Showing papers in "Journal of Applied Microbiology in 1980"

Lipids in the Classification and Identification of Coryneform Bacteria Containing Peptidoglycans Based on 2, 4-diaminobutyric Acid

Abstract: Strains of 2, 4-diaminobutyric acid-containing coryneform bacteria were degraded by acid methanolysis and the non-hydroxylated fatty acid esters released examined by thin-layer and gas chromatography. The major fatty acid structural types were straight-chain, anteiso- and iso-methyl branched-chain acids. Polar lipids of the test strains were examined by two-dimensional thin-layer chromatography. All strains possessed very characteristic polar lipid patterns consisting of diphosphatidylglycerol, phosphatidylglycerol and a number of uncharacterized glycolipids. Menaquinones (vitamin K) were the sole isoprenoid quinones detected in the test strains. Corynebacterium insidiosum, Cor. michiganense, Cor. nebraskense and Cor. sepedonicum contained unsaturated menaquinones with nine isoprene units, whereas unsaturated menaquinones with 10 isoprene units predominated in strains of Cor. iranicum and Cor. tritici and a strain labelled Arthrobacter sp. The single strain of Cor. aquaticum examined contained comparable amounts of menaquinones with 10 and 11 isoprene units whereas strains of Cor. mediolanum and Flavobacterium dehydrogenans contained major amounts of menaquinones with 11 and 12 isoprene units. The results of the present study indicate that lipid markers may be of considerable value in the classification and identification of 2, 4-diaminobutyric acid-containing phytopathogenic and saprophytic coryneform bacteria.

Relationships Amongst the Fast-growing Rhizobia of Lablab purpureus, Leucaena leucocephala, Mimosa spp., Acacia farnesiana and Sesbania grandiflora and their Affinities with Other Rhizobial Groups

Abstract: Colony characteristics, growth in litmus milk, precipitation in calcium glycerophosphate medium and utilization of carbon sources of the root-nodule bacteria isolated from the tropical legumes Leucaena, Mimosa, Acacia, Sesbania and Lablab were similar to fast-growing rhizobia of temperate legumes, particularly Rhizobium meliloti. In agglutination tests, isolates from each host shared antigens with one or more of five Rhizobium strains from Leucaena. Infective characteristics of the fast-growing rhizobia were studied in modified Leonard jars and in agar culture. Cross-infections by rhizobia between these plants were common and the association often effective. Lablab was effectively nodulated by its own fast-growing isolate but only formed root swellings, possibly ineffective pseudonodules, with the other isolates. Slow-growing rhizobia which were able to nodulate Macroptilium atropurpureus were unable to form nodules on these legumes except Lablab which was considered more akin to the cowpea group. All fast-growing isolates nodulated, often effectively, Vigna unguiculata and V. unguiculata ssp. sesquipedalis. The isolate from Lablab also effectively nodulated a number of other tropical legumes which have previously only been reported to nodulate with slow-growing nodule bacteria and it also produced ineffective nodulation on Medicago sativa. This is the first record of an effective fast-growing isolate from Lablab.

The Attachment of Bacteria to the Gastric Epithelium of the Pig and its Importance in the Microecology of the Intestine

Abstract: Some characteristics of the association between lactic acid bacteria and pig squamous epithelial cells were studied. Strains from several sources were tested for adhesion in vitro but only those from pigs and chickens attached. The adhesion rate of pig isolates was very variable and, of the isolates tested, strains of Lactobacillus fermentum and Streptococcus salivarius attached in largest numbers. These strains were selected for further study. They did not attach to columnar epithelial cells from the small and large intestine. Adhesion was reduced by sodium periodate or protease. Both strains had a microcapsule with fibrils which stained with ruthenium red. The adhesive bond between lactobacilli and squamous tissue was strong enough to resist washing 50 times but there was a persistent release of bacteria during the washing process. When the strains of both species or of L. fermentum alone were fed to artificially reared pigs there was a statistically significant reduction in the numbers of Escherichia coli in the stomach.

A Note on the Separation of Natural Mixtures of Bacterial Menaquinones using Reverse Phase Thin-layer Chromatography

Abstract: Natural mixtures of bacterial menaquinones were separated according to the length and degree of saturation of the polyisoprenyl side-chain using ready made Merck RP-18F254 reverse phase thin-layer chromatography plates. The system described affords a simple and rapid means of menaquinone characterization.

