BACTERIAL INDICATORS oF SHELLFISH WATER QUALITY

by Amar S. Menon (Environment Canada 2000)

INTRODUCTION

The objective of microbiological monitoring of the water quality in shellfish growing areas is to provide evidence of protection against the transmission of water-borne infectious diseases. The presence of any microbial pathogens in coastal waters presents potential health risks. However, seeking these pathogens in water is impractical for routine monitoring purposes, and thus an indirect approach of using indicator organisms to measure fecal contamination is developed. The term "Indicator Organisms" are employed only to serve as an indication of fecal pollution from warm-blooded animal wastes and the possible presence of pathogenic micro-organisms. They themselves are not pathogenic and, therefore, cannot be taken as an absolute criterion for the presence or absence of pathogenic organisms in waters. This indirect method of testing waters for the presence of pathogens is necessary because of the technology limitation and cost in the detection of pathogens. Besides, the use of pathogens as the sole indicator of fecal pollution is undesirable because of the variation in number and types of pathogens in sewage and the frequency occurrence of pathogens in water is highly variable. The failure to detect the presence of pathogens in a given water body does not necessarily ensure the safety of the water from all other bacterial and viral pathogens.

INDICATOR ORGANISMS

Since most disease outbreaks associated with consumption of shellfish originated with fecal contamination, a logical approach is to seek a microbial indicator group commonly found in the feces of all warm-blooded animals. Ideally an indicator organism should satisfy the following criteria:

1. It should always be present in waters whenever pathogens are present.

2. It should occur in much greater numbers than the pathogens.

3. It should be absent, or at least very few numbers in clean waters.

4. It should not be able to proliferate to any greater extent than pathogens in aquatic environment.

5. It should respond to natural environmental stress and wastewater treatment processes and disinfectants in a manner similar to the pathogens of interests.

6. Indicator density should bear some relation to the degree or extent of pollution.

7. It should be easy to isolate, identify, and enumerate by routine laboratory procedures.

Unfortunately, no organism meets all these criteria. It is doubtful if an ideal indicator exists or will ever be found for bacterial, protozoa, and viral pathogens. So we must, therefore, deal in terms of the best indicator available. The best indicator would obviously be the one whose density correlates best with the health hazards associated with fecal contamination.

A variety of groups of bacteria and viruses have been used or recommended to measure the sanitary quality of recreational and shellfish growing waters. These range from a broad spectrum group, such as the total plate counts, to a narrow spectrum group such as Escherichia coli and specific pathogens. Of these various groups the most commonly used are the total coliform, fecal coliform, and fecal streptococcus groups.

TOTAL COLIFORMS

The most widely accepted bacterial indicator of fecal pollution in water has been the coliform group of bacteria. The coliform bacteria has been chosen as indicator of water quality for many years primarily based on the work of Escherich in 1885, in which he identified Bacillus coli, from which the name "coli-form" is derived, as being characteristic of feces of warm-blooded animals. The presence of these organisms in water was assumed to indicate a potential health hazard because of their association in the gut with a variety of pathogenic microorganisms.

However, in recent years, it was shown that coliform bacteria was in fact a heterogeneous group of bacterial species composed of Escherichia, Klebsiella, Citrobacter, Enterobacter, and Aeromonas genera. While Escherichia coli is exclusively fecal in origin, the other four genera are widely distributed in nature and commonly found in soils, on vegetation, and in industrial wastes. The non-fecal coliform biotypes are frequently associated with surface runoff and have a tendency to multiply in nutrient-rich waters. Therefore, the presence of total coliforms in surface waters does not always imply fecal contamination and the sanitary significance of these bacteria becomes doubtful if no obvious pollution source is found. For this reason, more consideration as to the origin of the coliform organisms is in order.

FECAL COLIFORMS

The inadequacy of the total coliform test in differentiating coliform bacteria of fecal origin from non-fecal source has lead to the development of the fecal coliform test. Fecal coliform is a subgroup of the total coliform group which is capable of producing gas within 24 hours at 44.5°C in EC or A-1 medium. This group is considered to be more specific indicator for fecal contamination and was adopted by the NSSP in 1974 for the monitoring of shellfish growing waters. The advantages in using fecal coliforms as indicator are as follows:

1. The majority of coliform bacteria from the intestines of warm-blooded animals grow at elevated temperatures.

2. These organisms are of relatively infrequent occurrence except in association with fecal pollution.

3. These organisms are more resistant to the aquatic environment than are many enteric bacterial pathogens.

4. These organisms are shed in greater numbers than are most pathogens.

5. These organisms can be grown on simple media and quantified using routine laboratory procedures.

However, there are also some limitations associated with the use of fecal coliforms as an indicator group. The specifically of fecal coliforms suffers some of the same shortcomings of the total coliform group, in that it contains organisms whose source is not exclusively fecal. In fact, fecal coliforms is also a heterogeneous group that comprises of Escherichia coli, as well as thermotolerant Klebsiella biotypes. Klebsiella is not frequently present in human feces. They occurs in pulp and paper mill effluents, (1) textile processing plant wastes, (2) and other industrial sources in the absence of fecal contamination.(3) The ability of Klebsiella to multiply in polluted waters diminishes its usefulness as an indicator. Recent epidemiological study (4) indicate there is little correlation between fecal coliform density and swimming associated gastrointestinal illness in aquatic environment. Escherichia Coli and enterococci may be a better indicator for assessing health hazard risk in surface waters.

