Viruses in a 14th-Century Coprolite

Abstract

Coprolites are fossilized fecal material that can reveal information about ancient intestinal and environmental microbiota. Viral metagenomics has allowed systematic characterization of viral diversity in environmental and human-associated specimens, but little is known about the viral diversity in fossil remains. Here, we analyzed the viral community of a 14th-century coprolite from a closed barrel in a Middle Ages site in Belgium using electron microscopy and metagenomics. Viruses that infect eukaryotes, bacteria, and archaea were detected, and we confirmed the presence of some of them by ad hoc suicide PCR. The coprolite DNA viral metagenome was dominated by sequences showing homologies to phages commonly found in modern stools and soil. Although their phylogenetic compositions differed, the metabolic functions of the viral communities have remained conserved across centuries. Antibiotic resistance was one of the reconstructed metabolic functions detected. V iral metagenomics is a sequencing-based analysis of all of viral genomes isolated from a sample. It has promoted the characterization of viral community diversity. Viral metagenomics has already been successfully applied to the exploration of modern environmental specimens sampled from marine water, freshwater, stromatolites and thrombolites, and soil (1-4) and to modern human-associated specimens collected from the liver, blood, nasopharyngeal aspirates, and stool (5-9). The DNA viromes generated from modern stools have been demonstrated to be dominated by bacteriophages (10, 11) and to be less diverse than environmental samples (8,12).Viral metagenomics does not require culturing viruses or a priori knowledge of the sequences that will be targeted, which allows for the identification of new, unknown or unexpected viruses and for the global assessment of the virome. Viral metagenomics is thus particularly suitable for paleomicrobiological studies, as little is known about which viruses are characteristic of ancient specimens. Indeed, the majority of ancient DNA (aDNA) studies are based on the analysis of human and bacterial aDNA (13-15), and viral persistence and its detectability in ancient specimens remain unclear. Electron microscopy has previously revealed that viral particles can persist for over 400 years, but their viability was lost (16). Moreover, PCR amplifications yielded positive results for viral aDNA in ancient specimens, such as mummified soft tissues, bones, and teeth. The amplification products varied between 100 and 570 bp in size, which indicated that viral aDNA can be detected for at least 1,500 years (17-20).Here, we used electron microscopy and, for what may be the first time, viral metagenomics to characterize the viral community of an ancient stool specimen. A viral DNA metagenome was generated from a 14th-century coprolite sample that was recovered from a Middle Ages site in Namur, Belgium. MATERIALS AND METHODSVLP isolation, TEM, and DNA extraction. First, 5.8 g of the interior of the coprolite was aseptically removed and solubi...

A century of bacteriophage research and applications: impacts on biotechnology, health, ecology and the economy!

“…Now the favour is returned with novel molecular biotechniques such as clustered regularly interspaced short palindromic repeats (CRISPR)‐Cas9 being used to modify industrial biotechnologically important bacterial strains, such as Bacillus subtilis and E. coli , to increase their phage resistance during large‐scale fermentation processes . Also, viral metagenomics, complete phage genome sequencing, phage‐mediated efficient gene transfer and molecular characterization of phage lytic enzymes (endolysins) are now routinely studied in view of improved applications of phages to combat pathogens and spoilage bacteria in the medical and agri‐food sector …”

“…Evidence is accumulating for the coevolution of antibiotic resistance and antibiotic biosynthesis genes in soil bacteria such as Streptomyces species and similarly for a coevolution of bacteria and phage populations in nature as well as in experimental batch cultures and in chemostat microcosms . In this context it is essential to understand resistance build‐up in function of the bacterial composition or sequence and selection of the ‘best practice’ use of single phages or of phage cocktails …”

Ancient human microbiomes

“…Recently, shotgun NGS and scanning electron microscopy (SEM) was applied to medieval coprolites sealed within latrine barrels with the aim of retrieving ancient gut microbiome viruses (Appelt et al, 2014). By comparing the resulting metagenomic sequences to the NCBI RefSeq Viral Genomes database, several thousand DNA sequences were identified with homology to known viral families, including a few that infect eukaryotes, but mostly belonging to the Siphoviridae family of double-stranded DNA bacteriophages.…”

Preservation of viral genomes in 700-y-old caribou feces from a subarctic ice patch

“…The genome of Barley stripe mosaic virus RNA was detected by RT-PCR in 600-900 y before present (BP) barley grains from North Africa and further sequenced using the Illumina Hi-Seq platform (4). Descriptions of ancient DNA viruses include a bacteriophage from dehydrated feces in a closed barrel buried in a 14th-century latrine in Belgium and plankton-infecting coccolithovirus phage DNA detected by PCR from sediments as old as 7,000 y under the Black sea floor (5,6). A replicationcompetent pandoravirus-like Pithovirus with a 600-kb genome was reported isolated following Acantamoeba inoculation with 30,000-y-old thawed Siberian permafrost (7).…”