MCB 425 Environmental Microbiology Lecture Notes PDF

MCB 425 ENVIRONMNETAL MICROBIOLOGY (3 credit units) BY DR. FATOKUN, E.N. DEPARTMENT OF BIOLOGICAL SCIENCE AND BIOTECHNOLGY, COLLEGE OF PURE AND APPLIED SCIENCE CALE...

MCB 425 ENVIRONMNETAL MICROBIOLOGY (3 credit units) BY DR. FATOKUN, E.N. DEPARTMENT OF BIOLOGICAL SCIENCE AND BIOTECHNOLGY, COLLEGE OF PURE AND APPLIED SCIENCE CALEB UNIVERSITY, IMOTA, LAGOS STATE COURSE OUTLINE 1. UNIT I: Environmental pollution and degradation 2. UNIT II: Air pollutants and pollution 3. UNIT III: Soil pollutants and pollution 4. UNIT IV: Impact assessment of microbial contaminants: air 5. UNIT V: Impact assessment of microbial contaminants: soil 6. UNIT VI: Wastes disposal and management 7. UNIT VII: Domestic wastes and waste water treatment 8. UNIT VIII: Principles and standards of sanitary water quality 9. UNIT IX: Disease transmission by water 1 Introduction Environmental Microbiology is the study of microorganisms and the physical and chemical conditions influencing them. The environment comprises air, water and soil. Optimal environmental conditions are prerequisites to proliferation and the survival of living organisms including microorganisms and man. Past, present and potential global threat of environmental pollution and degradation is a major factor that affects the formation of a society’s environment. The primary goal of this course, therefore, is to provide knowledge of the environmental pollutants and pollution, microbial contamination and deterioration of the environment and its impact, water related diseases, as well as the disposal and management of wastes. Learning Objectives This course is aimed at realizing the following learning objectives: Familiarize with the concepts of environmental pollution and degradation Identify the difference between indigenous and incidental microbes in different environments viz: air, soil and water Explain different microbial pollutants specific to different environments, viz: air and soil Determination of the approaches to impact assessment of microbial contaminants of different environments, viz: air, soil and water Elaborate on the approaches of wastes disposal and management Identify the procedures of water and sewage treatment Elaborate on the principles and standards of sanitary water quality Evaluation of water quality in terms of Biological and Chemical Oxygen Demand (BOD and COD) Distinguish between the different types of water related diseases. Determination of different disease transmission by water. 2  UNIT I ENVIRONMENTAL POLLUTION AND DEGRADATION Learning Objectives:  Understand and identify what an environment is with examples  Familiarize with the concept of environmental pollution  Familiarize with the concept of environmental degradation Learning Outcomes:  The students should be able to define an environment and give examples of such.  The students should have familiarized themselves with the concepts of environmental pollution  The students should have familiarized themselves with the concepts of environmental degradation Environment: The environment can be defined as the total living and non-living surroundings of any organism needed for life and sustainability. The environment comprises air, water and soil, and any other area that can be inhabited by microbes. Environments are components of ecosystems. An ecosystem is a community of microorganisms and their physical and chemical environment that functions as an ecological unit. A major requirement for an environment is for the non-living components to be conducive for the living components to thrive, that is, live, grow and multiply. Ecosphere: The ecosphere or biosphere, constitute the totality of living organisms on Earth and the abiotic surroundings they inhabit. It can be divided into atmosphere, hydrosphere and lithosphere (litho-ecosphere) to describe the portions of the global expense inhabited by living things in air, water and soil environments respectively. 3 Microorganisms live within the habitats of the ecospheres. The habitat is one component of a broader concept of the ecological niche, which include not only where an organism lives but also what it does there. The niche is the functional role of an organism within an ecosystem. Microorganisms may be categorized as autochthonous (native/indigenous) or allochthonous (foreign/incident). Autochthonous (native/indigenous) microbes: these are always found in a given environment and are able to adapt to changes in the environmental conditions e.g. changes in quantity of available nutrients or normal seasonal change. Allochthonous (foreign/incident) microbes: these microorganisms inhabit temporarily in an environment, they multiply when growth conditions are favourable and disappear under unfavourable conditions. Life on the Earth is fragile, and every living being can continue to live only in the environmental conditions optimal for its life. Factors which can influence living organisms and life itself are both natural (e.g. plant’s toxins and poisons), and anthropogenic factors (e.g. products of synthesis or by - products in the process of production of other substances). Environmental Pollution: It is the introduction of undesirable substances in an environment which adversely affect living organisms. Such substances are called pollutants. Pollution occurs when in due course of time, the environment is unable to absorb and neutralize toxic byproducts of human activities (poisonous gas emissions) and pollutants by natural processes. There are four major types of pollution viz; air pollution, water pollution, land and soil pollution and noise pollution. Examples of pollutants include, inorganic pollutants such as heavy metals and their compounds (lead, mercury, copper, cobalt), organic pollutants including pesticides, polycyclic aromatic hydrocarbons (PAHs), etc., organic fertilizers, industrial by-products, etc., chemically inert compounds such as dust and aerosols in the air and suspensions in water and toxic micro-elements including iron, boron, arsenic, selenium, etc. Pollutants that exit for longer period in the environment are called persistent organic pollutants (POP). Bioaccumulation: is the process where pollutants such as toxic chemicals and pesticides concentration increases directly within the tissue and system of an organism e.g. human body. It can occur through the air, water, food consumption, or direct contact with the skin. As the toxic 4 compounds accumulate within the body, they increase the risk of chronic poisoning and other sever health disorders. Biomagnification: refers to the process where some types chemical substances or toxins build up at the higher levels of a food chain; that is, the toxins get transferred from one organism to another, at different levels of food chain. These substances include heavy metals, mercury, toxins and other harmful products. The substances tend to increase and accumulate as they move up the food chain. Table 1: differences between bioaccumulation and biomagnification S/N Bioaccumulation Biomagnification 1 The accumulation of toxic chemical is in the The concentration of a toxic chemical tissue of a specific organism increases as one moves up the food chain 2 The concentration of a substance increases The concentration increases as one moves up within an organism the food chain 3 The concentration of a pollutant increases The concentration of pollutants increases as within an organism they move from one trophic level to the next 4 Bioaccumulation occurs within a specific Biomagnification occurs between different trophic level trophic levels 5 Example is the buildup of a toxic element such Example is the transfer of pollutant from as lead in the human body due to exposure from plants to herbivores, and then to carnivores the environment Environmental degradation means that the environment becomes unusable for its designed purposes or that the development of living organisms and their communities in the environment is disturbed or changed in such a way as to be deleterious or undesirable. Environmental degradation involves the creation of an unfavourable conditions for living organisms, which limits the development of organisms and their communities. Degradation occurs when there is deterioration of the environment as a result of depletion of natural resources such as air, soil and water; the destruction of ecosystems and the extinction of wildlife. By degradation, the environment is progressively contaminated, over-exploited and destroyed. Examples of environmental degradation include: 5 Atmospheric degradation: air pollution by particles, depletion of ozone layer, etc. Water degradation: involves dumping of wastes in water bodies, e.g. industrial wastes in rivers or lakes, garbage into the ocean, and other indiscriminate acts of dumping in water bodies. Degradation of soil and land: this is brought about by soil erosion, poor agricultural practices, overuse of fertilizers and pesticides, landfill leaks, destruction of land by mining activities, deforestation, etc. Environment degradation factors are: Physical factors such as electromagnetic radiation, noise pollution and thermal pollution; Biological factors such as infectious agents, parasites and living organisms whose products of metabolism or decay products are harmful to humans or other living organisms. Effect of environmental degradation Atmospheric changes: environmental degradation could lead to alterations in naturally occurring processes such as water cycle, regular animal and plant activities. Impact on human health: Environmental degradation may result in lack of water, decrease in water and food quality which may lead to several sicknesses and diseases leading to death of millions of people. Scarcity of natural resources: environmental degradation can bring about shortage in resources including food crops, water, arable land, medicinal plants, etc. UNIT II Air pollutants and pollution Air is one of the essential factors making life on the Earth possible. Depending on the body constitution, a human being consumes 6–13 cubic metres of air daily or even more in cases of heavy physical loads. Consequently, trace amounts of harmful substances in the air may have an adverse effect on the human health. 