Environmental Microbiology Lecture Notes

Environmental Microbiology Definitions

• Microbial Ecology: Study of microorganisms' interactions within their environment (air, water, soil), encompassing Eukaryota, Archaea, Bacteria, and viruses.

• Environmental Microbiology: Study of microbial community composition and physiology in environments like soil, water, air, and sediments, including artificial settings like bioreactors.

The Need to Understand Environmental Microbiology

1. Emergence of New Pathogens: Waterborne and food-borne pathogens pose threats to human and animal health.

2. Waste Disposal Practices: Past practices contaminate surface and groundwater with chemicals.

3. DNA Technology: PCR development allows for better microbe measurement and analysis.

   • These factors prompted scientists to assess the safety of food and water supplies and develop tools for microbe detection and identification.

Classification of Environment

1. Physical/Abiotic: External factors like air, water, and soil.

2. Living/Biotic: All living organisms such as plants, animals, and microorganisms.

Components of Ecosystem (Environment)

• A. Abiotic Components:

  • I. Climatic factors: Temperature, light level, and water.

  • II. Soil factors: Mineral matter, texture, organic matter, and organisms.

• B. Biotic Components:

  • I. Producers (Autotrophs): Green plants with chlorophyll.

  • II. Consumers (Heterotrophs): Organisms lacking chlorophyll (e.g., cow, rabbit).

  • III. Decomposers (Saprotrophs): Bacteria and fungi living on dead organic matter.

Important Terms in Ecology

• Abiotic Environment: Non-living ecosystem components like temperature and rainfall.

• Acclimation: Gradual physiological adaptation to environmental changes.

• Aerobic: Respiration and metabolism in the presence of oxygen.

• Anaerobic: Respiration and metabolism without oxygen.

• Autotroph: Organism capable of photosynthesis, producing organic molecules from inorganic ones (self-dependent in nourishment).

• Biogeochemical Cycles: Natural cycles of elements and components, converting inorganic to organic and vice versa.

• Biotic Factors: Environmental factors resulting from living organisms' activities.

• Consumer: Organism feeding on other organisms.

• Decomposer: Organism obtaining food by breaking down complex organic compounds into simpler inorganic compounds.

• Food Chain: Energy transfer from one organism to another, each occupying a trophic level.

• Habitat: Physical environment where an organism lives.

• Heterotrophs: Organisms incapable of photosynthesis, dependent on others for food.

• Lentic: Ecology of standing water (lakes, ponds).

• Lotic: Ecology of running water (rivers, streams).

• Productivity: Amount of energy stored in a living body in a specific area and time.

Microorganisms

• Organisms of microscopic or submicroscopic size, especially bacteria or viruses.

• A. Bacteria: Unicellular prokaryotes, important in recycling materials; some cause disease, others are useful.

• B. Fungi: Unicellular or multicellular eukaryotes (yeasts, molds, mushrooms); important recyclers, some produce antibiotics and alcohols, some are pathogenic.

• C. Algae: Unicellular, colonial, or filamentous photosynthetic organisms; responsible for organic molecule production in aquatic environments; include Diatoms (Crysophyta), Dinoflagellates (Pyrrophyata), Euglenoids (Euglenophyta), Green algae (Chlorophyata), Brown algae (Phaeophyta), and Red algae (Rodophyta).

• D. Protozoa: Unicellular, non-photosynthetic organisms in aquatic environments and wet soils; classified by motility (pseudopodia, cilia, or flagella).

Microbiological Terms

• Microbicide: Chemical that kills microbial cells and spores (bactericide, fungicide, algaecide, insecticide).

• Microbistatic: Chemical that inhibits microbial growth.

• Disinfection: Destruction of vegetative forms, not spores; used for inanimate objects as effective concentrations may be toxic to human tissues.

• Antiseptics: Destroy or inhibit microorganisms on skin and other tissues.

• Infection: Entry, establishment, and multiplication of pathogens within a host.

Aquatic Microbiology

• Aquatic ecosystems are communities of organisms dependent on each other and their environment in a body of water.

• Aquatic microbiology studies microscopic living organisms in fresh or saltwater systems, including viruses, bacteria, Actinomycetes, and fungi.

• Microorganisms in aquatic environments must adapt to a wide range of physical conditions like salinity, temperature, pressure, sunlight, and water clarity.

