Comparative epidemiology of three maize viruses in Reunion Island in relation to the population dynamic of their vectors Cicadulina mbila and Peregrinus maidis

The simultaneous presence of maize streak virus (MSV), maize stripe virus (MStpV) and maize mosaic virus (MMV) in Réunion bas enabled us to study their comparative epidemiology. . Symptomatological ratings were taken through three years of culture on week1y sowings of the temperate hybrid INRA 508 and the composite variety IRAT 297. Fluctuations of emigrant populations of #C. mbila# and #P. mbila# and climatic faclors (temperature, rainfall and relative humidity) were measured and analysed by time series and stepwise regression analyses. Streak is the dominant disease, particularly during the warm rainy season and MMV was less frequent. Highest autocorrelations were observed with a time-lag of 12 months, confirming the annual periodicity of the fluctuation. Pattern of change of insect numbers was positively correlated with the change in disease incidence (correlations ranging from 0.65 to 0.87). Disease incidence and vector numbers always remained constant or increased slowly with temperature up 10 24°C and increased rapidly above 24°C. The relationship between rainfall, relative humidity, disease incidence and vector numbers is less clear. Depending on the series, from 63% to SO% of the variance of disease incidence was explained by the stepwise regression with vector numbers, and (sometimes) temperature, rainfall or relative humidity. Therefore. the close adjustement found between observed and data calculated with the resulting regression allowed us to propose a simple epidemiological model. Epidemiological data especially from Africa are compared in order to better understand the epidemiology of these viruses. Further work is needed to confirm the validity of the mode. The rate of plants infected by the three viruses and the number of# P. maidis# per plant were significantly lower (P = 0.0001) on IRAT 297 than on INRA 508. The resistance behavior of IRAT 297 under natural conditions points to its utilization as resistance donor in breeding program geared toward obtaining multi-resistance maize genotypes. (Texte intégral

Plant resistance-driven emergence of recombinant begomoviruses

The analysis of plant virus genomes reveals that many were shaped by recombination. However, the history of the emergence dynamics of these recombinants is mostly unknown as well as the underlying evolutionary forces that drove their frequency increase. The pivotal role of recombination in geminivirus evolution is supported by the detection of numerous recombination events in sequence data, and by their high propensity to recombine. These typical features were observed with Tomato yellow leaf curl virus (TYLCV), a tomato begomovirus that was extensively studied because of its global economic importance. TYLCV-IS76 is a recombinant TYLCV detected initially in Morocco. It inherited a 76-nt region of tomato yellow leaf curl Sardinia virus (TYLCSV) starting from the origin of replication (OR) towards the V2 gene. Based on field surveys carried out in Morocco and laboratory analysis, a real time emergence of TYLCV-IS76 has been reconstructed from its generation to the displacement of its parental viruses (1). Its emergence coincided with the deployment of Ty-1 resistant tomato cultivars, and a causal link was demonstrated with various competition tests in which positive selection of TYLCV-IS76 was observed in Ty-1 resistant plants (2). TYLCV-IS141 is a TYLCV recombinant detected in Italy (1, 3, 4, 5). It inherited a 141-nt region of TYLCSV between OR and the initiation codon of the V2 gene. TYLCV-IS141 and TYLCV-IS76 exhibit similar recombination profiles and fitness phenotypes in Ty-1 resistant plants. It was inferred from competition tests carried out with various natural and artificially generated TYLCV-IS76 and TYLCV-IS141 clones, that the fitness phenotype of these recombinants was determined by new beneficial intra genomic interaction rather than by a direct effect of specific mutations. Gene silencing is suspected to be involved in the positive selection of these recombinants because Ty-1 is a RNA dependent RNA polymerase gene

The viral content ratio between abdomen and head is informative of the relative efficiency with which Bemisia tabaci populations transmit begomoviruses. [P.12]

