Topic 6 Flashcards
Microorganisms
- Decompose organic matter
- respire -> releasing CH4 + CO2 (carbon cycle)
- in and on animals/plants, which die and the microorganisms then secrete enzymes to break them down
Steps to Estimating time of death
- Body temperature
- Degree of muscle contraction
- Forensic entomology
- Extent of decomposition
- Stage of succession
Time of Death: Body temperature
Heat produce form metabolic reactions in mammals
- humans = 37 C
TOD: metabolic relations slow down -> stop
- body temp fall -> equals surroundings = Algor Mortis
- human body cools at 1.5 C to 2.0 C per hour
-> cooling rate affected by: air temp, body weight, clothing, position
Algor Mortis
The process in which the body temperature falls, after time of death, to equal that of its surroundings (at an estimated rate of 1.5 to 2.0 C per hour)
Time of Death: Degree of Muscle Contraction
4-6 hrs after death -> contract & become stiff = Rigor Mortis
- muscle cells deprived of oxygen
- respiration still occurs -> anaerobic -> lactic acids builds up in muscle
- pH of cells decreases -> inhibits ATP producing enzymes
- No ATP -> myosin + actin bonds in muscle cells become fixed -> stiffen
- – smaller muscles in head contract first
- – large muscles in lower body = last
- 12-18hrs after TOD = every muscle contracted
- 24-36 hrs after TOD = wears off
Rigor Mortis
The process in which the body’s muscles begin to contract & become stiff 4-6 hrs after TOD.
- affected by:
- -degree of muscle development (less = faster)
- temperature (higher = faster)
- 12-18hrs after TOD = every muscle contracted
- 24-36 hrs after TOD = wears off
Time of Death: Forensic Entomology
Study of the insect colony present at TOD
- estimated by identifying type of insect present
- flies = few hrs after death
- beetles = later
- Stage of life cycle:
- blowfly larvae: hatch from egg 24 hrs after laid
Conditions that can affect life cycle:
- drugs
- humidity
- oxygen
- temperature
Blowfly larvae lifecyle
ngng
Time of Death: Extent of Decomposition
Bacteria + enzymes start to decompose
- hrs to a few days: cell + tissue breakdown
- skin = greenish hue (Putrefaction) - few days to a week: microorganisms decompose tissues
- gases produced (methane) -> body bloats
- skin blisters & falls off - few weeks: tissues begin to liquify
- seep into area around body - few months -> years: skeleton
- decades -> centuries: skeleton begins to disintegrate
note: affected by temperature + O2 availability
- e.g acidic peat bogs = preserve
Time of Death: Stages of Succession
Above ground, at TOD:
- favourable for bacteria -> decompose tissues
- flies & larvae -> fly larvae feed
- beetles suited
- dead body dries out -> flies leave, beetles remain to decompose
- no tissue -> (mostly) no organisms
note: for plants -> insects remain from stage to stage
location: sealed away = no insects
under water = fish / rodents etc
DNA profile
Unique genetic fingerprint (except for monozygotic twins)
DNA profile (PCR process)
- DNA sample taken (blood, saliva, epithelial cells)
- Acquisition: DNA separated from sample
- Replication: Polymerase Chain Reaction
- Separation: Gel Electrophoresis
Fingerprints
Types:
- arch
- tented arch
- whorl
- loop
Sweat + oil secretions placed into iron flakes then computerised
Dental Records
Teeth + filings decay slowly -> more resistant to burning
Surgical Implants
Recorded number on item that can be linked to surgery
DNA profile: Aquisition
DNA broken down in buffer solution
- small suspended particles separated via filitration/centrifuging
- Protease enzymes incubated with suspension -> removes proteins
- Cold ethanol added -> precipitate out DNA
- continuous washing of DNA with buffer
DNA profile: Restriction Endonucleosis
Enzyme = EcoRI
- cuts DNA out at specefic base sequences (4-6 bases long)
- cuts between G & A bases when ‘GAATC’ sequence occurs
- if the restriction site is either side of short tandem repeats, DNA fragment remains intact, but cut from rest of genome
DNA profile: Polymerase Chain Reaction
Uses DNA primers, which are marked with fluorescent tags
- DNA placed in reaction tube with DNA polymerase, primers & nucleotides
- once in PCR machine, undergoes temperature change cycle:
1. 95 C = separates double stranded DNA (breaks H bonds)
2. 55 C = optimises annealing of primer to target DNA sequence (start of STR), then polymerase attaches & replication occurs
3. 70 C = optimum for heat stable DNA polymerase - > lines up free DNA nucleotides along each template strand (complementary strand formed)
- > two new copies of DNA fragment
Cycle starts again, using all copies of DNA fragment (each cycle doubles DNA)
DNA profile: Gel Electrophoresis
Separates fragments according to length
