Urine culture: Everything you have to know

What is urine culture?

Urine culture is a test that can use to identify possible pathogens that cause urinary tract infections.
In the modern world, females and males both affect urinary tract infections. Females are mostly infecting because their urethra is short, and anus to urethra distance is short. When bacteria that cause UTI enter the urethra, they can move from the ureters to the bladder. At that time patient has to get treatment. If there are no treatments for patients, bacteria will grow in the bladder because urine is concentrate in the bladder. After that, bacteria can move into the kidney and infect the glomerulus. Finally, bacteria enter the blood. That is a difficult situation. So, early diagnosis of Urinary tract infection is essential.

Urine culture is the best test for Urinary pathogen diagnosis. Urine culture test performs using CLED agar.
CLED is a non-selective-differential plating medium. Today, Chrome agar also used to diagnose UTIs, causing pathogens.

What kind of urine samples used for a Urine culture?

  • Mid-stream sample
  • Clean catch sample
  • Suprapubic aspirate
  • Indwelling catheter

Specific features of the Urine collection container

The urine collection container should be sterile. That means Free of bacteria. Screw-capped containers help to prevent leaking urine. Wide mouth container help to the patient because the patient can easily put in urine into the container. The container should be dry.

How to collect samples for urine culture

Correct instruction of patients on how to collect urine is essential

Mid-stream, clean catch sample

Wash hand with soap and water before collecting the urine. After that, the patient should clean their genital area with soap and water. The patient should remove their first portion of urine into the toilet bowl. Collect the mid part of the urine flow into a sterile container after the initial flow of the urine has been passed.

Clean catch sample(pediatric sample)

Following breast/bottle feeding, the baby should be kept without a nappy(Perineal cleaning is recommended). Urine is collected straight from the stream into the container(Preferably midstream). The collection of a satisfactory specimen can be very difficult for babies. Instruct the mother to attempt collection soon after a feed because babies will urinate after feeding.

Suprapubic aspirate

This is an invasive procedure. Aseptic collection of urine directly from the urinary bladder using a needle and syringe. Need to be indicated on the request form.

Indwelling catheter

Catheterization is somewhat difficult for the patient. In that case, the catheter is gone through the urethra to the bladder. Clamp the tube and collect the urine aseptically using a needle and syringe. Insert the needle in head to toe direction.

Sample transportation and storage

  • The sample container should be labeled.
  • The specimen should be transported to the laboratory as soon as possible with the request form.
  • If there is a delay in transport keep the specimens at 4 Celsius
  • Refrigeration should be for less than 24 hours if transport takes more than two hours, transport in ice.
  • If boric acid containers are used(1.8% boric acid), only the amount of urine indicated on the bottle should be collected. Urine samples thus collected can be transported at room temperature.

Samples rejection criteria

  • Specimen kept at room temperature for more than two hours(Bacterial multiplication can occur)
  • Specimens transport without ice if the transport has taken more than two hours. (Bacterial proliferation can occur)
  • The specimen is refrigerated for more than 24 hours(Specimen refrigerated because of stop the bacterial multiplication. If samples refrigerated for more than 24 hours, bacteria will kill.
  • Unlabeled specimens(Specimens should be labeled otherwise patient cannot be identified)
  • Specimens were taken from a receptacle(Receptacle mean catheter bag or bedpan). High contamination can occur because the surface of the plastic bag makes biofilms on the surface.
  • Catheter tips(Catheter tips are not suitable for urine culture because high contamination can occur)
  • Unsterile container (Urine collection container should be a sterile container. Otherwise, contamination can occur as a result mixed growth can be seen in overnight urine culture plate)
  • Leaking specimens (Environmental contamination can occur because of leaking specimens)

Urine culture procedure

Urine culture requirements – Types of equipment

  • Incubator (35 Celsius)
  • Refrigerator (2-8 Celsius)
  • Bunsen burner
  • CLED agar
  • Sterile wire loop 0.001ml (1microleter)

Test procedure

  • CLED agar plate was divided into three sections, and it was labeled.(90mm plate – 4 sections), (140mm plate – 8 sections)
  • Firstly, Mix urine sample well.
  • Sterile one microliter wire loop was inserted vertically into the urine sample. Only a wire loop should be inserted, and the CLED plate was inoculated.
  • It was incubated at 35 Celsius at 16-18 hours(Overnight, ambient air)

Urine culture test results

2nd-day observation

  • Check whether their growth present or not
  • If there is any growth present, count the colony present
  • If you don’t know the colony count procedure – Interpretation and reporting procedure, click here
  • If there is no growth present, urine culture negative
  • When the urine culture positive, we have to identify the possible pathogen.

Possible pathogens could be,

Acute, uncomplicated UTIs
  • Escherichia coli- commonest
  • Proteus mirabilis- associated with renal tract abnormalities, particularly calculi
  • Staphylococcus saprophyticus – in young women
  • Enterococci
  • Group B streptococci
Complicated UTIs
  • E. coli – commonest
  • Klebsiella, enterobacter – usually associated with instrumentation or catheterization
  • Pseudomonas aeruginosa – associated with a structural abnormality or long-term catheterization
  • S. aureus- rarely causes UTI. Associated with a renal abnormality or as a secondary infection due to bacteremia, surgery or catheterization. It is also seen as a contaminant due to perineal carriage.
  • Coagulase-negative staphylococci – may cause complicated infections
  • Candida species – Candida albicans is the most frequently isolated species. Bladder colonization is associated with indwelling catheters but may also be present as contamination from the genital tract.

Possible pathogen identification

Firstly gram stain is done. – Results:- Gram-Negative Bacilli, Gram-positive cocci in clusters (Catalase Positive), Gram-positive cocci in chains(Catalase negative)

How to identify gram-negative bacilli that cause UTIs(Urinary tract infections)

  • Firstly You have to observe and identify the organism is Lactose-fermenter or Non-lactose-fermenter using CLED agar plate

Color change(CLED Agar)

  • Lactose fermenter bacteria – Yellow color
  • Non-lactose-fermenter bacteria – Blue color
  • Coliform – gram-negative bacilli, Lactose fermenter
  • Proteus spp – gram-negative bacilli, Non-lactose-fermenter, Oxidase Negative, Urea positive
  • Pseudomonas spp – gram-negative bacilli, Non-lactose-fermenter, Oxidase Positive
  • Some coliform – gram-negative bacilli, Non-lactose-fermenter, Oxidase Negative, Urea Negative(KIA if suspecting Typhoid)

How to identify Gram-positive cocci in clusters (Catalase Positive) that cause UTIs(Urinary tract infections)

  • Staphylococcus aureus – Gram-positive cocci in clusters (Catalase Positive), Coagulase Positive
  • S. saprophyticus – Gram-positive cocci in clusters (Catalase Positive), Coagulase Negative, Novobiocin Resistant(≤15mm)
  • Coagulase-negative staphylococcus – Gram-positive cocci in clusters (Catalase Positive), Coagulase Negative, Novobiocin Sensitive(>15mm)

How to identify Gram-positive cocci in chains (Catalase Negative) that cause UTIs(Urinary tract infections)

  • Enterococci – Gram-positive cocci in chains (Catalase Negative), Bile aesculin and growth in 6.5% NaCl Positive
  • Others – Gram-positive cocci in chains (Catalase Negative), Streptococcus species. Identify group B streptococcus in pregnant women.



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