A new SimPlate heterotrophic plate count (HPC) method (IDEXX Laboratories, Westbrook, Maine) was compared with the pour plate method at 35°C for 48 h. Six laboratories tested a total of 632 water samples. The SimPlate HPC method was found to be equivalent to the pour plate method by regression analysis (r ‫؍‬ 0.95; y ‫؍‬ 0.99X ؉ 0.06).Water utilities are required to maintain a detectable disinfection residual in water distribution systems or measure for heterotrophic plate count (HPC) bacteria (6). The standard HPC pour plate method is an approved U.S. Environmental Protection Agency USEPA method (5) for reporting HPC in lieu of testing for residual disinfectant concentration or for testing when residual disinfectant levels are less than 0.2 mg/ liter in finished waters (4). This method, as well as other HPC methods, such as membrane filtration or spread plating, may be also used to collect data for internal purposes (nonreporting). All the methods (1) for testing of heterotrophic bacteria require time-consuming preparation of media and can be difficult to read. Recently, the SimPlate total plate count method for determining the most probable number (MPN) of microorganisms in food was developed by IDEXX Laboratories, Westbrook, Maine (3), and approved by the Association of Official Analytical Chemists (AOAC) International Research Institute (2). The formulation was modified to allow for the detection of heterotrophic bacteria in water. The test known as SimPlate for HPC medium contains substrates that are hydrolyzed by microbial enzymes to release 4-methylumbelliferone, which fluoresces blue under a long-wavelength (365-nm) 6-W UV light after incubation for 48 h at 35°C. The bacteria are detected as fluorescent wells on the SimPlate. The bacterial density of a water sample is determined by determining the number of positive wells and by using the MPN table provided for SimPlate. This format will allow a MPN/milliliter value up to 738 without any dilution. The objective of this study was to compare the performance of SimPlate and the HPC pour plate method (1) for the enumeration of heterotrophic bacteria. Six laboratories in different regions of the United States participated in this study.Between May and June 1997, naturally occurring heterotrophic bacterium samples were collected in sterile vessels at each site. The samples consisted of chlorinated drinking waters (neutralized with sodium thiosulfate) (1), well waters, untreated natural (raw) waters (lakes and streams), and secondary chlorinated sewage effluents (neutralized with sodium thiosulfate). Since chlorinated drinking waters have relatively few or no heterotrophic bacteria, the sites prepared composites of raw waters and/or secondary effluent with neutralized chlorinated drinking water. The composite samples were prepared in ratios of 1:1, 1:2, and 2:1 to allow as broad a range of bacterial counts as possible to be represented in the study. Each site was requested to test 100 samples for the evaluation. Approximately 40% of the samples were to be from ...

It has been reported previously (Annan & Manson, 1969) that chromatography of the a s -casein complex of bovine milk on columns of sulphoethyl Sephadex C-50 resulted in fractionation of the complex into a main component, asj-casein and a number of minor components one of which was designated Os 0 -casein. The present communication describes the preparation of Os 0 -casein from unfractionated casein in an amount and state of purity which render possible its further characterization. EXPERIMENTAL Acid-precipitated whole casein. Milk was obtained from individual Ayrshire cows of the Institute herd which were in the middle period of lactation and were free from mastitis. They were also selected as yielding one only of the known genetic variants of a si -casein, viz. type B as determined by electrophoresis performed on milk samples using the procedure described below. Whole casein was prepared from milk thus selected, as described previously (Manson & Annan, 1971), and dissolved in distilled water by addition of 1 N-NaOH in such a way that the pH value never exceeded 7-5. The solution was clarified by filtration through a filter disk (HP/EKS obtained from Carlson-Ford Ltd, Ashton-under-Lyne, England) under an applied pressure of 50 kPa (0-5 kg/cm 2 ) and its protein content when determined by measurement of its absorbance at 278 nm was approximately 6 %. tx S0 -Casein. a S0 -Casein was prepared from solutions of acid-precipitated whole casein by chromatography on columns (50-0 x 5-0 cm) of sulphopropyl Sephadex C-50 (Pharmacia, Great Britain, Ltd, London) which were made up in and equilibrated with Na formate/formic acid buffer (4-0 g NaOH, 11-5 g formic acid/1) containing 7-5 M-urea and having a pH of 4-0. In an earlier paper (Annan & Manson, 1969) this buffer was wrongly described as containing 4-0 g Na formate/1. Before application to the column the whole casein was first reduced by treatment with 2-mercaptoethanol (ME) at pH 8-6 following the procedure described by Anfinsen & Haber (1961). The reduction was terminated by addition of formic acid to pH 4-0 and the mixture was adjusted in composition to approximately that of the chromatographic buffer by addition to each 100 ml of 67-5 g urea, 1-7 g formic acid and 0-6 g NaOH followed by distilled water to 150 ml. The mixture was then dialysed against formate/urea buffer after which NaCl was added to a final concentration of 0-04 M. In a typical fractionation performed at 20 °C this casein solution (800 ml; 32 g protein) was applied to the top of the column followed by formate/urea buffer containing 0-04 M-NaCl. The flow through the column was maintained at 60 ml/h and the effluent which was monitored spectrophotometrically for the presence of protein

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