Several dsRNA bands (approx. 0.6–7 kbp in size) were recovered from tissues of mosaic-diseased fig seedlings which contained the enveloped round structures known as double membrane bodies (DMBs). blast analysis of a 4353 and a 1120 nt sequence from the two largest RNA segments showed homology with the polymerase and the putative glycoprotein precursor genes of negative-sense single-stranded RNA viruses of the family Bunyaviridae. Negative- and positive-sense riboprobes designed from both RNA segments hybridized to two bands of approximately 7 and 2.3 kbp in Northern blots of dsRNAs. Thus, these segments were identified as putative RNA-1 and RNA-2 of a novel virus for which the name fig mosaic virus (FMV) is proposed. Identity levels of predicted amino acids of the protein encoded by FMV RNA-1 with those of species of the family Bunyaviridae and European mountain ash ringspot-associated virus (EMERaV) were 28 and 54 %, respectively. RNA-2 showed 38 % identity at the amino acid level only with EMARaV. RNA-1 segment contained five conserved motifs (A–E) and an endonucleolytic centre of comparable genes of L RNA of bunyaviruses and EMARaV RNA-1. In a phylogenetic tree constructed with RdRp sequences, EMARaV grouped with FMV in a clade distinct from those of all bunyavirus genera. The consistent association of DMBs with mosaic symptoms and the results of molecular investigations strongly indicate that DMBs are particles of FMV, the aetiological agent of fig mosaic disease.

Polyploidization is a fundamental process in plant evolution. One of the biggest challenges faced by a new polyploid is meiosis, particularly discriminating between multiple related chromosomes so that only homologous chromosomes synapse and recombine to ensure regular chromosome segregation and balanced gametes. Despite its large genome size, high DNA repetitive content and similarity between homoeologous chromosomes, hexaploid wheat completes meiosis in a shorter period than diploid species with a much smaller genome. Therefore, during wheat meiosis, mechanisms additional to the classical model based on DNA sequence homology, must facilitate more efficient homologous recognition. One such mechanism could involve exploitation of differences in chromosome structure between homologs and homoeologs at the onset of meiosis. In turn, these chromatin changes, can be expected to be linked to transcriptional gene activity. In this study, we present an extensive analysis of a large RNA-seq data derived from six different genotypes: wheat, wheat–rye hybrids and newly synthesized octoploid triticale, both in the presence and absence of the Ph1 locus. Plant material was collected at early prophase, at the transition leptotene-zygotene, when the telomere bouquet is forming and synapsis between homologs is beginning. The six genotypes exhibit different levels of synapsis and chromatin structure at this stage; therefore, recombination and consequently segregation, are also different. Unexpectedly, our study reveals that neither synapsis, whole genome duplication nor the absence of the Ph1 locus are associated with major changes in gene expression levels during early meiotic prophase. Overall wheat transcription at this meiotic stage is therefore highly resilient to such alterations, even in the presence of major chromatin structural changes. Further studies in wheat and other polyploid species will be required to reveal whether these observations are specific to wheat meiosis.

Polyploidization has played an important role in plant evolution. However, upon polyploidization, the process of meiosis must adapt to ensure the proper segregation of increased numbers of chromosomes to produce balanced gametes. It has been suggested that meiotic gene (MG) duplicates return to a single copy following whole genome duplication to stabilize the polyploid genome. Therefore, upon the polyploidization of wheat, a hexaploid species with three related (homeologous) genomes, the stabilization process may have involved rapid changes in content and expression of MGs on homeologous chromosomes (homeologs). To examine this hypothesis, sets of candidate MGs were identified in wheat using co-expression network analysis and orthology informed approaches. In total, 130 RNA-Seq samples from a range of tissues including wheat meiotic anthers were used to define co-expressed modules of genes. Three modules were significantly correlated with meiotic tissue samples but not with other tissue types. These modules were enriched for GO terms related to cell cycle, DNA replication, and chromatin modification and contained orthologs of known MGs. Overall, 74.4% of genes within these meiosis-related modules had three homeologous copies which was similar to other tissue-related modules. Amongst wheat MGs identified by orthology, rather than co-expression, the majority (93.7%) were either retained in hexaploid wheat at the same number of copies (78.4%) or increased in copy number (15.3%) compared to ancestral wheat species. Furthermore, genes within meiosis-related modules showed more balanced expression levels between homeologs than genes in non-meiosis-related modules. Taken together, our results do not support extensive gene loss nor changes in homeolog expression of MGs upon wheat polyploidization. The construction of the MG co-expression network allowed identification of hub genes and provided key targets for future studies.

