- Email: [email protected]
Urine analysis and urine culture: Revisited
Pediatric Infectious Disease 2012 December Volume 4, Number 4; pp. 178e182
Notes from the Lab
Urine analysis and urine culture: Revisited Tanu Singhal
INTRODUCTION Urine routine analysis and urine culture are one of the most commonly performed tests in children. It is extremely important that the tests are performed and interpreted properly so that urinary tract infections in children are neither missed nor over diagnosed. This communication discusses those aspects of urine analysis and culture that are relevant toward the diagnosis of urinary tract infections (UTI).
COLLECTION OF THE URINE SPECIMEN Perhaps the most challenging aspect of urine examination in children is collection of the specimen.1e3 Proper collection is crucial to avoid contamination and false positive results. Children who are older than 2 years and who are toilet trained can be instructed to provide a midstream urine specimen. Studies show that contamination of midstream specimens and false positive diagnosis of UTI can occur if the perineum is not cleaned and the labia not retracted/foreskin not pushed back. Specificity rates approach 100% if urine is collected after proper instruction (cleaning of the genitalia with soap and water and retraction of the labia/pushing back of the foreskin) and if 105/ml CFU and not 104 CFU/ml is used as a cutoff for diagnosing UTI. The options for collection in infants and younger children include suprapubic aspiration, urethral catheterization, bag and clean catch specimens. Suprapubic aspiration (SPA) is the gold standard method for collection of urine. The rate of contamination is 0% and thus specificity for excluding diagnosis of UTI is 100%. The disadvantages include substantial failure rates, invasiveness of the technique, resistance from parents and complications of the procedure. The success rate of the procedure varies from 25 to 98%. The success rate can be
enhanced by use of portable ultrasound or by waiting for 60 min after a baby has passed urine. The main complication is transient microscopic hematuria which is usually of no consequence; macroscopic hematuria has been noted in 0.5e2% and intestinal perforation in 0.2%. Transurethral bladder catheterization is another invasive method for collection of urine which has a reported specificity varying from 80 to 100% in exclusion of UTI when compared to SPA. The specificity is lower if the first few drops are not excluded from collection, if urine is obtained after several attempts, in uncircumcised boys and if 104 CFU/ml is used as a cutoff. Success rates approach 100% especially if portable ultrasound is used but the amount of urine obtained is lower than SPA. Complications include transient hematuria, catheter induced UTI and urethral stricture. The risk of catheter induced UTI has not been quantified but is assumed to be low. While collecting SPA and catheter samples, a container should be kept handy to collect urine which the baby may inadvertently pass. Bag specimens are considered unsuitable for culture but acceptable for urine routine analysis. When compared to SPA, they have a high contamination rate and thus have poor specificity (68%). The false positive rate depends of the prevalence of UTI; the false positive rate is as high as 85% if the pre-test prevalence of UTI is 5%. For febrile boys, with a prevalence of UTI of 2%, the rate of false-positive results is 95%; for circumcised boys, with a prevalence of UTI of 0.2%, the rate of false-positive results is 99%. Bag specimens are however useful for ruling out UTI; a negative culture of a bag specimen almost completely rules out UTI if the patient is not antibiotics. If used, the bag should be applied after proper cleaning of the perineum and removed promptly as soon as the urine is collected. Clean catch collection is the term when urine is collected by parents in children who are unable to void on command. When compared to SPA, sensitivity and specificity of 89%
Consultant Pediatrics and Infectious Disease, Kokilaben Dhirubhai Ambani Hospital and Medical Research Centre, India. email: [email protected] Received: 30.11.2012; Accepted: 7.12.2012; Available online: 20.12.2012 Copyright Ó 2013, Indian Academy of Pediatrics, Infectious Disease Chapter. All rights reserved. http://dx.doi.org/10.1016/j.pid.2012.12.004
Urine analysis and urine culture
Notes from the Lab
179
be processed immediately, it should be refrigerated at 4 C (this will inhibit bacterial multiplication but still cause degeneration of white cells). If transport to another laboratory is necessary, the sample should be transported on ice. Another option for handling delays in processing is to inoculate it at the bedside on dip slides, and keep at room temperature.
