Low G/C Vs High G/C Grame +ve Bacteria

• Gram-positive bacteria constitute a large and diverse group
• Approximately 50% of characterized bacterial species are gram-positive

Gram-Positive Division

• They divide into two phyla/groups based on GC content
• Low GC (less than 50%) bacteria belong to the Firmicutes phylum
• High GC (greater than 50%) bacteria belong to the Actinobacteria phylum
• Examples of High GC include Corynebacterium, Mycobacterium, Nocardia, and Streptomyces

Firmicutes: General Information

• Includes Nonsporulating Bacillales and Clostridiales.
• Includes Sporulating Bacillales and Clostridiales.
• Includes Lactobacillales
• They are endospore formers and lactic acid bacteria
• They are low G+C gram-positive bacteria

Tenericutes: General Information

• Includes mycoplasmas
• Lack a cell wall

Actinobacteria: General Information

• Includes Coryneform and Propionic Acid Bacteria
• Includes Mycobacterium, Filamentous Actinomycetes: Streptomyces, and Relatives
• Includes actinomycetes, primarily filamentous soil bacteria
• High G+C gram-positive bacteria are defined as genomic DNA having many GC base pairs

Low GC Subdivision - Firmicutes Details

• There are exceptions in this subdivision having G+C content as high as 55% (Geobacillus thermocatenulatus)
• They can be cocci or bacilli
• Many produce endospores (sporulating)
• Some produce energy through photosynthesis
• They play an important role in beer, wine, and cider spoilage

Firmicutes – Nonsporulating Bacillales and Clostridiales Key Genera

• Listeria
• Staphylococcus
• Sarcina

Listeria

• Is gram-positive, catalase-positive coccobacillus
• Facultatively aerobic chemoorganotroph
• Forms chains 3 to 5 cells long
• Found widely in soils and is an opportunistic pathogen
• Pathogenic Listeria monocytogenes causes listeriosis, a foodborne infection
• Often grows well at low temperatures, such as in refrigerated foods
• Obligate aerobe that requires microoxic conditions
• Produces acid but not gas from glucose

Staphylococcus: Genus Details

• Facultative aerobe that respires but can ferment
• Gram-positive coccus that grows in clusters
• Catalase positive
• Resistant to reduced water potential
• Tolerates high salt (NaCl) and drying
• Many species are pigmented
• Very similar to Micrococcus (Actinobacteria)
• Common commensals and parasites of humans

Staphylococcus epidermidis

• Non-pigmented and nonpathogenic
• Found on the skin or mucous membranes

Staphylococcus aureus

• Yellow pigmented and pathogenic
• Associated with boils, pimples, pneumonia, meningitis, and arthritis
• Causes a variety of suppurative (pus-forming) infections
• Causes superficial skin lesions such as boils, styes, and furuncles
• Can cause pneumonia, mastitis, phlebitis, meningitis, and urinary tract infections, as well as deep-seated infections, such as osteomyelitis and endocarditis.
• It is a hospital-acquired (nosocomial) infection of surgical wounds and infections associated with indwelling medical devices
• Causes food poisoning by releasing enterotoxins into food, and toxic shock syndrome by release of superantigens into the bloodstream

Sarcina Details

• Obligate anaerobe
• Divides in three planes resulting in packets of eight cells or more
• Catalase-negative
• Extremely acid-tolerant, as low as pH 2
• Can inhabit and grow in stomachs of monogastric animals
• Can be found in soil, mud, feces, and stomach contents
• Found in the stomach of persons suffering from pyloric ulcerations

Firmicutes: Lactobacillales

• Generally, aerotolerant anaerobes lacking an electron transport chain
• Include Lactobacillus
• Include Streptococcus
• Include Enterococcus
• Include Leuconostoc

Lactobacillales Subgroups

• One group, called homofermentative, produces a single fermentation product, lactic acid
• The other group, called heterofermentative, produces other products, mainly ethanol plus CO₂, as well as lactate

