ATP-Bioluminescence as a method to evaluated microbiological quality of UHT milk

New approaches are needed to quickly indicate possible contamination of UHT milk, among them the technique of ATP-Bioluminescence. Therefore, the aim of this study was to compare the results of culture methods with the results of ATP-Bioluminescence technique of 102 UHT whole milk samples incubated at 48, 72, and 168 hours. UHT milk samples were analyzed for the presence of mesophilic and psychrotrophic aerobic microorganisms using Plate Count Agar (PCA), Brain-Heart Infusion (BHI) media and PetrifilmTM Aerobic Count (AC) plates. The ATP-Bioluminescence technique was applied through the Microbial Luminescent Screening (MLS) system. Significant correlations were found between counts of aerobic mesophilic microorganisms on PCA, PetrifilmTM AC, BHI and results of ATP bioluminescence technique (P≤0.05). The ATP-Bioluminescence technique had higher correlation with counting method in PCA than BHI media. At lower pass/fail limits of Relative Light Units (60, 50, 45 and 40 RLU), the number of samples identified as positive increased and statistically agreed with aerobic mesophilic microorganism counts (P>0.05). For the dairy industry, the ATP-Bioluminescence technique may become an important tool that assists the official methods to quickly monitor the microbiological quality of UHT milk though this will likely require a threshold below 150 RLU.

A Gene from the Mesophilic Bacterium Dehalococcoides ethenogenes Encodes a Novel Mannosylglycerate Synthase

ABSTRACT Mannosylglycerate (MG) is a common compatible solute found in thermophilic and hyperthermophilic prokaryotes. In this study we characterized a mesophilic and bifunctional mannosylglycerate synthase (MGSD) encoded in the genome of the bacterium Dehalococcoides ethenogenes. mgsD encodes two domains with extensive homology to mannosyl-3-phosphoglycerate synthase (MPGS, EC 2.4.1.217) and to mannosyl-3-phosphoglycerate phosphatase (MPGP, EC 3.1.3.70), which catalyze the consecutive synthesis and dephosphorylation of mannosyl-3-phosphoglycerate to yield MG in Pyrococcus horikoshii, Thermus thermophilus, and Rhodothermus marinus. The bifunctional MGSD was overproduced in Escherichia coli, and we confirmed the combined MPGS and MPGP activities of the recombinant enzyme. The optimum activity of the enzyme was at 50°C. To examine the properties of each catalytic domain of MGSD, we expressed them separately in E. coli. The monofunctional MPGS was unstable, while the MPGP was stable and was characterized. Dehalococcoides ethenogenes cannot be grown sufficiently to identify intracellular compatible solutes, and E. coli harboring MGSD did not accumulate MG. However, Saccharomyces cerevisiae expressing mgsD accumulated MG, confirming that this gene product can synthesize this compatible solute and arguing for a role in osmotic adjustment in the natural host. We did not detect MGSD activity in cell extracts of S. cerevisiae. Here we describe the first gene and enzyme for the synthesis of MG from a mesophilic microorganism and discuss the possible evolution of this bifunctional MGSD by lateral gene transfer from thermophilic and hyperthermophilic organisms.

Characterization of Bacillus cereus Isolates from Raw Soybean Sprouts

Raw soybean sprouts, which are used as ingredients in cook-chilled products, were analyzed to evaluate the incidence of mesophilic aerobic microorganisms, psychrotrophic microorganisms, anaerobic microorganisms, coliforms, and spore-forming microorganisms Bacillus cereus, Clostridium botulinum, and Clostridium perfringens. Mesophilic microorganisms on raw soybean sprouts were present in large populations, 5.5 × 106 to 1.4 × 108 CFU/g, and psychrotrophic microorganisms were found to be more numerous than the other groups. Coliforms accounted for 15% of mesophilic microorganism counts on average, and the average for spore-forming microorganisms was 5.2 × 102 CFU/g. B. cereus was isolated from 12 of 17 soybean sprout samples, whereas C. botulinum and C. perfringens were not isolated. B. cereus was isolated in greater numbers at 30° C compared with other temperatures and was not isolated at 4° C. Of the 55 strains isolated from soybean sprouts, 52 were positive for starch hydrolysis, and only 3 strains did not hydrolyze starch. Among the 55 strains of B. cereus isolates, 53 showed the ability to produce diarrheal enterotoxin by CRET-RPLA.