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Products of thiocyanate oxidation by lactoperoxidase inhibit gram-positive bacteria that produce peroxide. We found these products to be bactericidal for such gram-negative bacteria as Pseudomonas species and Escherichia coli , provided peroxide is supplied exogenously by glucose oxidase and glucose. By the use of immobilized glucose oxidase the bactericidal agent was shown to be dialyzable, destroyed by heat and counteracted, or destroyed by reducing agents. Because the system is active against a number of gram-negative bacteria isolated from milk, it may possibly be exploited to increase the keeping quality of raw milk.

The effect of lactic cultures on the growth of psychrotrophic bacteria in milk stored at 3.5 and 7C was investigated. Psychrotrophs were those in raw milk, or those isolated from raw milk and added to sterile milk as pure cultures. Of the lactic cultures investigated, certain commercial multi-strain cultures (lactic acid-producing streptococci plus Leuconostoc cremoris ) were the most effective in restricting psychrotrophic growth. Lactic culture was added to milk at .5% in all experiments. Inhibition of psychrotrophs was due to a reduction in growth rate because reduction in the initial psychrotroph population usually was not observed. The degree of inhibition varied with the lactic culture, the initial population of psychrotrophs, the psychrotroph culture, storage temperature and time. Inhibition due to lactic culture was decreased by addition of catalase, suggesting that hydrogen peroxide was the inhibitor. Small amounts of hydrogen peroxide were detected in sterile milk inoculated with an inhibitory culture and stored at 3.5 or 7C. Hydrogen peroxide decreased in the presence of psychrotrophs, and hydrogen peroxide was not in raw milk.

Production of heat resistant proteases by psychrotrophs growing in milk, resistance of such proteases to ultrahigh temperature treatments and action of these enzymes on milk were studied. All of the psychrotrophs obtained from raw milk produced proteases that survived 149 C for 10s. Seventy to ninety percent of the raw milk samples contained psychrotrophs capable of producing heat resistant proteases. The protease chosen as a model was resistant to heat treatments at 110 to 150 C, and the inactivation parameters suggested that thermal destruction of heat resistant proteases would damage the milk severely. The casein content and pH of normal milk were suitable for protease action, and the protease was quite active at normal and elevated room temperatures. The protease rapidly spoiled sterile milk with the development of bitter flavor, clearing, or coagulation; and the susceptibility of sterile milk to protease increased during storage of the milk.

Spore spoilage in pasteurized milk and UHT and sterilized milk is reviewed with particular reference to cream. The problem of the incidence of bitty cream and the effectiveness of refrigeration in checking this is discussed. Results of studies on the bacterial quality of bulk raw milk and tanker reload milk using the thermoduric count are given in detail. Current methods for pasteurizing cream are described and studies on the effect of storage temperature on the shelf life of pasteurized cream are referred to in detail with particular reference to the lag in bacterial multiplication and the use of the Methylene Blue Test.

The Bristol, Virginia Division of Dairymen, Inc. is a storage and transfer operation for raw milk. As production increased and effluent quality regulations become more stringent, waste treatment progressed from land irrigation, to a two-stage biological system to the present four-stage biological system including: (a) full mixed anaerobic; (b) facultative-high rate trickling filter and non solids controlled aeration; (c) quiescent anaerobic; and (d) facultative-high rate trickling filter and activated sludge. Treatment efficiencies were

Studies were made with alkaline and acid detergent solutions at three temperatures (20, 45, and 70 C) and four milk concentrations (0.00, 0.0.1, 0.10, and 1.0%). Turbidity measurements were made continuously while the solutions were circulating through a piping system. The turbidity of milk-detergent-water solutions was found to be primarily influenced by the concentration of milk. As little as 0.01% milk caused a significant change in turbidity (1.4%), and the percentage of transmittance decreased an average of 78.6 when 1% milk was added to the water that contained the detergent. Detergent concentrations and temperature of the solutions had only minor effects on turbidity. Formation of precipitates by action of phosphates in hard water did not decrease the sensitivity of the method to added milk.

