2. INTRODUCTION
Urinary tract infection (UTI) is the most common type of hospital-
acquired or nosocomial infection. Most hospital-acquired UTIs happens
after Urinary Catheterization which is the placement of a catheter
through the urethra into the urinary bladder.
A catheter can act as nidus for bacteria from the urethra and allow them
into the bladder causing an infection to start.
Urine culture helps to determine the choice of treatment for the patient.
As soon as the sample is collected, its processed within 1-2 hrs.
The stability of urine specimens submitted for culture remains a challenge
for many laboratories because of delays in specimen transport.
3. If the specimen is not processed in two hours the microbial count
increases. By adding preservative in the specimen, the transport,
testing and storage of the specimen increases upto 24 hours at room
temprature. The shelf life of sample can be increased using
preservatives.
The urinalysis preservative is intended to inhibit the metabolism of
bacteria while maintaining their cellular integrity.
Without the presence of a preservative, the bacteria continue to
metabolize and reproduce, causing changes in the urine components.
4. To Investigate the usefulness of the BD Vacutainer Plus Urine C&S
Preservative Tube (BDU; Becton Dickinson, Franklin Lakes, NJ) and
compare its performance using urine specimens with and without
preservatives.
This study is designed to assess the need of transport tubes with
preservative for urine culture as it will enable us to compare the
presence of micro-organisms and their colony count in both the culture
system ie with and without preservative
To collect urine from catheterized patient suspected to have CAUTI in
container with and without preservative and process the both
simultaneously.
5. To perform routine examination of urine, colony counts in culture,
identification and sensitivity for all pathogen in urine as per standard
protocol.
To compare colony count of bacteria in urine from containers with
preservative and without preservatives.
To monitor time required in processing urine for culture.
6. The current guidelines for urine collection, transport, and culture
emphasize the need for using either transport tubes containing
preservatives or the need for not exceeding the 2-hour interval from
collection to processing.
If preservative tubes are not used and a transport time of less than 2
hours may not be achievable, refrigeration of the urine specimen has
been shown to also limit the overgrowth of organisms however it is
unrealistic to expect that no urine specimen will spend more than 2
cumulative hours unrefrigerated in most settings.
In the 2005 College of American Pathologists (CAP) Q-Probes study on
urine culture contamination, the investigators found that only a small
number of microbiology laboratories enforce the 2-hour cutoff rule for
limiting transport time of urine specimens.
7. Hence we decided to investigate the usefulness of the BD Vacutainer
Plus Urine C&S Preservative Tube (BDU; Becton Dickinson, Franklin
Lakes, NJ) and compare its performance using urine specimens with
and without preservatives.
This study is designed to assess the need of transport tubes with
preservative for urine culture as it will enable us to compare the
presence of micro-organisms and their colony count in both the culture
system ie with and without preservative
8. The preservatives like Sodium propionate, Ethyl paraben,
Chlorohexidine may help to maintain an acidic pH and prevent
bacterial proliferation.
The role of each chemical is as follows:
Ethyl paraben 5.6% is an antifungal preservative.
Sodium propionate 94% is an mold inhibitor.
Cholrohexidine 0.4% is antibacterial and so used in low concentration
because it has bacteriostatic effect.
9. Study Design: Prospective
Setting : Tertiary care: Superspeciality Hospital
Study Period : One month (January 2015)
Sample Size: 50 (random catheterized urine samples from patients
suspected of UTI)
Ethics permission: Exempt from Ethical review Board
A Total of 50 urine sample specimens were included in this study.
The catheterized urine sample was collected using standard
precautions with syringe in a sterile screw capped wide mouth
container and the urine container with preservative (BD Urinalysis
plus tubes) at the point of collection and transported to the laboratory
and into another urine container without preservative.
METHODOLOGY
10. The patient particulars, the time of collection of urine specimen and the
time of receipt in the laboratory were noted in the case record form.
The plates were incubated at 37 °C in ambient atmosphere and were read
after 18-24 hours of incubation.
The CFU were noted and the bacteria were identified using standard
microbiological procedures.
The antimicrobial susceptibility test was performed using automated
system (Vitek compact 2).
Reading are taken at zero hour and after 24 hr for both with and without
preservative and the urine specimen was processed according to standard
microbiology procedures.
The urine specimen was plated on blood agar ( T- method for semi
quantitative count) and Mac Conkey’s agar.
11. The Incubated agar plates are taken to count ,The colonies and to
study the colony characteristics of isolated colonies. It is also used to
interpret the bacterial load.