and 95% has been reported. The main drawback is the time and patience required. Success rates improve if the urine is collected 3e5 min after a feed and after holding the infant prone on a sterile container and stroking the back. For choice of method see Fig. 1
PROCESSING OF THE URINE SPECIMEN URINE ROUTINE ANALYSIS
The urine specimen should be processed for both routine and culture analysis within 1 h of collection.1 Delay in processing can lead to degeneration of white cells and increase in colony count due to bacterial multiplication. If it cannot
Though urine culture is the gold standard for the diagnosis of UTI, its results are not available for 24e48 h.1e3 Hence
Suspected UTI
Child can void on command
Yes
No
Collect mid stream sample and send for urine routine and culture
Child sick enough to require immediate antimicrobial therapy
Yes
Collect sample by SPA or catheter and send for routine and culture
No
Try to collect clean catch specimen and send for urine routine and culture
Not successful
Collect bag sample and send for urine routine analysis
Suggestive of UTI
Not suggestive of UTI Observation without antimicrobial therapy
Fig. 1 Algorithm for collection of urine specimen for UTI.
180
Pediatric Infectious Disease 2012 December; Vol. 4, No. 4
a routine analysis is important for rapid diagnosis of UTI. A routine urine analysis is also helpful in ruling out UTI in suspected cases especially if it is difficult to collect an appropriate sample for culture. As per current definitions of UTI, a positive urinalysis is also required to diagnose UTI along with significant bacteriuria. Urine collected by any method including bag is suitable for urine routine provided it is processed in less than 1 h if kept at room temperature and less than 4 h if refrigerated. Rapid techniques to predict UTI include urine dipstick tests for leukocyte esterase and nitrites, standard microscopy on a centrifuged specimen, high-powered microscopy with a hemocytometer, and Gram stain of unspun urine for organisms. Metanalysis on the sensitivities/specificities of each of these tests singly or in combination have been published. Table 1 reports the sensitivity and specificity of each of these tests either singly or in combination. The dipstick tests are useful as they are rapid, inexpensive, convenient and can be done at the bedside. Sometimes, they may be the only available tests. The performance of these tests depends on the method used for collection of urine, the definitions used to define positive cultures and the age of the patient. The sensitivity and specificity of the leukocyte esterase test (surrogate for pyuria and correlating well with a cutoff of 5 WBC/hpf) is around 80%; it can be false positive if there is pyuria from any other cause (e.g. Kawasaki disease). The nitrite test based on conversion of dietary nitrate present in urine to nitrite by most gram negative enteric bacteria has a specificity approaching 98%. If positive it almost always indicates a UTI. However, nitrite is frequently false negative (50%). False negative results are either due to infection with organisms that do not reduce nitrate to nitrite or if urine has not remained in the bladder for at least 4 h (since that is the time required to convert nitrate to nitrite). Hence the test is usually negative in infants who empty their bladder frequently. The sensitivity and specificity of either a positive nitrite or LE test for diagnosis of UTI is around 90%. Hence, 10% of children with UTI will
Singhal
have neither of these tests positive. If both LE and nitrite test are positive, the sensitivity improves to 72% (compared to nitrite alone) and specificity improves to 96% (compared to LE alone). Detection of pyuria by microscopy of centrifuged urine is one of the most commonly performed component of routine urine analysis in most labs around the country. A cutoff of 5 WBC/hpf is used to define a positive test; the reported specificity and sensitivity is 80%. False positive results are due to other causes of pyuria such as such as fever due to any cause, dehydration, hypercalciuria, renal stones and rare conditions such as Kawasaki disease. As per results of the metanalysis, 20% of UTI’s will not have pyuria. However there is an ongoing debate about the sensitivity of pyuria for diagnosis of UTI. Technically pyuria may be absent in a very early UTI since the inflammatory response is yet to occur. However, since pyuria denotes an inflammatory response, it should be present when a child has fever due to a UTI. It is currently believed that 80% sensitivity of pyuria, reported in earlier studies was due to inclusion of asymptomatic bacteriuria in study cases and lower sensitivity of microscopy of spun urine. Unspun urine seen in a hemocytometer (presence of more than 10 WBC/mm3 is used as a cutoff) has higher sensitivity as compared to standard microscopy (5 WBC/hpf corresponds to 25 WBC/mm3). Finding any bacteria/10 oil immersion fields on Gram stain of fresh unspun urine is a surrogate for a high colony count of more than 105/mm3 and has sensitivities and specificities of 80e90% for diagnosis of UTI. Enhanced urine analysis (positive cell count or a positive Gram stain/a positive cell count and a positive Gram stain) offers the combination of best sensitivity and specificity for diagnosis of UTI (Table 1). In a nutshell, finding of either a positive dipstick test or presence of more than 5 WBC/hpf of spun urine or presence of bacteria on the Gram stain of an unspun urine is suggestive though not diagnostic of a UTI. The diagnosis needs to be confirmed by urine culture.
