Lecture Notes - General Microbiology

Lecture notes from the Autumn 2006 semester at London Metropolitan University, Module BM1003N. Please cite me if you quote from my notes. I'd appreciate being told about this, as well (you don't have to, but it's nice to know).

Thursday, October 22, 2009

BG Additional

P C Culmer
Additional Information.

I have a good amount of experience in administration, having taken a basic level NVQ in the subject, and then worked as the project manager of one building company, and as the general administrator of another. These roles entailed the usual spectrum of administrative tasks, including fairly extensive bookkeeping in the latter. I have formally studied health and safety many times in various theatres, including laboratory, office, catering, and industrial settings.

I am highly literate and numerate, and familiar with the everyday use of Microsoft Office. I also have some experience of SPSS. Having recently graduated, I have plenty of recent experience of writing reports at varying lengths and levels of detail, and also of making presentations of information. I feel that my recent experience as a mature student may help me relate to students in my day to day work.

I believe in dealing with people as individuals, not as representatives of a class, gender, or culture, and feel that my experience in a disabled person’s organization may give me an enhanced understanding of the crucial difference between the meeting of a persons differing needs and condescension. I feel that anyone dealing with students, colleagues, the general public, or any other client or potential client, should have foremost in their mind that they are there to meet that person’s needs, and should act accordingly.

With regard to the specifics of the person profile:

Education: I have a Bsc (Hons) in Biochemistry. I am highly numerate, with a basic NVQ in Business Administration, and several years experience.

Knowledge and Skills: I am very familiar with Microsoft Office, experience in making presentations, and feel that I can communicate effectively in writing. I can provide examples of my work for your consideration. I am experienced in all aspects of general administration, and also in bookkeeping. I have some knowledge of database work, though this is not as extensive as I would prefer.

Experience: I have several years experience of working in an office, and of providing support and information.

Personal Attributes: My base line approach to work is to be conscientious, enthusiastic, client-centred, and loyal. The fact that I have Asperger Syndrome tends to make my default approach a methodical one, and I am more than willing to be flexible and to undertake as much additional training as is available.

I would wish for nothing less than total success for any organization of which I was a part, and to play my part in that success.

In short, I feel that my abilities and personal characteristics would be an asset to the Research Centre, and that being a part of it would benefit my personal development.

Tuesday, December 12, 2006

Week 11

Today's Topics:
○ Biotechnology

○ Exam: 16/1/07

Lecture Topic:

