Linear Lepidopteran ambidensovirus 1 sequences drive random integration of a reporter gene in transfected Spodoptera frugiperda cells

Background The Lepidopteran ambidensovirus 1 isolated from Junonia coenia (hereafter JcDV) is an invertebrate parvovirus considered as a viral transduction vector as well as a potential tool for the biological control of insect pests. Previous works showed that JcDV-based circular plasmids experimentally integrate into insect cells genomic DNA. Methods In order to approach the natural conditions of infection and possible integration, we generated linear JcDV-gfp based molecules which were transfected into non permissive Spodoptera frugiperda (Sf9) cultured cells. Cells were monitored for the expression of green fluorescent protein (GFP) and DNA was analyzed for integration of transduced viral sequences. Non-structural protein modulation of the VP-gene cassette promoter activity was additionally assayed. Results We show that linear JcDV-derived molecules are capable of long term genomic integration and sustained transgene expression in Sf9 cells. As expected, only the deletion of both inverted terminal repeats (ITR) or the polyadenylation signals of NS and VP genes dramatically impairs the global transduction/expression efficiency. However, all the integrated viral sequences we characterized appear “scrambled” whatever the viral content of the transfected vector. Despite a strong GFP expression, we were unable to recover any full sequence of the original constructs and found rearranged viral and non-viral sequences as well. Cellular flanking sequences were identified as non-coding ones. On the other hand, the kinetics of GFP expression over time led us to investigate the apparent down-regulation by non-structural proteins of the VP-gene cassette promoter. Conclusion Altogether, our results show that JcDV-derived sequences included in linear DNA molecules are able to drive efficiently the integration and expression of a foreign gene into the genome of insect cells, whatever their composition, provided that at least one ITR is present. However, the transfected sequences were extensively rearranged with cellular DNA during or after random integration in the host cell genome. Lastly, the non-structural proteins seem to participate in the regulation of p9 promoter activity rather than to the integration of viral sequences

Positive selection alone is sufficient for whole genome differentiation at the early stage of speciation process in the fall armyworm

International audienceBackground: The process of speciation involves differentiation of whole genome sequences between a pair of diverging taxa. In the absence of a geographic barrier and in the presence of gene flow, genomic differentiation may occur when the homogenizing effect of recombination is overcome across the whole genome. The fall armyworm is observed as two sympatric strains with different host-plant preferences across the entire habitat. These two strains exhibit a very low level of genetic differentiation across the whole genome, suggesting that genomic differentiation occurred at an early stage of speciation. In this study, we aim at identifying critical evolutionary forces responsible for genomic differentiation in the fall armyworm. Results: These two strains exhibit a low level of genomic differentiation (F ST = 0.0174), while 99.2% of 200 kb windows have genetically differentiated sequences (F ST > 0). We found that the combined effect of mild positive selection and genetic linkage to selectively targeted loci are responsible for the genomic differentiation. However, a single event of very strong positive selection appears not to be responsible for genomic differentiation. The contribution of chromosomal inversions or tight genetic linkage among positively selected loci causing reproductive barriers is not supported by our data. Phylogenetic analysis shows that the genomic differentiation occurred by sub-setting of genetic variants in one strain from the other. Conclusions: From these results, we concluded that genomic differentiation may occur at the early stage of a speciation process in the fall armyworm and that mild positive selection targeting many loci alone is sufficient evolutionary force for generating the pattern of genomic differentiation. This genomic differentiation may provide a condition for accelerated genomic differentiation by synergistic effects among linkage disequilibrium generated by following events of positive selection. Our study highlights genomic differentiation as a key evolutionary factor connecting positive selection to divergent selection

Centromeres, transposable elements, piRNAs and heterochromatin in Spodoptera frugiperda

International audienceHolocentric chromosomes have been reported in at least 768 species both in animal and plant kingdoms and appeared 12 times during evolution. The spread of the centromere on the whole length of chromosome arms has several consequences on functional organization of the genome in term of evolution rate, distribution of transposable elements (TE) and of chromatin domains, the need for adaptations during meiosis. From the analysis of the nematode C. elegans, the best characterized holocentric model, kinetochore protein composition seemed conserved between monocentrics and holocentrics. However, recent studies highlighted the absence of the ubiquitous histone H3 like CENP-A in several orders of holocentric insects including Lepidoptera as shown in the silkworm, showing diversity in kinetochore composition of holocentric species.In this talk, I will present our attempts to understand the functional organization of a major pest of agriculture, Spodoptera frugiperda through i) characterization of centromeric protein homologs by RNA interference in a cell line ii) annotation of TE elements in various genome assemblies as well as piRNAs, their non-coding RNA regulators iii) analysis of heterochromatin domains in vitro and in vivo

