Search

Preventing bacterial contaminations is a significant challenge in applications across a variety of industries, e.g. in food processing, the life sciences or biohazard detection. Here we present a fully automated lab-on-a-chip system wherein a disposable microfluidic chip moulded by polymeric injection is inserted into an operating device. Liquid samples, here obtained from an air sampler, can be processed to extract and lyse bacteria, and subsequently to purify their DNA using a silica matrix. After the washing and elution steps, the DNA solution is dispensed into a reaction vessel for further analysis in a conventional laboratory polymerase chain reaction (PCR) device. We demonstrate the workability and efficiency of our approach with results from a 9 ml liquid sample spiked with E. coli.

A wide range of analytical methods are available for the detection and identification of biological warfare agents. These technologies are often hampered in their performance when the inactivated samples are analyzed. To work with pathogens outside of biosafety level 3 laboratories, a complete inactivation is mandatory when appropriate protection equipment is unavailable. When methods of inactivation are used, the detection of bacteria becomes more difficult. In contrast to measuring viable organisms, inactivation steps can have a massive impact on the intrinsic cellular information. This study examined the effects of autoclaving and chemical inactivation methods on Bacillusspores using biological warfare detection setups like real‐time PCR and MALDI‐TOF‐MS. Here, the inactivation of Bacillus atrophaeusspores with formaldehyde, which is a suggested model for biological warfare spore agents, was compared with other inactivation reagents like Wofasteril®E400, a commercially available decontaminant based on peroxyacetic acid. With Wofasteril®E400 the critical factor of inactivation time was reduced to about 15 min and a limit of detection of 8500 spores by PCR was still measurable using five‐times‐washed spores. It has also been shown that MALDI‐TOF‐MS peak information can be hampered by inactivation methods.

After 10 days of cultivation at 37 °C, the herbicide resistance encoded by the chimaeric pat4l gene (coding region from Streptomyces viridochromogenesfused to the 823 bp CaMV35S promoter) was strongly reduced in all of the 27 independent transgenic Nicotiana tabacumSRI lines analyzed. This reversible reduction occurred in sterile and unsterile culture in the first and second generation and even when the overnight temperature was reduced to 24 °C. Neither the enzyme activity, the protein nor the pat4l specific RNA could be detected in the heat treated plants, regardless of the number of copies and the hemior homozygous state. In contrast to this, the expression of the synthetic patS coding region fused to the 534bp CaMV35S promoter and coding for essentially the same protein, was stable in heat treated plants. The exchange of the GC rich coding region of the pat4l gene by the AT rich synthetic DNA fragment carrying the patS coding region led to the stabilization of the specific RNA steady state level. However, the presence of the transgene-encoded protein at 37 °C could only be achieved by using specific 5′ and 3′ untranslated regions of the synthetic patS gene.