○ What is it? The use of microorganisms, usually on a large scale, to produce valuable products (eg beer, cheese, antibiotics)
○ These are usually enhancements of natural reactions, amplified for better yield.
○ Genetic manipulation now permits production of products that microorganisms do not naturally produce.
○ Main organisms used:
§ Fungi (yeasts & moulds)
§ Streptomyces spp.
§ Escherichia coli
§ Bacillus subtilis
○ Microorganisms are often altered by mutation or recombination, to achieve high metabolic specialisation.
○ Requirements:
§ Must be able to grow on large scale
§ Should grow rapidly, and yield product in a short time
§ Must be able to grow on cheap culture, ideally waste from another process.
§ Should be amenable to GM.
§ Must not be pathogenic
○ Examples:
§ Antibiotics
§ Vitamins
§ Enzymes
§ Alcohol
§ Vinegar
§ Bioconversion
○ Primary Metabolites
§ Produced during growth of culture (eg ethanol)
§ Result of growth
○ Secondary Metabolites
§ Produced at end of growth - at, near, or in, stationary phase.
§ Survival mechanism (eg antibiotics, to kill off competing organisms, or enzymes to free up more food)
§ Not essential for growth & reproduction
§ Dependent on growth conditions (especially medium)
§ Often possible to get massive overproduction.
○ Vessels used are called fermentors, and often hold hundreds of thousands of litres.
○ Two classes: ærobic and anærobic.
○ Antibiotics:
§ Substance produced by microorganisms that kill or inhibit others.
§ Most useful antibiotics produced by filamentous fungi and by Actinomycete group (eg Streptomyces spp.)
○ Penicillin
§ Original: Penicillium notatum - low yield (1-10μgcm-3)
§ 1943: Penicillium chrysogenum, further improved
§ Now Penicillium chrysogenum, in aerobic stirred fermentors (50,000μgcm-3)
○ β-lactam antibiotics: Penicillin & its relatives.
§ Biosynthetic penicillins: result of biosynthesis
§ Semisynthetic penicillins: chemically modified
§ Interfere with cell wall production.
○ Vitamins & amino acids
§ Most made by chemical synthesis
§ A few are too complex for this, eg B12 (Propionibacterium spp. & Pseudomonas spp.), and riboflavin (Ashbya gossypii)
§ Amino acids are regulated by inhibitory feedback - mutant species resist this inhibition, and thus overproduce. (Eg use of S-aminoethylcysteine to produce resistant species, where lysine will not inhibit production by aspartokinase).
○ Extracellular enzymes (excreted into medium)
§ Capable of digesting e.g. cellulose & starch
§ Useful in food, pharmaceutical & textile industries
§ Proteases and lipases from alkaliphilic Bacillus lichenformis used in detergents (e.g. biological washing powders)
§ Amylases and glucoamylases: production of glucose from starch.
○ Microbial bioconversion - biotransformation
§ Used for specific reactions beyond organic chemistry
§ E.g. Rhizopus nigricans converts progesterone to 11α-hydroxyprogesterone, which is then converted chemically to hydroxycortisone and then cortisone.
○ Genetically engineered microorganisms
§ E.g. use of E. coli to produce human growth hormone & insulin
§ Contain gene from another organism
§ Grow rapidly on simple media
§ Produce massive amounts of blood proteins, hormones, etc.
§ Main hosts: Escherichia coli, Bacillus subtilis, Saccharomyces cerevisiae.
§ Work from mRNA (to avoid introns), and use reverse transcriptase to make DNA, and then ligate this into a plasmid.
○ Vaccines
§ Killed or attenuated pathogens (or fractions) that produce immunity.
§ Body develops antibodies against surface proteins
§ Genetically engineered antigens
□ Cheaper
□ Easier to purify
□ Free from other proteins
§ E.g. Hepatitis B antigen cloned into S. cerevisiae
○ BT toxin
§ Ulcerates gut wall in larvae
§ Gene transferred to P. flourescens, which inhabits rhizosphere. Innoculation protects roots from pests.
§ Then transferred to plants
§ Constant exposure stimulated BT resistant pests.
○ Herbicide resistance:
§ E.g. gene for resistance to glyphosate transferred into plants, to allow glyphosate to be used to suppress pests without affecting crop.

Saturday, December 02, 2006

Colony type 1 - LM400

Colony type 1 - LM1000

Friday, December 01, 2006

Food Culture Plates - observations made 28/11/06

Culture from Salad, on MacConkey Agar, at 1E-3

Culture from Salad, at 1E-4

Culture from Salad, at 1E-5

Culture from Salad, at 1E-5 - culture surface of agar.

Culture from Salad, at 1E-6. I identified 3 cultures on this plate -

Colony type 1 was circular, raised, papillate, red, with an entire edge, and about 4mm in diameter. Under the microscope, it had strings of gram positive bacilli

Colony type 2 was irregular, flat, matt, pink, with an entire edge, and about 15mm in diameter. Under the microscope, it had individual, gram negative bacilli.

Both of these colonies tested positive for catalase, and negative for oxidase.

Colony type 3 was circular, convex, shiny, red, with an entire edge, and about 2mm in diameter.