- DNA placed on agrose/polyacrylamide gel (in wells)
- provides a stable medium through which the fragments move - Gel is submerged in a buffer solution
- Connected to electrodes which provide a potential difference across the gel
- -ve fragments migrate through the gel according to overall charge + size (smaller + less repeats = faster)
- smaller fragments end up closer to +ve electrode - reference sample with fragments of known length may be added to the gel = DNA ladder/marker
- fragment lengths are measured in base pairs
DNA profile: Southern Blotting
Gel placed directly onto nylon/nitrocellulose membrane, with wad of fry absorbent paper placed on top
- > acts as a wick to draw buffer solution up through the gel (carrying DNA fragments) onto the membrane
- fragments maintain position relative to each other
- denatured into single strands -> exposing base sequence
Buffer solution (DNA profiling)
Detergent + Salt that disrupts cell membrane
Introns
Non-coding regions in DNA
Exons
Coding regions in DNA
Short Tandem Repeats (STR)
Sequences of repeated bases
- 2 to 50 pairs, 5 to several hundred times
- same locus on homologous chromosome pairs
- STR quantities differ between individuals, this difference = identifier
DNA profile: Probe attachment + viewing (fluorescent or radioacitve
Nylon/nitrocellulose membrane incubated with excess of a labelled DNA probe
- allowing time for probe to bind to any complementary sequences (hybridising)
- > unbound probe washed away
If probe = radioactive (or labelled with radioactive phosphorous(
- membrane -> dried
- placed next to x-ray film -> blackens wherever probe has bound with DNA
If probe = fluorescent
- visualised on membrane under UV light
Radioactive Phosphorus
32^P
Labelled DNA Probe
Short section of DNA with a base sequence complementary to the target DNA sequence that needs to be located
DNA profile (Restiction endonucleosis process)
- DNA sample taken (blood, saliva, epithelial cells)
- Restriction Endonucleosis
- Gel Electrophoresis
- Southern Blotting
- Probing + visualisation
DNA profile: PCR analysis using Gel Electrophoresis
As DNA primers have fluorescent tags, the normal gel electrophoresis occurs, but then the system can be automated
- as DNA fragments move through gel, they pass a laser
- the dye in the tag fluoresces -> light detected
- gives the time taken for fragment to pass through gel
- > calibrated against known fragment lengths (determined by no. of base pairs)
Note: several STR loci can be analysed at once using tags that fluoresce at different wavelengths
Bacteria Structure (with diagram)
Single Celled prokaryotes
- few micrometers
(insert diagram)
Virus Structure (with diagram)
- Nucleic acids surrounded by proteins
- tiny (decimal of micrometers)
- Nucleic acid core (DNA or RNA)
- some carry proteins inside their capsid
- some have an envelope
- Attachment proteins
(insert diagram)
Virus’ Envelope
A stolen cell membrane from a previous host cell
Virus’ Capsid
A protein coat around the virus’s core
Virus’ Attachment proteins
Proteins that stick out from the capsid or envelope, and allow the virus to cling to a suitable host cell
Pathogens
Any organism that causes infectious diseases
e.g. some bacteria, some fungi, all viruses
HIV
Human Immunodeficiency Virus
- infects + destroys T helper cells
- uses them as host cells
- T helper cells usually activate other immune system cells
AIDS
Acquired Immune Deficiency Syndrome
HIV infection pathway (applicable to other viruses)
- Spread via infected bodily fluids
- contact MUCOSAL surfaces (genital tissue), damaged tissue, or is injected into bloodstream - (all viruses) reproduces inside cells of infected organism e.g. T-Helper cells (needs enzymes + ribosomes to replicate)
- attachment proteins attach to receptor molecule (on cell membrane)
- Capsid is inserted into cell, uncoats & releases RNA into cytoplasm
- reverse transcriptase used to make complementary strand of DNA from viral RNA template
- > double -stranded DNA -> inserted into human DNA
- host-cell enzymes make viral proteins (using viral DNA)
- viral proteins -> new viruses
- > bud from cell -> infect others
HIV initial infection & latency period
II: HIV replicates rapidly -> severe flu like symptoms
LP: HIV replication drops -> no symptoms; can last for several years
AIDS development
HIV infection eventually leads to AIDS = immune system deteriorates -> eventually fails (takes roughly 10 years to develop)
Classified as having AIDS when:
- Symptoms of failing Immune start to appear
- T helper cell count drops below certain level
- > leads to opportunistic infections
Symptom sequence:
- Minor infections of mucus membranes & recurring respirator infections (lower T helper count)
