Microbiology Chap. 10 and 11

Anoxygenic Phototrophs

§Earliest photosynthesizers likely anoxygenic phototrophs •Use hydrogen sulfide or organic compounds (not water) to make NADPH; do not generate O2 •Modern-day phylogenetically diverse •Live in bogs, lakes, upper layers of mud •Little or no O2, but light penetrates •Different photosystems than plants, algae, cyanobacteria •Use unique bacteriochlorophyll •Absorbs deep-penetrating wavelengths of light -Purple sulfur bacteria—Chromatium, Thiospirillum, Thiodictyon -Purple non-sulfur bacteria— Rhodobacter, Rhodopseudomonas,Rhodospirillum -Green sulfur bacteria—Chlorobium, Pelodictyon -Filamentous anoxygenic phototrophic bacteria—Chloroflexus -Others—Heliobacterium 6CO2 + 12H2S-------------> C6H12O6+12S+ 6H20 (Carbon source) (electron source)

Clostridium (Anaerobic Chemoorganotrophs—Fermentation)

•Are Gram-positive, endospore-forming rods •Common in soils; vegetative cells live in anaerobic microenvironments created by aerobes consuming O2 •Endospores tolerate O2, survive long periods of heat, drying, chemicals, irradiation •Germinate when conditions improve •Diverse metabolism; some cause diseases -Endospore-forming obligate anaerobes. Inhabitants of soil. Gram-positive. (Phylum: Firmicutes)

16S rDNA Sequence Analysis

•Comparisons revolutionized classification •Sequences highly conserved since function critical •Lack of mutations allows identification of distant relatedness •Certain regions relatively variable, can determine recent divergence •Some sequences will be similar even if organisms diverged long ago •Horizontal gene transfer appears rare •Culturing not necessary (can use PCR) •May not resolve at species level since closely related prokaryotes can have identical 16S rDNA sequences -DNA hybridization a better tool in these cases •Analysis of ribosomal proteins accounts for novel organisms; more gene products compared

Purple Bacteria

•Demonstrated requirement of light for growth •No evolution of oxygen •Oxidize hydrogen sulfide during CO2 fixation •Noted remarkable similarity of photosynthetic reactions in all photosynthetic organisms •Gram-negative; appear red, orange, or purple •Photosynthetic apparatus in cytoplasmic membrane •Unlike other anoxygenic phototrophs •Folds increase surface area

Propionibacterium (Anaerobic Chemoorganotrophs—Fermentation)

•Gram-positive pleomorphic rods •Produce propionic acid via fermentation •Can also ferment lactic acid •Important in dairy industry: Swiss cheese •Typical nutty flavor •CO2 creates signature holes •Also found in intestinal tract, anaerobic microenvironments on skin •acne -Obligate anaerobes that produce propionic acid as their main fermentation end product. Used in the production of Swiss cheese. Gram-positive. (Phylum: Actinobacteria)

Prokaryotes are metabolically diverse

•Numerous approaches to harvesting energy to produce ATP

Characterizing strains

-Foodborne illness -Diagnosing certain diseases -Forensic investigations of bioterrorism, biocrimes

Classification

Arranging organisms into smaller or related groups

Metabolic capabilities

A set of biochemical tests can be used to identify a microorganism. -Catalase test (Streptococcus is cat-) -Many rely on pH indicators -Sugar fermentation -Urease production -Basic strategy relies on dichotomous key (Flowchart of tests with positive or negative result, simultaneous inoculating speeds process) -Some tests accomplished without culturing (breath test to assay urease and identify Helicobacter pylori) -Commercial kits allow rapid identification via biochemical tests (API, Enterotube) -Biolog system

Microscopic Morphology

An important initial step. Size, shape, and staining characteristics can give suggestive information as to the identity of the organism. Further testing, however, is needed to confirm the identification. Sometimes enough to diagnose eukaryotic infections. Special stains useful. Gram staining suggested to sufficiently start appropriate therapy.

Antibiograms

Antibiotic susceptibility patterns can be used to characterize strains. -Antibiotic susceptibility patterns -Clearing zones around antibiotic discs -Largely replaced by molecular techniques

Prosthecate bacteria - Caulobacter, Hyphomicrobium (Thrives in Aquatic Environments)

Appendages increase their surface area. Gram negative (Phylum: Proteobacteria) •diverse group of Gram-negatives •Have extensions of cytoplasm and cell wall called prosthecae: provide increased surface area for nutrient absorption, some allow attachment •Caulobacter serves as model for cell differentiation •Hypomicrobium similar to Caulobacter •Have distinct method of reproduction, budding produces motile swarmer cells

Ferdinand Cohn (1870's)

