Genetic Variability of Natural Populations of Grapevine leafroll-associated virus 2 in Pacific Northwest Vineyards

Genetic variability of field populations of Grapevine leafroll-associated virus 2 (GLRaV-2) in Pacific Northwest (PNW) vineyards was characterized by sequencing the entire coat protein (CP) and a portion of the heat-shock protein-70 homolog (HSP70h) genes. Phylogenetic analysis of CP and HSP70h nucleotide sequences obtained in this study and corresponding sequences from GenBank revealed segregation of GLRaV-2 isolates into six lineages with virus isolates from PNW distributed in ‘PN’, ‘H4’, and ‘RG’ lineages. An estimation of the ratio of nonsynonymous substitutions per nonsynonymous site to synonymous substitutions per synonymous site indicated that different selection pressures may be acting on the two genomic regions encoding proteins with distinct functions. Multiple alignments of CP amino acid sequences showed lineage-specific differences. Enzyme-linked immunosorbent assay results indicated that GLRaV-2-specific antibodies from a commercial source are unable to reliably detect GLRaV-2 isolates in the RG lineage, thereby limiting antibody-based diagnosis of all GLRaV-2 isolates currently found in PNW vineyards. A protocol based on reverse-transcription polymerase chain reaction and restriction fragment length polymorphism analysis was developed for differentiating GLRaV-2 isolates belonging to the three lineages present in the region. The taxonomic status of GLRaV-2 is discussed in light of the current knowledge of global genetic diversity of the virus.

Subfamilies of cpmA, a gene involved in circadian output, have different evolutionary histories in cyanobacteria

The cpmA gene mediates an output signal in the cyanobacterial circadian system. This gene and its homologues are evolutionarily old, and occur in some non-photosynthetic bacteria and archaea as well as in cyanobacteria. The gene has two functional domains that differ drastically in their level of polymorphism: the N-terminal domain is much more variable than the PurE homologous C-terminal domain. The phylogenetic tree of the cpmA homologues features four main clades (C1–C4), two of which (C1 and C3) belong to cyanobacteria. These cyanobacterial clades match respective ones in the previously reported phylogenetic trees of the other genes involved in the circadian system. The phylogenetic analysis suggested that the C3 subfamily, which comprises the genes from the cyanobacteria with the kaiBC-based circadian system, experienced a lateral transfer, probably from evolutionarily old proteobacteria about 1000 million years ago. The genes of this subfamily have a significantly higher nonsynonymous substitution rate than those of C1 (2·13×10−10 and 1·53×10−10 substitutions per nonsynonymous site per year, respectively). It appears that the functional and selective constraints of the kaiABC-based system have slowed down the rate of sequence evolution compared to the cpmA homologues of the kaiBC-based system. On the other hand, the differences in the mutation rates between the two cyanobacterial clades point to the different functional constraints of the systems with or without kaiA.

Coevolution of theS-Locus GenesSRK,SLGandSP11/SCRinBrassica oleraceaandB. rapa

Brassica self-incompatibility (SI) is controlled by SLG and SRK expressed in the stigma and by SP11/SCR expressed in the anther. We determined the sequences of the S domains of 36 SRK alleles, 13 SLG alleles, and 14 SP11 alleles from Brassica oleracea and B. rapa. We found three S haplotypes lacking SLG genes in B. rapa, confirming that SLG is not essential for the SI recognition system. Together with reported sequences, the nucleotide diversities per synonymous and nonsynonymous site (πS and πN) at the SRK, SLG, and SP11 loci within B. oleracea were computed. The ratios of πN:πS for SP11 and the hypervariable region of SRK were significantly >1, suggesting operation of diversifying selection to maintain the diversity of these regions. In the phylogenetic trees of 12 SP11 sequences and their linked SRK alleles, the tree topology was not significantly different between SP11 and SRK, suggesting a tight linkage of male and female SI determinants during the evolutionary course of these haplotypes. Genetic exchanges between SLG and SRK seem to be frequent; three such recent exchanges were detected. The evolution of S haplotypes and the effect of gene conversion on self-incompatibility are discussed.