Host-plant adaptation as a driver of incipient speciation in the fall armyworm ( Spodoptera frugiperda )

Background Divergent selection on host-plants is one of the main evolutionary forces driving ecological speciation in phytophagous insects. The ecological speciation might be challenging in the presence of gene flow and assortative mating because the direction of divergence is not necessarily the same between ecological selection (through host-plant adaptation) and assortative mating. The fall armyworm (FAW), a major lepidopteran pest species, is composed of two sympatric strains, corn and rice strains, named after two of their preferred host-plants. These two strains have been hypothesized to undergo incipient speciation, based on ( i ) several lines of evidence encompassing both pre- and post-zygotic reproductive isolation, and ( ii ) the presence of a substantial level of genetic differentiation. Even though the status of these two strains has been established a long time ago, it is still yet to be found whether these two strains indeed exhibit a marked level of genetic differentiation from a large number of genomic loci. Here, we analyzed whole genome sequences from 56 FAW individuals either collected from pasture grasses (a part of the favored host range of the rice strain) or corn to assess the role of host-plant adaptation in incipient speciation. Results Principal component analysis of whole genome data shows that the pattern of divergence in the fall armyworm is predominantly explained by the genetic differentiation associated with host-plants. The level of genetic differentiation between corn and rice strains is particularly marked in the Z chromosome. We identified one autosomal locus and two Z chromosome loci targeted by selective sweeps specific to rice strain and corn strain, respectively. The autosomal locus has both increased D XY and F ST while the Z chromosome loci had decreased D XY and increased F ST . Conclusion These results show that the FAW population structure is dominated by the genetic differentiation between corn and rice strains. This differentiation involves divergent selection targeting at least three loci, which include a locus potentially causing reproductive isolation. Taken together, these results suggest the evolutionary scenario that host-plant speciation is a driver of incipient speciation in the fall armyworm

COI-based phylogenetic relationships of <i>Spodoptera</i> species.

<p>The tree on the left summarizes the results of ML analyses while the tree on the right summarizes the results of BI analyses. Black dots indicate nodes supported by bootstrap values ≥ 70% (ML tree) or supported by posterior probability ≥ 0.95 (BI tree).</p

List of evidences arguing in favor of a high level of differentiation between the two FAW strains.

<p>As indicated by the combination of pros and cons some results are more or less contradictory.</p><p>List of evidences arguing in favor of a high level of differentiation between the two FAW strains.</p

Putative species clusters corresponding to: (i) morphological delineations (column 1); (ii) PTP analyses using either the majority-rule consensus topology resulting from the BI analyses (column 2) or the best tree from the ML analyses (column 3); (iii) GMYC analyses based either on a strict clock and a BD model (column 4) or on a UCLN clock and a BD model (column 5).

<p>The reference tree used here corresponds to the tree resulting from the BI analyses carried out with MrBayes.</p

Complex multiple introductions drive fall armyworm invasions into Asia and Australia

Abstract The fall armyworm (FAW) Spodoptera frugiperda is thought to have undergone a rapid ‘west-to-east’ spread since 2016 when it was first identified in western Africa. Between 2018 and 2020, it was recorded from South Asia (SA), Southeast Asia (SEA), East Asia (EA), and Pacific/Australia (PA). Population genomic analyses enabled the understanding of pathways, population sources, and gene flow in this notorious agricultural pest species. Using neutral single nucleotide polymorphic (SNP) DNA markers, we detected genome introgression that suggested most populations in this study were overwhelmingly C- and R-strain hybrids (n = 252/262). SNP and mitochondrial DNA markers identified multiple introductions that were most parsimoniously explained by anthropogenic-assisted spread, i.e., associated with international trade of live/fresh plants and plant products, and involved ‘bridgehead populations’ in countries to enable successful pest establishment in neighbouring countries. Distinct population genomic signatures between Myanmar and China do not support the ‘African origin spread’ nor the ‘Myanmar source population to China’ hypotheses. Significant genetic differentiation between populations from different Australian states supported multiple pathways involving distinct SEA populations. Our study identified Asia as a biosecurity hotspot and a FAW genetic melting pot, and demonstrated the use of genome analysis to disentangle preventable human-assisted pest introductions from unpreventable natural pest spread