In September 2011, a high incidence of a new maize (Zea mays L.) disease was reported at lower elevations (1,900 m asl) in the Longisa division of Bomet County, Southern Rift Valley, Kenya. The disease later spread to the Narok South and North and Naivasha Districts. By March 2012, the disease was reported at up to 2,100 m asl. Diseased plants had symptoms characteristic of virus diseases: a chlorotic mottle on leaves, developing from the base of young whorl leaves upward to the leaf tips; mild to severe leaf mottling; and necrosis developing from leaf margins to the mid-rib. Necrosis of young leaves led to a “dead heart” symptom, and plant death. Severely affected plants had small cobs with little or no grain set. Plants frequently died before tasseling. All maize varieties grown in the affected areas had similar symptoms. In these regions, maize is grown continuously throughout the year, with the main planting season starting in November. Maize streak virus was present, but incidence was low (data not shown). Infected plants were distributed throughout affected fields, with heavier infection along field edges. High thrips (Frankliniella williamsi Hood) populations were present in sampled fields, but populations of other potential disease vectors, such as aphids and leafhoppers, were low. Because of the high thrips populations and foliar symptoms, symptomatic plants were tested for the presence of Maize chlorotic mottle virus (MCMV) (3) using tissue blot immunoassay (TBIA) (1). Of 17 symptomatic leaf samples from each Bomet and Naivasha, nine from Bomet and all 17 from Naivasha were positive for MCMV. However, the observed symptoms were more severe than commonly associated with MCMV, suggesting the presence of maize lethal necrosis (MLN), a disease that results from maize infection with both MCMV and a potyvirus (4). Therefore, samples were tested for the presence of Sugarcane mosaic virus (SCMV), which is present in Kenya (2). Twenty-seven samples were positive for SCMV by TBIA, and 23 of 34 samples were infected with both viruses. Virus identities were verified with reverse-transcription (RT)-PCR (Access RT-PCR, Promega) and MCMV or SCMV-specific primers. MCMV primers (2681F: 5′-ATGAGAGCAGTTGGGGAATGCG and 3226R: 5′-CGAATCTACACACACACACTCCAGC) amplified the expected 550-bp product from three leaf samples. Amplicon sequences were identical, and had 95 to 98% identity with MCMV sequences in GenBank. SCMV primers (8679F: 5′-GCAATGTCGAAGAAAATGCG) and 9595R: 5′-GTCTCTCACCAAGAGACTCGCAGC) amplified the expected 900-bp product from four leaf samples. Amplicon sequences had 96 to 98% identity, and were 88 to 96% identical with SCMV sequences in GenBank. To our knowledge, this is the first report of MCMV and of maize coinfection with MCMV and SCMV associated with MLN in Kenya and Africa. MLN is a serious threat to farmers in the affected areas, who are experiencing extensive to complete crop loss. References: (1) P. G. S. Chang et al. J. Virol. Meth. 171:345, 2011. (2) Delgadillo Sanchez et al. Rev. Mex. Fitopat. 5:21, 1987. (3) Jiang et al., Crop Prot. 11:248, 1992. (4) R. Louie, Plant Dis. 64:944, 1980.

The diagnosis of novel unidentified viral plant diseases can be problematic, as the conventional methods such as real‐time PCR or ELISA may be too specific to a particular species or even strain of a virus, whilst alternatives such as electron microscopy (EM) or sap inoculation of indicator species do not usually give species level diagnosis. Next‐generation sequencing (NGS) offers an alternative solution where sequence is generated in a non‐specific fashion and identification is based on similarity searching against GenBank. The conventional and NGS techniques were applied to a damaging and apparently new disease of maize, which was first identified in Kenya in 2011. ELISA and TEM provided negative results, whilst inoculation of other cereal species identified the presence of an unidentified sap transmissible virus. RNA was purified from material showing symptoms and sequenced using a Roche 454 GS‐FLX+. Database searching of the resulting sequence identified the presence of Maize chlorotic mottle virus and Sugarcane mosaic virus, a combination previously reported to cause maize lethal necrosis disease. Over 90% of both viral genome sequences were obtained, allowing strain characterization and the development of specific real‐time PCR assays which were used to confirm the presence of the virus in material with symptoms from six different fields in two different regions of Kenya. The availability of these assays should aid the assessment of the disease and may be used for routine diagnosis. The work shows that next‐generation sequencing is a valuable investigational technique for rapidly identifying potential disease‐causing agents such as viruses.

