Specimen collection and Handling Flashcards
Specimen Collection Timing
Collect during ACUTE/EARLY PHASE OF ILLNESS (2-3 days for viral infection). Collect before administration of antibiotic, antifungal, or antiviral.
Specimen Storage
Collect and store specimen in STERILE SCREW-CAP CONTAINER except for STOOL
Swab Use - General
POOR specimen if aspirate or biopsy can be obtained
Swab Use - Appropriate
nasal/nasopharyngeal, oropharyngeal, eyes, ears, or genitals
Swab Use - Not Appropriate
NOT FOR ANAEROBES AND FUNGI
Swab Shaft - Toxic to Chlamydia and Mycoplasma
Wooden
Swab Shaft - Used usually in a calcium alginate-tipped swab
Aluminum
Swab Shaft - General purpose shaft for bacteria and viruses
Plastic
Swab Tip - General use for bacteria and viruses
Dacron
Swab Tip - Excellent collection and release of samples
Rayon
Swab Tip - Toxic to Neisseria (unless with charcoal)
Cotton
Swab Tip - Toxic to enveloped virus & Neisseria, interferes with PCR methods
Calcium alginate
Swab Design - Can trap organisms, leading to decreased recovery/sensitivity
Woven/Traditional
Swab Design - Uses Nylon fibers for enhanced uptake and release of microbes
Flocked
Enhancing Microbe Recovery
Place swab in 0.5-1.0 mL of sterile broth/saline and vortex it
Specimen Transport - Container
Leak-proof plastic bag with separation for paperwork; has BIOHAZARD label
Specimen Transport - Time Frame
Transport within 2 hours to the lab, except sterile specimens (within 15 minutes)
SARS-CoV-2 Transport Medium
3mL of viral transport medium (VTM), Amies transport medium, phosphate buffered saline, or sterile saline
Bacteria Susceptible to Air Exposure
Anaerobes
Bacteria Susceptible to Temperature Changes
Neisseria meningitidis
Bacteria Susceptible to pH Changes
Shigella
Bacteria Susceptible to Glycerol
Vibrio
Shipment - Arrangement of Container
Sample > Primary container > Absorbent > Secondary container > Dry ice/Liq. nitrogen > Outer/Tertiary container
Shipment - Category A
Substances/microbes capable of permanent disability or life-threatening
Shipment - Category B
Substances/microbe incapable of permanent disability or life-threatening on its current form
Preservation - Urine
Use BORIC ACID (bacteriostatic for 3 days)
Preservation - For Parasites
Don’t use formalin or Polyvinyl alcohol (PVA)
Preservation - Stuart/Amies Medium
Contains charcoal to neutralize fatty acid toxic to bacteria with pili
Preservation - Anticoagulants
Prevent clotting in blood, bone marrow, synovial fluid
Preservation - SPS (Sodium Polyanethol Sulfonate)
0.025-0.050% concentration, anticoagulant, anticomplement, antiphagocytic
Preservation - SPS - Function
Neutralizes aminoglycoside, neutralize with 1% gelatin
Preservation - SPS - Anaerobe Sensitivity
0.025% concentration preferred for some anaerobes and Neisseria
Preservation - Heparin
Can be used for viral culture and Mycobacterium culture, inhibits some gram-positive bacteria and yeasts
Preservation - Unused Anticoagulants
Citrate, EDTA, and others – should never be used
Preparation - Plating
Sterile specimens are plated directly into the culture media
Preparation - Centrifuge
> 1 mL of fluid samples centrifuged at 3000 g for 20 minutes
Preparation - Tissue Homogenization
Tissues may be homogenized using a tissue grinder (disadvantages: destroys fungi)
Specimen for Abscess, Lesion, Wound, Ulcer container
