Most Probable Number (MPN) CE 370 - Lab Most Probable Number (MPN)

Introduction Many of the Bacteria that cause serious disease, such as typhoid fever, cholera, and dysentery, are discharged along with fecal wastes. These disease-causing organisms are called pathogens and are difficult to isolate and identify. Fortunately, less harmful, easily isolated bacteria called indicator organisms can be used indirectly to detect pathogens. Coliform bacteria is used as an indicator organisms. They live in the intestine of the human beings in large numbers, non disease-causing, easily detectable. The presence of coliform bacteria indicates that the water has been polluted with human or worm-blooded animals waste. It is a warning signal that more dangerous bacteria may be present.

Purpose of Experiment To learn enumeration technique of coliform bacteria

MPN Measurement A series of serial dilutions of wastewater sample is made (dilute by factors of 10) Transfer 1 mL sample from each of the serial dilutions to five fermentation test tubes containing lactose medium and inverted gas collection tube Incubate tubes for 24 hours at 35 ° C in a water bath Accumulation of gas (CO2) in inverted gas tubes is considered a +ve reaction (No gas –ve reaction)

The results of each dilution are reported as a fraction with the # of positive tubes over the total # of tubes (I.e., 3/5 means three + tubes in a five-tube sample) Test for fecal coliform is similar to total coliform assay except that the medium is different (lactose, tryptose, bile salts), tubes are incubated for 3 hr at 35 ° C and 44 for 21 h. Concentration of total or fecal coliform bacteria are reported as most probable number per 100 mL (MPN/100 mL)

MPN is based on the application of the Poisson distribution MPN is based on the application of the Poisson distribution. MPN is a statistical estimate of concentration. “Standard” method for calculation of MPN is to take 10, 1, and 0.1 mL of sample and dilute to 100 mL

Example A bacterial analysis of a surface water yielded the following results for the standard confirmed test of total coliform. Determine the coliform MPN using Poisson distribution.

Solution From Table F-1, Wastewater Engineering, Metcalf & Eddy, MPN /100 mL = 47 coliforms in 100 mL

Table 2. For 5 tubes each at 0. 1, 0. 01, and 0 Table 2. For 5 tubes each at 0.1, 0.01, and 0.001 g inocula, the MPNs and 95 percent confidence intervals. Pos. Tubes MPN/g Conf. lim. Pos. tubes Conf.lim. 0.1 0.01 0.001 Low High <1.8 -- 6.8 4 2 21 40 1 1.8 0.09 3 25 9.8 70 6.9 17 6 3.6 0.7 10 42 3.7 26 5.5 15 31 5.6 22 50 32 38 14 100 12 27 9.9 6.1 33 8.1 3.4 39 34 8.2 8.3 47 120 3.5 5 41 11 48 4.5 0.79 23 9.1 43 58 150 9.2 4.1 46 9.3 63 84 220 5.9 36 49 170 94 230 250 7.8 2.1 400 79 13 35 110 140 52 180 210 5.7 130 20 440 280 710 350 24 430 1,100 240 1100 540 1700 920 2600 1600 4600   >1600 700