Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) is a polyphagous pest indigenous throughout the Americas, which recently appeared in Africa, first reported from São Tomé, Nigeria, Bénin and Togo in 2016, and which we now report from Ghana. This species is recognised to comprise two morphologically identical but genetically distinct strains or species in the Americas, and we found both to be present in Ghana. We discuss possible routes of entry to Africa, of which the likeliest is adults and/or egg masses transported on direct commercial flights between the Americas and West Africa, followed by dispersal by adult flight within Africa. Identification of Lepidoptera is normally based on the markings and morphology of adults, and not on the larvae which actually cause the damage, and therefore larvae have to be reared through to adult for authoritative identification. We confirmed that the use of DNA barcoding allowed unequivocal identification of this new pest from Ghana based on the larvae alone. As authenticated barcodes for vouchered specimens of more pests become available, this approach has the potential to become a valuable in-country tool to support national capability in rapid and reliable pest diagnosis and identification.

Endophytic isolates of Trichoderma species are being considered as biocontrol agents for diseases of Theobroma cacao (cacao). Gene expression was studied during the interaction between cacao seedlings and four endophytic Trichoderma isolates, T. ovalisporum-DIS 70a, T. hamatum-DIS 219b, T. harzianum-DIS 219f, and Trichoderma sp.-DIS 172ai. Isolates DIS 70a, DIS 219b, and DIS 219f were mycoparasitic on the pathogen Moniliophthora roreri, and DIS 172ai produced metabolites that inhibited growth of M. roreri in culture. ESTs (116) responsive to endophytic colonization of cacao were identified using differential display and their expression analyzed using macroarrays. Nineteen cacao ESTs and 17 Trichoderma ESTs were chosen for real-time quantitative PCR analysis. Seven cacao ESTs were induced during colonization by the Trichoderma isolates. These included putative genes for ornithine decarboxylase (P1), GST-like proteins (P4), zinc finger protein (P13), wound-induced protein (P26), EF-calcium-binding protein (P29), carbohydrate oxidase (P59), and an unknown protein (U4). Two plant ESTs, extensin-like protein (P12) and major intrinsic protein (P31), were repressed due to colonization. The plant gene expression profile was dependent on the Trichoderma isolate colonizing the cacao seedling. The fungal ESTs induced in colonized cacao seedlings also varied with the Trichoderma isolate used. The most highly induced fungal ESTs were putative glucosyl hydrolase family 2 (F3), glucosyl hydrolase family 7 (F7), serine protease (F11), and alcohol oxidase (F19). The pattern of altered gene expression suggests a complex system of genetic cross talk occurs between the cacao tree and Trichoderma isolates during the establishment of the endophytic association.

BackgroundThe basidiomycete Moniliophthora roreri is the causal agent of Frosty pod rot (FPR) disease of cacao (Theobroma cacao), the source of chocolate, and FPR is one of the most destructive diseases of this important perennial crop in the Americas. This hemibiotroph infects only cacao pods and has an extended biotrophic phase lasting up to sixty days, culminating in plant necrosis and sporulation of the fungus without the formation of a basidiocarp.ResultsWe sequenced and assembled 52.3 Mb into 3,298 contigs that represent the M. roreri genome. Of the 17,920 predicted open reading frames (OFRs), 13,760 were validated by RNA-Seq. Using read count data from RNA sequencing of cacao pods at 30 and 60 days post infection, differential gene expression was estimated for the biotrophic and necrotrophic phases of this plant-pathogen interaction. The sequencing data were used to develop a genome based secretome for the infected pods. Of the 1,535 genes encoding putative secreted proteins, 1,355 were expressed in the biotrophic and necrotrophic phases. Analysis of the data revealed secretome gene expression that correlated with infection and intercellular growth in the biotrophic phase and invasive growth and plant cellular death in the necrotrophic phase.ConclusionsGenome sequencing and RNA-Seq was used to determine and validate the Moniliophthora roreri genome and secretome. High sequence identity between Moniliophthora roreri genes and Moniliophthora perniciosa genes supports the taxonomic relationship with Moniliophthora perniciosa and the relatedness of this fungus to other basidiomycetes. Analysis of RNA-Seq data from infected plant tissues revealed differentially expressed genes in the biotrophic and necrotrophic phases. The secreted protein genes that were upregulated in the biotrophic phase are primarily associated with breakdown of the intercellular matrix and modification of the fungal mycelia, possibly to mask the fungus from plant defenses. Based on the transcriptome data, the upregulated secreted proteins in the necrotrophic phase are hypothesized to be actively attacking the plant cell walls and plant cellular components resulting in necrosis. These genes are being used to develop a new understanding of how this disease interaction progresses and to identify potential targets to reduce the impact of this devastating disease.

