Introduction to Diagnostic Microbiology Flashcards
How differentiate Staphyloccocus aureus from other Staphylococcus species?
- -* coagulase positive, other species are coagulase negative
- beta-hemolytic, other species are non-hemolytic
What does catalase do?
- prevents bacteria oxidative damage by reactive oxygen species, by catalysing H2O2 to water and oxygen
- all Staphyloccus species are catalase-positive
What does coagulase do?
- leads to fibrin formation around bacteria, protecting it from phagocytosis
- found within Staphylococcus aureus
Where is Staphylococci found?
- ubiquitous (found everywhere)
- Coagulase-negative staphylocci are part of skin normal flora
: ~30% of normal healthy adults are persistant nasopharyngeal carriers of S. aureus
How is Staphylococci spread?
- person-to-person spread through direct contact or exposure to contaminated fomites (e.g. bed linens, clothing, etc.)
- among the most common causes of community and/or hospital acquired infections
(some strains are resistant to different antibiotics)
How do you differentiate Staphylococcus epidermidis from Staphylococcus aureus?
- S. epidermidis is coagulase-negative
- considered less virulent than S. aureus
Where is S. epidermidis usually associated with and what is its pathogenesis associated with?
- most commonly found associated with prosthetic devices
- pathogenesis associated with slime layer and biofilm
What causes hemolytic patterns?
- bacteria secrete proteins / metabolites that lyse red blood cells
What are the different hemolytic patterns and their characteristics?
alpha: incomplete lysis; green
beta: complete lysis; clear
gamma: no lysis
Mannitol-salt agar
- contains mannitol sugar and pH indicator (phenol red)
- if an organism can ferment mannitol, an acidic byproduct is formed that will cause the pheno red in the agar to turn yellow
Which is (are) the catalse-negative species?
a) enterococci
b) staphylococci
c) streptococci
d) micrococci
- a) enterococci
- c) streptococci
Description of Streptococcus
- gram-positive cocci in chains
- catalase-negative
What classifications can be used for Streptococcus?
1) Lancefield grouping
: based on cell wall carbohydrate antigens
- Group A-W (Strep test RAPID)
2) Hemolytic pattern
How can you differentiate the following in a lab:
- S. pyogenes*
- S. pneumoniae*
- S. agalactiae*
Viridans streptococcus
1) hemolytic pattern
- S. pneumoniae & Viridans strep are alpha-hemolytic
- S. pyogenes & S. agalactiae are beta-hemolytic
2) Optochin test & capsule presence
- S. pneumoniae is optochin sensitive & has a capsule
- Viridans strep is optochin resistant & does not have a capsule
3) Bacitracin test
- S. pyogenes is bacitracin sensitive
- S. agalactiae is bacitracin resistant
S. agalactiae
: Group?
: virulence factors?
: epidemiology?
: transmission?
- Group B streptococcus
- virulence factors: polysaccharide capsule
- epidemiology: colonise lower GIT & UGT
: commonly associated with infections of newborns (Meningitis, pneumonia)
- transmission: increased risk if mother is vaginally colonised
Viridins streptococci
- heterogenous collection of alpha-hemolytic and non-hemolytic streptococci
- colonises the oropharynx, GIT and UGT
- avirulent
: can cause dental carries (S. mutans)
: common cause of subacute endocarditis
Differentiation of S. pneumoniae and Viridans streps
optochin test
S. pneumoniae: sensitive
Viridans streps**: resistant
bile solubility test
S. pneumoniae: sensitive
Viridans streps**: resistant
Biology of Bacillus anthracis
- gram-positive rods; single or paired
- aerobic
- spore-forming
Virulence factors of B. anthracis
- polypeptide capsule
: consists of poly-D-glutamic acid
- toxins
: Protecting Antigen (PA)
: Lethal factor (LF)
: Edema factor (EF)
Epidemiology & Transmission of B. anthracis
- reservoir
- transmission
reservoir
- animals, skins, soil
- human infections via exposure to reservoirs
transmission
- innoculation (most common form of transmission)
- ingestion
- inhalation (i.e. Wool sorter’s disease)
potential for bioterrorism- anthrax
Diseases caused by B. anthracis
: cutaneous anthrax
- painless swelling
- progresses to ulcer surrounded by vesicles
- necrotic eschar, central, with erythematous border often with painful regional lymphadenopathy
Diseases caused by B. anthracis
: gastrointestinal anthrax
- rare
- symptoms are based on site of infection
- swelling
- vomiting, bloody diarrhea
Diseases caused by B. anthracis
: pulmonary anthrax
- life-threatening
- leads to shock if without treatment
- macrophages ingest bacteria and transport to lymph nodes
- fever, malaise, swelling, dyspnea
Diagnosis of B. anthracis
- gram-stain: gram-positive rods
- culture: non-hemolytic
- serology
- PCR
- public health reference lab (CDC select agent)
Treatment for B. anthracis
- Ciprofloxacin
- doxycycline
- vaccine: toxoid, AVA, high risk only
Bacillus cereus epidemiology & transmission
- ubiquitous (found in nearly all environments)
- transmission: foodbourne
: associated with fried rice
: food kept warm (not hot)
: buffets