Bacteria classified by shape

Biochemical typing

Biochemical tests are most commonly used to identify various species of bacteria, but in some cases they can be used to distinguish different strains. A group of strains that have a characteristic biochemical pattern is called a biovar or a biotype. -Vibrio cholerae El Tor strain can be tracked this way

Albert Kluyver (C.B. van Niel, 1930's)

Classification based on evolutionary relationships

Culture characteristics

Colony morphology can give initial clues to the identity of an organism. -Streptococci colonies generally fairly small -Serratia marcescens colonies often red at 22 degrees C -Pseudomonas aeruginosa often produces green pigment. Cultures have distinct fruity odor. -Differential media aids in identification -Streptococcus pyogenes (strep throat) yields B-hemolytic colonies on blood agar -E.coli (UTI) ferments lactose, forms pink colonies on MacConkey agar

Agrobacterium (Thrives in Terrestrial Environments)

Cuse plant tumors. Scientists use their Ti plasmid to move genes into plant cells. Gram-negative. (Phylum: Proteobacteria) •Gram-negative rods •Genetically modify plants via plasmid to form tumors •Enter through a plant wound •Ti plasmid in A. tumefaciens •DNA encodes ability to synthesize plant growth hormone; results in tumor •Also encodes synthesis of unusual amino acid derivative opine •Agrobacterium use opine as nutrient

Bdellovibrio (Thrives in Aquatic Environments)

Derive nutrients from other organisms. Predator of other bacteria. Gram-negative (Phylum: Proteobacteria) •highly motile Gram-negative curved rods •Prey on E. coli and other Gram-negatives •Strikes forcefully; prey propelled short distance •Parasite attaches, rotates, secretes digestive enzymes; forms hole in cell wall of prey

Genotypic Characteristics

Differences in DNA sequences can be used to determine the point in time at which two organisms diverged from a common ancestor

Azotobacter (Thrives in Terrestrial Environments)

For cysts. Notable for their ability to fix Nitrogen in aerobic conditions. Gram-negative. (Phylum: Proteobacteria) •Gram-negative pleomorphic rods •Fix nitrogen in aerobic conditions •High respiratory rate maintains low O2 in cell •Also protein binds nitrogenase, protects from O2 damage •Form resting cell called cyst •Negligible metabolic activity •Withstand drying and UV radiation •Not highly resistant to heat

Myxobacteria- Chondromyces, Myxococcus, Stigmatella (Thrives in Terrestrial Environments)

Group together to form fruiting bodies; ceels within these differentiate to form dormant microcysts. Gram-negative (Phylum: Proteobacteria) •group of aerobic Gram-negative rods that includes Chondromyces, Myxococcus, Stigmatella •Favorable conditions: secrete slime layer, form swarm •Nutrients depleted: cells congregate into fruiting body •Cells differentiate, form dormant microcysts •Microcysts resist heat, drying, radiation •Degraders of complex organic substances

Others—Heliobacterium (Anoxygenic Phototrophs)

Have not been studied extensively. Endospore formers (Phylum: Firmicutes) •Although green and purple bacteria most extensively studied, other anoxygenic phototrophs exist •Members include Heliobacterium •Gram-positive endospore-forming rods •Related to Clostridium

Anabaena, Synechococcus (Oxygenic Phototrophs—Cyanobacteria)

Important primary producers. Some fix N2. Gram-negative (Phylum: Cyanobacteria)

Taxonomy

Is the science that studies organisms to arrange them into groups, or taxa

Nucleic acid probes

Locate nucleotide sequence characteristic of species or groups. Most methods first increase DNA in sample ex.(inoculation on agar or in vitro DNA amplification (PCR)) -Fluorescence in situ hybridization (FISH) probes for 16S rRNA -No amplification needed, signature sequence characterizes certain group

The Enterobacteriaceae— Escherichia, Enterobacter, Klebsiella, Proteus, Salmonella, Shigella, Yersinia (Aerobic Chemoorganotrophs—Facultative Anaerobes)

Most reside in the intestinal tract. Those that ferment lactose are coliforms; their presence in water serves as an indicator of fecal pollution. Gram-negative. (Phylum: Proteobacteria) •enterics or enterobacteria are Gram-negative rods found in intestinal tract of humans, other animals; some thrive in soil •Facultative anaerobes that ferment glucose •Normal intestinal microbiota include Enterobacter, Klebsiella, Proteus, most E. coli strains •Diarrheal disease caused by Shigella, Salmonella enterica, and some E. coli strains •Life-threatening: typhoid fever (Salmonella enterica serotype Typhi) and bubonic and pneumonic plague (Yersinia pestis) •Lactose fermenters termed coliforms

Legionella (Thrives in Aquatic Environments)

Often reside within protozoa. Gram-negative (Phylum: Proteobacteria) •Gram-negative obligate aerobes •Use amino acids, but not carbohydrates, as source of carbon and energy •Often reside in protozoa; also found in water in air conditioners, and in produce misters •L. pneumophila can cause respiratory disease

