sRNA-based therapeutics for disease caused by A. pleuropneumoniae

Lead Research Organisation: University of Portsmouth
Department Name: Inst of Biomedical and Biomolecular Sc

Abstract

The bacterium Actinobacillus pleuropneumoniae (APP) causes is a highly contagious lung disease in pigs which affects herds in the UK and worldwide. Infection results in either rapid mortality or slow growth, breathing problems and suffering. APP is responsible for substantial economic losses to the worldwide pig industry.

After good husbandry practices (provision of good ventilation, all in/all out facilities, avoidance of temperature fluctuations and good hygiene) are taken into account there are two basic methods used to limit APP infection: namely vaccines and antibiotics. However, current vaccines have severe limitations and antibiotic resistance is of increasing concern, a recent survey suggesting that 70% of APP isolates in the UK are now resistant to one or more antibiotics. Consequently, there is an urgent need for effective therapeutic approaches to combat APP. Addressing such issues directly aligns to the key BBSRC strategic priorities of 'Animal Health', 'Welfare of Managed Animals' and 'Combating Antimicrobial Resistance'.

Traditional antibiotics seek to simply kill bacteria, but a novel therapeutic alternative is to target functions that are essential for the bacteria's ability to infect the host. Such an approach effectively disarms the bacteria, rather than killing them, thereby avoiding the selective pressure from which traditional antibiotic resistance originates. Bacterial cells, such as APP, have important switching molecules (sRNAs) which are responsible for turning on the mechanisms that enable host infection to occur. Our proposal plans to identify these crucial sRNAs in APP and then inhibit their function using compounds called peptide nucleic acids (PNAs). Targeting the sRNAs in this manner will lead to a new therapeutic approach for treating APP disease. More broadly, this research also provides the basis for exploring sRNA inhibition as an alternative therapeutic approach to bacterial diseases caused by other major pathogens of animals and man.

This work is innovative, timely and multi-disciplinary, building on recent work and employing the latest molecular tools and technological advances to deliver results. This is a joint collaborative venture which benefits from the synergy between world leading expertise in APP (Imperial College London) and an outstanding track record in sRNA research (University of Portsmouth). Within this context, the project has a substantial chance of success, standing to offer a step change in capability for treating APP, but with far reaching impact for the therapeutic treatment of bacterial diseases more broadly.

Technical Summary

Actinobacillus pleuropneumoniae (APP) causes acute and chronic lung disease in pigs and is of substantial economic importance to the worldwide swine industry. There is an urgent need for improved vaccines and therapeutics as antibiotic resistance is an increasing problem. The focus of this proposal is bacterial small non-coding RNAs (sRNAs) which, facilitated by Hfq, play a key role controlling mRNAs responsible for virulence. Our work has shown that the hfq-mutants of APP are severely attenuated and rapidly cleared from pigs. While Hfq could be targeted directly as a therapeutic approach, the presence of homologous eukaryotic proteins suggests that targeting the associated sRNAs will be a more effective strategy. In particularly, this approach will bring about a therapeutic effect without the risk of developing bacterial resistance, since no survival pressure, from which traditional antibiotic resistance originates, is imposed.

To date, our work has experimentally identified 15 Hfq-associated sRNAs in APP, and computationally a further 30 have been predicted. Our recent bioinformatic analysis confirms a number of these sRNAs target mRNAs with relevance to virulence. This proposal represents a major collaboration between Imperial College London and the University of Portsmouth, utilising the extensive experience in APP and Hfq-sRNA interactions, respectively. We plan to ascertain the identity of Hfq-associated sRNAs that are key in turning on virulence genes and determine whether inhibiting these sRNAs using peptide nucleic acids (PNAs) provides a novel therapeutic approach to treating APP disease. Using a combination of in vitro biochemical and biophysical molecular characterisation approaches, coupled with in vivo reporter and/or phenotypic assays, we will determine the viability of this therapeutic approach. Success would offer a step change in capability for combating APP, but with far reaching impact for the treatment of bacterial diseases more broadly.

Planned Impact

Please see main proposal.

Publications

10 25 50
 
Description This grant is on-going.
Exploitation Route This grant is on-going.
Sectors Healthcare,Pharmaceuticals and Medical Biotechnology

 
Description University of Portsmouth PhD Studentship (£ 60000; 2016-2019)
Amount £60,000 (GBP)
Organisation University of Portsmouth 
Sector Academic/University
Country United Kingdom
Start 10/2016 
End 09/2019
 
Description Collaboration with Prof. Denise Bazzolli at the Universidade Federal de Viçosa 
Organisation Federal University of Viçosa
Country Brazil 
Sector Academic/University 
PI Contribution My research team are undertaking molecular interaction studies guided by microbiology data provided by Prof. Bazzolli's team.
Collaborator Contribution Prof. Bazzolli's team are providing microbiology data and expertise.
Impact One of Prof. Bazzolli's team has secured funding to undertake a year-long placement in my research group to support this collaboration.
Start Year 2015
 
Description Collaboration with Prof. Jay Hinton at the University of Liverpool (2017 - Still Active) 
Organisation University of Liverpool
Country United Kingdom 
Sector Academic/University 
PI Contribution Myself and my team provide expertise in the research subject area.
Collaborator Contribution Prof. Hinton provides expertise in bioinformatics to support with data analysis.
Impact The grant and collaboration are on-going.
Start Year 2017
 
Description Collaboration with Prof. Paul Langford at Imperial College London 
Organisation Imperial College London
Country United Kingdom 
Sector Academic/University 
PI Contribution My research team are conducting in vitro molecular interaction studies, and exploring the applicability of utilising our patented novel array technology, to explore cellular mechanisms within bacteria.
Collaborator Contribution Prof. Langford's team provide microbiology expertise and access to in vivo testing studies.
Impact A joint BBSRC grant was secured to support this collaborative research.
Start Year 2015
 
Description Maintaining an Active Online Presence 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact My group has an active Twitter account with around 200 followers. We publish highlights from our research, outreach and engagement activities.
Year(s) Of Engagement Activity 2011,2012,2013,2014,2015
 
Description Promoting PG study 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Undergraduate students
Results and Impact Presentation by members of my research team at various departmental events for undergraduates to promote engagement in postgraduate study. this involved the individuals highlighting their research work, including their day to day work, opportunities for collaboration and engagement as well as their outputs and impact.
Year(s) Of Engagement Activity 2013,2014,2015,2016
 
Description Science Fairs 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Myself and my team have participated in supporting a number of Science Fairs in the region, engaging with attendees to promote science and the research we undertake.
Year(s) Of Engagement Activity 2013,2014,2015,2016
 
Description University Open Days 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Schools
Results and Impact Myself and my team regularly support University Open days. Activities can be many and varied, including giving talks, presenting posters, running hands-on laboratory demonstrations and engaging in question and answer sessions. There are usually a number of these events per year, with over 100 participants (schools and college students, sometimes accompanied by a parent/guardian) attending each event. Feedback from such events has highlighted our success in inspiring the next generation of scientists and has been specifically linked to an increase in the number of students applying to study Biochemistry over the last few years.
Year(s) Of Engagement Activity 2009,2010,2011,2012,2013