Begomoviruses (family Geminiviridae) are transmitted by the whitefly Bemisia tabaci in a circulative non propagative manner. B. tabaci is a species complex composed of at least 24 morphocryptic species which differ in host range, insecticide resistance, endosymbionts and virus transmission. Begomoviruses are supposed to cross the gut barrier at the midgut level and salivary gland barrier in the principal salivary gland (PSG) cells because of the highest virus concentrations in these organs. Thus, the critical steps of the virus circulation in the insect body are (i) the exit of virion from the midgut, (ii) their preservation in the hemocoel and (iii) their entry in the PSG. Thus, we proposed that the efficiency of viral transfer from midgut to PSG may be assessed by measuring the viral content in both compartments and that the deduced viral content ratio may be correlated to viral transmission efficiency by the vector. Our predicition was tested with two invasive B. tabaci species, Middle East-Minor Asia 1 (MEAM1), and Mediterranean (MED), and three begomoviruses: the invasive species Tomato yellow leaf curl virus-Mild (TYLCV-Mld), Tomato leaf curl Comoros virus (ToLCKMV), indigenous from Mayotte and R4, a recombinant between TYLCV and ToLCKMV. In a first approach, PSG and midgut were separated by a cross section through the prothorax and viral loads were estimated in both sections by measuring viral DNA using real time PCR. As the midgut of B. tabaci was reported to be sometimes pushed through the diaphragm separating the abdomen and the thorax, the estimation of the viral content ratio between PSG and midgut may be biased by thorax sectioning. The simple cross sectioning was however validated because the ratio determined with such sections and the ratio determined after a careful gut dissection was similar. Using the simple cross section, the viral content ratio between head and abdomen was higher for MEAM1 than for Med for the three begomoviruses. As predicted, the transmission efficiency was higher with MEAM1 than Med Q2 for the three viruses. These results indicate that viral content ratio may be a reliable predictor of the relative transmission efficiency between different B. tabaci populations. Measuring transmission efficiency is time consuming, involves technically difficult experiments with acquisition and inoculation steps and needs specialized cage and containment equipment. However measuring viral content ratios needs only a few cages for the acquisition step, a binocular lens and an access to the commonly used qPCR machines. This approach might be extended to estimate the relative transmission efficiency of other circulative non propagative viruses. (Résumé d'auteur

Multiple genetic and ecological outcomes following contacts between invading and indigenous populations of Bemisia tabaci

The whitefly Bemisia tabaci is a pest of vegetable, ornamental and field crops. It is also the vector of several economically important plant viruses as Tomato yellow leaf curl virus. The comparison of cytochrome oxydase 1 sequences of a global collection of B. tabaci has revealed 12 well resolved genetic groups with a strong geographic structure. Based on the considerable genetic diversity together with distinct biological parameters and particularly varying abilities to interbreed, it was proposed that B. tabaci represent a cryptic species complex. Due to human activities, a genetic group, believed to be originating from the sahel-like regions of Middle Eastern Mediterranean/ North Africa/ Asia Minor, was very largely dispersed outside its geographic origin. This group, mainly consisting of individuals inducing physiological disorders on infested plants, is commonly referred to as the biotype B. As B. tabaci is a worldwide pest, the dispersed biotype B was often introduced into agroecosystems containing indigenous populations of B. tabaci. The objective of this study was to analyze the genetic and ecological outcomes of such contacts between the invading biotype B and indigenous populations in different regions of the world. In the Caribbean islands we have observed the almost complete displacement of the indigenous populations. In Reunion Island, we have detected the introgression of genetic material from the indigenous populations into biotype B. In Morocco, we have observed the coexistence of the indigenous populations and biotype B without any clear evidence of gene flow. (Texte intégral

Is the assistance of satellite by TYLCV strictly cell autonomous?

