Clincal Microbiology Flashcards
What are the characteristics of bacteria
Multiply by fission
Form colonies
Bacteria morphology
Bacteria Shapes
Coccus
Bacillus
Spirochetes
Types of Gram stains
Gram positive appear blue-purple,
gram negative appear pink-red
Growth requirements
Medium provides nutrients for growing
microorganisms
○ Bacteria that can only grow on
specialized medium are fastidious
bacteria
○ Aerobic and anaerobic bacteria
what is Pathogens
bacteria that
cause disease through
infection
what is communicable disease
Diseases that spread from
person
Normal flora
bacteria
that are natural inhabitants
of the human body
Opportunistic pathogens
invade the body only
when immune defenses are
impaired
Bacteriological Procedures
Specimen collection
- Bacteria collected
- Find growth of
bacteria on a medium that
provides nutrients
Identifying bacteria
- can use Microscopic morphology, colony
appearance, reactions with Gram
stain, growth on media,
biochemical reactions, gene
probes, antibody reactions
Antibiotic susceptibility testing
- Find minimum inhibitory
concentration (MIC)
Reasons for Ordering Cultures
bad Throat
- Sore throat, fever, negative
rapid strep test
bad Nasal
- B Pertussis, MRSA,
rhinovirus, flu, RSV
bad Urine
- Recurring UTIs
bad Sputum
- Prolonged cough, fever, lung
infection, TB, pneumonia
bad Stool
- Diarrhea with blood and mucous,
prolonged bout of diarrhea
Signs of intestinal infection,
Salmonella, E. coli
bad Blood
- Suspected sepsis or
septicemia
throat culture
If negative for rapid test for strep throat, perform throat culture
● Sterile swab should be touched to tonsils or pharyngeal surfaces; tongue and
mouth will contaminate swab
● Streak immediately on blood agar plate if available
Nasal Culture
Nasopharyngeal swabs with flexible shafts, moistened with sterile saline; one
swab for each nasal cavity
● Swab inserted into anterior nostril 2 cm and rotated gently against nasal mucosa
● No rayon/Dacron swabs because they kill target organisms
● If pertussis (whooping cough) suspected, calcium alginate swab used with
Regan-Lowe medium transport
Urine Culture
Clean-catch collected in sterile container
● If urine can’t be cultured within an hour after collection, use sterile tube such as
BD Urine C&S Transport kit
Sputum Culture
Sputum is material coughed up from lungs and is examined to aid in diagnosis
of lower respiratory tract infection
● Collection instructions
○ Rinse mouth
○ Collect 2 separate specimens in sterile containers, such as BD Sputum
Collection System
Stool Culture
One specimen per day collected on 3 consecutive days
● 3 vials that contain medium such as Cary Blair or Amies
● Para-Pak vials for culture
Blood Culture
Collected by venipuncture; site must be cleansed first with alcohol then with a swab
containing 2% solution of tincture of iodine
● In order of draw, blood culture bottles are filled first; 2 bottles usually collected, aerobic bottle
filled first
● Bactec instrument inoculated blood cultures and senses changes in CO2, which is released as
microorganisms grow; smear of blood from positive bottle can be made and Gram-stained to
determine microorganism present
● Trek Diagnostic ESP EZ Draw draws 0.1 mL of blood
Aseptic technique
set of procedures used to prevent spread of infection during surgical
procedures
Aseptic technique (physical)
○ Protective clothing
○ Fluid-resistant lab coat
○ Disposable lab coats/gowns
Microbiological safety cabinets
Class I – protect worker and environment but not culture
Class II – provide protection to worker, environment, and culture
Disinfectants and antiseptics
Disinfectants are chemicals used to kill/control growth of microorganisms or inanimate objects
Bactericidal – kill bacteria
Bacteriostatic – slow growth of bacteria
Antiseptics are chemicals used to control growth of microorganisms on living tissue
Sterilization frees an article or area from all living organisms
Agar
derivative of seaweed used to solidify liquid
media, which grows and isolates bacteria
Primary medium
first inoculated with
specimen collected from patient
○ Most common is blood agar
Enriched medium
supports growth of
wide variety of organisms
Selective media
ingredients that inhibit the
growth of certain microorganisms while allowing the
growth of others
Indicator medium
detects metabolic activity of
particular microorganisms
Inoculation
Process of transferring microorganisms to a growth medium