6 Any chemical, biological or physical factor is called toxic if it causes an adverse biological reaction. Air Microflora Bioaerosols are particles of microorganisms living in or originate from living organisms emitted in the air. Microorganisms present in the air as bioaerosols could be in forms of vegetative cells, e.g. fungal and actinomycetes hyphae fragments, spores of fungi and bacteria; viruses; algae and protozoa cysts. Bacterial cells and cellular fragments, fungal spores and by-products of microbial metabolism, present as particulate, liquid or volatile organic compounds may be components of bio-aerosols. Bioaerosols constitute the air microflora. Air microflora is not autochthonous (native) to the atmosphere but is allochthonous populations transported from terrestrial and aquatic habitats into the atmosphere. Example, air microbes at 300- 1000 ft. above Earth’s surface originated from soils and were attached to leaves and dust particles blown by wind. They are dispersed in the air through wind, when bubbles break on surface of water containing microbes, or through dispersal of micro droplets containing soil microorganisms when bubbles form during rain impingement. Air microflora are of two types, outdoor microflora and indoor microflora. Outdoor microflora is the atmosphere outside the building; comprises mostly of fungal genera Aspergillus, Phytophthora, Alternaria and Erysiphae, basidiospores, ascospores of yeasts and Candida, fragments of mycelia and spores of molds. Bacterial genera include Clostridium, Corynebacteria, Achromobacter, Micrococcus, Sarcina and Bacillus. The number and varieties of these microorganisms are dependent on human population and activities. The indoor microflora is the atmospheric air inside the building. It consists mostly of fungal genera Penicillium and Aspergillus; and bacterial genera of Bacillus, Staphylococcus and Clostridium. Sources include furnishings and building materials; fungal contamination within wall, ceiling, and floor cavities by movement of cells, spores, and cell fragments via wall openings and gaps at structural joints. 7  Air Pollution Air pollution is the presence of chemicals in the atmosphere in quantities and duration that are harmful to human health and the environment; it is the presence/introduction of a substance which has harmful or poisonous effects into the air. These chemicals/substances are suspended in the air as particles called aerosols. Aerosols are particulates that are emitted into the atmosphere. Types of air pollutants: Primary pollutants and secondary pollutants Primary pollutants/aerosols - products of natural events (like fires and volcanic eruptions) and human activities added directly to the air Secondary pollutants/aerosols - formed by interaction of primary pollutants with each other or with normal components of the air Sources of Anthropogenic Pollution of Air The main sources of anthropogenic pollution that also affect the quality of air are: Energy production, heating, industrial production, transport and agriculture. Major Classes of Air Pollutants: Carbon oxides (CO & CO2) sources = incomplete combustion of fossil fuels transportation, industry, & home heating CO2 is an important greenhouse gas CO (carbon monoxide) the most abundant pollutant known to affect human health as it combines with hemoglobin and may create problems for infants, the elderly, and those with heart or respiratory diseases Sulfur oxides (mainly SO2, or sulfur dioxide) source = combustion of coal and oil (esp. coal) 8  SO2 released may come from: utilities, industrial manufacturing processes, industrial combustion, transportation, etc. can react with gases in atmosphere to form sulfuric acid ('acid rain') Exposure to SO2 can cause impairment of respiratory function, aggravation of existing respiratory disease (especially bronchitis), and a decrease in the ability of the lungs to clear foreign particles. It can also lead to increased mortality, especially if elevated levels of particulate matter (PM) are also present. Groups that appear most sensitive to the effects of SO2 include asthmatics and other individuals with hyperactive airways, and individuals with chronic obstructive lung or cardiovascular disease. Elderly people and children are also likely to be more sensitive to SO2. Nitrogen oxides - NO (nitric oxide) & NO2 (nitrogen dioxide) source = motor vehicles & industry (burning fossil fuels) can react with other gases in atmosphere to from nitric acid (HNO3) ('acid rain') Volatile organic compounds/aerosols or dust (hydrocarbons) - methane, benzene, propane, & chlorofluorocarbons (CFCs) source = motor vehicles (evaporation from gas tanks), industry, & various household products Concentrations of many VOCs are consistently higher indoors than outdoors. Eye and respiratory tract irritation, headaches, dizziness, visual disorders, and memory impairment are among the immediate symptoms that some people have experienced soon after exposure to some organics. Suspended particulate matter solid particles (e.g., dust, soot, & asbestos) & liquid droplets (e.g., pesticides) sources =power plants, iron/steel mills, land clearing, highway construction, mining, & other activities that disturb or disrupt the earth's surface 9  act as respiratory irritants; some are known carcinogens (e.g., asbestos) can aggravate heart/respiratory diseases Toxic compounds Trace amounts of at least 600 toxic substances (such as lead and mercury) are produced by human activities Mercury is an element that occurs naturally in the earth’s crust. Most people and wildlife can generally tolerate the extremely low levels of this naturally occurring substance. Once mercury enters the water it can be converted to its most toxic form, methyl mercury, by bacteria or chemical reactions. Methyl mercury is absorbed by tiny aquatic organisms, which are then eaten by small fish. The chemical is stored in the fish tissue and is passed on at increasing concentrations to larger predator fish; a process known as Bioaccumulation. People and wildlife at the top of the food chain are consequently exposed to elevated amounts of methyl mercury through the contaminated fish they consume; this process is called biomagnification. sources of mercury = burning coal and waste (such as medical wastes) Photochemical oxidants mainly ozone Because sunlight has a critical role in its formation, ozone pollution is principally a daytime problem in the summer months. The presence of hydrocarbons and nitrogen oxide in sunlight with little air movement leads to the generation of ozone. These two compounds are produced by cars, trucks, factories, and power- generating plants or wherever gasoline, diesel fuel, kerosene, oil, or natural gas is combusted. These gases combine together with sunlight, producing ozone. Urban areas with heavy traffic and large industrialized communities are primary areas for ozone problems. 10  The greatest concern about ozone pollution is the potential damage it may inflict on human health. High concentrations of ozone are especially hazardous to children, the elderly, and people with respiratory problems. Each year many food crops are damaged by ozone. Ozone also damages rubber, nylon, plastics, dyes, and paints. Smog forms from mixture of primarily nitrogen oxides (from vehicles), volatile organic compounds, & sunlight complex mixture of gases but primarily ozone more common in cities with sunny, dry and warm climates Effects of Air Pollution on Human Health Much evidence links air pollutants to respiratory and other diseases in humans. Examples of air pollution-related diseases:  Pulmonary irritation and impaired lung function leading to:  chronic bronchitis: a disease condition of inflammation of the airways (trachea, bronchioles) in the lungs and filled with mucus. It is characterized by constant coughing with mucus, shortness of breath, wheezing sound when breathing and fatigue.  emphysema: a lung disease which results from damage of the air sacs (alveoli). It is characterized by breathlessness, coughing and fatigue.  Cancer: e.g. liver cancer which may be due to an established biological occupational carcinogen from mycotoxins. Examples are aflatoxin from Aspergillus flavus is capable of causing liver cancer. Ochratoxin A produced by different species of Aspergillus and Penicillium causes kidney disease and also a possible human carcinogen. Exposure to aflatoxin and ochratoxin occurs by ingestion but can also occur by inhalation in industries such as peanut processing, livestock feed processing, or when grain dust exposure occurs.  Systemic toxicity from heavy metals including lead and mercury can be harmful to reproductive, neurological and respiratory systems. 11   Increased susceptibility to diseases such as asthma, pneumonia and respiratory viral diseases.  Short-term exposure to air pollution can cause headaches, nausea and dizziness. Effects of Air Pollution on other animals and plants: Wild and domestic animals are probably affected in the same ways as humans Plants are damaged by ozone, sulfur dioxide, & acids:  ozone - weakens pine needles & makes them more susceptible to insects and diseases  sulphur dioxide - suppresses growth  acid - damages leaves & needles & also removes nutrients Pollution-Effects on Microorganisms and Microbial Activity in the Environment Microorganisms play a dominant role in transforming pollutants that reach the environment. This ability results from the fact that microorganisms exhibit an extremely wide metabolic diversity and are thus able to degrade an equally wide variety of chemical compounds. However, since many pollutants are cellular poisons it is not surprising to find that they also inhibit both microbial growth and activity. Microbial activity is responsible for the major cycling of elements in the environment so any impairment of microbial growth will automatically have negative effects on microbial activity. Major transformations, such as nitrogen fixation, nitrification and carbon mineralization may be impaired as a result of pollutant impact. However, microbes use pollutants as nutrient sources, a fact that can lead to major, often unchecked and uncontrolled growth of microbial biomass. The increase in microbial populations following pollution impact can also be used as an indicator that pollution is occurring; the classic example being of course the increase in bacterial (coliforms) following sewage pollution of water courses. Therefore, in terms of the likely disturbance impact of a pollutant on the environment we can therefore recognize negative disturbance, where a pollutant inhibits an essential microbial process, and positive disturbance impact where a process is stimulated by pollutant impact. 