Genome-wide association analysis and differential expression analysis of resistance to Sclerotinia stem rot in Brassica napus

Brassica napus is one of the most important oil crops in the world, and stem rot caused by the fungus Sclerotinia sclerotiorum results in major losses in yield and quality. To elucidate resistance genes and pathogenesis-related genes, genome-wide association analysis of 347 accessions was performed using the Illumina 60K Brassica SNP (single nucleotide polymorphism) array. In addition, the detached stem inoculation assay was used to select five highly resistant (R) and susceptible (S) B. napus lines, 48 h postinoculation with S. sclerotiorum for transcriptome sequencing. We identified 17 significant associations for stem resistance on chromosomes A8 and C6, five of which were on A8 and 12 on C6. The SNPs identified on A8 were located in a 409-kb haplotype block, and those on C6 were consistent with previous QTL mapping efforts. Transcriptome analysis suggested that S. sclerotiorum infection activates the immune system, sulphur metabolism, especially glutathione (GSH) and glucosinolates in both R and S genotypes. Genes found to be specific to the R genotype related to the jasmonic acid pathway, lignin biosynthesis, defence response, signal transduction and encoding transcription factors. Twenty-four genes were identified in both the SNP-trait association and transcriptome sequencing analyses, including a tau class glutathione S-transferase (GSTU) gene cluster. This study provides useful insight into the molecular mechanisms underlying the plant's response to S. sclerotiorum.

Pasteurella multocida Expresses Two Lipopolysaccharide Glycoforms Simultaneously, but Only a Single Form Is Required for Virulence: Identification of Two Acceptor-Specific Heptosyl I Transferases

Lipopolysaccharide (LPS) is a critical virulence determinant in Pasteurella multocida and a major antigen responsible for host protective immunity. In other mucosal pathogens, variation in LPS or lipooligosaccharide structure typically occurs in the outer core oligosaccharide regions due to phase variation. P. multocida elaborates a conserved oligosaccharide extension attached to two different, simultaneously expressed inner core structures, one containing a single phosphorylated 3-deoxy-D-manno-octulosonic acid (Kdo) residue and the other containing two Kdo residues. We demonstrate that two heptosyltransferases, HptA and HptB, add the first heptose molecule to the Kdo1 residue and that each exclusively recognizes different acceptor molecules. HptA is specific for the glycoform containing a single, phosphorylated Kdo residue (glycoform A), while HptB is specific for the glycoform containing two Kdo residues (glycoform B). In addition, KdkA was identified as a Kdo kinase, required for phosphorylation of the first Kdo molecule. Importantly, virulence data obtained from infected chickens showed that while wild-type P. multocida expresses both LPS glycoforms in vivo, bacterial mutants that produced only glycoform B were fully virulent, demonstrating for the first time that expression of a single LPS form is sufficient for P. multocida survival in vivo. We conclude that the ability of P. multocida to elaborate alternative inner core LPS structures is due to the simultaneous expression of two different heptosyltransferases that add the first heptose residue to the nascent LPS molecule and to the expression of both a bifunctional Kdo transferase and a Kdo kinase, which results in the initial assembly of two inner core structures.

Wet-dry cycling extends seed persistence by re-instating antioxidant capacity

Seeds in the field experience wet-dry cycling that is akin to the well-studied commercial process of seed priming in which seeds are hydrated and then re-dried to standardise their germination characteristics. To investigate whether the persistence (defined as in situ longevity) and antioxidant capacity of seeds are influenced by wet-dry cycling, seeds of the global agronomic weed Avena sterilis ssp. ludoviciana were subjected to (1) controlled ageing at 60% relative humidity and 53.5°C for 31 days, (2) controlled ageing then priming, or (3) ageing in the field in three soils for 21 months. Changes in seed viability (total germination), mean germination time, seedling vigour (mean seedling length), and the concentrations of the glutathione (GSH) / glutathione disulphide (GSSG) redox couple were recorded over time. As controlled-aged seeds lost viability, GSH levels declined and the relative proportion of GSSG contributing to total glutathione increased, indicative of a failing antioxidant capacity. Subjecting seeds that were aged under controlled conditions to a wet-dry cycle (to −1 MPa) prevented viability loss and increased GSH levels. Field-aged seeds that underwent numerous wet-dry cycles due to natural rainfall maintained high viability and high GSH levels. Thus wet-dry cycles in the field may enhance seed longevity and persistence coincident with re-synthesis of protective compounds such as GSH.

Seminal plasma proteome of eletroejaculated Bos indicus bulls

The present study describes the seminal plasma proteome of Bos indicus bulls. Fifty-six, 24-month old Australian Brahman sires were evaluated and subjected to electroejaculation. Seminal plasma proteins were separated by 2-D SDS-PAGE and identified by mass spectrometry. The percentage of progressively motile and morphologically normal sperm of the bulls were 70.4±2.3 and 64±3.2%, respectively. A total of 108 spots were identified in the 2-D maps, corresponding to 46 proteins. Binder of sperm proteins accounted for 55.8% of all spots detected in the maps and spermadhesins comprised the second most abundant constituents. Other proteins of the Bos indicus seminal plasma include clusterin, albumin, transferrin, metalloproteinase inhibitor 2, osteopontin, epididymal secretory protein E1, apolipoprotein A-1, heat shock 70kDa protein, glutathione peroxidase 3, cathelicidins, alpha-enolase, tripeptidyl-peptidase 1, zinc-alpha-2-glycoprotein, plasma serine protease inhibitor, beta 2-microglobulin, proteasome subunit beta type-4, actin, cathepsins, nucleobinding-1, protein S100-A9, hemoglobin subunit alpha, cadherin-1, angiogenin-1, fibrinogen alpha and beta chain, ephirin-A1, protein DJ-1, serpin A3-7, alpha-2-macroglobulin, annexin A1, complement factor B, polymeric immunoglobulin receptor, seminal ribonuclease, ribonuclease-4, prostaglandin-H2 D-isomarase, platelet-activating factor acetylhydrolase, and phosphoglycerate kinase In conclusion, this work uniquely portrays the Bos indicus seminal fluid proteome, based on samples from a large set of animals representing the Brahman cattle of the tropical Northern Australia. Based on putative biochemical attributes, seminal proteins act during sperm maturation, protection, capacitation and fertilization.