Search

The possibility of producing toxic effects in day-old chicks with mycotoxins from Phomopsis sp. or Diaporthe phaseolorum var. sojae isolates was studied. Evidence was found that Phomopsis sp., but not Diaporthe, produces hepatic necrosis with high mortality in chicks. Enzymatically, the hepatotoxin produced by Phomopsis sp. caused an increase in the level of liver glucose-6-P dehydrogenase. Mycotoxicosis caused by Phomopsis sp. was compared with lupinosis.

Extracellular accumulation of lysine and threonine was investigated in modified whey permeate by using Brevibacterium lactofermentum ATCC 21086 and Escherichia coli ATCC 21151. Whey permeate was prepared from whey by membrane ultrafiltration, and lactose was hydrolyzed by treating permeate with HCl or beta-galactosidase. The highest amount of lysine (3.3 g/liter) was produced from a mixture of acid-hydrolyzed whey permeate and yeast extract (0.2%). The highest amount of threonine (3.6 g/liter) was produced from a mixture of whey permeate, (NH(4))(2)SO(4) (1.4%), yeast extract (0.1%), and Na(2)CO(3) (0.3%).

Cholesterol metabolism by Mycobacterium species ATCC Number 19652 was studied in defined media. Whole cells were found to take up 91% of the total cholesterol when incubated five days at 34 degrees C in media of pH 6.8-7.4. Uptake of cholesterol by whole cells could be significantly inhibited by 2,4-dinitrophenol and dicyclohexylcarbodiimide. Growth media supernates as well as isolated microbial cell walls were found to contain cholesterol hydrolysing activity. This activity was extractable by Triton X-100 and appeared to have a molecular weight of approximately 100-200,000.

A study was conducted in which microwave irradiation and conventional waterbath treatment were compared as to their efficiency for heat-activating Bacillus spores. Spore suspensions were prepared from B. brevis, B. cereus, B. licheniformis, a lysogenic strain of B. megaterium (NRRL-B-3695), two strains of B. stearothermophilus, and B.Subtilis. Suspensions were either irradiated for 30 sec in a microwave oven, or conventionally heat-treated in the waterbath for 60 min at 60 degrees C, the serially diluted and plated onto nutrient agar. Colonies of each species from each treatment were isolated, and cultures were inoculated into several biochemical media. Spore suspensions heat-activated by microwave irradiation resulted in plate counts that were from 3% to 24% greater than from suspension heat-activated by conventional mean (60 degrees C for 60 min). There were no observed alterations in biochemical activities in any of the representative colonies from either of the two treatments. No induction of bacteriophage from lysogenic B. megaterium NRRL-B-3695 was observed in colonies from either of the two treatments. Microwave irradiation appears to be more efficient, less time-consuming, and at least as effective as heat activation by conventional waterbath treatment for Bacillus spores.

Beef loaf prepared according to four treatment options occurring in commissary food service was evaluated for microbiological and sensory quality. General acceptability was scored significantly (p of less than 0.01) higher for beef loaf portions held hot for 1 hour than for those held 2 hours, or those chilled 24 hours and heated in a convection oven. No significant (p of less than 0.01) differences for any quality characteristic scored were found between a loaf held hot for 1 hour and that chilled for 24 hours, heated, and then held hot for 1 hour. Appearance of beef loaves held hot for 1 hour was significantly (p of less than 0.05) better than that of loaves heated in the convection oven but was not significantly better than that of loaves given any of the other treatments. Microbiological analyses revealed the presence of relatively small numbers of microorganisms and no significant differences among treatments. The lowest total plate count was found in loaves baked, chilled for 24 hours, heated in a gas deck oven, and then held in insulated plastic trays for 1 hour. The highest counts were for loaves which were baked, placed immediately in insulated plastic trays, and held for 1 hour. Increases in total microbial counts and clostridial counts were observed as meat was held chilled for 24 hours. But no increases were observed when meat was held for 2 hours in insulated plastic trays. Although these microbiological findings were not significant, data may have important implications for an actual operation, where conditions are not so carefully controlled.

