The University of Iowa (UIHC)
Department of Pathology
LABORATORY SERVICES HANDBOOK


Bacterial Culture

Order Form: A-1a Clinical Microbiology Laboratory or Epic Req
  Microbiology
BT 6004
356-2591 (0700-2300)
Bacteriology/Virology Section

356-3527 (2300-0700)
Core Lab
Delivery Instructions:
Deliver to laboratory immediately after collection.
Testing Schedule:
0700-2200, 7 days a week, including holidays.
Comments:
Gram stains are automatically performed on fluids and exudates other 
than blood and urine.  Do not send sterile body fluids in plastic red 
top tubes.  These tubes contain a clot activator which may affect 
testing.  Therefore, tests may be unreliable.

Label transport tube with two patient identifiers, date and time of 
collection.

Transport at room temperature unless otherwise specified.
A. Abscess - Tissue or aspirates are always superior to swab specimens.
   Remove surface exudate by wiping with sterile saline or 70% alcohol.
   Aspirate with needle and syringe. Cleanse rubber stopper of
   anaerobic transport device (907464) with alcohol; push needle
   through septum and inject all abscess material on top of agar. If a
   swab must be used, pass the swab deep into the base of the lesion
   to firmly sample the fresh border. Transport time less than or
   equal to 2 hours.

B. Anaerobic cultures - Tissue or aspirates are preferred rather than
   swabs. Fluid collections should be aspirated through disinfected
   tissue or skin. For superficial ulcers, collect material from below
   the surface (after surface debridement or use a needle and
   syringe). Submit specimens using anaerobic transport media: BBL
   Port-A-Cul anaerobic collection jar (907722) for tissue or ACT II
   tube system for fluid and swab specimens (907464). A sterile screw-
   cap container (907069) may be used for tissue if transported to the
   microbiology lab immediately (add drops of sterile saline to keep
   small pieces of tissue moist). Deliver all specimens to the
   laboratory immediately after collection.

C. Blood
   a. Adult - Cleanse skin with ChloraPrep one-step 1.5 mL Frepp
      Applicator (907672):
      1) Holding the applicator sponge downward, pinch wings on
         applicator to break ampule and release the antiseptic.
      2) Use a side-to-side motion to scrub the site with the friction
         pad for a full 30 sec; allow site to dry completely (at least
         30 sec) before venipuncture. Do not touch site after prep.
      3) Remove overcaps from bottles (1 aerobic 924171 and 1 anaerobic
         924172) and cleanse each rubber septum with separate 70%
         alcohol swabs. Allow septum to dry for 1 min before
         inoculating.
      4) Draw 20 mL of blood and inoculate each bottle with 10 mL of
         blood.  Do not vent or overfill bottles. Adding low (<8 mL)
         or high (>10 mL) volumes may adversely affect the recovery of
         organisms.  Transport time <2 h.
      5) For adults with a suspected bloodstream infection (BSI),
         collect two initial sets of blood cultures sequentially from
         separate phlebotomy procedures followed by a third and a
         fourth set at 4-6 hour intervals (will detect >99% of BSIs).
         Three sets of blood cultures collected within a 24 hour
         period will detect 96.9 - 98.3% of BSIs. A single set of
         blood cultures to detect BSIs in adults is inadequate (only
         73% sensitivity); two sets of blood cultures will allow
         detection of 87.7-89.7% of BSI episodes.  (J Clin Microbiol
         2007; 45:3546).
      6) If patient is allergic to chlorhexidine, prep site with a
         povidone iodine swab stick (907172) applied in concentric
         circles (start at center). Allow to dry at least 1 min before
         venipuncture. If patient is allergic to iodine, cleanse site
         with 70% alcohol for 60 sec.
   b. Pediatric - Prepare skin and bottles as for adult.  Collect as
      much blood as possible up to 10 mL per bottle.

D. Bone marrow aspirate - Prepare puncture site as for surgical
   incision.  Inoculate blood culture bottle or lysis centrifugation
   tube.  Transport time <2 hours.  Routine bacterial culture of bone
   marrow is rarely useful.

E. Burn - Clean and debride burn. Place tissue in sterile screw-cap
   container (907069). Transfer aspirates to a sterile container. These
   are processed for aerobic culture only. Quantitative culture may or
   may not be valuable. A 3 to 4 mm punch biopsy specimen is optimum
   when quantitative cultures are ordered. Cultures of surface samples
   can be misleading.