Serological testing

Proteins and polysaccharides of prokaryotic cells can serve as identifying markers. These can be detected using specific antibodies. -Most useful include surface structures of cell wall, capsule, flagella, pili -Some Streptococcus species contain unique carbohydrate in cell wall

Sigurd Orla-Jensen (1908)

Physiology rather than morphology

Strains

Population of cells descended from a single cell

Identification

Process of characterizing in order to group

Carl Woese (late 1970's)

Prokaryotes divided into two major groups based upon ribosomal RNA sequences (Led to current 3 domain system: Bacteria, Archaea, and Eukarya)

Roger Stainer (1970's)

Relationships determined by comparing physical traits, nucleotide sequences

Streptomyces (Thrives in Terrestrial Environments)

Resemble fungi in their pattern of growth: produce antibiotics. Gram-positive. (Phylum: Actinobacteria) •aerobic Gram-positive bacteria •Growth resembles fungi: form mass of branching hyphae called mycelium •Chains of spores (conidia) develop at tips •Conidia resistant to drying; easily spread by air currents •Produce •extracellular enzymes for degradation of organic cmpds •also geosmins (earthy smell) •medically useful antibiotics including streptomycin, tetracycline, erythromycin

Bioluminescent bacteria- Aliivibrio fischeri, Vibrioharveyi, and Photobacterium phosphoreum (Thrives in Aquatic Environments)

Some form symbiotic relationships with specific types of fish and squid. Gram negative (Phylum: Proteobacteria) •Symbiotic relationships with certain fish, squid •Help with camouflage, confuse predators and prey •Bioluminescence under control of quorum sensing •Luciferase enzyme •Gram-negative rods (Vibrio are curved rods) •Facultative anaerobes, marine environments

Spirillum (Thrives in Aquatic Environments)

Spiral-shaped microaerophilic bacteria. Gram-negative (Phylum: Proteobacteria) •Gram-negative spiral-shaped microaerophilic bacteria •S. volutans stores phosphate as volutin granules •Metachromatic granules (methylene blue)

Nomenclature

System of assigning names

Hydrogen-oxidizing bacteria— Aquifex, Hydrogenobacter (Aerobic Chemolithotrophs)

Thermophilic bacteria that oxidize hydrogen gas as an energy source. One of the earliest bacterial forms to exist on earth. (Phylum: Aquifacae) •Aquifex, Hydrogenobacter among few hydrogen-oxidizing bacteria that are obligate chemolithotrophs •Thermophilic; typically inhabit hot springs •Some Aquifex have maximum growth at 95ºC •Deeply branching in phylogenetic tree, believed one of earliest bacterial forms to exist on earth •O2 requirements low, possibly available early on in certain niches due to photochemical processes that split water H2 + (1/2)O2-------------> H2O (Energy source) (terminal electron acceptor)

Thermus (Aerobic Chemoorganotrophs—Obligate Aerobes)

Thermus aquaticus is the source of Taq polymerase (used in PCR). Stains Gram-negative. (Phylum: Deinococcus-Thermus) •Thermus and Deinococcus: related genera •Have unusual cell walls •Thermus are thermophilic, valuable for their heat-stable enzymes, Taq polymerase in PCR reaction •Stain Gram neg

Vibrio (Aerobic Chemoorganotrophs—Facultative Anaerobes)

Typically found in marine environments because most species require at least low levels of Na+ for growth. Gram-negative. (Phylum: Proteobacteria) •found in marine water, include the agent of cholera (V. cholerae)

Epulopiscium (Thrives in Aquatic Environments)

Very large cigar-shaped bacteria that multiply by releasing several daughter cells; each cell has thousands of copies of the genome. Gram-positive. (Phylum: Firmicutes) •Gram-positive cigar-shaped bacteria •Reside in intestinal tract of surgeon fish •Larger than most prokaryotes (600 x 80 μm) •Thousands of copies of genome allow protein synthesis throughout organism •Instead of binary fission, they enlarge considerably and then lyse to release up to seven daughter cells

Horizontal gene transfer

complicates DNA comparisons. -Bacterium Thermotoga maritima appears to have acquired 25% of genes from archael species

Green Bacteria

•Gram-negative; typically green or brownish

Cornelius B. van Niel (1897-1985)

•Ph.D. from Technological University in Delft, Holland •Accepted position at Hopkins Marine Station, CA •Studied photosynthetic purple bacteria

Photosynthetic reactions

•Purple sulfur bacteria use hydrogen sulfide instead of H2O •Produce oxidized sulfur compounds instead of O2 •Indicated O2 from plants and algae comes from water, not CO2