12 Acid Rain Source of acid Rain Acid rain, or acid deposition, is a broad term that includes any form of precipitation with acidic components, such as sulfuric or nitric acid that fall to the ground from the atmosphere in wet or dry forms. This can include rain, snow, fog, hail or even dust that is acidic. Acid rain results when sulfur dioxide (SO2) and nitrogen oxides (NOX) are emitted into the atmosphere and transported by wind and air currents. The SO2 and NOX react with water, oxygen and other chemicals to form sulfuric and nitric acids. These then mix with water and other materials before falling to the ground. While a small portion of the SO2 and NOX that cause acid rain is from natural sources such as volcanoes, most of it comes from the burning of fossil fuels. The major sources of SO2 and NOX in the atmosphere are: Nitrogen oxide & sulfur dioxide released primarily from electric power plants & motor vehicles SO2 + water vapor + ozone ---> H2SO4 NO + sunlight + O2 ---> NO2 + various atmospheric gases ---> HNO3 Acid Transport Prevailing winds transport the compounds, sometimes hundreds of miles, across state and national borders. This makes acid rain a problem for everyone and not just those who live close to these sources. Effects of Acid Deposition acidification of lakes and streams Acid rain causes a cascade of effects that harm or kill individual fish, reduce fish population numbers, completely eliminate fish species from a waterbody, and decrease biodiversity. As acid rain flows through soils in a watershed, aluminum is released from soils into the lakes and streams located in that watershed. So, as pH in a lake or stream decreases, aluminum levels increase. Both low pH and increased 13  aluminum levels are directly toxic to fish. In addition, low pH and increased aluminum levels cause chronic stress that may not kill individual fish, but leads to lower body weight and smaller size and makes fish less able to compete for food and habitat. contributes to damage of trees i. Acid rain does not usually kill trees directly. Instead, it is more likely to weaken trees by damaging their leaves, limiting the nutrients available to them, or exposing them to toxic substances slowly released from the soil. Quite often, injury or death of trees is a result of these effects of acid rain in combination with one or more additional threats. Acidic water dissolves the nutrients and helpful minerals in the soil and then washes them away before trees and other plants can use them to grow. At the same time, acid rain causes the release of substances that are toxic to trees and plants, such as aluminum, into the soil. ii. Forests in high mountain regions often are exposed to greater amounts of acid than other forests because they tend to be surrounded by acidic clouds and fog that are more acidic than rainfall. Frequent rain of acid fog on plants, essential nutrients on their leaves and needles are stripped away. This loss of nutrients in their foliage makes trees more susceptible to damage by other environmental factors. Accelerates the decay of building materials and paints, including irreplaceable buildings, statues, and sculptures that are part of our nation's cultural heritage. Controlling Acid Deposition Clean up smoke stacks and exhaust pipes Acid deposition is caused by two pollutants that are released into the atmosphere, or emitted, when these fuels are burned: sulfur dioxide (SO2) and nitrogen oxides (NOx). Sulfur is present in coal as an impurity, and it reacts with air when the coal is burned to form SO2. In contrast, NOx is formed when any fossil fuel is burned. Consider options for reducing SO2 and NOx emissions 14  Using coal containing less sulfur, washing the coal, and using devices called scrubbers to chemically remove the SO2 from the gases leaving the smokestack. Power plants can also switch fuels; for example, burning natural gas creates much less SO2 than burning coal. Catalytic converters reduce NOx emissions from cars, and it is important to keep them working properly. Use alternative energy sources: these are energy that sources not popularly used and is usually environmentally sound. They are other sources of electricity besides fossil fuels, also known as renewable energy. They include: hydropower, wind energy, geothermal energy, solar energy, biomass, wave and tidal energy. There are also alternative energies available to power automobiles, including natural gas-powered vehicles, battery-powered cars, fuel cells, and combinations of alternative and gasoline powered vehicles. Take action as individuals: Individuals can contribute directly by conserving energy, since energy production causes the largest portion of the acid deposition problem. For example, you can:  Turn off lights, computers, and other appliances when not in use.  Use energy efficient appliances such as lighting, air conditioners, heaters, refrigerators, washing machines, etc.  Keep your thermostat at 68 ᵒF in cold season or and 72 ᵒF in warmer season.  Reduction of carpool by using public transportation, or better yet, walk or bicycle whenever possible Global Warming Global warming is primarily a problem of too much carbon (iv) oxide (CO2) in the atmosphere, which acts as a blanket, trapping heat and warming the planet. Furthermore, as fossil fuels/coal, oil and natural gas are burnt for energy; as forests are cut down and burnt to create pastures and plantations, carbon accumulates and overloads the atmosphere. In addition, certain waste 15 management and agricultural practices aggravate the problem by releasing other potent global warming/greenhouse gases such as methane and nitrous oxide. Some greenhouse gases occur naturally in the atmosphere, while others result from human activities. Naturally occurring greenhouse gases include water vapor, carbon dioxide, methane, nitrous oxide, and ozone. Certain human activities, however, add to the levels of most of these naturally occurring gases such as CO2, methane (emitted during the production and transport of coal, natural gas, and oil. Methane emissions also result from the decomposition of organic wastes in municipal solid waste landfills, and the raising of livestock), nitrous oxide, Hydrofluorocarbons (HFCs), perfluorocarbons (PFCs), and sulfur hexafluoride (SF6). These compounds are potent greenhouse gases. Sulfur hexafluoride, itself, is the most potent greenhouse gas that has been evaluated. Furthermore, CO2 levels in the atmosphere have risen substantially, thereby increasing their global warming effect. Possible Effects of a Warmer World Changes in food production: Reductions in biodiversity: Plants and animals generally react to consistently warmer temperatures by moving to higher latitudes and elevations. Recent studies reveal that some species have already started to shift their ranges, consistent with warming trends. Many populations and species may become more vulnerable to declining numbers or extinction if warming occurs faster than they can respond or if human development presents barriers to their migration. Rise in sea level: Warmer temperatures increase melting of mountain glaciers and cause ocean water to expand. Largely as a result of these effects, global sea level has risen 4 to 10 inches over the past 100 years. More extreme weather: this will lead to climate change that will result in more hurricanes, floods, and droughts. 16  Threats to human health: As temperatures rise, disease-carrying mosquitoes and rodents move into new areas, infecting people in their wake. Global warming will likely put as much as 65% of the world's population at risk of infection—an increase of 20%. Approaches to Slowing Global Warming Cut fossil fuel use:  Car makers could intensively increase the fuel economy of their cars and trucks.  Converting electric utilities to burn cleaner natural gas instead of using coal to produce electricity. Improve energy efficiency: Energy efficiency is the cleanest, safest, most economical way to begin to curb global warming. This can be achieved by:  Installing the best current technology in our cars and light trucks, home appliances and power plants.  Also, by saving energy in our homes and office buildings using energy efficient lighting, heating and air-conditioning system. This could keep millions of tons of carbon dioxide out of our air each year. Reduce deforestation and plant more trees:  Terrestrial ecosystems offer an opportunity to absorb and store (sequester) a significant amount of carbon dioxide from the atmosphere. By planting trees, preserving forests, and changing cultivation practices to increase soil carbon, for example, it is possible to increase the size of carbon sinks. Slow human population growth: by encouraging population growth control measures References https://www.vedantu.com/chemistry/effects-of-environmental-pollution UNIT III 17  Soil pollutants and pollution Learning objectives: To acquaint the students with the terms in soil pollution To enlighten the students on different causes of soil pollution To understand the sources of soil pollution To familiarize students with the effects of soil pollution To acquaint them with the different measures to soil pollution control Soil Pollution Definitions:  Soil pollution is defined as the "adding of substances to the soil that has a negative impact on the physical, chemical, and biological aspects of the soil and lowers its productivity."  It's a build-up of poisonous compounds, chemicals, salts, radioactive materials, or disease- causing agents in the soil that harm plant development and human and animal health.  Soil pollution can also be defined as when the levels of contaminants in soil surpass the levels that should be present naturally. This is because soil naturally contains a few harmful chemicals however, the amounts of contaminants found naturally in soil are not high enough to constitute a risk. Sources of Soil Pollution Industrial Wastes The untreated discharge of industrial pollutants into the soil causes soil pollution. It contains a high level of hazardous pollutants, resulting in soil pollution. Industrial wastes contain varying amounts of harmful substances and dangerous compounds, which, when deposited in soil, influence the topsoil layer strength. Soil fertility is influenced by the mining of minerals from the earth. The by-products are contaminated, whether iron ore or coal, and they are disposed of in an unsafe manner. 18   The three categories of industries that generate the most industrial waste are metallurgy, non-metallurgy, and food processing.  The waste generated varies in every industry.  It has high toxic contaminants such as dirt and gravel, masonry and concrete, scrap metal, oil, solvents, chemicals, scrap lumber, and even vegetable matter from restaurants.  Dumping of these industrial wastes into the environment without being treated, would result in major environmental issues.  This will surely reduce soil productivity and have a negative impact on agricultural production in the surrounding area. Agriculture Agricultural activities are the most common cause of soil pollution. To increase agricultural productivity, farmers frequently employ fertilizers and very toxic pesticides to clear their crops of insects, fungi, and bacteria. Long-term usage of these insecticides and pesticides can lead to soil pollution. They are packed with compounds that do not occur naturally and cannot be broken down by them. As a result, when they mix with water, they seep into the ground and gradually deplete the soil's fertility. Many of these pesticides are absorbed by plants, which then decompose and pollute the soil. i. Pesticides  Pesticides are toxic chemicals that are used to destroy pests.  Pesticides have the potential to alter the composition, diversity, and basic function of essential soil microorganisms.  Some pesticides have been shown to interfere with enzyme production by soil microbes, suppressing some while promoting others, affecting soil fertility, nutrient cycling, and metabolism.  Pesticides have also been shown to have an impact on larger animals that assist in preserving the soil's structure and fertility. ii. Fertilizers and Manures 19   Manures come from natural sources, whereas fertilizers are generated synthetically in factories.  