Use of a precooked frozen entree, hamburger patties, in a satellite foodservice system was assessed as related to time and temperature conditions and microbiological and sensory quality to identify critical phases in utilizing a precooked frozen entree. Time for product storage at the service location was approximately 3 days, and average time at room temperature during assembly was 2 h. Temperature conditions were generally variable with internal temperatures after heating food for service ranging from 152 F (67 C) to 192 F (89 C). Except for some of the low internal temperatures, conditions were acceptable from a food safety standpoint. Mean scores for sensory quality characteristics evaluated ranged from 5.1 to 6.9 (9-point scale) and total plate counts indicated that microbial quality was good. Genera of pathogenic microorganisms were identified, including Clostridium and Staphylococcus ; therefore, the potential exists for public health hazards if precooked food is subsequently mishandled in a system of this type. Critical problems may become apparent in the control of variability within the system, particularly, at the point of heating food for service.

The present study was conducted to determine the effects of metabolized aflatoxin B1 (via a mammalian liver mixed-function oxidase system) and native aflatoxin B1 upon induction of bacteriophage in Bacillus subtilis and growth of B. subtilis. A lysogenic strain of B. subtilis (BDS-1, phi 105) and an indicator strain of this species (DBS-1) were utilized in the present experiment. Lysogenic cultures were incubated for various lengths of time in the presence of either native or metabolized toxin, and plaque-forming units were determined. Identical experiments were conducted with the indicator strain and colony-forming units were determined. At a native toxin concentration of 25 micrograms/ml of medium, the maximum number of plaque-forming units was induced in lysogenic cells. However, when cells were incubated in the presence of aflatoxin B1 and mixed-function oxidase, neither plaque-forming units nor colony-forming units could be detected from lysogenic or indicator cells, respectively. This organism is apparently much more susceptible to inhibition of cellular replication than to lysis via bacteriophage induction. Thus, from the present study and from previously reported research, differences in susceptibility to native and metabolized aflatoxin B1 exist among species of Bacillus.

The distribution and metabolism of [(14)C]aflatoxin B(1) in chicken tissues were further investigated. Previously dried and frozen ethyl acetate extracts of liver, heart, gizzard, breast, leg, blood, and fecal samples were obtained from either layer or broiler chickens fed subclinical levels of [(14)C]aflatoxin B(1). Treatment of these extracts with either carboxypeptidase A, leucine aminopeptidase, pepsin, or trypsin revealed that an average of 50% of the (14)C detected in the acetate extracts was a liberated peptide (or amino acid) conjugate of [(14)C]aflatoxin B(2a). When a prepared standard of B(2a) was made by incubation of B(1) with cold dilute aqueous HCl, the R(f) values and absorbance maxima were identical with those of the tissue extracts after enzymatic treatment.

Attempts were made to use the osmotic shock procedure established for Gram-negative micro-organisms on a Gram-positive micro-organisms, Bacillus megaterium ATCC 19213. The procedure did not appear to shock the cells osmotically. The procedure, in fact, appeared to enhance binding of both glucose and fructose by the organism. The binding of glucose was greater by cells, whether grown on fructose or glucose, than it was for fructose. A procedure was developed for removing and isolating a protein capable of binding glucose. This involved solubilizing the cytoplasmic membrane of the organism with the nonionic detergent Triton X-100, and then isolating it by gel filtration on a Bio-Gel A 1.5 column with 0.05 M pyrophosphate containing 1% (v/v) Triton X-100 as the eluting buffer. This is the first isolation of such a protein in a Gram-positive organism. Isolated protein was demonstrated to have a specificity only for glucose. The protein did not bind to either the monosaccharides fructose and galactose, or the disaccharide sucrose. This was in comparison to whole cells which demonstrated ability to take up fructose. The protein was estimated to have a molecular weight between 24,000 and 30,000 daltons by gel filtration techniques.

To investigate the role of biotin in lysine production, Brevibacterium lactofermentum ATCC 21086 was grown in an acid-hydrolysed whey permeate medium with and without added biotin. Added biotin stimulated lysine production and growth of B. lactofermentum. Five micrograms of biotin/100 ml was the optimum level of addition. Biotin increased the uptake of 14C-glucose and affected fatty acid composition of cell wall lipids. Cell walls of test organisms contained less 16:0 and more 18:2 fatty acids than did those from control cells. However, biotin did not substantially affect the phospholipid content of cell walls and whole cells, and the intracellular free lysine level. It was concluded that the promotive effect of biotin in lysine production might be due to the stimulatory effect of biotin on the growth of B. lactofermentum. Alteration of the cell surface caused by biotin did not appear to affect the release or accumulation of lysine.