DNA Hybridization

•Relatedness of organisms can be determined by similarity of nucleotide sequences •Sequence homology measured by DNA hybridization -How completely do they hybridize with each other •Extent of hybridization reflects degree of similarity •Complementary base pairing of single strands •If high percentage, considered related -70% similarity often considered same species -But Shigella and Escherichia should be grouped in same species based on DNA hybridization

Lactic acid bacteria—Streptococcus, Enterococcus, Lactococcus, Lactobacillus, Leuconostoc (Anaerobic Chemoorganotrophs—Fermentation)

•produce lactic acid •Most can grow in aerobic environments; only ferment •Lack catalase •Streptococcus inhabit oral cavity; normal microbiota •Some pathogenic (for example, β-hemolytic S. pyogenes) •S. thermophilus used to make yogurt •Lactococcus species used to make cheese •Enterococcus inhabit human, animal intestinal tract •Lactobacillus rod-shaped, common in mouth, vagina •Break down glycogen deposited in vaginal lining •Resulting low pH helps prevent vaginal infections •Also present in decomposing materials •Important in production of fermented foods -Gram positive Lactobacillus and Streptococcus in yogurt -Produce lactic acid as the major end product of their fermentative metabolism. Aerotolerant anaerobes. Several genera are used by the food industry. Gram-positive. (Phylum: Firmicutes)

Ammonium oxidizers- Nitrosomonas, Nitrosococcus,

NH4+ + 1(1/2)O2-------------> NO2- + H2O + 2H+ (Energy source) (terminal electron acceptor)

Evolutionary Chronometers

DNA sequences. Provide relative measure of time elapsed since divergence from common ancestor -Random mutations accumulate over time -DNA sequencing allows construction of phylogenetic tree

Micrococcus (Aerobic Chemoorganotrophs—Obligate Aerobes)

Widely distributed; common laboratory contaminants. Gram-positive. (Phylum: Actinobacteria) •Gram-positive cocci •Found in soil, dust particles, inanimate objects, skin •Pigmented colonies •Tolerate dry, salty conditions

Anaerobic Chemotrophs

-Atmosphere anoxic for first ~1.5 billion years that prokaryotes inhabited earth •Early chemotrophs likely used anaerobic respiration •Terminal electron acceptors like abundant CO2 or S •Others may have used fermentation •Passed electrons to organic molecule like pyruvate §Today anaerobic habitats common •Aerobes contribute by depleting O2 •Mud, tightly packed soil limit diffusion of gases •Aquatic environments can become limiting for oxygen •Human body (especially intestinal tract) •Anaerobic microenvironments in skin, oral cavity

Anaerobic Chemoorganotrophs—Fermentation

-Clostridium -Lactic acid bacteria—Streptococcus, Enterococcus, Lactococcus, Lactobacillus, Leuconostoc -Propionibacterium •Numerous anaerobic bacteria ferment •This is not the same thing as anaerobic respiration! •ATP via substrate-level phosphorylation (not ETC) •Many different organic energy sources, end products glucose + pyruvate-------------> lactic acid (Energy source) (Terminal electron acceptor)

Aerobic Chemoorganotrophs—Facultative Anaerobes

-Corynebacterium -The Enterobacteriaceae— Escherichia, Enterobacter, Klebsiella, Proteus, Salmonella, Shigella, Yersinia -Vibrio

Bergey's Manual of Systematic Bacteriology

-Describes all known species -Newest edition in 5 volumes -Classifies according to genetic relatedness -Previous edition grouped according to phenotype, so some major differences -Names given according to International Code of Nomenclature of Bacteria

Aerobic Chemolithotrophs

-Filamentous sulfur oxidizers—Beggiatoa, Thiothrix -Unicellular sulfur oxidizers—Thiobacillus, Acidithiobacillus -Nitrifiers—Nitrosomonas, Nitrosococcus, Nitrobacter, Nitrococcus -Hydrogen-oxidizing bacteria— Aquifex, Hydrogenobacter -§gain energy by oxidizing reduced inorganic chemicals •Sulfur-oxidizing bacteria: Gram-negative rods, spirals •Energy from oxidation of sulfur, sulfur compounds including H2S, thiosulfate •O2 is terminal electron acceptor; generates sulfuric acid •Important in sulfur cycle •Filamentous and unicellular lifestyles •Filamentous forms may interfere with wastewater treatment facilities •Unicellular forms can affect environmental pH S + 1(1/2)O2 + H2O-------------> H2SO4 (Energy source) (terminal electron acceptor)

Informal groupings of Bacteria

-Lactic acid bacteria -Anoxygenic phototrophs -Endospore-formers -Sulfate reducers

Aerobic Chemoorganotrophs—Obligate Aerobes

-Micrococcus -Mycobacterium -Pseudomonas -Thermus -Deinococcus §oxidize organic compounds for energy •Some inhabit specific environments, others ubiquitous •Obligate Aerobes organic compounds + O2-------------> CO2+H2O (Energy source) (terminal electron acceptor)