Table 1 Sensitivity and specificity of components of urine analysis alone or in combination.1,3 Parameter Leukocyte esterase (LE) test Nitrite test LE or nitrite positive Microscopy WBC 5/hpf of spun urine Cell count (10/mm3) of unspun urine Gram stain (GS) of bacteria in unspun urine Enhanced urine analysis (cell count and GS) Enhanced urine analysis (cell count or GS) LE test or nitrite or microscopy positive
Sensitivity % (range) 83 53 93 73 77 81 85 95 99.8
(67e94) (15e82) (90e100) (32e100) (57e92) (16e99) (75e88) (94e96) (99e100)
Specificity % (range) 78 98 72 81 89 83 99 89 70
(64e92) (90e100) (58e91) (45e98) (37e95) (91e100) (99) (84e93) (60e92)
Urine analysis and urine culture
URINE CULTURE The urine culture is the gold standard for diagnosis for UTI.1,2 Specimen sent for culture should be either clean catch/midstream/SPA/catheter sample. Bag samples are inappropriate. The sample should be inoculated within 1 h of voiding. UTI is confirmed or excluded based on the number of colony-forming units that grow on the culture media. The definition of significant colony count depends on the clinical status of the patient and the method use for collection of urine. For urine collected by clean catch or midstream method, a significant number of bacteria same as those that cause infection will be found colonizing the distal urethra and perineum. Hence a colony count of 105 is used as a cutoff for defining a significant colony count. Earlier guidelines suggested a similar cutoff of 105/ml for significant colony count in catheter samples. The 2011 guidelines from the American Academy of Pediatrics have argued that this colony count cutoff was obtained by analysis of early morning sample of adult women and may be inappropriate for diagnosis of UTI in children below age 2 years.1 This is because the colony count is a determinant of the time the urine resides in the bladder which is lower in infants and children. They propose reduction of this cutoff to 50,000 per ml for defining a significant colony count if urine is collected by catheterization. This will increase the sensitivity but lower the specificity for diagnosis of UTI. However, because the new proposed criteria for UTI now include a positive urine analysis in addition to positive culture results, infants with “positive” culture results alone will be recognized as having asymptomatic bacteriuria rather than a true UTI. These 2011 guidelines do not discuss revising the cutoff for urine obtained by the clean catch/midstream method. Growth of any gram negative bacilli in urine obtained by suprapubic puncture is considered significant (1 colony on plate is equal to 1000 CFU/ml) and growth of gram positive cocci is considered significant if growth is more than a few thousand. Definitions of positive and negative cultures are operational and not absolute. Significance also depends on the identification of the isolated organism as a pathogen. Organisms such as Lactobacillus species, coagulase-negative staphylococci, and Corynebacterium species are not considered clinically relevant urine isolates in the otherwise healthy 2-month to 2-year-old. Isolation of fungi in the urine is usually not significant, it just indicates colonization in patients who are on broad spectrum antibiotics. False negative results can occur with urine culture if the patient is on antibiotics, if there is rapid rate of urine flow with reduced incubation time or if there is obstruction
Notes from the Lab
181
interfering with discharge of urine in the bladder. Demonstration of a significant colony count on urine culture does not equate with a UTI especially in the absence of a negative urine routine analysis. Demonstration of a significant colony count on urine culture does not equate with a UTI especially in the absence of a negative urine routine analysis. Significant colony count can also be attributed to asymptomatic bacteriuria, contamination during collection and delayed processing of the urine sample. In a child with suspected UTI who has a positive urine culture but negative urine analysis, repeat urine routine and culture should be sent. If the repeat urine is also negative on urine analysis, a diagnosis of asymptomatic bacteriuria should be made. Alternative culture methods such as the dipslide may have a place in the office setting; sensitivity is reported in the range of 87%e100%, and specificity, 92%e98%. Dip slides do not identify the organism or give information on antimicrobial sensitivities; hence the isolate will need to be sent to a certified lab for further processing.