Key message: The meiotic recombination geneDmc1on wheat chromosome 5D has been identified as a candidate for the maintenance of normal chromosome synapsis and crossover at low and possibly high temperatures. We initially assessed the effects of low temperature on meiotic chromosome synapsis and crossover formation in the hexaploid wheat (Triticum aestivumL.) variety ‘Chinese Spring’. At low temperatures, asynapsis and chromosome univalence have been observed before in Chinese Spring lines lacking the long arm of chromosome 5D (5DL), which led to the proposal that 5DL carries a gene (Ltp1) that stabilises wheat chromosome pairing at low temperatures. In the current study, Chinese Spring wild type and 5DL interstitial deletion mutant plants were exposed to low temperature in a controlled environment room during a period from premeiotic interphase to early meiosis I. A 5DL deletion mutant was identified whose meiotic chromosomes exhibit extremely high levels of asynapsis and chromosome univalence at metaphase I after 7 days at 13 °C, suggesting thatLtp1is deleted in this mutant. Immunolocalisation of the meiotic proteins ASY1 and ZYP1 onltp1mutants showed that low temperature results in a failure to complete synapsis at pachytene. KASP genotyping revealed that theltp1mutant has a 4-Mb deletion in 5DL. Of 41 genes within this deletion region, the strongest candidate for the stabilisation of chromosome pairing at low temperatures is the meiotic recombination geneDmc1.Theltp1mutants were subsequently treated at 30 °C for 24 h during meiosis and exhibited a reduced number of crossovers and increased univalence, though to a lesser extent than at 13 °C. We therefore renamed ourltp1mutant ‘ttmei1’ (temperature-tolerantmeiosis1) to reflect this additional loss of high temperature tolerance.

Although most flowering plants are polyploid, little is known of how the meiotic process evolves after polyploidisation to stabilise and preserve fertility. On wheat polyploidisation, the major meiotic gene ZIP4 on chromosome 3B duplicated onto 5B and diverged (TaZIP4-B2). TaZIP4-B2 was recently shown to promote homologous pairing, synapsis and crossover, and suppress homoeologous crossover. We therefore suspected that these meiotic stabilising effects could be important for preserving wheat fertility. A CRISPR Tazip4-B2 mutant was exploited to assess the contribution of the 5B duplicated ZIP4 copy in maintaining pollen viability and grain setting. Analysis demonstrated abnormalities in 56% of meiocytes in the Tazip4-B2 mutant, with micronuclei in 50% of tetrads, reduced size in 48% of pollen grains and a near 50% reduction in grain number. Further studies showed that most of the reduced grain number occurred when Tazip4-B2 mutant plants were pollinated with the less viable Tazip4-B2 mutant pollen rather than with wild type pollen, suggesting that the stabilising effect of TaZIP4-B2 on meiosis has a greater consequence in subsequent male, rather than female gametogenesis. These studies reveal the extraordinary value of the wheat chromosome 5B TaZIP4-B2 duplication to agriculture and human nutrition. Future studies should further investigate the role of TaZIP4-B2 on female fertility and assess whether different TaZIP4-B2 alleles exhibit variable effects on meiotic stabilisation and/or resistance to temperature change.