Eukarya

-Peptidoglycan cell wall: No -Cytoplasmic membrane lipids: Fatty acids linked to glycerol by ester linkage -Ribosomes: 80S -Presence of Introns: Yes -Membrane bound nucleus: Yes

Archaea

-Peptidoglycan cell wall: No -Cytoplasmic membrane lipids: Hydrocarbons (not fatty acids) linked to glycerol by ether linkage -Ribosomes: 70S -Presence of Introns: Sometimes -Membrane bound nucleus: No

Bacteria

-Peptidoglycan cell wall: Yes -Cytoplasmic membrane lipids: Fatty acids linked to glycerol by ester linkage -Ribosomes: 70S -Presence of Introns: No -Membrane bound nucleus: No

Sequencing Ribosomal RNA genes

-Ribosomal RNA (rRNA's) or encoding DNA (rDNAs) -Sequence relatively stable - Ribosome would not function with too may mutations -16S rRNA most useful because of moderate size -1500 nucleotides -16S (18S in eukaryotes) RNAs are small subunit rRNA's -Sequence compared with extensive databases -Can identify organisms that cannot be grown in culture

Detecting Specific Nucleotide Sequences

-Tests can identify sequences unique to species or group -Nucleic acid probes -Nucleic acid amplification tests (NAATs) -Limitation in each detects only single possibility -NEWER OPTION: Multiplex PCR has different primers for different organisms. More complex to design this type of test and expensive -Need to run multiple probes if organism being tested could be one of multiple different species or related groups

Oxygenic Phototrophs—Cyanobacteria

Anabaena, Synechococcus •Earliest oxygenic phototrophs •Introduction of O2 began ~3 billion years ago •Use water as source of electrons for reducing power •Still play essential role as primary producers •Harvest sunlight to convert CO2 (fix carbon) into organic compounds •Initially though to be algae, were called blue-green algae (but they are bacteria!) •Diverse group of Gram-negative bacteria •More than 60 genera •Many convert N2 to ammonia: nitrogen fixation •Morphologically diverse •Unicellular: cocci, rods, spirals •Multicellular: filamentous associations: trichomes •May be in sheath •Motile trichomes glide as unit •May have gas vesicles for vertical movement in water •Large numbers can accumulate in freshwater habitats •Called a bloom •Sunny, hot weather can lyse cells, create scum •Photosystems like those in chloroplasts of algae, plants, which evolved from ancestral cyanobacteria •Also have phycobiliproteins •Absorb additional wavelengths -Phycobilisome structure •Nitrogen-fixing cyanobacteria critical ecologically •Incorporate N2 and CO2 into organic material •Form usable by other organisms •Nitrogenase destroyed by O2, must be protected •Anabaena form specialized heterocysts •Lack photosystem II so no water-splitting •A. azollae fixes N2 in special sac of fern •Synechococcus fix N2 in dark when no photosynthesis (temporal separation) 6CO2 + 6H20-------------> C6H12O6+6O2 (Carbon source) (electron source)

Sheathed bacteria- Sphaerotilus, Leptothrix(Thrives in Aquatic Environments)

Aquatic environments lack steady nutrient supply Form chains of cells closed within a protective sheath. Swarmer cells move to new locations. Gram-negative (Phylum: Proteobacteria) •form chains of cells within tube •Sheaths protect, help bacteria attach to solid objects •Often seen streaming from rocks in water polluted by nutrient-rich effluents; may clog pipes •Include Gram-negative rods Sphaerotilus, Leptothrix •Motile swarmer cells exit open end of sheath, move to new surface, attach

Endospore-formers- Bacillus, Clostidium (Thrives in Terrestrial Environments)

Bacillus species include both obligate aerobes and facultative anaerobes; Clostridium species are obligate anaereobes. Gram positive. (Phylum: Firmicutes) •most resistant to environmental extremes •Bacillus, Clostridium most common •Gram-positive rods •Bacillus include obligate aerobes and facultative anaerobes •Some medically important: B. anthracis

Sequence analysis of ribosomal components

Certain regions of the SSU rDNA (16S in bacteria and archaea and 18S in eukaryotes) can be used to determine distant relatedness of diverse organisms; other regions can be used to determine more recent divergence. Amino acid sequences of ribosomal proteins can also be compared

Filamentous anoxygenic phototrophic bacteria—Chloroflexus (Anoxygenic Phototrophs)

Characterized by their filamentous growth. Gram-negative. (Phylum: Chloroflexi) •Form multicellular arrangements •Exhibit gliding motility •Many have chlorosomes •16S rDNA shows unrelated to green sulfur bacteria •Metabolically diverse •Some preferentially use organic compounds to generate reducing power, can grow in dark aerobically using chemotrophic metabolism •Chloroflexus best studied, especially thermophilic strains found in hot springs