Lactobacillus Details

• Key genera are Lactobacillus and Streptococcus
• Nonsporulating, oxidase- and catalase-negative rods or cocci that are exclusively fermentative
• Fermentative bacteria that produce lactic acid
• Widely used in food production and preservation
• Lack porphyrins and cytochromes, so no oxidative phosphorylation
• Obtain energy by substrate-level phosphorylation
• Are aerotolerant anaerobes, so not sensitive to oxygen
• Most get energy from the metabolism of sugars
• Rod-shaped and grow in chains
• Commonly founded in dairy products
• Resistant to acidic conditions, growing in pH as low as 4
• Can be homofermentative

Streptococcus Details

• Coccus-shaped and grows in chains or tetrads
• Homofermentative
• Plays important roles in the production of buttermilk and silage
• Some species form dental caries
• Some Streptococcus species are pathogenic
• Two groups of pathogenic Streptococci are: pyogenes (strep throat) and viridans (S. mutans-dental caries)
• Blood agar hemolysis is diagnostic
• Beta-hemolysis (complete red blood cell hemolysis) is diagnostic for pyogenes
• Incomplete hemolysis is diagnostic for viridans
• Immunological groups (A, B, C, F, G)

Other Lactobacillales Genera

• Lactococcus: genus of dairy significance and is homofermentative
• Enterococcus: genus of fecal origin and can be pathogenic
• Leuconostoc: heterofermentative lactococci that produce flavors (diacetyl and acetoin).
• Peptococcus and Peptostreptococcus: obligate anaerobes that ferment protein ('pepto') instead of sugar

Firmicutes – Sporulating Bacillales and Clostridiales Key Genera

• Bacillus
• Clostridium
• Sporosarcina
• All endospore-forming bacteria are Bacillales or Clostridiales
• Distinguished on the basis of cell morphology, shape, cellular position of endospore, relationship to O2, and energy metabolism
• Generally found in soils, therefore exist in a very variable environment of nutrient levels, temperature and water activity
• Endospores are advantageous for soil microorganisms due to variations in nutrients, temperature and water activity

Endospores:

• Type of dormant cell that are cryptobiotic
• Formed by a few groups of bacteria as intracellular structures, but ultimately released as free endospores
• Highly resistant to environmental stresses such as high temperature, irradiation, strong acids, and disinfectants
• Formed by vegetative cells in response to environmental stresses
• Germinate to become vegetative cells when the environmental stress is relieved, thus a mechanism of survival, rather than a mechanism of reproduction

Bacillus and Paenibacillus Details

• Aerobic or facultatively aerobic rods that grow well on many carbon sources
• Many produce extracellular hydrolytic enzymes that break down polymers such as polysaccharides, nucleic acids and lipids
• Many bacilli produce antibiotics during sporulation including bacitracin (B. subtilis), and polymyxin (B. polymyxa)
• Paenibacillus popilliae and Bacillus thuringiensis produce insect larvicides, for example, Bt toxin

Bacillus: Biopesticides

• Several Bacillus species produce insect larvicide
• Bacillus thuringiensis strains are used as biological insecticides as they are specific for lepidoptrans (moths), dipterans (mosquitos), and coleopterans (beetles)
• Bt toxin genes have been isolated and introduced to plants through genetic engineering to result in plants that are “naturally" resistant to insects

Bacillus: Pathogenic

• Bacillus anthracis and B. cereus are the predominant pathogens of medical importance
• Paenibacillus alvei, B. megaterium, B. coagulans, Brevibacillus laterosporus, B. subtilis, B. sphaericus, B. circulans, Brevibacillus brevis, B. licheniformis, P. macerans, B. pumilus and B. Thuringiensis have also been occasionally isolated from human infections
• B. anthracis is the causative agent of anthrax
• B. cereus causes food poisoning
• Non-anthrax Bacillus species can also cause a wide variety of other infections and are being recognized with increasing frequency as pathogens in humans