ANTIMICROBIAL SENSITIVITY TESTING In the current era of antimicrobial resistance, susceptibility testing of the isolate assumes great importance. Certain studies from India estimate the prevalence of UTI due to extended spectrum beta lactamase (ESBL) producing organisms to amount to 40% of all UTI’s in adults.4 Laboratories should be asked to follow standard guidelines for reporting results of susceptibility tests as resistant, intermediate and sensitive and not report as þ, þþ and þþþ. Equal care should be taken to interpret results of the susceptibility tests. There may be discordance between in vitro and in vivo susceptibility results. In certain cases the isolate is resistant in vitro but there is a clinical response. This is because the breakpoints used to define susceptibility are based on serum levels of antibiotics; however certain antibiotics such as beta lactams, quinolones and aminoglycosides are concentrated in urine and thus may exceed the minimum inhibitory concentration of the organism. If the urine culture is sent after starting antibiotics, we may isolate resistant flora which have been selected out due to concurrent antibiotic use. Conversely, there may be clinical failure with use of certain antibiotics despite in vitro susceptibility. This may occur due to use of antibiotics which do not achieve good urinary levels such as tetracyclines, or due to inadequate tissue levels as it happens with use of nitrofurantoin to treat upper urinary tract infections.
182
Pediatric Infectious Disease 2012 December; Vol. 4, No. 4
Singhal
CONCLUSIONS d
d
d
d
d
A urine analysis should be asked for only in those children where UTI is strongly suspected and when other localizing symptoms for fever such as runny nose etc., are absent. This is to avoid false positive diagnosis (even in an upper respiratory tract infection pyuria will be there and a significant colony count may be obtained due to contamination during collection or asymptomatic bacteriuria). Clean catch, midstream, SPA, catheterization and bags are the commonly used methods for urine collection in infants and children. All these are appropriate for urine routine analysis but bag samples are not recommended for culture. The processing of sample for urine routine/culture should be within 1 h of collection. If there is a delay the sample should be refrigerated. Transport to other labs should be on ice. Urine routine analysis should include dipstick tests, microscopy for pyuria, Gram stain and if possible cell count. The presence of any abnormality in any of these tests is suggestive but not confirmatory of a UTI. Sample should be sent for culture (to exclude false positive tests) and antimicrobial therapy initiated. False negative results on routine analysis are rare. If the routine analysis (including dipstick, microscopy) is negative and child is not sick and appropriate sample for culture cannot be collected, the child can be followed up without initiating antimicrobial therapy. Urine culture confirms diagnosis of UTI and gives information of antimicrobial susceptibility of isolate. As per recent guidelines, a CFU count of 50,000 is defined as significant bacteriuria in children aged 2 months to 2 years for urine collected by catheterization and CFU count of 100,000/ml for those in whom urine is collected by midstream/clean catch technique. Growth
d
d
of any gram negative isolate is considered significant in urine obtained by SPA. The diagnosis of UTI should be based on both a positive urinalysis and significant bacteriuria on culture. If routine analysis is negative and urine culture shows significant bacteriuria, the routine should be repeated. If the second routine analysis is also negative, it indicates asymptomatic bacteriuria and largely excludes UTI as a cause of fever. Results of antimicrobial susceptibility testing should be interpreted carefully.
CONFLICTS OF INTEREST The author has none to declare.