Psychrotrophic bacteria were enumerated in raw milk by the microscopic colony count and counts were compared with those obtained by standard methods. Incubation conditions were 7 C for 10 days for the standard procedure and 7 C for 48 and 72 h or 21 C for 13.5 and 16.5 h for the microscopic colony count. Though each of the microscopic colony counts differed from the standard psychrotroph count (P < 0.01), there was a good general relationship in each instance (correlation coefficients ≥ 0.94). Wide variations in ratios of microscopic colony counts to standard psychrotroph counts made interpretations difficult. Generally, microscopic colony counts after 48 and 72 h at 7 C were less than standard psychrotroph counts. Incubation at 21 C yielded microscopic colony counts that were similar to standard psychrotroph counts for most samples; however, some microscopic colony counts at this temperature were substantially higher than the standard psychrotroph counts.

Summary Thirty samples of Damietta cheese, purchased over a period of one year in retail markets, and 10 samples from laboratories of the Ministry of Public Health were examined quantitatively for coagulase-positive staphylococci on three recommended selective media. Nine out of the thirty market samples (30 %) and all the laboratory samples contained coagulase-positive staphylococci in numbers that ranged from 1,000 to several million per g. A correlation was found between acidity, salt content, and the numbers of staphylococci detected, which depended also on the age of the cheese. A comparison of selective media for isolating coagulase-positive staphylococci was favoured by Mannitol Salt Agar and Staphylococcus Medium 110, although the former showed the highest percentage of recovery. Ludlam's Medium was not recommended, for it seemed too inhibitory to be used for enumeration of staphylococci in cheese.

The geometric mean bacterial count of 131 raw milk samples, using plates incubated 48 h at 30 C, was 15% higher than the geometric mean when the bacterial counts were determined at 32 C incubation. When 72 h of incubation were used, the geometric mean of samples using the lower incubation temperature was 20% higher. The mean for all 72-h raw milk counts was 31% higher than that for 48 h counts. There was no significant difference between geometric means of 51 pasteurized milk samples using plates incubated at 30 or 32 C after 48 h incubation. After 72 h of incubation, the geometric mean obtained at 30 C was 10% higher. For all 72-h counts on pasteurized milk the mean was 55% higher than for 48 h. A number of both raw and pasteurized samples showed little or no increase in count following Preliminary Incubation at 12.8 C for 18 h while others “blew up” to high levels, suggesting undesirable contamination. The International Dairy Federation procedures tended to reflect the latter better than the SPC.

Today's consumer determines the acceptance of milk by flavor and shelf life. Rancid, oxidized, and strong feed tastes of raw milk need to be eliminated. To prevent rancid and psychrotrophic flavor development, milk should be processed within 24 h of collection. Hot water sterilization of processing and filling equipment is essential. Product temperatures below 40 F at the filler extend shelf life. Proper handling during delivery and in stores is necessary to maintain good taste and keeping quality. Temperatures below 40 F, minimal fluorescent light and daylight exposure, and proper rotation are the key factors. Containers vary in their ability to protect the flavor and change in milk composition. Holding quality programs of 14 days at 45 F should be adopted by industry. Ten-day coding or open dating should be practiced by all processors. Regulatory agencies need to permit tests which will indicate shelf life. More emphasis must be placed on factors which determine consumer acceptability. These are flavor and shelf life.