Clostridium Details

• Lacks a respiratory chain and is anaerobic
• Unlike Bacillus, Clostridium obtains ATP only by substrate level fermentation
• Some Clostridium can fix nitrogen
• Saccharolytic (ferments sugars) to produce acetone and butyric acid and one group ferments cellulose
• Proteolytic: ferments amino acids or amino acid pairs (via Stickland reactions)
• Mainly found in anaerobic pockets in the soil
• Also lives in mammalian intestinal tracts
• Some species are pathogenic, and diseases include botulism, tetanus, and gangrene: C. botulinum, C. tetani, C. perfringens, respectively

Firmicutes: Clostridium Perfringens Details

• Produces a huge array of invasins and exotoxins
• Causes wound and surgical infections that lead to gas gangrene, in addition to severe uterine infections
• Clostridial haemolysins and extracellular enzymes such as proteases, lipases, collagenase and hyaluronidase, contribute to the invasive process
• Produces an enterotoxin and is an important cause of food poisoning
• Organism is usually encountered in improperly sterilized (canned) foods in which endospores have germinated

Firmicutes: Clostridium Difficile Details

• Causes antibiotic-associated diarrhea (AAD) and more serious intestinal conditions such as colitis and pseudo membranous
• Colitis is due to overgrowth of Clostridium difficile in the colon, usually after the normal flora has been disturbed by antimicrobial chemotherapy
• C. difficile produces two toxins
• Toxin A is an enterotoxin because it causes fluid accumulation in the bowel
• Toxin B is an extremely lethal (cytopathic) toxin

Firmicutes: Clostridium Tetani Details

• Clostridium tetani is the causative agent of tetanus
• Mortality rates reported vary from 40% to 78%
• The disease stems does not stem from invasive infection but from a potent neurotoxin (tetanus toxin or tetanospasmin) produced when spores germinate and vegetative cells grow after gaining access to wounds
• Most cases of tetanus result from small puncture wounds or lacerations which become contaminated with C. tetani spores that germinate and produce toxin
• The toxin is produced during cell growth, sporulation and lysis
• Toxin migrates along neural paths from a local wound to sites of action in the central nervous system
• Symptoms/signs: severe painful spasms and rigidity of the voluntary muscles
• The characteristic symptom of "lockjaw" involves spasms of the masseter muscle

Firmicutes: Clostridium Botulinum Details

• Seven toxigenic types of Clostridium botulinum designated include: A, B, C1, D, E, F, and G
• Toxin is ingested with food in which spores have germinated and the organism has grown
• Absorbed by the upper part of the GI tract and passes into the bloodstream by which it reaches the peripheral neuromuscular synapses
• Toxin binds to the presynaptic stimulatory terminals and blocks the release of the neurotransmitter acetylcholine
• Symptoms of botulism begin within 18-36 hours and include weakness, dizziness and dryness of the mouth
• Nausea and vomiting may occur
• Neurologic features soon develop: including blurred vision, inability to swallow, difficulty in speech, descending weakness of skeletal muscles and respiratory paralysis

Sporosarcina Details

• Unique among endospore formers because cells are cocci instead of rods
• Strictly aerobic and contain spherical cells
• Form tetrads or packets of 8 or more
• Common in soils
• The major species is Sporosarcina ureae
• Actively decomposes very high levels of urea unlike other soil bacteria that are very sensitive to urea- very alkaline-tolerant.
• Proposed that it is a major urea degrader in nature

