Section: New Results

Modelling and analysing a network of individuals, or a network of individuals' networks

Computationally investigating co-phylogenetic reconstructions and co-evolution

Despite an increasingly vast literature on co-phylogenetic reconstructions for studying host-symbiont associations, understanding the common evolutionary history of such systems remains a problem that is far from being solved. Most algorithms for host-symbiont reconciliation use an event-based model, where the events include in general (a subset of) co-speciation, duplication, loss, and host-switch. All known parsimonious event-based methods then assign a cost to each type of event in order to find a reconstruction of minimum cost. The main problem with this approach is that the cost of the events strongly influences the reconciliation obtained.To deal with this problem, we developed an algorithm, called Coala , for estimating the frequency of the events based on an approximate Bayesian computation approach [4] . The benefits of this method are twofold: (1) it provides more confidence in the set of costs to be used in a reconciliation, and (2) it allows estimation of the frequency of the events in cases where the dataset consists of trees with a large number of taxa. We evaluated our method on simulated and on biological datasets. We showed that in both cases, for the same pair of host and parasite trees, different sets of frequencies for the events lead to equally probable solutions. Moreover, often these solutions differ greatly in terms of the number of inferred events. It appears crucial to take this into account before attempting any further biological interpretation of such reconciliations. More generally, we also showed that the set of frequencies can vary widely depending on the input host and parasite trees. Indiscriminately applying a standard vector of costs may thus not be a good strategy. This work had been indicated as submitted in 2014.

Once such a cost vector has been inferred, one can proceed analysing the possible co-evolution of host-symbiont associations, phylogenetic tree reconciliation is the approach of choice for investigating the co-evolution of sets of organisms such as hosts and parasites. It consists in a mapping between the parasite tree and the host tree using event-based maximum parsimony. Given a cost model for the events, many optimal reconciliations are however possible. Only two algorithms existed that attempted such enumeration; in one case not all possible solutions are produced while in the other not all cost vectors are currently handled. We developed a polynomial-delay algorithm, Eucalypt , for enumerating all optimal reconciliations that address these two issues [15] . We showed that in general many solutions exist. We gave an example where, for two pairs of host-parasite trees having each less than 41 leaves, the number of solutions is 5120, even when only time-feasible ones are kept. To facilitate their interpretation, those solutions are also classified in terms of how many of each event they contain. The number of different classes of solutions may thus be notably smaller than the number of solutions, yet they may remain high enough, in particular for the cases where losses have cost 0. In fact, depending on the cost vector, both numbers of solutions and of classes thereof may increase considerably (for the same instance, to respectively 4080384 and 275). To further deal with this problem, we introduced and analysed a restricted version where host-switches are allowed to happen only between species that are within some fixed distance along the host tree. This restriction allowed us to reduce the number of time-feasible solutions while preserving the same optimal cost, as well as to find time-feasible solutions with a cost close to the optimal in the cases where no time-feasible solution is found. This work had been indicated as submitted in 2014.

Evolution and metabolic complementation of organisms leaving inside the cells of another (endosymbionts)

Insect cells host many endosymbiotic bacteria, which are in general classified according to their importance for the host: “primary” symbionts are by definition mandatory and synthesise essential nutrients for the insects that feed on poor or unbalanced food sources, while “secondary” symbionts are optional and use mutualistic strategies and/or manipulation of reproduction to invade and persist within insect populations.

Hamiltonella defensa is a secondary endosymbiont that established two distinct associations with phloemophagous insects. In aphids, it protects the host against parasitoid attacks. Its ability to infect many host tissues, notably the hemolymph, could promote its contact with parasitoid eggs. Despite this protective phenotype, the high costs associated with its presence within the host prevent its fixation in the population. In the whitefly Bemisia tabaci however, this symbiont is found only in cells specialised in hosting endosymbionts, the bacteriocytes. In these cells, it cohabits with other symbiotic species, such as the primary symbiont Portiera aleyrodidarum, a proximity that favours potential exchanges between the two symbionts. It is fixed in populations of B. tabaci, which suggests an important role for the consortium, probably nutritious.