Pseudomonas (Aerobic Chemoorganotrophs—Obligate Aerobes)

Common environmental bacteria that, as a group, can degrade a wide variety of compounds. Gram-negative (Phylum: Proteobacteria) •Gram-negative rods; oxidase positive •Motile by polar flagella; often produce pigments •Most are strict aerobes; no fermentation •Extreme metabolic diversity important in degradation •Ability sometimes from plasmids •Widespread: soil, water •Most harmless •Some pathogens: P. aeruginosa common opportunistic pathogen

G+C content

Determining the G + C content offers a crude comparison of genomes. Organisms with identical G + C contents can be entirely unrelated, however. G + C content is the percentage of G-C base pairs in DNA •If deviates by more than a few percent, organisms not related •Conversely, similarity of GC content does not guarantee relatedness

Phylogeny

Evolutionary relatedness. Yields 3 domain system based on Carl Woese. Replaces R. H. Whittaker's five- kingdom system. Illustrates relatedness of organisms based on rRNA sequence data that supports 3 domain system.

3 Domain System

Expanded by comparing amino acid sequences of ribosomal proteins

Rhizobia- Rhizobium, Sinorhizobium, Bradyyrhizobium, Mesohizobium, Azorhizobium (Thrives in Terrestrial Environments)

Fix nitrogen; form a symbiotic relationship with legumes. Gram-negative (Phylum: Proteobacteria) •Gram-negative rods that often fix nitrogen •Includes Rhizobium, Sinorhizobium, Bradyrhizobium, Mesorhizobium, Azorhizobium •Live in nodules on roots of legumes (peas, beans) •Plants synthesize leghemoglobin, which binds and controls O2 levels to yield microaerobic conditions •Allows bacteria to fix nitrogen

Purple sulfur bacteria—Chromatium, Thiospirillum, Thiodictyon (Anoxygenic Phototrophs) (Purple Bacteria)

Form colored masses in sulfur-rich aquatic habitats and use sulfur compounds as a source of electrons when making reducing power. Gram-negative. (Phylum: Proteobacteria) •Large cells; some motile •May have gas vesicles to control depth •Most store sulfur in intracellular granules •Preferentially use H2S to generate reducing power •Other inorganic (H2) or organic (pyruvate) compounds •Some can grow aerobically in absence of light •Oxidize reduced inorganic, organic compounds

Green sulfur bacteria—Chlorobium, Pelodictyon (Anoxygenic Phototrophs)

Found in habitats similar to those preferred by the purple sulfur bacteria. Gram-negative. (Phylum: Chlorobi) •Habitats similar to purple sulfur bacteria •Form granules outside of cell, use H2S •Accessory pigments located in chlorosomes attached to cytoplasmic membrane •Lack flagella •May have gas vesicles •Strict anaerobes •None are chemotrophic

Molecular Typing

Gel electrophoresis can be used to detect restriction fragment length polymorphisms (RFLPs). Whole-genome sequencing (WGS) is increasingly being used to detect differences. -Cut DNA samples with same restriction enzyme -Separate via gel electrophoresis -Patterns called restriction fragment length polymorphisms (RFLPs) - Different RFLPs indicate different strains

Purple non-sulfur bacteria— Rhodobacter, Rhodopseudomonas, Rhodospirillum(Anoxygenic Phototrophs) (Purple Bacteria)

Grow in aquatic habitats, preferentially using organic compounds as a source of electrons for reducing power. Many are metabolically versatile. Gram-negative. (Phylum: Proteobacteria) •Moist soils, bogs, paddy fields •Preferentially use organic molecules instead of H2S as source of electrons (but some can still use H2S) •Lack gas vesicles •May store sulfur; granules form outside cell •Remarkably diverse metabolism •Many use H2 for e- and some can use H2S (like purple sulfur bacteria) •Most can grow aerobically in absence of light using chemotrophic metabolism

Free-living spirochetes- Spirochaeto, Leptospira (some species)(Thrives in Aquatic Environments)

Long spiral shaped bacteria that move by means of endoflagella. Gram-negative (Phylum: Spirochaetes) •group of Gram-negatives with spiral shape •Flexible cell wall •Endoflagella or axial filament contained within periplasm allows corkscrew-like motion •Able to move through viscous environments like mud •Spirochaeta thrive in muds, anaerobic waters •Leptospira are aerobes; some free-living, others inhabit animals •L. interrogans causes leptospirosis

Protein profile

MALDI-TOF MS separates and sorts an organism's proteins by mass (mass spectrophotometer), generating a profile that provides a fast way to identify an organism grown in culture. -Laser beam vaporizes and ionizes sample Time of flight: small ions travel faster than larger ones in tube -Mass spectrum a "fingerprint" or profile of the proteins and other macro-molecules in the cell