Tenericutes: The Mycoplasmas Details

• Key genera include Mycoplasma and Spiroplasma
• Lack a cell wall
• Many species require sterols and lipoglycans to strengthen and stabilize their membranes
• Key components of peptidoglycan are missing
• Resistant to osmotic lysis, partially because of sterols, which makes cytoplasmic membranes more stable than other bacteria
• Some mycoplasmas contain lipoglycans that stabilize the cytoplasmic membrane and facilitate attachment to receptors
• Pleomorphic with cells that may be cocci or filaments of various lengths
• A single culture may exhibit small cocci, larger swollen forms, filamentous forms which may be highly branched
• Do not stain Gram positive but are related to Firmicutes
• Contain a very small genome (500 - 1,100 bp)
• Some of the smallest organisms (0.1-0.24 µm) capable of autonomous growth
• Parasitic, inhabiting animal and plant hosts
• Several are pathogenic for humans, other animals, and plants, for example, M. pneumoniae
• Mycoplasma colonies show a characteristic "fried-egg" appearance
• Media for the culture of mycoplasmas are typically quite complex
• Mode of growth varies between liquid and agar cultures
• Energy metabolism varies (strict aerobes, facultative aerobes, obligate anaerobes)
• Not affected by antibiotics targeting cell wall

Spiroplasma Details

• Helical or spiral-shaped lacking cell wall and flagella
• Exhibit rotary screw motility or undulating motility
• Isolated from ticks, haemolymph and gut of insects, vascular plant fluids, and insects that feed on the fluids
• Cause some plant and insect diseases

Tenericutes: Phytoplasma

• Obligate parasites of plant phloem tissue and transmit using insect vectors
• They cannot be cultured in vitro in cell-free media
• Pathogens of agriculturally important plants, including coconut, sugarcane and sandalwood, causing a wide variety of symptoms ranging from mild yellowing to the death of infected plants
• They are most prevalent in tropical and sub-tropical regions of the world
• Need a vector to be transmitted from plant to plant using sap-sucking insects like leaf hoppers

Actinobacteria

• Includes Actinomyces
• Includes Corynebacterium
• Include Frankia
• Includes Gardnerella
• Includes Mycobacterium
• Includes Nocardia
• Propionibacterium
• Includes Streptomyces

Actinobacteria: General Information

• Corynebacterium, Arthrobacter, Propionibacterium, Rod-shaped to filamentous and usually aerobic Inhabitants of soil and plant materials
• Mostly harmless commensals with the exception of Mycobacterium
• Coryneform bacteria are valuable for antibiotics and certain fermented dairy products
• Coryneform bacteria are aerobic, rod-shaped bacteria that form irregularly shaped, club-shaped, or V-shaped (snapping division) cell arrangements.

Corynebacterium Details

• Gram-positive, aerobic, nonmotile, and rod-shaped
• Form club-shaped, irregularly shaped, or V-shaped with a snapping division cell arrangement
• Extremely diverse including animal and plant pathogens, saprophytes, and some pathogens

Arthrobacter Details

• Primarily exists in soils
• Undergoes conversion from rod to coccus and back
• Remarkably resistant to desiccation and starvation even though they do not form spores or other resting cells
• Are a heterogeneous group with considerable nutritional versatility
• Strains have been isolated that decompose herbicides, caffeine, nicotine, phenols, and other unusual compounds

Propionic Acid Bacteria Details

• First discovered in Swiss cheese, where CO2 production forms holes and propionic acid adds flavor
• Gram-positive anaerobes
• Ferment lactate, carbohydrates, and polyhydroxy alcohols
• Produces large amounts of Propionic acid
• Products are primarily propionate, acetate and CO2
• Uses metabolic strategy called secondary fermentation
• Obtains energy from fermentation products, for example, lactate produced by other bacteria

