A Spatially Explicit Metapopulation Model and Cattle Trade Analysis Suggests Key Determinants for the Recurrent Circulation of Rift Valley Fever Virus in a Pilot Area of Madagascar Highlands

Rift Valley fever (RVF) is a vector-borne zoonotic disease that causes high morbidity and mortality in ruminants. In 2008–2009, a RVF outbreak affected the whole Madagascar island, including the Anjozorobe district located in Madagascar highlands. An entomological survey showed the absence of Aedes among the potential RVF virus (RVFV) vector species identified in this area, and an overall low abundance of mosquitoes due to unfavorable climatic conditions during winter. No serological nor virological sign of infection was observed in wild terrestrial mammals of the area, suggesting an absence of wild RVF virus (RVFV) reservoir. However, a three years serological and virological follow-up in cattle showed a recurrent RVFV circulation. The objective of this study was to understand the key determinants of this unexpected recurrent transmission. To achieve this goal, a spatial deterministic discrete-time metapopulation model combined with cattle trade network was designed and parameterized to reproduce the local conditions using observational data collected in the area. Three scenarios that could explain the RVFV recurrent circulation in the area were analyzed: (i) RVFV overwintering thanks to a direct transmission between cattle when viraemic cows calve, vectors being absent during the winter, (ii) a low level vector-based circulation during winter thanks to a residual vector population, without direct transmission between cattle, (iii) combination of both above mentioned mechanisms. Multi-model inference methods resulted in a model incorporating both a low level RVFV winter vector-borne transmission and a direct transmission between animals when viraemic cows calve. Predictions satisfactorily reproduced field observations, 84% of cattle infections being attributed to vector-borne transmission, and 16% to direct transmission. These results appeared robust according to the sensitivity analysis. Interweaving between agricultural works in rice fields, seasonality of vector proliferation, and cattle exchange practices could be a key element for understanding RVFV circulation in this area of Madagascar highlands.     

High Prevalence of West Nile Virus in Domestic Birds and Detection in 2 New Mosquito Species in Madagascar

West Nile virus is an arthropod-borne zoonosis transmitted by a large number of mosquito species, and birds play a key role as reservoir of the virus. Its distribution is largely widespread over Africa, Asia, the Americas and Europe. Since 1978, it has frequently been reported in Madagascar. Studies described a high seroprevalence level of the virus in humans in different areas of the island and a human fatal case of WNV infection was reported in 2011. Despite these reports, the epidemiology of WNV in Madagascar, in particular, viral circulation remains unclear. To explore the transmission of WNV in two rural human populations of Madagascar, we investigated local mosquitoes and poultry for evidence of current infections, and determined seroprevalence of candidate sentinel species among the local poultry. These 2 areas are close to lakes where domestic birds, migratory wild birds and humans coexist. Serological analysis revealed WNV antibodies in domestic birds (duck, chicken, goose, turkey and guinea fowl) sampled in both districts (Antsalova 29.4% and Mitsinjo 16.7%). West Nile virus nucleic acid was detected in one chicken and in 8 pools of mosquitoes including 2 mosquito species (Aedeomyia madagascarica and Anopheles pauliani) that have not been previously described as candidate vectors for WNV. Molecular analysis of WNV isolates showed that all viruses detected were part of the lineage 2 that is mainly distributed in Africa, and were most closely matched by the previous Malagasy strains isolated in 1988. Our study showed that WNV circulates in Madagascar amongst domestic birds and mosquitoes, and highlights the utility of poultry as a surveillance tool to detect WNV transmission in a peri-domestic setting.     

Genetic Differentiation and Demographic History of the Northern Rufous Mouse Lemur (<Emphasis Type="Italic">Microcebus tavaratra</Emphasis>) Across a Fragmented Landscape in Northern Madagascar