Inhibition of growth and uptake processes in bacteria by some chemical food preservatives.

Abstract: The effect on growth and uptake processes of some common chemical food preservatives [benzoate, sorbate, propionate and alkyl esters of p-hydroxybenzoic acid (parabens)] has been studied in strains of Escherichia coli, Bacillus subtilis and Pseudomonas aeruginosa. For parabens. the inhibitory action on growth and amino acid uptake in whole cells and bacterial membrane vesicles followed similar dose-response curves. Growth inhibition caused by parabens appears to be a consequence of transport inhibition. For benzoate, sorbate and propionate, uptake inhibition seems inadequate to explain growth inhibition.

Fatty acid and isoprenoid quinone composition in the classification of Bacteroides melaninogenicus and related taxa.

Abstract: The fatty acid and isoprenoid quinone composition of 37 Bacteroides strains was examined. In a single strain of Bacteroides melaninogenicus subsp. levii neither menaquinones nor ubiquinones was detected. Menaquinones were the only isoprenoid quinones found in the remaining 36 Bacteroides strains examined. Unsaturated menaquinones with six isoprene units were the main components isolated from B. ochraceus. The menaquinone components of B. asaccharolyticus strains enable them to be divided into two subgroups containing menaquinones with nine and ten isoprene units respectively. Bacteroides vulgatus strains contained menaquinones with ten isoprene units as the major component, whereas B. fragilis, B. thetaiotaomicron and B. melaninogenicus subsp. melaninogenicus were characterized by comparable amounts of menaquinones with 10 and 11 isoprene units. Unsaturated menaquinones with 11 isoprene units were found in B. melaninogenicus subsp. intermedius and with 11 and 12 isoprene units in rumen strains of B. ruminicola subsp. brevis and B. ruminicola subsp. ruminicola. By contrast, oral strains of B. ruminicola subsp. brevis were characterized by major amounts of menaquinones with 12 and 13 isoprene units. Strains at present designated B. oralis, however, were heterogeneous with respect to menaquinone composition. The non-hydroxylated fatty acids of all the Bacteroides strains examined consisted predominantly of straight-chain, anteiso- and iso-methyl branched-chain acids. Monounsaturated fatty acids were found in only small amounts. The major fatty acid in B. ochraceus and B. asaccharolyticus strains was 13-methyltetradecanoic acid (iso C15) whereas 12-methyl-tetradecanoic acid (anteiso C15) predominated in B. melaninogenicus subsp. melaninogenicus. B. melaninogenicus subsp. intermedius. B. melaninogenicus subsp. levii, B. oralis and oral isolates of B. ruminicola subsp, brevis. The rumen strains of B. ruminicola subsp. brevis and B. ruminicola subsp. ruminicola were characterized by major amounts of anteiso C15 and pentadecanoic (C15:0) fatty acids. The accumulated lipid data correlate well with the major trends in Bacteroides taxonomy and appear to be of value in the classification of this group.

Effect of High Concentrations of Carbon Dioxide on Growth Rate of Pseudomonas fragi, Bacillus cereus and Streptococcus cremoris

Abstract: The maximum specific growth rates of Pseudomonas fragi, Bacillus cereus and Streptococcus cremoris were studied over a wide range of carbon dioxide concentrations. The growth rate compared with a control was reduced to 50% in Ps. fragi at 0–5 atm CO2, in B. cereus at 1—3 atm and in Strep, cremoris at 8–6 atm. B. cereus and Strep, cremoris were completely inhibited at 3 and 11 atm CO2, respectively. The growth rate of the aerobic Ps. fragi at 0–99 atm CO2 (0–01 atm oxygen) was reduced to about 20% of that in air. The growth rate of Ps. fragi was decreased at oxygen concentrations lower than 0–01 atm. When Ps. fragi was grown at oxygen limitation (0.0025 atm oxygen) and exposed to 0.99 atm CO2, the inhibiting effect of the CO2 was added to that of the oxygen limitation. No indications of a synergistic effect between CO2 inhibition and oxygen limitation were noted. B. cereus and Strep, cremoris were tested under anaerobic conditions.

