by Abigail Curtis
My first niche microbiological interest came freshman year of high school, when I discovered that viruses don't solely infect animals. Indeed, viruses are pervasive in every ecosystem, including the smallest ones in the ground. The aspect of viruses in microbial ecosystems that I found most fascinating was the concept of giant viruses, and the enigmatic character of their genome and importance. Ever since, I've tried my best to keep up to date on the subject. I find the recent turn to sequencing technologies as an approach to the discovery of the world of viruses below our feet incredibly exciting.
The genus Mimiviridae contains some of the largest viruses known to humankind. In fact, a 2010 STC post by Merry Youle highlighted the discovery of Acanthamoeba polyphaga mimivirus (APMV), shattering previous records with a genomic size of ~1.2 Mbp and capsid size of ~500 nm. However, since the time that post – "A Giant Among Giants" – appeared, there have been increased efforts and improved technologies in the mission to discover and map the viral composition of terrestrial environments.
No longer are scientists required to cultivate viruses in hosts. The combination of next generation sequencing technologies, open access sequence databases, and programs that specifically mine databases to identify viral genomes, such as VirSorter and VirFinder, now permit the quick identification of viral genomes. Of note is a new approach referred to as "mini-metagenomics." Rather than isolate genetic material from all of the microbes within a sample, a mini-metagenomic approach first sorts microbes into smaller sub samples through parallelization, a process carried out using microfluidic devices. Samples that contain in the order of 100 microbes serve to generate the "mini-metagenomic" sequences. This approach is best for analyzing small samples with high microbial diversity, as it provides metagenome-assembled genomes (MAGs) with a greater degree of accuracy. With the development of both bulk metagenomics (which yield "bulk MAGs") and mini-metagenomics (which yield "sorted-MAGs"), investigators can now discover more viruses without having to know their host.
Using both approaches, a group studying soil samples from Harvard Forest discovered sixteen new soil giant viruses. Mini-metagenomics yielded 15 genomes, while bulk metagenomics yielded only one. In addition, the higher quality sequences obtained from mini-metagenomics allowed for more accurate virus classification. The presence of hallmark genes encoding the major capsid protein (MCP), paralogous genes, and packaging ATPases allowed them to unambiguously classify the new viruses as nucleocytoplasmic large DNA viruses (NCLDVs). The authors were able to place 10 of their 16 viral genomes within the genus Mimiviridae. Three genomes seemed to form a new group, while the remaining genomes were not complete enough to allow classification.
New sequencing technologies have not only enhanced abilities to discover new viruses. They also provide new knowledge regarding the role and importance of giant viruses in their environments, along with confirming the radical nature of their genomes. In a study analyzing 501 NCLDV MAGs, viruses within Mimiviridae and Phycodnaviridaefamilies contain genes encoding enzymes essential for glycolysis and the citric acid cycle. Most prevalently among them, the glycolytic glyceraldehyde-3-phosphate dehydrogenase (G3P), phosphoglycerate mutase (PGM), phosphoglycerate kinase (PGK), and the tricarboxylic aconitase and succinate dehydrogenase (SDH). One MAG had 7 out of 10 genes necessary for glycolysis. These findings indicate giant viruses' extraordinary genomic independence from their hosts (termed “quasi-autonomous” viruses) and highlights a potential for pathogens to reconstruct metabolism in virocells.
Another unique giant virus is the tupanvirus, found in oceanic and lake sediments, which contains the most comprehensive set of genes involved in translation, including genes for 20 aminoacyl tRNA synthetases, 11 transcription factors, and up to 70 tRNAs in one genome. The 'soda lake' strain of Tupanvirus is also unique amongst giant viruses in its ability to infect several protist hosts, such as several Acanthameoba species, and numerous phytoplankton strains.
Giant viruses continue to excite and intrigue, as members contradict another universal characteristic of viruses: capsid proteins. While the known members of the Mimiviridae family have capsid shells in nearly a perfect icosahedral shape, there are notable giant viruses that do not. Pandoraviruses, Cedratviruses, and Orpheoviruses lack a capsid gene; in its place a tegument-adjacent envelope. The Pithoviruses stand out because, although they contain genes coding for a capsid protein, do not present one phenotypically. In contrast, faustoviruses have two protein shells, the outermost layer consisting of a capsid protein found in classical viruses, while the inner casing is hexameric.
All the viruses aforementioned were discovered in the past 15 years, suggesting that scientists have barely begun to understand Earth's viral diversity. Since viral biodiversity is incredibly vast, and the overlap of genes between currently known species is slim, constructing a comprehensive phylogenetic tree remains difficult. Yet, theories abound regarding their evolutionary origins. One theory is that giant viruses formed before the ribosome formation, but after the evolution of aminoacyl tRNA synthetases, and they have since acquired metabolic genes from hosts via lateral gene transfer.
Megavirales' challenge against pre-existing classifications of microbes has led scientists to think outside of the box, resulting in the term “Things Resisting Uncompleted Classifications”(TRUC). Forming a fourth branch of the microbial world, TRUC consists of microbes defying traditional classification. Within this category, Megavirales, Pandoraviruses, and other giant viruses form a "supergroup" due to their unusual mixture of genomic and phenotypic features.
But why should we care about viruses from soil? We don’t live in the soil, and soil viruses are not likely to cause infection in humans. Still, these creatures play important roles in soil ecology, exemplified in carbon cyclings. In the permafrost thaw gradient of northern Sweden, Trouble et al. (2018), utilized viral operational taxonomic units (vOTUs), finding linkages to lineages of Acidobacteria, Deltaproteobacteria, and Verrucomicrobia that resulted in the association of 30 auxiliary metabolic genes (AMGs) presumed to play a role in carbon metabolism. More recently, data from soil samples from Santa Fe and Pajarito (both in New Mexico), suggests that increased bacterial abundance can, via altering dynamics between fellow microbes, change both carbon and nutrient cycling. Although terrestrial microbiomes do not share "the viral shunt" of their aquatic counterparts, viral infection (caused by RNA viruses) triggering apoptosis is hypothesized to mobilize cell carbon that plays a vital role in carbon cycling. Without soil viruses, the delicate ecosystem upon which much of our food supply rests would deteriorate. With technologies like mini-metagenomics, the continued identification of new viruses and increased understanding of viral function and phylogeny, advances in the field of soil microbiology can not only be hoped for, but expected.
Abby is an incoming first-year student at Harvard College with an intended joint concentration in molecular and cellular biology and physics; post-grad, she hopes to become a microbiologist. She is currently a summer researcher in the Isberg lab at Tufts Medical School, working with Legionella pneumophila.