Expression and delivery of an endolysin to combat Clostridium perfringens

Clostridium perfringens is a cause for increasing concern due to its responsibility for severe infections both in humans and animals, especially poultry. To find new control strategies to treat C. perfringens infection, we investigated the activity and delivery of a bacteriophage endolysin. We identified a new endolysin, designated CP25L, which shows similarity to an N-acetylmuramoyl-l-alanine amidase domain and is distinct from other C. perfringens endolysins whose activity has been demonstrated in vitro. The cp25l gene was cloned and expressed in Escherichia coli, and the gene product demonstrated lytic activity against all 25 C. perfringens strains tested. The probiotic strain Lactobacillus johnsonii FI9785 was engineered to deliver the endolysin to the gastrointestinal tract. The integration of the nisRK two-component regulatory system from the Lactococcus lactis nisin A biosynthesis operon into the chromosome of L. johnsonii allowed constitutive expression of the endolysin under the control of the nisA promoter (P(nisA)), while the use of a signal peptide (SLPmod) led to successful secretion of the active endolysin to the surrounding media. The high specificity and activity of the endolysin suggest that it may be developed as an effective tool to enhance the control of C. perfringens by L. johnsonii in the gastrointestinal tract

Discovery of a novel lantibiotic nisin O from Blautia obeum A2-162, isolated from the human 2 gastrointestinal tract

The Institute of Food Research is funded by the BBSRC (strategic core grants IFR/08/1 BB/J004529/1); SHD and HJF acknowledge support from the Scottish Government Food Land and People programme. DH and CGF received BBSRC PhD studentship grants. Acknowledgements We are grateful to Kathryn Cross for the EM analysis and Neil Rigby for helpful advice.Peer reviewedPostprin

Acceptance of Assisted Living Technologies in Europe

A recent European Commission report revealed the discrepancies in e-health adoption amongst older people between various European countries (Kubitschke, 2010). Uptake rates, in terms of the percentage of people aged over 65 using e-health technologies, differ for example, for social alarms between around 3% in Germany and the Netherlands, 6-10% the Nordic countries and 15% in the United Kingdom. Even within individual countries there are areas with vastly different adoption rates (Cooney, 2010). Considering that the European population is ageing and the life expectancy at birth is still rising significantly (European Commission, 2011), there is a need for government policy on ageing in place to keep people at home as long as possible and there are chances for producers and providers of ALT solutions to increase their sales. The most important barrier to overcome is the low adoption rate of ALT solutions. Governments, companies and society as a whole need to find out why individuals prefer not to use ALT products and/or services even though they can be seen as potential users

Complete Genome Sequence of Lactobacillus johnsonii FI9785, a Competitive Exclusion Agent against Pathogens in Poultry

Wegmann U, Overweg K, Horn N, et al. Complete Genome Sequence of Lactobacillus johnsonii FI9785, a Competitive Exclusion Agent against Pathogens in Poultry. JOURNAL OF BACTERIOLOGY. 2009;191(22):7142-7143.Lactobacillus johnsonii is a member of the acidophilus group of lactobacilli. Because of their probiotic properties, including attachment to epithelial cells, immunomodulation, and competitive exclusion of pathogens, representatives of this group are being intensively studied. Here we report the complete annotated genome sequence of Lactobacillus johnsonii FI9785, a strain which prevents the colonization of specific-pathogen-free chicks by Clostridium perfringens

Enhanced Secretion of Biologically Active Murine Interleukin-12 by Lactococcus lactis▿

The novel signal peptide SLPmod was used for the secretion of murine interleukin-12 (mIL-12) by Lactococcus lactis. A >4-fold increase in secretion was observed when SLPmod was used instead of the Usp45-derived secretion signal. Oral delivery of this cytokine using the autoinducible host L. lactis FI5876 utilizing SLPmod resulted in a significant increase in mIL-12 plasma levels in mice

Spontaneous mutation reveals influence of exopolysaccharide on Lactobacillus johnsonii surface characteristics.

As a competitive exclusion agent, Lactobacillus johnsonii FI9785 has been shown to prevent the colonization of selected pathogenic bacteria from the chicken gastrointestinal tract. During growth of the bacterium a rare but consistent emergence of an altered phenotype was noted, generating smooth colonies in contrast to the wild type rough form. A smooth colony variant was isolated and two-dimensional gel analysis of both strains revealed a protein spot with different migration properties in the two phenotypes. The spot in both gels was identified as a putative tyrosine kinase (EpsC), associated with a predicted exopolysaccharide gene cluster. Sequencing of the epsC gene from the smooth mutant revealed a single substitution (G to A) in the coding strand, resulting in the amino acid change D88N in the corresponding gene product. A native plasmid of L. johnsonii was engineered to produce a novel vector for constitutive expression and this was used to demonstrate that expression of the wild type epsC gene in the smooth mutant produced a reversion to the rough colony phenotype. Both the mutant and epsC complemented strains had increased levels of exopolysaccharides compared to the wild type strain, indicating that the rough phenotype is not solely associated with the quantity of exopolysaccharide. Another gene in the cluster, epsE, that encoded a putative undecaprenyl-phosphate galactosephosphotransferase, was deleted in order to investigate its role in exopolysaccharide biosynthesis. The ΔepsE strain exhibited a large increase in cell aggregation and a reduction in exopolysaccharide content, while plasmid complementation of epsE restored the wild type phenotype. Flow cytometry showed that the wild type and derivative strains exhibited clear differences in their adhesive ability to HT29 monolayers in tissue culture, demonstrating an impact of EPS on surface properties and bacteria-host interactions