Genetic variability and evolutionary implications of RNA silencing suppressor genes in RNA1 of sweet potato chlorotic stunt virus isolates infecting sweetpotato and related wild species.

BACKGROUND: The bipartite single-stranded RNA genome of Sweet potato chlorotic stunt virus (SPCSV, genus Crinivirus; Closteroviridae) encodes a Class 1 RNase III (RNase3), a putative hydrophobic protein (p7) and a 22-kDa protein (p22) from genes located in RNA1. RNase3 and p22 suppress RNA silencing, the basal antiviral defence mechanism in plants. RNase3 is sufficient to render sweetpotato (Ipomoea batatas) virus-susceptible and predisposes it to development of severe diseases following infection with unrelated virus. The incidence, strains and gene content of SPCSV infecting wild plant species have not been studied. METHODOLOGY/PRINCIPAL FINDINGS: Thirty SPCSV isolates were characterized from 10 wild Ipomoea species, Hewittia sublobata or Lepistemon owariensis (family Convolvulaceae) in Uganda and compared with 34 local SPCSV isolates infecting sweetpotatoes. All isolates belonged to the East African (EA) strain of SPCSV and contained RNase3 and p7, but p22 was not detected in six isolates. The three genes showed only limited genetic variability and the proteins were under purifying selection. SPCSV isolates lacking p22 synergized with Sweet potato feathery mottle virus (SPFMV, genus potyvirus; Potyviridae) and caused severe symptoms in co-infected sweetpotato plants. One SPCSV isolate enhanced accumulation of SPFMV, but no severe symptoms developed. A new whitefly-transmitted virus (KML33b) encoding an RNase3 homolog (<56% identity to SPCSV RNase3) able to suppresses sense-mediated RNA silencing was detected in I. sinensis. CONCLUSIONS/SIGNIFICANCE: SPCSV isolates infecting wild species and sweetpotato in Uganda were genetically undifferentiated, suggesting inter-species transmission of SPCSV. Most isolates in Uganda contained p22, unlike SPCSV isolates characterized from other countries and continents. Enhanced accumulation of SPFMV and increased disease severity were found to be uncoupled phenotypic outcomes of RNase3-mediated viral synergism in sweetpotato. A second virus encoding an RNase3-like RNA silencing suppressor was detected. Overall, results provided many novel and important insights into evolutionary biology of SPCSV.

Sugar-mediated semidian oscillation of gene expression in the cassava storage root regulates starch synthesis.

Starch branching enzyme (SBE) activity in the cassava storage root exhibited a diurnal fluctuation, dictated by a transcriptional oscillation of the corresponding SBE genes. The peak of SBE activity coincided with the onset of sucrose accumulation in the storage, and we conclude that the oscillatory mechanism keeps the starch synthetic apparatus in the storage root sink in tune with the flux of sucrose from the photosynthetic source. When storage roots were uncoupled from the source, SBE expression could be effectively induced by exogenous sucrose. Turanose, a sucrose isomer that cannot be metabolized by plants, mimicked the effect of sucrose, demonstrating that downstream metabolism of sucrose was not necessary for signal transmission. Also glucose and glucose-1-P induced SBE expression. Interestingly, induction by sucrose, turanose and glucose but not glucose-1-P sustained an overt semidian (12-h) oscillation in SBE expression and was sensitive to the hexokinase (HXK) inhibitor glucosamine. These results suggest a pivotal regulatory role for HXK during starch synthesis. Abscisic acid (ABA) was another potent inducer of SBE expression. Induction by ABA was similar to that of glucose-1-P in that it bypassed the semidian oscillator. Both the sugar and ABA signaling cascades were disrupted by okadaic acid, a protein phosphatase inhibitor. Based on these findings, we propose a model for sugar signaling in regulation of starch synthesis in the cassava storage root.

Unravelling the genetic diversity of the three main viruses involved in Sweet Potato Virus Disease (SPVD), and its practical implications.

SUMMARY Sweetpotato (Ipomoea batatas) is a widely grown food crop, in which the most important diseases are caused by viruses. Genetic variability of three widely distributed sweetpotato viruses was analysed using data from 46 isolates of Sweet potato feathery mottle virus (SPFMV), 16 isolates of Sweet potato mild mottle virus (SPMMV) and 25 isolates of Sweet potato chlorotic stunt virus (SPCSV), of which 19, seven and six isolates, respectively, are newly characterized. Division of SPFMV into four genetic groups (strains) according to phylogenetic analysis of coat protein (CP) encoding sequences revealed that strain EA contained the East African isolates of SPFMV but none from elsewhere. In contrast, strain RC contained ten isolates from Australia, Africa, Asia and North America. Strain O contained six heterogeneous isolates from Africa, Asia and South America. The seven strain C isolates from Australia, Africa, Asia, and North and South America formed a group that was genetically distant from the other SPFMV strains. SPMMV isolates showed a high level of variability with no discrete strain groupings. SPCSV isolates from East Africa were phylogenetically distant to SPCSV isolates from elsewhere. Only from East Africa were adequate data available for different isolates of the three viruses to estimate the genetic variability of their local populations. The implications of the current sequence information and the need for more such information from most sweetpotato-growing regions of the world are discussed in relation to virus diagnostics and breeding for virus resistance.

Incidence of Viruses and Virus like Diseases of Sweetpotato in Uganda.

Sweetpotato plants were surveyed for viruslike diseases and viruses in the four major agroecological zones of Uganda. Testing of 1,260 sweetpotato plants, of which 634 had virus-like symptoms, showed that virus disease incidence ranged from 2.7% (Soroti district, short grassland-savannah zone) to 20% (Mukono district, tall grass-forest mosaic zone). Sweet potato chlorotic stunt virus (SPCSV), Sweet potato feathery mottle virus (SPFMV), Sweet potato mild mottle virus (SPMMV), and sweet potato chlorotic fleck virus (SPCFV) were serologically detected and positive results confirmed by immunocapture reverse transcriptase polymerase chain reaction (IC-RT-PCR) and subsequent sequence analyses of the amplified fragments, except SPCFV, which lacked sequence information. SPCSV and SPFMV were detected in all the 14 districts surveyed, whereas SPMMV and SPCFV were detected in 13 and 8 districts, respectively. Logistic regression analysis revealed that SPCSV and SPFMV, SPFMV and SPMMV, and SPFMV and SPCFV more frequently occurred together than any other virus combinations or as single virus infections. Co-infections of SPCSV with SPFMV and/or SPMMV were associated with more severe and persistent symptoms than infections with each of the viruses alone. Several plants (11%) displaying viruslike symptoms did not react with the virus antisera used, suggesting that more viruses or viruslike agents are infecting sweetpotatoes in Uganda.