Swab in Amie’s or Stuart’s medium
Common Isolates Abscess, Lesion, Wound, Ulcer - Macule
Dermatophytes, T. pallidum
Common Isolates -Abscess, Lesion, Wound, Ulcer Papule
HPV, S. aureus, Pox
Common Isolates -Abscess, Lesion, Wound, Ulcer Nodule
C. diphtheriae, Sporothrix, M. marinum
Common Isolates - Abscess, Lesion, Wound, Ulcer Pustule
Candida, Herpes, N. gonorrhoeae, S. aureus, VZV
Common Isolates - Abscess, Lesion, Wound, Ulcer Bulla
C. perfringens, HSV, S. aureus, V. vulnificus
Common Isolates - Abscess, Lesion, Wound, Ulcer Ulcer
B. anthracis, H. ducreyi, T. pallidum
Blood - Adult Volume
≥ 20mL/set
Blood - Pediatric Volume
1-20mL/set
Blood - Culture Bottle
Both aerobic and anaerobic
Blood - When to Draw
During febrile episodes, 2-3 sets per 24 hours, Do not draw >4 sets in 24 hours
ARD (Antimicrobial Removal Device) - Manual
1:10 blood to broth ratio, Growth detected in pellicle/turbidity/hemolysis/bubbles
ARD - Automated
1:5 blood to broth ratio, Growth detected by BacT/Alert (5 days TAT)
Common Blood Isolates
Staphylococcus aureus, E. coli, Staphylococcus epidermidis, Enterococcus, B-hemolytic streptococci, Candida albicans, Pseudomonas aeruginosa, Klebsiella pneumoniae, Enterobacter cloacae, Proteus, Bacteroides, Clostridium
Bone Marrow - Culture Bottle
May use blood culture bottle if volume is sufficient
Common Bone Marrow Isolates
Brucella, Histoplasma
Amniotic, Peritoneal, Pericardial, Pleural Fluids container
Sterile screw cap or blood culture bottle, Transport within 15 minutes at room temperature
Common Fluid Isolates - Pericardial
Coxsackie, Adenovirus, Influenza
Common Fluid Isolates - Arthritis
S. aureus, Streptococci, H. influenzae, Bacteroides, N. gonorrhoeae, Candida, Cryptococcus
CSF/Spinal Fluid - Transport Temperature
22-25°C, delayed testing at 20-24°C or 35°C
CSF/Spinal Fluid - Viral Studies
Transport at 4°C, Transport within 15 minutes at room temperature
CSF/Spinal Fluid - Direct Examination
India ink and Gram stain
CSF Collection - Number of Bottles
3 or more bottles collected: 2nd bottle → Micro, if only 1 bottle collected: Micro (1st) → Hema (2nd) → Chem/Serology (3rd)
CSF - Cytocentrifugation
Use acridine orange for best sensitivity and organism recovery
Meningitis - Newborn
S. agalactiae, E. coli
Meningitis - Children
H. influenzae
Meningitis - Adult
N. meningitidis
Meningitis - Geriatric
S. pneumoniae
Meningitis - Immunocompromised
L. monocytogenes
Meningitis/Encephalitis - Additional Causes
P. cantonensis, N. fowleri, Acanthamoeba, Coxsackie, HIV, T. solium, Toxoplasma
Ear Specimen - Transport Container
Sterile screw cap container
Ear Specimen - Anaerobic Transporter
Inner ear aspirate
Otitis Externa - Acute
S. aureus, S. pyogenes, P. aeruginosa (Swimmer’s ear/Acute diffuse otitis externa)
Otitis Externa - Chronic
P. aeruginosa, Anaerobes
Otitis Media - Acute
S. pneumoniae, H. influenzae, M. catarrhalis, S. pyogenes, RSV, Influenza virus, coronavirus, enterovirus, rhinovirus
Otitis Media - Chronic
Anaerobes (Bacteroides, Fusobacterium, Prevotella, Porphyromonas, Peptostreptococcus)
Eye Conjunctiva or Corneal Scraping - Transport Method
Direct inoculation to BAP and CAP
Eye Conjunctiva or Corneal Scraping - Transport Time
Transport within 15 mins at room temperature
Corneal Scraping - Incubation
28°C in SDA
Common Eye Isolates
Chlamydia trachomatis, Acanthamoeba, Herpes simplex virus
