Standard Plate Count Agar Research Articles

Characterization of dysgonic, heterotrophic bacteria from drinking water.

Only a small percentage of the heterotrophic bacteria encountered in water distribution systems are identifiable, because of these organisms fail to grow on the conventional media used for biochemical characterization. Organisms that would not subculture from the same standard plate count agar used for initial isolation were successfully subcultured on a low-nutrient medium, R3A. These cultures were then inoculated to a modified O/F base medium containing specific substrates. This, combined with a lower incubation temperature (30 degrees C), increased the enzymatic activity of many of the organisms. These reactions established a groundwork for tentative taxonomy.

Detection of post-pasteurization contamination of cream by impedimetric methods.

Impedimetric methods for evaluating post-pasteurization contamination and shelf-life of cream were assessed. Over 94% of the samples tested were in agreement, using selected cut-offs of 20 h for detection time measured at 21 degrees C with creams containing inhibitors for the growth of Gram positive bacteria on standard plate count agar as growth media, and 3.2 X 10(7) cfu/g for plate counts obtained on cream which had been pre-incubated in the presence of inhibitors for the growth of Gram positive organisms, and on cream stored at 6 degrees C for 7 d. Agreement between the impedimetric method and plate count was not as good if either Brain Heart Infusion or Milk Agar was used in place of Plate Count Agar in the former technique. A poor correlation was obtained between plate count methods for enumerating post-pasteurization contamination and keeping quality with impedimetric measurements on cream alone. It was possible, with a reasonable degree of certainty, to determine if cream had suffered post-pasteurization contamination within 20 h of production.

Microbiological Survey of Adirondack Lakes with Various pH Values

Nine high-altitude oligotrophic Adirondack lakes in upstate New York having water of pH 4.3 to 7.0 were surveyed for total bacterial numbers and possible adaptation of the microbial communities to environmental pH. The number of heterotrophic bacteria from water samples recoverable on standard plate count agar were low (10 to 10 per ml) for most of the lakes. Acridine orange direct counts were approximately two orders of magnitude higher than plate counts for each lake. Sediment aerobic heterotrophs recovered on standard plate count agar ranged from 1.4 x 10 to 1.3 x 10 per g of sediment. Direct epifluorescence counts of bacteria in sediment samples ranged from 3.0 x 10 to 1.4 x 10 per g. Low density values were consistent with the oligotrophic nature of all the lakes surveyed. There were no apparent differences in numbers of bacteria originally isolated at pH 5.0 and pH 7.0 between circumneutral lakes (pH > 6.0) and acidic lakes (pH < 5.0). Approximately 1,200 isolates were recultured over a range of pH from 3.0 to 7.0. Regardless of the original isolation pH (pH 5.0 or pH 7.0), less than 10% of the isolates grew at pH < 5.0. Those originally isolated at pH 5.0 also grew at pH 6.0 and 7.0. Those originally isolated at pH 7.0 preferred pH 7.0, with 98% able to grow at pH 6.0 and 44% able to grow at pH 5.0. A chi-square contingency test clearly showed (P < 0.005) that two distinct heterotrophic populations had been originally isolated at pH 5.0 and pH 7.0, although there is undoubtedly some overlap between the two populations.

Evaluation of Methods for Estimation of Viable Bacterial Count of Raw Meat

An improved method was developed for the estimation of viable bacterial counts for raw meat stored at low temperature, and was used to analyze the influence of various factors affecting the viable bacterial count. The viable bacterial counts of 40 samples each of commercial cut beef, pork, and chicken meat stored refrigerated or frozen in the raw state were estimated comparatively by using three methods of sample inoculation, i. e., the surface plate method and the pour plate method poured at 50 and 60°C with standard plate count agar, at six incubation temperatures (7, 25, 30, 32, 35, and 37°C).1) Among the three methods of inoculation, the viable bacterial count was the highest in the surface plate method, regardless of temperature of culture. As regards the temperature of culture, the count was the highest in the culture at 25°C, regardless of the method of inoculation used. Therefore, the count was the highest when the surface plate method was performed at 25°C.2) The viable bacterial count was significantly higher when estimated by the surface plate method at 25°C than when estimated by the method prescribed in the Official Guideline of Sanitary Tests of Food with culture at 35°C.3) There was a significant correlation between the viable bacterial count estimated by the surface plate method at 25°C and that at any other temperature.4) The mode of storage (refrigeration or freezing) was the most influential factor upon the viable bacterial count. The next most important factor was the kind of meat (beef, pork, or chicken), followed by the temperature of culture, and the method of inoculation in decreasing order of influence upon the count.

Quantitative microbiological monitoring of hemodialysis fluids: evaluation of methods and demonstration of lack of test relevance in single-pass hemodialysis machines with automatic dialysate proportioning with reverse osmosis-treated tap water.