REFERENCES 1. Improvement and Management Subcommittee on Urinary Tract InfectionSteering Committee on Quality. Management of the initial UTI in febrile infants and children 2 to 24 months. Pediatrics. 2011;128:595e610. 2. Committee on Quality ImprovementSubcommittee on Urinary Tract Infection. Practice parameter: the diagnosis, treatment, and evaluation of the initial urinary tract infection in febrile infants and young children. Pediatrics. 1999;103:843e852. 3. Zorc Joseph J, Kiddoo Darcie A, Shaw Kathy N. Diagnosis and management of pediatric urinary tract infections. Clin Microbiol Rev. 2005;18:417e422. 4. Akram Mohammed, Shahid Mohammed, Khan Asad U. Etiology and antibiotic resistance patterns of communityacquired urinary tract infections in J N M C Hospital Aligarh, India. Ann Clin Microbiol Antimicrob. 2007;6:4. http://dx.doi. org/10.1186/1476-0711-6-4.
Notes from the Lab
Urine analysis and urine culture: Revisited Tanu Singhal
INTRODUCTION Urine routine analysis and urine culture are one of the most commonly performed tests in children. It is extremely important that the tests are performed and interpreted properly so that urinary tract infections in children are neither missed nor over diagnosed. This communication discusses those aspects of urine analysis and culture that are relevant toward the diagnosis of urinary tract infections (UTI).
COLLECTION OF THE URINE SPECIMEN Perhaps the most challenging aspect of urine examination in children is collection of the specimen.1e3 Proper collection is crucial to avoid contamination and false positive results. Children who are older than 2 years and who are toilet trained can be instructed to provide a midstream urine specimen. Studies show that contamination of midstream specimens and false positive diagnosis of UTI can occur if the perineum is not cleaned and the labia not retracted/foreskin not pushed back. Specificity rates approach 100% if urine is collected after proper instruction (cleaning of the genitalia with soap and water and retraction of the labia/pushing back of the foreskin) and if 105/ml CFU and not 104 CFU/ml is used as a cutoff for diagnosing UTI. The options for collection in infants and younger children include suprapubic aspiration, urethral catheterization, bag and clean catch specimens. Suprapubic aspiration (SPA) is the gold standard method for collection of urine. The rate of contamination is 0% and thus specificity for excluding diagnosis of UTI is 100%. The disadvantages include substantial failure rates, invasiveness of the technique, resistance from parents and complications of the procedure. The success rate of the procedure varies from 25 to 98%. The success rate can be
enhanced by use of portable ultrasound or by waiting for 60 min after a baby has passed urine. The main complication is transient microscopic hematuria which is usually of no consequence; macroscopic hematuria has been noted in 0.5e2% and intestinal perforation in 0.2%. Transurethral bladder catheterization is another invasive method for collection of urine which has a reported specificity varying from 80 to 100% in exclusion of UTI when compared to SPA. The specificity is lower if the first few drops are not excluded from collection, if urine is obtained after several attempts, in uncircumcised boys and if 104 CFU/ml is used as a cutoff. Success rates approach 100% especially if portable ultrasound is used but the amount of urine obtained is lower than SPA. Complications include transient hematuria, catheter induced UTI and urethral stricture. The risk of catheter induced UTI has not been quantified but is assumed to be low. While collecting SPA and catheter samples, a container should be kept handy to collect urine which the baby may inadvertently pass. Bag specimens are considered unsuitable for culture but acceptable for urine routine analysis. When compared to SPA, they have a high contamination rate and thus have poor specificity (68%). The false positive rate depends of the prevalence of UTI; the false positive rate is as high as 85% if the pre-test prevalence of UTI is 5%. For febrile boys, with a prevalence of UTI of 2%, the rate of false-positive results is 95%; for circumcised boys, with a prevalence of UTI of 0.2%, the rate of false-positive results is 99%. Bag specimens are however useful for ruling out UTI; a negative culture of a bag specimen almost completely rules out UTI if the patient is not antibiotics. If used, the bag should be applied after proper cleaning of the perineum and removed promptly as soon as the urine is collected. Clean catch collection is the term when urine is collected by parents in children who are unable to void on command. When compared to SPA, sensitivity and specificity of 89%
Consultant Pediatrics and Infectious Disease, Kokilaben Dhirubhai Ambani Hospital and Medical Research Centre, India. email: [email protected] Received: 30.11.2012; Accepted: 7.12.2012; Available online: 20.12.2012 Copyright Ó 2013, Indian Academy of Pediatrics, Infectious Disease Chapter. All rights reserved. http://dx.doi.org/10.1016/j.pid.2012.12.004
Urine analysis and urine culture
Notes from the Lab
179
be processed immediately, it should be refrigerated at 4 C (this will inhibit bacterial multiplication but still cause degeneration of white cells). If transport to another laboratory is necessary, the sample should be transported on ice. Another option for handling delays in processing is to inoculate it at the bedside on dip slides, and keep at room temperature.