However, the fertilizers and manures when applied in an inappropriate time and quantity and when they contain harmful chemicals will pollute the soil and affect its nature and composition drastically.  They can contribute to soil acidity and soil crust, lowering organic matter, humus content, and beneficial species, limiting plant growth, modifying soil pH, increasing pests, and even causing greenhouse gas emissions.  Nitrogen fertilizers applied in high quantities to fields over time destroy the equilibrium between the three macronutrients, N, P, and K, resulting in lower crop yields. Radioactive Wastes  Any material that is naturally radioactive or has been polluted by radioactivity and is assessed to have no further utility is considered radioactive waste.  Nuclear reactors, fuel processing plants, hospitals, and research centers all produce radioactive (or nuclear) waste.  Disposal into the ground has occasionally proven to be the most practical and simplest technique.  However, in many cases, the radionuclides enter the soil either directly or indirectly through water seeping via the soil and leaching pollutants from the surface of solid waste buried without adequate protection.  The soil and its associated pore become contaminated regardless of how the release happens. They lose their potential to produce high-quality agricultural products and are therefore categorized as degraded. Pollution due to Urban Activities  Urbanization and increasing population densities in cities, result in soil pollution due to urban activities. 20   Deforestation in urban areas occurs as a result of the construction of buildings, roads, and other structures, resulting in erosion and changes in land quality.  Pollutants are typically dispersed throughout cities or concentrated in industrial districts or landfills.  Regular construction, as well as a lack of effective waste disposal, can create severe soil degradation due to a lack of proper drainage and surface run-off. Pollution from domestic wastes  Household waste typically consists mostly of food waste that will gradually decompose. A collection of solid wastes in one place or scattered around is unsightly and might smell.  This produces a bad odour and attracts insects and rats, both of which contribute to the transmission of disease.  As the waste decomposes it produces a liquid called leachate which trickles down into the soil.  Leachate is a highly concentrated liquid pollutant that may contain toxic chemicals and pathogenic micro-organisms as well as high levels of organic compounds.  Rainwater falling on, and washing through solid waste adds to the problem. Effects of Soil Pollution On the Environment  Soil pollution is harmful to the ecosystem and has implications for its biotic component.  Improper agriculture methods that deplete soil organic matter can allow toxins to enter the food chain more easily.  For example, contaminated soil can percolate pollutants into groundwater, which accumulate in plant tissue and are subsequently passed on to grazing animals, birds, and humans that consume the plants and animals. On the Soil  A chain reaction occurs when soil pollution occurs. 21   It affects soil biodiversity, lowers soil organic matter, and reduces soil filtering capacity.  It also contaminates water held in the soil and groundwater, resulting in nutrient imbalances in the soil. On Agriculture  Soil pollution has an influence on food security because it impairs plant metabolism, lowering crop production.  They make crops dangerous for animals and humans to eat.  Pollutants also impair soil biodiversity and fertility by directly harming beneficial soil microbes and larger soil-dwelling species.  The soils' ability to cope with pollutants will also become too low. On Human Health  Contaminants from soil can get to humans through eating of soil (geophagia), inhalation, through the skin and indirect contact.  Pollutants in the soil, groundwater, and food supply can cause a wide range of diseases and deaths in humans.  Short-term health effects from exposure to such soils include headaches, coughing, chest pain, nausea, and skin/eye irritation.  Long-term exposure to contaminated soil can cause central nervous system depression, damage to essential organs (such as the liver, kidney), and even cancer.  Soil pollution can lead to spread of antimicrobial resistance genes from soil microbes to pathogens thereby affecting health by increasing human resistance to antimicrobials. Measures to Control Soil Pollution Reduce the use of Chemical Fertilizers  Although fertilizers could improve soil fertility at the right levels, too many of them actually damage the soil. 22   A surplus of chemical fertilizers may contaminate the soil in a number of ways. The soil's pH levels might be affected.  The beneficial microbes in the soil can potentially be wiped off. Additionally, the runoff from such soils pollutes the water as well.  Employing chemical fertilizers has a drawback therefore organic fertilizer may be considered as alternative. Encourage Reforestation and Afforestation Soil erosion, which is brought on by deforestation, is one of the key causes of soil pollution. Reforestation of a deforested area ought to be encouraged in order to stop this from happening. Recycle and Reuse Products  Recycle and reuse of products ensure that soil pollution is decreased while simultaneously reducing waste generation.  Currently, a considerable amount of the waste that is produced is made of plastic. These wastes are typically buried in landfills.  These plastics and other items degrade slowly in these landfills and emit hazardous substances into the soil.  These poisonous compounds are a significant cause of soil contamination and are extremely detrimental to the health of the soil.  Preventing more waste from being disposed of in these landfills can be done by reusing and recycling items, which would lessen soil degradation. UNIT IV IMPACT ASSESSMENT OF MICROBIAL CONTAMINANTS: AIR Learning objectives To be able to differentiate between passive and active air sampling 23 To be conversant with different approach to assessing microbial air contaminants To understand the different methods of enumerating air microbes Poor indoor air can have both immediate and long-term health effects, such as Sick building syndrome (SBS) and Building-related illnesses (BRIs). SBS refers to a collection of symptoms reported by the occupants or workers of a given building and is generally not attributed to a particular cause. These may include immediate or short-term effects such as irritation of the eyes, nose, and throat, headaches, dizziness, fatigue, affect the nervous and cardiovascular systems and cause reduced fertility and congenital disabilities in the long term. BRI’s on the other hand refers to medical conditions such as hypersensitivities, asthma, and respiratory infections such as pneumonia, linked to a specific cause. Generally, exposure to microbes or their components is associated with three main groups of illness: toxicity, infections, and allergic reactions, including respiratory infections and other related diseases. Neonates, young children, older adults, and especially people suffering from co-morbid conditions are highly vulnerable to IAP. A. Sampling of Air Through air sampling, it is possible to evaluate microbial contamination in environments at high risk of infection. Various kinds of passive and active sampling techniques are widely and routinely used for the assessment of microbial contamination of indoor air. Passive sampling Passive sampling is performed by using ‘agar settle plates’. This sampling method is called Koch's sedimentation/Gravitation method. The air sample is collected under the force of gravity onto exposed Petri plates, the plates are incubated, and results are expressed as CFU/m2. In this procedure the optimal duration of exposure should give a significant and readily countable number of well isolated colonies, for example about 30-100 colonies. Usually it depends on the dustiness of air being sampled. In occupied rooms and hospital wards the time would generally be between 10 to 60 m. During sampling it is better to keep the plates about 1 metre above the ground. Immediately after exposure for the given period of time, the plates are closed with the lids. Then the plates are incubated for 24 hrs at 37°C for aerobic bacteria and for 3 days at 22°C for saprophytic bacteria. For molds incubation temperature varies from 10-50°C for 1-2 weeks. After 24 incubation the colonies on each plate are counted and recorded as the number of bacteria carrying particles settling on a given area in a given period of time. The passive method has been standardized by the Index of Microbial Air contamination (IMA). The standard IMA for the measurement of microbial air contamination in environments at risk quantifies the microbial flow directly related to the contamination of surfaces coming from microbes that reach critical points by falling on to them. The index of microbial air contamination is based on the count of the microbial fallout on to Petri dishes left open to the air according to the 1/1/1 scheme (for 1h, 1m from the floor, at least 1m away from walls or any obstacle). The index of microbial air contamination has proved to be a reliable and useful tool for monitoring the microbial surface contamination settling from the air in any environment. The IMA can be expressed also as CFU/m2/time. Advantages of Koch’s sedimentation (gravitation) method  Koch’s sedimentation (gravitation) method gives comparable results  Requires no special powered instruments or personnel  It is not influenced by engineering factors.  It provides a valid risk assessment if passive sampling is performed in an operation theater or near a surgical site. Limitation of Koch's sedimentation (gravitation) method Though the method has the advantage of simplicity, it has certain limits. In this method only the rate of deposition of large particles from the air, not the total number of bacteria carrying particles per volume, is measured. Growth of bacteria in the settled particles may be affected by the medium used since not all microorganisms are growing well on all media. Moreover, since air currents and any temporary disturbances in the sampling area can affect the count, many plates have to be used. Active sampling  Active sampling is based on sampling techniques such as filtration, impingement, impaction, and centrifugation, each with its own sets of advantages and limitations. This approach collects airborne microorganisms present in inhalable dust in the indoor 25  environment. In this, an air sampler physically draws a measured volume of air with the help of a pump, through or over a particle collection device into a liquid or solid culture medium or a nitrocellulose membrane. The microbial count is given as colony forming units per cubic metre air (cfu/m3 of air). There are various types of active air samplers such as Anderson, Active Casella slit, Surface air system, and Reuter centrifugal air samplers that are available commercially. This sampling technique is mainly applicable when the concentration of microorganisms is not very high, such as in an operating theatre of a hospital. Limitation Some disadvantages of active sampling methods such as the impact method are:  A decline in the microbes’ viability caused by the shock of a sudden collision with culture media  Possibility of the nutrient culture getting overgrown in cases of high air pollution.  