Phylogeographic barriers, together with habitat loss and fragmentation, contribute to the evolution of a species’ genetic diversity by limiting gene flow and increasing genetic differentiation among populations. Changes in connectivity can thus affect the genetic diversity of populations, which may influence the evolutionary potential of species and the survival of populations in the long term. We studied the genetic diversity of the little known Northern rufous mouse lemur (Microcebus tavaratra), endemic to Northern Madagascar. We focused on the population of M. tavaratra in the Loky–Manambato region, Northern Madagascar, a region delimited by two permanent rivers and characterized by a mosaic of fragmented forests. We genotyped 148 individuals at three mitochondrial loci (D-loop, cytb, and cox2) in all the major forests of the study region. Our analyses suggest that M. tavaratra holds average genetic diversity when compared to other mouse lemur species, and we identified two to four genetic clusters in the study region, a pattern similar to that observed in another lemur endemic to the region (Propithecus tattersalli). The main cluster involved samples from the two mountain forests in the study region, which were connected until recently. However, the river crossing the study region does not appear to be a strict barrier to gene flow in M. tavaratra. Finally, the inferred demographic history of M. tavaratra suggests no detectable departure from stationarity over the last millennia. Comparisons with codistributed species (P. tattersalli and two endemic rodents, Eliurus spp.) suggest both differences and similarities in the genetic clusters identified (i.e., barriers to species dispersal) and in the inferred demographic history. These comparisons suggest that studies of codistributed species are important to understand the effects of landscape features on species and to reconstruct the history of habitat changes in a region.     

Effect of O. porcinus Tick Salivary Gland Extract on the African Swine Fever Virus Infection in Domestic Pig

African swine fever is a haemorrhagic disease in pig production that can have disastrous financial consequences for farming. No vaccines are currently available and animal slaughtering or area zoning to restrict risk-related movements are the only effective measures to prevent the spread of the disease. Ornithodoros soft ticks are known to transmit the African swine fever virus (ASFV) to pigs in farms, following the natural epidemiologic cycle of the virus. Tick saliva has been shown to modulate the host physiological and immunological responses during feeding on skin, thus affecting viral infection. To better understand the interaction between soft tick, ASFV and pig at the bite location and the possible influence of tick saliva on pig infection by ASFV, salivary gland extract (SGE) of Ornithodoros porcinus, co-inoculated or not with ASFV, was used for intradermal auricular inoculation. Our results showed that, after the virus triggered the disease, pigs inoculated with virus and SGE presented greater hyperthermia than pigs inoculated with virus alone. The density of Langerhans cells was modulated at the tick bite or inoculation site, either through recruitment by ASFV or inhibition by SGE. Additionally, SGE and virus induced macrophage recruitment each. This effect was enhanced when they were co-inoculated. Finally, the co-inoculation of SGE and virus delayed the early local spread of virus to the first lymph node on the inoculation side. This study has shown that the effect of SGE was powerful enough to be quantified in pig both on the systemic and local immune response. We believe this model should be developed with infected tick and could improve knowledge of both tick vector competence and tick saliva immunomodulation.     

Forest Type Influences Population Densities of Nocturnal Lemurs in Manompana,Northeastern Madagascar

Forest loss, fragmentation, and anthropization threaten the survival of forest species all over the world. Shifting agriculture is one of these threatening processes in Madagascar. However, when its cycle is halted and the land is left to regenerate, the resulting growth of secondary forest may provide a viable habitat for folivorous and omnivorous lemur species. We aimed to identify the response of nocturnal lemurs to different successional stages of regenerating secondary, degraded mature, and mature forest across a mosaic-type landscape. We surveyed four nocturnal lemur species (Avahi laniger, Microcebus cf. simmonsi, Allocebus trichotis, and Daubentonia madagascariensis) in four forest types of varying habitat disturbance in northeastern Madagascar. We estimated densities in mature and regenerating secondary forest for the eastern woolly lemur (Avahi laniger) and mouse lemur (Microcebus cf. simmonsi), two sympatric species with folivorous and omnivorous diets respectively. We did not estimate densities of Allocebus trichotis and Daubentonia madagascariensis owing to small sample size; however, we observed both species exclusively in mature forest. We found higher population densities of A. laniger and M. cf. simmonsi in secondary than in mature forest, showing the potential of regenerating secondary forest for lemur conservation. Several environmental factors influenced the detectability of the two lemur species. While observer and habitat type influenced detection of the eastern woolly lemur, canopy height and vine density influenced detection of mouse lemurs. Understanding how different species with different diets interact with anthropogenically impacted habitat will aid future management decisions for the conservation of primate species.     

