M. Manafi

New developments in chromogenic and fluorogenic culture media.

This review describes some recent developments in chromogenic and fluorogenic culture media in microbiological diagnostic. The detection of beta-D-glucuronidase (GUD) activity for enumeration of Escherichia coli is well known. E. coli O157:H7 strains are usually GUD-negative and do not ferment sorbitol. These characteristics are used in selective media for these organisms and new chromogenic media are available. Some of the new chromogenic media make the Salmonella diagnostic easier and faster. The use of chromogenic and fluorogenic substrates for detection of beta-D-glucosidase (beta-GLU) activity to differentiate enterococci has received considerable attention and new media are described. Rapid detection of Clostridium perfringens, Listeria monocytogenes, Bacillus cereus and Staphylococcus aureus are other application of enzyme detection methods in food and water microbiology.

Evaluation of CHROMagar Candida for rapid screening of clinical specimens for Candida species.

CHROMagar Candida is a new differential culture medium that allows selective isolation of yeasts and simultaneously identifies colonies of Candida albicans, Candida glabrata, Candida tropicalis and Candida krusei. We evaluated this medium and compared it with a reference medium, Sabouraud glucose agar, for the presumptive identification of yeast species isolated directly on the medium from 1150 clinical specimens. A total of 731 specimens showed no growth, 299 isolates (70.2%) showed growth to the same extent on both media. Forty mixed cultures were detected on both media. More than one isolate was detected in 30 of the tested specimens on either CHROMagar (26 specimens) or Sabouraud glucose agar (four specimens). We found a sensitivity of 98.8% and a specificity of 100% for C. albicans, 66.7% and 99.8% for C. tropicalis, 100% and 100% for C. krusei, and 98% and 95.7% for C. glabrata. Regarding these results, CHROMagar Candida is recommended as a useful isolation medium capable of the presumptive identification of yeasts and better detection of mixed cultures in clinical specimens.

Fluorogenic and chromogenic enzyme substrates in culture media and identification tests

Rapid detection and identification of microorganisms is extremely important in many fields of applied and research microbiology. In general, fluorogenic and chromogenic substrates have proved to be a powerful tool, utilizing specific enzymatic activities of certain microorganisms, either in parallel with or instead of traditional methods. By incorporation of synthetic fluorogenic or chromogenic substrates into primary selective media, enumeration and detection can be performed directly on the isolation plate. The introduction of many of these media and identification tests has led to improved accuracy and faster detection of target organisms, often reducing the need for isolation of pure cultures and confirmatory tests.

Quantitative determination of total coliforms and Escherichia coli in marine waters with chromogenic and fluorogenic media

This study compared the performance of LMX® broth (LMX), Chromocult Coliform® agar (CC) and Chromocult Coliform agar plus cefsulodin (10 μg ml−1) (CC‐CFS), with standard methods multiple tube fermentation (MTF), for the enumeration of total coliforms and Escherichia coli from marine recreational waters. LMX and CC are two media designed to concurrently detect total coliform (TC) bacteria and E. coli by the specific action of β‐galactosidase (total coliforms) and β‐glucuronidase (E. coli). Overall results for the TC test showed that LMX, CC and MTF recovered 2.63, 1.95 and 1.90 times as many TCs as CC‐CFS, respectively. Data from the multiple range test showed significant differences (P < 0.05) between TC counts on CC‐CFS and LMX. The traditional MTF was less sensitive for E. coli enumeration. However, there was no statistically significant differences between LMX, CC, CC‐CFS and the MTF method for E. coli enumeration. Background interference was reduced on CC‐CFS and the counts obtained reflected more accurately the number of TCs. Therefore, the contribution of β‐galactosidase positive, non coliform bacteria (Aeromonas spp. and Vibrio spp.) to TC counts should not be neglected.

Comparative evaluation of different chromogenic/fluorogenic media for detecting Escherichia coli O157:H7 in food