Magnetospirillum (Thrives in Aquatic Environments)

Magnetic crystals allow them to move in water and sediments. Gram-negative. (Phylum: Proteobacteria) •Magnetotactic bacteria contain magnetic crystals that align cells with Earth's magnetic fields •Allows movement up or down in water or sediments •Likely allows location of their required microaerophilic habitats •Magnetospirillum are Gram-negative; spiral-shaped

PulseNet

Makes it easier to use RFLP data to track foodborne disease outbreaks -Established by CDC -Labs can submit RFLP patterns to database and quickly receive information about other isolates -Whole genome sequencing (WGS) can trace outbreaks caused by closely related strains -Genomic Trakr established by FDA -Network of labs that use WGS data for tracking outbreaks

Methanogens—Methanospirillum, Methanosarcina (Anaerobic Chemolithotrophs)

Members of the Archaea that oxidize hydrogen gas, using CO2 as a terminal electron acceptor to generate methane. (Phylum: Euryarchaeota)

Anaerobic Chemolithotrophs

Methanogens—Methanospirillum, Methanosarcina •Chemolithotrophs oxidize reduced inorganic chemicals to obtain energy •For example, H2 •Anaerobes use alternative electron acceptor •For example, CO2, sulfur •Relatively few anaerobic chemolithotrophs discovered •Some are bacteria •Acetogens (anaerobic chemolithotrophs in termite gut) •There are a number of aerobic bacterial chemolithotrophs (later in chapter) •Most anaerobic chemolithotrophs in domain Archaea •Methanogens are one group •Volta experiment •Methanogens are group of methane-producing archaea •Oxidize H2 gas to generate ATP (oxidize means to lose e-) •Alternatives include formate, methanol, acetate •CO2 as terminal electron acceptor •Smaller energy yield than other electron acceptors •Very sensitive to O2 (Hard to study) •Sewage, swamps, marine sediments, rice paddies, digestive tracts •Cows produce ~10 ft3/day 4H2 + CO2 -------------> CH4 + 2H20 (Energy source) (Terminal electron acceptor)

Nitrite Oxidizers- Nitrobacter, Nitrococcus

NO2- + (1/2)O2-------------> NO3- (Nitrate) (Energy source) (terminal electron acceptor)

Nitrifiers—Nitrosomonas, Nitrosococcus, Nitrobacter, Nitrococcus (Aerobic Chemolithotrophs)

Oxidize ammonia or nitrite as energy sources. This converts certain fertilizers to a form easily leached from soils, and depletes O2 in waters polluted with ammonia-containing wastes. Genera that oxidize nitrite prevent the toxic buildup of nitrite. Gram-negative. (Phylum: Proteobacteria) •are diverse group of Gram-negatives •Oxidize inorganic nitrogen compounds for energy •Concern to farmers using ammonium nitrogen (why?) •Can deplete water of O2 (hypoxia) if wastes high in ammonium •Two groups; usually grow in close association •Ammonium oxidizers: Nitrosomonas, Nitrosococcus NH4+ + 1(1/2)O2-------------> NO2- +H2O + 2H+ (Energy source) (terminal electron acceptor) •Nitrite oxidizers: Nitrobacter, Nitrococcus NO2- + (1/2)O2-------------> NO3- (Nitrate) (Energy source) (terminal electron acceptor)

Filamentous sulfur oxidizers—Beggiatoa, Thiothrix (Aerobic Chemolithotrophs)

Oxidize sulfur compounds as energy sources. Found in sulfur springs and sewage-polluted waters. Gram-negative. (Phylum: Proteobacteria) •Beggiatoa, Thiothrix: sulfur springs, sewage-polluted waters, surface of marine and freshwater sediments •Store sulfur as intracellular granules •Beggiatoa filaments move by gliding motility •Thiothrix filaments immobile; progeny cells detach, move via gliding motility

Unicellular sulfur oxidizers—Thiobacillus, Acidithiobacillus (Aerobic Chemolithotrophs)

Oxidize sulfur compounds as energy sources. Some species produce enough acid to lower the pH to 1.0. Gram-negative. (Phylum: Proteobacteria) •Acidithiobacillus: terrestrial and aquatic habitats •Oxidize metal sulfides, can be used for bioleaching •for example, oxidation of gold sulfide produces sulfuric acid; lower pH converts metal to soluble form •Can oxidize sulfur in fuels to sulfate; removal helps prevent acid rain •Can produce damaging acid runoff as low as pH 1.0

