Alexandre Hassanin

Bats host major mammalian paramyxoviruses

The large virus family Paramyxoviridae includes some of the most significant human and livestock viruses, such as measles-, distemper-, mumps-, parainfluenza-, Newcastle disease-, respiratory syncytial virus and metapneumoviruses. Here we identify an estimated 66 new paramyxoviruses in a worldwide sample of 119 bat and rodent species (9,278 individuals). Major discoveries include evidence of an origin of Hendra- and Nipah virus in Africa, identification of a bat virus conspecific with the human mumps virus, detection of close relatives of respiratory syncytial virus, mouse pneumonia- and canine distemper virus in bats, as well as direct evidence of Sendai virus in rodents. Phylogenetic reconstruction of host associations suggests a predominance of host switches from bats to other mammals and birds. Hypothesis tests in a maximum likelihood framework permit the phylogenetic placement of bats as tentative hosts at ancestral nodes to both the major Paramyxoviridae subfamilies (Paramyxovirinae and Pneumovirinae). Future attempts to predict the emergence of novel paramyxoviruses in humans and livestock will have to rely fundamentally on these data. Supplementary information The online version of this article (doi:10.1038/ncomms1796) contains supplementary material, which is available to authorized users.

Evidence for Multiple Reversals of Asymmetric Mutational Constraints during the Evolution of the Mitochondrial Genome of Metazoa, and Consequences for Phylogenetic Inferences

Mitochondrial DNA (mtDNA) sequences are comonly used for inferring phylogenetic relationships. However, the strand-specific bias in the nucleotide composition of the mtDNA, which is thought to reflect assymetric mutational constraints, combined with the important compositional heterogeneity among taxa, are known to be highly problematic for phylogenetic analyses. Here, nucleotide composition was compared across 49 species of Metazoa (34 arthropods, 2 annelids, 2 molluscs, and 11 deuterosomes), and analyzed for a mtDNA fragment including six protein-coding genes, i.e., atp6, atp8, cox1, cox2, cox3, and nad2. The analyses show that most metazoan species present a clear strand assymetry, where one strand is biased in favor of A and C, whereas the other strand has reverse bias, i.e. in favor of T and G. the origin of this strand bias can be related to assymetric mutational constraints involving deaminations of A and C nucleotides during the replication and/or transcription processes. The analyses reveal that six unrelated genera are characterized by a reversal of the usual strand bias, i.e., Argiope (Araneae), Euscorpius (Scorpiones), Tigrioupus (Maxillopoda), Branchiostoma (Cephalochordata) Florometra (Echinodermata), and Katharina (Mollusca). It is proposed that assymetric mutational constraints have been independantly reversed in these six genera, through an inversion of the control region, i.e., the region that contains most regulatory elements for replication and transcription of the mtDNA. We show that reversals of assymetric mutational constraints have dramatic consequences on the phylogenetic analyses, as taxa characterized by reverse strand bias tend to group together due to long-branch attraction artifacts. We propose a new method for limiting this specific problem in tree reconstruction under the Bayesian approach. We apply our method to deal with the question of phylogenetic relationships of the major lineages of Arthropoda, This new approach provides a better congruence with nuclear analyses based on mtDNA sequences, our data suggest that Chelicerata, Crustacea, Myriapoda, Pancrustacea, and Paradoxopoda are monophyletic.

Phylogenetic Analysis of Invertebrate Lysozymes and the Evolution of Lysozyme Function

Abstract. We isolated and sequenced the cDNAs coding for lysozymes of six bivalve species. Alignment and phylogenetic analysis showed that, together with recently described bivalve lysozymes, the leech destabilase, and a number of putative proteins from extensive genomic and cDNA analyses, they belong to the invertebrate type of lysozymes (i type), first described by Jollès and Jollès (1975). We determined the genomic structure of the gene encoding the lysozyme of Mytilus edulis, the common mussel. We provide evidence that the central exon of this gene is homologous to the second exon of the chicken lysozyme gene, belonging to the c type. We propose that the origin of this domain can be traced back in evolution to the origin of bilaterian animals. Phylogenetic analysis suggests that i-type proteins form a monophyletic family.

The 'evolutionary signal' of homoplasy in protein-coding gene sequences and its consequences for a priori weighting in phylogeny.

