Tuong Nguyen

Fecal Coliform in Areas with Impervious Cover

Is the presence of coliform bacteria in waterbodies a result from urbanized area that has impervious cover?

19 April 2011 Tuong Nguyen

Working group #4: Focus on the rapid increase of impervious land cover due to urbanization and how it affects water resources around Sunset Valley, TX

Tuong Nguyen 4.1 Abstract

Fecal Coliform in Areas with Impervious Cover

The population of Sunset Valley, TX has been increasing over the past decades, resulting in rapid increase in urbanization and residencies. The first thing that was altered is land use, which leads to the change in land cover. This affects other resources such as water quality. I am trying to find a connection between the effect of increasing urbanization and its impact on water quality. I tested for fecal coliform population in waterbodies around Sunset Valley and compared that to the percent of impervious cover. My findings were not as good as I had hoped. I was unable to make any connection between the percent impervious cover and fecal coliform population due to many limitations that arose.

4.2 Introduction

The 14th Edition of Standard Methods defines total coliform as a group of bacteria which is aerobic and facultative anaerobic, gram-negative, nonspore-foaming, rod shaped bacteria which ferment lactose with gas formation within 48 hours at 35C. This definition includes the following bacteria: Escherichia coli, Escherichia aurescens, Escherichia freundii, Escherichia Intermedia, Aerobacter aerogenes, and Enterobacter cloacae. These bacteria are found in the intestines of warm blooded animals, thus are present in surface waters, vegetation, soils, and sewage. This group of bacteria is intended to be an indicator of fecal contamination. The indicator organism by itself is not considered directly harmful to man or animals, however, its presence usually indicate the existence of pathogenic or disease-causing organisms. There are problems with

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Fecal Coliform in Areas with Impervious Cover

using the total coliform group as an indicator. Only bacteria of the coliform group that are of fecal origin may be used for analyses. Fecal coliforms are one of those bacteria that are of fecal origin, but human feces also include other types from the total group which are regarded as non-fecal segments of the coliform group. Fecal coliform can be isolated from other coliforms bacteria by setting temperatures higher than 35C during incubation. Fecal coliform have the ability to ferment carbohydrates at 44.5C within 24 hours while the rest of the total coliform group will not (Greenberg et al. 1976). My working group focused on the rapid increase of impervious land cover due to urbanization and its effects on water resources. My duty is to assess whether or not an increase in impervious cover affects the population of fecal coliform. According to the U.S. Census Bureau, the population of City of Sunset Valley, TX has almost tripled from 365 in 2000 to 840 in 2009. The rapid increase of demographics leads to the increase in urbanization such as more roads and shopping malls. The development of this city is ideal for my research because I want to address if fecal coliform levels in waterbodies correlate with increasing level of urbanization. This question is important to ask because the presence of these organisms, good or bad, has an impact on water quality. Increase in impervious cover due to urbanization changes the quality of streams. In addition to imperviousness, runoff from urbanized surfaces as well as municipal and industrial discharges results in increased loading of nutrients, metals, pesticides, and other contaminants to streams. These changes result in consistent declines in the richness of organisms in urban streams (Meyer and Paul 2001). I tested the relationship between impervious cover percent and fecal coliform population. It is important to know what is present in the water and what methods are needed to control its contamination level.

Tuong Nguyen

Fecal Coliform in Areas with Impervious Cover

The appearance of fecal coliform in water may not be directly harmful (Erickson and Doyle 2006); however, it is important to monitor its presence because it may indicate that the water has been contaminated with the fecal material of humans or other animals. Fecal coliform bacteria can enter rivers through direct discharge of waste from mammals and birds, from agricultural and storm runoff, and from human sewage. There has been some research in the past concerning factors that contribute to the population of fecal coliform. Higher pH correlated to the lower concentration of coliforms (Chordi et al. 1991) and higher fecal coliform densities are associated with discharges from wastewater treatment plants (Hendricks and Psaris 1981). Here I examine the effect of land cover due to urbanization on fecal coliform population. I observed the sensitivity of the coliform bacteria to impervious cover percentages. I took equal water samples at three different sites with different impervious cover. I quantified and compared the population of the fecal coliform bacteria to the level of imperviousness of the surrounding land at that particular location. To see if there is any other correlation to the fecal coliform population other than land cover, I took note on the pH level, the oxidation reduction potential, and the temperature of the water sample. Since E. coli is a good fecal coliform indicator, I counted them as a representation of fecal coliform for this project. The U.S. Environmental Protection Agency (EPA) determines that the geometric mean of E. coli should not be over 126 CFU(colony-forming units)/100 mL and not to exceed 235 CFU/100 mL (USEPA 2003) for healthy recreational water.

