Bacteria, Fungi Viruses lecture Flashcards
What does dipstick urinalysis look for (some examples)
Cells
RBCs
WBCs (leucocytes)
Bacteria, Fungi, Yeast, Parasites (Culture on agar +blood)
Glucose, Ketones, nitrates, crystals,
When culturing micro samples what medium should be used
Culture on agar: blood/chocolate/CLED)
How long does it take to find out specific pathogen by micro culture
24-48hrs
24 any info
48 specific info (antibiotic resistance/culture specificity)
3 types of AGAR +/- nutrient plates
blood (BAP, blood agar plate)
enriched, bacterial growth medium. Fastidious organisms, such as streptococci, do not grow well on ordinary growth media but grow on blood agar. … Blood agar consists of a base containing a protein source (e.g. Tryptones), soybean protein digest, sodium chloride (NaCl), agar, and 5% sheep blood
chocolate (CHOC/CBA chocolate agar plate)
variant of the blood agar plate, containing red blood cells that have been lysed by slowly heating to 80°C. Chocolate agar is used for growing fastidious respiratory bacteria
CLED (cystine–lactose–electrolyte-deficient agar)
It supports the growth of urinary pathogens and contaminants but prevents undue swarming of Proteus species due to its lack of electrolytes.
Things that can be added to agar (5 examples)
Blood Potato starch Sugars / Salts Antibiotics pH indicators
What does CLED plate show
for urinary cultures
CLED agar produces a yellow colour if the bacteria ferments lactose to produce acid
- typical of E. coli and not some other urinary bacteria
What is measured on agar plate to show antibiotic resistance
antibiotic susceptibility via zones of inhibition
resistant vs sensitive
What are the benefits of diagnostic microbiology
Corrects your guesses
Allows you to focus your treatment
Informs the epidemiology
Laboratory Techniques used in diagnostic microbiology
Microscopy
Culture inc on Nutrient and selective agar plates
and for difficult-to-culture organisms;
Serological Techniques
Molecular Techniques (PCR)
Tissue culture
Staining
When might gram staining/microscopy not be useful?
Can be useful from ‘sterile sites’
Cerebrospinal fluid (CSF), joint fluid
In many cases NOT useful from ‘non-sterile’ sites
e.g. Sputum from a patient with a ? chest infection
What are the 5 basic shapes of bacteria
spherical (cocci), rod(bacilli), spiral(spirilla), comma (vibrios) or corkscrew (spirochaetes).
what are the 6 shapes of spherical (cocci) bacteria
Micrococci
Diplococci
Streptococci
Staphylococci
Tetracocci
Sarcine
They can exist as single cells, in pairs, chains or clusters.
what are the 3 shapes of rod shaped (bacillus) bacteria
Bacillus
Steptobacillus
Coccobacillus
What is the best way to make the diagnosis of Pneumocystis jiroveci (carinii)pneumonia
to perform a Gomori methenaminesilver(GMS)stainon the lung tissue or bronchoalveolar lavage (BAL) fluid.
What is BAL fluid
BAL=bronchoalveolar lavage (saline flush of lungs during bronchoscopy)
What is GMS stain?
Grocott-Gomori methenamine silver (GMS) is a special stain to detect fungi - HIVE +ve/AIDS (susceptible to unusual pathogens)
What is a Zeihl Neelson Stain
Ziehl-Neelsen stainingis a type of acid-fast stain, mainly Mycobacteria
What is Auramine phenol stain
Auramine phenol stain can identify tuberculosis mycobacteria
Parasites - what is an example from lecture that can be visualised directly
Pinwormsare tiny, narrow worms. They’re white in colour and less than a half-inch long.
How can you visualise viruses by microscopy?
can be seen by Electron microscopy
What type of organism can be directly visualised
Parasites and ova (parasitic eggs) stool samples mainly
Pros and cons of bacterial culture
Slow but provides a lot of information
Can quantify bacteria
Purification
Provisional identification
Antibiotic susceptibility testing by disc testing
Bacterial identification
Traditionally basis of bacterial metabolism
What’s an API test
when is it used
API (analytical profile index) strips give accurate identifications based on extensive databases and are standardized, easy-to-use test systems.
use to sub classify gram -ve bacteria
What technique is MALDI-TOF
Mass spectrometry is an analytical technique in which samples are ionized into charged molecules and ratio of their mass-to-charge can be measured. In MALDI-TOF mass spectrometry, the ion source is matrix-assisted laser desorption/ionization (MALDI), and the mass analyzer is time-of-flight (TOF) analyzer.
Culture-based diagnostics: summary
Good for detecting bacteria you can stain and grow
Reliable identification and resistance testing
Very slow and labour intensive
what is Serology (serum-ology)
Serology is the scientific study of serum and other body fluids. In practice, the term usually refers to the diagnostic identification of antibodies in the serum. Such antibodies are typically formed in response to an infection, against other foreign proteins, or to one’s own proteins.
IgM Found mainly in blood and lymph fluid, this is the first antibody the body makes when it fights a new infection.
IgG This is the most common antibody. It’s in blood and other body fluids, and protects against bacterial and viral infections. IgG can take time to form after an infection or immunisation.
what does IgM indicated in terms of infection
acute infection innate?
what does IgG indicate in terms of infection
chronic adaptive response
What viruses to consider in glandular fever
Epstein Barr Virus (EBV)
Cytomegalovirus (CMV)
Symptoms of glandular fever
Fever
Sweats
Sore throat
Swollen lymph glands in her neck
If person has specific IgG are they immune or susceptible to the pathogen?
IgG+ve = immunity (previous infection)
If person has specific IgM to pathogen what does that indicate?
Active/acute infection
PCR three steps
denature (92-95)
anneal primers (50-70 primer dependant)
extension of dsDNA (72 taq pol)
approx 30-35 cycles
What is latency in infection
ability of infection to remain dormant for long periods of time before reactivation
e.g.
Chicken pox reactivating as Herpes zoster (shingles)
Cold sores (HSV)
CMV & EBV (subclinical unless immune suppressed)
Tuberculosis
Mechanisms and sites of latency poorly understood
Genetic material present but not causing disease
When is Whole genome sequencing used on bacteria
Sequencing entire DNA of bacteria
Compare strains to see how related they are/evolve
Interrogate for antibiotic resistance genes
Set to revolutionise diagnosis of infection
Rapid (hours)
Cheap (comparable to culture)
Desktop Platforms
When is PCR used?
Viral encephalitis Viral infection of brain tissue Fever, headache, confusion, fits Many causes Enteroviruses most common Herpes viruses (HSV, VZV, CMV, EBV) PCR commonly used to diagnose viral encephalitis Good negative predictive value Poor positive predictive value so backed up by brain imaging, cells in spinal fluid
What is HIV mainly diagnosed by
HIV mainly diagnosed by serology
PCR diagnosis in early HIV infection
To diagnose resistance
Monitoring response to treatment (’viral load’)
what is tissue culture used for in diagnostic micro
Intracellular organisms
Viruses
Mycoplasma
Chlamydia
how is tissue culture used for in diagnostic micro
Two main ways:
Cytopathic effects on cells
Expression of viral proteins detected at the cell surface
What is rubella
Rubella(German measles) is a rare illness that causes a spotty rash. Serious in pregnant women.
What is CPE
what lab test identifies this
Cytopathic effect(CPE), structuralchangesin a host cell resulting from viral infection. CPE occurs when the infecting virus causes lysis (dissolution) of the host cell or when the cell dies without lysis because of its inability to reproduce.
Tissue culture
Immunofluoresence in micro
fluorescent ab/ag labelling of e.g.viral surface proteins