- AIDS progresses -> T helper count drops further
- > more serious infections:
- chronic diarrohea
- severe bacterial infections
- tuberculosis - Late stages = v.low T helper count
- > serious infections
- toxoplasmosis of the brain (parasite)
- candidiasis of the respiratory system (fungal)
- > causes death (secondary infection)
Tuberculosis infection pathway
Mycobacterium Tuberculosis causes TB
- droplets (tiny) inhaled into lungs
- bacteria taken up by phagocyte (wbc) in lungs (inhaled)
- bacteria replicate in phagocyte
- infected phagocytes -> sealed in tubercles (structures in lungs)
- bacteria -> dormant (no obvious symptoms)
- dormant bacteria -> reactivate -> overcome immune system -> TB
- more likely in weakened immune systems (e.g. AIDS)
- length of time between infection -> TB varies (weeks to years)
TB sequence of Symptoms
- fever, general weakness, severe coughing (lung inflammation) = initial
- progresses -> damages lungs -> (untreated) respiratory failure -> death
- can spread e.g. brains + kidneys
- untreated -> organ failure -> death
Pathogen routes (of infection)
- cuts in skin
- digestive system (contaminated food + drink)
- respiratory system (droplet inhalation)
- mucosal surfaces (nose, mouth, genitals)
Barriers to prevent infection
- Stomach acid = acidic conditions -> kill
- some pass into intestines -> invade gut wall cells - Skin = physical barrier
- blood clots extend protection
- margin to enter - Gut + skin flora = billions of harmless microorganisms
- compete with pathogens for nutrients + space - Lysozymes = enzyme (kills bacteria by damaging cell wall)
- mucosal surfaces -> secretions = contain lysozyme
Foreign antigens ->
Trigger immune response
1. non specific
or
2. specific
Antigen
molecules found on cell surface
Non-specific immune response (stages)
Happens in the same way for all microorganisms , and occurs straight away:
- Inflammation at infection site
- Production of Interferons
- Phogocytosis + Lysozyme action
Inflammation at infection site: Non-specific immune response
- immune system cells recognise foreign antigens
- > release histomine molecules (trigger inflammation) - Triggers vasodilation; increasing blood flow around infection site
- Also increases permeability of blood vessels
- immune system cells flood to site
- due to increased permeability -> move into infected tissue - start to destroy pathogen
Production of interferons: Non-specific immune response
- cells infected by virus -> produce interferons
- > prevent virus spreading to other cells
- prevent viral replication
- inhibit production of viral proteins - Activate cells involved in specific immune response
- kills infected cells - Activate other Non-specific immune responses
- e.g. promote inflammation
Interferons
Anti-viral proteins
Phagocytosis + Lysozyme action: Non-specific immune response
- Phagocyte recognises pathogens antigens
- Cytoplasm moves around pathogen -> engulfs it
- Pathogen contained in phagocytic vacuole
- Lysosyme fuses with phagocytic vacuole
- lysozyme breaks pathogen down - Phagocyte presents pathogens antigens
- sticks antigens on surface to activate other immune system cells
- antigen presenting cell
Specific Immune response (brief)
Antigen Specific (occurs after non-specific if necessary)
- Phagocytes activate T Cells
- T helper cells activate B cells
Phagocytes activate T Cells: Specific Immune response
T cell = white blood cell
- surface covered in receptors
- receptors bind to antigens
- > displayed by antigen presenting cells (macrophages; type of phagocyte)
- each T cell = different shaped surface receptors
- receptor + complimentary antigen -> bind
- > activate T Cell -> divides to clone
Different T cells = different functions:
- T helper = substance released to activate
- B Cells
- T killer cells
- macrophages - T-Killer = attach to antigen of infected cell -> kill cell
- T-memory
T Helper cells activate B Cells: Specific Immune response
B cells = white blood cells
- covered by proteins = antibodies
- each B cell = different shaped antibodies
- antibodies + antigens = antigen-antibody complex
- antibody + complimentry antigen -> bind
- binding + substances form T helper cells -> activates B cells
B cell activated -> divides (mitosis) to form:
- plasma cells = B effector cells
- B memory cells
Plasma Cells
= B cell clones
-> secrete antibodies
Antibodies (structure)
- 4 polypeptide chains
- 2 heavy
- 2 light
- each chain has a variable + constant region - variable regions = antigen binding site (complementary shape)
- hinge region = provides flexibility (accommodates binding)
- constant region = allows for binding to receptors on immune system cells