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Fecal Coliform in Areas with Impervious Cover

4.3 Materials and Methods

There are three different methods in detecting fecal coliform in water. The methods are S-MF (standard aerobic incubation of membrane filters), A-MF (anaerobic incubation of membrane filters), and MPN (most probable number method). S-MF and A-MF are based on direct estimation while MPN is based on statistical analysis. The SMF and A-MF techniques are more sensitive than the MPN technique for counting the total number of coliforms (Cangamella et al. 1988). The procedures in each method are not simple. Due to limited resources and time, I used an alternate method involving the Coliscan Easygel. The Coliscan is patented by Micrology Laboratories in Goshen, IN. This method incorporates two special chromogenic substrates which are acted upon by the presence of the enzymes galactosidase and glucuronidase to produce pigments of contrasting colors. All I have to do to identify the presence and numbers of coliforms is to add the sample water into the medium, pour it in a petri dish and incubate it at a controlled temperature. General coliforms will produce the enzyme galactosidase and the colonies that grow in the medium will be in pink. E. coli will produce both galactosidase and glucuronidase and will produce a dark blue to purple colonies in the medium. I only have to count the blue/purple colonies which indicate the number of E. coli per sample. The pink colonies were omitted because they indicate the general coliform. Any non-colored colonies are not coliforms, but may come from the family Enterobacteriaceae. Because the Coliscan contains inhibitors, most other bacterial types did not grow. The results were available after 24-28 hours of incubation.

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Fecal Coliform in Areas with Impervious Cover

Figure 1: Shows the Coliscan bottle and a sample petri dish after incubation. Note that the E. coli is in purple-blue color.

I carried out my experiment from March of 2011 to April of 2011. I collected a total of 60 mL water sample from each site. The original plan was to take water samples right after rain where runoff carries sediments, nutrients, and other contaminants along with it into drains and ponds. I wanted to take water samples at the same sites as Michael. Unfortunately there was not enough rain thus all the water was dried up when I arrived at the sites. I was fortunate enough to find water from sites in and around Sunset Valley (Figure 2.0). Water samples were taken from A (Figure 2.2): a drain near the intersection of Independence Dr and Armur Dr (Southeast of Sunset Valley), B

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Fecal Coliform in Areas with Impervious Cover

(Figure 2.3): from a pond behind of the apartment complex near 2804 W William Cannon Dr (South of Sunset Valley), and C (Figure 2.4): from a big drain behind Banfield the Pet Hospital near the intersection of Ernest Robles Way and Sycamore Tr (East of Sunset Valley). For each of those sites, I took a total of 60 mL of water samples, measured its pH, ORP (oxidation-reduction potential), and temperature using the ultrameter. I poured 5 mL of the sample water to each Coliscan bottle and then to the treated petri dish. This results in 12 petri dishes per site. I allowed the liquid inside each dish to solidified, and then I cover the edge of the dish with parafilm and incubate it at approximately 37.5C. After at least 24 hours, I came back and count the total number of fecal coliform bacteria per 5 mL of water.

Figure 2.0: Shows the City of Sunset Valley and its boundary.

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Fecal Coliform in Areas with Impervious Cover

Figure 2.1: Shows an aerial overview of three sample sites. Site A is Southeast of Sunset Valley, site B is South of Sunset Valley, and site C is inside of Sunset Valley.

Figure 2.2: Shows an aerial overview of site A.

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Fecal Coliform in Areas with Impervious Cover

Figure 2.3: Shows an aerial overview of site B.

Figure 2.4: Shows an aerial overview of site C.

Tuong Nguyen 4.4 Data/Results

Fecal Coliform in Areas with Impervious Cover

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I did not expect the mean value of CFU/5 mL to be that close. The standard deviation is also higher than my expectation. The standard deviation became even more problematic when I convert my CFU/5 mL into CFU/100 mL. This means that there is possible error in my counting or on the quality of water itself. According to Table 1.1, the UCF/100 mL in all three sites exceeded the EPA standard for healthy recreational water. Fortunately the samples were taken from drains and pond, thus they were not meant to be used for recreational purposes. The pH of the three sites is not too far from each other, they are at around the pH of normal water. Temperature varies a little bit, but it is nothing too dramatic. The ORP values are also not too different, it ranges from 168 ppm to 230 ppm.

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Fecal Coliform in Areas with Impervious Cover

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Site pH ORP (ppm) T(C) Dish 1 2 3 4 5 6 7 8 9 10 11 12 Average Std. Dev.