and 95% has been reported. The main drawback is the time and patience required. Success rates improve if the urine is collected 3e5 min after a feed and after holding the infant prone on a sterile container and stroking the back. For choice of method see Fig. 1
PROCESSING OF THE URINE SPECIMEN URINE ROUTINE ANALYSIS
The urine specimen should be processed for both routine and culture analysis within 1 h of collection.1 Delay in processing can lead to degeneration of white cells and increase in colony count due to bacterial multiplication. If it cannot
Though urine culture is the gold standard for the diagnosis of UTI, its results are not available for 24e48 h.1e3 Hence
Suspected UTI
Child can void on command
Yes
No
Collect mid stream sample and send for urine routine and culture
Child sick enough to require immediate antimicrobial therapy
Yes
Collect sample by SPA or catheter and send for routine and culture
No
Try to collect clean catch specimen and send for urine routine and culture
Not successful
Collect bag sample and send for urine routine analysis
Suggestive of UTI
Not suggestive of UTI Observation without antimicrobial therapy
Fig. 1 Algorithm for collection of urine specimen for UTI.
180
Pediatric Infectious Disease 2012 December; Vol. 4, No. 4
a routine analysis is important for rapid diagnosis of UTI. A routine urine analysis is also helpful in ruling out UTI in suspected cases especially if it is difficult to collect an appropriate sample for culture. As per current definitions of UTI, a positive urinalysis is also required to diagnose UTI along with significant bacteriuria. Urine collected by any method including bag is suitable for urine routine provided it is processed in less than 1 h if kept at room temperature and less than 4 h if refrigerated. Rapid techniques to predict UTI include urine dipstick tests for leukocyte esterase and nitrites, standard microscopy on a centrifuged specimen, high-powered microscopy with a hemocytometer, and Gram stain of unspun urine for organisms. Metanalysis on the sensitivities/specificities of each of these tests singly or in combination have been published. Table 1 reports the sensitivity and specificity of each of these tests either singly or in combination. The dipstick tests are useful as they are rapid, inexpensive, convenient and can be done at the bedside. Sometimes, they may be the only available tests. The performance of these tests depends on the method used for collection of urine, the definitions used to define positive cultures and the age of the patient. The sensitivity and specificity of the leukocyte esterase test (surrogate for pyuria and correlating well with a cutoff of 5 WBC/hpf) is around 80%; it can be false positive if there is pyuria from any other cause (e.g. Kawasaki disease). The nitrite test based on conversion of dietary nitrate present in urine to nitrite by most gram negative enteric bacteria has a specificity approaching 98%. If positive it almost always indicates a UTI. However, nitrite is frequently false negative (50%). False negative results are either due to infection with organisms that do not reduce nitrate to nitrite or if urine has not remained in the bladder for at least 4 h (since that is the time required to convert nitrate to nitrite). Hence the test is usually negative in infants who empty their bladder frequently. The sensitivity and specificity of either a positive nitrite or LE test for diagnosis of UTI is around 90%. Hence, 10% of children with UTI will
Singhal
have neither of these tests positive. If both LE and nitrite test are positive, the sensitivity improves to 72% (compared to nitrite alone) and specificity improves to 96% (compared to LE alone). Detection of pyuria by microscopy of centrifuged urine is one of the most commonly performed component of routine urine analysis in most labs around the country. A cutoff of 5 WBC/hpf is used to define a positive test; the reported specificity and sensitivity is 80%. False positive results are due to other causes of pyuria such as such as fever due to any cause, dehydration, hypercalciuria, renal stones and rare conditions such as Kawasaki disease. As per results of the metanalysis, 20% of UTI’s will not have pyuria. However there is an ongoing debate about the sensitivity of pyuria for diagnosis of UTI. Technically pyuria may be absent in a very early UTI since the inflammatory response is yet to occur. However, since pyuria denotes an inflammatory response, it should be present when a child has fever due to a UTI. It is currently believed that 80% sensitivity of pyuria, reported in earlier studies was due to inclusion of asymptomatic bacteriuria in study cases and lower sensitivity of microscopy of spun urine. Unspun urine seen in a hemocytometer (presence of more than 10 WBC/mm3 is used as a cutoff) has higher sensitivity as compared to standard microscopy (5 WBC/hpf corresponds to 25 WBC/mm3). Finding any bacteria/10 oil immersion fields on Gram stain of fresh unspun urine is a surrogate for a high colony count of more than 105/mm3 and has sensitivities and specificities of 80e90% for diagnosis of UTI. Enhanced urine analysis (positive cell count or a positive Gram stain/a positive cell count and a positive Gram stain) offers the combination of best sensitivity and specificity for diagnosis of UTI (Table 1). In a nutshell, finding of either a positive dipstick test or presence of more than 5 WBC/hpf of spun urine or presence of bacteria on the Gram stain of an unspun urine is suggestive though not diagnostic of a UTI. The diagnosis needs to be confirmed by urine culture.