Aerobic Metabolism of Brochothrix thermosphacta Growing on Meat Surfaces and in Laboratory Media

Abstract: Acetoin and acetic, isobutyric and isovaleric acids are major end-products, and important components of the spoilage odours, of Brochothrix thermosphacta growing aerobically on meat surfaces or in tryptone-based medium containing glucose, ribose or glycerol. Acetoin and acetic acid are probably derived entirely from the carbohydrates and isobutyric and isovaleric acids from valine and leucine respectively. Glucose and pH are both important factors in controlling the relative amounts of end-products, low glucose and near neutral pH favouring fatty acid formation, high glucose and lower pH values favouring acetoin formation.

New Strains of Bacillus licheniformis and Bacillus coagulans producing Thermostable α-Amylase Active at Alkaline pH

Abstract: Two species of Bacillus producing thermostable α-amylase with activity optima at alkaline pH are reported here. These organisms were isolated from soil and have been designated as Bacillus licheniformis CUMC 305 and B. coagulans CUMC 512. The enzymes released by these two species were partially purified up to about 81- and 72-fold respectively of the initial activity. The enzyme from B. licheniformis showed a wide temperature-range of activity, with optimum at 91°C. At this temperature it remained stable for 1 h. It retained 40–50% activity at 110°C and showed only 60% of its activity at 30°C. The enzyme showed a broad pH range of activity (4–10) retaining substantial activity on the alkaline side. The optimum pH was 9·5. The enzyme of B. coagulans showed activity up to 90°C, with optimum at 85°C and had a wide pH range with optimum at 7·5–8·5. The hydrolysis pattern of the substrate starch by these enzymes indicated that glucose, maltose, maltotriose and maltotetraose are the principal products rather than higher oligosaccharides.

Growth Factors of Bacterial Origin for the Culture of the Rumen Oligotrich Protozoon, Entodinium caudatum

Abstract: Attempts were made to develop an artificial medium suitable for axenic culture of Entodinium caudatum. Agnotobiotic cultures of the protozoon were established as stock cultures for testing the suitability of various growth media. A cell-free extract of mixed bacteria isolated from the rumen was shown to contain one or more growth factors for the protozoon when supplied with activated charcoal as a carrier. The medium (CYSE medium), which supported the growth of the protozoon in the presence of 50 μg/ml each of penicillin and chloramphenicol, consisted of activated charcoal (20 mg), heat-treated yeast (Y) (80 mg), 13%β-sitosterol-coated rice starch (S) (120 mg), and cell-free extract of rumen bacteria (1 ml) in 40 ml buffer solution. When culturing the protozoon, the CYSE medium was supplemented daily with 20 mg each of Y and S and half of the medium was replaced with fresh medium once every 5 d. The possible use of this method to establish an axenic culture of E. caudatum is discussed.

Yersinia enterocolitica in Danish pigs

Abstract: Yersinia enterocolitica serotype 0:3, the predominating pathogenic serotype in Danish pigs, was isolated consistently from the tonsils of pigs in six farms but not from those in another four farms during a one-year survey, indicating a herd-wise distribution. Only one positive culture was obtained from four specific-pathogen-free herds. The organisms were not recovered from samples of fodder, water and faeces from any of the infected farms. Strains of Y. enterocolitica were tested for sensitivity to antimicrobial agents.

Growth of bacteria on meat at room temperatures.

Abstract: The development of spoilage flora and the growth of individual psychrotrophs and pathogens on meat held at 20 or 30°C were studied. Under aerobic conditions psychrotrophic pseudomonads accounted for 60% of the spoilage flora at 20°C, but <20% at 30°C where they were displaced by species of Acinetobacter and Enterobacteriaceae which included both mesophilic and psychrotrophic strains. Mesophiles dominated the anaerobic spoilage flora at 30°C when clostridia were the major species, but at 20° the flora contained mesophiles and psychrotrophs in similar proportions and was dominated by Enterobacteriaceae. These results were largely predictable from the growth rate data for individual organisms. Interactions between species occurred more frequently at 20°C than at 30°C. When pathogenic species were grown at 20 or 30°Cin competition with equal numbers of psychrotrophic spoilage organisms, no interactions were observed. When pathogens were grown in competition with high numbers of psychrotrophs, only Lactobacillus growing anaerobically inhibited Salmonella typhimurium and Escherichia coli, but other pathogens were inhibited to varying degrees depending on the competing species and the incubation conditions. In general, the degree of inhibition was greater at 20 than at 30°C and facultative organisms were more susceptible under anaerobic than aerobic conditions. It appears that the cumulative stresses of low pH, suboptimal temperatures and competition with large numbers of saprophytic organisms can inhibit many of the pathogens likely to be present on meat. The organisms least affected by the conditions on meat surfaces, Salmonella and Esch. coli, are likely to be the main hazards on meat of normal pH held at room temperatures.