A 7.14 168 24.2 CFU/5 CFU/ 100 mL mL 34 680 22 440 42 840 36 720 23 460 25 500 31 620 24 480 37 740 38 760 25 500 33 660 30.833 616.667 6.807 136.137

B 6.93 230 25.6 CFU/5 CFU/ 100 mL mL 46 920 39 780 53 1060 27 540 30 600 23 460 33 660 42 840 31 620 28 560 48 960 35 700 36.250 725.000 9.353 187.059

C 6.73 189 23.4 CFU/5 CFU/ 100 mL mL 36 720 27 540 30 600 29 580 33 660 42 840 30 600 28 560 35 700 26 520 40 800 28 560 32.000 640.000 5.222 104.447

Table 1.1: Shows the water properties from sites A, B and C. For E. coli, the CFU/5 mL and CFU/100 mL are also shown.

Tuong Nguyen 4.5 Discussion

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From Table 1.1 and figure 3.1, temperature had no effect on the population of E. coli. This was to be expected because I did not collect enough water samples with a wide range of temperature. The pH value did not correlate well with the E. coli population (Figure 3.2). The mean E. coli population peaked at approximately 6.9 pH and decreased when it became more basic or acidic. The average E. coli population for site B was higher than that of site A and C although water at site A has a higher pH and water at site C has a lower pH, respectively. Once again, I need more water samples to be able to make a sound conclusion. One other correlation that arose from Table 1.1 and Figure 3.3 was that the ORP value affected the average E. coli population by a small amount. The data suggested that higher ORP value yielded a higher average fecal coliform population. Although I only had three points to analyze, I can confirm that ORP does have a positive effect on E. coli population.

Figure 3.1: Shows the relationship between temperature and mean CFU/5 mL.

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Fecal Coliform in Areas with Impervious Cover

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Figure 3.2: Shows the relationship between pH and mean CFU/5 mL.

Figure 3.3: Shows the relationship between ORP and mean CFU/5 mL.

Site A and B are type III residential areas with an average 9% impervious cover and site C is a commercial non-residential area with an average 55% impervious cover

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Fecal Coliform in Areas with Impervious Cover

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(Table 1.2, last column for average percentage of impervious cover). According to Figure 3.4, site C has its mean UCF/5 mL close to site A with a much higher impervious cover percentage. Additionally, site B has higher mean UCF/5 mL than site A and C despite its low impervious cover. There was not enough evidence to make a solid conclusion based on my finding above. I expected the average population of fecal coliform/E. coli to be higher with higher impervious cover percentage but this was not the case.

Single-Family

Total Parcels

Average Impervious Area (SqFt) 5,117 2,646 5,035 8,459 234,820 89,000 169,877 186,360 1,046,329 25,640

Total Impervious Area (SqFt) 1,355,880 116,424 926,485 312,971 6,105,317 89,000 3,737,300 186,360 2,092,657 7,461,197

Average Parcel Size (SqFt) 46,128 13,593 44,904 90,901 429,065 469,795 307,010 937,796 1,496,942 80,342

Total Land Area (SqFt) 12,223,790 598,073 8,262,370 3,363,347

Average Percentage of Impervious Cover 11% 19% 11% 9% 55% 19% 55% 20% 70% 32%

265 I II III < 3,253 ft 2 2 44 184 37 26 1 22 1 2 291

3,253-6,980 ft

6,980 ft2 - 11,966 ft2

Non-Residential Civic Commercial Multifamily AISD Total

11,155,690 469,795 6,754,216 937,796 2,993,883 23,379,480

Table 1.2: Shows impervious cover and land data in Sunset Valley (taken from the Final Report of the Drainage Utility Assessment City of Sunset Valley. July 28, 2010).

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Fecal Coliform in Areas with Impervious Cover

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Figure 3.4: Shows the relationship between mean UCF/5 mL and percent impervious cover.

Tuong Nguyen 4.6 Conclusion

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Although my research failed to make a connection between fecal coliform population and impervious cover percentage, I cannot ignore the fact that the highest concentration of coliform bacteria occurred in the urban headwaters (Andersen et al. 2007). The results should be more conclusive if more sites are available to me. If I still cannot come up with a firm correlation, it could be due to the fact that coliform and E. coli population estimate techniques measure from 5 to 100% of the potential population (Dutka, Kuchma, and Kwan 1979). The errors for my findings can be minimal to substantial. In order to get the best result possible, I may have to use other methods such as the S-Mf, the A-MF, and MPN with more water samples from each site. Unfortunately I do not have enough time and resource to work on my project with extreme precision. This project has potential and it is unlucky that I was not able to see the results I wanted. There should still be worries about the issue of contamination in water resources due to increasing urbanization. There will be build-up of organic matter and nutrients on soils if urbanization continues to increase. If left unchecked, these organic matter and nutrients will have an effect on the fecal indicator bacteria including total coliforms and E. coli. Accumulation and survival of the fecal indicator bacteria has a negative impact on surface water quality. In surface water, sediments may make up a reservoir of different pollutants including inorganic and organic compounds and microorganisms. The survival of fecal bacteria, once released in the aquatic environment, is determined by many environmental factors including temperature variation, salinity, oxygen levels,