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Endophytic fungi were isolated from healthy stems and pods of cacao ( Theobroma cacao ) trees in natural forest ecosystems and agroecosystems in Latin America and West Africa. These fungi were collected for screening as a potential source of biocontrol agents for the basidiomycetous pathogens of cacao in South and Central America, Moniliophthora roreri (frosty pod rot) and Moniliophthora perniciosa (witches' broom). Many of these isolates were morphologically unidentifiable as they failed to form fruiting structures in culture, or only produced arthrosporic stages. Affinities with basidiomycetes were suspected for many of these based on colony morphology. Fifty-nine of these morphologically unidentifiable isolates were selected for molecular identification by DNA extraction and sequence analysis of nuclear ribosomal DNA (rDNA). The large subunit (LSU) was chosen for initial sequencing because this region has been used most often for molecular systematics of basidiomycete fungi, and comprehensive LSU datasets were already available for sequence analyses. Results confirmed that the majority of the isolates tested belonged to the Basidiomycota, particularly to corticoid and polyporoid taxa. With LSU data alone, identification of the isolates was resolved at varying taxonomic levels (all to order, most to family, and many to genus). Some of the isolates came from rarely isolated genera, such as Byssomerulius , whilst the most commonly isolated basidiomycetous endophyte was a member of the cosmopolitan genus Coprinellus (Agaricales). The role of these fungi within the host and their potential as biological control agents are discussed.

In October 2001, a wilting disorder of new aetiology was reported affecting banana ( Musa sp.) within the Mukono district of Uganda. The disorder was characterized by a rapid yellowing and wilting of the younger leaves, a discoloration of the internal vascular vessels, occasionally a dieback initiating from the male floral parts with internal rotting of banana fruits. These symptoms were notably distinct from fusarium wilt ( Fusarium oxysporum ) and 'Matooke wilt' (a wilt-like disorder of unknown aetiology), but strongly resembled Moko disease caused by Ralstonia solanacearum , although this particular pathogen had not previously been recorded on banana in Africa.A bacterium was isolated on nutrient agar that was identified by fatty acid (Microbial ID Inc. [MIDI]) and metabolic (Biolog, Inc, Hayward, CA, USA) analyses as Xanthomonas axonopodis (ID probability score < 0·4) and Xanthomonas campestris (ID probability score ∼ 0·9), respectively. The presence of the Xanthomonas specific fatty acids 11:0 ISO, 11:0 ISO 3OH and 13:0 ISO 3OH was recorded. Pathogenicity tests on disease-free tissue culture-derived banana plantlets by stem inoculation with a bacterial suspension induced wilt symptoms consistent with field observations after 3 weeks. Reisolation and identification, as outlined above, confirmed Koch's postulates. Reference to the literature suggested the bacterium was Xanthomonas campestris pv. musacearum (Yirgou & Bradbury, 1968). However, this bacterium is relatively poorly described and not contained within either the MIDI or Biolog databases. To support the identification, rep-PCR (Louws et al ., 1994) using ERIC and BOX primers was performed on the Ugandan banana isolate, cultures of X. campestris pv. musacearum from Ensete and Musa in Ethiopia (IMI 349461, IMI 349986, IMI350025) and other cultures of Xanthomonas spp. from Africa. These analyses revealed an identical DNA fingerprint for all isolates from Musa and Ensete , but distinct fingerprints for the isolates from other hosts. This is the first report of X. campestris pv. musacearum outside of Ethiopia, where it is recorded as a pathogen of ensete and, to a lesser extent, banana. Accordingly, this pathogen has been given the common name of ensete bacterial wilt, although the aptness of this now looks questionable. The risk posed by this new disease record to the contiguous banana plantation of Uganda is undetermined, but significant spread is already being observed. The causative organism for this new disease record has been deposited within the CABI Genetic Resource Collection as IMI 386970. References

Comparative analyses were undertaken to characterize Xanthomonas campestris pv. musacearum, the causal agent of a wilt of enset and banana, and to assess its relatedness to other xanthomonads by fatty acid methyl esters, genomic fingerprinting using rep-PCR and partial nucleotide sequencing of the gyrase B gene. The results from all three analyses indicated that strains of X. campestris pv. musacearum are homogeneous and very similar to X. vasicola strains isolated from sugarcane and maize from Africa. Pathogenicity studies indicated that strains of X. vasicola pv. holcicola and X. vasicola from sugarcane induced no symptoms on banana, whereas X. campestris pv . musacearum produced severe disease. These data will support a future proposed reclassification of X. campestris pv. musacearum as X. vasicola pv . musacearum when more data are available.

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