Microbial Nutrition in Aquatic Environments

1. Photosynthesis: Some bacteria near the surface produce food through photosynthesis.

2. Chemosynthesis: Bacteria at hydrothermal vents produce food through chemosynthesis using preformed organic carbon.

3. Decomposition: Many microorganisms obtain nutrition from the breakdown of organic matter.

Importance of Aquatic Microorganisms

• They play a vital role in nutrient cycling and are a crucial part of the food chain.

• Nutrient Cycling: Microorganisms break down organic matter, releasing nutrients usable by plants, especially nitrogen, phosphorus, and carbon.

• Food Source: Many microorganisms become food for animals.

• Human Use: Bacteria are used in sewage treatment plants to break down human wastes.

Disadvantages of Aquatic Microorganisms

• Some are pathogenic and can cause serious diseases.

• Examples: Salmonella typhi, S. paratyphi, and Norwalk virus in sewage-contaminated water.

• Fecal coliform (E. coli) and Enterococcus bacteria indicate the presence of disease-causing microorganisms.

Specific Zonation in Water Column

• Based on light and temperature variations.

• A. Water Surface and Epilimnion:

  • Temperature: 22-25°C (Water surface), 20-22°C (Epilimnion).

  • Microbial flora: Photosynthetic bacteria, Cyanobacteria, Mesophilic contaminating bacteria, Psychrotrophic or psychrophilic bacteria.

  • Microbial activity: Photosynthesis, aerobic nitrogen fixation, aerobic decomposition of organic matter.

• B. Thermocline (Metalimnion):

  • Temperature: 7°C.

  • Microbial flora: Psychrophilic facultative anaerobic bacteria.

  • Microbial activity: Aerobic and anaerobic decomposition.

• C. Hypolimnion:

  • Temperature: -5°C.

  • Microbial flora: Extreme psychrophilic anaerobic bacteria.

  • Microbial activity: Anaerobic nitrogen fixation (Clostridium pasteurianum), anaerobic decomposition, production of $CH<em>4$, $H</em>2S$, and $NH_3$.

• D. Sediments:

  • Microbial flora: Barophilic bacteria, anaerobic bacteria, $H<em>2S$ and $CH</em>4$ producing bacteria.

  • Microbial activity: Anaerobic decomposition.

Metabolic Rate and Temperature

• Metabolic rates increase exponentially over a temperature range.

• Below this range, there is little metabolic activity; above it, enzymes denature.

• Metabolic rates double for each 10°C increase (Q10).

• Enzyme kinetics depend on temperature: slow at low temperatures, maximum rate at optimal temperature, less efficient above optimal temperature.

• Optimal temperature range for organism activity and reproduction.

• Table (2-1): Temperature tolerances of different bacteria.

  • Psychrophilic: Minimum (-10)-(+5)°C, Optimum (+10)-(+20)°C, Maximum (+20)-(+30)°C

  • Mesophilic: Minimum (+10)-(+15)°C, Optimum (+30)-(+40)°C, Maximum (+40)-(+50)°C

  • Thermophilic: Minimum (+25)-(+45)°C, Optimum (+50)-(+75)°C, Maximum (+75)-(+93)°C

• Psychrophilic: Optimum growth below 15°C, minimum below 0°C.

Factors Affecting Enzyme Functioning

1. Enzymes are protein complexes, sensitive to temperature factors.

   • Temperatures over 80°C, heavy metals, or high H+ or OH- concentrations denature them.

   • Active chemicals such as chlorine and corrosive agents destroy proteins.

   • Extreme limits range from -2°C to 70°C; most cells thrive from 20°C to 40°C.

   • Temperatures below optimal slow enzyme action;

   • Temperatures above 75°C destroy most enzymes, except in bacterial endospores.

   • Sulfolobus sp. grow in temperature ranged from 70- 90 ºC.

2. Enzymes are sensitive to pH variations;

   • Optima generally range from pH 4.5 to pH 8.5, with optimal pH near 7.0 (neutrality).

3. Other factors include hydrostatic and osmotic pressures, ultraviolet light, and other radiations.

Gases and Aquatic Microorganisms

• Major gases: oxygen and carbon dioxide.

• Other gases: nitrogen, hydrogen, and methane.

Solubility of Gases

1. The higher the temperature, the less gas is dissolved.

   • Oxygen concentration differs by a factor of 2.1 between 0°C and 35°C.

2. Air or water pressure affects saturation concentration.

   • High elevation lakes have lower oxygen concentrations due to lower air pressure.

• Saline water holds less oxygen than pure water.

  • Ocean's salt decreases oxygen saturation at 4°C from 13.1 mg/L (pure water) to 8.6 mg/L.