Pathogenesis & Disease of B. cereus
eye infections
gastroenteritis (most severe form)
- heat-stable form
- heat-labile form
Heat-stable form of B. cereus
- emetic (vomitting) form associated with enterotoxins
: ingestion of enterotoxins, not bacteria
: incubation period short (1-6 hours)
: illness duration short (~24 hours)
: vomitting, nausea, abdominal cramps
Heat-labile form of B. cereus
- diarrheal form
: stimulates adenylate cylate-cAMP system in intestinal epithelial cells
: profuse watery diarrhea
: true infection with bacteria
: contaminated meat and sauces
Diagnosis of B. cereus
culture and gram-stain
- clinical specimens
- implicated foods
Treatment of B. cereus
- vancomycin (glycopeptide)
- ciprofloxacin (quinolone)
- gentamicin (amino glycoside)
Biology of Corynebacterium
- Gram-positive; irregular, “club-shaped” rods
- aerobic
- catalase-positive
- non-spore forming
Virulence factors of Corynebacterium diphtheriae
diphtheria toxin
- lysogenic bacteriophage
- beta-phage
- introduces exotoxin encoded on tox gene
- toxin inhibits protein synthesis by inactivating elongating factor 2 (EF-2)
Epidemiology & Transmission of C. diphtheriae
- found worldwide
- maintained in population by asymptomatic carriage in oropharynx & skin
- transmission via respiratory droplets
- immunisation program in US controls disease
Pathogenesis & Disease of C. diphtheriae
- respiratory
: sudden onset, malaise, sore throat
: exudative pharyngitis progresses into psuedomembrane (whitish plaque)- bacteria, dead cells, fibrin: firmly adheres to underlying tissue
: systemic complications (myocarditis, recurrent laryngeal nerve palsy)
Diagnosis of C. diphtheriae
- clinical diagnosis
- culture: selective medium- cysteine-tellurite blood agar (CTBA)
: tellurite inhibits growth of most URT bacteria and gram-negative rods
: reduced by C. diphtheriae- produces gray/black colour on agar
- PCR: detect tox gene
- toxigenicity testing: producing of exotoxin
: Elk test
Diagnosis of C. diphtheriae with Elk test
- sterile paper with diphtheria antitoxin imbedded into agar medium
- C. diphtheriae isolates are streaked across plate ar 90 degrees to the antitoxin strip
- toxigenic C. diphtheriae is detected because secreted toxin diffuses from area of growth and reacts with antitoxin to form lines of precipitin
Prevention & treatment of C. diphtheriae
- penicillin or vancomycin
- antitoxin
- education on vaccination
- DPT vaccine (diphtheria, pertussis, tetanus antigens)
Myobacterium biology
- aerobic rods
- cell wall is rich in lipids
: mycolic acids- increases resistance to desiccation and many chemicals
- acid-fast bacilli
Myobacterium tuberculosis biology
- facultative intracellular pathogen (can live/reproduce inside/outisde of cell)
- aerobic rod
virulence factors M. tuberculosis
- dormant in macrophages
- cord factor
: 2 mycolic acids together with a disaccharide (2 molecules of glucose)
: inhibits neutrophil migration and damages mitochondria
- sulfatides
: prevent phagosome fusion with lysosome containing bacterocidal enzymes
Epidemiology & transmission of M. tuberculosis
- reservoir: human lungs
- transmission via respiratory droplets
: coughing, sneezing
: ventilation & isoloationi important to prevent transmission
pathogenesis & disease of M. tuberculosis
- enters airways and penetrates alveoli
- phagocytosed by alveolar macrophages
- bacteria multiply (days-weeks) in macrophages
- infected macrophages travel to regional lymph nodes (hilar, mediastinal) and tissues
- alveolar macrophages release cytokines (IL-12 and TNF-alpha) that increases local inflammation and recruits T-cells
- T-cells are activated and secrete IFN-gamma, which activates macrophages to increase phagosome-lysosome fusion and intracellular killing of bacteria (phagosome has bacteria)
granuloma of M. tuberculosis
- prevents further bacteria spread
- necrotic mass surrounded by dense wall of CD4, CD8 and NK cells and macrophages
- bacteria can remain dormant in this stage or can be reactivated years later when / if the patient’s immunity decreases
clinical manifestations of M. tuberculosis
- initially, non-specific complaints of malaise, weight loss, cough and night sweats
: primary infection with resolution or with progression
- reactivation
- disseminated (military) tuberculosis
- sputum may be scant or bloody and purulent (pus)
: cavitary disease with tissue destruction is associated with sputum production with hemoptysis
How is clinical diagnosis of M. tuberculosis supported
- radiographic evidence of pulmonary disease
- positive skin test reactivity
- lab dectection
Lab Diagnostics of M. tuberculosis
- Auramine-rhondamine staining bacilli (fluroscent apple green)
- Acid fast stain (Ziehl Neelson method)
- growth is slow on Lowenstein Jensen medium
Acid fast stain for Mycobacterium
Ziehl Neelson Method
- carbolfuschin (primary stain- red colour)
- acid fast organisms resist decolourization with acid alcohol
- after decolourisation, methyelene blue (secondary stain) is added to organisms to counterstain any material that is not acid fast