Begomoviruses are circular single stranded DNA (css) plant viruses with bipartite (A and B) or monopartite (A-like component) genomes. They are sometimes associated with satellites, cssDNA molecules, namely alphasatellites and betasatellites. Like the B component of bipartite begomoviruses, satellites depend on the A or A-like component for their replication (betasatellite) and encapsidation (alphasatellites and betasatellites). Although Tomato yellow leaf curl virus (TYLCV) was only rarely reported with satellites, alphasatellites and betasatellites of various geographic origins are readily assisted by TYLCV in experimental conditions. This result was consistent with the observation that satellite DNA contents were mostly higher than that of TYLCV (Conflon et al., 2018). The ease with which satellites can be assisted with TYLCV was supported further by FISH observations, which showed that the frequency of TYLCV-infected cells that were co-infected with a satellite exceeded 85% for an alphasatellite, and 95% for a betasatellite. Interestingly, a substantial number of cell nuclei were positive only for the satellite, suggesting that the assistance seems to be possible, even with a low amount of TYLCV DNA, and possibly no TYLCV DNA. This later possibility that need to be confirmed with further tests, is according to the “multicellular way of life” theory proposed recently for Faba bean necrotic stunt virus, a multipartite nanovirus with eight separately encapsidated components (Sicard et al.,2019)

Molecular evidence for the association of a strain of Uganda variant of East African cassava mosaic virus to symptom severity in cassava (Manihot esculenta Crantz) fields in Togo

Abstract: Problem statement: This study was carried out to demonstrate that the severity of Cassava Mosaic Disease (CMD) in Togo, is not only influenced by synergism between cassava Begomoviruses in presence, but essentially by recombination between the different Begomoviruses infecting cassava. Approach: Foliar samples presenting typical biological features of Begomoviruses infection were collected from cassava and wild infected plants from different regions of Togo and analysed by PCR targeting the Coat Protein (CP). The PCR products obtained from different isolates of two major Begomoviruses species infecting cassava in Togo were then sequenced and compared with the sequenced of the African cassava mosaic Begomoviruses identified to date and available in NCBI GenBank database by phylogenetic analysis. Results: The results indicate that not only the two major Begomoviruses could be in synergistic interaction in infected cassava in Togo as it has been shown between African Cassava Mosaic Virus (ACMV) and East African Cassava Mosaic Virus (EACMV) elsewhere, but could also create recombinants which would be highly interfering in the development of symptom severity in the country. Conclusion/Recommendations: The study confirmed the assumption that the symptom severity in cassava fields in Togo is rather caused by recombination between different Begomoviruses in presence than by synergistic interaction. More investigations should be done to give insight to this founding. (Résumé d'auteur

Assessing the possible maintenance of TYLCV-satellite association. [O.21]

Viruses of the genus Begomovirus (Family Geminiviridae) are frequently detected with half genome size DNA molecules, either defective DNAs or satellite DNA (? or ?). Whereas some begomoviruses, like Tomato yellow leaf curl virus (TYLCV) were never detected with satellite DNAs, other begomoviruses, like Cotton leaf curl virus (CLCuV), depend upon a betasatellite for their infectivity. Besides the CLCuV-type begomoviruses which may be considered as bipartite begomoviruses, most of the begomoviruses detected with satellites were shown to be infective without their satellites. The alphasatellite was rarely proved to have any impact on the helper virus but the betasatellite was often shown to increase the virulence of its helper virus. Although satellites were never detected with TYLCV in natural conditions, TYLCV was reported as a helper virus for both satellites in artificial conditions and its virulence was dramatically increased when co-inoculated with betasatellites. We have confirmed these results with the Cotton leaf curl Gezira betasatellite (CLCuGB) and two alphasatellites, Cotton leaf curl Gezira alphasatellite (CLCuGA) and Okra leaf curl Burkina Faso alphasatellite (OLCA). If the co-infection of TYLCV and a betasatellite would occur in natural conditions, tomato production may be severely affected. As the probability of such a scenario mainly depends on the maintenance of TYLCV-satellite associations over time, we have studied various factors potentially determining this maintenance: (i) the relative intra-plant accumulation of TYLCV and the satellites, (ii) the cellular co-infection level of TYLCV and satellites, and (iv) the transmission efficiency of satellites by the vector Bemisia tabaci. These various factors were analyzed with CLCuGB, CLCuGA and OLCA. Besides the specific question of the possible maintenance of satellites with TYLCV, the results of our study are expected to provide a new insight on begomoviruses detected in co-infection with satellites in natural conditions, but which were proved to be infectious without satellites. (Résumé d'auteur