The methods are usually not cheap.  Active samplers require trained operators and a power supply, which might constrain their use in remote areas. B. Enumeration of Microorganisms in Air Various methods commonly applied for enumeration and detection of microorganisms can be subdivided into: Microscopic methods Culture methods Combination of both 1. Microscopic Methods These consist of: Letting air through a membrane filter or placing a glass coated with a sticky substance (e.g. vaseline), in the path of air Staining of the trapped microorganisms with acridine orange and examination and 26  Microscopic testing consisting of cell counting under a fluorescence microscope The final result is given as a total number of microbes in 1 m3 of air. The advantage of this method is that it allows the detection of live and dead microbes in air, as well as those, which do not abundantly flourish in culture media. Due to this, the number of microbes determined is usually higher by one order of magnitude than in culture methods. In addition, it is possible to detect and identify other biological agents e.g. plant pollen, allergenic mites, abiotic organic dust (fragments of skin, feathers, plants, etc.). However, the method has a serious drawback: inability to determine the species of microbes (bacteria, fungi, viruses). 2. Culture Methods These methods consist of transferring microbes from air onto the surface of the appropriate culture medium. After a period of incubation at optimal temperature, the colonies formed are counted and the result is given as colony forming units per cubic metre air (cfu/m3 of air). Because a colony can form not only from a single cell, but also from a cluster of cells, the air may contain more microbes than suggested by the cfu result. Besides, the method allows the detection of only the cells that are viable and those which are able to grow upon the medium used. Microbes transferred to the culture medium require resuscitation as they were subjected to the influence of unfavourable conditions. Therefore, it is recommended to supplement the culture with components such as betaine and catalase. 3. Combination of both However, testing of viruses differs significantly from the methods utilized for other organisms because: They may develop only in living cells, therefore they require tissue cultures (e.g. the epithelium of human trachea or monkey's kidney) or, in the case of bacteriophages, bacterial cultures, Species identification of detected viruses is meticulous and, among other things, consists of performing electrophoresis or utilizing antiserum that contains antibodies of common viruses, 27  Drawing large quantities of air is essential (over 1000 dm3, at least one order of magnitude higher than in the case of bacteria), as the amount of viruses in air is rather small (this especially concerns the enteroviruses). After transferring the viruses onto the surface of a single-layer culture, the viruses penetrate the cells, reproduce in them, and after their destruction attack the neighboring cells. Consequently, the areas around the initial places of the cell infections get cleared of cells – this clearing is called plaques. Therefore, the number of viruses detected is given as the number of units that form the plaques, in short pfu/m3 (plaque forming units). It has to be pointed out though, that such a method only allows the detection of viruses capable of infecting the utilized cells. C. Control of microorganisms in the air In order to reduce bio-aerosol loads in indoor environments, certain control measures can be followed. Several methods may be used to reduce the number of microorganisms in the air. These include physical treatments and chemical agents or a combination of both. Filtration This is one of the most used methods of producing sterile air with higher assurance of sterility. Filtration is the removal of particles, including microorganisms, from the air. Air filtration has greatest particle potential of all the separation methods. Depth filters are made of cellulose, glass fiber mixtures with resin, or acrylic binders. Laminar air flow The use of laminar air flow with high efficiency particulate air (HEPA) filters is highly recommended at critical points (35,36,40). HEPA filters remove 100% of particles that are greater than 0.3µm in size and consequently will remove all viable bacteria. Bactericidal Aerosols Periodical use of disinfectants and biocides is one of the methods to ensure controlled bio-aerosol concentrations. Air in the operating rooms and other critical areas like isolation rooms can be disinfected by fumigation using various microbicidal agents. Ultra-violet (UV) radiation 28 Ultra-violet radiation can be used to decrease airborne microflora, but its use is limited by worker safety. Solar heating Solar heating, which can result in a 7-log reduction in viable bacteria over 6 hours, is a possible disinfection consideration, for developing countries. UNIT V IMPACT ASSESSMENT OF MICROBIAL CONTAMINANTS: SOIL Soil monitoring The main objective of soil monitoring is to prevent and lessen contamination by substances with the potential to exert an adverse effect on the soil itself, and on air, water and organisms that may contact the soil. Assessment of soil pollutants In determining the relative risk posed by chemicals or contaminants in soils, one must also address the risk-assessment pattern, the pathways by which human health and the environment can be affected, and the availability of the released chemical for transport and adverse impact. Environmental risk is defined as the likelihood of injury, disease, death, or adverse impact resulting from human or environmental exposure (real or potential) to chemicals under site specific circumstances. The negative biological effects of pollutants present in all kinds of environmental samples can be assessed using different living organisms or cells as ‘analytical devices’. The biological response following the exposure of living organisms or cells to such environmental samples gives an information on toxicity, genotoxicity, estrogenicity, etc. According to the technical principle, methods of biological monitoring can be classified into: Bioassays, biosensors, immunoassays, estrogenicity test and ecological methods, but there also exist other types of classification, for example, division to biomarkers, whole cell biotests and early warning biological systems. 29 Bioassay or ecotoxicity assay is an experiment in which living test-species are exposed directly to an environmental sample such as surface water, ground water, wastewater, sediments and soil, or extracts of environmental sample to measure a potential biological effect due to the presence of potential contaminants. Microbial bioassays can roughly be divided as (general) toxicity assays and genotoxicity assays. The purpose of ecotoxicity bioassays is to assess the significant effect of an environmental sample on general physiological state of the test-species, while genotoxicity tests specifically show the effects resulting in changes of genetic material. Bioassays can be subdivided into acute and chronic, based on the exposure time of the test-species to the sample under investigation. The former is where exposure time does not exceed 96h, while the latter subgroup includes tests with longer exposure time. Acute bioassay calculates EC50 and LC50 where; EC50 is an estimated toxicant concentration in a sample at which 50% of the test organisms show an effect following a given exposure time. LC50 is an estimated toxicant concentration in a sample at which 50% of the test organisms die following a given exposure time. Chronic bioassay test parameter is: NOEC (No-Observed-effect concentration) value which is the highest toxicant concentration at which no significant effect can be detected when compared to the control sample. LOEC (Lowest-Observed-Effect Concentration) value is the lowest tested toxicant concentration that is significantly different from the control, the lowest concentration where the effects observed in the treated group imply an adverse effect on the subject. Most common parameters measured by microbial toxicity assays are population growth, substrate consumption, respiration, ATP luminescence and bioluminescence inhibition. Examples of bioassays using bacteria include Vibrio fischeri bioluminescence inhibition; inhibition of β- galactosidase activity in E. coli, and in Bacillus sp.: dehydrogenase activity assay. Other bioassays example is growth inhibition of green microalgae Scenedesmus supspicatus (a unicellular algae) following 72h exposure to pollutant chemical. 30 Assessment of soil microbial contaminants A range of bacterial pathogens, introduced through contaminated irrigation water or manure, are capable of surviving for long periods in soil and water where they have the potential to contaminate crops in the field and create some potential public health problems due to direct interaction with microbial contaminated soil. Assessment of the effect of these pathogens on soil may not be direct. Rather, a correlation between microbial contamination and registered occurrences of contagious diseases among hosts can be established. UNIT VI WASTES DISPOSAL AND MANAGEMENT Introduction A clear understanding of the quantities and characteristics of the waste being generated is a key component in the development of robust and cost-effective solid waste management strategies. Learning objectives: To identify different classifications and types of wastes. To understand the approach to an effective solid waste management. To familiarize the students with the key components of solid waste management. Definition: Waste can be defined as unwanted or undesired material leftover after the completion of a process. It is also any substance or object which the holder discards or intends to discard. A waste product is regarded as a pollutant when it damages the environment. However, pollutants are generally wastes but all wastes are not pollutants. Wastes can be classified in several forms, depending on the point of consideration: 31 In terms of form, they classified as solid, liquid and gaseous wastes. In terms of source, there are municipal, industrial and biomedical wastes. According to nature wastes may be grouped as biological, chemical or physical. In terms of properties, there are biodegradable and non-biodegradable. Biodegradable wastes are typically from plants or animal sources, which may be broken down by other living organisms. With proper treatment biodegradable wastes can be used for composting, animal feed, or converted to energy. Common types are food waste, garden waste, paper and cardboard wastes and biodegradable plastics. Non-biodegradable are wastes that cannot be broken down by other living organisms e.g. plastics, styrofoam (polystyrene), metal, glass, some chemicals and toxins. In terms of treatment wastes may grouped into biodegradable, recyclable and inert (e.g. construction and demolition waste, rocks, dirt, debris, etc.). In terms of safety, there are hazardous and non-hazardous waste. SOLID WASTES Solid wastes are defined as unwanted/undesirable organic and inorganic leftover materials produced by various activities of the society and which have lost their value to the first user. These wastes come from human and animal activities that are normally discarded as useless or unwanted. They are generated from several sources such as domestic wastes, commercial wastes, institutional wastes and industrial wastes. Solid waste can be grouped according to their source as Municipal solid waste, Industrial solid waste and Biomedical solid waste. Municipal solid waste: this usually includes all of wastes generated in a community, wastes from domestic wastes, commercial wastes, institutional wastes and building materials wastes. Industrial solid waste: is the generated by manufacturing or industrial processes. This types include cafeteria garbage, trash, masonry and concrete, scrap metals, dirt and gravel, oil, chemicals, solvents, wood and scrap lumber, weed grass and trees, etc. Industrial solid waste which may be solid, liquid, gaseous or sludges held in containers could be hazardous and non-hazardous waste. 