To analyse independently homoplasy for the six possible types of substitution (i.e., A-G, C-T, A-C, A-T, C-G and G-T) at each of the three codon-positions of the cytochrome b gene, two approaches were used: the first is based on the consistency index which measures the amount of homoplasy, and the second is based on the saturation analysis which describes graphically the distribution of homoplasy within the taxonomic sampling. The results obtained from a data set of 32 sequences of Artiodactyla indicate that evolution of the cytochrome b is governed by differential constraints: 1) between the six substitutions-types, 2) between the three codon-positions, and 3) between the two mtDNA strands. Moreover, we find that non-synonymous sites can be more homoplastic than synonymous sites when the possibilities of substitutions are severely restricted because of the functional requirements of hydrophobicity. Most weighting schemes applied to protein-coding genes are elaborated from unjustified assumptions. We propose to weight each substitution-type at each codon-position according to its homoplasy content evaluated either with the consistency index or with an index representing the level of mutational saturation.

Isolates of Zaire ebolavirus from wild apes reveal genetic lineage and recombinants

Over the last 30 years, Zaire ebolavirus (ZEBOV), a virus highly pathogenic for humans and wild apes, has emerged repeatedly in Central Africa. Thus far, only a few virus isolates have been characterized genetically, all belonging to a single genetic lineage and originating exclusively from infected human patients. Here, we describe the first ZEBOV sequences isolated from great ape carcasses in the Gabon/Congo region that belong to a previously unrecognized genetic lineage. According to our estimates, this lineage, which we also encountered in the two most recent human outbreaks in the Republic of the Congo in 2003 and 2005, diverged from the previously known viruses around the time of the first documented human outbreak in 1976. These results suggest that virus spillover from the reservoir has occurred more than once, as predicted by the multiple emergence hypothesis. However, the young age of both ZEBOV lineages and the spatial and temporal sequence of outbreaks remain at odds with the idea that the virus simply emerged from a long-established and widespread reservoir population. Based on data from two ZEBOV genes, we also demonstrate, within the family Filoviridae, recombination between the two lineages. According to our estimates, this event took place between 1996 and 2001 and gave rise to a group of recombinant viruses that were responsible for a series of outbreaks in 2001–2003. The potential for recombination adds an additional level of complexity to unraveling and potentially controlling the emergence of ZEBOV in humans and wildlife species.

Evolutionary affinities of the enigmatic saola (Pseudoryx nghetinhensis) in the context of the molecular phylogeny of Bovidae

To elucidate the systematic status of the enigmatic saola (Pseudoryx nghetinhensis), a new bovid genus recently discovered in Vietnam, and to investigate phylogenetic relationships within the family Bovidae, four distinct DNA markers were sequenced. Complete mitochondrial cytochrome b (1143 bp) and 12S rRNA (956 bp) genes and non–coding regions from the nuclear genes for aromatase cytochrome P–450 (199 bp) and lactoferrin (338 bp) have been compared for 25 bovid species and three Cervidae and Antilocapridae outgroups. Independent and/or combined analyses of the four nucleotide matrices through maximum parsimony and maximum–likelihood methods indicated that Bovidae consists of two major lineages, i.e. Bovinae which contains the tribes Bovini, Boselaphini and Tragelaphini, and Antilopinae which encompasses all other bovids. Within Bovinae, the tribe Bovini is divided into buffalo Bovini (Bubalus and Syncerus) and cattle Bovini (Bos and Bison) and Tragelaphini are possibly related to Boselaphini. Pseudoryx is shown to be (i) robustly nested within Bovinae; (ii) strongly associated with Bovini; and (iii) tentatively sharing a sister–group relationship with cattle Bovini. Within Antilopinae, three robust clades are in evidence: (i) Hippotragus and Damaliscus are linked to Ovis; (ii) Aepyceros joins Neotragus; and (iii) Cephalophus clusters with Oreotragus.

Captured retroviral envelope syncytin gene associated with the unique placental structure of higher ruminants

Syncytins are envelope genes of retroviral origin that have been co-opted for a role in placentation and likely contribute to the remarkable diversity of placental structures. Independent capture events have been identified in primates, rodents, lagomorphs, and carnivores, where they are involved in the formation of a syncytium layer at the fetomaternal interface via trophoblast cell–cell fusion. We searched for similar genes within the suborder Ruminantia where the placenta lacks an extended syncytium layer but displays a heterologous cell-fusion process unique among eutherian mammals. An in silico search for intact envelope genes within the Bos taurus genome identified 18 candidates belonging to five endogenous retrovirus families, with one gene displaying both placenta-specific expression, as assessed by quantitative RT-PCR analyses of a large panel of tissues, and conservation in the Ovis aries genome. Both the bovine and ovine orthologs displayed fusogenic activity by conferring infectivity on retroviral pseudotypes and triggering cell–cell fusion. In situ hybridization of placenta sections revealed specific expression in the trophoblast binucleate cells, consistent with a role in the formation—by heterologous cell fusion with uterine cells—of the trinucleate cells of the cow and the syncytial plaques of the ewe. Finally, we show that this gene, which we named “Syncytin-Rum1,” is conserved among 16 representatives of higher ruminants, with evidence for purifying selection and conservation of its fusogenic properties, over 30 millions years of evolution. These data argue for syncytins being a major driving force in the emergence and diversity of the placenta.

