Detection of Bacillus cereus group bacteria from cardboard and paper with real-time PCR

Abstract

The aim of this study was to develop a PCR-based rapid method to detect Bacillus cereus group cells from paper and cardboard. Primers targeting the 16S rDNA and real-time PCR with SYBR green I detection were used in order to be able to also quantify the target. Both autoclaved cardboard samples spiked with B. cereus vegetative cells or spores and naturally contaminated paper and cardboard samples were studied. Results were compared with culturing verified by commercial (API) tests. Several different methods were tested for DNA isolation from the paper and cardboard samples. Two commercial kits intended for soils, the UltraClean soil DNA kit and the FastDNA spin kit for soil, gave the most reproducible results. In spiked samples, the average yield was 50% of added vegetative cells, but spore yield was only about 10%. PCR results from adding vegetative cells correlated with added colony-forming unit (cfu) values ( r=0.93, P <0.001) in the range 100-10,000 cfu g(-1). Three out of nine studied paper and cardboard samples contained B. cereus group bacteria, based both on culturing and real-time PCR. The numbers were 10(2)-10(3) bacteria g(-1); and PCR gave somewhat higher results than culturing. Thus, real-time PCR can be used as a rapid semi-quantitative method to screen paper and cardboard samples for contamination with B. cereus group bacteria.

Quantitative contributions of bacteria and of Deinococcus geothermalis to deposits and slimes in paper industry

“…The signiWcance of D. geothermalis in the bioWlm industrial deposits may be higher than appears from its minority presence, if its laboratory-shown properties also apply for mill conditions: (1) ability to adhere to steel or glass surface so Wrmly that the bioWlm cannot be removed by hot NaOH (pH 12) or SDS wash [14]; (2) D. geothermalis colonised steel surface attracts several paper machine Bacillus species, including the undesirable pathogen B. cereus [23,25,31,32], which otherwise would not produce bioWlm in paper machine waters [13]; (3) D. geothermalis bioWlm cells adhere to each other and the nonliving surface by large number of glycoproteins of type IV pili [27] but can nevertheless spread along the nonliving surface by gliding motility [14,22,27]; (4) D. geothermalis bioWlm colonies do not produce loosely attached slime or capsule [7,14], but, instead, are protected by a number of tightly attached glycoconjugates; [22] (5) the colonies are nonporous [22] and may thus exclude biocides and other microbe killing agents [12]. The industrial survey executed in the present study showed that D. geothermalis is in paper industry a frequent coloniser, not occurring in bulk quantity.…”

Detection and quantitation of colored deposit-forming Meiothermus spp. in paper industry processes and end products

“…To our knowledge, this is the Wrst time the connection between the bacterial DNA content and spots in end products is demonstrated. DNA methods have previously been used with paper industry samples by Priha et al [21]. They applied Bacillus cereus-speciWc primers [7] to assess the presence of this species in paper industry products.…”

Chemical and Microbiological Aspects of the Interaction Between Food and Food Packages

“…The presence of B. cereus over 100 CFU per gram of cardboard (containers for milk) can be the cause of food poisoning [4]. Another research has indicated that 26.3 % of isolated life forms from food packaging boards appear to be foodborne pathogens such as B. cereus [11,12].…”