F. Catheter Tips - Only intravascular catheter tips from pediatric
   patients and peritoneal dialysis catheters are routinely accepted
   for culture. Send 5 cm of distal tip in sterile screw-cap container
   (907069). Transport time is less than or equal to 15 min. Foley
   catheters are not accepted for culture since growth represents
   distal urethral flora.

G. Cerebrospinal Fluid (CSF) - Aseptically collect CSF from a lumbar
   puncture into sterile tubes (907131). Send second tube (greater
   than or equal to 3 mL) to the Microbiology Laboratory. Transport
   time less than or equal to 15 min. Cerebrospinal fluid for
   bacterial culture should never be refrigerated.

H. Decubitus ulcer - A swab is not the specimen of choice. Cleanse
   surface with sterile saline. Submit tissue or aspirate inflammatory
   material from the base of the ulcer in a sterile tube or anaerobic
   system.  Transport time <2 hours.

I. Ear
   a. Inner ear - Tympanocentesis should be reserved for complicated,
      recurrent, or chronic persistent otitis media. For intact
      eardrum, clean ear canal with soap solution and collect fluid
      via syringe aspiration. Submit in sterile container. For
      ruptured eardrum, collect fluid on flexible shaft swab via an
      auditory speculum.  Transport time <2 hours.
   b. Outer ear - Use moistened swab to remove any debris or crust from
      ear canal. Obtain sample by firmly rotating swab in outer canal.
      For otitis externa, vigorous swabbing is required - surface
      swabbing may miss streptococcal cellulitis.

J. Eye
   a. Conjunctiva - Sample each eye with separate swabs (premoistened
      with sterile saline) by rolling over conjunctiva. When only one
      eye is infected, sampling both can help distinguish indigenous
      microflora from true pathogens.
   b. Corneal scrapings - Collected by ophthalmologist. Using sterile
      spatula, scrape ulcers and lesions; inoculate scraping directly
      onto media (BHI with 10% sheep blood, chocolate, and inhibitory
      mold agar). Prepare 2 smears by rubbing material onto 1-2 cm
      area of slide. Transport time less than or equal to 15 min.
   c. Vitreous fluid - Prepare eye for needle aspiration of fluid.
      Transfer fluid to sterile tube. Transport time less than or
      equal to 15 min.

K. Feces - see stool.

L. Fistula - see abscess.

M. Fluids - see sterile body fluids.

N. Genital - Cultures for Neisseria gonorrhoeae should be collected
   with a sterile swab and inoculated directly to a Jembec plate
   (obtain from laboratory; place white tablet in hole of Jembec plate
   to provide CO2 enriched atmosphere, close top of the plate tightly
   and place in Ziploc bag provided, keep at room temperature, deliver
   to lab as soon as possible). If a Jembec plate is unavailable, an
   aerobic culturette swab (922349) may be used if transported to
   laboratory immediately.
   a. Endocervical - Remove cervical mucus with swab and discard.
      Insert a second swab into endocervical canal and rotate against
      walls. Allow time for organisms to absorb onto the swab surface.
   b. Urethral - Collect urethral specimens at least 1 h after patient
      has urinated. Insert small swab 2-4 cm into urethral lumen,
      rotate, leave for 2s to facilitate absorption.

O. Pilonidal cyst - see abscess.

P. Respiratory, lower - Transport time is less than or equal to
   2 hours.
   a. Bronchoalveolar lavage or brush, endotracheal aspirate – Collect
      fluid in a sputum trap; place brush in sterile container with
      1 mL saline.
   b. Sputum, expectorated - Patient should rinse mouth and gargle with
      water prior to collection; instruct patient to cough deeply.
      Collect specimens in sterile transport containers (907069).
   c. Sputum, induced - Have patient brush gums and teeth, then rinse
      mouth thoroughly with water.  Using a nebulizer, have the patient
      inhale 20-30 mL of 3 to 10% sterile saline.  Collect sputum in
      sterile container.