Tuesday, November 28, 2006

Week 9

Once again, this weeks notes are largely copied from Dr Matewele's notes.
Lecture Topic:
Medical microbiology
○ Microbial flora may produce products such as bacteriocins that inhibit other organisms, and so protect us from pathogens
○ Microbial flora also stimulate the immune system, e.g. antibodies against "friendly" E coli also protect against pathogens such as E coli O157:H7.
○ Stem cells can be described a pluripotent haemopoietic (blood making) cells that can form different type of blood cells, eg white blood cells and red blood cells. T cells develop in a primary lymphoid organ called the thymus, and B cells develop and mature in another called the bone marrow. Immune responses take place in secondary lymphoid organs, e.g. lymph nodes for responses in tissues, and the spleen for responses in blood.
○ Animals reared in sterile environments do not develop the lymphoid system properly, as it is not challenged.
○ Colonisation: where microorganisms multiply in the body - if these are non pathogenic, they become part of the microbial flora. If they are pathogenic they cause an infection - this does not necessarily mean a disease. A disease develops when the body functions are affected.
○ Sources of pathogens:
§ Human to human - eg S aureus
§ Animal to human - eg rabies, influenza - also known as zoonotic infections. - note: influenza has about 8 chromosomes, which recombine - eg from dogs & poultry
§ Soil to humans - e.g. Clostridium tetani
§ Water to human - eg Vibrio cholerae causes cholera, Legionella pneumophila causes Legionnaires Disease.
§ Air to humans - e.g. Mycobacterium tuberculosis
§ Insect vectors - Malaria - transmitted by anopheles mosquitoes - Plasmodium.
○ Epidemiology - practiced by epidemiologists, who study and monitor the incidence and control of diseases. Incidence is classified as:
§ Outbreak - where it is sudden, within a segment of a population.
§ Epidemic - unexpected occurrence of a disease in a large population. May be common source, sudden but limited in extent (e.g. cholera from a contaminated pump) or propagated, gradual growth (e.g. Common cold or Spanish flu).
§ Pandemic - world wide.
§ Endemic - constantly at low level in community.
○ The pathogenicity of the microorganism is associated with virulence factors, such as capsules, which resist phagocytosis, toxins, e.g. in Corynebacterium diptheriae, often produced by bacteriophages that infect bacteria.
○ The microorganisms initially interact with the host either by attaching to epithelial cells ( e.g. in the genitourinary tract for Neisseria gonorroea, or digestive tract for Salmonella spp.)
○ Invasive microorganisms may produce enzymes e.g. protease to break down protein, the popular term "flesh eating bacteria" for Streptococcus pyogenes. In addition it produces streptokinase, a fibrinolysin, to dissolve clots.
Staphylococcus aureus produces coagulase, which produces coagulation. The fibrils in the coagulant can then be used for protecting the microorganism. It also produces a protein that binds to the fc (fraction crystallisable) region of antibodies, preventing an immune response.
Clostridium perfringens produces lecithinase, which breaks down lecithin in cell membranes, causing cytolysis, and hyaluronidase, which breaks down hyaluronic acid, the cement binding cells together.
○ Thus, some organisms, such as Strep pyogenes, Staph Aureus, and C perfringens are invasive.
○ Others, such as C perfringens produce a theta toxin, which causes diarrhoea, and S aureus produces toxic shock syndrome.
○ Beneficial effects:
§ Vaccines
§ Antibiotics
§ Human proteins (e.g. insulin)
§ Transformation of steroids
§ Food Production (Brewing, Cheese, etc)
§ Probiotics can incorporate "Friendly bacteria" into animal feeds, e.g. Lactobacillus to reduce or control Salmonella,
§ DNA vaccines, where the dna has a code for the parasite proteins
§ Bacteriophages to control pathogens such as E coli O157
§ Prevention of ulcers by vaccination against Helicobacter pylori
§ An understanding of how prions cause disease can help to identify molecules that inhibit prion mechanism.

Thursday, November 23, 2006

Results of Mouthwash Assay - observations made 21/11/06

Mouthwash A: zones of inhibition: 9mm, 7mm, 8mm
Mouthwash B: zones of inhibition: 12mm, 13mm, 11mm
Mouthwash C: zones of inhibition: 14mm, 16mm, 14mm

Week 8

Today's Topics:
○ Protozoa

Lecture Topic:
○ Single celled microorganisms
○ No cell wall
○ Shape maintained by ectoplasm
○ My undergo encystation in adverse conditions - not a cell wall, but similar
○ Parasitic protozoa may also encyst when changing hosts (different life parts of life cycle) (eg Plasmodium)
○ Chemoheterotrophs.
○ Use phagocytosis to take up nutrients.
○ Classification based on shape, presence or absence of cilia or flagella.
○ Some may also use locomotive pseudopodia.
○ May reproduce sexually and/or asexually.
○ Example: Cryptosporidium spp. - found in cattle. Oocysts defaecated, and can end up in water supply if raw or insufficiently treated. Cryptosporidosis can cause diarrhoea when merozoites multiply within cytoplasmic membrane (not intracellularly). This can be persist in immunocompromised individuals.
○ Example: Toxoplasmosis, caused by Toxoplasma gondii. Usual life cycle is oocysts defaecated by cats, and after 3-4 days, sporozoites develop, which are eaten by mice. Sporozoites become haploid trachyzoites, which infect cats when they eat the mice. Exposure to oocysts can infect humans, causing problems in pregnancy or if immunocompromised. Effects on foetus may include congenital blindness, brain damage, or even abortion.