32 Hazardous waste: is a waste with properties that make it dangerous or capable of having a harmful effect on human health or the environment. Hazardous waste is generated from many sources, ranging from industrial manufacturing process wastes, some commercial products such as batteries, cleaning fluids, paints or pesticides discarded by commercial establishments or individuals. Non-hazardous wastes are any waste that causes no harm to human or environmental health, but may still be subject to certain management requirements. Biomedical solid waste: this is also known as infectious or medical waste and is defined as solid waste generated during diagnosis, testing, treatment, research or production of biological products for humans or animals. SOLID WASTE DISPOSAL AND MANAGEMENT Solid waste management is the collection, transport, processing, disposal, managing and monitoring of waste materials. The term usually relates to materials produced by human activity, and the process is generally undertaken to reduce their effect on health, the environment or aesthetics. For an effective solid waste management in an affected area, the following steps are to be considered:  Identify the types of waste  Identify the sources of waste  Determine the potential health hazards from waste  Determine the volume of waste generated  Identify safe collection method(s)  Identify safe transportation method(s)  Identify safe disposal methods However, the key components of solid waste management are generation, storage, collection, transportation and disposal. 33 i. Generation of solid waste: this is the stage at which materials become valueless, of no use to the owners and wish to get rid of them. Items considered to be valueless to an individual may not necessarily be so to another. ii. Storage: is the system of keeping materials after they have been discarded, and before collection and final disposal. Storage may not be necessary where people practice on-site disposal such as discarding of items directly into family pits. iii. Collection: solid waste collection refers to how the waste is collected for transportation to the final disposal site. Collection system should be carefully planned as to ensure the storage facilities are not overloaded. iv. Transportation: this is the stage when solid waste is transported to the final disposal site. Types of transportation are human-powered, animal-powered and motorised. v. Disposal: a safe disposal is the final stage of solid waste management where associated risks are minimized. There are four main methods of disposal: Land application- burial or landfilling- waste is placed in a large excavation (pit or trench) in the ground, which is back-filled with excavated soil each day waste is tipped. It is a sanitary disposal method if properly managed but limited by requirement for reasonably large area. Composting- composting of vegetables and other organic wastes can be applied in many situations, organic wastes can be dug into the soil to add humus and fertilizer. Advantaged by being environmentally friendly and beneficial for crops, but limited by the need for intensive management and experienced personnel for large-scale operations. Burning or incineration- often used for combustible waste disposal, and should only be done off- site or a considerable distance downwind of dwellings. May be used to reduce waste volume and appropriate where there is limited space for burial and landfill. Advantages are it reduces volume of waste and appropriate in off-site pits to reduce scavenging. Limited by production of smoke and possibility of fire hazard. Recycling (resource recovery)- plastics, bags, containers, tins and glass are often recycled. In most developing countries there exists a strong tradition of recycling leading to lower volumes of waste 34 than in many more developed societies. Advantages is that it is environmentally friendly but constrained by limited potential in most emergency situations, and is expensive to set up. UNIT VII DOMESTIC WASTES AND WASTE WATER TREATMENT Learning objectives To understand the concept of wastewater To familiarize the students with wastewater parameters testing To understand the purpose of wastewater treatment To identify the stages in wastewater treatment process Wastewater is any water that has been adversely affected in quality by anthropogenic influence and comprises liquid waste discharged by domestic residences, commercial properties, industry, and/or agriculture and can contain a wide range of potential contaminants and concentrations of organic matter from human faeces, urine and gray water. Gray water results from washing, bathing and meal preparation. Water from various industries may also enter the system. People excrete 100 – 500 grams wet weight of faeces and 1– 1.3 litres of urine per person per day. Sewage is the part of wastewater that is contaminated with feces or urine, but when the term is used as wastewater, sewage refers to wastewater from sources including domestic, municipal, or industrial liquid waste products disposed of, usually via a pipe or sewer system. Untreated sewage may contain water; nutrients (nitrogen and phosphorus); solids (including organic matter); pathogens (including bacteria, viruses and protozoa); helminthes (intestinal worms and worm-like parasites); oils and greases; runoff from streets, parking lots and roofs; heavy metals (including mercury, lead, copper) and many toxic chemicals including polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons (PAHs), pesticides, phenols and chlorinated organics. The cloudiness of sewage is caused by suspended particles, which in untreated wastewater, is from 100 to 350 mg/l. Organic components (which also measures the strength) of wastewater is determined by three major parameters: 35  (i) Total Organic Carbon (TOC): specifies the amount of carbon contained in organic compounds. It is determined by oxidation of the organic matter with heat and oxygen, followed by measurement of the carbondioxide liberated with an infrared analyzer. (ii) Chemical Oxygen Demand (COD): is the amount of oxygen required to oxidize all organic compounds chemically completely to carbondioxide and water. COD is measured by oxidation with potassium dichromate (K2Cr2O7) in the presence of sulphuric acid and silver. (iii) (iii) Biochemical Oxygen Demand (BOD): is the amount of dissolved oxygen (DO) consumed by organisms during the biochemical oxidation of organic (Carbonaceous compounds) and inorganic matter. The 5-day BOD test (BODs) is a measure of the amount of oxygen (O2) consumed by a mixed population of heterotrophic bacteria in the dark at 20℃ over a period of 5 days. The major objective of domestic waste water treatment is to ensure that water released into the environment do not pose environmental and health hazards. This is done by the reduction or removal of the pollutants which may be either in form of solids (suspended or soluble matter), and kill disease causing organisms. The essential goal of wastewater treatment is the removal and degradation of organic matter under controlled conditions. The Wastewater Treatment Process Sewage treatment is grouped into 3 major steps. Primary treatment Secondary treatment Tertiary treatment 36 Fig. 1. Stages in wastewater treatment (8 Wastewaster Treatment Process Steps & Stages - Cole- Parmer (coleparmer.com) PRIMARY TREATMENT Stage One — Bar Screening: Removal of large items from the influent to prevent damage to the facility’s pumps, valves and other equipment. The process of treating and reclaiming water from wastewater (any water that has been used in homes, such as flushing toilets, washing dishes, or bathing, and some water from industrial use and storm sewers) starts with the expectation that after it is treated it will be clean enough to reenter the environment. Stage Two — Screening: Removal of grit by flowing the influent over/through a grit chamber. Fine grit that finds its way into the influent needs to be removed to prevent the damage of pumps and equipment downstream (or impact water flow). Too small to be screened out, this grit needs to be removed from the grit chamber. Stage Three — Primary Clarifier: Initial separation of solid organic matter from wastewater. Solids known as organics/sludge sink to the bottom of the tank and are pumped to a sludge digester or sludge processing area, dried and pulled away. After grit removal, the influent enters large primary clarifiers that separate out between 25% and 37 50% of the solids in the influent. The solids that fall to the bottom of the clarifier are known as sludge and pumped out regularly to ensure it doesn’t impact the process of separation. The sludge is then discarded after any water is removed and commonly used as fertilizer. SECONDARY TREATMENT Stage Four — Aeration: Air is pumped into the aeration tank/basin to encourage conversion of ammonia to nitrate (NH3 to NO3) and provide oxygen for bacteria to continue to propagate and grow. Once converted to NO3, the bacteria remove/strip oxygen molecules from the nitrate molecules and the nitrogen (N) is given off as nitrogen gas (N2↑). The primary function of the aeration tank is to pump oxygen into the tank to encourage the breakdown of any organic material, and the growth of the bacteria, as well as ensure there is enough time for the organic material to be broken down. Dissolved oxygen monitoring at this stage of the plant is critical. Stage Five — Secondary Clarifier: Treated wastewater is pumped into a secondary clarifier to allow any remaining organic sediment to settle