Diversity of Giant Viruses Infecting Vermamoeba vermiformis.

The discovery of Acanthamoeba polyphaga mimivirus in 2003 using the free-living amoeba Acanthamoeba polyphaga caused a paradigm shift in the virology field. Twelve years later, using another amoeba as a host, i.e., Vermamoeba vermiformis, novel isolates of giant viruses have been discovered. This amoeba-virus relationship led scientists to study the evolution of giant viruses and explore the origins of eukaryotes. The purpose of this article is to review all the giant viruses that have been isolated from Vermamoeba vermiformis, compare their genomic features, and report the influence of these viruses on the cell cycle of their amoebal host. To date, viruses putatively belonging to eight different viral taxa have been described: 7 are lytic and 1 is non-lytic. The comparison of giant viruses infecting Vermamoeba vermiformis has suggested three homogenous groups according to their size, the replication time inside the host cell, and the number of encoding tRNAs. This approach is an attempt at determining the evolutionary origins and trajectories of the virus; therefore, more giant viruses infecting Vermamoeba must be discovered and studied to create a comprehensive knowledge on these intriguing biological entities.

Pacmanvirus S19, the Second Pacmanvirus Isolated from Sewage Waters in Oran, Algeria.

Acanthamoeba castellanii is an amoeba host that was used to isolate a novel strain named pacmanvirus S19. This isolate is the second strain reported and belongs to the extended Asfarviridae family. Pacmanvirus S19 harbors a 418,588-bp genome, with a GC content of 33.20%, which encodes 444 predicted proteins and a single Ile-tRNA.

The Kaumoebavirus LCC10 Genome Reveals a Unique Gene Strand Bias among "Extended Asfarviridae".

Kaumoebavirus infects the amoeba Vermamoeba vermiformis and has recently been described as a distant relative of the African swine fever virus. To characterize the diversity and evolution of this novel viral genus, we report here on the isolation and genome sequencing of a second strain of Kaumoebavirus, namely LCC10. Detailed analysis of the sequencing data suggested that its 362-Kb genome is linear with covalently closed hairpin termini, so that DNA forms a single continuous polynucleotide chain. Comparative genomic analysis indicated that although the two sequenced Kaumoebavirus strains share extensive gene collinearity, 180 predicted genes were either gained or lost in only one genome. As already observed in another distant relative, i.e., Faustovirus, which infects the same host, the center and extremities of the Kaumoebavirus genome exhibited a higher rate of sequence divergence and the major capsid protein gene was colonized by type-I introns. A possible role of the Vermamoeba host in the genesis of these evolutionary traits is hypothesized. The Kaumoebavirus genome exhibited a significant gene strand bias over the two-third of genome length, a feature not seen in the other members of the "extended Asfarviridae" clade. We suggest that this gene strand bias was induced by a putative single origin of DNA replication located near the genome extremity that imparted a selective force favoring the genes positioned on the leading strand.

Comparative Genomics Unveils Regionalized Evolution of the Faustovirus Genomes.

Faustovirus is a recently discovered genus of large DNA virus infecting the amoeba Vermamoeba vermiformis, which is phylogenetically related to Asfarviridae. To better understand the diversity and evolution of this viral group, we sequenced six novel Faustovirus strains, mined published metagenomic datasets and performed a comparative genomic analysis. Genomic sequences revealed three consistent phylogenetic groups, within which genetic diversity was moderate. The comparison of the major capsid protein (MCP) genes unveiled between 13 and 18 type-I introns that likely evolved through a still-active birth and death process mediated by intron-encoded homing endonucleases that began before the Faustovirus radiation. Genome-wide alignments indicated that despite genomes retaining high levels of gene collinearity, the central region containing the MCP gene together with the extremities of the chromosomes evolved at a faster rate due to increased indel accumulation and local rearrangements. The fluctuation of the nucleotide composition along the Faustovirus (FV) genomes is mostly imprinted by the consistent nucleotide bias of coding sequences and provided no evidence for a single DNA replication origin like in circular bacterial genomes.

A porous graphitized carbon LC-ESI/MS method for the quantitation of metronidazole and fluconazole in breast milk and human plasma.

Information on drug transfer into the breast milk is essential to protect the infant from undesirable adverse effects of maternal consumption of drugs and to allow effective pharmacological treatment of breastfeeding mothers. Metronidazole and fluconazole are two drugs frequently used in nursing women to treat various infections, thus questioning infant's safety due to drug exposure through breast milk. In this article a porous graphitized carbon LC/ESI-MS assay was developed for the quantitation of metronidazole and fluconazole in breast milk and human plasma. The assay was based on the use of 150 µL of biological samples, following acetonitrile precipitation of proteins and filtration that enabled injection into the LC/ESI-MS system. All analytes and the internal standard, ropinirole, were separated by using a porous graphitized carbon analytical column (150 × 2.1 mm i.d., particle size 5 µm) with isocratic elution. The mobile phase consists of 55% acetonitrile in water acidified with 0.1% concentrated formic acid and pumped at a flow rate of 0.25 mL min-1. The assay was linear over a concentration range of 0.1 to 15 µg mL-1 for all analytes in both biological samples. Intermediate precision was found to be <8.4% over the tested concentration ranges. A run time of <5 min for each sample made it possible to analyze a large number of biological samples per day. The method is the first reported application for the analysis of metronidazole and fluconazole in both breast milk and human plasma and it can be used to support a wide range of clinical studies.