Seit einigen Jahren last sich ein zunehmendes Interesse der Molkereiwirtschaft an Fragen der Rohstofferfassung beobachten. Der Strukturwandel beim Milcherzeuger, erweiterte Erfassungsgebiete, der technische Fortschritt und die Veranderung der Kostensituation haben die traditionellen Erfassungsformen verandert. Heute wird der uberwiegende Teil der Milch mit Tanksammelwagen erfast. Aus Wirtschaftlichkeitsgrunden gilt es nun, das kapitalintensive Instrument Tanksammelwagen optimal einzusetzen. Zur Erhohung der Wirtschaftlichkeit genugt aber nicht nur eine Senkung der Kosten. Auch die die Erlosseite beeinflussende Rohmilchqualitat bedarf besonderer Aufmerksamkeit. Collection of Milk – A Problem of Adjustment in the Dairy Industry Since a few years, the dairy industry is showing considerable interest in the problems of collection of raw materials. Structural changes at the level of milk producer, extension of areas for collection, technical progress and changes in price situation have altered the traditional forms of collection. The major part of milk is collected at present in tank cars. Due to economic reasons, efforts are made towards optimum use of the rather expensive tank cars. Reduction of costs for economic reasons should not be the only aim, but also special attention should be given to the quality of raw milk.

A vapor pressure osmometer requiring a 5- to 7-microliter sample to saturate a 0.64 cm filter paper disc fixed a digital readout of milliosmolality in 110 sec. A coefficient of variability of 0.70 was obtained on a raw milk sample tested 25 times when an acetone impregnated tissue was used to clean the sample chamber between tests. Two hundred individual cow milk samples from 20 herds averaged 280.0 ± 3.0 milliosmols. Milk samples containing up to 25% added water were evaluated on both the vapor pressure osmometer and a thermistor cryoscope with a resultant correlation coefficient of 0.991. A collaborative study involving eight hospital and industry laboratories was conducted. When the results of two laboratories were discarded, due to instrument maintenance problems, there were no significant differences among the laboratories in their abilities to quantitate added water in milk.

This paper reports effects of concentration of milk, concentration of individual ingredients of detergents, and temperature on transmittance of light at 527 nm through solutions of milk, water, and individual detergent ingredients. Milk in soft water caused the major portion of the variation, more than 99% of the total sum of squares, for each ingredient except sodium hydroxide and wetting agent. Milk contributed 95.7 and 17.9% to the sums of squares for the sodium hydroxide and wetting agent, respectively. Concentration of detergent ingredient and temperature were of practical significance only with wetting agent. Effects of milk on transmittance were significantly different among milk concentrations for all detergent ingredients, except trisodium phosphate in hard water at the lowest concentrations of milk. Although precipitates contributed to turbidity of solutions containing trisodium phosphate in hard water, milk was responsible for 95.9% of the variance in the sum of squares. Additionally, in hard water milk accounted for 98.9, 99.1 and 99.7% of the sums of squares in analyses of effects on turbidity of sodium tripolyphosphate, sodium hexametaphosphate, and tetrasodium pyrophosphate, respectively. Turbidity increased with time of holding some phosphate solutions, however, this did not appear to pose serious problems in measuring content of milk.

Raw milk samples were diluted with distilled water or distilled water with added phosphate buffer as recommended by Standard Methods for the Examination of Dairy Products. The standard plate counts were higher in diluent without phosphate buffer with both high and low count milk. The higher counts were significant when analyzed by a nonparametric sign test or a t-test of differences but were not significant with an analysis of variance technique. Reproducibility was not statistically different in the two diluents. It is suggested that the use of phosphate buffer for raw milk bacteria counts be discontinued until information showing definite advantages is provided.

Milk is nature's most nearly perfect food and to utilize its nutritive values milk must be free from all pathogenic microorganisms. Pathogenic bacteria present, if any, in raw milk supplies are destroyed by the pasteurization process. Milk and its fluid products, so treated, possess on an average of 7–10 days shelflife, which in modem sophisticated distribution systems, is not adequate. Commercial sterilization in conjunction with aseptic packaging extend the shelflife of milk products to 90+ days. These products, excellent in performance, are not yet popular in the dairy case but, if consistent efforts are continued for their promotion, will win consumer's confidence. Increasing application of the sterilization process in the dairy industry has prompted marketing of numerous types and brands of sterlilizing equipment. Every sterilization operation, due to differences in equipment and range of products manufactured, is faced with various problems in commercial applications of the process. Besides technical problems, label declaration and mode of distribution of these products cause legal complications in many areas of this country. Furthermore, no definition for commercially sterilized milk products aseptically packaged in flexible containers has been officially established by federal, state, and local regulatory agencies. Regardless of problems and challenges in commercial application of the sterilization process in the milk industry, potential opportunities in this country for milk products, thus processed, are great.