We studied the specificities of each of these systems [27] . First, in the bacteriocytes of B. tabaci, we identified a partitioning of the synthetic capacities of two endosymbionts, H. defensa and P. aleyrodidarum, in addition to a potential metabolic complementation between the symbionts and their host for the synthesis of essential amino acids. We proposed a key nutritive role for H. defensa, which would indicate a transition to a mandatory status in relation to the host and would explain its fixation in the population.

We also focused on the genomic evolution of the genus Hamiltonella, by comparing the strains infecting B. tabaci with a strain infecting the aphids [29] . We highlighted the specialisation of the symbionts to their hosts, and found that the genomes of the endosymbionts reflected their respective ecology. The aphid strain thus possesses many virulence factors and is associated with two partners, a bacteriophage and a recombination plasmid. These systems, inactive in the symbiont of B. tabaci, are directly related to the protection against and arms race with parasitoids. Conversely, the presumed avirulence of whitefly endosymbionts is consistent with their nutritional phenotype and a transition to a mandatory status to the host.

Finally, we studied the phenomenon of “accelerated mutation rate” in H. defensa, compared to its sister species Regiella insecticola, which is also a clade of protective endosymbionts of aphids. After excluding the assumption that the transition to the intracellular life occurred independently in the two lineages, we tried to establish a link between these differences in terms of evolvability in the endosymbionts and of their gene contents, particularly for genes involved in ecology and DNA repair. All the results obtained have provided insight into the evolution of the species H. defensa, since the last ancestor to the present species, by establishing a link between bacterial.

These results were part of the PhD of Pierre-Antoine Rollat-Farnier, co-supervised by Laurence Mouton (LBBE, UMR5558), Marie-France Sagot (Inria and LBBE, UMR5558) and Fabrice Vavre (LBBE, UMR5558) and defended on November 24th, 2014. The results had been indicated as submitted in 2014.

Insights on the virulence of swine respiratory tract mycoplasmas through genome-scale metabolic modelling

The respiratory tract of swines is colonised by several bacteria among which are three Mycoplasma species: Mycoplasma flocculare, Mycoplasma hyopneumoniae and Mycoplasma hyorhinis. While colonisation by M. flocculare was shown to be virtually asymptomatic, M. hyopneumoniae is known to be the causative agent of enzootic pneumonia and M. hyorhinis to be present in cases of pneumonia, polyserositis and arthritis. Nonetheless, the elevated genomic resemblance among these three mycoplasmas combined with their different levels of pathogenicity is an indication that they have unknown mechanisms of virulence and differential expression. We performed whole-genome metabolic network reconstructions for these three mycoplasmas and were able to show that overall they have similar metabolic capabilities. The metabolic differences that were observed include a wider range of carbohydrate uptake in M. hyorhinis, which in turn may also explain why this species is a widely known contaminant in cell cultures. Moreover, the myo-inositol catabolism is exclusive to M. hyopneumoniae and may be an important trait for virulence. However, the most important difference seems to be related to glycerol conversion to dihydroxyacetone-phosphate, which produces toxic hydrogen peroxide. This activity, missing only in M. flocculare, may be directly involved in cytotoxicity, as already been described for two lung pathogenic mycoplasmas, namely Mycoplasma pneumoniae in human and Mycoplasma mycoides subsp. mycoides in ruminants. Metabolomic data suggest that even though these mycoplasmas are extremely similar in terms of their genome and metabolism, different products and reaction rates may be the result of differential expression in each of them. We were able to infer from the reconstructed networks that the lack of pathogenicity of M. flocculare if compared to the highly pathogenic M. hyopneumoniae may be related to its incapacity to produce cytotoxic hydrogen peroxide. Moreover, the ability of M. hyorhinis to grow in diverse sites and even in different hosts may be a reflection of its enhanced and wider carbohydrate uptake. Altogether, the metabolic differences highlighted in silico and in vitro provide important insights to the different levels of pathogenicity observed in each of the studied species.

These results were part of the PhD of Mariana Galvão Ferrarini, co-supervised by Arnaldo Zaha (Federal University of Rio Grande do Sul and Marie-France Sagot (Inria and LBBE, UMR5558). and defended on December 10th, 2015. These results have been submitted to a journal. The PhD manuscript will be made available in HAL in early 2016.