○ Eukaryotes vs. prokaryotes
§ Nuclear membrane
§ Histones & non histones
§ Mitochondria
§ Golgi
§ Endoplasmic reticulum
§ 80s ribosomes instead of 70s
§ Can have sexual reproduction (meiosis to produce haploid cells)
□ Diploid cell --> meiosis --> haploid cells --> fusion: plasmogamy --> dikaryon --> karyogamy --> diploid zygote.
○ Algae
§ Cell walls: cellulose, pectin
§ Chlorophylls
§ Other pigments (phycobilins)
§ Generally multicellular except Euglynophyta spp.
§ Storage products (lipids, starch, etc)
○ Protozoa
§ Alternate hosts
§ No cell walls
§ Classified by morphology (eg flagella, cilia, pseudopodia)
○ Fungi
§ Cell walls: chitin
§ Named according to spores in perfect (sexual) stage.

Medical Microbiology
○ Only 10% of cells in body are human.
○ Microbial flora:
§ Staphylococcus epidermidis - skin
§ Escherichia coli - colon - synthesises vitamins
§ Relationships:
§ Mutualism (both host and microorganism benefit)
§ Commensalism: (latin: at the same table) host neither benefits, nor is harmed. (can be argued to be mutualistic, as presence prevents colonisation by pathogens)
§ Parasitism: microorganism benefits at expense of host. This is associated with pathogens (Greek: suffering makers)
§ Some microbial flora may be opportunistic pathogens if they colonise or replicate in usually sterile areas (e.g. blood, brain, csf, deep tissues)

Tuesday, November 14, 2006

Week 7

Note: most of the latter part of this week's notes were taken down verbatim from Dr Paul Matewele's onscreen notes, with a little contraction in places.

Today's Topics:
○ eukarya

Lecture Topic:
○ Differences:
§ Eukarya
□ "True nut"
□ Organelles
□ Golgi
□ Mitochondria
□ Endoplasmic Reticula
□ Nuclear membrane
□ Nucleus
□ Histones
□ Non histones
□ Multiple chromosomes (mainly)
□ Use mitosis & meiosis
□ 80s ribosomes
§ Prokarya
□ "Before nut"
□ Single chromosome (mainly)
□ 70s ribosomes
○ In meiosis, progeny is haploid, and diploid status is restored by sexual reproduction. Prokarya do not use sexual reproduction.
○ Mitosis - non sexual reproduction. Diploid parent produces two diploid offspring. Mitosis (genetic reproduction) is followed by cytokinesis (cell division). Mitosis --> Cytokinesis --> G1 phase (growth) --> S phase (DNA synthesis) --> G2 phase (more growth) --> Mitosis.
○ Meiosis - sexual reproduction. Gametes fuse (cytoplasm only) - plasmogamy - forms dikaryon (two nuclei) --> Nuclei fuse - karyogamy - forms diploid zygote --> meiosis - forms four haploid gametes --> gametes fuse.
○ Plasmogamy and karyogamy may be referred to collectively as syngamy.

○ Different types of eukaryotes:
○ Fungi, protozoans, algae.
○ Fungi and algae have cell walls. Fungi generally have chitin, whilst algae have cellulose or cellulose and pectin. Protozoans have no cell wall. The morphology of protozoa is maintained by an ectoplasm - gelatinous material the cell membrane - this is inside cell membrane, unlike the slime capsule that bacteria use.
○ Fungi
§ Can be divided into two classes - fungi perfecti and fungi imperfecti.
§ Fungi perfecti have a sexual stage, aka the perfect stage, whereas fungi imperfecti have no sexual reproduction.
§ The species or genus names are also associated with the type of spores produced during sexual reproduction, e.g. Bacidiomycetes produce basidiospores. An example is Agaricus bisporus, the common mushroom. Ascomycetes produce ascospores; examples include Penicillium notatum and Aspergillus niger. Oomycetes produce oospores; examples include Lagenidium callinectes and Phytophthora infestans.
§ An example of fungi imperfecti is the class of dermatophytes, which cause skin infections such as Trichophyton rubrum, which causes athletes foot.
§ Characterisation of fungi is generally based on the types of spores produced and the spore bearing structures.
§ Spore grows into a germ tube, which then becomes a hypha. Mycelia branch off of the hyphae, and spore bearing structures grow on the mycelia.
§ Example: P. infestans, an oomycete. (see diagram in lecture notes book, page 21).
○ Algae
§ These can carry out photosynthesis.
§ The type of chlorophyl - a / b / c / c1 / c2 / d - can be used in their classification.
§ Algae in deep oceans cannot absorb light directly; they have pigments called phycobilins. Some are red - phycoerythrins, and others are blue - phycocyanins.
§ The storage products can also be used in classification, e.g. starch, lipids, and glycogen.
§ The cell wall is predominately cellulose.
§ Some algae are flagellated, and have various habitats. This is another source of classification.
§ Example: Ulva spp. (see diagram in lecture notes book, page 22)

Week 6

Today's Topics:
○ Bacterial Taxonomy

Lecture Topic:
○ 2 major functions: identification and classification
○ Basic unit: species
○ Species are grouped into a genus
○ Genera are grouped into a family
○ Families are grouped into an order.