out of treated water flow. As the influent exits the aeration process, it flows into a secondary clarifier where, like the primary clarifier, any very small solids sink to the bottom of the tank. These small solids are called activated sludge and consist mostly of active bacteria. Part of this activated sludge is returned to the aeration tank to increase the bacterial concentration, help in propagation, and accelerate the breakdown of organic material. The excess is discarded. TERTIARY TREATMENT Stage Six — Chlorination (Disinfection): Chlorine is added to kill any remaining bacteria in the contact chamber. With the enhanced concentration of bacteria as part of the aeration stage, there is a need to test the outgoing effluent for bacteria presence or absence and to disinfect the water. This ensures that higher than specified concentrations of bacteria are not released into the environment. Chlorination is the most common and inexpensive type of disinfection but ozone and UV disinfection are also increasing in popularity. Stage Seven — Water Analysis and Testing: Testing for proper pH level, ammonia, nitrates, 38 phosphates, dissolved oxygen, and residual chlorine levels to conform to the plant’s permissible discharge limit. Although testing is continuous throughout the wastewater treatment process to ensure optimal water flow, clarification and aeration, final testing is done to make sure the effluent leaving the plant meets permit specifications. Plants that do not meet permit discharge levels are subject to fines and possible incarceration of the operator in charge. Stage Eight — Effluent Disposal: After meeting all permit specifications, clean water is reintroduced into the environment. Final testing is done to make sure the effluent leaving the plant meets permit specifications. Plants that do not meet permit discharge levels are subject to fines and possible incarceration of the operator in charge. UNIT VIII PRINCIPLES AND STANDARDS OF SANITARY WATER QUALITY Determination of sanitary quality of domestic water Natural waters are subject to important changes in their microbial quality that arise from agricultural use, discharges of sewage or wastewater resulting from human activity or storm water runoff. Sewage effluents contain a wide variety of pathogenic microorganisms that may pose a health hazard to the human population when the effluents are discharged into recreational waters. The density and variety of these pathogens are related to the size of the human population, the seasonal incidence of the illness, and dissemination of pathogens within the community. Learning objectives: To familiarize the students with the concept of indicator organisms To understand the types and implication of indicator organism in water To be acquainted with different approaches to water quality testing Indicator organisms 39 Many waterborne pathogens are difficult to detect and/or quantify and the specific methodology to detect them in environmental water samples has still to be developed. Indicator microorganisms are used to predict the presence of and/or minimize the potential risk associated with pathogenic microbes. Indicator organisms are useful in that they circumvent the need to assay for every pathogen that may be present in water. Ideally, indicators are nonpathogenic, rapidly detected, easily enumerated, have survival characteristics that are similar to those of the pathogens of concern, and can be strongly associated with the presence of pathogenic microorganisms. Rationale for the use of indicator organisms The key criteria for ideal bacterial indicators of faecal pollution are:  They should be universally present in large numbers in the faeces of human and other warm-blooded animals.  They should also be present in sewage effluent, be readily detectable by simple methods.  They should not grow in natural waters.  They should also be of exclusive faecal origin and be present in greater numbers than faecally transmitted pathogens. No single indicator organism fulfills all these criteria, but the member of the coliform group that satisfies most of the criteria for the ideal indicator organism in temperate climates is E. coli. The presence of E. coli in a sample of drinking water may indicate the presence of intestinal pathogens. The absence of E. coli cannot be taken as an absolute indication that intestinal pathogens are also absent. Other Indicator organisms include Enterococci and spores of sulphite-reducing clostridia, typified by Clostridium perfringens. Enterococci do not multiply in the environment and can occur normally in faeces. Numbers of enterococci in humans are greatly outnumbered by E. coli bacteria. Coliform bacteria Coliform testing results, together with other information obtained from engineering or sanitary surveys, provide the best assessment of water-treatment effectiveness and the sanitary quality of source water. 40 Coliform bacteria are defined as facultatively anaerobic, Gram-negative, non-spore-forming, rod-shaped bacteria that ferment lactose to produce acid, gas, or both in the presence of bile salts within 48 h at 35 °C, possess the enzyme β-galactosidase and are oxidase-negative. Coliform bacteria belong to the family Enterobacteriaceae and share similar cultural characteristics. Typical of these genera encountered in water supplies are Citrobacter, Enterobacter, Escherichia, Hafnia, Klebsiella, Serratia and Yersinia. Thermotolerant (Faecal) coliform bacteria Thermotolerant coliform bacteria sometimes referred to as faecal coliform possess the characteristics of coliform bacteria but are able to carry out lactose fermentation with production of gas and acid at 44.5 ± 0.2°C within 48 hours. For this reason, the term “thermotolerant coliforms” rather than “faecal coliforms” is a more accurate name for this group. Thermotolerant coliforms include strains of the genera Klebsiella and Escherichia. Certain Enterobacter and Citrobacter strains are also able to grow under the conditions defined for thermotolerant coliforms. E. coli is, however, the only biotype of the family Enterobacteriaceae that is almost always faecal in origin. Therefore, the thermotolerant coliform group when used should ideally be replaced by E. coli as an indicator of faecal pollution. Total coliform comprises bacteria of faecal origin, thermotolerant coliform, and some other bacteria from environmental sources. Hence, incidence of total coliforms may or may not suggest faecal contamination but may be due to contaminants from soil or organic matter. Escherichia coli (E. coli) E. coli has long been used as an indicator of fecal pollution. E. coli is a coliform bacterium and has historically been regarded as the primary indicator of faecal contamination of both treated and untreated water. Most of the E. coli strains possess the enzyme β-glucuronidase, which can be detected using specific fluorogenic or chromogenic substrates. For the purpose of water testing, most E. coli can be confirmed by a positive indole test and by their inability to use citrate (as the only carbon source) in the culture medium. Alternatively, E. coli can be distinguished easily enzymatically by the lack of urease or presence of β-glucuronidase enzymes. Intestinal enterococci (Faecal streptococci and enterococci) 41 Intestinal enterococci are defined as Gram-positive cocci that tend to form in pairs and chains. They are non-spore forming, oxidase-negative, catalase-negative, possess Lancefield’s Group D antigen and hydrolyse aesculin. They can grow aerobically and anaerobically in the presence of bile salts, and in sodium azide solutions, concentrations of which are inhibitory to coliform bacteria and most Gram-negative bacteria. Enterococcus faecalis and some related species can reduce 2,3,5 triphenyltetrazolium chloride to the insoluble red dye, formazan. The taxonomy of this group, comprise species of two genera Enterococcus and Streptococcus. Although several species of both genera are included under the term enterococci, the species most predominant in polluted aquatic environments are Enterococcus faecalis, E. faecium and E. durans. Faecal streptococci have received widespread acceptance as useful indicators of faecal pollution in natural aquatic ecosystems. These organisms show a close relationship with health hazards (mainly for gastrointestinal symptoms) associated with bathing in marine and freshwater environments. They are not as ubiquitous as coliforms, they are always present in the faeces of warm-blooded animals, and it is believed that they do not multiply in sewage-contaminated waters. Clostridium perfringens C. perfringens is an enteric, gram-positive, anaerobic, spore-forming, pathogenic bacterium found in human and animal feces. Clostridium perfringens is a member of the sulphite-reducing clostridia which is non-motile and is capable of fermenting lactose, reducing nitrate and liquefying gelatin. Most clostridia are strictly anaerobic, but a few species are capable of limited growth in the presence of low levels of oxygen. Clostridium perfringens is the key species of the sulphite- reducing clostridia. It produces environmentally resistant spores that survive in water and in the environment for much longer periods than the vegetative cells of E. coli and other faecal indicators. Spores of C. perfringens are largely faecal in origin, they are always present in sewage (about 104 -105CFU/100 ml), they are highly resistant in the environment and appear not to reproduce in aquatic sediments (which appear to be the case with thermotolerant coliforms). It is important to note that spores of C. perfringens do not act as an indicator for non-sewage or animal faecal contamination in general, and therefore they are only suitable as indicator organisms for parasitic protozoa (e.g. Cryptosporidium parvum, Giardia lamblia) and viruses from sewage-impacted waters. Heterotrophic plate count 42 The heterotrophic plate count includes all of the micro-organisms that are capable of growing in or on a nutrient-rich solid agar medium. Two incubation temperatures and times are used: 37 °C for 24 hours to encourage the growth of bacteria of mammalian origin, and 22 °C for 72 hours to enumerate bacteria that are derived principally from environmental sources. The main value of colony counts lies in comparing the results of repeated samples from the same source. If levels increase substantially from normal values, there may be cause for concern. The rationale for enumerating heterotrophic plate counts has been to assess the general bacterial content of the water and to monitor trends or rapid changes in water quality. ANALYTICAL METHODS Coliform bacteria have long been used as water-quality indicators based on the premise that, because these organisms are present in the intestines of warm-blooded animals, their presence in water could indicate that recent fecal contamination has occurred. The standard test for the coliform group may be carried out by i). the multiple-tube fermentation technique or presence- absence procedure, ii). the membrane filter (MF) technique, or iii). the enzymatic substrate coliform test. Each technique is applicable within the limitations specified and with due consideration of the purpose of the examination. MOST PROBABLE NUMBER (MULTIPLE TUBE FERMENTATION TECHNIQUE) The MPN technique is generally conducted in three sequential phases (presumptive, confirmatory, and complete), each phase requiring 1 to 2 days of incubation. The results of the multiple fermentation tube test for coliforms is reported as a most probable number (MPN) index. This is an index of the number of coliform bacteria that, more probably than any other number, would give the results shown by the test. It is not a count of the actual number of indicator bacteria present in the sample. The MPN index is determined by comparing the pattern of positive results (the number of tubes showing growth at each dilution) with statistical tables. MPN test is completed in three steps: A. Presumptive test B. Confirmed test C. Completed test 43 Presumptive Test 1. Prepare MacConkey purple broth media of single and double strength in test tubes with Durham’s tube and autoclave it. 2. Take three sets of test tubes (totaling 15) containing five tubes in each set; one set with 10 ml of double strength (DS) other two containing 10 ml of single strength (SS). 