Methylene blue in the absence of oxygen is widely used to measure ('reductase" activity of bacteria cells or of cell-free enzyme preparations. A modification of the "methylene blue technique," frequently called the '' reductase test," has long been applied to milk containing dissolved 0, as a means of estimating its sanitary history and keeping quality. The test is usually considered an indirect quantitative index of the viable bacterial content at the start of incubation. The substitution of resazurin for methylene blue was recommended by Pesch and Simmert (1928) and has since been used by many milk control laboratories.Recently Scott and Gillespie (1943, 1914) applied the resazurin test to the quality grading of egg "pulp" prepared in various Australian states and found that the rate at which resazurin is reduced to pink color afforded a reliable index of the bacterial content of the pulp. Berry and Wolford (1945), using the technique described in this paper, examined stored frozen-egg emulsions and obtained reduction values approximating those of Scott and Gillespie. Johns (1944) investigated the usefulness of the reductase test in a study of the microbiology of egg powder using methylene blue and resazurin, and reported that the tests involved endpoint difficulties and long incubation periods.Preliminary investigations on the application of the resazurin-dye test to inoculated egg liquid and powder indicated that the rate of dye reduction by inoculated lyophilized powders was not related to the plate count of such powders but appeared to be related to the number of viable bacteria in the melange prior to drying. The object of the work reported here, therefore, was to investigate the effect of dehydration by lyophilization and of storage on the rate of resazurin-dye reduction by inoculated and uninoculated egg emulsions in relation to their viable count. METHODSThe eggs used were U. S. Grade A. The shells were broken aseptically by the method tentatively suggested f o r shell egg in "Standard Methods for the Examination of Dairy Products" (1941). Half of each lot of egg emulsion was inoculated with Pseudomonas f l .~o r e s c e n s .~ This organism was University. The organism was cultivated in broth containing 0.3 per cent beef extract, 0.5 per cent peptone, 0.5 per cent dextrose, and 0.1 per cent yeast extract, which was incubated at 3OoC.(86"F.) for 48 hours and centrifuged: the cells from 200 ml. of broth were resuspended in 2.5 ml. of sterile distilled H20. The organisms from 1200 ml. of broth were used t o inoculate each lot of pulp collected from 12 eggs. 372
GROWTH OF Trich;op/ayto.Fz interdigitde ON WOOL 729 for agglutinins. Of the 8 carp that survived until the tenth week, at death, 2 had titers of 120, 2 had titers of 1 :@, one had a titer of 1 $30, and one of 1 640 (see table). The remaining fish died during the first 2 weeks of the experiment and had no agglutinins.In earlier experiments it has been shown that carp in nature do not have these agglutinins.The other fish, adult rainbow and brown trout, held in 10°C running water, were subjected to weekly injections of 1 ml of Bwt. sdnzlonicida4 vaccine containing approximately 5 billion heat-killed virulent bacteria. Because of the conditions under which they were held, few trout survived longer than 5 weeks, and most of them died during the second and third weeks. Post mortem blood samples taken after the first week showed agglutinins, the results being recorded in the table. It will be noted that a number of trout had no agglutinins; but most of these died early in the study. Three of 74 control animals were shown to have titers as high as 1:80.This does not necessarily indicate the presence of natural agglutinins. The presence of agglutinins for Bmt. salnzonicidu in these trout, which had received no vaccine injections, is more readily explained by the fact that these animals have been exposed to trout furunculosis throughout most of their several years of life. It is very possible that the antibody has been developed in response to this natural exposure of the trout to the infection.These experiments indicate that agglutinin production takes place at 10°C. It is believed that earlier investigators failed to detect this phenomenon because their animals were not given the antigen over a long enough period of time.Experiments are now in progress with gold fish, which tolerate extremes of teniperature, to determine the effect of temperature on the rate of agglutinin production.
Little work appears to have been done on the influence of bacteria on the loss of powder quality as determined by the usual quality tests of palatability, fluorescence, solubility, etc. DeBord (1925) noted that a somewhat rancid odor developed in powders during storage, particularly when those powders were prepared from low-grade eggs and stored at 37°C. (98.6"F.). He stated that the counts of viable bacteria furnished little basis for estimating the quality of the product. Thistle, Pearce, and Gibbons (1943) found that moisture content and bacterial plate count are independent measures of quality, and that the slight degree of association between bacterial count and palatability in prime-quality powders was presumably fortuitous. Gibbons and Fulton (1943) stated : "Although there is as yet no direct evidence that any correlation exists between bacterial content and quality of egg powders, the number of bacteria gives some indication of sanitation in the . . . processing plants. " Recently Stuart, Goresline, Smart, and Dawson (1945) showed that spray-dried egg powder of low sanitary quality decreased in solubility during storage to a greater extent than powder of high sanitary quality.Marked differences in deteriorative changes during storage, as measured by the usual quality criteria, in commercially spray-dried and in lyophilized whole-egg powders have been observed in this laboratory. Some of the differences in storage behavior were quite probably related to differences in drying procedures and some to the quality of the shell egg used. It is obvious that microbial growth in the egg liquid might cause undesirable changes which would affect the quality of the powdered product. It is reported in a companion communication by Hirschmann and Lightbody (1947) that catalysts or reducing substances of microbial origin survived lyophilization and retained the power to reduce resazurin dye. It appeared possible that these agents might accelerate changes and shorten storage life of egg powder. The object of this study, therefore, was to determine whether large numbers of Pseudomonas jluorescem, incorporated in egg liquids prior to drying by lyophilization, contributed to deterioration of the low-moisture powders during storage, as estimated by the usual criteria of quality. METHODSLiquid egg pulp was collected aseptically and one half of each lot inoculated with Ps. fluorescens. The liquids were dried by lyophilization, and the powders stored in sealed glass containers. Details concerning inoculum, distribution, lyophilization, and storage and the methods used for moisture
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.