Pseudomonas syringae on plants in Iceland has likely evolved for several million years outside the reach of processes that mix this bacterial complex across earth’s temperate zones

Funding Information: Funding: This research was funded by (i) the Campus France/ Partenariat Hubert Curien Jules Verne Franco-Icelandic Exchange Program project 40885YF, (ii) the Ranis Icelandic Research Fund project 206801–051 and (iii) French National Research Agency (ANR) project SPREE-17-CE32-0004-01. Publisher Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland.Here we report, for the first time, the occurrence of the bacteria from the species complex Pseudomonas syringae in Iceland. We isolated this bacterium from 35 of the 38 samples of angiosperms, moss, ferns and leaf litter collected across the island from five habitat categories (boreal heath, forest, subalpine and glacial scrub, grazed pasture, lava field). The culturable populations of P. syringae on these plants varied in size across 6 orders of magnitude, were as dense as 107 cfu g−1 and were composed of strains in phylogroups 1, 2, 4, 6, 7, 10 and 13. P. syringae densities were significantly greatest on monocots compared to those on dicots and mosses and were about two orders of magnitude greater in grazed pastures compared to all other habitats. The phylogenetic diversity of 609 strains of P. syringae from Iceland was compared to that of 933 reference strains of P. syringae from crops and environmental reservoirs collected from 27 other countries based on a 343 bp sequence of the citrate synthase (cts) housekeeping gene. Whereas there were examples of identical cts sequences across multiple countries and continents among the reference strains indicating mixing among these countries and continents, the Icelandic strains grouped into monophyletic lineages that were unique compared to all of the reference strains. Based on estimates of the time of divergence of the Icelandic genetic lineages of P. syringae, the geological, botanical and land use history of Iceland, and atmospheric circulation patterns, we propose scenarios whereby it would be feasible for P. syringae to have evolved outside the reach of processes that tend to mix this bacterial complex across the planet elsewhere.Here we report, for the first time, the occurrence of the bacteria from the species complex Pseudomonas syringae in Iceland. We isolated this bacterium from 35 of the 38 samples of angiosperms, moss, ferns and leaf litter collected across the island from five habitat categories (boreal heath, forest, subalpine and glacial scrub, grazed pasture, lava field). The culturable populations of P. syringae on these plants varied in size across 6 orders of magnitude, were as dense as 107 cfu g−1 and were com-posed of strains in phylogroups 1, 2, 4, 6, 7, 10 and 13. P. syringae densities were significantly greatest on monocots compared to those on dicots and mosses and were about two orders of magnitude greater in grazed pastures compared to all other habitats. The phylogenetic diversity of 609 strains of P. syringae from Iceland was compared to that of 933 reference strains of P. syringae from crops and environmental reservoirs collected from 27 other countries based on a 343 bp sequence of the citrate synthase (cts) housekeeping gene. Whereas there were examples of identical cts sequences across mul-tiple countries and continents among the reference strains indicating mixing among these countries and continents, the Icelandic strains grouped into monophyletic lineages that were unique compared to all of the reference strains. Based on estimates of the time of divergence of the Icelandic genetic lineages of P. syringae, the geological, botanical and land use history of Iceland, and atmospheric circulation patterns, we propose scenarios whereby it would be feasible for P. syringae to have evolved outside the reach of processes that tend to mix this bacterial complex across the planet elsewhere.Peer reviewe

A short motif in the N-terminal part of the coat protein is a host-specific determinant of systemic infectivity for two potyviruses.

International audienceAlthough the biological variability of Watermelon mosaic virus is limited, isolates from the three main molecular groups differ in their ability to infect systemically Chenopodium quinoa. Mutations were introduced in a motif of three or five amino acids located in the N-terminal part of the coat protein, and differing in isolates from group 1 (motif: lysine-glutamic acid-alanine (Lys-Glu-Ala) or KEA, systemic on C. quinoa), group 2 (Lys-Glu-Thr or KET, not systemic on C. quinoa) and group 3 (KEKET, not systemic on C. quinoa). Mutagenesis of KEKET in an isolate from group 3 to KEA or KEKEA was sufficient to make the virus systemic on C. quinoa, whereas mutagenesis to KET had no effect. Introduction of a KEA motif in Zucchini yellow mosaic virus coat protein also resulted in systemic infection on C. quinoa. These mutations had no obvious effect on the disorder profile or potential post-translational modifications of the coat protein as determined in silico