SummaryStarch-gel electrophoresis (SGE) and formol titration methods for detecting proteolysis in cold-stored raw milk have been studied to establish their value as quality indices. When examined by SGE, the first evidence of proteolysis in raw milks stored at 5°C was the formation of para-κ-casein. However, this fraction could not be detected on the starch gels until the total bacterial count (TBC) exceeded 107/ml. The SGE method appeared more reliable than the previously discussed tyrosine value method. Formol titration did not appear to have any application in the screening of cold-stored raw milks with TBC < 107/ml, but would detect some milks of poorer bacteriological quality.

The many factors involved in the contamination of milk with B. cereus cells and the importance of minimizing the number of spores at the time of pasteurization are discussed. Methods of controlling the level of B. cereus spores and the use of heat in the cleaning and sterilizing operation are shown, and studies on the level of aerobic spores in raw milk supplies showing a seasonal effect evaluated.

Cold raw milk with .1 ppm of added copper was pumped at a rate of 2 liters/h through a bed of 20 g of glass-bound trypsin for 5 h daily for 14 days. After each daily run the reactor was back washed with distilled water, flooded with 20% ethanol in phosphate buffer, and refrigerated. Samples were pasteurized, stored 6 days, and tested for oxidation. All copper-containing samples were protected from oxidation by the treatment. The bound enzyme showed no loss of activity over the 14 days as determined by N-benzoyl-L-arginine ethyl ester hydrolysis. Standard plate counts of ethanol solution after storage, aqueous washing, and the milk before and after the treatment showed that 20% ethanol was effective in limiting microbial growth, that there was no microbial build-up in the reactor during the 5 h of continuous milk passage, and that milk was not substantially contaminated by the treatment. Flow rate decreased with time whether the reactor was used as a packed column or a fluidized bed due to plugging of the column or the retaining screen. No increase in nonprotein nitrogen in the treated samples was detected. However, when casein was used as the substrate, increases in nonprotein nitrogen and free amino groups were appreciable.

Sporeforming bacteria are present in practically all raw milk, but usually in small numbers if the milk is produced under modern sanitary conditions. There appears to be no relationship between total bacteria and spore counts, nor between total bacteria and the incidence of any given species of sporeforming organism. Bacillus species account for about 95% of the total sporeforming bacteria in milk, with Clostridium species comprising the remainder. In the United States, approximately 43% of the Bacillus organisms are B. licheniformis and 37% are B. cereus. Reports from some other countries indicate a predominance of B. cereus, or essentially a reversal in the prevalance of these two species. Spores of both these organisms survive low temperature pasteurization, some persisting through even the lower temperatures in the ultra-high range, and recent reports indicate that some sporeformers are psychrotrophic. B. cereus has been implicated in outbreaks of food-borne illness, but this is still controversial. Should B. cereus be proven capable of growing in the psychrotrophic range, and also responsible for food-borne illness, then the significance of spores in milk would take on added importance.

Summary Standard plate and psychrophilic bacterial counts, on 312 samples of milk collected from farm bulk tanks and milk cans, were compared with methylene blue reduction times using the standard procedure or preliminary incubation of samples at 55 F for 18 hr prior to testing. When psychrophilic bacterial counts were less than 100,000/ml, according to standard plate counts a high proportion of methylene blue reduction times were properly classified. However, psychrophilic bacterial counts greater than 100,000/ml did not significantly decrease reduction times, thus a high proportion of such milks was not properly classified by methylene blue reduction times. The proportion of misclassification of samples having standard plate counts in excess of 200,000/ml and methylene blue reduction times of 5 and 6 hr was determined.