3. Using sterile pipettes, transfer 10 ml of water to each of the DS broth tubes. Transfer 1 ml of water sample to each of 5 tubes of one set of SS broth and transfer 0.1 ml water to five tubes of remaining last set of SS broth tubes. 4. Incubate the tubes at 37°C for 24 hours. 5. After incubation, observe the gas production in Durham’s tube and the color change of the media. 6. Record the number of positive results from each set and compare with the standard chart to give presumptive coliform count per 100 ml water sample. Result Positive: The formation of 10% gas or more in the Durham tube within 24 to 48 hours, together with turbidity in the growth medium and the color change in the medium constitutes a positive presumptive test for coliform bacteria, and hence for the possibility of fecal pollution. Negative: No growth or formation of gas in Durham’s tube. Note: The test is presumptive only because under these conditions several other microorganisms other than coliforms also produce acid and gas from lactose fermentation. A. Confirmed Test Inoculation of the lactose-broth In order to confirm the presence of coliform, a confirmatory test is done. 44  1. For this, a loopful of suspension from a positive tube is inoculated into a 3 ml lactose-broth or brilliant green lactose fermentation tube 2. Incubate the inoculated lactose-broth fermentation tubes at 37°C and inspect gas formation after 24 ± 2 hours. 3. If no gas production is seen, further incubate up to a maximum of 48 ±3 hours to check gas production. Inoculation on agar plate (EMB agar or Endo Agar) or media slants 1. Take a loopful of suspension from a positive tube and inoculate it on EMB agar and nutrient agar slants. 2. The agar slants should be incubated at 37°C for 24± 2 hours. 3. Colonies must be examined macroscopically. Result Positive: Formation of gas in lactose broth and the demonstration of a coliform-like colony on the EMB agar indicate the presence of a member of the coliform group in the sample examined. Coliforms produce colonies with a greenish metallic sheen which differentiates them from non- coliform colonies (show no sheen). The presence of typical colonies at high temperatures (44.5 ±0.2) indicates the presence of thermotolerant E. coli. Negative: The absence of gas formation in lactose broth or the failure to demonstrate coliform- like colonies on the EMB agar. B. Completed Test 1. Transfer a typical coliform colony from the agar plate into a tube of brilliant green bile broth (BGLB) with Durham’s tube inside and on the surface of a nutrient agar slant. 2. Incubate at 35°C for 24 hours. 3. After 24 hours, check the broth for the production of gas, and perform Gram staining for organisms on the nutrient agar slant. 45 Positive: The presence of gas in the brilliant green bile broth tube and Gram-negative, non-spore- forming rods on NA slant constitutes a positive completed test for the presence of coliform bacteria, which, in turn, infer that a member of the coliform group is present. Negative: Absence of growth and gas formation in the broth. Absence of gram-negative, non- sporing rods on Gram staining. Fig. 2. Steps in multiple tube fermentation technique of water quality analysis Uses of MPN Test  It is commonly used in estimating microbial populations in soils, waters, food, agricultural products.  The technique is particularly useful with samples that contain particulate material that interferes with plate count enumeration methods.  It has also been suggested as a consideration for an alternate method to trend environmental monitoring studies.  It is also useful for counting bacteria that reluctantly form colonies on agar plates or membrane filters but grow readily in liquid media. Advantages of MPN Test  Ease of interpretation, either by observation or gas emission 46   Effective method of analyzing all types of water, highly turbid samples such as sediments, sludge, mud, etc.  Does not require correlation with other methods for use. Limitations of MPN Test  The method is to be considered only when other counting methods are inappropriate.  Laborious and expensive in terms of materials, glassware, and incubator space.  It has relatively a large margin of error.  Requires longer time for determination of results. MEMBRANE FILTRATION TECHNIQUE The membrane filtration (MF) technique is based on the entrapment of the bacterial cells by a membrane filter (pore size of 0.45 µm). After 100 mL of test water sample is filtered, the membrane is placed on an appropriate medium and incubated. Discrete colonies with typical appearance are counted after 24-48 hours, and the population density of the target bacteria, usually described as colony forming unit (cfu) per 100 ml in the original sample, can be calculated from the filtered volumes and dilutions used. This technique is more precise than the MPN technique. Limitations are however, it can only be used for low-turbidity waters with low concentrations of background micro-organisms; it cannot be used for chlorinated primary effluent, cannot be used with samples with toxic wastes and cannot be used for samples with large amounts of algae. UNIT IX DISEASE TRANSMISSION BY WATER Water associated diseases are caused by pathogenic microorganisms, most commonly transmitted in contaminated fresh water. Infection commonly results during bathing, washing, drinking, in preparation of food or by consumption of food that is infected. Waterborne disease can be caused by protozoa, viruses, bacteria and intestinal parasites. Learning objectives: 47 To be familiarized with differences in water associated diseases To understand the mode of transmission, symptoms and diseases of selected water related pathogens. Classification of Disease transmission by water Water associated diseases are classified according to their mode of transmission and the form of infection caused. They are classified as follows: i. Waterborne diseases ii. Water-washed diseases iii. Water-based diseases iv. Water-related diseases I. Waterborne diseases are caused by contamination of water by faecal matter and urine. Infection is by direct ingestion of water or indirectly by any ingestion of pathogens (e.g. food; Typhoid Mary). It results in widespread outbreak of disease amongst those using the same source of water. Carriers may be asymptomatic. Example: Cholera, infectious hepatitis, paratyphoid, tularaemia, typhoid, amoebic dysentery, bacillary dysentery. Weil’s diseases (leptospirosis) caused by skin contact with infected rat urine found in sewage, flood water and contaminated surface water. II. Water-washed diseases are caused by poor personal hygiene due to water shortages. Spread of infection is reduced by additional water supply. Microbial quality may be unimportant as it is used for cleaning/washing not ingestion. Diseases are normally skin infections, mucous membranes and eyes. Diseases are non-faecal in origin, (e.g. bacterial skin sepsis, scabies, and cutaneous fungal infections). Diseases also include those spread by fleas, ticks and lice (e.g. epidemic typhus, rickettsial typhus and louse-borne fever). Some water borne diseases can be contracted due to poor personal hygiene (e.g. shigellosis). Other examples include ascariasis, conjunctivitis, leprosy, skin sepsis. III. Water-based diseases: these are caused by pathogens that have complex life cycle requiring an aquatic intermediate host. Disease cannot be contracted by ingestion or contact with pathogen as excreted by infected person. They are all caused by parasitic worms with 48  severity of infection dependent on number of worms infecting host. Schistosomiasis, caused by trematode Schistosoma spp. has aquatic snails as intermediate host; the cercarias penetrate skin of humans and are normally spread via irrigation schemes. Eggs leave humans via urine. Over 200 million people get infected globally. Other examples: Dracunculus medimensis - guineaworm (nematode) - Cyclops spp. intermediate host. IV. Water-related diseases are pathogens carried by insect vectors living near water. Diseases are all severe and difficult to control: Viral diseases; Yellow fever – mosquito Aedes spp. Dengue – mosquito Aedes aegypti; Protozoan diseases Gambian sleeping sickness (trypanosomiasis) caused by river tsetse fly Glossina spp. Malaria caused by the protozoan Plasmodium spp. Carried by the vector mosquito Anopheles spp. Table 1. Waterborne diseases and their microbial agents Microbial Microbial agent Mode of transmission and Symptoms infection sources in water supply Cryptosporidiosis Cryptosporidium Oral. Collects on water Flu--like symptoms, watery parvum filters and membranes that diarrhea, loss of appetite, cannot be disinfected, substantial loss of weight, animal manure, seasonal bloating, increased gas runoff of water. stomach. Giardiasis Giardia lamblia Hand-to-mouth/Oral. Diarrhea, abdominal Untreated water, poor discomfort, bloating, disinfection, pipe breaks, flatulence and pains. leaks, groundwater contamination, campgrounds where humans and wildlife use same source of water Amoebiasis Entamoeba histolytica Sewage, non-treated Abdominal discomfort, drinking water, flies in fatigue, weight loss, water supply diarrhea, bloating, fever 49 Cyclosporiasis Cyclospora Sewage, non-treated cramps, nausea, vomiting, cayetanensis drinking water muscle aches, low--grade fever, and fatigue Dysentery Shigella, Salmonella Fecal-oral. Contaminated Frequent stooling with spp. food and drinking water blood and/or mucous and sometimes vomiting of blood Cholera Vibrio cholerae Fecal-oral/ Hand-to-mouth Diarrhea, (rice-water stool), vomiting Legionellosis Legionella spp. From warm aquatic Fever, chills, pneumonia (especially L. environment by inhalation (with cough), anorexia, pneumophila) of contaminated aerosols of muscle aches, malaise and water spray, mists of occasionally diarrhea and