Actinobacteria: Mycobacterium Details

• First discovered by Robert Koch
• Rod-shaped organisms, exhibit acid-fastness due to mycolic acids which are complex, highly branched lipids on their surface
• Not readily stained by Gram stain due to high surface lipid content
• Cells are somewhat pleomorphic and may grow filamentous
• Separated into two groups: slow and fast growers
• Classified into three groups based on pigmentation
• Mycobacteria form tight, compact, wrinkled colonies on agar
• Classified into three groups based on pigmentation, some produce yellow carotenoid pigments that aid in identification and may protect against oxidative damage
• Have simple nutritional requirements
• Some are human pathogens, where virulence of Mycobacterium tuberculosis is correlated to cordlike structure
• Cordlike structure is related to glycolipid on cell surface Acid Fastness of mycobacteria is where their growth cycle possesses the distinctive staining property called
• Due to the presence of mycolic acids on the surface of the mycobacterial cell wall -Mycolic acid is found only in the genus Mycobacterium
• The carboxylic group of the mycolic acid reacts with fuchsin dye causing acid fast microorganisms to appear red
• Mycobacterium can be pleomorphic, and may undergo branching or filamentous growth
• These filaments become fragmented into rods or coccoid elements on the slightest disturbance.

Filamentous Actinobacteria

• Key genera: Streptomyces, Actinomyces and Nocardia
• Include Streptomyces and Actinomyces as the key genera
• Filamentous, aerobic Gram-positive bacteria
• Commonly found in soil
• Most actinomycetes form spores called conidia
• The manner of spore formation varies and is used in separating subgroups
• Mycelial spore-forming is thus both of phylogenetic and taxonomic importance Produce hyphae (filaments) and mycelium, which is an aerial network of filaments with spores for survival and dispersal

Streptomyces Details

• Over 600 species
• Streptomyces spores are callled conidia
• Strict aerobes that produce colored colonies
• Streptomyces filaments are typically 0.5-1.0 µm in diameter, are of indefinite length, and often lack cross walls in the vegetative phase
• Streptomyces grow at the tips of the filaments, often with branching
• The vegetative phase thus consists of a complex, tightly woven matrix, resulting in a compact, convoluted mycelium and subsequent colony
• 50% of all isolated Streptomyces produce antibiotics with over 500 distinct antibiotics produced by Streptomyces
• Some produce more than one antibiotic
• Genomes are typically quite large (8 Mbp and larger) due to many genes required for antibiotic synthesis
• Primarily soil microorganisms, responsible for earthy odor of soil (geosmin)
• Alkaline/neutral soils more favorable
• Found more in well-drained than waterlogged soils
• Knowledge of the ecology of Streptomyces remains poor
• Streptomyces produce extracellular enzymes
• They are nutritionally versatile chemoorganotrophs, growing on a variety of carbon sources including fats, proteins, sugars, alcohols, organic acids, and some aromatic compounds and rarely need growth factor requirements
• Strict aerobes whose growth in liquid culture is stimulated by forced aeration
• Sporulation only occurs when grown on the surface of a solid substrate or when organisms form a pellicle on the surface of an unshaken liquid culture.

Geosmins

• Group of metabolites produced by Streptomycetes in the soil which cause the familiar earthy odor of soil
• Are sesquiterpenoid compounds (unsaturated ring compounds containing carbon, oxygen and hydrogen)
• Example, trans-1, 10-dimethyl-trans-9-Decalol
• Produced by some cyanobacteria

Antibiotics Produced by Streptomyces

• Often on agar plates with mixed cultures including streptomyces where adjacent colonies of other bacteria show zones of inhibition
• 50% of all Streptomyces isolated shown to be antibiotic producers
• Over 500 distinct antibiotics have been shown to be produced by Streptomyces, where some strains produce more than one
• Over 60 streptomycete antibiotics used in human & veterinary medicine
• Among the most popular antibiotics are streptomycin & tetracycline

Streptomycin

• An aminoglycoside that is produced by S. griseus
• Aminoglycosides contain amino sugars linked by glycosidic bonds
• Active against most gram-negative bacteria
• Targets the 30S sub-unit of the ribosome thus inhibiting protein synthesis
• Not widely used due to serious side effects such as neurotoxicity and nephrotoxicity