Applied Sciences

Clostridium difficile causes outbreaks of infectious diarrhoea, most commonly occurring in healthcare institutions. Recently, concern has been raised with reports of C. difficile disease in those traditionally thought to be at low risk i.e. community acquired rather than healthcare acquired. This has increased awareness for the need to track outbreaks and PCR-ribotyping has found widespread use to elucidate epidemiologically linked isolates. PCR-ribotyping uses conserved regions of the 16S rRNA gene and 23S rRNA gene as primer binding sites to produce varying PCR products due to the intergenic spacer (ITS1) regions of the multiple operons. With the explosion of whole genome sequence data it became possible to analyse the start of the 23S rRNA gene for a more accurate selection of regions closer to the end of the ITS1. However the following questions must still be asked: (i) Does the chromosomal organisation of the rrn operon vary between C. difficile strains? and (ii) just how conserved are the primer binding regions? Eight published C. difficile genomes have been aligned to produce a detailed database of indels of the ITS1's from the rrn operon sets. An iPad Filemaker Go App has been constructed and named RiboTyping (RT). It contains detail such as sequences, ribotypes, strain numbers, GenBank numbers and genome position numbers. Access to various levels of the database is provided so that details can be printed. There are three main regions of the rrn operon that have been analysed by the database and related to each other by strain, ribotype and operon: (1) 16S gene (2) ITS1 indels (3) 23S gene. This has enabled direct intra-and inter-genomic comparisons at the strain, ribotype and operon (allele) levels in each of the three genomic regions. This is the first time that such an analysis has been done. By using the RT App with search criteria it will be possible to select probe combinations for specific strains/ribotypes/rrn operons for experiments to do with diagnostics, typing and recombination of operons. Many more incomplete C. difficile whole genome sequencing projects are recorded in GenBank as underway and the rrn operon information from these can also be added to the RT App when available. The RT App will help simplify probe selection because of the complexity of the ITS1 in C. difficile even in a single genome and because other allele-specific regions (16S and 23S genes) of variability can be relationally compared to design extra probes to increase sensitivity.

Although premenstrual exacerbation of vulvovaginal symptoms attributed to Candida spp. is well documented, the causation of these symptoms is not well understood. This study describes the daily vaginal colonization of Candida in three women. A single pilot study was designed to test the methodology of the proposed randomized controlled trial, Garlic and Candida. This study reports the colonization of Candida spp. in three women. Ten women aged 18-50 who reported at least one episode of vulvovaginal candidiasis were recruited by the University of Melbourne. Each participant took daily vaginal swabs for 2 weeks during the luteal phase of their menstrual cycle, which were analysed for quantitative colony counts of Candida spp. Of these, three women were colonized with Candida spp. For the first time, to our knowledge, daily colonization of Candida during the luteal phase of the menstrual cycle is described in three women, demonstrating an increase in the colony count preceding symptom development. This small study demonstrated the colonization of Candida spp. during the luteal phase of the menstrual cycle in three women. Candida colonization is poorly understood, yet investigating the relevance of the link between symptom exacerbation and the menstrual cycle in those women who experience recurrent episodes of vulvovaginal candidiasis may influence the management of this condition.

Objective Garlic is effective against Candida species in vitro, and along with other alternative therapies, is used by women with vulvovaginal candidiasis. The objective of this study was to ascertain whether oral garlic reduced vaginal candida counts during the second half of the menstrual cycle in asymptomatic women colonised with Candida species.

Direct plant regeneration was obtained from fresh c h r y s a n t h e m u m (Chrysanthemum morifolium Ramat. cv. Royal Purple) stem segments cultured on M u r a s h i g e and S k o o g ' s (1962) basal m e d i a supplemented with 6-benzylaminopurine (BAP, 0.5-2.0 mg/1) and a-naphthaleneacetic acid (NAA, 012-2.0 mg/1). The morphogenetic potential varied with the developmental stage of the stem explant. The highest percentage of shoot formation (100%) and greatest average number of shoots per explant (14.6) were observed on stem segments taken from the top of the cutting. This organogenetic capacity decreases in the more mature stem. Normal, flowering plants were obtained three to four months after culture.

FtsZ1-1 and MinD plastid division-related genes were identified and cloned from Brassica oleracea var. botrytis. Transgenic tobacco plants expressing Bo-FtsZ1-1 or BoMinD exhibited cells with either fewer but abnormally large chloroplasts or more but smaller chloroplasts relative to wild-type tobacco plants. An abnormal chloroplast phenotype in guard cells was found in BoMinD transgenic tobacco plants but not in BoFtsZ1-1 transgenic tobacco plants. Transgenic tobacco plants bearing the macro-chloroplast phenotype had 10 to 20-fold increased levels of total FtsZ1-1 or MinD, whilst the transgenic tobacco plants bearing the mini-chloroplast phenotype had lower increased FtsZ1-1 or absence of detectable MinD. We also described for the first time, plastid transformation of macro-chloroplast bearing tobacco shoots with a gene cassette allowing for expression of green fluorescent protein (GFP). Homoplasmic plastid transformants from normal chloroplast and macro-chloroplast tobacco plants expressing GFP were obtained. Both types of transformants accumulated GFP at *6% of total soluble protein, thus indicating that cells containing macro-chloroplasts can regenerate shoots in tissue culture and can stably integrate and express a foreign gene to similar levels as plant cells containing a normal chloroplast size and number.

Echinacea, native to the Canadian prairies and the prairie states of the United States, has a long tradition as a folk medicine for the Native Americans. Currently, Echinacea are among the top 10 selling herbal medicines in the U.S. and Europe, due to increasing popularity for the treatment of common cold and ability to stimulate the immune system. However, the genetic relationship within the species of this genus is unclear, making the authentication of the species used for the medicinal industry more difficult. We report the construction of a novel Subtracted Diversity Array (SDA) for Echinacea species and demonstrate the potential of this array for isolating highly polymorphic sequences. In order to selectively isolate Echinaceaspecific sequences, a Suppression Subtractive Hybridization (SSH) was performed between a pool of twenty-four Echinacea genotypes and a pool of other angiosperms and non-angiosperms. A total of 283 subtracted genomic DNA (gDNA) fragments were amplified and arrayed. Twenty-seven Echinacea genotypes including four that were not used in the array construction could be successfully discriminated. Interestingly, unknown samples of E. paradoxa and E. purpurea could be unambiguously identified from the cluster analysis. Furthermore, this Echinacea-specific SDA was also able to isolate highly polymorphic retrotransposon sequences. Five out of the eleven most discriminatory features matched to known retrotransposons. This is the first time retrotransposon sequences have been used to fingerprint Echinacea, highlighting the potential of retrotransposons as based molecular markers useful for fingerprinting and studying diversity patterns in Echinacea.

received This paper establishes a theoretical basis for the implementation of governance systems in Occupational Health and Safety (OHS). This arose when a complex government organization reviewed the usage of its OHS computer system. This review precipitated several cycles of Action Research (AR). The AR allowed the recognition that the organization needed to define its OHS strategy, clarify its commitment to OHS, and examine its OHS systems in order to provide an effective OHS management and corporate governance system. As part of the review, a Viable Systems Diagnosis was done on the OHS function. It was recognized that Stafford Beer provided valuable insights into the information needs and communication flows of OHS information throughout this complex organization. Action research appears to be a particularly useful method for examining and implementing OHS management systems. KEY WORDS: action research in OHS; OHS management systems; OHS corporate governance; viable systems diagnosis. Pub: Kindly provide the article acceptance information.