Microorganisms and Oxygen Requirements

1. Obligatory aerobes

2. Microaerophilic organisms

3. Facultative aerobes and anaerobes

4. Obligatory anaerobes

Salinity

• Water ecosystem divided by salt content:

  1. Freshwater habitat: salinity near zero.

  2. Marine water: salinity from 33-37 g/kg.

• Halophilic microorganisms: Marine organisms living in saline habitats.

  • Weakly halophilic: 2-4%

  • Moderately halophilic: 5-20%

  • Extremely halophilic: 20-30%

• Halophobic microorganisms: Intolerant of saline habitats.

• Halotolerant microorganisms: Grow in freshwater but tolerate high salinity.

• Elevation in salinity increases generation time and impacts morphology and physiology.

  • Some bacteria become elongated or filamentous with increased salinity.

Biogeochemical Cycles Role of Microorganisms

• Nature recycles atoms (C, H, O, N, S, Fe) from inorganic to organic matter, and microorganisms are crucial.

The Carbon Cycle

• Photosynthesis and respiration are related:

  • Respiration releases carbon dioxide used in photosynthesis.

  • Photosynthesis releases oxygen used in respiration.

• On land, plants uptake carbon dioxide via photosynthesis.

• In aquatic ecosystems, carbon dioxide combines with water to form bicarbonate ions, a carbon source for algae. The amount of bicarbonate in water is in equilibrium with amount of carbon dioxide in air.

• Autotrophs fix carbon, heterotrophs produce $CO<em>2$. $CO</em>2 + H<em>2O \rightleftharpoons CH</em>2O$ (organic material) (autotrophs)
  $CH<em>2O + O</em>2 \rightleftharpoons CO<em>2 + H</em>2O$ (heterotrophs)

• Autotrophs are primary producers, converting carbon for heterotrophs.

• Cyanobacteria, lithotrophs, and methanogens fix $CO_2$ in the global carbon cycle.

Methanogens

• Inhabit anaerobic environments where $CO<em>2$ and $H</em>2$ occur.

  • 5% of $CO_2$ is reduced to cell material in autotrophic growth.

  • 95% is reduced to $CH<em>4$ (methane) during energy generation. $CO</em>2 + H<em>2 \rightleftharpoons CH</em>2O$ (cell material) + $CH_4$ (methanogenesis)

• Methane accumulates in rocks as fossil fuel, in animal guts, sediments, swamps, landfills, and sewage digesters.

• Under aerobic conditions, methylotrophs oxidize methane and its derivatives.

  • Example: Methanococcus sp.

Biodegradation

• Decomposition of organic material ($CH<em>2O$) to $CO</em>2$ and $H_2O$.

• Fungi and prokaryotes are important in soil habitats.

• Process: Depolymerization of biopolymers (cellulose, lignin, proteins) by extracellular enzymes, followed by oxidation (fermentation or respiration) of the monomeric subunits.

• End products: $CO<em>2$, $H</em>2O$, $H<em>2$, $NH</em>3$, $H_2S$ scarfed up by lithotrophs and autotrophs for recycling.

• Prokaryotes: Actinomycetes, Clostridia, Bacilli, Arthrobacters, and Pseudomonads.

• Polymers (e.g. cellulose) monomers e.g. glucose (Depolymerization)

• Monomers fatty acids (e.g. lactic acid, acetic acid, propionic acid) + $CO<em>2 + H</em>2$ (Fermentation)

• Monomers + O2 $CO<em>2 + H</em>2O$ (Aerobic respiration)

• Microbes can degrade any natural compound.

• Plastics, insecticides, pesticides, and poisons are degraded slowly.

Nitrogen Cycle

• Moves atmospheric nitrogen into organic N, then ammonia N, then nitrate N, and back to atmospheric N.

• Nitrogen is 78% of the atmosphere as di-nitrogen gas ($N_2$); essential for proteins and nucleic acids.

• Most nitrogen (98%) is in rock, sediment, and soils, but rock or mantle N is not readily available.

• Volcanic eruptions periodically release ammonia ($NH<em>3$) and gaseous nitrogen ($N</em>2$).

Nitrogen Fixation

• Conversion of $N<em>2$ to $NH</em>3$ or organic nitrogen, catalyzed by nitrogenase.
  $N<em>2 + 6e^- + 8H^+ \xrightarrow{\text{Nitrogenase and Fe, Mo}} 2NH</em>3 + H_2$

• Ammonia is combined with organic acids to form amino acids and proteins.

