Bou Khalil, J. Y. | Search bioRxiv

bioRxiv2020

Advantages and limits of metagenomic assembly and binning of a giant virus

Giant viruses have large genomes, often within the size range of cellular organisms. This distinguishes them from most other viruses and demands additional effort for the successful recovery of their genomes from environmental sequence data. Here we tested the performance of genome-resolved metagenomics on a recently isolated giant virus, Fadolivirus, by spiking it into an environmental sample from which two other giant viruses were isolated. At high spike-in levels, metagenome assembly and binning led to the successful genomic recovery of Fadolivirus from the sample. A complementary survey of viral hallmark genes indicated the presence of other giant viruses in the sample matrix, but did not detect the two isolated from this sample. Our results indicate that genome-resolved metagenomics is a valid approach for the recovery of near-complete giant virus genomes given that sufficient clonal particles are present. Our data also underline that a vast majority of giant viruses remain currently undetected, even in an era of terabase-scale metagenomics.

microbiology↗

bioRxiv2019

High Content Screening, a reliable system for Coxiella burnetii isolation from clinical samples

Q fever, caused by Coxiella burnetii, is a worldwide zoonotic disease that may cause severe forms in humans and requires a specific and prolonged antibiotic treatment. Although the current serological and molecular detection tools enable a reliable diagnosis of the disease, culture of C. burnetii strains is mandatory to evaluate their antibiotic susceptibility and sequence their genome in order to optimize patient management and epidemiological studies. However, cultivating this fastidious microorganism is difficult and restricted to reference centers as it requires biosafety-level 3 laboratories and relies on cell culture performed by experienced technicians. In addition, the culture yield is low, which results in a small number of isolates being available. In this work, we developed a novel high content screening (HCS) isolation strategy based on optimized high-throughput cell culture and automated microscopic detection of infected cells with specifically-designed algorithms targeting cytopathic effects. This method was more efficient than the shell-vial assay when applied to both frozen specimens (7 isolates recovered by HCS only, sensitivity 91% vs 78% for shell-vial) and fresh samples (1 additional isolate using HCS, sensitivity 7% vs 5% for shell-vial). In addition, detecting positive cultures by an automated microscope reduced the need for expertise and saved 24% of technician working time. Application of HCS to antibiotic susceptibility testing of 12 strains demonstrated that it was as efficient as the standard procedure that combines shell-vial culture and quantitative PCR. Overall, this high-throughput HCS system paves the way to the development of improved cell culture isolation of human viruses.

microbiology↗