Serological Typing

Proteins and carbohydrates that vary among strains can be used to differentiate strains. A group of strains that have a characteristic serological type is called a serovar or a serotype. -E.coli distinguished by antigenic type of flagella, capsules, lipopolysaccharide molecules -E.coli 0157:H7 (O antigen is lipopolysaccharide; H antigen is flagella) -K is capsule -Group with characteristic antigens: serovar, serotype

Deinococcus (Aerobic Chemoorganotrophs—Obligate Aerobes)

Resistant to the damaging effects of gamma radiation. Stains Gram-positive. (Phylum: Deinococcus-Thermus) •are extraordinarily radiation resistant •Survive dose several thousand times that lethal to a human •Dose shatters genome into many fragments, yet enzymes repair damage •Scientists hope to genetically engineer species to help clean up radioactive wastes •Stain Gram pos

Phage Typing

Strains of a given species sometimes differ in their susceptibility to various types of bacteriophages. -Relies on differences in susceptibility to bacteriophages -Susceptibility pattern can be determined with bacteria and different bacteriophage suspensions -Largely replaced by molecular methods (still useful in labs lacking equipment for genomic testing)

Ecophysiology

Study of the adaptations of physiological mechanisms that prokaryotes use to live in terrestrial and aquatic environments As a group, prokaryotes show remarkable diversity in their physiological adaptations to a wide range of habitats, including the bodies of animals.

Anaerobic Chemoorganotrophs—Anaerobic Respiration

Sulfur- and sulfate-reducing bacteria—Desulfovibrio •Chemoorganotrophs oxidize organic compounds (for example, glucose) to obtain energy •Anaerobes often use sulfur, sulfate as electron acceptor •Sulfur- and Sulfate-Reducing Bacteria •Produce hydrogen sulfide (rotten-egg smell) •H2S is corrosive to metals •Important in sulfur cycle •At least a dozen recognized genera •Desulfovibrio most studied •Gram-negative curved rods •Some archaea also •Hyperthermophiles organic comp. + S -------------> CO2 + H2S (Energy source) (Terminal electron acceptor)

Species

The basic unit. Group of morphologically similar organisms capable of producing fertile offspring. Definition problematic for prokaryotes. Is a group of closely related isolates or strains.

DNA hybridization

The extent of nucleotide sequence similarity between two isolates can be determined by measuring how completely single strands of their DNA hybridize to one another.

Phenotypic characteristics

Traditionally, relatedness of different bacteria has been decided by comparing phenotypes. These characteristics, however, do not necessarily reflect the evolutionary relatedness of organisms

Sulfur oxidizing, nitrate-reducing marine bacteria- Thioplaca, Thiomargarita (Thrives in Aquatic Environments)

Use novel mechanisms, to compensate for the fact that their energy source (reduced sulfur compounds) and terminal electron acceptor (nitrate) do not coexist (Phylum: Proteobacteria) •Some store sulfur (energy source) and nitrate (terminal electron acceptor) not found in same environment • Thioploca species form long sheaths; cells shuttle between sulfur-rich sediments and nitrate-rich water •Thiomargarita namibiensis cells have nitrate storage vacuole occupying ~ 98% of cell; surrounded by cytoplasm containing sulfur globules •cell diameter can reach 0.75 mm

Sulfur- and sulfate-reducing bacteria—Desulfovibrio (Anaerobic Chemoorganotrophs—Anaerobic Respiration)

Use sulfate as a terminal electron acceptor, generating hydrogen sulfide. Found in anaerobic muds rich in organic material. Gram-negative (Phylum: Proteobacteria)

Phenotypic characteristics

Used to help identify microorganisms (prokaryotes). -Microscopic Morphology -Culture characteristics -Metabolic capabilities -Serology -Fatty acid analysis -Protein profile

Nucleic Acid Amplification Tests (NAATs)

Used to increase number of copies of specific DNA sequences -Allows detection of small numbers of organisms -Often from body fluids, soil, food, water -Detection of organisms that cannot be cultured -Polymerase chain reaction (PCR) common technique

Mycobacterium (Aerobic Chemoorganotrophs—Obligate Aerobes)

Waxy cell wall resists staining; acid-fast. (Phylum: Actinobacteria) •are acid-fast bacteria •Mycolic acid in cell wall prevents Gram-staining •Acid-fast staining used •Generally pleomorphic rods •Notable pathogens: M. tuberculosis, M. leprae •More resistant to disinfectants, often resistant to antimicrobial drugs •Related Nocardia species also acid-fast

Corynebacterium (Aerobic Chemoorganotrophs—Facultative Anaerobes)

Widespread in nature. Gram-positive. (Phylum: Actinobacteria) •Gram-positive pleomorphic rods •Wide-spread •Often club-shaped and form V shapes or palisades •Referred to as coryneforms or diphtheroids •Generally facultative anaerobes; some strict aerobes •Some can ferment •Many harmless •C. diphtheriae causes diphtheria