How to do swab agar plate
Gently roll specimen swab onto one quadrant of agar plate, then prep bacterial smear by
rolling swab across sterile glass slide
○ Inoculating loop spreads inoculated material over agar plate to produce isolated colonies
○ Accomplished by streaking 4 quadrants, decreasing amount of culture material as
streaking proceeds into each quadrant
○ Loop Is sterilized and cooled to spread material into each quadrant
Incubation
Usually kept in 35-37 C
incubator
● Plates are inverted to
prevent formation of
condensate inside lid
● For aerobic specimens,
keep lid open a little bit
and incubate in aerobic
incubator (ambient air)
● Pathogens that grow in
CO2 are kept in CO2
incubator with 5-10% CO2
● Anaerobic specimens can
be incubated in sealed
bags/containers
What to do after Observation After 24 Hours
Growth on blood agar should be observed for hemolysis, the lysis of RBCs by bacteria
growing on blood agar
Beta hemolysis
completely lyses blood cells and makes surrounding area transparent
Alpha hemolysis
incompletely lyses RBCs and makes area green
how to do Gram stain
○Primary stain
■ Crystal violet poured on slide
■ All bacteria stain purple
○ Gram’s iodine
■ Gram’s iodine, a mordant, allows
dye to adhere
■ All bacteria remain purple
○ Decolorizer
■ Alcohol added briefly to slide when
it is tilted downward at 30 degrees
■ Gram-negative bacteria will be
colorless
○ Counterstain
■ Slide flooded with counterstain
safranin
■ Gram-negative bacteria turn pink
Performing the Throat Culture
Collecting the specimen
○ Pharyngeal surfaces swabbed with sterile
Dacron/rayon swab; passed across surfaces
of both tonsils or surfaces of fossae and
back of throat
○ Don’t touch tongue or inside of mouth
● Inoculating the media
○ Throat swab immediately used to
inoculate a quadrant of the blood agar
plate
○ Paper disk with antibiotic bacitracin can
be placed on concentrated streak in 1st
quadrant
● Incubating the culture
○ CO2 concentration of 10%
● Reading throat culture plate
○ Beta hemolysis should show up in stabs
○ Presence of zone of inhibition around
bacitracin disk and presence of
beta-hemolytic colonies shows Group A
Strep
Why are urine cultures collected?
Culture requested when patient
has UTI symptoms
What contributes to STDs?
Escherichia coli - gram-negative rod part of flora of
intestinal tract
How to perform urine culture
Clean-catch; must be inoculated within 1 hour of collection. Put into a blood agar. Inhibit growth of gram-positive so gram neg grows. Incubate at 35-37 degrees. Colony color, and hemolysis should be noted. Count of 100 000/mL or greater is evidence of UTI
What are some common gram positive bacteria
Staphylococcus and Streptococcus
Staphylococcus
Large, white or yellow
○ On Gram stain, appear spherical and in grape-like clusters
Streptococci
Small, colorless
○ Spherical, chains or pairs
Catalase test
used to perform on gram positive. staph is positive and strep negative
Coagulase test
can differentiate between Staph aureus and other
staph
Identification of Gram-negative Bacteria
Klebsiella form bubblegum pink, mucoid colonies. Gram negative bacteria will grow on EMB and MAC. Reported as gram-negative rods, coliform by EMB, where coliform refers to intestinal bacteria
Antibiotic Susceptibility Tests
Tests which antibody works against certain viruses
Minimum inhibitory concentration (MIC)
Instruments perform bacterial identification. Swab bacteria on an agar plate and put antibodies in each section of an agar plate. The antibody that works the best when it has a circle around the antibody
vaginitis,
an infection of vagina
Common microorganisms in vaginitis
Gardnerella vaginalis, Mobiluncus species, Streptococcus
group B, Chlamydia trachomatis, Neisseria gonorrhoeae, the protozoan
Trichomonas and the yeast Candida albicans
waht is Saline wet mount prep
Mix drop of vaginal specimen with drop of 0.85%
saline on slide
KOH prep of vaginal secretions for yeasts and fungi
2 drops vaginal material with 1 drop 10% KOH
solution
■ Fungal elements appear as hairs or threads
Gram stain of endocervical secretions of bacteria and
yeasts
G Vaginalis is gram-variable; scattered over
constituents of smear
■ Yeast stains dark purple
■ Examined for N gonorrhoeae, which causes
gonorrhea; gram-negative, kidney bean-shaped
urethritis
an inflammation of urethra
If midstream urine contains RBCs, bacteria,
WBCS, or protein,?