Aflatoxin production by a toxigenic strain of Aspergillus flavus was greatly reduced by benzoic acid and sodium benzoate in synthetic media. The reduction was accompanied by the appearance of a yellow pigment. Spectral analyses partially characterized this pigment as closely related to an acetyl derivative of a versiconal-type compound. A cell-free extract prepared from A. flavus grown in synthetic media was active in converting this yellow compound into aflatoxin B1 in the presence of reduced nicotinamide adenine dinucleotide phosphate at 25 degrees C (pH 7.4). In the presence of benzoic acid and its salt or autoclaved cell-free extract, conversion of yellow compound to aflatoxin B1 was prevented. These results suggest that the yellow compound is an intermediate in the secondary metabolic cycle involved in aflatoxin B1 production. Benzoic acid, sodium benzoate, or autoclaving the cell-free extract appear to have respectively blocked or denatured an enzymatic step late in the biosynthetic pathway of aflatoxin B1.

The effects of sodium chloride, sodium acetate, benzoic acid, sodium benzoate, malonic acid, and sodium malonate on growth and aflatoxin production by Aspergillus flavus were investigated in synthetic media. Sodium chloride at concentrations equivalent to or greater than 12 g/100 ml inhibited growth and aflatoxin production, while at 8 g or less/100 ml, growth and aflatoxin production were stimulated. At 2 g or less/100 ml, sodium acetate also stimulated growth and aflatoxin production, but reduction occurred with 4 g or more/100 ml. Malonic acid at 10, 20, 40, and 50 mM reduced growth and aflatoxin production (over 50%) while sodium malonate at similar concentrations but different pH values had the opposite effect. Benzoic acid (pH 3.9) and sodium benzoate (pH 5.0) at 0.4 g/100 ml completely inhibited growth and aflatoxin production. Examination of the effect of initial pH indicated that the extent of inhibitory action of malonic acid and sodium acetate was a function of initial pH. The inhibitory action of benzoic acid and sodium benzoate appeared to be a function of undissociated benzoic acid molecules. Aflatoxin reduction was usually accompanied by an unidentified orange pigment, while aflatoxin stimulation was accompanied by unidentified blue and green fluorescent spots but with lower Rf values that aflatoxins B1, G1, B2, and G2 standards.

Studies were conducted to determine possible interactions between aflatoxin B1 and deoxyribonucleic acid (DNA) and induction of bacteriophage formation in lysogenic Bacillus megaterium NRRL-B-3695. At pH 7.4, the spectrophotometric characteristics of B1 with various concentrations of calf thymus DNA were altered with both a hypochromic shift and a shift of the wavelength for maximum absorbance of the toxin. Aflatoxin B1 apparently interacted with DNA as indicated by spectrophotometric analysis and ultrafiltration studies. Under alkaline conditions (pH 10.0), spectrophotometric characteristics of this interaction were altered from those observed at pH 7.4. Toxin incubated at pH 7.4 could induce bacteriophage formation but failed to do so at pH 10. Preincubation of toxin with calf thymus DNA did not block subsequent induction. It is proposed that a bacterial DNA-toxin interaction, involving an alkaline-labile bond of aflatoxin B1, is necessary to cause induction of bacteriophage formation in lysogenic B. megaterium.

The effects of administering low levels of aflatoxin B(1)-(14)C by crop intubation daily for 14 days to broiler chickens were determined. Studies on the distribution of (14)C in the blood, selected organs, tissues, and excreta were conducted. No toxic effects were observed in broiler chickens during the 14 days of the experiment. The broiler chickens excreted 90.64% of the (14)C administered. Of the (14)C retained, 11.04, 9.83, 4.30, 12.52, 31.66, and 30.63% were detected in the blood, liver, heart, gizzard, breast, and leg, respectively. Chemical assay of those samples demonstrating radioactivity revealed that 81.2% of the radioactivity in these substrates was not extractable by classical extraction procedures while approximately 10% was extractable. Treatment of aqueous extracts for conjugated steroids by treatments with beta-glucuronidase revealed that 31.5% of the (14)C detected in the aqueous extract was a liberated glucuronide conjugate of aflatoxin M(1)-(14)C.