PubMed · 12617120
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Mindy Goldman. 2002. Challenges in developing a bacterial detection system.. https://doi.org/10.1111/j.1423-0410.2002.tb05284.x
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AIMS: To quantify the influence of the growth phase, storage temperature and nutritional quality of the plate count medium on the apparent viability of Mannheimia haemolytica during storage at different temperatures. METHODS AND RESULTS: Mannheimia haemolytica was grown in shake flasks and in aerobic continuous culture to investigate factors affecting cell viability during storage, which was determined using plate counts on different media and epifluorescence microscopy. The high specific death rates of cells harvested after cessation of exponential growth and stored at 22, 4, -18 and -75 degrees C could be related to the rapid onset of exponential death in batch cultures. Yeast extract supplementation of the culture medium increased the viability of cells at most of the above-mentioned storage temperatures. Of the total cell count in continuous culture, only 48% could be recovered on brain-heart infusion agar, whereas supplementation of the agar medium with foetal calf serum increased the plate count to 71% of the total count. CONCLUSIONS: Mannheimia haemolytica cells harvested from the exponential growth phase had the highest survival rate during storage at low temperatures. Plate count values also depended on the nutritional quality of the agar medium. SIGNIFICANCE AND IMPACT OF THE STUDY: Results presented here impact on the procedures for culture preservation and plate count enumeration of this fastidious animal pathogen.
AIMS: This study examined the suitability of three analytical methods for isolating and enumerating Escherichia coli from conventionally treated sewage sludge. METHODS AND RESULTS: Crude sewage, mesophilic anaerobic digested (MAD) sludge, and final product sludge samples were taken from six sewage treatment works for analysis. Two of the three methods tested were membrane filtration techniques, utilizing chromogenic E. coli/coliform (CEC) media and membrane-lactose glucuronide agar (MLGA); the third method was a most probable number (MPN) technique utilizing Colilert in Quantitray 2000 (Idexx). The methods were evaluated for variation, consistency, false-positive and false-negative results, as well as method correlation. The methods gave good and consistent recovery of E. coli for a range of conventionally treated sewage matrices. All of the methods had a false-positive rate of <3%, although MLGA had a high false-negative rate (35.5%) compared with Colilert (3.81%) and the CEC method (6.75%). This resulted in slightly lower presumptive counts but comparable numbers of confirmed counts. CONCLUSIONS: The three detection methods tested, chromogenic, MLGA and Colilert gave comparable recoveries, and did not vary by greater than one order of magnitude (1 log). SIGNIFICANCE AND IMPACT OF THE STUDY: Forthcoming revisions to the Use of Sludge in Agriculture Regulations (1989) will categorize sewage sludge as untreated, conventionally treated or enhanced treated in accordance to microbiological standards. The standard will be based upon numbers of E. coli removed through the sludge treatment process and the numbers remaining in the final product. It is recommended that the Colilert 2000 (Idexx, Westbrook, Maine) and CEC methods would be equally suitable to assess the reduction of indigenous E. coli in conventionally treated sludges, and that MLGA be used with follow-up confirmatory testing.
The BD ProbeTec ET system for identification of Mycobacterium tuberculosis complex (MTBC) isolates from BACTEC 12B culture vials was evaluated in comparison with BACTEC NAP (p-nitro-alpha-acetylamino-beta-hydroxy-propiophenone) differentiation. Of 145 mycobacterial isolates tested, comprising 89 MTBC and 56 non-tuberculous mycobacteria (NTM), BD ProbeTec ET correctly identified 87 MTBC and 56 NTM but missed two MTBC. Three NTM were misidentified when NAP was incubated at 37 degrees C only. Overall sensitivity, specificity and positive and negative predictive values were respectively 97.8, 100, 100 and 96.6 % for the BD ProbeTec ET system and 100, 94.6, 96.7 and 94.6 % for NAP.