Interview with the author: Using host transcriptomics to sample blood parasites

Hosts offer diverse habitat for an incredibly rich array of microbial groups. Genomic resources for many groups residing within hosts (‘infra-communities’) are poor often due to the difficulty in isolating the DNA from the microbe from that of the host, particularly for species living within host cells. In this interview we go behind the scenes with Spencer Galen as he guides us through his transcriptomic approach he developed with colleagues to sample blood parasites such as malaria. Given how ubiquitous and important these parasites can be for animal health, this resource has the potential to pave the way for important advances in disease ecology. Read the paper here.

Avian blood transcriptomes revealed that hosts often have far more complex parasite communities than traditionally thought. For instance, the transcriptome of this Baltimore oriole (Icterus galbula) revealed at least six malaria parasite infections from three malaria parasite genera. The blood smear image from this bird shows the three genera in close contact within the host bloodstream. L: Leucocytozoon, PL: Plasmodium, PA: Parahaemoproteus.
Credit: Spencer Galen

What led to your interest in this topic / what was the motivation for this study? 

This study began with two classic ingredients of scientific discovery: a lot of frustration mixed with a bit of inspiration from other researchers. The frustration was born from a lack of available genetic resources for malaria parasites and other blood parasites, which I felt was hindering the kind of research that I wanted to do. The inspiration came during the first year of my PhD, when several papers were published within a span of just a few months showing that researchers were passively generating large quantities of blood parasite genomic data by sequencing the transcriptomes of their vertebrate hosts. My PhD advisor Susan Perkins and I thought that designing a study to explore this approach in more detail could solve some of my frustrations and help the field of blood parasite research at large.

What difficulties did you run into along the way? 

When we started this project there was always the looming possibility that we would sequence a number of host transcriptomes that were infected with blood parasites and simply not recover any useful parasite data. Even a small-scale transcriptomic project is not a trivial matter financially, and so I will admit that I lost some sleep wondering if this project was a bad idea. Fortunately, field and lab work went quite smoothly, and the results of my first scan for parasites within our initial test transcriptomes exceeded my wildest expectations. And so in reality the biggest challenge was my own self-doubt – if I had paid too much attention to those thoughts, this project might not have gotten off the ground.

What is the biggest or most surprising innovation highlighted in this study? 

We were astounded by just how prevalent blood parasite transcripts can be within host transcriptomes. For instance, in one bird (Vireo plumbeus sampled in the mountains of New Mexico) we found that nearly 17% of all contigs generated from the initial Trinity assembly were derived from a parasite that was infecting just 0.75% of all blood cells. A second surprising finding was the degree to which many of the birds that we sampled were infected with complex communities of parasites that we did not detect using traditional microscopic and DNA barcoding methods. Across all samples we found that transcriptomes revealed about ~20% more infections than the methods that are typically used to study these parasites. This included one individual bird that was infected by three different genera and at least six species of malaria parasite.

Moving forward, what are the next steps in this area of research?

While it is exciting to find that a transcriptomic approach can improve our ability to study the genomic diversity and abundance of wildlife blood parasites, it still remains a rather inefficient approach – at the end of the day, the majority of transcripts from each sample came from the host organism that was not the focus of our study. The next step will be to apply single-cell and other advanced RNA sequencing techniques that have successfully been applied to model systems to provide greater resolution to studies of blood parasite gene expression and host-parasite interactions.   

What would your message be for students about to start developing or using novel techniques in Molecular Ecology? 

At risk of sounding overly pessimistic, be prepared for things to fail the first time around and have a plan B in place. It is wonderful to have a lot of confidence, but pessimism does tend to favor preparedness. Small actions within this frame of mind can save you a lot of grief in the long run, and can be as simple as testing a new method on a sample that isn’t important before you start your project or taking the time to visit a lab to learn a technique before you try it yourself. I naturally assume everything I try in the lab will fail, so each time things work (and they actually often do!) it is a pleasant surprise.

What have you learned about methods and resources development over the course of this project? 

I think that there is a difference between producing a resource, and producing a resource that is easily accessible to the broader research community in practice. As a result, I spent a lot of time thinking about how my colleagues would most directly benefit from the data that we had generated. In the end we made the data from this study available in as many formats as we thought might be useful to other researchers (raw sequences, assemblies from before and after parasite identification, curated alignments, DNA barcodes, etc.). The amount of time that it took to prepare these datasets was extremely small relative to the length of the entire project, and I think will go a long way towards making these data as useful as possible.

Describe the significance of this research for the general scientific community in one sentence.

This study improves our ability to research the ecology and evolution of wildlife blood parasites, a cosmopolitan and ubiquitous group that is widely relevant to global health.

Describe the significance of this research for your scientific community in one sentence.

The methodological framework that we present in this study profoundly improves the genomic resource base that is available to research understudied blood pathogens of wildlife, as well as better detect multi-species parasite communities within hosts.

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