populations there is little or no interaction. At somewhat higher populations,
positive interaction may predominate and the growth rate may increase. At maximum
population, negative interaction like nutrient exhaustion and production of toxic
metabolites, begin to predominate (20).This interaction between populations is also
indirectly related to the movement of microorganisms through the subsurface and needs to
be taken into consideration when evaluating contaminant movement.

CSIR STUDIES

Aim
To demonstrate whether the survival of non-soil bacteria occurs in real life situations at a
specific site, in this case Atlantis, where enough background information is available to
verify the results in a fully integrative project.

Methodology
During phase one, a laboratory study on the movement of Escherichia coli through sand
columns was investigated. During phase two, groundwater samples were collected in an
area where artificial groundwater recharge, with treated sewage, is practised (Tredoux and
Cave, 2002).
During phase one, sterile water containing Escherichia coli was filtered through columns
filled with soil of different grades. The filtrate was sampled and analysed for Escherichia
coli, by membrane filtration using mFC agar and MUG supplement.

The experimental apparatus consists of the following components (see Photo 1 for better
visualisation):

# four clear PVC cylinders (950mm high, 350 mm diameter, 0.09 m3 volume;
# twelve 15 mm PVC taps;
# sixteen water permeable membranes;
# four 15 L plastic buckets;
# four 80 L plastic buckets; and
# one 15 L plastic bucket with 1300 one millimetre holes in the bottom.

Photo1: Images of experimental apparatus

To each cylinder, three 15 mm PVC taps were connected as shown in the Figure 1. Four
of the sixteen water permeable membranes are placed in each of the 15 L plastic buckets.
One 15 L plastic bucket is placed in each of the 80 L buckets. One cylinder each is then
placed in each of the 15 L buckets on top of the four water permeable membranes and
filled with sand of a specific characteristic. Sterile tap water was used to simulate rainwater
and was distributed onto the sand in the cylinders with the 15 L plastic bucket with the
1300 1 mm holes in the bottom.

The Escherichia coli was cultured in tryptone water. The bacterial count in the stock
solution before dosage was 1.63 x 1014 cfu per 100 mL.

The four cylinders was filled with sand with different grain sizes: No. 1 = 0.42 to 0.47 mm;
No. 2 = 0.60 to 0.65 mm; No. 3 = 250 to 355 mm; and No. 4 = 250 to 275 mm. The total
rainfall, 25.5 L, as calculated for a cylinder was "rained" in succession onto each of the
four cylinders as follows; first watering, 10 L of sterile water containing 10 mL inoculant;
second watering, 8 L of sterile water; and third watering, 7.5 L of sterile water. The
cylinders was again rained on with sterile water (without inoculant) on day, 14, 20, 21, 29,
34, 36, 42 and 43. The experiment was prematurely stopped due to unexpected fieldwork.
 Figure 1: Schematic layout of experimental apparatus

Phase 2 was conducted where artificial recharge is practised since 1982. A schematic
outlay of the area and sampling points are shown in Figure 2.

Inflow
Stilling basin
Inflow
Pond 7

G33135

Surface 1
Augerhole 2 Augerhole 1

Aquifer

Figure 2: Schematic outlay of Phase 2 sampling points

The water used for infiltration, flows from a collection basin by pipeline into a settling basin.
Out of this basin it flows over a flow-measuring devise into an area called "Pond 7". From
this pond the water naturally enters the subsurface to finally reach the aquifer from where
it is abstracted for treatment and use. A handheld auger was used to drill two holes to the
water table. One hole, Auger 1, is five metres from Pond 7 and the other hole, Auger 2, is
100 metres from Pond 7. The water table below ground level is at 1.0 metre in Auger 1
and at 1.5 metre in Auger 2. The closest borehole, G33134, is 50 metres away and
"downstream" of Pond 7. This is an observation borehole and therefore not pumped. The
borehole is 33 metres deep and the water level below surface is 0.3 metre. Moles have
created "flowpaths" through the western weir of Pond 7, resulting in a semi-flooded area
around borehole G33134 causing the high water table. Eight water samples were
collected from the inflow, outflow, surface water, auger water and borehole water. The
water was analysed for faecal coliform bacteria including Escherichia coli.