then patient may have UTI
instead of STD
how to do Urethral culture
Urethral discharge collected using urogenital
swab and cultured for gonorrhea
○ Inoculated on MTM plate to identify
gonorrhea
what does a Urethral gram stain do
Examined for WBCs and bacteria
Identification
of Gonorrhea
N. gonorrhoeae appear as tiny,
shiny, grayish colonies growing
along steak pattern. Oxidase test can be used to aid in
identifying colonies, which is
positive. Purple-black color forms when
oxidase-positive colonies are
exposed to reagent
Types of herpes
Herpes simplex virus, type 1 (HSV-1)
is responsible for cases of oral
herpes and herpes simplex virus, type 2 (HSV-2) causes genital
herpes
Identification of
Herpes (HSV)
If herpes lesion present, vesicle can
be broken with swab or needle and
fluid collected with swab, which is
then sent to lab for culture
● Serum titers of IgG and IgM can
also be measured
Identification of
Chlamydia
Caused by C. trachomatis
● DNA probes use specific nucleic
acids, called probe DNA, that is
added to patient specimen; if
patient is positive,
color/luminescence is produced
● In females, Chlamydia can cause
cervix to bleed easily, cervix said to
be friable
Identification of
Syphilis
Syphilis is venereal disease caused by
Treponema pallidum, a spirochete, or spiral
bacteria
● Primary syphilis characterized by skin lesions,
organ/tissue damage in secondary stage,
cardiovascular/CNS affected in late-stage
● Patient with T. pallidum produces non-specific
antibodies called reagin; RPR (rapid plasma
reagin) test has carbon-containing cardiolipin
antigen that reacts with reagin; also flocculation
test
How do pandemics
re-emerge and spread?
Environmental Change
Population concentrations
international FLights
Public health
Pathogen adaptation
Natural disasters
West Nile Virus
In 2002, it caused largest WNV epidemic and animal epizootic ever reported, with 4000 human
cases and 284 deaths in the US
● Normally transmitted by mosquitoes
West Nile Virus Symptoms
fever, hemorrhaging caused by organ damage
waht are Viral Hemorrhagic Fevers
Symptoms: fever, hemorrhaging caused by organ damage
● Caused by RNA viruses in 4 families: arenaviruses, filoviruses,
bunyaviruses, flaviviruses
What are examples of Viral Hemorrhagic Fevers
Rift Valley Fever
Ebola and Marburg viruses
Severe Acute Respiratory Syndrome
Avian influenza
Rift Valley Fever
Rift Valley fever occurs in livestock and humans in Africa
○ Caused by virus of Bunyaviridae family and transmitted by
mosquitoes
Ebola and Marburg viruses
Ebola and Marburg viruses are filoviruses that cause high
mortality
○ WHO documented 17 outbreaks in humans
Severe Acute Respiratory Syndrome
Mycobacterium tuberculosis causes tuberculosis
○ As HIV has increased worldwide, incidence of TB has also
increased
Avian influenza
Reservoir for influenza A are birds
○ Outbreak in May 2006 in Indonesia was possible pandemic
What are biological weapons?
Diseases used againest peopl in war
