posted on 14 Feb 2022 | Author DR. JAVEED KAKROO
Urinary tract infections (UTIs) include infections limited to the bladder (cystitis), which are extremely common in women and can cause pain when urinating, as well as more serious infections that also affect the kidneys (pyelonephritis). If you are a clinician, you are likely familiar with the process of requesting urine specimens from patients with UTI symptoms, and also familiar with receiving and acting on results. But what exactly happens to that urine, and the organisms that may grow from it, between the time it leaves the bladder and the time the report appears in the medical record?
From bladder to cup to bench
One of the most important variables in the urine culture process is the collection method. The bladder itself is generally considered a sterile environment (although, as we will see later, this is not always the case), but the external genitalia are colonized by commensal bacteria which can contaminate blood samples. urine and eventually grow in culture. A suprapubic aspiration, in which a needle is inserted directly through carefully cleaned skin into the bladder, is the most effective way to avoid the risk of urogenital contamination, but this method is relatively invasive and rarely used. For infants, young children, and other people who are unable to urinate directly into a specimen container (for example, people with neurogenic bladder), urine can be collected using a Foley catheter, which is inserted through the urethra into the bladder; this method also limits contamination.
Older children and adults who are able to do this can simply provide a “flushed” urine sample: that is, they urinate into a cup. The mid-stream clean capture approach is recommended for discharged urine specimens to reduce the likelihood of contamination. However, there is no way to entirely prevent the possibility of contamination, and recent evidence suggests that cleaning and using a mid-stream sample does not reduce contamination at all. (In contrast, the much-maligned “urine bag” method of collection sometimes used in infants, in which urine is collected in a plastic bag taped to the perineal region, may not be as prone to contamination clinically significant than is generally assumed). As we will see, the relative likelihood of contamination with different specimen collection methods becomes important in the clinical interpretation of urine culture results.
Once a urine sample has been collected, it must be transported to the laboratory. Since bacterial quantity is an important factor in evaluating the potential clinical significance of any organism present in the specimen, it is important to limit bacterial growth between the time of specimen collection and plating. for culture. If the urine specimen is stored at room temperature, it should be smeared within 2 hours of collection. The time between collection and inoculation can be extended to 24 hours if the sample is stored in the refrigerator or transported in a container containing boric acid as a preservative.
The preview: urinalysis
It usually takes about a day for the bacteria in a urine sample to grow to a sufficient quantity to be detected and identified using standard clinical microbiology laboratory techniques, and therefore it takes also at least this time to determine that the bacteria are not present in the culture. However, valuable information about the likelihood of a UTI can be obtained quickly through urinalysis. White blood cells in the urine, which reflect the typical inflammation of an infection, can be detected and quantified by urinalysis. The presence of ≥ 10 white blood cells per μL (or > 5 per high power field) is almost always seen in people with a UTI. A urinalysis can also test for the presence of nitrites, which are produced by gram-negative bacterial species capable of reducing nitrates to nitrites; these species include Escherichia coli, the most common cause of UTIs. A point-of-care urine dipstick can provide preliminary information about these tests in minutes, while a microscopic urinalysis provides more quantitative and sensitive results. Urinalysis results also provide information about other parameters in the urine, including pH and the presence of red blood cells, proteins, and other materials that can be indications of a variety of kidney disease unrelated to infection.
If urinalysis of a person with symptoms of a UTI confirms a probable UTI, a doctor may start empiric antibacterial therapy based on the most likely causative organisms while awaiting culture results to tailor the therapy. On the other hand, a normal urinalysis suggests that a UTI is less likely to be the cause of the symptoms. A number of diagnostic stewardship programs have evaluated the implementation of “reflex” urine culture protocols, in which a culture is only performed if urinalysis is suggestive of a UTI. Studies of these approaches indicate that they can be effective in safely reducing unnecessary antibiotic consumption. It is also important to note that asymptomatic bacteriuria, or the presence of bacteria in the urine of a person who does not have symptoms of a UTI, does not require treatment in most cases (pregnant women are an exception), so urine cultures generally should not be obtained. in people without symptoms of UTI.
The main characteristic: the culture
Once the urine sample reaches the clinical microbiology laboratory, it is usually plated on 2 types of media: a MacConkey agar plate, which inhibits the growth of gram-positive bacteria and also allows early predictions about the identity of gram-negative bacteria, and blood agar, which allows the growth of a lactose-fermenting organism, such as Escherichia coli, on a biplate of urine with MacConkey agar (L) and blood agar from sheep (R).
From almost all bacteria that cause urinary tract infections. The vast majority of UTIs are caused by gram-negative bacteria, most commonly E. coli, which grows as pink colonies on MacConkey agar due to its ability to ferment lactose. Other Enterobacteriaceae, such as Klebsiella and Proteus species, can also cause UTIs, as can a few types of Gram-positive bacteria, including Enterococcus species and Staphylococcus saprophyticus. Urine biplates, in which each of the 2 types of agar fill half of the plate, allow for more efficient plating. Plates are incubated at 35-37°C and examined at 8 p.m. and, if there is no growth at this point, can be incubated for an additional day and re-examined. Urine cultures are streaked quantitatively, using a calibrated inoculation loop that collects 1 or 10 μL of urine; when colonies grow on the agar, the number of colony forming units per milliliter (CFU/mL) can be calculated by multiplying by 1000 or 100, respectively.
Quantification of bacteria in urine cultures is essential, especially for voided specimens because, as noted above, contamination of urine specimens with urogenital flora is common. The identity of all growing organisms, the amount in which they grow, and the types of specimens are all considered when interpreting culture results. Clinical microbiology labs use detailed algorithms to determine which bacteria are reported to the clinician and how they are described in the report. The full reporting algorithms are complex and vary to some degree from lab to lab, but some principles are common to all:
(1) When only 1 or 2 types of bacteria grow and are present in large quantities (i.e. ≥ 10,000 CFU/mL), they are almost always identified to the species level and reported as such .
(2) When 3 or more types of bacteria grow and none predominates (i.e. none present at >100,000 CFU/mL), results may be reported as a “ mixed bacterial flora”.
(3) When bacteria are present in smaller amounts (i.e.
The goal of these algorithms is to ensure that disease-causing bacteria are accurately flagged so that patients can be treated, while avoiding unnecessary flagging of bacteria that are very unlikely to cause UTI in order to avoid excessive use of antibiotics. Of course, there may be certain clinical situations in which it might be appropriate to further evaluate a mixed culture or a culture with an organism that is not generally a uropathogen (for example, if a patient has urinary material indwelling), so, as always, clinicians can call the microbiology lab to ask for more details about exactly what grew on a particular plaque or to request that additional workup be done in specific cases.
The relative ease of obtaining a urine sample and the rapid growth of most uropathogens in culture means that UTIs are often a seemingly straightforward diagnosis. However, interpreting cultures from a sample that must pass through the dense microbiota of the urogenital region before reaching the sample container requires careful work in the clinical microbiology laboratory, where medical laboratory professionals, using together their experience in colony recognition with detailed algorithms, must balance the need for diagnosis with the risk of too much (clinically irrelevant) information.
(The author is Certified Microbiologist Infection Control Auditor, Kidney Hospital Srinagar. firstname.lastname@example.org)