A Rift Valley fever (RVF) outbreak occurred in Madagascar from January to May 2008. The objectives of this study were (1) to assess the current and past circulation of RVF virus (RVFV) in livestock in Madagascar and (2) to evaluate the extent and magnitude of the 2008 RVF outbreak in livestock. The results of a country-wide serosurvey conducted in August 2008 on small and large ruminants are reported here. The study included 3437 cattle and 989 small ruminants (227 sheep and 762 goats) sampled in 30 of the 111 Malagasy districts, selected to be representative of the different ecozones and livestock density areas. Sera of animals were tested for the detection of immunoglobulins M (IgM) and G (IgG) against RVFV using commercial enzyme-linked immunosorbent assays kits. Recent infections (presence of IgM against RVFV) were detected in only 9 cattle (0.3% [0.1-0.4]) and 33 small ruminant (3.3% [2.2-4.5]) samples. Past infections (presence of IgG and absence of IgM against RVFV) were detected in 887 cattle (25.8% [24.3-27.3]) and 244 small ruminant (24.7% [22.0-27.4]) samples. Past infections were detected in all sampled sites. All ecozones were affected. In the southern and northwestern areas, the prevalence of cattle showing evidence of past infection with RVFV increased with the age of the animals. Our results suggest that there has been country-wide circulation of RVFV in 2008 in Madagascar, including in parts of the country where no clinical illness, either in animals or in humans, was reported. The data also suggest that the southern and northwestern areas may be endemic for RVFV, and that the virus may spread when ecological conditions are favorable for its amplification.

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 18th century.

The performance of Whatman 3-MM filter papers for the collection, drying, shipment and long-term storage of blood at ambient temperature, and for the detection of African swine fever virus and antibodies was assessed. Conventional and real-time PCR, viral isolation and antibody detection by ELISA were performed on paired samples (blood/tissue versus dried-blood 3-MM filter papers) collected from experimentally infected pigs and from farm pigs in Madagascar and Côte d'Ivoire. 3-MM filter papers were used directly in the conventional and real-time PCR without previous extraction of nucleic acids. Tests that performed better with 3-MM filter papers were in descending order: virus isolation, real-time UPL PCR and conventional PCR. The analytical sensitivity of real-time UPL PCR on filter papers was similar to conventional testing (virus isolation or conventional PCR) on organs or blood. In addition, blood-dried filter papers were tested in ELISA for antibody detection and the observed sensitivity was very close to conventional detection on serum samples and gave comparable results. Filter papers were stored up to 9 months at 20-25°C and for 2 months at 37°C without significant loss of sensitivity for virus genome detection. All tests on 3-MM filter papers had 100% specificity compared to the gold standards. Whatman 3-MM filter papers have the advantage of being cheap and of preserving virus viability for future virus isolation and characterization. In this study, Whatman 3-MM filter papers proved to be a suitable support for the collection, storage and use of blood in remote areas of tropical countries without the need for a cold chain and thus provide new possibilities for antibody testing and virus isolation.

African swine fever (ASF) suspected clinically in Madagascar (1998-9) was confirmed by polymerase chain reaction (PCR) and nucleotide sequencing, following virus isolation. No haemadsorption or cytopathic effect could be detected following leukocyte inoculation, but viral growth in cells was confirmed by PCR. Detection of ASF virus genome was carried out by amplification of a highly conserved region coding for the p72 protein. Nucleotide sequencing of the amplicon revealed 99.2% nucleotide identity between the recent Malagasy strains and a virus recovered from the 1994 outbreak in Mozambique (SPEC265). A serological survey performed on 449 sera, revealed that only 5.3% of the sera taken from pigs between 1998 and 1999 were positive.

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.