Escherichia coli O157:H7 is a serious and common human pathogen that can cause diarrhoea, haemorrhagic colitis, and haemolytic uraemic syndrome (HUS). This study evaluated the enrichment, detection and confirmation procedures for the isolation of E. coli O157:H7 from raw ground beef and raw drinking milk. The purpose of this investigation was to compare Rainbow Agar O157 (RB; Biolog, Hayward, USA), Biosynth Culture Medium O157:H7 (BCM O157:H7; Biosynth, Staad, Switzerland) and Fluorocult HC (HC; Merck, Darmstadt, Germany) with the conventional Sorbitol MacConkey Agar (SMAC, Merck) using mEC + n (raw ground beef) and mTSB + n (raw milk) enrichment media. Single-path GLISA test (Gold Labeled Immuno Sorbent Assay; Merck) was used as the confirmation test. Growth of 466 strains of gram-negative rods isolated from food samples and 46 known E. coli strains from type culture and other collections (34 E. coli O157:H7 strains and 12 serotypes other than E. coli O157:H7) was examined on the agar media. The E. coli O157:H7 strains could readily be isolated and recognized uniquely by their typical black/grey colonies on RB and blue/black colonies on BCM O157:H7. Examination of the 46 known strains of E. coli reference strains showed false negative results on BCM O157:H7 (3.0%), RB (8.8%), HC (5.9%) and SMAC (5.9%) agars. On BCM O157:H7 no false negative results were found with the typical E. coli O157:H7 (beta-D-glucuronidase and sorbitol negative strains). One of two atypical E. coli O157:H7 strains (beta-D-glucuronidase positive) showed similar colouration to the typical strains and was mis-identified by each of the three media (RB, BCM O157:H7, and SMAC agar media). None of the 60 food samples tested yielded E. coli O157:H7. Examination of the food samples, showed that RB gave the lowest number of false positives. The percentages were RB (2.1%), BCM O157:H7 (3.3%), HC (6.2%), and SMAC (57.3%).

Performance of Candida ID, a New Chromogenic Medium for Presumptive Identification of Candida Species, in Comparison to CHROMagar Candida

ABSTRACT Candida ID agar allows identification of Candida albicans and differentiation of other Candidaspecies. In comparison with CHROMagar Candida, we evaluated the performance of this medium directly from 596 clinical specimens. In particular, detection of C. albicans after 24 h of incubation was easier on Candida ID (sensitivity, 96.8%) than on CHROMagar (sensitivity, 49.6%).

Rapid identification of Candida albicans by fluoroplate candida agar

150 clinical isolates of Candida species, Trichosporon species and Pichia anomala as well as 32 yeasts isolated from dairy products were tested in order to evaluate the utilization of a specific fluorogenic substrate in direct detection of Candida albicans. Detection of N-acetyl-β-d-galactosaminidase (NAGase) was performed with fluoroplate candida agar (Merck, Darmstadt, FRG), which contained 4-methylumbelliferyl-N-aceryl-β — d-galactosaminide (4-MUAG). The API 20C yeast identification system was used as a reference identification method for all isolates. The results of our investigations showed that 99% of C. albicans and all strains of C. stellatoidea examined were NAGase-positive. There were two false positive reactions of four C. tropicalis isolates and one false positive reaction of three Trichosporon beigelii strains.

A new plate medium for rapid presumptive identification and differentiation of enterobacteriaceae

A new selective differential agar medium for rapid presumptive identification of Enterobacteriaceae from water and food samples is described (EMX ID agar). By a combination of fluorogenic and chromogenic substrates, the medium detects the presence of beta-D-glucuronidase, beta-D-galactosidase, beta-D-xylosidase, tryptophane deaminase and H2S; additionally, cytochrome-oxidase and indole production can be demonstrated. This medium provides an inexpensive means for simple and rapid presumptive identification of E. coli and coliforms and for the differentiation within the Klebsiella-Enterobacter and the Proteus-Providencia-Morganella group. Furthermore, it allows to distinguish between the H2S-positive Enterobacteriaceae Citrobacter freundii, Salmonella spp., S. arizonae, Edwardsiella, Proteus mirabilis, P. vulgaris and some oxidase-positive bacteria.

Comparison of three rapid screening methods for Salmonella spp.: ‘MUCAP Test, MicroScreenR Latex and Rambach Agar’

Three new rapid methods for detection of Salmonella spp. have been studied. The fluorogenic MUCAP test (Biolife, Italy), the MicroScreenR latex slide agglutination test (Mercia, UK) and the Rambach agar test (Technogram, France) were compared for their sensitivity and their specificity. Some 175 strains, incuding 74 Salmonella strains and 101 non‐Salmonella strains were included in the study. The sensitivities of the MUCAP, the MicroScreenR and Rambach agar tests were 100%, 96% and 91%, respectively, and their specificities 80%, 96% and 100%, respectively.

Comparison of a New Commercial Test, GLABRATA RTT, with a Dipstick Test for Rapid Identification of Candida glabrata

ABSTRACT This study compares the performance of a 3-h dipstick trehalose test with GLABRATA RTT, a new commercially available 20-min test for the rapid identification of Candida glabrata. With the exception of blood agar, GLABRATA RTT gave reliable results with all media tested and was always superior to the dipstick test.