○ Identification needs a pure culture.
○ Procedure:
§ Examine morphology of culture
§ Examine morphology of cells (eg methylene blue stain) - note: use lowercase for morphology, to distinguish from genera.
□ Single sphere - cocci
□ Pairs - diplococci
□ Chains - streptococci
□ Cluster - staphylococci
□ Single rod - bacilli
□ Curved rod (comma) - vibrio
□ Wavy rod (fixed) - spirillum
□ Wavy rod (flexible) - spirochæte
□ Determine composition & structure of cell wall - gram stain and/or acid fast stain
□ Acid fast species are gram+, but do not take gram stain well due to lipids in cell wall. Good for identifying tb and leprosy, which can be distinguished by differing acid resistance.
□ There are also stains that will differentially stain features such as flagella, spores, etc.
§ Biochemical tests, eg:
□ Sugar tests - can species use a particular carbohydrate?
¨ Liquid culture containing only one carbon source - individual CHO
¨ pH indicator for eg lactic acid
¨ Inverted tube (durham tube) to collect gas (eg CO2)
□ Gelatin tube
¨ Stab innoculation
¨ some organisms metabolise gelatin
¨ Different growth patterns
□ Nitrate reduction (to nitrite or nitrogen)
§ You can then select from:
§ Physiological tests
□ Factors such as
□ optimum temperature eg:
¨ Mycobacterium tuberculosis prefers 37C
¨ M. avium prefers 42C
¨ M. marinium prefers 25C
□ pH
§ Pathological tests:
□ Which organisms does it affect?
¨ Staphylococcus epidermidis - rarely pathogenic
¨ S. aureus - can cause several diseases
¨ (don't actually need for these two - se is white, sa is golden)
§ Immunological tests, eg for species of Salmonella
□ Test for reaction with specific antibodies.
○ Standard system:eg:
§ Escherichia coli
§ Genus: Escherichia
§ Species: coli.
§ Conventions:
□ written in italics or underlined.
□ Genus has initial capital.
□ Genus may be abbreviated to initial, or shorter version where this will not cause confusion.
□ Several species from one genus may be referred to as, eg Escherichia spp.
§ Current bacterial classification: Bergey's Manual of Systematic Bacteriology, 2nd ed (2001)
□ Volume 1: Archaea
¨ Eg Halobacterium spp., Thermococcus spp.
□ Volume 2: Proteobacteria (gram negative)
¨ Eg enteric bacteria & Pseudomonas spp.
¨ P. florescens - fluorescent
¨ P. aeruginosa - cause some effects of cystic fibrosis
□ Volume 3: Low G+C Gram Positive bacteria
¨ Eg lactic acid bacteria (such as Lactobacillus spp.), Bacillus spp., Clostridium spp., Streptococcus spp.
¨ B. subtilis
¨ C. sporogenes
□ Volume 4: High G+C Gram Positive bacteria
¨ Eg Mycobacterium Spp., Streptomyces spp.
¨ Streptomyces spp. Are good sources of antibiotics.
□ Volume 5: Everything else
¨ Eg spirochaetes, Bacteriodes spp.

