Rajiv Gandhi University of Health

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, KARNATAKA, BANGALORE-41

PROFORMA FOR REGISTRATION OF SUBJECT FOR DISSERTATION

NAME OF CANDIDATE LEWAKA PLAIN

ADDRESS LEWAKA PLAIN MSC. MLT STUDENT (MICROBIOLOGY) ST.JOHN’S MEDICAL COLLEGE HOSPITAL BANGALORE-34 COURSE OF STUDY MSC. MLT

SUBJECT MICROBIOLOGY

DATE OF ADDMISSION TO 04-10-2008 COURSE

Title: - Effects Of Boric Acid On Colony Yield In Urine Specimens Need For Study:-

 Urine is a good culture medium. Delay in delivery to the laboratory may lead to an increase in bacterial content from an initial number of no significance to a total which may be taken as significant.¹

 Collection and preservative of urine for analytical testing must follow a carefully prescribed procedure to ensure valid results. A Clean-Catch, midstream is most desirable for bacteriology examination and suprapubic aspiration is used to obtained anaerobic organism.¹⁰

 The main objective for adding preservative in urine is to preserve the bacterial count. Stable bacterial counts in urine specimens before culture are necessary to assure the accurate diagnosis of U.T.I.³These preservatives maintain the original colony count during ambient temperature transport until culture can be done. Preservation by refrigeration is the best method but cannot be used for the purpose of isolating mycobacteria or fungi. It is needed to observe if these preservatives interfere with the colony yield in urine culture.

 The different preservatives of urine are sodium fluoride, thymol, hydrochloric acid, boric acid-glycerol-sodium formate, lyophilized preservatives, boric acid, etc. Boric acid is selected as a preservative for the study because preservatives, other than boric acid, are found to have interfered in many of the chemical test done for urine. They cannot be used for fungi and mycobacteria culture. From the previous studies done, boric acid gives satisfactory preservation compare to other preservatives¹. It is also non toxic.⁵ It is cost effective, bacteriostatic and can be used as a preservative for fungi and mycobacteria. Review Of Literature:-

 The different preservatives used in microbiology for urine culture are:-

1. Sodium Fluoride : Cannot be used for culture of yeast¹⁰

2. Fluid preservatives : Contains boric acid-glycerol-sodium formate.²

3. Transport tubes : Sage urine culture tube and Becton-Dickenson urine culture kit.³

4. Lyophilized preservatives

5. Boric acid: Most abundantly used preservative for urine.

 For culture, sterile containers should be used for collection of the urine sample.⁹

 Preservatives like Thymol and HCl cannot be used for culture of urine specimen.⁸

 Brian .A. Lauer, L. Barth Reller and Stanley Mirett found that the Becton- Dickinson urine culture kit maintains a stable bacterial population in urine for up to 24 hours and is as reliably as refrigeration.²

 Melvin .P. Weinstein found that when liquid preservative is used, the number of bacteria in urine may be lower than that of the number in initial plating time after 18 to 24 hours. With lyophilized preservative, though it allows apparent growth of micro-organism during the 24 hours holding period, it is much promising as a preservative.⁴  Kim .K. Mickander, Carol .J. Shanholtzer and Lance .R.R Peterson found that the Becton- Dickinson urine culture kit is toxic to E.Coli and Klebsiella pneumonia in 2ml of urine, and the sage urine culture tube maintained the number of bacteria in 1 to 4.5ml of urine but is toxic to E.Coli when held for 24 hours.³

 I.A. Porter and J. Brodie on their study of Boric acid preservation of urine samples by comparing the culture yield of urine sample transported over a distance by the dip-inoculum transport medium (D.I.T.M), ice-box and boric acid preservation with “natural” urine specimens, found that boric acid when used in a concentration of 1.8% gives satisfactory preservation.¹