Summary A maximum flash temperature of 168°F. may be used in heating milk for the manufacture of cheddar cheese. Higher temperatures give a weak texture, a scorched flavor and sometimes allow a bitter flavor to develop. Heating milk to 160° to 168°F. improves the quality of the cheese in the case of poor milk, although uniform improvement is not noticed in the case of milk of good quality. Gassy milk pasteurized to 176°F. produces cheese of better quality than if the raw milk were used. Greater uniformity in the cheese is obtained by heating the milk. A yield of 9.49 pounds of cheese for each 100 pounds of milk testing 3.34 per cent butterfat was obtained. The loss of butterfat in the whey was 0.053 per cent lower when the milk was heated.

Details are given of two simple tests for efficiency of pasteurisation. The first, test A, may be carried out completely within half an hour, and will detect any gross errors of pasteurisation technique. The second, test B, is more searching, and will reveal minor errors of technique as follows:(a) 1½° F. below the minimum temperature for pasteurisation, or (b) heating (at 145° F.) for 20 instead of 30 min., or (c) admixture of 0·25 per cent, of raw milk with properly pasteurised milk. Taken together, the two tests form an efficient method for the laboratory diagnosis of faulty pasteurisation.

1. Spray-dried (Kestner process), roller-dried and evaporated milks were prepared commercially from one batch of raw milk.2. The biological value of the proteins (nitrogen), vitamin B1and the growth-promoting properties of the milks were measured by tests on rats.3. Vitamin A and carotene, riboflavin and vitamin C were measured by physical and chemical methods at intervals in the course of storage for a year.4. No significant difference was found between the biological values of the proteins (nitrogen) of the processed milks. The values ranged from 80 for evaporated milk to 84 for spray-dried milk.5. The proteins (nitrogen) of evaporated milk were significantly less digestible (true digestibility 91) than those of spray-dried milk (true digestibility 94). The difference between either of those and roller-dried milk (true digestibility 93) was not significant.

1. The experiments described in this part of the report were designed to obtain information about the effect of time and temperature of pre-test storage on test results and, in particular, about the possibility of a practical method of compensating test results to eliminate or minimize their dependence on the time and temperature of pre-test storage.2. Four separate experiments were planned and carried out in three widely separated areas in England and Wales. Samples of morning milk, brought to the laboratory with the minimum of delay, were divided into a series of subsamples which were subsequently stored at controlled temperatures ranging from 3 to 24°C, and at shade atmospheric temperature, for periods ranging from 7 to 30 hr. before examination by one or more of the following tests: keeping quality at 18, 22 and 24° C. using clot-on-boiling (C.O.B.), and precipitation with 68% alcohol (A.P.T.) or 50% alcohol; methylene-blue test at 37–5° C. and at 18, 22 or 24° C. The results for the different tests were the times to end-point after transfer from the storage to the incubation baths.

Summary Plate loop counts and standard plate counts on each of several manufacturing grade raw milk samples (handled in cans or in farm bulk tanks) have been compared. On the average, the plate loop count (PLC) was lower than the standard plate count (SPC) regardless of the type of handling of milk on the farm, can or bulk tank. Agreement between the SPC and PLC seemed to depend upon the bacterial-count levels present in milk. Statistical analyses indicated significant differences, at 1% level of probability, between the average bacterial count by SPC and PLC methods regardless of count level (≤100,000/ml or >100,000/ml) in case of can milk samples. On the other hand, in case of farm bulk tank milk samples, no significant differences, at the 1% level of probability, between the average bacterial count by SPC and PLC methods were obtained, when the counts were equal to or less than 100,000 per ml; when the counts exceeded 100,000 per ml, significant differences were present. Since the bacterial counts of manufacturing grade raw milk samples are likely to exceed 100,000/ml, the data presented in this investigation indicate that, until the bacteriological quality of manufacturing grade milk supplies undergoes substantial improvement, the PLC method does not appear to be a suitable substitute for the SPC method for routine bacteriological examination of such milk supplies.

