Microbiology

Antonio Rivas PA-C

Feb 2008

Microbiology

• Clinical microbiology encompasses– Bacteriology– Parasitology– Virology– Mycology

Bacteriology

• Characteristics of bacteria– Multiply by fission– Grow in colonies

• Morphology– Coccus– Bacillus– Spiral– Growth patterns

Specimen collection

• Collect before antimicrobials are given• Specimen collected where the organism is

most likely to be found w/o external contamination

• Stage of the disease– Enteric pathogens are present in higher numbers

during the acute or diarrheal phase of the infection

• Sufficient quantity of specimen • Prompt delivery to the lab

Aseptic Technique

Techniques Culture

Culture Techniques

Bacteriological Growth Media

• Agar plates contain different components that enhance or inhibit the growth of certain microorganisms

• Examples:– MAC-selective media - inhibits Gram

Positive organism– HE- selective media- inhibits Gram

negative organism

Bacteriological Growth Media

Incubation - Plates are incubated for :24-48 hrs at different temperature and O2 concentration-

- Most human pathogens grow best at 35-37 degree C

Culture Techniques

• Observing culture after 24 hours– Colony characteristics– Presence (or absence) of hemolysis

Staphylococcus aureus Beta hemolyticus strept.

Colony Characteristics• Colony size, shape, moist or dry, color, smell

• Pseudomonas sp. Smells like grapes• Neisseria gonorrhea smells like sweaty tennis shoe• Shigella is mucoid and purple

– Presence of hemolysis in blood agar plate (Strep.pyogenes-throat inf.-beta hemolysis)

• Beta-hemolysis:• Lysis of the red cell present in the media showing clear

area around the bacterial growth

Automated ID

Gram Stain• Bacterial species divided into two groups

according to how they take Gram Stain– Gram positive : take up the crystal violet basic

stain (stain purple)– Gram negative : allow the crystal violet to easily

washout with acetone and take up the Safranin dye (stain pink)

– Related to the cell membrane composition

Gram Stain

• Important for– Bacterial identification– Antibiotic susceptibility testing– Preparing a smear

• From a swab• From a culture

Preparing the Bacterial Smear

• Heat-fixing the smear– Smear must be dry– Affixes bacteria to slide– Do not use excessive heat

Gram Stain

• Primary stain

• Gram’s iodine– Mordant

• Decolorizer

• Counterstain

Observe the Stained Smear

• Oil immersion

• Gram reactions– Gram (+) → purple– Gram (-) → pink

Microbiology

• Bacteremia : transient release of bacteria to the blood stream, indicates the presence of a focus of disease

• Septicemia or Sepsis : a situation in which bacteria and their products are causing harm to the host

• Terms are used interchangeable

• Portals of entry for septicemia:– GU– Respiratory– Abscesses– Surgical wound infections– Biliary tract– Other sites

• Organisms most commonly isolated from blood are:– Gram positive cocci– Coagulase neg staphylococci– Staphylococcus aureus– Enterococcus sp.

• Meningitis: infection in the subarachnoid space, between the Pia-mater and the Arachnoid

• Dx by PE, CSF analyisis, and cultures • Hematogenous spread• Children <5 years

– Haemophilus influenza

• Neonates– Group B streptococci– E.coli– Listeria sp.

• Most common bacteria causing meningitis in children age 6 years and up– Haemophilus influenza– Neisseria meningitidis – Streptococcus pneumoniae

• Meningitis in adults– Neisseria meningitidis(young adults crowded

conditions)– Pneumococci – Listeria monocytogenes– Staphylococcus aureus

Staphylococcus saprophyticus

• Gram positive cocci• Second cause of UTI in young females after

E.coli• Symptoms :• Burning when urinating• Increased urge to urinate• Dripping effect• Razor like pain in the lower abdomen and

during intercourse• Treated with Quinolones in the US

Enterococcus faecalis

• Gram positive bacilli

• Inhabitant in GI tract humans and animals

• Life threatening nosocomial infections

• High level antibiotic resistance

• Can cause endocarditis, bladder, prostate and epididymal infections

Streptococcus pyogenes

• Gram positive cocci in chains• Group A, beta hemolytic• Strept throat-pharyngitis• Impetigo• Cellulitis• Necrotizing fasciitis• Toxic shock• Rheumatic fever• Glomerulonephritis• Sensitive to penicillin

Candida albicans

• Yeast, fungi

• Opportunistic oral and genital infection in immunocompromised patients

• Live in human mouth and GI

• May occur in blood

• Thrush – immunocompromised

Cryptococcus neoformans

• Encapsulated yeast-like fungus

• Cryptococcal meningitis in immunocompromissed patients

• India ink stain used for Dx in CSF

E.coli

• Gram negative rod

• Virulent strains can cause gastroenteritis, UTI, neonatal meningitis

• Ascending UTI with fecal contamination