A simple, rapid and efficient way to obtain infectious clones of potyviruses

International audienceThe availability of an infectious cDNA clone is a prerequisite for genetic studies on RNA viruses. However, despite important improvement in molecular biology techniques during the last decades, obtaining such clones often remains tedious, time-consuming and rather unpredictable. In the case of potyviruses, cDNA clones are frequently unstable due to the toxicity of some viral proteins for bacteria. The problem can be overcome by inserting introns into the viral sequence but this requires additional steps in the cloning process and depends on the availability of suitable restriction sites in the viral sequence or adjunction of such sites by mutagenesis. Homologous recombination in yeast rather than in vitro restriction and ligation can be used to build infectious clones or other viral constructs. This paper describes how, by using recombination in yeast and fusion PCR, infectious intron-containing clones were obtained within a few weeks for two strains of watermelon mosaic virus (WMV, Potyvirus), whereas previous attempts using “classical” cloning techniques had failed repeatedly. Using the same approach, intronless infectious clones of two other potyviruses, zucchini yellow mosaic virus (ZYMV) and papaya ringspot virus (PRSV), were obtained in less than two week

Une enquête riche d'enseignements: Melon/courgette

National audienc

Results of a survey for cucurbit viruses in France and new insights on Watermelon mosaic virus and Zucchini yellow mosaic virus epidemiology

International audienc

Do recombinants appearing in natural populations of watermelon mosaic virus represent new agronomic threats?

International audienceWatermelon mosaic virus (WMV, Potyvirus) is very common in France where it has been present for at least 40 years. Since the early 2000s, new “emerging” (EM) strains of WMV, highly divergent molecularly from the “classic” (CL) strains present before and probably originating from recent introductions, were detected in southeastern France. Since both types of strains did not appear to be present in the same geographic locations before, this situation constituted a unique opportunity to study the frequency of appearance and the potential spread of recombinants in the few years following the introduction of the new strains. Analyzing isolates from experimental plots in Montfavet (Southeastern France) as well as from epidemiological surveys performed from 2004 to 2008 all around France (about 2000 WMV isolates) revealed at least 7 independent recombination events, either between CL and EM strains or between different EM subgroups. Most recombinants were found in a few plants from the same field, but, with one notable exception, did not seem to spread or be maintained locally for several years. Mixed infections of CL and EM isolates were also performed in experimental conditions in order to compare the frequency and nature of recombination events to those of natural situation. The fitness of natural recombinants relative to potential “parental” strains was also tested in controlled conditions. The epidemiological and evolutionary consequences of recombination in WMV populations will be discussed. This work represents one of the first estimation for the frequency of appearance of recombinants in natural populations of a plant RNA virus

Do recombinants appearing in natural populations of watermelon mosaic virus represent new agronomic threats?

International audienceWatermelon mosaic virus (WMV, Potyvirus) is very common in France where it has been present for at least 40 years. Since the early 2000s, new “emerging” (EM) strains of WMV, highly divergent molecularly from the “classic” (CL) strains present before and probably originating from recent introductions, were detected in southeastern France. Since both types of strains did not appear to be present in the same geographic locations before, this situation constituted a unique opportunity to study the frequency of appearance and the potential spread of recombinants in the few years following the introduction of the new strains. Analyzing isolates from experimental plots in Montfavet (Southeastern France) as well as from epidemiological surveys performed from 2004 to 2008 all around France (about 2000 WMV isolates) revealed at least 7 independent recombination events, either between CL and EM strains or between different EM subgroups. Most recombinants were found in a few plants from the same field, but, with one notable exception, did not seem to spread or be maintained locally for several years. Mixed infections of CL and EM isolates were also performed in experimental conditions in order to compare the frequency and nature of recombination events to those of natural situation. The fitness of natural recombinants relative to potential “parental” strains was also tested in controlled conditions. The epidemiological and evolutionary consequences of recombination in WMV populations will be discussed. This work represents one of the first estimation for the frequency of appearance of recombinants in natural populations of a plant RNA virus

Les virus des cucurbitacées

National audienc

Evolution et dispersion d'un virus des cucurbitacées, le Watermelon mosaic virus. Résumé

National audienc

Evolution of populations of cucurbit-infecting potyviruses in France: insights into the mechanisms of plant virus emergence

International audienceWatermelon mosaic virus (WMV) and Zucchini yellow mosaic virus (ZYMV) are among the major cucurbit-infecting viruses worldwide. In France, WMV has a high prevalence every year throughout the growing season; ZYMV, although it can present locally a very high agronomic impact, induces epidemics that are very irregular both in spatial distribution and timing of infection. Large-scale epidemiological surveys followed by molecular analysis of all collected samples were performed during 5 years in France, and revealed the recent appearance of new, “emerging” (EM) isolates of both viruses in south-eastern France. In the case of WMV, four subgroups of EM isolates were defined, probably resulting from several recent introductions. They presented a strong geographic structure that remained stable over 5 years, and tended to replace rapidly the original “classic” isolates in areas where both types of strains were present. For ZYMV, at least two subgroups of new isolates were also detected in the last years, although the geographic structure of infections and evolution of populations was less obvious than for WMV. In order to characterize the evolution of viral populations and study the mechanisms favouring or limiting viral emergence and spread, several complementary approaches were developed: multilocal and multiscale epidemiological studies, molecular analyses and population genetics, as well as estimation of viral fitness in experimental condition and modeling. The risks of emergence of recombinants between “classic” and EM populations in the few years following their contact were also assessed