Molecular phylogenetics unveils the ancient evolutionary origins of the enigmatic fairy armadillos.

Fairy armadillos or pichiciegos (Xenarthra, Dasypodidae) are among the most elusive mammals. Due to their subterranean and nocturnal lifestyle, their basic biology and evolutionary history remain virtually unknown. Two distinct species with allopatric distributions are recognized: Chlamyphorus truncatus is restricted to central Argentina, while Calyptophractus retusus occurs in the Gran Chaco of Argentina, Paraguay, and Bolivia. To test their monophyly and resolve their phylogenetic affinities within armadillos, we obtained sequence data from modern and museum specimens for two mitochondrial genes (12S RNA [MT-RNR1] and NADH dehydrogenase 1 [MT-ND1]) and two nuclear exons (breast cancer 1 early onset exon 11 [BRCA1] and von Willebrand factor exon 28 [VWF]). Phylogenetic analyses provided a reference phylogeny and timescale for living xenarthran genera. Our results reveal monophyletic pichiciegos as members of a major armadillo subfamily (Chlamyphorinae). Their strictly fossorial lifestyle probably evolved as a response to the Oligocene aridification that occurred in South America after their divergence from Tolypeutinae around 32 million years ago (Mya). The ancient divergence date (∼17Mya) for separation between the two species supports their taxonomic classification into distinct genera. The synchronicity with Middle Miocene marine incursions along the Paraná river basin suggests a vicariant origin for pichiciegos by the disruption of their ancestral range. Their phylogenetic distinctiveness and rarity in the wild argue in favor of high conservation priority.

Multiple host-switching of Haemosporidia parasites in bats

BackgroundThere have been reported cases of host-switching in avian and lizard species of Plasmodium (Apicomplexa, Haemosporidia), as well as in those infecting different primate species. However, no evidence has previously been found for host-swapping between wild birds and mammals.MethodsThis paper presents the results of the sampling of blood parasites of wild-captured bats from Madagascar and Cambodia. The presence of Haemosporidia infection in these animals is confirmed and cytochrome b gene sequences were used to construct a phylogenetic analysis.ResultsResults reveal at least three different and independent Haemosporidia evolutionary histories in three different bat lineages from Madagascar and Cambodia.ConclusionPhylogenetic analysis strongly suggests multiple host-switching of Haemosporidia parasites in bats with those from avian and primate hosts.

Comparisons between mitochondrial genomes of domestic goat (Capra hircus) reveal the presence of numts and multiple sequencing errors

Materials and methods. In the present study, we amplified and sequenced the complete mitochondrial genome from a Vietnamese domestic goat (Capra hircus). The data were compared with mtDNA sequences available in the nucleotide databases. Results. The results revealed many problems in the goat mitochondrial reference genome (GenBank accession number NC_005044). Firstly, the authors did not sequence the complete genome, simply 44.5% of its total length. Secondly, two fragments (representing 1201 and 2384 nt) contained an unusually high percentage of sequencing errors. Thirdly, a segment of 1881 nt, covering most of nd5 and the 5′ part of nd6, was shown to be a nuclear sequence of mitochondrial origin (Numt). Surprisingly, a similar Numt was also detected in four other goat mitochondrial genomes available in GenBank (GU22978-81). Two primers were designed specially to amplify ∼960 nt of the Numt identified in goat mtDNA genomes. After cloning, two Numts were detected for C. hircus. Several Numts, most of them with stop codon or frameshift mutations, were also found in Hemitragus jemlahicus (Himalayan tahr) and Pseudois nayaur (bharal). Phylogenetic analyses suggest that a nuclear integration occurred in the common ancestor of Ammotragus, Arabitragus, Capra, Hemitragus and Pseudois, followed by several subsequent duplication events. Conclusion. As poor-quality sequences can produce misleading interpretations of both phylogeny and molecular evolution, we propose including a new link to each accession number in the nucleotide databases, named “external expertise”, which could be openly and continually updated by non-anonymous searchers in order to validate good-quality data, or, conversely, to indicate possible problems in the sequence, such as DNA contamination or sequencing errors. This information could prove very useful over time to select good-quality sequences for in silico analyses.