Two methods for estimating the quantity of microorganisms present in hemodialysis fluid, a blood agar surface-spread plate method and a total-count water tester device impregnated with modified standard plate count agar (Millipore Corp., Bedford, Mass.), were evaluated. Both methods exhibited comparable precision; however, colony counts obtained with the total-count water tester were consistently and unacceptably low. The need for routine quantitative microbiological monitoring of hemodialysis fluids such as that recommended by the American Public Health Association was not supported by the results of this study. Such testing was not of value in predicting untoward reactions for patients undergoing hemodialysis, nor did quantitative testing of hemodialysis fluids identify the buildup of potentially hazardous levels of contamination within hemodialysis systems. Finally, the kinds of organisms found in hemodialysis systems, i.e., gram-negative water-borne bacilli, were elucidated.

An estuarine agar medium for enumeration of aerobic heterotrophic bacteria associated with water, sediment, and shellfish.

A plate count agar was formulated for use in bacteriological analysis of estuarine samples and was tested together with standard plate count agar and an estuarine salts yeast extract agar for growth of aerobic, heterotrophic bacteria in water, sediment, and oysters. The estuarine agar was found to be efficient for enumerating aerobic, heterotrophic bacterial populations of water, sediment, and oysters, and is recommended for plate counts of estuarine samples.

Bacterial Contamination of Drinking Water Supplies in a Modern Rural Neighborhood

On six occasions during a 15-month period, the private well and spring water supplies in a modern rural neighborhood of 78 households were examined for total coliforms, fecal coliforms, Staphylococcus aureus, and standard plate count bacteria. More than one-third of the water supplies were unsatisfactory on at least one occasion as judged by standard plate counts over 10(3)/ml and the presence of coliforms, fecal coliforms, and/or S. aureus. Citrobacter freundii, Klebsiella pneumoniae, and Escherichia coli were the most frequently isolated total coliforms. At least 12 other genera of bacteria were identified from standard plate count agar. Coliform contamination was found to be higher after periods of rainfall, and high standard plate counts were more prevalent during warmer weather. These observations probably reflect leakage of surface water into improperly sealed wells or aquifer contamination during winter and the lack of chlorination to control microbial regrowth during the warm season. An inverse correlation was found between the presence of high standard plate counts and incidence of coliforms. Consumer education and at least a twice yearly monitoring of private water supplies (winter and summer) are suggested methods to signal that treatment may be necessary to reduce the risk of waterborne disease.

Effect of Delayed Heading on Some Quality Attributes of Penaeus Shrimp

To determine the effect of delayed heading on shrimp quality, shrimp were stored on ice with and without heads for 10 days. Some shrimp were delay-headed after 5 days and returned to ice for the remainder of the storage period. Microbiological studies were conducted at 0, 5 and 10 days of storage. Total aerobic plate counts were done using Standard Plate Count agar with an added 0.5% NaCl. Incubation was at 20 C for 5 days. Analyses indicated similar counts on shrimp tails stored with or without heads and those delayed-headed. Counts ranged from 2.4 × 106 bacteria/gram at 0 day to 1.6 × 109 bacteria/gram on the 10th day. Identification of the flora present revealed that the same major groups of organisms predominated on shrimp tails subjected to the different storage treatments and the head did not alter development of the usual flora. Flavobacterium, Pseudomonas, Planococcus, Moraxella and the Vibrio/Aeromonas group were the major genera encountered. A shift in bacterial populations was observed during storage. Flavobacterium species predominated during the first 5 days of storage; however, after the fifth day Pseudomonas species predominated. Sensory panel data revealed no differences in acceptability between shrimp tails stored with or without heads and those delay-headed.

INHIBITION OF GROWTH OF AIRBORNE COLIFORMS AND OTHER BACTERIA ON SELECTIVE MEDIA

Coliforms when sampled from air were inhibited or restricted in growth on regular selective media. For trials, growth on Standard Plate Count Agar (SPC) was used as 100%. The percentage of colony growth of Escherchia coli and Enterobacter aerogenes on modified selective media or with SPC were respectively as follows: violet red bile/violet red bile (VRB/VRB) (overlay on base) <4, 15; desoxycholate/desoxyeholate (DES/DES) < 1, 17; tergitol/tergitol (TER/TER) 23,49; eosine methylene blue/eosine methylene blue (EMB/EMB) 122, 86; endo/endo (END/END) 40, 104; MacConkey/MacConkey (MAC/MAC) <2, 10; SPC/VRB <2, 22; SPC/DES <2, 23; VRB/SPC 54, 60; TER/SPC 72, 119; EMB/SPC 97, 119; END/SPC 95, 90; 1 VRB: 1 SPC, as overlay and base 14, 50. Recovery percentages of Pseudomonas aeruginosa and Pseudomonas fragi on modified selective media were greater than those of the coliform bacteria. Fewer Serratia marcescens colonies grew on DES or SPC/DES and more grew on the other modified selective media than did coliform colonies. Growth of airborne Salmonella newbrumwick ranged from <1% on SS agar to 118% on MAC/SPC. Aerosolization of coliforms with skim milk compared to distilled water resulted in growth of more colonies on selective media. Desoxycholate, Bile Salts No. 3, Tergitol No. 7, and crystal violet in the media definitely limited growth of airborne coliforms.