• Cyanobacteria (Nostoc and Anabaena sp.) fix N non-symbiotically and symbiotically.

• Bacteria associated include Clostridium pasteurianum, Desulphovibrio sp., Pseudomonas sp., Aeromonas sp., Achromobacter sp., Flavobacterium sp.

Ammonification

• Release of ammoniacal nitrogen from nitrogen-containing organic compounds.

• Bacteria decompose organic nitrogen forms to ammonium.

• Process:
  $RNH<em>2$ (Organic N) Poly peptide Peptide Amino acid Amino acid $NH</em>3$
  Protein Proteinase Polypeptidase Peptidase Proteolytic

• Microorganisms include: Clostridium histoluticum, C. sporogenes, Proteus sp., Pseudomonas sp., Bacillus sp.

Nitrification

• Oxidation process mediated by Nitrosomonas species:
  $NH<em>4^+ + \frac{3}{2} O</em>2 \longrightarrow HNO<em>2 + H^+ + H</em>2O$

• Step mediated by Nitrobacter species:
  $HNO<em>2 + \frac{1}{2} O</em>2 \longrightarrow NO_3^- + H^+$

Denitrification

• Conversion of $NO<em>3^-$ to $N</em>2$ gas in low oxygen conditions.
  $C<em>6H</em>{12}O<em>6 + 4NO</em>3^- \longrightarrow 6CO<em>2 + 6H</em>2O + 2N<em>2 text{(gas)} + NO + NO</em>2$

• Bacterial denitrification involves microbial reduction of $NO<em>3^-$ to $NO</em>2$ or N.

• Examples: Pseudomonas denitrificans, Micrococcus denitrificans, Thiobacillus denitrificans, Pseudomonas aeruginosa.

Phosphorus Cycle

• Weathering of rocks makes phosphate ions ($PO<em>4$= and $HPO</em>4$=) available to plants.

• Apatite $(Ca<em>3(PO</em>4)_2)$ contains phosphorus.

• Organisms utilize phosphate in phospholipids, ATP, teeth, bones, and shells.

• Phosphorus is a limiting nutrient.

Uses and Causes of Phosphorus

• Humans mine phosphate ores for use in fertilizer, as an animal feed supplement, and for detergents.

• Detergents, untreated human and animal wastes, and fertilizers from cropland add excess phosphate to water, causing algal blooms.

Forms of Phosphorous

• Main forms of phosphorous: Inorganic (orthophosphate $PO<em>4$, hydrogen phosphate $HPO</em>4$, apatite $Ca<em>3 (PO</em>4)_2$ and organic phosphorous

• The phosphorus cycle is comparatively simple. Inorganic phosphate exists in only one form (apatite $Ca<em>3 (PO</em>4)_2$). It is interconvert from an inorganic to an organic form and back again, and there is no gaseous intermediate.

Phosphorus in Biological Systems

• Phosphorus is an essential element in biological systems because:

  • it is a constituent of nucleic acids, (DNA and RNA)

  • it occurs in the phospholipids of cell membranes.

  • Phosphate is also a constituent of ADP and ATP which are universally involved in energy exchange in biological systems.

• Dissolved phosphate $(PO_4)$ finally ends up in the oceans. It is returned to land by shore animals and birds that feed on phosphorus containing sea creatures and then deposit their feces on land.

• Dissolved $PO_4$ is also returned to land by a geological process, the uplift of ocean floors to form land masses, but the process is very slow.

• Phosphorus is highly reactive and likely to bond with other elements.

• Microorganisms absorb insoluble phosphorus compounds and turn them into soluble phosphates through the action of acids.

• Algae and green plants absorb these phosphates and, in turn, are eaten by animals. When they die, the animals release the phosphates back into the soil.

Sulfur Cycle

• Main forms: inorganic S, $SO<em>4$, $H</em>2S$, $SO<em>3$, $S</em>2O_3$ and organic forms.

• Sulfur is a component of vitamins and essential metabolites and it occurs in two amino acids, cysteine and methionine.

• Microbes transform sulfur from its most oxidized form ($SO<em>4$) to its most reduced state ($H</em>2S$).

Sulfur-Oxidizing Prokaryotes

• Oxidize $H<em>2S$ to S and S to $SO</em>4$.

• The first is the oxygenic photosynthetic purple and green sulfur bacteria (Like Chromatium) that oxidize $H_2S$ as a source of electrons for cyclic photophosphorylation.

