BACTERIA

Bacteria on the point of a pin.

DOMAIN BACTERIA – Kingdom Eubacteria

•Single-celled prokaryote•Have peptidoglycan (protein carbohydrate) cell walls•Can form colonies of clumps or filaments.•Three basic shapes: Cocci (round), bacilli (rod), spirilla (spiral)

Strepto – occurs in chains

Staphylo – occurs in clusters

Cell wall – protects the cell & gives it shape.

Outer membrane – protects the cell against some antibiotics (only present in gram-negative)

Cell membrane – regulates movement of materials into & out of the cell; contains enzymes important tocellular respiration.

Plasmid – circular piece of DNA that contains some genes obtained throughgenetic recombination.

Capsule & Slime layer – protect the cell & assist in attaching the cell to other surfaces.

Classifying Prokaryotes

DOMAIN ARCHAEA – Kingdom Archaebacteria

•Genes that resemble eukaryotic genes & some thatresemble prokaryotes.

•Have unusual lipids in their cell membranes

•Found in EXTREME environments – but also in non-extreme

•Cell walls lack peptidoglycan

•Have introns in their DNA

•Single-celled prokaryote

Archaebacteria

Methanogens•Convert H2 & CO2 into methane CH4

•Anaerobic bacteria•Found in bottoms of swamps, sewage, & intestinaltracts of animals.

Extreme Halophiles•Salt loving•Found in Great Salt Lake & Dead Sea

Thermoacidophiles•Live in extreme acidic & hot environment

Prokaryote Evolution

•Fossil Evidence indicates bacteria existed about 3.5billion years ago.

•Eukaryotes existed about 2.5 billion years ago.

•Bacteria evolved to adapt to almost any environment,from ocean trenches to thermal vents.

Identifying Prokaryotes

Prokaryotes are identified by characteristics such as:

•the way they obtain energy

•shape

•type of cell wall

•the way they move

Identifying Prokaryotes

Gram Staining

Staining bacteria

Imagine trying to see and study 2 distant white golf balls against a white background. This would be similar to trying to view bacteria without staining them.

Staining increases contrast and resolution.

Most microorganisms are colorless and difficult to view with bright-field microscopes (simple and compound microscopes). Stains are used to make microorganisms and their parts more visible because the stains increase contrast between structures and between the specimen and its background.

Staining – coloring specimens with stain or dye.

Before stain can be applied, a microbiologist must attach the specimen to the slide. This is called making a smear and fixing it to the slide. If the organisms are growing in a liquid, a small drop of the liquid is spread across the surface of the slide. If the organisms are growing on a solid surface, such as agar, they are mixed with a drop of water on the slide.

In either case, a smear is formed. This is a thin film on the slide.

The smear is dried and then attached or fixed to the surface of the slide using either heat or chemicals.

Robert Koch developed heat fixation more than a hundred years ago.

In this process, the slide is passed (smear side up), through the flame of a bunsen burner.

Chemical fixation involves applying methyl alcohol or formalin to the smear for one minute.

Simple stains – these are composed of a single basic dye such as crystal violet, safranin, or methylene blue. They are considered “simple” because they require nothing more than soaking the smear in the dye for 30-60 seconds and then rinsing with water. A properly fixed slide will remain attached to the slide during this treatment.

After carefully blotting the slide dry, the smear can be observed under the microscope. Simple stains are used to determine size, shape, and arrangement of cells

Gram stain – in 1884 the Danish scientist Hans Christian Gram developed the most frequently used differential stain, which now bears his name. The Gram stain differentiates between two large groups of microorganisms:

Purple staining : Gram positive cells

Pink staining : Gram negative cells

These two types of cells differ greatly in the chemical and physical structure of their cell walls. Typically a Gram stain is the first step a medical laboratory technologist performs to identify a pathogen.

Steps in Gram Staining

1. Flood the smear with the basic dye crystal violet for 1 minute. Rinse with water. Crystal violet is the primary stain and colors all cells.

2. Flood the smear with iodine for 1 minute. Rinse with water. Iodine is a mordant – a substance that binds to a dye and makes it less soluble. After this step, all cells remain purple.

3. Rinse the smear with a solution of ethanol and acetone for 10-30 seconds and rinse with water. This solution acts as a decolorizing agent. It breaks down the thin cell wall of Gram-negative cells, allowing the stain and mordant to be washed away. These cells are now colorless. Gram-positive cells will still be purple.

4. Flood the smear with safranin for 1 minute. Rinse with water. This red counterstain provides a contrast to the primary stain. Gram negative cells now are pink but Gram-positive cells remain purple.

In step 4 the safranin is probably absorbed by all cells, but the pink color is masked by the darker purple dye already present in Gram-positive cells.

Gram staining works best with young cells. Older Gram-positive cells bleach more easily than young ones and can stain pink which makes them appear to be Gram-negative. This is known as a false Gram-negative reaction. Smears should, therefore, be made using freshly grown bacteria approximately 24 hours old.

Some variations exist for staining, including:

Using 95% ethanol instead of the ethanol-acetone mixture.

Dissolving the safranin in the ethanol and simultaneously decolorizing and counterstaining.