NON-LETHAL INJURY AND LIMITATIONS OF RECOVERY OF COLIFORM ORGANISMS ON SELECTIVE MEDIA

Cell injury as a factor in the enumeration of coliform bacteria with selective media was evaluated. Non-lethal injury reduced the ability of cells to produce outgrowth. Escherichia coli and Aerobacter aerogenes were equally sensitive. Brilliant green lactose bile broth, desoxycholate lactose agar, and violet red bile agar were similar in inhibitory effect.Cell injury occurred with sub-lethal heat treatments, exposure to chlorine, exposure to sodium chloride, and freezing and thawing cycles. Circumstances to inflict injury are found in common environmental conditions of the food industry. Presence of injured cells may provide a count with a selective medium that is only 10% of the total count obtained with standard plate count agar. The limiting factor for recovery in selective media was associated with the surfactant.An understanding of these limitations in the use of selective media in quality control and public health applications of the tests should contribute to proper interpretation of results.

Factors Influencing the Occurrence of High Numbers of Iodine-Resistant Bacteria in Iodinated Swimming Pools

It has been shown that, although iodinated swimming-pool waters are usually free from coliform bacteria and enterococci, the total counts frequently become relatively high. Pseudomonas alcaligenes and Alcaligenes faecalis have been shown to account for most of these high counts. It was of interest, therefore, to compare the microbial flora of four alternately chlorinated and iodinated swimming pools. By means of the membrane filter method and suitable selective media, examinations were made for total viable counts, coliform bacteria, enterococci, staphylococci, Streptococcus salivarius , and P. aeruginosa . Colonies also were picked from membrane filters incubated on standard plate count agar and identified. The results showed that, although viable counts were significantly higher during the iodinated periods, the specific types of bacteria determined were either fewer than or the same as in chlorinated periods. During chlorination, the predominant microbial flora consisted of staphylococci and members of the genus Bacillus . During iodination, however, the P. alcaligenes-A. faecalis group accounted for 92 to 99% of the microbial flora. The accumulation of high numbers of these bacteria was shown to be due to their iodine resistance and their ability to grow rapidly in pool water in the absence of free iodine.

The microflora of raw bulk tank milk.

Milk samples from 141 farms were plated on standard plate count agar and the colony count determined after incubation for 48 hours at 32 °C. Twenty colonies were picked at random from plates containing between 30 and 300 colonies. The isolates were inoculated into litmus milk and subsequently characterized on the basis of shape, Gram-staining reaction, catalase production, lactose fermentation, and the ability to form spores.Certain general trends in the flora were observed. In milk of a colony count less than 2 × 104 per ml, micrococci were the dominant group of organisms, and as the colony count of the milk increased the percentage of micrococci decreased and the percentage of Gram-negative rods and streptococci usually increased. In spite of these general trends a study of the flora of individual samples showed that quite marked variations did occur.

Factors Influencing the Occurrence of High Numbers of Iodine-Resistant Bacteria in Iodinated Swimming Pools

It has been shown that, although iodinated swimming-pool waters are usually free from coliform bacteria and enterococci, the total counts frequently become relatively high. Pseudomonas alcaligenes and Alcaligenes faecalis have been shown to account for most of these high counts. It was of interest, therefore, to compare the microbial flora of four alternately chlorinated and iodinated swimming pools. By means of the membrane filter method and suitable selective media, examinations were made for total viable counts, coliform bacteria, enterococci, staphylococci, Streptococcus salivarius, and P. aeruginosa. Colonies also were picked from membrane filters incubated on standard plate count agar and identified. The results showed that, although viable counts were significantly higher during the iodinated periods, the specific types of bacteria determined were either fewer than or the same as in chlorinated periods. During chlorination, the predominant microbial flora consisted of staphylococci and members of the genus Bacillus. During iodination, however, the P. alcaligenes-A. faecalis group accounted for 92 to 99% of the microbial flora. The accumulation of high numbers of these bacteria was shown to be due to their iodine resistance and their ability to grow rapidly in pool water in the absence of free iodine.

&lt;I&gt;Lactobacillus bulgaricus&lt;/I&gt;の栄養要求とそれにもとずく培地の改良について

Cultures of Lactobacillus bulgaricus often fail to grow on media commonly used for lactic acid bacteria such as standard plate count agar. This is considered dus to deficiencies of some growth factors required by the organisms. To determine the factors deficient in these media, a survay of growth requirements of six cultures of L. bulgaricus in chemically defined media was done.All of six cultures required “tween 80” and enzymatic digest of casein for their growth. “Tween 80” could be replaced with “tween 40” plus oleic acid, lecithin or cephalin.Three cultures required pantethine in a medium containing 0.4γ/ml of calcium pantothenate. However, pantethine could be replaced with higher amounts (e.g., 1γ/ml) or calcium pantothenate. When cysteine level in a medium was reduced to 10γ/ml (one-tenth of usual synthetic media) and cystine was omitted, calcium pantothenate in a concentration of 2γ/ml failed to replace pantotheine. This suggests that “activity coefficient” pantethine/pantothenate is a function of cysteine level in a medium.One culture identified asThermobacterium jugurtOLRA-JENSEN did not grow in the absence of lactose even after prolonged incubation period.None required erotic acid or thymidine, and the latter seemed to be inhibitory to some cultures.