Cassava begomovirus species diversity changes during plant vegetative cycles.

Cassava is a root crop important for global food security and the third biggest source of calories on the African continent. Cassava production is threatened by Cassava mosaic disease (CMD), which is caused by a complex of single-stranded DNA viruses (family: Geminiviridae, genus: Begomovirus) that are transmitted by the sweet potato whitefly (Bemisia tabaci). Understanding the dynamics of different cassava mosaic begomovirus (CMB) species through time is important for contextualizing disease trends. Cassava plants with CMD symptoms were sampled in Lake Victoria and coastal regions of Kenya before transfer to a greenhouse setting and regular propagation. The field-collected and greenhouse samples were sequenced using Illumina short-read sequencing and analyzed on the Galaxy platform. In the field-collected samples, African cassava mosaic virus (ACMV), East African cassava mosaic virus (EACMV), East African cassava mosaic Kenya virus (EACMKV), and East African cassava mosaic virus-Uganda variant (EACMV-Ug) were detected in samples from the Lake Victoria region, while EACMV and East African mosaic Zanzibar virus (EACMZV) were found in the coastal region. Many of the field-collected samples had mixed infections of EACMV and another begomovirus. After 3 years of regrowth in the greenhouse, only EACMV-like viruses were detected in all samples. The results suggest that in these samples, EACMV becomes the dominant virus through vegetative propagation in a greenhouse. This differed from whitefly transmission results. Cassava plants were inoculated with ACMV and another EACMV-like virus, East African cassava mosaic Cameroon virus (EACMCV). Only ACMV was transmitted by whiteflies from these plants to recipient plants, as indicated by sequencing reads and copy number data. These results suggest that whitefly transmission and vegetative transmission lead to different outcomes for ACMV and EACMV-like viruses.

Genome segment ratios change during whitefly transmission of two bipartite cassava mosaic begomoviruses.

Cassava mosaic disease is caused by a complex of whitefly-transmitted begomoviruses, which often occur in co-infections. These viruses have bipartite genomes consisting of DNA-A and DNA-B that are encapsidated into separate virions. Individual viruses exist in plants and whitefly vectors as populations comprising both genome segments, which can occur at different frequencies. Both segments are required for infection, and must be transmitted for virus spread to occur. Cassava plants infected with African cassava mosaic virus (ACMV) and/or East African cassava mosaic Cameroon virus (EACMCV), in which the ratios of DNA-A:DNA-B titers differed between plants, were used to examine how titers of the segments in a plant relate to their respective probabilities of acquisition by whiteflies and to the titers of each segment acquired and subsequently transmitted by whiteflies. The probabilities of acquiring each segment of ACMV did not reflect their relative titers in the source plant but they did for EACMCV. However, for both viruses, DNA-A:DNA-B ratios acquired by whiteflies differed from those in the source plant and the ratios transmitted by the whitefly did not differ from one - the ratio at which the highest probability of transmitting both segments is expected.

Identification and characterization of suppressors of plant cell death (SPD) effectors from Magnaporthe oryzae.

Phytopathogenic microorganisms, including the fungal pathogen Magnaporthe oryzae, secrete a myriad of effector proteins to facilitate infection. Utilizing the transient expression of candidate effectors in the leaves of the model plant Nicotiana benthamiana, we identified 11 suppressors of plant cell death (SPD) effectors from M. oryzae that were able to block the host cell death reaction induced by Nep1. Ten of these 11 were also able to suppress BAX-mediated plant cell death. Five of the 11 SPD genes have been identified previously as either essential for the pathogenicity of M. oryzae, secreted into the plant during disease development, or as suppressors or homologues of other characterized suppressors. In addition, of the remaining six, we showed that SPD8 (previously identified as BAS162) was localized to the rice cytoplasm in invaded and surrounding uninvaded cells during biotrophic invasion. Sequence analysis of the 11 SPD genes across 43 re-sequenced M. oryzae genomes revealed that SPD2, SPD4 and SPD7 have nucleotide polymorphisms amongst the isolates. SPD4 exhibited the highest level of nucleotide diversity of any currently known effector from M. oryzae in addition to the presence/absence polymorphisms, suggesting that this gene is potentially undergoing selection to avoid recognition by the host. Taken together, we have identified a series of effectors, some of which were previously unknown or whose function was unknown, that probably act at different stages of the infection process and contribute to the virulence of M. oryzae.

Oxygenase coordination is required for morphological transition and the host-fungus interaction of Aspergillus flavus.

Oxylipins, a class of oxygenase-derived unsaturated fatty acids, are important signal molecules in many biological systems. Recent characterization of an Aspergillus flavus lipoxygenase gene, lox, revealed its importance in maintaining a density-dependent morphology switch from sclerotia to conidia as population density increased. Here, we present evidence for the involvement of four more oxylipin-generating dioxygenases (PpoA, PpoB, PpoC, and PpoD) in A. flavus density-dependent phenomena and the effects of loss of these genes on aflatoxin production and seed colonization. Although several single mutants showed alterations in the sclerotia-to-conidia switch, the major effect was observed in a strain downregulated for all five oxygenases (invert repeat transgene [IRT] strain IRT4 = ppoA, ppoB, ppoC, ppoD, and lox). In strain IRT4, sclerotia production was increased up to 500-fold whereas conidiation was decreased down to 100-fold and the strain was unable to switch into conidial production. Aflatoxin (AF) production for all mutant strains and the wild type was greatest at low population densities and absent in high populations except for strain IRT4, which consistently produced high levels of the mycotoxin. Growth on host seed by both IRT4 and IRT2 (downregulated in ppoA, ppoB, and ppoD) was marked by decreased conidial but increased AF production. We propose that A. flavus oxygenases and the oxylipins they produce act in a highly interdependent network with some redundancy of biological function. These studies provide substantial evidence for oxylipin-based mechanisms in governing fungus-seed interactions and in regulating a coordinated quorum-sensing mechanism in A. flavus.