Results and Discussion

The flow through the cylinders during phase one was observed and the following
differences were noted. In cylinder No.1, ponding and flow were equal (see Photo 2),
fingering was fast with wetting front closeup and spreading through the whole sand column
almost immediately. Water flow through the sand in the cylinder, pass through the four
water permeable membranes, filling the 15 L bucket from the bottom overflowing into the
80 L bucket. The total outflow measured in the 80 L bucket was 20.5 L. Therefore, 5 L
water was contained in the sand column. Three samples were collected and analysed.

Photo 2: Flow through sand column in Cylinder 1

In cylinder No.2, ponding on top of the sand occurred, fingering was slow with no outflow;
Due to the ponding, overflow over the top of the cylinder into the 15 L bucket happened,
and contaminating possible outflow. Therefore no samples could be collected from this
cylinder;
In cylinder No.3, ponding occurred, very slow fingering with the wetting front only halfway
through the sand column after the first watering and outflow only after the third watering,
The total outflow measured in the 80 L bucket was 13.5 L. Therefore, 12 L water was
contained in the sand column.

In cylinder No.4, ponding occurred, very slow fingering as shown in Photo 2. No outflow
with wetting front only one quarter through the sand column after first watering. Slow
outflow 10 minutes after third watering. The total outflow measured in the 80 L bucket was
13.5 L. Therefore, 12 L water was contained in the sand column.

Photo 2: Slow flow and ponding in Cylinder 4

In order to determine the die-off or survival in the sand columns, the cylinders were left
alone for a period of 14 days. Watering and sampling times with results are set out in
Table 1.

Table 1: Escherichia coli counts (cfu/100 mL) in water from cylinders on different
days

Cylinder 1 15 20 21 29 34 36 42 43
No. 1 S1 110 0 - - - 0 0 0 0
S2 900 4900 - - - 0 0 - -
S3 1250 - - - - - - - -
No. 2 S1 - - - - - 0 0 0 0
S2 - - - - - 0 0 - -
No. 3 S1 750000 - - - - 1200 700 1000 29
S2 1200000 - - - - 1400 3400 - -
No. 4 S1 900000 144000 25300 43400 1900 3000 161 100 5700
S2 1410000 192000 112000 46000 10900 12800 21300 - -

The results were a bit confusing. One would have assumed that with the fast flow through
the Cylinder 1, the bacterial count in the outflow water would have been high. However,
the opposite was found with low numbers in the fast flow cylinder and high numbers in the
slow flow cylinders. Due to the contamination of the receiving bucket in cylinder No. 2, this
cylinder was only used from Day 34. At that stage no Escherichia coli was found. The
Escherichia coli count in the first sampling is always lower than the count in the second
sample. Comparing the 1st sample with the 2nd sample it appears that although the
bacteria is spread through the sand column, higher numbers occur higher-up in the sand
column. From the results it is also evident that the counts are getting lower over time
although there are higher counts in between at later days. Why the bacteria disappeared in
the outflow water from cylinder No. 1 after 15 days is still a mystery. They are either locked
in a biomass in the cylinders, or washed out after sampling or most of them died during the
race through the sand column. The results are also presented in Figure 3 for better
visualisation.

10000000

1000000

100000

4S2
10000
4S1
3S1
1S3
CFU / 100 mL

1000
1S2

100 1S1
3S1

10

0
2S1
2S2

1 15 20 21 29 34 36 42 43

Sampling days

Figure 3: Escherichia coli in sand columns over time

The results from phase 2 are presented in Table 2.

Table 2: Microbiological analyses results (per 100 mL) of Atlantis samples

Sample ID Inflow Outflow Surface 1 Surface 2 Auger Auger Auger G33134

1 1 2
Faecal coliforms 370 131 75 1314 24600 2990 780 1
Escherichia coli 163 32 7 1000 15000 1960 160 1

