Overview

Project Highlights:

 

  • Long read viral metagenomics
  • Opportunities to collect samples from tropical rainforests
  • Isolation of novel bacteriophages

 

Tropical rainforests are known to supply the greatest diversity of plants and animals on the planet, yet they only cover a very small percentage of the Earth`s surface. The most abundant living organisms on the planet are bacteria that drive all biogeochemical cycles. Despite the importance of rainforests as reservoirs of animal and plant diversity, virtually nothing is known of the microbial community found within them. Given the vast plant and animal diversity, it would be expected that microbial diversity will also be high. This project will focus on understanding the diversity of bacteriophages in the Belum (Malaysia) and Amazon rainforests.

Bacteriophages are viruses that specifically infect bacteria and are the most numerous biological entities on the planet. Bacteriophages are known to drive the evolution of the bacterial hosts and in other ecosystems directly influence the amount of C02 that is fixed.

Given the vast diversity of micro-environments within rainforests that drive plant speciation, the diversity of bacteria associated with these plants and co-occurring phages is also likely to be high. Understanding bacterial-plant interactions can be complex given the vast number of different bacterial and metabolic plasticity bacteria display. Bacteriophages often carry auxiliary metabolic genes that are used by the phage to augment host metabolism to maximise phage replication. These genes often overcome metabolic bottlenecks encountered by their bacterial hosts and point towards the most important limiting factor in an ecosystem. Amazingly they have not been examined in the rainforest and such genes may be of key importance to understand ecosystem function from a bottom up perspective.

Therefore by studying bacteriophages in rainforests, not only can we expand the known diversity of bacteriophages that is poorly sampled. We can begin to understand important factors in bacterial-plant interactions, as bacteriophages encode genes to overcome metabolic bottlenecks. Therefore, we hypothesise: phage associated with different plant species will harbour different auxiliary metabolic genes.

This project will use both culture-based and culture-independent methods to study bacteriophages from Belum rainforest and from the Amazon that are associated with hallmark plant species. The project will specifically investigate the viral communities in soil surrounding different plant species.

Selected Samples sites in Belum rainforest. 
A) waterfall. B) hilltop site C)site of Rafflesia D) TEM of bacteriophages

Methodology

Using a targeted approach the student will isolate novel bacteriophages and bacteria from Belum/Amazon samples. There will be a focus to isolate phages that reflect novel bacteria. 

In parallel using culture-independent methods they will use long read viral metagenomics – minION and short reads Illumina sequencing to capture the diversity of viruses found in these areas.

The student will receive state-of-the-art bioinformatics training in order to describe the diversity of these phages, and relate this to the context of known phages. They will contribute to our ongoing programme (http://130.226.24.116/phagecompass/index.html)

Transmission Electron Microscopy – TEM will be carried out to examine the morphology of both isolated and total viral samples

Isolated phages will be characterised to identify for their novel properties and phenotypes – this work will be directed by information from their genomes

 

Training and Skills

The students will receive training in many key areas within microbiology. These are:

Bacterial isolation and bacteriophage isolation

Bioinformatic analysis of viral metagenomes from short read data sets

Training in analysis of long read viral metagenomics generated by nanopore technology

Genome annotation and comparison tools

Genome contextualisation and phylogeny

Timeline

Year 1: Isolation of bacterial samples from water and soil samples from sites of interest

Fieldwork and isolation of bacteriophages that target novel bacteria and development of phage-bacteria model systems

Collection of samples for viral metagenomics

Year 2:

Sequencing and analysis of novel viruses from the targeted approach

Characterisation of the viral fraction from the culture independent methods – using both Nanopore (long read) and Illumina (short-read) approaches

Identification of the most novel and interesting aspects of the genomes for mechanistic characterisation – this includes identifying novel auxiliary metabolic genes that dictate phage behaviour in nature.

Year 3: Experimental testing of predicted function of identified novel genes.

Partners and collaboration (including CASE)

The work will be done in collaboration with partners in AMIST University Malaysia and the Pulau Banding Foundation who we already work with and with Guillherme Olivera at the Instituto Tecnológico Vale in Brazil. In addition current collaborators at the University of Copenhagen will also be involved, who will provide expertise on AI based approaches to phage predictions.

Further Details

Websites: A Milllard – lab website http://millardlab.org/ . https://www2.le.ac.uk/departments/genetics/people/andrew-millard/dr-andrew-millard

Prof Clokie https://www2.le.ac.uk/departments/genetics/people/martha-clokie