Q. Respiratory, upper - Transport time <2 hours.
   a. Oral - remove oral secretions and debris from surface of lesion
      with a swab.  Use a second swab to vigorously sample lesion,
      avoiding normal tissue.  Superficial swab specimens should not
      be submitted.  Tissue or needle aspirates are preferred.
   b. Nasal swabs (R/O MRSA) - Insert a sterile swab (use Copan dual
      swab 26200 for PCR; 922349 for culture) into the nose until
      resistance is met at the level of the turbinates (approximately
      2-3 cm into the nose).  Rotate the swab against the nasal
      mucosa.  Repeat the process on the other side.
   c. Sinus aspirates - Aspirate with needle and syringe.  Cleanse
      rubber stopper of anaerobic transport device (907464) with
      alcohol; push needle through septum and inject specimen on top
      of agar.
   d. Throat - Routine throat cultures will be processed only for
      growth of beta-hemolytic Streptococcus species.  Contact
      Microbiology Lab (356-2591) to arrange for provision of special
      media if culture for other organisms (Corynebacterium
      diphtheria, Neisseria gonorrhoeae) is desired.  Do not obtain
      throat samples if epiglottis is inflamed, as sampling may cause
      serious respiratory obstruction.  Sample the posterior pharynx,
      tonsils, and inflamed areas with a sterile swab.

R. Sterile body fluids (other than CSF) - Disinfect overlying skin with
   iodine or chlorhexidine preparation.  Obtain specimen with needle
   and syringe.  Transfer fluid to anaerobic transport system
   (907464) - cleanse rubber stopper with alcohol; push needle through
   septum and inject fluid on top of agar, sterile screw-cap container
   (907069), or aerobic blood culture bottle (923387). Amniotic and
   culdocentesis fluids should always be transported in an anaerobic
   system. Submit as much fluid as possible.  NEVER submit a swab
   dipped in fluid.  If blood culture bottle is inoculated, submit
   separate aliquot in sterile container for preparation of
   cytocentrifuged Gram stain.  Transport time is less than or equal
   to 15 min.

S. Stool - Submit 10-20 g in sterile container. Transport time is less
   than or equal to 1 hour. Refrigerate if transport is delayed. Stools
   are cultured to isolate bacterial causative agents of diarrheal
   illness; Salmonella, Shigella, Campylobacter, and Shiga toxin
   producing    E. coli. Routine stool culture includes EIA for Shiga
   toxin from E. coli. Cultures for Yersinia are performed by special
   request. Stools for C. difficile toxin detection must be
   transported to the laboratory immediately or refrigerated if
   transport is delayed. Surveillance cultures may be ordered on Bone
   Marrow transplant and other immunocompromised patients to detect
   overgrowth of normal flora by Staph aureus, yeast or a gram
   negative bacillus.

T. Tissue - Submit in anaerobic collection jar (907722) or sterile
   screw-cap container (907069); add drops of sterile saline to keep
   small pieces of tissue moist.  Transport time is less than or equal
   to 15 min.

U. Urine - Collect 1-10 mL of urine in a sterile specimen container
   (907069). Transport urine specimens to the microbiology laboratory
   or refrigerate within 30 minutes. Refrigerated specimens should be
   delivered to the lab as soon as possible, and may be rejected if not
   received within 24 hours of collection.
   a. Midstream clean catch method: Patients should be instructed to
      wash hands prior to collection and offered exam gloves.
      1. Female patients should be instructed to sit on toilet with
         legs apart and spread labia with one hand. First void in
         toilet and then, continuing to void, hold specimen container
         in "midstream" to collect sample.
      2. Male patients should be instructed to retract foreskin if
         uncircumcised. First void in toilet and then, continuing to
         void, hold specimen container in "midstream" to collect
         sample.
   b. Straight catheter: Thoroughly cleanse the urethral opening with
      soap and water. Rinse area with wet gauze pads. Aseptically
      insert catheter into the bladder. After discarding initial 15 to
      30 mL of urine, collect urine for submission in a sterile
      container.
   c. Indwelling catheter: Clamp catheter below port and allow urine to
      collect in tubing. Disinfect the catheter collection port with
      70% alcohol. Use needle and syringe to aseptically collect
      5-10 mL freshly voided urine though catheter port. Transfer to
      sterile container. Do not collect urine from collection bag.
   d. Ileal conduit: Remove the external device and discard urine
      within device. Gently cleanse the stoma with 70% alcohol
      followed by povidone-iodine swab stick (907172). Using sterile
      technique, insert a double catheter into the cleansed stoma, to
      a depth beyond the fascial level, and collect the urine into a
      sterile container. Use of a double catheter helps to minimize
      contamination of the specimen with skin flora.

V. Wound - See abscess.
 
See Additional Information:
Microbiology Specimen Collection and Transport
Normal (Indigenous) Flora of Human Body
Specimens Requiring Immediate Delivery

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Updated: 10/06/2009

Note: The information contained in this handbook is for use by personnel of University of Iowa Health Care. No other use is implied or intended.