Drinking Water: The Importance of Microbiological Control

Worldwide, about three out of ten people – out of a total of 2.1 billion – do not have access to safe drinking water at home. These data are from the 2017 World Health Organization (WHO) and United Nations Children’s Fund (UNICEF) report. As a result, 361,000 children under 5 die each year from diarrhea every year . Thus, poor sanitation and contaminated water are directly linked to the transmission of diseases such as cholera, hepatitis A and typhoid.

The lack of potable water is not always noticeable to sight or smell. Thus, a laboratory analysis is necessary to detect it. The microbiological control of water becomes important precisely because it is a vehicle for the transmission of bacteria, namely total and thermotolerant coliforms, protozoa, viruses and fungi that cause numerous diseases to man.


The abundance of water on the planet causes a false sense of inexhaustible resource.
According to environmental experts, 97.5% of the water available on Earth is salty, being unsuitable for consumption. Only 2.493% is sweet, but it is inaccessible in glaciers or underground regions (aquifers). Only 0.007% of the water found in rivers, lakes and the atmosphere available for consumption remains. In addition, data provided by the UN estimates that by 2050, about 50 countries will not have enough water for their entire population.


Water contributes a lot to human health, and these two resources, water and health, associated, can improve the prospects for development. And this relationship between water, hygiene and health is a concept that has been with humanity since the beginning of civilization.

Water can be contaminated at the point of origin. For example, untreated sewage that is released into rivers and lakes or landfills that affect groundwater. Also, during its distribution and, finally, in private reservoirs, whether of companies or households. The most frequent causes of contamination are inadequate sealing and lack of a regular and periodic cleaning and disinfection program.


For the water to be considered potable, after conventional treatment, the physical-chemical and microbiological parameters must be in accordance with Ordinance No. 36, of the Ministry of Health, of January 19, 1990. In its Annex it presents the norms and the standard of drinking water for human consumption, to be observed in the national territory.

Adequate basic sanitation and treated water systems can reduce the incidence of infectious diseases by 20% to 80%. It inhibits its generation and interrupts its transmission.

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The pathogenicity of microorganisms is relative. They are often associated with their host’s immunity, infectivity characteristics and toxin production.

Any microorganism is potentially pathogenic if it finds a weakened host. However, only a limited number of microbial species can cause disease in a significant portion of normal people.

Microorganisms are introduced into our body through the skin or through ingestion of contaminated water or food.

To ensure that the water is free of pathogenic microorganisms, it must undergo a disinfection process. Chlorination is the most commonly used method in most countries. Chlorine, in the form of chlorine or hypochlorite gas, added to water, aims to destroy or inactivate target organisms. Chlorination is a method of simple application, relatively low cost and, above all, very reliable.
Thus, the water to be distributed to the population must contain a certain residual chlorine content. In this way, it prevents further contamination during the distribution process.

In the analysis or monitoring of water quality, specific biological indicators are used, such as bacteria in the coliform group. In Brazil, according to Ordinance no. 518, of 2004, from the Ministry of Health / ANVISA, the water is considered potable, from a microbiological point of view, when it is in accordance with the following conformity: absence of total and thermotolerant coliforms in 100 ml of drinking water sample, thus being considered harmless to man’s health.


Widely distributed in nature, coliforms spread more frequently in water. Especially, the thermotolerant coliforms, of fecal origin, which have received great attention from public health. Thermotolerant coliforms are associated with a high number of pathologies whose etiologic agents are isolated in clinical microbiology laboratories and directly considered the reason for most known human intestinal infections. The most important pathogen in this group is Escherichia coli . Designated as thermotolerant, it has no free life in the environment, indicating that when present in water, it is contaminated by feces.

Some other genera, such as Enterobacter, Citrobacter, Klebisiella and Serratia , live in water, in the soil and also constitute man’s intestinal microbiota, as well as that of other warm-blooded animals, these being characterized as total coliforms.

(Image / ref: E. Coli 10000x – Agricultural Research Service)


The samples must be collected in sterile plastic bottles of 100 ml, containing 10mg of sodium thiosulfate to neutralize the action of residual chlorine. The temperature must be maintained at 4 ° C and must be analyzed within 24 hours.

The multiple tube technique is the most traditional for the analysis of coliforms (total or thermotolerant) and E.coli. This methodology allows quantification by “most likely number” (NMP) of microorganisms. It is divided into two phases, one presumptive and the other confirmatory. The latter is only carried out if there is positive growth in the presumptive stage.

Multiple tube technique for determining the Most Likely Coliform Number

Presumptive phase: consists of homogenizing and transferring aliquots and / or dilutions of the sample to test tubes containing, at the bottom, an inverted tube for gas collection (Durhan tube), in lauryl tryptose broth. All tubes are incubated at 35 ° C for 24 to 48 hours and subsequent identification of those with positive growth of total coliforms. The result is identified by the occurrence of an acid reaction (yellowish color) or gas production (retained in the Durhan tube).

Confirmatory phase: the transfer (transfer of aliquots with platinum loop) from positive presumptive tubes to tubes prepared in the same way as in the previous one, but containing bright green broth, is carried out. All tubes are incubated at 35 ° C for 24 to 48 hours and subsequent identification of those with positive growth of total coliforms, identified by the occurrence of gas production in the Durhan tubes.

According to the number of positive tubes in each of the dilutions and phases used, the most likely number (PWN) is determined, based on statistical tables.


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