Gloria Mosquera

Detection of the Cassava Bacterial Blight Pathogen, Xanthomonas axonopodis pv. manihotis, by Polymerase Chain Reaction

Cassava bacterial blight, caused by Xanthomonas axonopodis pv. manihotis, is of significant concern wherever cassava is grown. The movement of infected, asymptomatic stems is a major means of pathogen dispersal. A reliable and sensitive diagnostic procedure is necessary for the safe movement of cassava planting material. We used a cloned and sequenced pathogenicity gene of X. axonopodis pv. manihotis to develop a polymerase chain reaction (PCR) test for this pathogen. A set of primers directed the amplification of an 898-bp fragment in all 107 pathogenic strains of X. axonopodis pv. manihotis tested. PCR products were not observed when genomic DNA was tested for 27 strains of other xanthomonads, for saprophytic bacteria, or for five nonpathogenic strains of X. axonopodis pv. manihotis. The primers worked well for pathogen detection in direct PCR assays of X. axonopodis pv. manihotis colonies grown on liquid medium and in PCR assays of extracts from leaf and stem lesions. The minimum number of cells that could be detected from cassava stem and leaf lesions was 3 × 102 to 104 CFU/ml. The PCR assays proved to be relativyel sensitive and could become very useful in detecting the pathogen in cassava planting material.

Identification of genes in cassava that are differentially expressed during infection with Xanthomonas axonopodis pv. manihotis.

SUMMARY The cDNA-amplified fragment length polymorphism approach was used to identify differentially expressed transcripts from cassava infected by Xanthomonas axonopodis pv. manihotis (Xam). Approximately 3600 transcript-derived fragments (TDFs) were screened of which 340 were isolated. The nucleotide sequences of 250 TDFs were analysed and assembled into contigs and singletons. The amino acid sequences of their predicted products were compared with entries in databases and 63 of these clones showed homology to known plant genes. Of these, 32 showed similarity to plant defence proteins. Fifty-one TDFs corresponded to proteins of unknown function and 106 did not match any sequence in the public databases. Quantitative reverse transcription PCR was carried out with a selected set of gene transcripts that demonstrated an increase of expression during the infection. These results point out candidate genes that are associated with cassava resistance to Xam and reinforce the idea of a complex process occurring during this plant-pathogen interaction.

Comparative Analysis of Two Emerging Rice Seed Bacterial Pathogens

Seed sterility and grain discoloration limit rice production in Colombia and several Central American countries. In samples of discolored rice seed grown in Colombian fields, the species Burkholderia glumae and B. gladioli were isolated, and field isolates were compared phenotypically. An artificial inoculation assay was used to determine that, although both bacterial species cause symptoms on rice grains, B. glumae is a more aggressive pathogen, causing yield reduction and higher levels of grain sterility. To identify putative virulence genes differing between B. glumae and B. gladioli, four previously sequenced genomes of Asian and U.S. strains of the two pathogens were compared with each other and with two draft genomes of Colombian B. glumae and B. gladioli isolates generated for this study. Whereas previously characterized Burkholderia virulence factors are highly conserved between the two species, B. glumae and B. gladioli strains are predicted to encode distinct groups of genes encoding type VI secretion systems, transcriptional regulators, and membrane-sensing proteins. This study shows that both B. glumae and B. gladioli can threaten grain quality, although only one species affects yield. Furthermore, genotypic differences between the two strains are identified that could contribute to disease phenotypic differences.

Resistance to Multiple Temperate and Tropical Stem and Sheath Diseases of Rice

Reaction to sheath blight, stem rot, and aggregated sheath spot were tested in 641 tropical japonica and indica rice lines. Disease resistance was mapped independently from flowering time and plant height. Quantitative trait loci of major effect for resistance to the three diseases were found. A multiple disease resistance quantitative trait locus was found on chromosome 9 across tropical japonica and indica populations.

Allelic variation for broad-spectrum resistance and susceptibility to bacterial pathogens identified in a rice MAGIC population

Summary Quantitative trait loci (QTL) that confer broad‐spectrum resistance (BSR), or resistance that is effective against multiple and diverse plant pathogens, have been elusive targets of crop breeding programmes. Multiparent advanced generation intercross (MAGIC) populations, with their diverse genetic composition and high levels of recombination, are potential resources for the identification of QTL for BSR. In this study, a rice MAGIC population was used to map QTL conferring BSR to two major rice diseases, bacterial leaf streak (BLS) and bacterial blight (BB), caused by Xanthomonas oryzae pathovars (pv.) oryzicola (Xoc) and oryzae (Xoo), respectively. Controlling these diseases is particularly important in sub‐Saharan Africa, where no sources of BSR are currently available in deployed varieties. The MAGIC founders and lines were genotyped by sequencing and phenotyped in the greenhouse and field by inoculation with multiple strains of Xoc and Xoo. A combination of genomewide association studies (GWAS) and interval mapping analyses revealed 11 BSR QTL, effective against both diseases, and three pathovar‐specific QTL. The most promising BSR QTL (qXO‐2‐1, qXO‐4‐1 and qXO‐11‐2) conferred resistance to more than nine Xoc and Xoo strains. GWAS detected 369 significant SNP markers with distinguishable phenotypic effects, allowing the identification of alleles conferring disease resistance and susceptibility. The BSR and susceptibility QTL will improve our understanding of the mechanisms of both resistance and susceptibility in the long term and will be immediately useful resources for rice breeding programmes.