The spore forming Bacillus cereus (66) was screened for the induction of β-lactamase in presence of an inducer using iodometric assay. A significant induction in marker enzyme was observed in B. cereus 66 at maximum residual limit (MRL) of penicillin, ampicillin, cloxacillin, amoxicillin, cefalexin, and cephazolin belonging to β-lactam group of antibiotics. A microbial based assay, where enzyme induction was optimized at pH 7.0, temperature 30°C, and whey powder (0.25%) after 4 h of incubation. The spore based assay was tested with milk samples spiked with 6 different β-lactam antibiotics. The results were 100 and 83.33% in correlation with microbial receptor and inhibition based assay, respectively. Overall, spore based assay can be a useful and cost effective tool for the specific detection of β-lactam group of antibiotics in milk.

Aim:To investigate the effect of oral administration of two Bacillus strains on fecal coliforms, Lactobacillus and Bacillus spp. in rat animal model.Materials and Methods:An in vivo experiment was conducted for 49-day period on 36 adult male albino Wister rats divided equally into to four groups. After 7-day adaptation period, one group (T1) was fed on sterile skim milk along with basal diet for the next 28 days. Second (T2) and (T3) groups received spore biomass of Bacillus coagulans B37 and Bacillus pumilus B9, respectively, suspended in sterilized skim milk at 8-9 log colony-forming units/ml plus basal diet for 28 days, while control group (T4) was supplied with clean water along with basal diet. There was a 14-day post-treatment period. A total of 288 fecal samples (8 fecal collections per rat) were collected at every 7-day interval starting from 0 to 49 days and subjected to the enumeration of the counts of coliforms and lactobacilli and Bacillus spores using respective agar media. In vitro acid and bile tolerance tests on both the strains were performed.Results:The rats those (T2 and T3) received either B. coagulans B37 or B. pumilus B9 spore along with non-fermented skim milk showed decrease (p<0.01) in fecal coliform counts and increase (p<0.05) in both fecal lactobacilli and Bacillus spore counts as compared to the control group (T4) and the group fed only skim milk (T1). In vitro study indicated that both the strains were found to survive at pH 2.0 and 3.0 even up to 3 h and tolerate bile up to 2.0% concentration even after 12 h of exposure.Conclusions:This study revealed that oral administration of either B. coagulans B37 or B. pumilus B9 strains might be useful in reducing coliform counts accompanied by concurrent increase in lactobacilli counts in the intestinal flora in rats.

Spore germination based assay involves the transformation of dormant spores of Bacillus stearothermophilus 953 into active vegetative cells. The inhibition of germination process specifically in presence of antibiotic residues was used as a novel approach for monitoring target contaminants in milk. The indicator organism i.e., B. stearothermophilus 953 was initially allowed to sporulate by seeding in sporulation medium and incubating at 55 °C for 18 ± 2 h. The spores exhibited a typical chain behavior as revealed through phase contrast microscopy. The minimal medium inoculated with activated spores was incubated at 64 °C for 2-3 h for germination and outgrowth in presence of specific germinant mixture containing dextrose, whey powder and skimmed milk powder added in specific ratio along with reconstituted milk as negative control and test milk samples. The change in color of the medium from purple to yellow was used as criteria for detection of antibiotic residues in milk. The efficiency of the developed assay was evaluated through a surveillance study on 228 samples of raw, pasteurized and dried milks and results were compared with AOAC approved microbial receptor assay. The presence of antibiotic level was 10.08 % at Codex maximum residual limit having false positive result only in 0.43 % of the samples. The results of the present investigation suggest that developed spore based assay can be a practical solution to dairy industry for its application at farm level, milk processing units, independent testing and R & D centres in order to comply with the legal requirements set by Codex.

An experiment was conducted for the 56 d period in thirty six adult male albino Wister rats divided equally into four groups. After 7 d adaptation period, all the groups were fed with cholesterol enriched diet for 14 d. Thereafter, one group (T1) was fed on sterile skim milk along with cholesterolemic diet for the next 21 d. The two experimental groups (T2 and T3) received cholesterolemic diet plus spore biomass of B. coagulans B37 and B. pumilus B9, respectively, suspended in sterilized skim milk @ 8-9 log cfu/ ml for the next 21 d. The control group was supplied with clean water along with cholesterolemic diet for 21 d. All the animals were fed on the basal diet along with water during a 14 d post-treatment period. After the adaptation period, the blood samples were collected weekly from the experimental rats up to 42 d, and plasma separated, and subjected to the determination of different plasma lipid fractions. The oral administration of both B. coagulans B37 and B. pumilus B9 resulted a decrease (P Less Than 0.05) in plasma cholesterol, low density lipoprotein (LDL)-cholesterol concentrations and atherogenic index in the experimental rats.

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The present study explored the possibilities of using Bacillus coagulans as a probiotic culture in vacuum-dried milk powder. The operational drying temperature at 63 AE 2°C under 0.7 kg/cm 2 pressure emerged as the best temperature at which to prepare dried milk powder within 4.5 h with a mean (AESEM) Bacillus coagulans B37 spore count of 8.78 AE 0.03 log cfu/g and moisture content of 4.82 AE 0.12%. The total spore counts in vacuum-dried milk powder did not change (P > 0.10) at storage temperatures of 7°C, 37°C or 45°C over a three-month period.

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