Studies on Acid Tolerance of Rhizobium trifolii in Culture and Soil

Abstract: Several experiments were carried out in an attempt to isolate and adapt strains of Rhizobium trifolii to growth at low pH on citrate-phosphate buffered yeast-extract mannitol agar (CPYEM). In another experiment, two streptomycin resistant strains (7A strr and 16 strr) with contrasting tolerances to acidity in culture were evaluated for survival in acid upland soils. No acid tolerant clones were isolated from three strains on CPYEM suggesting that there were no cells with greater tolerance of low pH present. The stepwise subculture of several strains on media of decreasing pH also failed to increase their acid tolerance. Growth initiation on CPYEM at pH 4–4 was a function of the number of viable cells in the inoculum and was accompanied by a rise in pH. In three acid soils, strain 7A strr was more persistant than 16 strr, whereas in culture the converse was true. Survival of these strains in the acid soils was inversely related to acid production on the medium of Norris (1965) in agreement with Norris's hypothesis. Ameliorating soil pH with lime improved the persistency of both strains. Prior exposure of 7A strr and 16 strr to the acid soils failed to improve subsequent survival in these soils.

Lactose Hydrolysing Enzymes in Streptococcus lactis and Streptococcus cremoris and also in some other Species of Streptococci

Abstract: Two types of Streptococcus lactis could be identified: cheese starter strains, which contain β-phosphogalactosidase and ferment lactose rapidly to lactate, and non-dairy strains, which contain both β-galactosidase and β-phosphogalactosidase and ferment lactose slowly to a variety of end products. All strains had homolactic glucose fermentations and heterolactic galactose fermentations. Other species of streptococci were examined for lactose hydrolysing enzymes and found to contain β-phosphogalactosidase, except Strep, thermophilus and Strep. faecium which had high levels of β-galactosidase. Discrepancies were found in the lactose hydrolysing enzymes content when the cells were treated in different ways.

The Occurrence of Bacillus cereus in some Dried Foods Including Pulses and Cereals

Abstract: In a survey of 39 dried food samples which represented 12 different pulses and cereals. 22 (56%) were found to be contaminated with Bacillus cereus. The numbers varied between 1 times 102 and 6 times 104 organisms/g. During normal cooking procedures and on storage at room temperature, the B. cereus resident on red lentils and kidney beans increased to a level at which enterotoxin production could become significant. Some physiological characteristics including deoxyribonuc-lease (DNase) and ribonuclease (RNase) secretion and salicin fermentation did not correlate with the ability or otherwise of a strain to cause food poisoning. Nevertheless, serotype 8 strains were prevalent on these foods and these have been implicated in both the diarrhoeal and vomiting-type food poisoning syndromes.

Aerobic Deterioration of Wheat, Lucerne and Maize Silages Prepared with Lactobacillus acidophilus and a Candida spp.

Abstract: Aerobic deterioration of lucerne, maize and wheat silages was characterized by rapid increases in yeast and mould flora which oxidized lactic and volatile acids resulting in increased temperature and pH. While populations of yeasts and moulds were similar, temperature increases were slightly greater for silages inoculated with Lactobacillus acidophilus and Candida spp. After 48 h the pH of the inoculated silages was higher in general and concentrations of acids were lower than controls. Bacterial growth was slight although continued lactic acid production was probable. In contrast to lucerne and maize silages, the pH of wheat silage remained stable during this period because of high butyric levels, but temperature and yeast populations increased. After 48 h the pH rose above 5 in maize and lucerne, and bacterial growth and metabolic activity resumed resulting in volatile and non-volatile acid production from carbohydrate fermentation and deamination of amino acids. During this phase of aerobic deterioration yeast growth slowed or stopped, but temperatures remained high and pH continued to climb probably because of production of ammonia. The changes in gross composition of the silages did not follow any particular pattern. Losses in dry matter were small (2.5–4.0%) and changes in individual components probably reflect this loss rather than substantial changes. Protein availability in the lucerne silages undoubtedly decreased, as protein losses were high. It is concluded that the aerobic deterioration of silage is enhanced by the addition of L. acidophilus and Candida spp. at ensiling.