• The second is the "colorless sulfur bacteria" like Rhodospirillum (which oxidize $H<em>2S$ and S as sources of energy. In either case, the organisms can usually mediate the complete oxidation of $H</em>2S$ to $SO<em>4$. $H</em>2S$ (oxidation) S $SO_4$ (litho or phototrophic sulfur oxidation)

Characteristics of Sulfur-Oxidizing Prokaryotes

• Frequently thermophiles in hot volcanic springs and deep-sea thermal vents rich in $H_2S$.

• May be acidophiles, acidifying their environment by producing sulfuric acid.

Sulfate Reducing Bacteria

• Use $SO<em>4$ and S as electron acceptors for respiration, producing $H</em>2S$ during anaerobic respiration similar to denitrification.
  $SO<em>4 + H^+$ (anaerobic condition by sulfate reducing bacteria) $H</em>2S$

• Occurs in anaerobic bogs, soils, and sediments, making the distinctive odor of $H_2S$.

• Sulfur assimilated by bacteria and plants as $SO_4$ for use and reduction to sulfide.

• Animals and bacteria remove sulfide from proteins during decomposition.

Photosynthetic Purple and Green Bacteria

• Conduct an-oxygenic photosynthesis, also called bacterial photosynthesis.

• Bacterial photosynthesis differs from plant-type (oxygenic) photosynthesis in several ways.

  • Bacterial photosynthesis does not produce $O_2$, it only occurs under anaerobic conditions.

  • Bacterial photosynthesis utilizes a type of chlorophyll other than chlorophyll a, and only one photosystem, photosystem I.

  • The electron donor for bacterial photosynthesis is never H2O but may be $H<em>2$, $H</em>2S$ or $So$, or certain organic compounds.

  • The light-absorbing pigments of the purple and green bacteria consist of bacterial chlorophylls and carotenoids. Phycobilins, characteristic of the cyanobacteria, are not found.

  • Many purple and green sulfur bacteria store elemental sulfur as a reserve material that can be further oxidized to $SO_4$ as a photosynthetic electron donor.

• The purple and green bacteria may use $H<em>2S$ during photosynthesis in the same manner that cyanobacteria or algae or plants use $H</em>2O$ as an electron donor for autotrophic $CO_2$ fixation (the "dark reaction" of photosynthesis). Or they may utilize organic compounds as electron donors for photosynthesis.

• For example, Rhodobacter can use light as an energy source while oxidizing succinate or butyrate in order to obtain electrons for $CO<em>2$ fixation. Butyrate (CH3CH2CH2COO -) + $O</em>2\xrightarrow{\text{oxidation by Rhodobacter}}$CO2$(fix.)+$H2O$</p></li></ul><h3 collapsed="false" seolevelmigrated="true">Sulfur (Sulfate) Reduction</h3><ul><li><p>Divided into Assimilatory and Dissimilatory:</p><ul><li><p>Assimilatory: Sulfate is converted to a protein containing Sulfur by:<br>$SO_4^{2-} + PADP \longrightarrow PADPS$(phosphoadenosine-5-phosphosulfate)<br>$PADPS$+ reduction Sulfite (SO3) Thiosulphate (S2O3) Sulfide Sulfate must be 'activated' initially by PAP before the reaction can proceed. Assimilatory sulfate reduction occurs anaerobically as well as aerobically. This yields methionine which yields sulfur-containing protein.</p></li><li><p>Dissimilatory: In soils that become deficient in oxygen, usually the result of flooding, the sulfide level will increase to relatively high concentrations. The basic reaction:</p></li></ul></li><li><p>Oxidizable C (electron donor) + 4$H2$+$SO4^{-2}$(electron acceptor)$S^{-2}$+ 4$HOH$</p></li><li><p>The formation of sulfide by sulfate reduction in nature is enhanced in: warm, wet, or water logged soils with a pH of above 6.0.</p></li><li><p>Sulfide accumulation may be particularly pronounced in sulfate-rich saline areas in which plant excretions (release of carbon compounds) serve as the oxidizable carbon source in addition to the hydrogen.</p></li><li><p>Thus, like denitrification, an oxidizable carbon source serves as the electron donor, while the sulfate serves as the electron acceptor.</p></li><li><p>The metabolic dissimilatory process is similar to the assimilatory sulfate reduction in that the sulfate must be first activated by a molecule called ADP (adenosine-5- phosphate).<br>Sulfate$(SO4)$+ ADP ADPS (adenosine-5-phosphosulfate) sulfite (SO3) sulfide$(H2S)$$