Endogeic earthworms shape bacterial functional communities and affect organic matter mineralization in a tropical soil

Priming effect (PE) is defined as a stimulation of the mineralization of soil organic matter (SOM) following a supply of fresh organic matter. This process can have important consequences on the fate of SOM and on the management of residues in agricultural soils, especially in tropical regions where soil fertility is essentially based on the management of organic matter. Earthworms are ecosystem engineers known to affect the dynamics of SOM. Endogeic earthworms ingest large amounts of soil and assimilate a part of organic matter it contains. During gut transit, microorganisms are transported to new substrates and their activity is stimulated by (i) the production of readily assimilable organic matter (mucus) and (ii) the possible presence of fresh organic residues in the ingested soil. The objective of our study was to see (i) whether earthworms impact the PE intensity when a fresh residue is added to a tropical soil and (ii) whether this impact is linked to a stimulation/inhibition of bacterial taxa, and which taxa are affected. A tropical soil from Madagascar was incubated in the laboratory, with a ¹³C wheat straw residue, in the presence or absence of a peregrine endogeic tropical earthworm, Pontoscolex corethrurus. Emissions of ¹²CO₂ and ¹³CO₂ were followed during 16 days. The coupling between DNA-SIP (stable isotope probing) and pyrosequencing showed that stimulation of both the mineralization of wheat residues and the PE can be linked to the stimulation of several groups especially belonging to the Bacteroidetes phylum.     

Comprehensive Phylogenetic Reconstructions of African Swine Fever Virus: Proposal for a New Classification and Molecular Dating of the Virus

African swine fever (ASF) is a highly lethal disease of domestic pigs caused by the only known DNA arbovirus. It was first described in Kenya in 1921 and since then many isolates have been collected worldwide. However, although several phylogenetic studies have been carried out to understand the relationships between the isolates, no molecular dating analyses have been achieved so far. In this paper, comprehensive phylogenetic reconstructions were made using newly generated, publicly available sequences of hundreds of ASFV isolates from the past 70 years. Analyses focused on B646L, CP204L, and E183L genes from 356, 251, and 123 isolates, respectively. Phylogenetic analyses were achieved using maximum likelihood and Bayesian coalescence methods. A new lineage-based nomenclature is proposed to designate 35 different clusters. In addition, dating of ASFV origin was carried out from the molecular data sets. To avoid bias, diversity due to positive selection or recombination events was neutralized. The molecular clock analyses revealed that ASFV strains currently circulating have evolved over 300 years, with a time to the most recent common ancestor (TMRCA) in the early 18^th century.     

Genetic and morphological diversity of mouse lemurs (Microcebus spp.) in northern Madagascar: The discovery of a putative new species?

Tropical forests harbor extremely high levels of biological diversity and are quickly disappearing. Despite the increasingly recognized high rate of habitat loss, it is expected that new species will be discovered as more effort is put to document tropical biodiversity. Exploring under‐studied regions is particularly urgent if we consider the rapid changes in habitat due to anthropogenic activities. Madagascar is known for its extraordinary biological diversity and endemicity. It is also threatened by habitat loss and fragmentation. It holds more than 100 endemic primate species (lemurs). Among these, Microcebus (mouse lemurs) is one of the more diverse genera. We sampled mouse lemurs from several sites across northern Madagascar, including forests never sampled before. We obtained morphological data from 99 Microcebus individuals; we extracted DNA from tissue samples of 42 individuals and amplified two mitochondrial loci (cytb and cox2) commonly used for species identification. Our findings update the distribution of three species (Microcebus tavaratra, Microcebus arnholdi, and Microcebus mamiratra), including a major increase in the distribution area of M. arnholdi. We also report the discovery of a new Microcebus lineage genetically related to M. arnholdi. Several complementary approaches suggest that the newly identified Microcebus lineage might correspond to a new putative species, to be confirmed or rejected with additional data. In addition, morphological analyses showed (a) clear phenotypic differences between M. tavaratra and M. arnholdi, but no clear differences between the new Microcebus lineage and the sister species M. arnholdi; and (b) a significant correlation between climatic variables and morphology, suggesting a possible relationship between species identity, morphology, and environment. By integrating morphological, climatic, genetic, and spatial data of two northern Microcebus species, we show that the spatial distribution of forest‐dwelling species may be used as a proxy to reconstruct the past spatial changes in forest cover and vegetation type.