Table 1 Sensitivity and specificity of components of urine analysis alone or in combination.1,3 Parameter Leukocyte esterase (LE) test Nitrite test LE or nitrite positive Microscopy WBC 5/hpf of spun urine Cell count (10/mm3) of unspun urine Gram stain (GS) of bacteria in unspun urine Enhanced urine analysis (cell count and GS) Enhanced urine analysis (cell count or GS) LE test or nitrite or microscopy positive
Sensitivity % (range) 83 53 93 73 77 81 85 95 99.8
(67e94) (15e82) (90e100) (32e100) (57e92) (16e99) (75e88) (94e96) (99e100)
Specificity % (range) 78 98 72 81 89 83 99 89 70
(64e92) (90e100) (58e91) (45e98) (37e95) (91e100) (99) (84e93) (60e92)
Urine analysis and urine culture
URINE CULTURE The urine culture is the gold standard for diagnosis for UTI.1,2 Specimen sent for culture should be either clean catch/midstream/SPA/catheter sample. Bag samples are inappropriate. The sample should be inoculated within 1 h of voiding. UTI is confirmed or excluded based on the number of colony-forming units that grow on the culture media. The definition of significant colony count depends on the clinical status of the patient and the method use for collection of urine. For urine collected by clean catch or midstream method, a significant number of bacteria same as those that cause infection will be found colonizing the distal urethra and perineum. Hence a colony count of 105 is used as a cutoff for defining a significant colony count. Earlier guidelines suggested a similar cutoff of 105/ml for significant colony count in catheter samples. The 2011 guidelines from the American Academy of Pediatrics have argued that this colony count cutoff was obtained by analysis of early morning sample of adult women and may be inappropriate for diagnosis of UTI in children below age 2 years.1 This is because the colony count is a determinant of the time the urine resides in the bladder which is lower in infants and children. They propose reduction of this cutoff to 50,000 per ml for defining a significant colony count if urine is collected by catheterization. This will increase the sensitivity but lower the specificity for diagnosis of UTI. However, because the new proposed criteria for UTI now include a positive urine analysis in addition to positive culture results, infants with “positive” culture results alone will be recognized as having asymptomatic bacteriuria rather than a true UTI. These 2011 guidelines do not discuss revising the cutoff for urine obtained by the clean catch/midstream method. Growth of any gram negative bacilli in urine obtained by suprapubic puncture is considered significant (1 colony on plate is equal to 1000 CFU/ml) and growth of gram positive cocci is considered significant if growth is more than a few thousand. Definitions of positive and negative cultures are operational and not absolute. Significance also depends on the identification of the isolated organism as a pathogen. Organisms such as Lactobacillus species, coagulase-negative staphylococci, and Corynebacterium species are not considered clinically relevant urine isolates in the otherwise healthy 2-month to 2-year-old. Isolation of fungi in the urine is usually not significant, it just indicates colonization in patients who are on broad spectrum antibiotics. False negative results can occur with urine culture if the patient is on antibiotics, if there is rapid rate of urine flow with reduced incubation time or if there is obstruction
Notes from the Lab
181
interfering with discharge of urine in the bladder. Demonstration of a significant colony count on urine culture does not equate with a UTI especially in the absence of a negative urine routine analysis. Demonstration of a significant colony count on urine culture does not equate with a UTI especially in the absence of a negative urine routine analysis. Significant colony count can also be attributed to asymptomatic bacteriuria, contamination during collection and delayed processing of the urine sample. In a child with suspected UTI who has a positive urine culture but negative urine analysis, repeat urine routine and culture should be sent. If the repeat urine is also negative on urine analysis, a diagnosis of asymptomatic bacteriuria should be made. Alternative culture methods such as the dipslide may have a place in the office setting; sensitivity is reported in the range of 87%e100%, and specificity, 92%e98%. Dip slides do not identify the organism or give information on antimicrobial sensitivities; hence the isolate will need to be sent to a certified lab for further processing.
