Micro binfie podcast

87 Nextstrain, SARSCOV2 and dealing with a data deluge by Microbial Bioinformatics

86 What's next for nextstrain? by Microbial Bioinformatics

We are joined again by Prof Mark Pallen who takes us through his early experiences in high-throughput microbial genomics. Mark was pleased that he persuaded Nick Loman to join him in Birmingham. Mark tells us how they worked with George Weinstock to perform the first genome sequence analyses of Gram-negatives for genomic epidemiology—in this case of multi-drug resistant Acinetobacter baumannii.

After winning an Ion Torrent sequencer in a competition, Mark and Nick then contributed some pioneering genomic analyses of the German STEC outbreak. One of their studies involved crowd sourced approaches, primed by Twitter and was published in the New England Journal of Medicine; the other provided a performance comparison of newly launched bench top sequencing platforms. Mark, Lee and Nabil discuss how this outbreak overturned dogmas concerning the archetypal status of pathotypes of E. coli. The conversation then moves on to the need for evidence trails and challenging assumptions, whether annotating proteins or quoting Darwin (see https://colinpurrington.com/2012/02/darwin-on-the-floor-lhao/). Nabil recalls the excitement of realtime analysis of an epidemic and acknowledges the legacy of Mark and Nick's work in 2011 to current approaches to the Covid pandemic.

Mark describes his exciting experiences exploiting metagenomics in clinical and ancient DNA contexts, including analysis of disease-associated stool samples and of 200-year TB genomes in Hungarian mummies. Yet again this led to overturning of assumptions--in this case that people only get infected with a single strain of M. tuberculosis. It turns out that multiple infections were the norm 200 years ago.

Shortly afterwards, Pallen helped assemble a team that analysed undersea sediments shedding light on the Neolithic transition in England and culminating in a Science paper. Mark then takes us through his recent metagenomics analyses of critically ill patients and of the chicken gut, emphasising the excitement of finding hundreds of new species in such a commonplace setting. Mark finishes off by sharing his excitement that there is still so much of the microbial world left for us to discover using sequencing and bioinformatics analyses. We are just 2% of the way there!

Mark Pallen explains how exciting it was to be in microbial bioinformatics around the turn of the millennium, as we gained genomes for the first time from model organisms and fearsome pathogens. He recounts working with his hero David Relman on the genome sequencing of the strange slow-growing organism called Tropheryma whipplei in competition with a French team.

Mark moved to Belfast in late 1999 collaborating with another Englishman working on the island of Ireland, Tim Foster in Dublin. Pallen describes the addictive exhilaration of using PSI-BLAST to find new sortases and sortase substrates across a range of new genomes—for him this was the bioinformatics equivalent of crack cocaine. He quotes the philosopher Alfred North Whitehead in saying that the goal of every scientist is to seek simplicity but distrust it. What Mark found was that in most organisms sortases were behaving quite differently from the rather simple scenario seen in Staphylococcus aureus. He made similar observations on the WXG100 proteins and type VII secretion, which he found in many new contexts quite different from the original context of ESAT-6 as an antigen in Mycobacterium tuberculosis. Mark makes clear that we still don't really fully understand the role of ESAT-6 twenty years on.

The focus of Pallen's work then shifted to E. coli, where he described vestigial gene clusters for non-functional type III secretion systems in this model organism. He came to realize that E. coli K-12 was not handed to microbiologist by God as a model organism but was just another strain of E. coli and nothing special. Many of the earliest genomes to be sequenced came from worn-out lab strains. To counter this problem, Gordon Dougan at the Wellcome Trust Sanger Institute moved the focus to genome-sequencing freshly isolated minimally passaged isolates. With Brendan Wren, Pallen wrote a review article for nature, emphasizing the importance of adopting an eco-evo perspective when trying to interpret bacterial genomes.

Around that time, Scott Beatson joined Pallen's group. Mark managed to persuade Scott to work on type III secretion in E. coli rather than Pseudomonas aeruginosa. The result was the discovery of dozens of new type III secretion effectors, tying together bioinformatics and lab work to culminate in a PNAS paper.

References - https://microbinfie.github.io/2022/06/09/bioinformatics-in-the-noughties.html

We discuss mobile genetic elements in bacteria and find, its really hard. Its just a short chat as Lee lost power, but we will be back with a part 2 sometime soon.

In this episode we talk to Professor Mark Pallen, who discusses the highlights from his long career as a medical microbiologist turned bioinformatician. His bioinformatics journey began in 1977, the year Fred Sanger invented DNA sequencing-as-we know-it, when Mark was tasked with assembling some amino acid sequences under exam conditions. Mark explains how little was know about sequences at the time. Luckily he managed to gain a grasp of molecular biology and joined a group in the late 1908s at Barts Hospital in London, where he met Brendan Wren. Mark's first eureka moment followed shortly afterwards, when he analysed sequences encoding the key enzyme urease from Helicobacter pylori. He also got very excited when he analysed genes from a clostridial butanol fermentation pathway, which he explains, played a central role in the formation of the state of Israel. His next big break came when he got the chance to do a PhD under Gordon Dougan. During this time, Mark not only improved his lab and bioinformatics skills, but captained a winning team in University Challenge and introduced the medical profession to the Internet. He recalls with excitement the moment when he first heard the news that a bacterial genome had been sequenced. Shortly afterwards he recruited an 18-year old gap year student, Nick Loman, to come and work with him analysing the very first Campylobacter jejuni genome. We close this episode just as the new millennium begins, with much more excitement to follow in the next episode.