Gastric Wash or Lavage - Transport Time
Transport within 15 mins at room temperature
Gastric Aspirates - Indications
Infants or AFB, process within 4 hours or neutralize with 10% sodium carbonate
Common Gastric Isolates
Campylobacter jejuni, Helicobacter pylori, Mycobacterium tuberculosis, Ascaris larvae
Rectal Swab - Transport Medium
Enteric transport medium
Rectal Swab - Inoculation Method
Insert 1-1.5 cm deep, feces visible on swab, inoculate in enteric media
Common Rectal Swab Isolates
Pathogenic Enterobacteria, Vibrio, Campylobacter, Helicobacter, Parasites (stool sample), Norwalk, Rotavirus, C. difficile
Urine - Best Collection Method
Mid-stream clean catch (Ambulatory/Outpatient/Non-invasive)
Urine - Catheterized Collection Method
Non-Ambulatory/In-Patient/Invasive, Clean urethra with soap and water, collect remainder after first 15mL
Urine - Suprapubic Aspiration
For Anaerobic culture, disinfect skin first
Common Urine Isolates
Escherichia coli (UPEC), Klebsiella, Staphylococcus saprophyticus, Enterococci, Proteus, Pseudomonas, Candida, Acinetobacter
Urine - Quantitative Counting
1 µL Loop (1 colony x 1000 = CFU/mL), 10 µL Loop (1 colony x 100 = CFU/mL)
Urethra - Collection Method
Swab in Amie’s or Stuart’s medium, collect 1 hour after patient has urinated
Urethra - Direct Smear
Gram stain
Urethra - Collection Time
Do not refrigerate, collect if >1 hour after urination
Common Urethral Isolates
N. gonorrhoeae, Chlamydia trachomatis, Ureaplasma, Mycoplasma
Cervix - Collection Method
Swab in Amie’s or Stuart’s medium, Remove mucus before collection
Cervix - Collection Instructions
Do NOT use lubricant on speculum, Swab deeply into endocervical canal
Cervix - Swab Material
Do NOT use wooden shaft or Calcium alginate swab
Common Cervix Isolates
N. gonorrhoeae, C. trachomatis, Ureaplasma
Vagina - Collection Method
Swab in Amie’s or Stuart’s medium, Remove exudate
Vagina - Collection Instructions
Swab secretions and mucous membrane
Vagina - Bacterial Vaginosis Diagnosis
Culture is not recommended
Vagina - Group B Streptococcus Screening
Use LIM broth
Common Vagina Isolates
Lactobacillus, G. vaginalis, Mobiluncus, S. agalactiae
Prostate - Collection Method
Swab in Amie’s or Stuart’s medium, Clean urethral meatus with soap and water
Prostate - Collection Instructions
Massage prostate through rectum and swab the secretions, Plate immediately
Common Prostate Isolates
E. coli, other enteric organisms, P. aeruginosa, Mycoplasma genitalium
IUD - Collection Method
Sterile screw cap container, Disinfect skin before removal
IUD - Transport Instructions
Transport within 15 mins at room temperature, Plate immediately
Common IUD Isolates
Actinomyces infections
IV Catheter - Collection Method
Sterile screw cap container, Disinfect skin with alcohol before removal
IV Catheter - Collection Instructions
Do not accept foley catheters, Roll segment back and forth for 4x using sterile forceps
IV Catheter - Quantitative Culture
≥ 15 colonies are significant
Common IV Catheter Isolates
S. epidermidis, S. aureus, other CONS, Enterobacteria, P. aeruginosa, Candida, Corynebacterium
Nasopharyngeal Swab - Collection Method
Swab in Amie’s or Stuart’s medium, Rotate the swab for 5 seconds
Nasopharyngeal Swab - Transport Instructions
Transport within 15 mins if no transport media, or within 2 hours at 20-24°C