ANTIMICROBIAL SENSITIVITY TESTING In the current era of antimicrobial resistance, susceptibility testing of the isolate assumes great importance. Certain studies from India estimate the prevalence of UTI due to extended spectrum beta lactamase (ESBL) producing organisms to amount to 40% of all UTI’s in adults.4 Laboratories should be asked to follow standard guidelines for reporting results of susceptibility tests as resistant, intermediate and sensitive and not report as þ, þþ and þþþ. Equal care should be taken to interpret results of the susceptibility tests. There may be discordance between in vitro and in vivo susceptibility results. In certain cases the isolate is resistant in vitro but there is a clinical response. This is because the breakpoints used to define susceptibility are based on serum levels of antibiotics; however certain antibiotics such as beta lactams, quinolones and aminoglycosides are concentrated in urine and thus may exceed the minimum inhibitory concentration of the organism. If the urine culture is sent after starting antibiotics, we may isolate resistant flora which have been selected out due to concurrent antibiotic use. Conversely, there may be clinical failure with use of certain antibiotics despite in vitro susceptibility. This may occur due to use of antibiotics which do not achieve good urinary levels such as tetracyclines, or due to inadequate tissue levels as it happens with use of nitrofurantoin to treat upper urinary tract infections.
182
Pediatric Infectious Disease 2012 December; Vol. 4, No. 4
Singhal
CONCLUSIONS d
d
d
d
d
A urine analysis should be asked for only in those children where UTI is strongly suspected and when other localizing symptoms for fever such as runny nose etc., are absent. This is to avoid false positive diagnosis (even in an upper respiratory tract infection pyuria will be there and a significant colony count may be obtained due to contamination during collection or asymptomatic bacteriuria). Clean catch, midstream, SPA, catheterization and bags are the commonly used methods for urine collection in infants and children. All these are appropriate for urine routine analysis but bag samples are not recommended for culture. The processing of sample for urine routine/culture should be within 1 h of collection. If there is a delay the sample should be refrigerated. Transport to other labs should be on ice. Urine routine analysis should include dipstick tests, microscopy for pyuria, Gram stain and if possible cell count. The presence of any abnormality in any of these tests is suggestive but not confirmatory of a UTI. Sample should be sent for culture (to exclude false positive tests) and antimicrobial therapy initiated. False negative results on routine analysis are rare. If the routine analysis (including dipstick, microscopy) is negative and child is not sick and appropriate sample for culture cannot be collected, the child can be followed up without initiating antimicrobial therapy. Urine culture confirms diagnosis of UTI and gives information of antimicrobial susceptibility of isolate. As per recent guidelines, a CFU count of 50,000 is defined as significant bacteriuria in children aged 2 months to 2 years for urine collected by catheterization and CFU count of 100,000/ml for those in whom urine is collected by midstream/clean catch technique. Growth
d
d
of any gram negative isolate is considered significant in urine obtained by SPA. The diagnosis of UTI should be based on both a positive urinalysis and significant bacteriuria on culture. If routine analysis is negative and urine culture shows significant bacteriuria, the routine should be repeated. If the second routine analysis is also negative, it indicates asymptomatic bacteriuria and largely excludes UTI as a cause of fever. Results of antimicrobial susceptibility testing should be interpreted carefully.
CONFLICTS OF INTEREST The author has none to declare.
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