- same on all antibodies - Disulphide bridges = hold chains together
Antibodies (Structural diagram)
Insert Image
How antibodies clear infection
- Agglutinate pathogens
- antibodies = 2 binding sites -> 2 pathogens at a time (clump together)
- phagocytes bind to antibodies -> phagocytoses clump - Neutralising toxins
- bind to toxins, inhibiting effect (neutralised)
- entire complex phagocytosed - Prevent pathogen binding to human cells
- antibody binds to antigen
- block cell surface receptors needed to bind to host cell
- pathogen cannot attach -> prevents infection
Types of antibodies
- Membrane bound
2. Secreted
Membrane bound antibodies
Attached to B cell membrane
- extra protein section on the heavy chains to anchor to membrane
- > copied into mRNA; protein synthesised, but modified before translation to provide a different heavy chain for secreted vs membrane bound antibodies (more than one protein from same gene)
Secreted antibodies
Free floating (come from plasma)
- heavy chain coded for by a single gene
- > copied into mRNA; protein synthesised, but modified before translation to provide a different heavy chain for secreted vs membrane bound antibodies (more than one protein from same gene)
mRNA modified before translation because…
- genes contain sections that do not code for amino acids (introns)
- Transcription = exons + introns copied into mRNA
= pre-mRNA - Introns removed = splicing
- exons joined -> mRNA strands
- occurs in Nucleus
= post-transcriptial change - Sometimes exons are also removed
- different mRNA strands = alternative splicing (possible for around 95% of genes)
- allows for more more than one sequence -.> multiple proteins possible from 1 gee=ne
Memory Cells
- Pathogen enters body 1st time -> antigens activate immune system
- NSI -> SI response = primary response; slow as few B cells for antibodies
- enough antibodies produced -> infection overcome (symptoms) - After exposure, T + B cells produce memory cells
- remain in body
- T memory remember specific antigen
- B memory record antibody recquired
= immune (able to respond quickly) - 2nd pathogen exposure
- > quicker + stronger response = secondary response
- T memory cells divide into correct T cell type to kill antigen carrying cell
- B memory cells divide into B effector cells to produce antibodies (few/no symptoms)
Types of Immunity
Active:
- Natural = Catch a disease
- Artificial = Vaccine
Passive:
- Natural = baby receives antibodies from mother (placenta + milk)
- Artificial = injected with antibodies (e.g. tetanus shot)
Active Immunity
Organisms own immune system makes antibodies after being stimulated by an antigen
Passive Immunity
Antibodies made by a different organism are provided, allowing for short-term, immediate immunity
Primary/Secondary immune response diagram
Insert diagram
Vaccines
Contain antigens -> stimulate primary response
- no disease
- leads to immunity
Protect (usually) against different strains
- caused by antigenic variation
HIV’s Evasion mechanism
- Kill infected immune system cell
- reduced cell no. -> reduces detection - HIV = high mutation rate in genes coding for antigens
- change antigen structure = Antigenic Variation
- > primary response for each new strain - Disrupts antigen presentation in infected cells
- prevents recognising + killing
Mycobacterium Tuberculosis evasion mechanism
- Engulfed by phagocyte -> produces substance
- prevents lysosyme fusing with phagocytic vacuole
- bacteria not broken down
- continues multiplying undetected - Disrupts antigen presentation
- stops recognising + killing
Types of Antibiotics
- Bacteriocidal
2. Bacteriostatic
Bacteriocidal Antibiotics
Inhibits enzymes required tom make chemical bonds in cell walls
- prevents growth -> cell death
- weakens cell wall + pressure (water via osmosis_
- > bursts
Bacteriostatic Antibiotics
Inhibit protein production by binding to bacterial ribosomes -> no enzymes
- stops metabolic processes (e.g. growth)
Antibiotics
Chemicals that affect metabolic processes (such as growth) of bacteria, and can cause their destruction
HATs
Hospital Acquired Infections
- Patients being treated in hospital more likely to receive them due to traffic, other patients & weakened immune systems
Transmitted Via poor hygiene
- non-contained coughs/sneezes
- surfaces not regularly disinfected
- no hand washing
HATs treatment/containing
People with HATs moved to isolated ward (reduces likelihood of transmission)
Some HATs = antibiotic resistant, so to prevent + control spread:
- no antibiotics for minor or viral infections
- no preventative antibiotics
- yes to narrow spectrum antibiotics
- yes to rotating use of different antibiotics
- courses must be completed (otherwise increases likelihood of resistance)