Common Nasopharyngeal Isolates
B. pertussis, SARS-CoV, H. influenzae, N. meningitidis, Rhinovirus, Coronavirus, Adenovirus, Influenza, Klebsiella ozaenae
Nasal Swab - Collection Method
Swab in Amie’s/Stuart’s, Premoisten with sterile saline, insert 1-2 cm into nares and rotate
Common Nasal Isolate
MRSA screening
Throat Swab - Collection Method
Sterile screw cap, Clean genitals with soap and water
Common Throat Isolates
S. pyogenes (Grp A Streptococcus), Pharyngitis – S. pyogenes, Group C/G streptococcus, Arcanobacterium, Fusobacterium, N. gonorrhoeae, Corynebacterium ulceran
Sputum - Collection Method
Sterile screw cap, Brush teeth then rinse or gargle with water
Sputum - Aerosol-induced Sample
Use 15% NaCl or 10% glycerin
Common Sputum Isolates
M. tuberculosis, Corynebacterium, Mycoplasma pneumoniae, Chlamydia trachomatis, Chlamydia pneumoniae, Bordetella pertussis, Legionella, Pneumocystis, Nocardia, Histoplasma, Coccidioides, RSV, Coronavirus, Influenza/Parainfluenza
Tissue - Collection Method
Anaerobic transporter, Disinfect skin, Do not allow the specimen to dry, May need to be homogenized
Common Tissue Isolates
Anaerobes, Gram-negative bacilli, Gram-positive organisms, Fungi
Specimen Transport for Viruses, Chlamydia, Mycoplasma
Transport in appropriate media at 4°C (ice pack or liquid nitrogen)
Transport Time for Majority of Samples
Transport within 2 hours at room temperature
Exceptions to Transport Time
Normally sterile samples (within 15 minutes), gastric biopsy (within 1 hour)
Specimen Storage at Room Temperature (20-24°C)
Hair, Nails, Skin scraping for fungi (<2 hrs)
Specimen Storage Up to 24 Hours at Room Temperature
Abscess, lesion, wound, pustule, ulcer, Nasopharyngeal swab, Oropharyngeal swab, Sterile body fluids (Peritoneal, Pleural, Pericardial), Inner ear sample, Eye samples (Conjunctiva, Cornea, Vitreous fluid), Rectal swab, Genital samples (cervix, vagina, cul-de-sac, prostate, Urethral swab), Bone marrow, Tissues, Preserved stool (in holding media), Preserved urine (in holding media), CSF (or at 35°C)
Specimen Storage at 4°C (Refrigerator Temperature) Up to 48 Hours
Feces for Clostridium difficile (culture or molecular)
Specimen Storage at 4°C (Refrigerator Temperature) Up to 24 Hours
Feces for E. coli O157:H7, Outer ear sample, Gastric aspirate/biopsy, Respiratory samples (BAL, Bronchial washing, Sputum), Unpreserved/Fresh urine, Unpreserved/Fresh stool
Specimen Storage at 4°C (Refrigerator Temperature) Up to 2 Hours
IV catheters, Pins
Specimen Storage at 35°C (Incubator Temperature) Within 6 Hours
CSF
Specimen Storage Upon Receipt at the Lab
Blood, Sterile body fluids in blood culture bottle
Level 1 Immediate Critical/Invasive Specimen
Amniotic fluid, Blood, Heart valve, Brain, CSF
Level 2 Unpreserved Specimen
Body fluids (not listed in Level 1), Bone, Wound Drainage, Feces, Sputum, Tissue
Level 3 Quantitation Required Specimen
Catheter tip, Urine (midstream/suprapubic/catheterized)
Level 4 Preserved Specimen
Preserved Urine, Preserved Stool, Swabs in holding/transport medium
Rejected Samples - Criteria
Non-matching label and requisition, Specimen transported at wrong temperature or wrong media, QNS, Leaking sample, Exceeds 2-hour transport, Preserved with fixative (formalin), Swabs for anaerobic culture, Calcium alginate swab for molecular assay, Dried sample/swab, Foley catheter tip