A Note on Scanning Electron Microscopy of Micro‐organisms associated with the Kefir Granule

Abstract: Scanning electron microscopy was used to examine the micro-organisms on kefir granules. The most richly colonized part was that nearest to the exterior of the granule; it contained mainly bacteria and a few yeasts. The latter increased towards the centre of the granule where few bacteria were present. The kefir grain has a spongy fibrillar structure with a reticular lamellar matrix and a fibre mass which, especially in the centre of the grain, shows branching and interconnections with long cordons.

Pseudomonas tolaasi in Mushroom Crops: A Note on Primary and Secondary Sources of the Bacterium on a Commercial Farm in England

Abstract: Primary sources of Pseudomonas tolaasi Paine on a mushroom farm were the peat and limestone used in the casing process. The pathogen could not be detected in the farm soil, water supply, the mushroom spawn used, or in compost after spawning, but was isolated from the casing (peat/limestone mixture) layer of symptom-free mushroom beds and both the casing layer and compost of beds bearing diseased mushrooms. Secondary sources were numerous once the pathogen was present in mushroom beds. These included symptomless and diseased mushrooms, the fingers and shoes of people handling the crop, their baskets, knives and ladders. Ps. tolaasi could be isolated from dust in the air in infected houses and also from floors. Spores of infected mushrooms may transport the bacterium, as did sciarid flies and mites which are common pests of mushroom crops.

Comparison of the Effects of Liquid Medium Repair and the Incorporation of Catalase in MacConkey Type Media on the Recovery of Enterobacteriaceae Sublethally Stressed by Freezing

Abstract: Nine pure cultures of species of Enterobacteriaceae were stressed by rapid freezing in tryptone soya broth (TSB) to — 22°C and subsequent storage at that temperature for 7 d. About one to two log cycles kill and at least one additional log cycle sublethal impairment was achieved. Numbers of colonies of these cultures in poured plates of violet red bile glucose (VRBG) agar, with 67 u/ml of catalase added at 47°C, were only slightly higher than those in plain VRBG, both incubated overnight at 30°C. Two hours incubation of TSB suspensions at 17–25° C resulted in almost complete restoration of the ability of cells to develop colonies in VRBG, without, however, leading to any significant multiplication. Similar experiments with 32 samples of frozen minced meat, 27 samples of frozen surface water, 18 of frozen chicken liver and 14 of fresh sausage substantiated the results obtained in the studies on pure cultures. In the experiments with the nine pure cultures the influence of the nutrient composition of the solid enumeration media: ‘minimal’ agar, TSB agar (TSBA) and Mueller-Hinton agar with Polyvitex nutrient supplement (MHA), on the recovery of Enterobacteriaceae stressed by freezing was also studied. Colony numbers in TSBA and MHA were virtually identical. The glucose mineral salts medium led to lower recovery, indicating that so-called ‘minimal medium recovery’ of stressed bacterial populations is not a common phenomenon.

Soft rot Erwinia bacteria in the atmospheric bacterial aerosol.

Abstract: Using a Casella High Volume Airborne Bacteria Sampler, the soft rot coliform bacteria Erwinia carotovora var. carotovora and E. carotovora var. atroseptica were caught from the open air during rainfall in mid to late summer, autumn and early winter, but not in late winter, spring or early summer. They were not found every time it rained, and never when the weather was dry. The bacteria were caught close to potato crops, but there were several occasions when they were found at sites where there were no such crops or at times of year after potato crops had been harvested. The sources of the organisms are uncertain, but those obtained close to potato crops may have originated there. The results are considered to provide further evidence that airborne spread and subsequent deposition of viable bacteria could cause contamination of Erwinia-free potato stocks, and suggest that rainfall is a major generator of the general atmospheric bacterial aerosol.

A note on bile acids transformations by strains of Bifidobacterium.

Abstract: The hydrolysis of sodium taurocholate and glycocholate was a common feature among 52 strains from 14 species belonging to the genus Bifidobacterium. Forty-eight strains were able to hydrolyse both these conjugated bile acids, yet four strains failed to split the amide bond of either. Twenty-eight strains were checked for the ability to transform sodium cholate, chenodeoxycholate, deoxycholate and lithocholate; only 13 of these strains formed minimal quantities of monochetoderivatives from cholic acid, while none of them was able to transform the other tested bile acids.