FECAL STREPTOCOCCUS

The third group of bacteria, the fecal streptococcus, has been suggested as a useful indicator of fecal contamination because they are present in large numbers in feces. They do not multiply in surface waters and are more resistance to adverse environmental conditions. Unfortunately, the fecal streptococcus, like the coliform group, also includes several biotypes which are widely distributed in nature and are of limited sanitary significance. The most valuable application of the fecal streptococcus indicator system in water quality is the development of fecal coliform to fecal streptococcus ratios that may be used to asses the source of fecal pollution.(5) If the ratio of the number of fecal coliform to fecal streptococcus is greater than 4, this generally indicates a human origin, and a ration of less than 0.7 indicates animal source. These ratios must be applied carefully as environmental factors will influence the fecal coliform and fecal rate of each in the receiving waters. For these reason, the ratios for water samples are valid only during the initial 24 hours from the source of pollution. For after 24 hours, problems with die-off tend to obscure meaningful results.

Microbiological Methods

Bacterial indicator densities in water are commonly determined either by a multiple-tube-fermentation (MPN) procedure or by a membrane filter (MF) technique. Detail description of these two methods are found in the current edition of Standard Methods for the Examination of Water and Wastewaters.(6)

Each method has its own advantages and disadvantages, but for the most part, they are considered to give comparable results, especially for fresh waters. From a statistical aspect, the MF method gives direct counts, better reproducibility of results, produces results in 24-48 hours and allows the examination of large volumes of samples than the MPN method. However, the presence of high suspended solids, heavy ions, algae, high densities of non-coliform organisms or other interfering substance in seawater may limit the application of the MF test in shellfish growing water. Presently, NSSP criteria specify the use of MPN procedure for the bacteriological examination of shellfish growing waters.

There is no world-wide agreement as to what criteria should be used to measure the safety of shellfish. It has been suggested that the shellfish standard should be based on the direct examination of the shellfish and not on the growing waters which is being practised by some countries like France and the United Kingdom. They argue that safety standards based upon shellfish product testing would be more equitable to industry since imports and domestic products would then be subject to the same standards. They also argue there is no direct correlation between water and shellfish quality and shellfish are the product that is consumed.

The basic concept of the NSSP is to control the safety of shellfish by preventing contamination of its environment, not to determine whether or not shellfish become contaminated after the fact. It is built on the premise that safe and high quality water will result in safe shellfish. It is believed that shellfish product standards are less effective than water quality standard in the classification of shellfish harvesting areas because of the following factors:

1. Various shellfish species concentrate bacteria at different rates and levels, and the same product standard probably could not be used for all species.

2. The same species of shellfish will concentrate bacteria at different rates and levels under different pumping and feeding regimes caused by diverse environmental conditions such as water temperature, suspended matter, and salinity.

3. Shellfish species will eliminate bacteria at different rates than viruses when their pumping is reduced due to low water temperatures.

4. Repeated sampling of shellfish shows wider variation in results and less homogeneity than does water sampling in the same area.

5. Shellfish in a shellfish growing area are harder to collect and may not be available at the estuary locations chosen to assess the impact of pollution sources.

6. Negative results in shellfish analysis may give a false sense of security because some health risks (e.g. viruses) can go undetected using traditional bacterial screening methods.

CONCLUSION

Bacteriological measurement of shellfish growing water quality must be based on the detection of fecal contamination by all warm-blooded animals. The fecal coliform test is the best method now available for detecting fecal contamination in waters. The fecal coliform standard are based upon the public health assumption that the presence of fecal material in estuarine constitute a potential risk to shellfish consumption. It must be emphasized that the detection and enumeration of indicator organisms should be interpreted only as what they are intended to indicate. The presence of indicator organisms would only indicate that pollution has occurred. The qualitative determination of bacterial indicators are never intended to be the sole information to judge the health hazard associated with a particular water. A detail knowledge of the sanitary conditions of the study area is essential to make proper judgement. Bacteriological examination of shellfish growing waters should be used only as an adjunct to the sanitary survey to show the extent of fecal contamination affecting an area. Fecal contamination is often intermittent and may not be revealed by the bacteriological examination of a single water sample. The most a bacteriological report can prove is that, at the time of examination, pollution indicating organisms did or did not grow in a selected medium under laboratory conditions from a sample of water. Therefore, if a sanitary survey shows that the waters in a shellfish growing area are obviously subject to contamination from direct fecal wastes, radionuclides, or toxic substances, the shellfish area should be closed irrespective of the results of bacteriological analyses.

NB: Footnote references cannot be located at time of publishing.

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