Standardization of an efficient technique for the reporting of CFU / mL in urine samples with bacterial cell cytolysis

Abstract

Urinary tract infections (UTIs) are common in women due to the anatomy of the urethra and its proximity to the anus.1 Bacterial cell cytolysis (BCC) describes the entry of bacteria into the cells of the UTIs. The most frequent BCC is by Gram-negative bacilli, colonizing, adhering and penetrating cells of the urinary tract. BCC is not considered in the CFU / mL count in urine culture using the conventional procedure. The drawback of the technique is that after BCC an innumerable number of bacteria can be located in a single cell and since the release of intracellular bacteria is not promoted, only a single CFU is quantified and not the real set that remains inside the cell. Such under-notification is significant given that the decision to offer an antimicrobial treatment to the patient depends on the count. This study aims to standardize an efficient technique for reporting CFU / mL in urine samples with BCC. Urine sample was collected. Urochemical profile, urinalysis, cytopathology and a plate count were performed to define the initial amount of bacteria, for later comparison against the post-treatment cell lysis bacterial count. To demonstrate the best cell lysis treatment: a randomized 3x2x3x2 full factorial design was used, in triplicate, whose factors were: Lysis method (Vortex®, Low frequency ultrasound (LFU) and Chemical Buffer), speed and time of exposure, and finally material of the sample container (glass or plastic). The technique that generated the greatest recovery and that reduced the quantification error was; LFU of the sample contained in glass, for 30s. However, regardless of the lysis technique, the use of only one substantially improves (63-734%) the bacterial bacillary recovery of urine samples with bacterial cell cytolysis.