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).

Tuesday, November 14, 2006

Week 4

Today's Topics:
○ Viri

Announcements:
○ 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

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