The AOAC Roese-Gottlieb, Babcock, and Rapid Detergent (TeSa) Methods for the determination of butterfat in raw milk were compared in three separate collaborative studies, two involving identical samples sent to six laboratories (Studies I and 2) and one in which the laboratories collected their own samples (Study 3). Only in the case of the TeSa method in Study 3 was there a significant difference in the average butterfat content as found by the three methods. The TeSa test showed the highest standard deviation in Studies 1 and 2 and intermediate in Study 3. Significantly lower results were obtained by the TeSa and Roese- Gottlieb methods when analyses were repeated on successive days (—0.11 and —0.03%, respectively) in Study 2. The slope of the regression line of the TeSa test versus Roese-Gottlieb differed significantly from a 1:1 relationship in Studies 1 and 3. The TeSa test is recommended only as a rapid field test for use with raw milk less than 24 hours old.

SummarySince Indian law requires curds to be made from heat-treated milk, a means for distinguishing between curds made from raw and from heat-treated milks was sought. Curds contain whey proteins that are partly or completely denatured by heat treatment; the kinetics of the reaction are first order. The concentrations of native whey proteins in raw and heat-treated milks and in curds made from those milks were measured. No difference was found between the concentrations in heattreated milk and in the curds made from it, so that identical values were obtained for the half-life on heating and for the activation energy of denaturation. The acidity and microbial growth occurring during the production of curd did not affect the whey proteins. On electrophoresis, curd from raw milk gave a few whey protein bands, but curd from milk boiled for 10 min gave none, showing complete denaturation of the whey proteins.

Summary A selective plating medium for the isolation and identification of the enterococcus group of streptococci has been developed. The method is based upon the ability of the enterococci to utilize sodium citrate as an available carbon source, to convert ditetrazolium chloride to a blue diformazan and to grow in the presence of 0.01 per cent sodium azide. Results are presented which show that the selective plating medium can be used to isolate and estimate the numbers of enterococci in raw milk.

1. Samples of milk from 183 cows were examined, and seventeen (9·2 per cent.) yielded Br. abortus.2. Two hundred and two samples of butter, margarine, cheese, and icecream failed to yield Br. abortus, although samples of locally prepared butter and cheese both contained tubercle bacilli.3. Examination of the serum from slaughtermen and men working in allied trades showed some evidence of latent infection, but no indication of much real illness due to Br. abortus.4. Examination of the sera of sheep failed to show any evidence that these animals become infected with Br. abortus.5. The examination of sera and organs from pigs failed to show any evidence that the porcine type of Br. abortus occurs.

1. Clean milk has been “flash” pasteurised at 165°F. Portions of the same raw milk have been treated with small amounts respectively of starter culture and lactic acid, so as to raise the acidity, before being similarly pasteurised.2. Comparison of the whey obtained by rennetting the three raw portions has shown that those from the acidified milk contained more soluble protein than the original.3. The differences between the raw and pasteurised milk in their coagulation time with rennin have been found to be proportionately much greater in the acidified portions.4. These differences in coagulation time have been shown to be correlated with decreases in the amount of soluble protein found in. the whey separated from the coagulum.5. These experimental results, together with those of other investigators, have been made the basis of a suggestion that the decreased coagulability of heated milk with rennin is partly due to flocculation of soluble milk protein upon the casein micelles.