Escherichia coli was found in all the samples collected. With this limited information it is
not possible to evaluate or to scrutinise the results in depth. However, one can
hypothesise that Escherichia coli can survive in the environment and move through the
soil-water matrix. As mentioned previously the water used for infiltration is collected from
stormwater and secondary treated domestic sewage. The Escherichia coli in the inflow is
rather low and it is assumed that the die-off occurs in the collection basins before it
reaches Pond 7. It is also true in the case of the outflow sample. Flow between the inflow
and outflow is controlled by the settling basin. Birds and other animal life in and around
Pond 7 obviously add Escherichia coli to Pond 7. It is also assumed that the differences in
Escherichia coli numbers in Pond 7 are due to many factors. The dilution factor created by
the volume of water in Pond 7 during the rainy season may be a main factor. From the
Augerhole results it is further assumed that there are more Escherichia coli in the
saturated soil than in the water from these augered holes. At Augerhole 1 Escherichia coli
numbers varies by 1-log when the water was allowed to clear. The Escherichia coli in the
borehole may be because of the fact that the water is standing and the borehole is not
pumped.

CONCLUSIONS

Literature study
In most cases it appears that 60 - 90 days are sufficient for reduction of pathogens to
negligible numbers once they have been through the soil, although survival times as long
as 5 years have been reported.

Some factors that control movement of bacteria are: basic flow through porous media;
saturated porous medium; saturated flow; clogging/deposition; entrainment/declogging;
straining; adsorption; anion exclusion; diffuse double layer; hydrophobia and
sedimentation.

The survival of microorganisms is affected by a number of environmental factors such as

the bacterial type, sunlight, rainfall, soil moisture & holding capacity, temperature, soil
composition, pH, presence of oxygen and nutrients and the availability of organic matter
and the antagonism from soil micro-flora.

Some of the factors that may influence the removal efficiency of viruses by soil are; flow
rate; cations; clays; pH & Iso-electric point; soluble organics and soil chemical
composition.

CSIR studies
From the laboratory study the following conclusions were arrived at:

there are a decline in numbers over time;

Escherichia coli survived for more than 40 days;
it is not known if there was die-off or just washing out; and
it is not known for how long Escherichia coli will stay viable under these
conditions.

From the field study the following assumptions were made:

Escherichia coli does survive in the environment;

Escherichia coli numbers in the soil is higher than in the subsurface water; and
Escherichia coli can move from surface water to groundwater.

Although the data from Phase 2 are very limited, the fact that Escherichia coli was found
viable in the environment during both phases, is an important factor for the supply of
potable water. This should be taken into account where groundwater, vulnerable to
pollution, is used without disinfection. This may have serious repercussions if it is ignored
by government departments and local authorities. One such a case, is the statement in a
document, graveR.doc, sent out by the Director General of the Department of Water
Affairs and Forestry to his regional directors with regards to cemeteries. The following is a
paragraph from this document:

"The process of decay of human bodies is much slower than, for example, the
degradation of waste in a waste disposal site, and most disease-causing
bacteria usually do not survive long outside a living human body, and will expire
within days from burial, before it comes into contact with groundwater or even
surface water. When degradation products of decaying bodies does come in
contact with water, it will therefore have a low risk of containing disease-causing
bacteria and, although high in nutrients, will be small in volume when compared
with waste from, for example, a leaking sewerage pipeline".

The statement that bacteria will "expire within days" is very interesting and it seems that
they have information that is not available to us.