Week 5

Today's Topics:
○ Prokaryote Cell Biology

Lecture Topic:
During the lecture, take notes here.
○ Viable count calculation checker on WebCT
○ Average adult has approximately 1014 cells.
○ 10% of these are human cells
○ 10% are bacteria
○ Bacteria on skin prevent colonisation by pathogens.
○ Approx 2000 people die per year in UK from undiagnosed infections.
○ Some bacteria have multiple chromosomes - most eukaryotes have linear chromosomes.
○ Plasmids are rare in eukaryotes - yeast has one.
○ Flagella
§ Movement is termed motility - something that can move is motile.
§ Bacteria swim in straight lines - flagella in cooperation - if not in cooperation, bacteria tumble.
§ Flagella arrangements:
□ Single, at one end, or at both ends - polar
□ All over cell - peritrichous
§ Not all species are motile - flagella are not universal.
§ Protein
○ Pili
§ Generally found in gram-negative bacteria
§ Allow bacterium to adhere to solid surfaces (e.g. in alimentary canal)
§ Sex pilus allows transfer of genetic bacteria (covered in detail in genetics II)
§ Protein
○ Capsule / slime layer
§ A matter of degree - a slime layer is a thin capsule
§ Thickness varies by species and environment
§ Protects cell from desiccation
§ Allows cell to adhere to solid surface
§ Allows cell to evade immune system - resists phagocytosis.
§ Generally associated with pathogens.
§ Protein and/or Carbohydrate
○ Cell wall
§ Found in almost all species
§ Gram staining depends on nature of cell wall.
§ Gram positive cells homogenous 20-80 nm wall.
§ Gram negative cell walls in layers:
□ Outer membrane - 8-10 nm
□ (periplasmic space) - widely variable or absent
□ Rigid layer - 1-3 nm
§ Peptidoglycan is only in bacteria, not in eukarya or archea. (aka mucopeptide).
§ PG made of n-acetyl glucosamine & n-acetyl muramic acid - alternating units in linear chains, cross-linked with short proteins, (about 10aa), in a 3d mesh.
§ Teichoic Acid - mixture of sugar alcohols, cross-liked with phosphates
§ Lipopolysaccharides - often toxic, and can be cause of symptoms of gram negative infections. - trigger immune response.
○ Plasma membrane
§ Much the same as in eukaryotes
§ Energy generation is carried out on PM, as no mitochondria.
○ Mesosome
§ Infolding of membrane to increase area for energy generation.
○ Storage Granules
§ Volutin - phosphorus store - stained pink by methylene blue
§ Polysaccharides - iodine stains starch blue , glycogen red-brown
§ Also sulphur, gases
○ Endospores - survival mechanism - allow cell to survive adverse conditions (not reproduction)

Week 4

Today's Topics:
○ Viri

○ Today's practical - video on lab and microbial techniques, aseptic technique, etc.
○ Group F - benches 23/24

Lecture Topic:
During the lecture, take notes here.
○ Viral Infection of cell
§ Attachment
§ Penetration/ infection

○ HIV replication
§ Attachment - GP120:CD4
§ Fusion/ endocytosis
§ Uncoating
§ Reverse Transcription (cf Central Dogma)
§ RNA-->DNA (Reverse Transcriptase)
§ Integration of DNA into genome (integrase)
§ Transcription (viral RNA & proteins)
§ Assembly
§ Release (budding)
§ 1 T-lymphocyte can produce 10M HIV virions in 36 hours, before dying from exhaustion
○ Outcome of infection
§ HIV: 2-3 years
§ Karposi's Sarcoma - tumour of blood vessel lining
§ Long-term latency - eg Chicken Pox & Shingles - Herpes Zoster (Varicella zoster)
§ Possible effects:
□ Tumour (eg Rous sarcoma or papilloma)
□ Lysis (eg polio)
□ Persistent infection (eg HIV)
□ Latent infection (eg Epstein Barr virus or herpes simplex)

○ Subviral agents
§ Viroids:
□ Naked RNA
□ circular
□ 240-400 nucleotides
□ cause at least 20 plant diseases (eg potato spindle tuber or coconut kdang kdang)
□ Only infect plants.
§ Virusoids
□ Larger than viroids
□ 1600-2000 nucleotides
□ Need helper virus (need capsid, but don't have one)
□ Hepatitis D ("The Delta Agent") is a virusoid, helped by Hepatitus B.
□ A parasite of a parasite.
§ Prions
□ Cause slow neurological conditions (transmissible spongiform encephalopathies)
□ Eg: Kuru, scrapie, BSE, CJD
□ Misfolded brain protein.
□ Proteins form fibril plaques which are cytotoxic.
□ Cell death leaves holes in tissue.
§ Article - bacterium living in insect cells, has lost ability for independent reproduction.
○ Assessment of bacterial growth - Bacterial Enumeration.
○ Usually consider populations rather than individual cells
○ Number of cells, or biomass (weight of cells)
○ Bacterial numbers:
§ Total count (haemocytometer or coulter counter): every cell.
□ Haemocytometer - blood cell counter - thicker slide, with well. Graticule under well, to assist counting.
□ Coulter counter - automated particle counter.
□ Usually expressed, eg as cells per cm3.
□ Direct count
§ Viable count - only live cells. (eg colony count or most probable number)
□ Dilute to allow spread, then innoculate petri dish. Incubate, then count colonies. - known volume of known dilution. Ideal count is 30 - 300 colonies for best accuracy; more are hard to count, less may not be representative.
□ Viable count = colony count x 1/dilution x 1/volume (cm3)
□ Volume is usually 0.1 cm3 or 0.2 cm3 (100 or 200μl)
□ Incubation in nutrient broth in series of tubes - count proportion of tubes that have growth.
□ Usually expressed as colony forming units per cm3.
□ Indirect count
○ Biomass
§ Wet and dry weight measure
1) Weigh centrifuge tube
2) Add culture
3) Weigh
4) Centrifuge
5) Remove supernatant
6) Weigh pelleted cells (wet weight)
7) Dry pellet in oven/incubator at >100°C
8) Weigh dried cells (dry weight)