-If UTI is present, then colony forming unit decides antibiotic therapy
is required or not depending on the load of bacteria present.
-If CFU/ml is insignificant then we do not provide antibiotic therapy to
the patients.
Colony forming unit
Loop diameter= 4.4mm = 0.01ml
1 colony = 100 CFU/ml =10^2 CFU/ml
10 colony = 1000 CFU/ml =10^3 CFU/ml
100 colony= 10000 CFU/ml =10^4 CFU/ml
1000 colony= 100000 CFU/ml =10^5 CFU/ml
COLONY FORMING UNIT
12. Fig 1a- Sterile wide mouth container for urine specimen collection
containing urine without preservative
PICTURES
19. A Total of 50 urine samples were included in the study. 20 urine
samples were obtained from female patients (40%) and 30 urine
samples from male patients (60%). Patients ranged in age from 10 to
90 years, with an average of 51-60 years.
The average time between the collection of the urine specimen and its
arrival in the laboratory was 30 minutes (Range , 0 – 1 hrs). Out of 50
samples, 15 samples had no growth and 35 samples showed the growth
of various organisms.
The readings obtained were similar for urine sample with and
without preservative after processing within 2 hours and after 24
hours. Increase colony counts of organism were observed for non-
preserved urine sample and these changes were statistically significant.
There were no difference in urine samples containing preservatives
after 24 hours.
RESULTS
20. 33% patients range in the age from 51-60 years. 35 samples exhibit growth
of following Gram negative organisms ie Pseudomonas aeruginosa (n=10),
Pseudomonas putida (n=1), Escherichia coli(n=7), Klebsiella pneumonia
(n=9), Proteus mirabilis (n=1), Providencia stuartii (n=1), Enterobacter
cloacae (n=1), Enterococcus faecalis (n=2) after 24 hours study period. In
addition, 1 culture was positive for yeast ie Candida albicans and 2 cultures
were positive for Gram positive bacteria ie Enterococcus faecium (n=1).
All organism were identified using standard microbiology techniques using
automated identification system.
No statistically significant difference was seen in the number of cultures
with more than or equal to104 pathogens but less than 105 CFU/ml between
the time interval of within 2 hours and after 24 hours.
The sample of urine containing preservative the number of cultures having
105 CFU/ml pathogens in non-preservative urine sample that were stored at
room temperature changes significantly by 24 hours after collection with
the numbers of sample having CFU count more than 105CFU/ml.
26. URINE CULTURE
SYSTEM WITHOUT
PRESERVATIVES
URINE CULTURE
SYSTEM WITH
PRESERVATIVES
Time interval Avg. Growth detected
CFU/ml
Avg Growth detected CFU/ml
Within 2 hours 104 104
After 24 hours > 105 104
Difference >101 0
27. The CAP Q-Probes study lead us to question the accuracy of the
assumption that patients’ urine samples are kept at 4°C (refrigeration)
after processing in the laboratory. It is therefore unlikely that urine
specimens will be continuously refrigerated after culture setup in
routine clinical and laboratory settings.
The results of this study demonstrate that bacterial growth in urine
samples without preservative will significantly change even after a
short period of exposure to ambient/room temperature during
specimen transport.
Considering these constraints of everyday clinical and laboratory
practice, the use of some form of preservative, such as the Sodium
propionate, Ethyl paraben, Chlorohexidine, as an additive to urine
collection tubes is highly desirable and should perhaps be even
mandatory for specimens sent from a distant specimen collection site.
DISCUSSION
28. Most studies, including the present study, demonstrate that the
majority of urine specimens sent for microbiologic examination and
culture will be negative for growth, show some growth of mixed
urogenital flora, or at best are positive for a uropathogen at
significantly less than 104 CFU/m.
If such nonpreserved urine specimens were to present, however, with
higher colony counts because of delays in specimen transport and
processing and if the false-positive rate could be in excess of 15% as
suggested by the CAP Q-Probes study, reporting of these urine culture
results would have significant implications on patient care, antibiotic
use, and other economic health care–related cost.
29. The BD tubes with preservative is an effective approach for
maintaining the qualitative as well as semi quantitative
assessment of urine specimens for culture for up to 24 hours
after specimen collection.
The results of the preserved urine samples were equal as those
using the approach of refrigeration.
Therefore the use of urine collection and transport tubes
containing a preservative such as Sodium propionate, Ethyl
paraben, Chlorohexidine is highly recommended and should
perhaps be mandatory when specimens are sent from a distant
collection site or routinely exceed a 2-hour specimen transport
time.
CONCLUSION
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