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Fecal Coliform in Areas with Impervious Cover

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nutrient deficiencies, predation, and ultraviolet irradiation (McFeters and Singh 1991; Davies et al. 1995; Thomas et al. 1999; Hughes 2003; Craig et al. 2004). There is a correlation between the higher growth and lower decay of fecal indicator bacteria in sediments with high level of organic matter and nutrients (Amedegnato et al. 2009). I understand that I did not produce the result I wanted, but if I have more opportunity to test more sites, there will be a positive correlation between percent impervious cover and fecal coliform population. For future studies, I will definitely do a more thorough research about my site and the availability of water in and around it. It is always best to collect samples after a rain event, but if rain is an issue, there should always be waterbodies nearby to provide backup samples. Furthermore, I will collect water at as many sites as possible. The amount of data will determine how strong or weak the correlation between all of the possible factors that influence the population of fecal coliform.

Tuong Nguyen Acknowledgement

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I thank Nicholas Kuzola, Michael Nelson, Matthew Hubbard, and Ladislaus (Dan) Perenyi for providing me with information and insights about my project. I also thank Stephen Bond for providing me with equipment and materials necessary for me to carry out my experiment. I especially thank Dr. Poteet for her help in guiding my project to the right direction and also for fully funded my project.

Tuong Nguyen References

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Amedegnato, E., Haller, L., Pot , J., and Wildi, W. 2009. Influence of freshwater sediment characteristics on persistence of fecal indicator bacteria. Water, Air, & Soil Pollution 203:217-227 Andersen, C. B., Haney, D. C., Lewis, G. P., Liao, M., and Sargent, K. A. 2007. Urban influences on stream chemistry and biology in the big brushy creek watershed, South Carolina. Water, Air, & Soil Pollution 182(1-4):303-323 Chordi, A., Fernandez, and A., Tejedor, C. 1991. Influence of pH on the elimination of fecal coliform bacteria in waste stabilization ponds. Water, Air, & Soil Pollution 39(3-4):317-320. Cangamella, F., Cesaroni, D., Grandi, M., Poda, G., and Trovatelli, L.D. 1988. Coliform detection from river waters: Comparison between MPN and MF techniques. Water, Air, & Soil Pollution 43(1-2):135-145. Craig, D. L., Fallowfield, H. J., and Cromar, N. J. 2004. Use of microcosms to determine persistence of Escherichia coli in recreational coastal water and sediment and validation with in situ measurements. Journal & Applied Microbiology 96:922 930. Davies, C. M., Long, J. A. H., Donald, M., and Ashbolt, N. 1995. Survival of fecal microorganisms in marine and freshwater sediments. Applied & Environmental Microbiology 61:18881896. Dutka, B. J., Kuchma, S., Kwan, K. K. 1979. Fecal coliform and E. coli estimates, tip of the iceberg. Water, Air, & Soil Pollution 11(3):349-362.

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Espey Consultants, Inc. 2010. Drainage utility assessment City of Sunset Valley. Projcect no. 7074.06. Erickson, M.C. and Doyle, M.P. 2006. Closing the door on the fecal coliform assay. Microbe 1(4):162-163. Greenberg, A. E, Rand, M. C., and Taras, M. J. 1976. Standard methods for the examination of water and wastewater, 14th edition. American Public Health Association. Washington, D.C. 1193 pp. Hendricks, D.W., and Psaris, P.J. 1981. Fecal coliform densities in a Western watershed. Water, Air, & Soil Pollution 17(3):253-262. Hughes, K. A. (2003). Influence of seasonal environmental variables on the distribution of presumptive fecal coliforms around an Antarctic research station. Applied & Environmental Microbiology 69:48844891. McFeters, G. A., and Singh, A. 1991. Effects of aquatic environmental stress on enteric bacteria. The Journal of Applied Bacteriology 66:559569. Meyer, J. L., and Paul, M. J. 2001. Streams in the urban landscape. Annual Review of Ecology and Systematics 32:333-365. Thomas, C., Hill, D. J., and Mabey, M. 1999. Evaluation of the effect of temperature and nutrients on the survival of Campylobacter spp. in water microcosms. Journal of Applied Microbiology 86:10241032. USEPA. 2003. Bacterial water quality standards for recreational waters (freshwater and marine waters) status report. EPA-823-R-03-008 Micrology Laboratories The U.S. Census Bureau