Effects of Sulphate, Iron and Hydrogen on the Microbiological Conversion of Acetic Acid to Methane*

Abstract: Sulphate stimulated the rate of conversion of acetic acid to methane in enriched cultures as well as in liquid from anaerobic fermenters fed with wastes, but the effect depended on the soluble iron content of the culture. At low soluble iron contents, i.e. 0–02–0–05 mM, sulphate had little or no immediate (after 7–10 d) effect, but at 0–5 mM or higher soluble iron content, sulphate (0–6 mM or more) significantly increased the rate of acetic acid conversion over that obtained with 0–5 mM soluble iron without sulphate. Long-term continuous culture tests (100–160 d) showed that 0–6 mM or more sulphate increased the maximum growth rate of the culture from 0–07–0–08 d without sulphate to 0–14–0–16/d, even at iron levels of 0–02–0–05 mM. Hydrogen was inhibitory to the conversion of acetic acid to methane under all conditions tested, but stimulated the reduction of sulphate when present.

The Effect of Hydrogen Peroxide and Ultraviolet Irradiation on Non-sporing Bacteria

Abstract: A kill of 99.99% was obtained in cell suspensions of Escherichia coli and Streptococcus faecalis by incubation with hydrogen peroxide 1.0% (w/v) for 75 and 180 min respectively. The same kill was produced by 30 s irradiation with ultraviolet (u.v.) light in the presence of hydrogen peroxide 1.0% (w/v). This simultaneous treatment with u.v. and hydrogen peroxide produced a synergistic kill at least 30-fold greater than that produced by irradiation of cell suspensions of Esch. coli with or without subsequent incubation with hydrogen peroxide.

Hydroxylations of Biphenyl by Fungi

Abstract: Sixty-six species of fungi were screened for their ability to hydroxylate biphenyl. Thin-layer and gas chromatographic analyses indicated that seven organisms actively hydroxylated this substrate. Of particular interest were Cunninghamella echinulata ATCC 9244 which produced 4-hydroxybiphenyl as the major metabolite, Helicostylum piriforme QM 6945 which produced exclusively 2-hydroxybiphehyl, and Aspergillus parasiticus ATCC 15517 which produced a good yield of 4, 4′-dihydroxybiphenyl. These results were confirmed by isolation of the phenolic metabolites following preparative-scale conversions and comparisons of the isolated phenols with authentic reference materials.

Quality control of solid culture media: a comparison of the classic and the so-called ecometric technique.

Abstract: Some selective culture media, prepared from commercially available dried preparations function poorly, and the same applies to an occasional non-selective culture medium. This calls for systematic monitoring of media before they are used in actual diagnostic work. Classic plating or dilution-to-extinction techniques are often found too cumbersome. Hence a simple streaking technique (‘ecometric’ evaluation) was developed earlier for this purpose. It was subsequently simplified further and its accuracy and precision were assessed in this study. After it had been found that the simplified ecometric procedure allowed a reasonably accurate analysis of the selective and productive properties of media, it was used to evaluate 16 selective media, currently used in food microbiology. Results obtained agreed well with observations on the functioning made during routine examination of various foods. Finally an attempt was made to assess whether ecographic inoculation would allow the study of micro-organisms under repressive conditions, particularly refrigeration temperature. It appeared that this was indeed possible.

The Structure and Function of the Spore Outer Membrane in Dormant and Germinating Spores of Bacillus megaterium

Abstract: Attempts to demonstrate the presence of the spore outer membrane in mature, dormant spores of a strain of Bacillus megaterium are described. The outer, integument, layers of this organism were found to contain one-third of the total spore cytochrome content, several enzymes of the electron transport chain (specifically NADH oxidase, dehydrogenase, cytochrome c reductase and NADPH dehydrogenase) and a large number of polypeptides extractable with sodium dodecylsulphate in the presence of dithiothreitol and protease inhibitors. These all suggest the presence of a membraneous element. Electron microscopic evidence is presented on the structure of the dormant integument enzymes. Changes in the integument enzymes and in the gel electrophoresis profile of the extractable integument polypeptides which occur during spore gemination, are described and compared with those that take place in the spore inner membrane. The heat sensitivity of the integument enzymes is compared with that of the inner membrane enzymes and the implications for theories of spore heat resistance discussed.