Roll-tube Method Research Articles

Isolation of Anaerobic Bacteria from Human Gingiva and Mouse Cecum by Means of a Simplified Glove Box Procedure1

An anaerobic glove box constructed of clear flexible vinyl plastic is described. It is sufficiently inexpensive and simple in operation to be used not only in research but also in a clinical laboratory by technicians without special training. Conventional bacteriological techniques may be used inside the glove box for culturing and transferring anaerobic bacteria. The box may be heated to 37 C and thus serve as an anaerobic incubator as well, permitting inspection of cultures at any time. Media may be prepared and agar plates may be poured on the laboratory bench in the conventional manner. An overlay of trace amounts of palladium black catalyst over plated agar media reduces the medium to an oxidation-reduction (O-R) potential of - 300 mv within 2 days after introduction into the glove box. In spite of its greater simplicity, the system matched or excelled the roll tube method with respect to all parameters tested, including O-R potential obtainable in the media, O(2) concentration in the gas phase, and efficiency in isolating anaerobic bacteria from the mouse cecum. Comparative studies indicate that the conventional anaerobic jar method was inadequate for the isolation of strict anaerobes from human gingival specimens and from the mouse cecum. This was due to the exposure of specimens and media to air during plating on the open laboratory bench. Anaerobic jars were adequate for maintaining the proper conditions for growth of anaerobic bacteria once these had been established in the glove box.

INVESTIGATIONS ON THE USE OF THE ROLL TUBE METHOD FOR COUNTING ESCHERICHIA COLI I IN WATER

SUMMARYThe roll tube method was used for the enumeration of Escherichia coli I in water and was found to be subject to less variation than the accepted method. Different media were compared and it was found that MacConkey's broth containing 3% of agar gave the best results. Pre‐incubation at 37% for 2 hr improved the selectivity and gave higher colony counts.

THE REMOVAL OF BACTERIA FROM GLASS SURFACES WITH CALCIUM ALGINATE, GAUZE AND ABSORBENT COTTON WOOL SWABS

SUMMARY: Calcium alginate wool swabs were no more efficient or reliable than gauze or absorbent cotton wool swabs when tested on drinking glasses infected with known numbers of bacteria. The number of bacteria left on the glass after swabbing was determined by a modified roll tube method.

Device for preparing roll-tube cultures

The roll-tube method for estimating bacterial counts was devised by Esmarch(1), but was soon afterwards superseded by the introduction of Petri plates which have several advantages for cultural and counting purposes. The roll-tube method, however, has been revived by several workers (2,3,4), and where a simple, rapid and economical method is required it is very useful, especially if low counts are being dealt with. For this reason it is particularly suitable for yeast and mould counts in butter and has been so used in this laboratory for many years. In addition to its regular use for this purpose, several series of comparisons have shown that the roll-tube method is capable of giving results which compare very well with Petri plate counts upon milk, using the usual dilution technique in both cases.

The Drying and Preservation of Bacterial Cultures

SUMMARY: Working details are given for keeping as desiccates a collection of some 1500 strains of bacteria. The efficacy of drying, the various methods of freezing, and the effect of storage were tested by viable counts, using a spinning bottle modification of the roll-tube method. The survival rate of bacteria suspended in broth or other protective colloids, and subjected to freezing at − 78°, varied with the species from about 100 % with the resistant Staphylococcus aureus to about 10 % with the sensitive Neisseria gonorrhoeae, and from about 100 to 1·5 % or less, respectively, when the organisms were suspended in saline. The percentage of organisms which survive the freeze-drying process was found to vary with the species, from 100 % to less than 1·0 % when the organisms were suspended in broth and from 100 to no survivors when the organisms were suspended in saline. The storage loss of dried cultures was found to be a function of the storage temperature; however, suitably dried cultures could be kept at room temperature for very long periods. By far the most important factor influencing loss on storage was the presence of traces of moisture, and to ensure optimal survival the cultures must be as dry as possible. It appears that even with adequately dried cultures the presence of oxygen is deleterious. The survival rate of bacteria in a sample of dried culture after heating to 80° for 1 hr., determined by plating a suspension of the heated sample in broth, provides a simple measure of the capacity of the particular batch of the dried culture to remain viable on storage at ordinary temperatures.

229. Bacteriological testing of butter. New and simplified routine methods

Simple and rapid methods have been devised which utilize the minimum of apparatus and material for the routine bacteriological testing of butter. These involve the preparation of microplate cultures to obtain counts of total bacteria, “heat-resistant” bacteria, lipolytic organisms, yeasts, moulds; Grimes's method as a measure of the coliform content is also used.The method for counting colonies of lipolytic bacteria is based upon their ability to produce round each colony a halo of white solid fatty acids in place of the clear glistening fat globules.The new methods have been exhaustively compared with those hitherto accepted, and have been found to give satisfactory and reliable results.A variety of comparisons has been carried out to improve still further the methods in use and to provide additional information about the nature of the bacteria contamination present in the butter. The roll-tube method seems preferable to the microplate for mould counts, and is quite suitable for yeast counts.The results obtained confirm the view expressed by Wood & Thornton(6) that yeast and mould counts provide an inadequate criterion of factory sanitation, and should be supported at least by the total count of bacteria.The employment of these methods has enabled large numbers of samples to be dealt with. Thus samples from most of the principal New Zealand butter factories have been regularly tested. The information so obtained has been of material value to the Dairy Division's Instructors in drawing attention to factories where sanitary conditions left room for improvement.