 When P.D. Meers and C.K. Chow did a study on bacteriostatic and bactericidal actions of boric acid against bacteria and fungi commonly found in urine, they found that at concentrations between 10 and 20g/l, boric acid was bacteriostatic or fungistatic. At 10g/l, it was found to be weakly bactericidal for some strains of Acinetobacter Calcoaceticus and Pseudomonas aeruginosa. They concluded that boric acid is rarely toxic, and when it is, the effect is usually sufficiently delayed to be only theoretical importance.⁵

 Boric acid inhibits some enterococci and Pseudomonas strains but preserving urine using boric acid is less expensive and urine can be examined microscopically. The amount of boric acid added for urine is 0.1g/10 ml of urine.⁶ Objectives OF Study:-  To standardize the percentage (%) of boric acid as a preservative on known urine pathogens.  To compare the effect of boric acid on the colony count in urine specimens for culture with that of the colony count on refrigerated urine sample. Materials And Methods:-  Using boric acid as the preservative in the urine specimen received for culture at diagnostic laboratory department of microbiology during the period of 2009-2010 at St. John’s Medical College Hospital, Bangalore, will be studied.

 Standardization of percentage (%) of boric acid is done on known urine pathogens, with the different concentration of boric acid, starting at 0, 2.5,5,10,20 and 40 g/l.

 The comparison between boric acid and refrigerated sample is done by comparing the colony count obtained after inoculating the samples (with and without boric acid) on receiving, with that of overnight refrigerated sample and overnight sample with boric acid kept at room temperature. Methods Of Collection And Data:-  Collection and processing of samples will be done according to the recommended procedure by CLSI guidelines. Inclusion Criteria:-  Urine culture done from urine specimens having boric acid as the preservative.  Urine culture with refrigerated urine specimens. Exclusion Criteria:-  Preservatives other than boric acid.

 Unrefrigerated urine sample without preservative.

References:- 1. I.A. Porter, M.D, M.C. Pathology; J. Brodie, M.D, D.P.H, F.C. Pathology. Boric Acid Preservation of Urine samples. British Medical Journal 1969,2,353-355.

2. Brian .A. Lauer, L. Barth Reller and Stanley Mirrett. Evaluation of Preservative fluid for urine collected for culture. Journal of Clinical Microbiology, July 1979, p.42-45.

3. Kim .K. Nikander, Carol .J. Shanholtzer and Lance .R. Peterson. Urine Culture Transport Tubes: Effect of sample volume on a Bacterial Toxicity of the preservative. Journal of Clinical Microbiology. April 1982. p.593-595

4. Melvin .P. Weinstein. Evaluation of Liquid and Lyophilized Preservative for urine culture. Journal of Clinical Microbiology. October 1983. p.912-916.

5. P.D. Meers, C.K. Chow. Bacteriostatic and Bactericidal actions of boric acid against bacteria and fungi commonly found in urine. Journal of Clinical Pathology 1990; 43; 484-487.

6. Monica Cheesbrough. District Laboratory practice in Tropical Countries– Part 2. 2002, p.106.

7. Connie .R. Mahon and George Manuselis. Textbook of Diagnostic Microbiology. Second Edition. 2000.p.1024 8. Monica Cheesbrough. District Laboratory practice in Tropical Countries– Part 1. 1999, p.316.

9. Ramnik Sood. Text book of medical Laboratory Technology. First edition. 2006, p.51.

10.Richard .A. Mc Pheison, Matthew .R. Pinius. Henry’s Clinical diagnosis and management by laboratory methods. 21 edition. 2007, p.27. Signature Of The Candidate:

REMARKS OF THE GUIDE

NAME AND DESIGNATION OF (IN BLOCK LETTERS)

Guide DR. RAM KUMAR

LECTURER,

DEPT. OF MICROBIOLOGY,

ST. JOHN’S MEDICAL

COLLEGE,

BANGALORE-560034

Signature

Head Of Department DR. H. SRINIVASA

PROFESSOR AND HEAD

DEPT. OF MICROBIOLOGY,

ST. JOHN’S MEDICAL

COLLEGE,

BANGALORE-560034

Signature REMARKS OF THE

Chairman And Principal

Signature