1. The results discussed in this part of the report were obtained by examination of milk samples taken at approximately fortnightly intervals throughout a period of one year from 298 farms in eight widely separated areas of England and Wales. The samples were examined by the routine resazurin test, the methylene-blue (4½/5½ hr.) test, the methylene-blue (½ hr.) test and the temperature-compensated keeping quality test using clot-on-boiling (c.o.b.) to determine the end-point. For the first three tests, the treatment of the samples and the standards applied conformed to those prescribed officially for these tests in England and Wales. Details are given of the treatment of samples and testing procedure for the temperature-compensated c.o.b. test. With this test two standards, equivalent to a residual keeping quality at 22° C. of 21 and 24 hr. from 9 a.m. on the day of collection of the milk from the farms, for both evening and morning milk were used.2. The results are analysed on the basis of the proportion of farms and of samples passing or failing the different tests, the effect of shade atmospheric temperature on the incidence of failures and the relationship between the standards imposed by the c.o.b. tests and the official methylene-blue (4½/5½ hr.) test.The proportion of farms and of samples passing each test was much higher in winter than in summer and was directly related to the prevailing shade atmospheric temperature. However, a proportion of farms was consistently able to comply with the standards. No more than 5% of the total failures by any test could be ascribed to the 60 (20%) best farms, whereas the 60 (20%) worst farms were responsible for as many as 37–60% of the total failures from the 298 farms.

Summary Two series of 20 samples each were prepared by adding 75mg. or 150 mg. of ascorbic acid to a liter of raw cow's milk. The samples were stored in 500-cc. flasks in the dark at 10° C. As aliquots were removed day by day for analysis, the volume of milk decreased and the volume of air in the flasks increased correspondingly. For the series of samples of milk to which 75mg. of ascorbic acid per 1. was added, the loss was 11 per cent per day for the first 3 days and 5 per cent per day for the remaining 7 days, or 7 per cent per day for the entire period. For the series of samples of milk to which 150mg. of ascorbic acid per 1. was added, the loss was 6 per cent per day for the first 4 days and 1 per cent per day for the remaining 6 days, or an average of 3 per cent per day for the 10 days of storage.

SUMMARY: A spore ‘spectrum’ is described of aerobic mesophiles capable of resisting different heat treatments. It is shown that B. licheniformis is the most common spore former found in bulk milk but since its spores are rapidly destroyed at 100°, the more heat resistant B. subtilis is the dominant surviving spore former in commercial sterilized milk. The thermal resistance of strains of B. subtilis and B. licheniformis isolated from different sources has been investigated and the strains of B. subtilis typed according to the behaviour of their spores when heated at 100°. All strains of B. licheniformis were destroyed more rapidly by boiling for 2 min than strains of B. subtilis but only those strains of the latter which showed some degree of heat activation were more resistant than B. licheniformis. The ‘resistant’ and heat activated strains of B. subtilis appear to be sparsely distributed in nature and were only isolated from sterilized milk where the heat treatment applied would tend to eliminate other strains. The spore content of bovine faeces was similar to that in bulk milk and the total spore content varied seasonally, the spore content of faeces being on the average a hundred times greater during indoor feeding than during the period when the cattle were fed outside. A faecal infection of the milk in the ratio of 1:104 would infect the milk with spores at about the same concentration as they are found in bulk raw milk, and it is suggested that bovine faeces could be a primary source of spore formers in milk supplies.

1. A method has been developed for measuring the intensity of colour produced in the Aschaffenburg & Mullen phosphatase test in which use is made of permanent colour standards.2. Using this method, the relationship between this phosphatase technique and the official Kay-Graham test has been investigated.3. The limiting standards corresponding to 2.3 Lovibond Blue Units in the KayGraham test have been established for the 30 min. and 2 hr. procedures of Aschaffenburg & Mullen.4. The advantages of the improved technique of the Aschaffenburg & Mullen test are discussed.

A total of 194 milk samples were obtained from 35 bulk tank grade-A milk producers in the Manhattan, Kansas, milk-shed during different seasons, and examined for standard, coliform, and psychrophile plate counts both before and after preliminary incubation (P.I.). Samples also were tested for inhibitory substances. Farms were ranked I, II, or III in general sanitation on the basis of visible conditions, with rank I most desirable. Except for one doubtful sample, all tests for inhibitory substances were negative. With standard plate counts, when no consideration was given to initial count level, there was no statistically significant relation between P.I. counts and season or sanitation rank, although summer growth ratios were always lowest. When data were grouped on the basis of initial count range, P.I. counts and growth ratios increased with increased sanitation rank in the lower count ranges. In initial count ranges > 50,000/ml the relationship between growth ratio and sanitation rank declined. With coliform counts, considerable inconsistency occurred in P.I. counts among seasons and sanitation ranks. Initial and P.I. coliforms counts showed little relationship with standard plate counts (except in summer) or with psychrophile counts. With psychrophile counts, growth ratios during P.I. were influenced by season, being highest in winter and lowest in summer. P.I. counts generally increased with increase in sanitation rank. The relation between growth ratios and sanitation rank was statistically significant (P