Metabolic Diversity

Chemoheterotrophs – organisms that must take in organic molecules for both energy and a supply of carbon.

Photoheterotrophs – organisms that are photosynthetic, but still need to take in organic compounds as a carbon source.

Chemoautotrophs – make organic carbon molecules fromcarbon dioxide and other compounds using energy fromchemical reactions.

Photoautotrophs – use energy from sunlight to convert carbon dioxide and water to carbon compounds.

Releasing Energy

Obligate aerobes – organisms that require a constant supply of oxygen in order to live. EX: Mycobacterium tuberculosis, the bacterium that causes tuberculosis.

Obligate anaerobes – do not require oxygen; some may be killed by O2! EX: Clostridium botulinum, found in soil and can grow in canned food causing fatal food poisoning.

Facultative anaerobes – can survive with or without O2.EX: E. coli, can live in the large intestines or contaminated water.

Growth & Reproduction

Conjugation – the process by which two living bacteria bind together and one bacterium transfers genetic information to the other.

Growth & Reproduction

Endospore – protects the cell against harsh environmental conditions, such as heat and drought. May allow the bacterium to survive for thousands of years.

Importance of Bacteria

Decomposers – break down the nutrients in dead matter.

Nitrogen – Fixing Bacteria (Nitrogen fixation)•Live freely & symbiotically with plants•Rhizobium; converts N2 to a form of nitrogen plants can use.•Found in legumes (bean type plants)

Importance of Bacteria

Phylum Cyanobacteria

•Photosynthetic

•Encased in jelly-like substance & cling together

•Contain enzymes for fixing atmospheric nitrogen & makeit available to plants.

•Some thrive on phosphates & nitrates in lakes & ponds.

Importance of Bacteria

Phylum Cyanobacteria

Eutrophication or Population Bloom – the suddenincrease in the number of cyanobacteria due to a highavailability of nutrients.

When cyanobacteria die, they are decomposed byheterotrophic bacteria. This increase in heterotrophicbacteria depletes O2 in the water, harming otherorganisms that live there.

Human Uses of Bacteria

•Food – baking & beverages•Clean up oil spills, rivers & streams•Make drugs•Aid in digestion

Bacteria & Disease

Pathogen – bacteria that cause disease or a disease causing agent.

Exotoxins – toxic proteins secreted by bacterial cells, includes some of the most potent poisons known.

Clostridium botulinum – one gram of the exotoxin that causes botulism could kill 1,000,000 people!

Staphylococcus aureus – harmless, found on skin; if it enters the body through a wound it can cause layers of skin to slough off, vomiting, severe diarrhea & deadly toxic shock syndrome.

Bacteria & Disease

Endotoxins – are NOT secretions; but components of cellwalls in bacteria: glycolipids, which are large molecularcomplexes of polysaccharides & lipids.

All endotoxins induce the same general symptoms: fever,aches and sometimes a dangerous drop in bloodpressure (shock).

Salmonella – produces endotoxins that cause food poisoning & typhoid fever.

Bacteria & Disease

Lyme Disease

•Most widespread pest carried disease in U.S.•Caused by Borrelia burgdorferi, a bacterium carried byticks that live on deer & field mice.•Antibiotics can cure the disease if administered within amonth of exposure.•If untreated, it can lead to arthritis,heart disease & nervous disorders.

Bacteria & Disease

Yersinia pestis – the bacterium that caused bubonicplague throughout Europe.

It also played a role in battle, when armies hurled thebodies of plague victims into enemy ranks.

Bacteria & Disease

Bacillus anthracis

•Found naturally in soil where grazing animals such as sheep, cattle & goats are located.•Easy to obtain, grow & it forms hardy endospores that can survive for years.•Upon entering the bloodstream, anthrax will actively metabolize & multiply.•Anthrax will release 3 proteins that combine to form a toxin that destroys body tissues & cells of the immune system.•Modes of entry: cutaneous, inhalation, ingestion

Bacteria & Disease

Bacillus anthracis

Antibiotics

Antibiotics are drugs that combat bacteria by interfering with various cellular functions

Some bacteria are antibiotic-resistant and destroy antibiotics, or prevent entry of the antibiotic into the cytoplasm.Penicillins such as penicillin and amoxicillin

Cephalosporins such as cephalexin (Keflex)

Macrolides such as erythromycin (E-Mycin), clarithromycin (Biaxin), and azithromycin (Zithromax)

Fluoroquinolones such as ciprofloxacin (Cipro), levofloxacin (Levaquin), and ofloxacin (Floxin)

Sulfonamides such as co-trimoxazole (Bactrim) and trimethoprim (Proloprim)

Tetracyclines such as tetracycline (Sumycin, Panmycin) and doxycycline (Vibramycin)

Aminoglycosides such as gentamicin (Garamycin) and tobramycin (Tobrex)

Antibiotics

Bacteria can be testedfor their sensitivity toantibiotics by growingthem in a petri dish withpaper disks containingdifferent antibiotics.

As the antibiotics diffuseinto the agar, thebacteria’s growth will beinherited by theantibiotics if the bacteriaare sensitive to thatantibiotic.