The fall armyworm (FAW), Spodoptera frugiperda, is a destructive pest native to America and has recently become an invasive insect pest in China. Because of its rapid spread and great risks in China, understanding of FAW genetic background and pesticide resistance is urgent and essential to develop effective management strategies. Here, we assembled a chromosome-level genome of a male FAW (SFynMstLFR) and compared re-sequencing results of the populations from America, Africa, and China. Strain identification of 163 individuals collected from America, Africa and China showed that both C and R strains were found in the American populations, while only C strain was found in the Chinese and African populations. Moreover, population genomics analysis showed that populations from Africa and China have close relationship with significantly genetic differentiation from American populations. Taken together, FAWs invaded into China were most likely originated from Africa. Comparative genomics analysis displayed that the cytochrome p450 gene family is extremely expanded to 425 members in FAW, of which 283 genes are specific to FAW. Treatments of Chinese populations with twenty-three pesticides showed the variant patterns of transcriptome profiles, and several detoxification genes such as AOX, UGT and GST specially responded to the pesticides. These findings will be useful in developing effective strategies for management of FAW in China and other invaded areas.

Cassava brown streak disease (CBSD) caused by Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) is causing severe losses in cassava production in Kenya, Tanzania and Uganda. Two real-time RT-PCR assays based on TaqMan chemistry capable of detecting and distinguishing these two viruses are described. These assays were used to screen 493 cassava samples collected from western and coastal Kenya, the main cassava regions of Uganda and inland Tanzania. Both viruses were found in all three countries and across regions therein. Association of CBSD leaf symptom status with CBSV and UCBSV assay results was weak, confirming the need for a diagnostic assay. For leaf samples that were observed with CBSD-like leaf symptoms but shown as CBSV and UCBSV negative by the RT-PCR assay, deep sequencing using a Roche 454 GS-FLX was used to provide additional evidence for the absence of the viruses. The probability of the CBSD associated diagnostics detecting a single CBSV or UCBSV positive sample amongst other non-CBSD samples was modelled. The results of this study are discussed in the context of the application of diagnostics of CBSD-associated viruses under the Great Lakes Cassava Initiative and the need to minimize the risk of further spread of the viruses with cassava multiplication material. It is shown that high throughput testing undertaken at Fera of 300 cassava leaves taken from fields for seed multiplication, when analysed in pools of 10, has given a 95% probability of detecting 1% infected plants in the field.

The complete genome sequence for an isolate of the Ugandan and Tanzanian strain types of Cassava brown streak virus have been determined using the novel approach of non-directed next generation sequencing. Comparison of the genome sequences revealed that CBSV is highly heterogeneous at the isolate level as well as the strain level. The isolate of the Ugandan strain was found to have a genome 9,070 nucleotides long coding for a polypeptide with 2,902 amino acid residues. The isolate of the Tanzanian strain was 9,008 nucleotides long and coded for a polypeptide with 2,916 amino acid residues. Nucleotide identity between the isolates across the genome was 76%, with protein encoding regions 57-77% and individual proteins had 65-91% amino acid similarity. In addition between the two strains four protein products (PIPO, CI, NIa-Vpg and coat protein) varied in size and an unusual HAM1-like protein, whilst of identical nucleotide length, was found to have the lowest homology. The implication of diversity of CBSV is discussed in the context of speciation, evolution, development of diagnostics, and breeding for resistance.