Genetic diversity and identification of symbiotic community of Q Bemisia tabaci from Mediterranean countries

Introduction: We investigated a) the genetic diversity and population structure, b) the secondary symbiotic bacterial community and c) the frequencies of target site mutations conferring resistance to pyrethroids and organophospates in Q Bemisia tabaci collected from 5 Mediterranean countries. Material and Methods: Polymorphism at seven microsatellite loci and a 676-bp fragment of the cytochrome oxidase I (COI) gene were investigated for the genetic analysis of B. tabaci populations. Simple PCR based detection assays were used for the discrimination of B. tabaci subgroups and the detection of mutations associated with target site insecticide resistance (iAChE, kdr). Secondary symbionts were detected using specific primers. Wolbachia strains were characterized by Multilocus Sequence Typing (MLST) analysis. Results: Analyses of the 676-bp COI fragment distinguished seventeen haplotypes which delineated two groups (Q1 and Q2), within the Q B. tabaci. Both Q1 and Q2 were found in Spain and France, whereas only Q1 was observed in Greece, Morocco and Tunisia. The analyses of the microsatellite loci polymorphism revealed a high level of genetic differentiation even between some neighbouring samples belonging to the same sub-group within a country. In B. tabaci from Greece, Bayesian analysis revealed two main genetic groups within Q1, the first with populations from South Crete, and the second with populations from continental Greece and North Crete. Genetic differentiation was not correlated with host plant species or habitat. The secondary symbionts Wolbachia and Hamiltonella were present at high frequency while Arsenophonus, Cardinium and Rickettsia were absent. MLST analysis identified two Wolbachia strains which were found together in most of the Greek populations but never in the same host individual. The frequency of pyrethroid resistance mutations L925I and T929V in the para sodium channel gene and the organophosphate resistant mutation F331W in the ace1 gene was determined. The higher frequencies of the resistance mutations were found in France, Spain, and Greece, where in some samples the resistant alleles were fixed. In Morocco and Tunisia, the resistant alleles were less frequent and in some localities even undetected. Conclusion: Our findings suggest that the Mediterranean Q B. tabaci is more diverse and structured than reported so far and suggest that human activities play a major role in the genetic structure and as well as in the dynamics of resistance genes. The role of the symbionts, and in particular of Wolbachia W1 and W2 strains in the shaping of the population genetic structure, is currently under investigation. (Résumé d'auteur

Mise au point de deux tests PCR multiplexes pour détecter simultanément des formes recombinantes et non-recombinantes du Tomato yellow leaf curl virus au Maroc

Le Tomato yellow leaf curl virus (TYLCV, genre Begomvirus, famille Geminiviridae) a été isolé pour la première fois au Maroc en 1998 (Peterschmitt et al., 1999) et le Tomato yellow leaf curl Sardinia virus (TYLCSV) en 1999 (Monci et al., 2000). La gravité des attaques de la tomate par ces virus a obligé les agriculteurs à cultiver la tomate sous serre insectproof et à progressivement remplacer les variétés traditionnelles par des variétés tolérantes. Le TYLCV et le TYLCSV étant des virus hautement recombinogènes (Garcia Andres et al., 2007), le risque d'une émergence de recombinants TYLCV/TYLCSV dans une zone infestée par les deux virus était considéré comme très élevé. Pour confirmer ce risque, nous avons mis au point deux tests PCR multiplexes permettant l'identification de tels recombinants. De plus, pour obtenir une première estimation du potentiel d'émergence de ces recombinants ? les recombinants sont-il détectés seuls ou en co-infection avec les parents ? ? les tests ont été conçus pour cibler simultanément les recombinants potentiels et les virus parentaux. Outre la description de la technique, nous présenterons aussi quelques résultats préliminaires qui ont permis de valider la technique. (Texte intégral