Use of Candida albicans and its oestrogen binding protein as a bio-recognition element for detection of oestrogen

Compared with bacteria, yeast have rarely been studied for use as a biocomponent for biosensors. Yeast are easy to culture and are eukaryotes, which means their biochemistry in many respects is similar to that of higher organisms. C. albicans and some other yeast species are known to have an oestrogen binding protein (EBP), which oxidises NAD(P)H to NAD(P)+ Oestrogen, when present, binds to the NAD(P)H oxidation site which leads to an accumulation of NAD(P)H (Madani, et al. 1994). Previous research has shown that oestrogen can be quantified using S. cerevisiae whole cells as the detection element and measuring NAD(P)H with a double mediator electrochemical system. This thesis employs the mediated electrochemical systems to investigate the influence of growth phase on EBP production in C. albicans and the response of cells of different ages to different concentrations of oestrogen. A cell not known to possess EBP (Arxula adeninivorans) was also investigated for its response to 17β-oestradiol. As expected, A. adeninivorans did not show a detectable response to 17β-oestradiol but surprisingly, its catabolism was inhibited. By using C. albicans cell lysate in the oestrogen detecting assay, utility was systematically increased and the complexity of the whole cell assay was decreased. In this assay, only a hydrophilic mediator was used, removing the need for a lipophilic mediator. The assay was used successfully in a complex medium, the upper detection limit was raised to 100 nM of 17β-oestradiol, and the assay period was reduced to 20 min. The electrodes were modified to directly detect NAD(P)H in cell lysate at a lower potential to avoid interference by oxidants such as ascorbic acid. Furthermore, EBP was purified using 17β-oestradiol affinity chromatography, and the protein was used with NADH in an oestrogen bioassay. In this assay NADH was electrochemically detected directly and could differentiate ‘with’ and ‘without oestrogen’ samples. This research also showed that a mediator can interact directly with EBP, i.e. without the use of NADH and further, direct electron transfer from EBP to both glassy carbon and pyrolyzed photoresist film (PPF) electrodes were demonstrated (i.e. without the use of a mediator). This research has further simplified the assay and will facilitate the development of rapid oestrogen detecting protein biosensor

The potential of Lactobacillus spp. for modulating oxidative stress in the gastrointestinal tract

The gastrointestinal (GI) tract is crucial for food digestion and nutrient absorption in humans. However, the GI tract is usually challenged with oxidative stress that can be induced by various factors, such as exogenous pathogenic microorganisms and dietary alterations. As a part of gut microbiota, Lactobacillus spp. play an important role in modulating oxidative stress in cells and tissues, especially in the GI tract. Oxidative stress is linked with excessive reactive oxygen species (ROS) that can be formed by a few enzymes, such as nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (NOXs). The redox mechanisms of Lactobacillus spp. may contribute to the downregulation of these ROS‐forming enzymes. In addition, nuclear factor erythroid 2 (NFE2)‐related factor 2 (Nrf‐2) and nuclear factor kappa B (NF‐κB) are two common transcription factors, through which Lactobacillus spp. modulate oxidative stress as well. As oxidative stress is closely associated with inflammation and certain diseases, Lactobacillus spp. could potentially be applied for early treatment and amelioration of these diseases, either individually or together with prebiotics. However, further research is required for revealing their mechanisms of action as well as their extensive application in the future

Elastic light scatter pattern analysis for the expedited detection of Yersinia species in pork mince: Proof of concept

Isolation of the pathogens Yersinia enterocolitica and Yersinia pseudotuberculosis from foods typically rely on slow (10–21 day) “cold enrichment” protocols before confirmed results are obtained. We describe an approach that yields results in 39 h that combines an alternative enrichment method with culture on a non-selective medium, and subsequent identification of suspect colonies using elastic light scatter (ELS) analysis. A prototype database of ELS profiles from five Yersinia species and six other bacterial genera found in pork mince was established, and used to compare similar profiles of colonies obtained from enrichment cultures from pork mince samples seeded with representative strains of Y. enterocolitica and Y. pseudotuberculosis. The presumptive identification by ELS using computerised or visual analyses of 83/90 colonies in these experiments as the target species was confirmed by partial 16S rDNA sequencing. In addition to seeded cultures, our method recovered two naturally occurring Yersinia strains. Our results indicate that modified enrichment combined with ELS is a promising new approach for expedited detection of foodborne pathogenic yersiniae

Arcobacter cryaerophilus isolated from New Zealand mussels harbor a putative virulence plasmid

A wide range of Arcobacter species have been described from shellfish in various countries but their presence has not been investigated in Australasia, in which shellfish are a popular delicacy. Since several arcobacters are considered to be emerging pathogens, we undertook a small study to evaluate their presence in several different shellfish, including greenshell mussels, oysters, and abalone (paua) in New Zealand. Arcobacter cryaerophilus, a species associated with human gastroenteritis, was the only species isolated, from greenshell mussels. Whole-genome sequencing revealed a range of genomic traits in these strains that were known or associated virulence factors. Furthermore, we describe the first putative virulence plasmid in Arcobacter, containing lytic, immunoavoidance, adhesion, antibiotic resistance, and gene transfer traits, among others. Complete genome sequence determination using a combination of long- and short-read genome sequencing strategies, was needed to identify the plasmid, clearly identifying its benefits. The potential for plasmids to disseminate virulence traits among Arcobacter and other species warrants further consideration by researchers interested in the risks to public health from these organisms