Summary The butyryl, caproyl, and caprylyl esters of 2-naphthol-6-sulfonic acid (sodium salt) have been synthesized and purified. These esters, particularly the butyryl, possess a reasonable degree of water solubility and are all hydrolyzed by some component of raw milk, presumably the lipase. 2-naphthol-6-sulfonic acid (sodium salt), one of the products of hydrolysis of these esters, can be estimated photometrically by using Folin and Ciocalteu's phenol reagent to develop color. A photometric method for estimating the lipase content of whole milk is presented which makes use of these esters as substrates and in which color is developed with the above mentioned reagent.

This epidemiological investigation revealed raw milk to be a source of mycobacteria, many of which resembled strains pathogenic for man and other animals. Eighty-three mycobacterial strains were isolated from 77 of 123 raw milk samples. The resemblance of these mycobacteria to known mycobacterial species and groups was as follows: 45, Mycobacterium phlei; 14, M. fortuitum; 1, M. smegmatis; and 23, slow growers (14 scotochromogens and 9 nonchromogens). Certain of the nonchromogenic slow growers may prove to be M. avium or M. bovis upon further bacteriological scrutiny.

During this investigation single samples were collected from 773 milk cans, three samples from each of 416 farm bulk tanks, six samples from each of 158 farm pick-up trucks, single samples from 49 storage vats, twelve samples from each of 98 transport trucks, and single samples from 77 storage tanks. Of the samples collected, 96% had counts under 200,000; 1.9% had counts of 200,000 – 300,000; 0.7% had counts of 300,000 to 400,000 and 1.4% had counts in excess of 400,000 per ml. The bacterial count finding on pumped milk indicated that milk passage through pumps did not appear to contribute to subsequent high counts through the break-up of bacterial clumps which may have been present. When bacterial counts were correlated with observations of the sanitary conditions of milk handling, the data reaffirmed that laxity in cleaning, bactericidal treatment or cooling will be reflected in the bacterial count and may result in the shipment of raw milk having counts in excess of 200,000 per ml.

A medium has been developed that permits the viable count of milk bacteria to be combined with the determination of biochemical properties likely to be important in milk itself. This has involved the modification of standard glucose-tryptone skim-milk agar by incorporation of two indicators to detect alkali as well as acid production, substitution of lactose for glucose, and increasing the quantity of skim milk for the detection of proteolysis and casein precipitation. The medium has proved particularly valuable in the study of the thermoduric flora of pasteurized milk. The phenomenon of casein precipitation is, however, less reliably determined than are changes in hydrogen-ion concentration and proteolysis.

Summary This investigation is concerned with the nature of bacterial inhibitory zones of milk, their incidence, and a technique for measuring the antibacterial property of raw milk. The disc assay test for antibiotics in milk was adaptable for the detection of natural inhibitors in raw milk. By substituting lyophilized milk tablets for the saturated paper discs, the sensitivity of the test was increased. Generally, raw milk produced two clearly defined zones, labelled primary and secondary, respectively, against Bacillus subtilis . A primary zone of inhibition first appeared after 12 hr. at 72° F. The secondary zone only appeared after the plates were incubated, refrigerated, and then reincubated. This secondary phenomenon caused disappearance of the bacterial colonies like that associated with lysis. It appears that the presence of the natural inhibitor in milk is independent of the breed of the cow. The initial zone size varied seasonally, and tended to decrease in diameter or disappear during the winter months, whereas secondary zones were less affected seasonally.