REFERENCES

1. POWELSON, David K. and GERBA, Charles P., 1995. Fate and transport of micro-organisms
in the vadose zone in Handbook of Vadose zone characterisation & monitoring. Ed. L G
Wilson, L G Everett and S J Cullen. Geraghty & Miller Environmental science and engineering
series. ISBN 0-87371-610-8.
2. ENGELBRECHT, J.F.P., 1993. An assessment of health aspects of the impact of domestic
and industrial waste disposal activities on groundwater resources. ISBN 1-86845-028-7.
3. GHIORSE, W.C. and WILSON, J.T., 1988. Microbial ecology of the Terrestrial Sub-surface.
Advances in Applied Microbiology. Vol. 33. Academic Press, Inc., 1988.
4. McNABB, J.F. and DUNLAP, W.J., 1975. Subsurface biological activity in relation to
groundwater pollution. Ground Water Vol. 13 no. 1. January - February 1975.
5. DITTHORN, F., and LUERSSEN, A., 1909. Experiments on the passage of bacteria through
soil. Eng. Rec. 60, 23, 642, 1909. In KRONE, R.B., ORLOB, G.T and HODGKINSON, C.,
1957. Movement of coliform bacteria through porous media. Sewage and Industrial Wastes,
Vol. 30. January 1958, pp 1-13.
6. ROMERO, J. C., 1970. The movement of bacteria and viruses through porous media.
Groundwater, 8, 37-48, 1970.
7. BAARS, J.K., 1957. Trael of pollution and purification en route, in sandy soils. Bulletin of the
World Health Organisation, Vol 16, 4, 727-747, 1957. In GERBA, C.P., WALLIS, G. and
MELNICK, J.L., 1975. Fate of wastewater bacteria and viruses in soil. Journal of the irrigation
and drainage division. American Society of Civil Engineering, Vol. 101, pp 157-174.
8. KRONE, R.B., ORLOB, G.T and HODGKINSON, C., 1957. Movement of coliform bacteria
through porous media. Sewage and Industrial Wastes, Vol. 30. January 1958, pp 1-13.
9. VIRARAGHAVAN, T., 1978. Travel of microorganisms from a septic tile. Water, Air, and Soil
Pollution, Vol. 9, pp 355-362.
10 GOYAL, S.M., KESWICK, B.H., and GERBA, C.P., 1984. Viruses in Groundwater beneath
sewage irrigated cropland. Water Research, 18, 299-302, 1984. In Powelson and Gerba
(1995), In POWELSON, David K. and GERBA, Charles P., 1995. Fate and transport of micro-
organisms in the vadose zone in Handbook of Vadose zone characterisation & monitoring. Ed.
L G Wilson, L G Everett and S J Cullen. Geraghty & Miller Environmental science and
engineering series. ISBN 0-87371-610-8.
11. GERBA, C.P., WALLIS, G. and MELNICK, J.L., 1975. Fate of wastewater bacteria and viruses
in soil. Journal of the irrigation and drainage division. American Society of Civil Engineering,
Vol. 101, pp 157-174.
12. MOE, C.L., COGGER, C.G. and SOBSEY, M.D., 1984. Viral and bacterial contamination of
groundwater by on-site wastewater treatment systems in sandy coastal soils. Second
International conference on groundwater quality research. Editors N.D. Durham and A.E.
 Redelfs. Oklahoma State University, Oklahoma. National center for groundwater research, pp
132-134.
13. FONTES, D.E., MILLS, A.L., HORNBERGER, G.M. and HERMAN, J.S., 1991. Physical and
chemical factors influencing transport of microorganisms through porous media. Applied and
Environmental Microbiology, Vol. 57, no. 9, September, pp 2473-2481.
14. HAGEDORN, C., HANSEN, D.T. and SIMONSON, G.H., 1978. Survival and movement of
fecal indicator bacteria in soil under conditions of saturated flow. Journal of Environmental
Quality, Vol. 7, no. 1, pp 55 - 59.
15. KIBBEY, H.J., HAGEDORN, C. And McCOY, E.L., 1978. Use of Streptococci as indicators of
pollution in soil. Applied and Environmental Microbiology, Vol 35 no 4. Apr., pp 711-717.
16. RICE, E.W., JOHNSON, C.H., WILD. D.K. and REASONER, D.J., 1992. Survival of
Escherichia coli O157 : H7 in drinking water associated with a waterborne disease outbreak of
hemorrhagic colitis. Letters in Applied Microbiology, Vol 15, pp 38 - 40.
17. FILIP, Z., KADDU-MULINDWA, D. And MILDE, G., 1988. Survival of some pathogenic
bacteria in groundwater. Wat. Sci. Tech., Vol 20, no, 3, pp227 -231.
18. KOTT, Y., 1988. Movement and survival of bacteria in porous media. Water, Science and
Technology, Vol. 20, no. 3. pp 61 -65.
19. ENTRY, J.A., HUBBARD, R.K., THIES, J.E. and FUHRMAN, J.J., 2000. The influence of
vegetation in riparian filterstrips on coliform bacteria: 1. Movement and survival in water.
Journal of Environmental Quality, Vol. 29, pp1206 - 1214.
20. UPDEGRAFF, David M., 1991. Background and practical applications of microbial ecology in
Modelling the environmental fate of microorganisms, Ed, C J Hurst. American Society for
microbiology, ISBN 1-55581-031-4, 1991.