Summary The rice XA21‐mediated immune response is activated on recognition of the RaxX peptide produced by the bacterium Xanthomonas oryzae pv. oryzae ( Xoo ). The 60‐residue RaxX precursor is post‐translationally modified to form a sulfated tyrosine peptide that shares sequence and functional similarity with the plant sulfated tyrosine (PSY) peptide hormones. The 5‐kb raxX‐raxSTAB gene cluster of Xoo encodes RaxX, the RaxST tyrosylprotein sulfotransferase, and the RaxA and RaxB components of a predicted type I secretion system. To assess raxX‐raxSTAB gene cluster evolution and to determine its phylogenetic distribution, we first identified rax gene homologues in other genomes. We detected the complete raxX‐raxSTAB gene cluster only in Xanthomonas spp., in five distinct lineages in addition to X. oryzae . The phylogenetic distribution of the raxX‐raxSTAB gene cluster is consistent with the occurrence of multiple lateral (horizontal) gene transfer events during Xanthomonas speciation. RaxX natural variants contain a restricted set of missense substitutions, as expected if selection acts to maintain peptide hormone‐like function. Indeed, eight RaxX variants tested all failed to activate the XA21‐mediated immune response, yet retained peptide hormone activity. Together, these observations support the hypothesis that the XA21 receptor evolved specifically to recognize Xoo RaxX.

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Worldwide, virus infections in grapevines are of concern due to the potential for economic loss. Although the grape industry in Mexico is relatively small and focused mainly on the local market, production dates back to the time of the Spanish colonization. This manuscript discusses the findings on grapevine viruses in Mexico. Nine viruses have been identified in the last fifty years, including grapevine red blotch virus (GRBV), grapevine leafroll-associated virus 3 (GLRaV-3), grapevine fanleaf virus (GFLV), and grapevine virus A (GVA). Important information is provided about these viruses and viral pathogens that have not yet been reported in Mexico, but represent an ongoing threat to plant health and grapevine production in other viticultural regions of the world. Strategies for virus control in vineyards are described. The information discussed here should be shared with growers and stakeholders to prevent future negative impacts on the Mexican grapevine industry and to save ancient grapevine accessions.

The genus Vitivirus in the family Betaflexiviridae includes eleven viruses known to infect grapevine: grapevine vitiviruses A, B, D, E, F, G, H, I, J, L and M (GVA-GVM). Three of these viruses, GVA, GVB and GVD, have been associated with the etiology of rugose wood disease in grapevine and cause agronomically significant losses. The other vitiviruses were more recently discovered and their effects on grapevine are undetermined. To certify grape material for propagation as virus tested, an updated reverse transcription PCR (RT-PCR) assay to detect all known vitiviruses is desirable. To accomplish this, multiple grapevine vitivirus sequences were aligned at the amino acid level to search for conserved motifs. Two highly conserved motifs were found at an ideal distance for RT-PCR detection in the RNAdependent RNA polymerase region of the replicase protein. The amino acid motifs were back translated to create degenerate primers and used to successfully amplify all eleven grapevine vitiviruses. The RT-PCR primers were used to test a panel of vitivirus-infected vines for inclusivity as well as vines infected with closely related viruses in the Betaflexiviridae family (i.e. grapevine pinot gris virus and grapevine rupestris stem pitting-associated virus) for exclusivity. Broader use of these primers to detect vitiviruses in other plant hosts was investigated. In summary, an end-point RT-PCR assay that detects all the known grapevine vitiviruses and potentially other members of the genus Vitivirus has been developed. The universal assay represents an alternative to individual assays to reduce the work associated with the diagnosis of vitiviruses, including for regulatory purposes.

Running head: raxX-raxSTAB gene cluster in Xanthomonas spp. SummaryThe rice XA21-mediated immune response is activated upon recognition of the RaxX peptide 1 produced by the bacterium Xanthomonas oryzae pv. oryzae (Xoo). The 60 residue RaxX 2 precursor is posttranslationally modified to form a sulfated tyrosine peptide that shares sequence 3 and functional similarity with the plant sulfated tyrosine (PSY) peptide hormones. The five kb 4 raxX-raxSTAB gene cluster of Xoo encodes RaxX, the RaxST tyrosylprotein sulfotransferase, 5 and the RaxA and RaxB components of a predicted type one secretion system. The identified the 6 complete raxX-raxSTAB gene cluster is present only in Xanthomonas spp., in five distinct 7 lineages in addition to X. oryzae. The phylogenetic distribution of the raxX-raxSTAB gene 8 cluster is consistent with the occurrence of multiple lateral transfer events during Xanthomonas 9 speciation. RaxX variants representing each of the five lineages, and three Xoo RaxX variants, 10 fail to activate the XA21-mediated immune response yet retain peptide hormone activity. These 11 RaxX variants contain a restricted set of missense mutations, consistent with the hypothesis that 12 selection acts to maintain peptide hormone-like function. These observations are also consistent 13 with the hypothesis that the XA21 receptor evolved specifically to recognize Xoo RaxX. 14 rax gene cluster These results suggest that RaxX recognition by XA21 is restrained by different sequence and 53 length requirements compared to its recognition by the root growth promoting receptor(s) for 54 PSY hormone(s). It also suggests that recognition of RaxX by XA21 is specific to Xoo, whereas 55 PSY mimicry is a general feature of RaxX from other Xanthomonas spp. Accordingly, we 56 hypothesized that PSY hormone mimicry is the original function of RaxX, whereas immune 57 recognition by XA21 evolved later in response to Xoo (Pruitt et al., 2017). 58Two general predictions derive from this hypothesis. The first prediction, that PSY hormone 59 mimicry is broadly selective, is supported here by the presence of the raxX-raxSTAB gene cluster 60Xanthomonas spp., and by the ability of all RaxX variants tested to stimulate root growth in an 61 assay for PSY function. The second prediction, that recognition by XA21 is restricted to X. 62 oryzae lineages, is validated here by the observation that XA21-mediated immunity is not 63 activated by RaxX variants from other Xanthomonas spp. These results illustrate how a 64 pathogen protein has evolved to retain its ability to modulate host physiology without being 65 recognized by the host immune system. 66 rax gene cluster RESULTSThe raxX-raxSTAB gene cluster is present in a subset of Xanthomonas spp.We searched databases at the National Center for Biotechnology Information to identify bacterial 67 genomes with the raxX-raxSTAB gene cluster. We found the intact raxX-raxSTAB gene cluster 68 exclusively in Xanthomonas spp., and ultimately detected it in more than 200 unique genome 69 sequences (File. S1) ...