§ Spectrophotometric measurement
□ Measure turbidity of culture

Week 2

Today's Topics:
○ Origin of life on Earth & microbial evolution

Lecture Topic:
During the lecture, take notes here.
○ Life cannot exist without liquid water
○ The atmosphere was anoxic when life first evolved, so the first life must have been anaerobic.
○ Temperatures were higher, so early life must have been thermophilic, living in environments such as hydrothermal systems.
○ Photosynthetic bacteria were probably not the first organisms, but are thought to have evolved early. First Pb were probably not oxygen producing.
○ Archea are not photosynthetic.
○ PS thought to have evolved in purple-green bacteria.
○ Cyanobacteria developed oxygenic photosynthesis, about 2.5-3.0 billion years ago. By 2.5 to 1.5 billion years ago free oxygen was present in atmosphere, as a result.
○ This had a profound effect - aerobic respiration provides massively more energy than anaerobic, used by almost all life today.
○ Also lead to development of ozone layer, reducing UV at surface and its mutagenic effects.
○ This permitted evolution of more complex forms of life.
○ Anaerobic prokarya: 4.0 billion years ago.
○ Anoxygenic photosynthetic bacteria: 3.0-3.5 billion years ago.
○ Eukarya appeared about 2 billion years ago
○ Microbial Nutrition
○ Energy: Light, inorganic, or organic compounds?
○ Carbon: CO2 or organic compounds
○ Photoautotrophic: obtain energy from light, and carbon from CO2.
○ Photoheterotrophic: obtain energy from light, and carbon from organic compounds. (Cannot fix from CO2.)
○ Chemoautotrophic: Energy from oxygenation of inorganic compounds, egg ammonia or H2S, and carbon from CO2.
○ Chemoheterotrophic: obtain both energy and carbon from organic compounds.
○ Possible early metabolic types
○ Oparin argues that initial metabolic type was simple heterotrophic bacterium. - only needs a few enzymes.
○ Autotrophic organisms are more complex, as they need multiple pathways, and so would have evolved later.
○ Others argue that early life forms may have been hydrogen bacteria, obtaining energy from oxidation of hydrogen. Both bacteria and archea types are known, using CO2 for carbon.
○ Many of these are anaerobic, and so could have fit in with early conditions.
○ Temperature and growth
○ Cardinal temperatures:
§ Optimum: fastest replication: range 0-75C
§ Minimum: no growth below: enzymes inactive, membranes solidify; al low as -20C
§ Maximum: no growth above: cell may die - enzymes denatured, membranes break down; as high as 120C.
§ Pressure can permit fluid water at unusual temps.
○ Upper limit for protozoa: 50C
○ Upper limit for algae & fungi: 55-60C
○ Classification by temp:
§ Psychrophile: Optimum temperature <15C
§ Psychrotroph (facultative psychrophiles): 20-30C eg: fungi, etc.
§ Mesophiles: 20-45C eg most human pathogens
§ Thermophiles: 55-65C
§ Hyperthermophiles: 80-113C
○ Oxygen and growth
○ Obligate aerobes - completely dependent on oxygen - includes almost all multicellular organisms
○ Facultative aerobes - do not need oxygen, but grow better with it.
○ Aerotolerant anaerobes - grow equally well with or without oxygen
○ Obligate anaerobe - killed by oxygen.
○ Microaerophiles - damaged by normal concentration of oxygen (20%)
○ pH and growth
○ Habitats range fro 0.2-2.0 at the acid end to 9-10 at the alkaline.
○ Acidophiles optimum 0-5.5
○ Neutrophiles optimum 5.5-8.0
○ Alkaliphiles optimum 8.0-11.5
○ Eg fungi prefer 4-6; malt extract agar is used to culture.
○ Most bacteria and protozoa are neutrophiles.
○ Osmotic concentration
○ Nonhalophile:adversely affected by high concentration.
○ Halotolerant: Able to grow over a range of concentrations.
○ Moderate Halophile: prefers high levels of NaCl - above 0.2M to grow. (eg Staphyllococcus)
○ Extreme Halophile: requires high levels - 2M to saturation (about 6.2M)
○ Microorganisms and Ecosystems
○ Primary producers: eg Algae - make organic matter from CO2 and water.
○ Primary consumers: eg Protozoa - feed on bacteria and fungi.
○ Decomposers: eg Fungi - break down organic matter into inorganic compounds.
○ Prefered conditions:
○ Protozoa prefer wet, fungi prefer dry.
○ Some prefer rhizosphere (area around roots) - eg nitrogen fixing bacteria (no eukarya can fix nitrogen)
○ Symbiosis: eg e. coli in gut -make vitamins that we cannot.
○ Impact of Bacteria and Archea on biogeochemical cycles
○ Carry out significant reactions that are crucial to operation of biosphere
○ Nitrogen, iron, sulphur, carbon cycles.
○ Carry out steps that no eukarya can.
○ Carbon cycle: carbon fixation (photosynthesis), methanogenesis, CO oxidation.
○ Nitrogen cycle: nitrogen fixation, nitrification, nitrate reduction, mineralisation.
○ Microbial Interactions
○ Mutualism: eg aphid & buchnera aphidicola - cannot reproduce independantly - 60-80 bacterisides, containing buchnera, which produce nitrogen compounds, vitamins, and amino acids not provided by diet.
○ Tube worm - trophosomes contain microorganisms that oxidise H2S and fix CO2.
○ Rumen ecosystem - read up for coursework.
○ Commensalism:eg e.coli in the gut - one gets advantage, other is neutral (some list as mutualistic - we get vitamin b from e coli)
○ Parasitism one is at expense of other. (pathogens)
○ No archea are parasitic
○ Predation: one eats another - bdellovibrio, vampirococcus, daptobacter.