Relevant links:

Butanol - https://academic.oup.com/femsle/article/124/1/61/486499

Tree-like thinking for genes, languages and gospel manuscripts - https://www.youtube.com/watch?v=8Ykj5wQs7vU

Further references - https://microbinfie.github.io/2022/05/12/bioinformatics-moments-before-the-millennium.html

We bring on Lingzi Xiaoli and Jill Hagey to talk about their benchmark datasets for SARS-CoV-2. Find out more at https://github.com/CDCgov/datasets-sars-cov-2. See our previous episode for part 1 of the conversation.

* Previous paper for bacterial datasets can be found at https://peerj.com/articles/3893/

* Jill can be found on Twitter at @JillHagey and jvhagey.github.io

* Lingzi can be found on LinkedIn at https://www.linkedin.com/in/lingzi-xiaoli-27b87174/

We bring on Lingzi Xiaoli and Jill Hagey to talk about their benchmark datasets for SARS-CoV-2. Find out more at https://github.com/CDCgov/datasets-sars-cov-2

* Previous paper for bacterial datasets can be found at https://peerj.com/articles/3893/

* Jill can be found on Twitter at @JillHagey and jvhagey.github.io

* Lingzi can be found on LinkedIn at https://www.linkedin.com/in/lingzi-xiaoli-27b87174/

Dr Erin Young from the Utah Department of Health and Dr Kelsey Florek from the Wisconsin State Laboratory of Hygiene join us to talk about

StaPH-B containers for public health bioinformatics. Its basically how to make biology easier for everyone!

Github: https://github.com/StaPH-B

Dr. Erin Young and Dr Kelsey Florek join us to talk about StaPH-B, a US state public health bioinformatics group. They also give some insights into the popular SARS-CoV-2 pipeline cecret.

Website: staphb.org/

Cecret Pipeline: github.com/CDCgov/SC2CLIA

Kelsey explains that StaPH-B was created to facilitate collaborations between bioinformaticians in state public health laboratories, especially those just getting started with sequencing and understanding the data generated. The organization provides a conduit of communication and expertise among different laboratories, feeding into projects funded by the NIH, CDC, and other grant agencies.

Erin highlights StaPH-B's diverse membership with different levels of expertise, which provides excellent learning opportunities. The organization uses a Slack workspace, with almost 400 members and over 50 channels dedicated to different activities related to bioinformatics, providing a valuable resource for bioinformaticians to seek out answers to questions and ideas.

The hosts ask Kelsey about who can join StaPH-B, and Kelsey clarifies that while it was initially founded for state public health bioinformaticians, it's open to everyone, and the content is focused on state public health activities. They discuss some of the achievements of StaPH-B, with Kelsey hailing the Slack workspace, collaborations on GitHub, Docker, and collaborative workflows as hugely successful. Additionally, Erin thinks that StaPH-B's training activities, including the Staph-B Toolkit, training sessions, and videos, ensure that knowledge and expertise are shared.

The conversation moves towards the Cecret pipeline, one of Erin's bioinformatics pipelines for SARS-CoV-2. She explains that the pipeline was developed during the pandemic, with the idea of using the Arctic group's protocol for sequencing SARS-CoV-2 on the Nanopore sequencing platform. However, Erin needed a bioinformatic pipeline that was Illumina-based, as it would have been easier to sequence SARS-CoV-2 on the MiSeq, rather than the Nanopore sequencing platform. Cecret pipeline was developed using BWA as the default aligner and is for viral-based sequencing with a known, reliable reference.

Erin points to the SEQret pipeline tutorials and the monthly videos produced by StaPH-B that outline various state laboratory projects as tips for people entering the field. Lastly, Kelsey emphasizes the importance of finding a use case to start building a centralized source of expertise in bioinformatics and making knowledge accessible by having a common resource that's easy to access.

In a previous episode, the guest speakers discussed the evolution of COVID genome analysis workflows and how they have changed over time due to the increasing amount of data being analyzed. They mentioned the use of different workflows such as Secret, NF Core, Monro, and Next Flow Optic Pipeline, each with their own unique features and popularity. Erin, the creator of Secret, talked about how paranoid she was when sharing her workflow publicly and how she would track every fork of her repository to ensure that the changes made to her code were scientifically sound.

The workflow has undergone gradual changes and fewer bugs since its creation, with no dramatic turning point. Its name, “Secret,” was inspired by a hiking landmark in Northern Utah that Erin found meaningful. The speakers emphasized the importance of managing and working with the increasing amounts of COVID data being analyzed, as well as connecting it to public health.

In conclusion, StaPH-B and workflows such as Secret are playing a significant role in the field of bioinformatics and COVID genome analysis. Collaborations and resources like StaPH-B are essential in sharing knowledge and expertise among different laboratories, allowing for the successful completion of projects funded by the NIH, CDC, and other grant agencies.