Week 1

Today's Topics:
o Introduction and Historical Perspectives

Read "system of work for use of microbiology laboratories", page 19

Lecture Topic:
o Module details on WebCT (or will be)
o Attendance Requirement on practicals.
o Record of Results Booklet must be signed by a member of the teaching team at the end of each practical session.
o Coursework 1: Microbiological Problems - due 10th November. (20%)
o Coursework 2: Record of Results booklet - due 8th December (20%)
o Lab coats are now supplied.
o Impact:
• Nitrogen fixing, etc
• Food preservation, fermentation
• Disease
• Biofuels
• Bioremediation
• Microbial Mining
• Biotech - gmfoods, pharmaceuticals, gene therapy
o 20 times more bacteria than human cells in body
o Effects on civilisation
• Spread of disease in and by armies
• Urbanisation - cramped, unhygienic conditions
• World trade and travel - rapid spread of disease
• Bubonic plague - killed 25M of 100M in europe & north africa in 1 year.
• Last century ha seen massive fall in disease as killer
• 50% of children died from infectious disease by age 10 befoe C20.
o Jenner
o Pasteur
o Robert Koch - developed techniques still used today - worked with anthrax.
• Pure culture methods
• Platinum loop
• Agar (Fannie Hesse - husband worked in lab - tried for jams but was too hard; suggested he tried for cultures)
• Petri Dish (RJ Petri)
• Team confirmed 15 significant diseases caused by microbes.

o Lister - antiseptics (Koch used perchloride of mercury; this is now deprecated as it is a pollutant)
o Anton De Bary - extended to plants. (potato blight - ref famine)
o Iwanowski - identified potato mosaic virus by filtration (too small to see under optical microscope)
o Rous
o Fleming
o Evolution (ref foundation biology)
o 50's Stanley Tyler & Elso Arghoorn - fossil stromatolites.
o Prokaryotes vs Eukaryotes
o Escherichia Coli can reproduce in 20 min, in ideal conditions
o Archea - have ether linked membrane lipids - discovered by Carl Woese, in 1977.