Quantitative real-time PCR detection of Zika virus and evaluation with field-caught mosquitoes

BACKGROUND Zika virus (ZIKV), a mosquito borne flavivirus is a pathogen affecting humans in Asia and Africa. ZIKV infection diagnosis relies on serology-which is challenging due to cross-reactions with other flaviviruses and/or absence or low titer of IgM and IgG antibodies at early phase of infection- virus isolation, which is labor intensive, time consuming and requires appropriate containment. Therefore, real-time RT-PCR (rRT-PCR) is an appealing option as a rapid, sensitive and specific method for detection of ZIKV in the early stage of infection. So far, only one rRT-PCR assay has been described in the context of the outbreak in Micronesia in 2007. In this study, we described a one step rRT-PCR for ZIKV which can detect a wider genetic diversity of ZIKV isolates from Asia and Africa. RESULTS The NS5 protein coding regions of African ZIKV isolates were sequenced and aligned with representative flaviviruses sequences from GenBank to design primers and probe from conserved regions. The analytical sensitivity of the assay was evaluated to be 32 genome-equivalents and 0.05 plaque forming unit (pfu). The assay was shown to detect 37 ZIKV isolates covering a wide geographic in Africa and Asia over 36 years but none of the 31 other flaviviruses tested showing high analytical specificity. The rRT-PCR could be performed in less than 3 hours. This method was used successfully to detect ZIKV strains from field-caught mosquitoes. CONCLUSION We have developed a rapid, sensitive and specific rRT-PCR for detection of ZIKV. This assay is a useful tool for detection of ZIKV infection in regions where a number of other clinically indistinguishable arboviruses like dengue or chikungunya co-circulate. Further studies are needed to validate this assay in clinical positive samples collected during acute ZIKV infection

Larval ecology of mosquitoes in sylvatic arbovirus foci in southeastern Senegal

BACKGROUND: Although adult mosquito vectors of sylvatic arbovirus [yellow fever (YFV), dengue-2 (DENV-2) and chikungunya (CHIKV)] have been studied for the past 40 years in southeastern Senegal, data are still lacking on the ecology of larval mosquitoes in this area. In this study, we investigated the larval habitats of mosquitoes and characterized their seasonal and spatial dynamics in arbovirus foci. METHODS: We searched for wet microhabitats, classified in 9 categories, in five land cover classes (agriculture, forest, savannah, barren and village) from June, 2010 to January, 2011. Mosquito immatures were sampled monthly in up to 30 microhabitats of each category per land cover and bred until adult stage for determination. RESULTS: No wet microhabitats were found in the agricultural sites; in the remaining land covers immature stages of 35 mosquito species in 7 genera were sampled from 9 microhabitats (tree holes, fresh fruit husks, decaying fruit husks, puddles, bamboo holes, discarded containers, tires, rock holes and storage containers). The most abundant species was Aedes aegypti formosus, representing 30.2% of the collections, followed by 12 species, representing each more than 1% of the total, among them the arbovirus vectors Ae. vittatus (7.9%), Ae. luteocephalus (5.7%), Ae. taylori (5.0%), and Ae. furcifer (1.3%). Aedes aegypti, Cx. nebulosus, Cx. perfuscus, Cx. tritaeniorhynchus, Er. chrysogster and Ae. vittatus were the only common species collected from all land covers. Aedes furcifer and Ae. taylori were collected in fresh fruit husks and tree holes. Species richness and dominance varied significantly in land covers and microhabitats. Positive associations were found mainly between Ae. furcifer, Ae. taylori and Ae. luteocephalus. A high proportion of potential enzootic vectors that are not anthropophilic were found in the larval mosquito fauna. CONCLUSIONS: In southeastern Senegal, Ae. furcifer and Ae. taylori larvae showed a more limited distribution among both land cover and microhabitat types than the other common species. Uniquely among vector species, Ae. aegypti formosus larvae occurred at the highest frequency in villages. Finally, a high proportion of the potential non-anthropophilic vectors were represented in the larval mosquito fauna, suggesting the existence of unidentified sylvatic arbovirus cycles in southeastern Senegal

Rift Valley fever outbreak, Mauritania, 1998: seroepidemiologic, virologic, entomologic, and zoologic investigations.

A Rift Valley fever outbreak occurred in Mauritania in 1998. Seroepidemiologic and virologic investigation showed active circulation of the Rift Valley fever virus, with 13 strains isolated, and 16% (range 1.5%-38%) immunoglobulin (Ig) M-positivity in sera from 90 humans and 343 animals (sheep, goats, camels, cattle, and donkeys). One human case was fatal

Détermination du débit de filtration glomérulaire (DFG) au cours du diabète : Cockroft et Gault, MDRD ou CKD-EPI ?

Plusieurs paramètres peuvent être étudiés pour évaluer le rein. Parmi ceux-ci, le débit de filtration glomérulaire (DFG) a été déterminé avec les formules de Cockroft et Gault (CG), du Modification of Diet in Renal Disease (MDRD) et du Chronic Kidney Disease EPIdemiology Collaboration (CKD-EPI) et la formule la mieux adaptée pour le diabétique a été recherchée. Chez 59 diabétiques de type 1 (DT1) et 70 diabétiques de type 2 (DT2), le DFG a été déterminé avec les formules de CG, du MDRD et du CKD-EPI. Avec l’analyse statistique, les seuils de significativité ont été fixés pour p<0,05 ; T0α>1,96 et Z0α>1,96. Le MDRD est superposable au CKD-EPI chez les DT1 et DT2. Chez les DT1, le DFG moyen et la corrélation entre 1/créatininémie et DFG ne varient pas si CG ou CKD-EPI ; cependant, les sujets à DFG réduit (< 90 ml/min/1,73 m²) sont plus nombreux avec CG plutôt qu’avec CKD-EPI (66,10% vs 47,46% ; T0α=2,05). Chez les DT2, le DFG moyen et la proportion de sujets à DFG réduit sont indépendants de la formule utilisée, mais la corrélation entre 1/créatininémie et DFG est plus forte si CKD-EPI que CG (0,961 vs 0,632 ; Z0α=7,02). Ainsi, la formule la mieux adaptée pour la détermination du DFG serait CG chez les DT1 et CKD-EPI chez les DT2, sachant que CKD-EPI est équivalent à MDRD quel que soit le type de diabète.Mots clés : Cockroft et Gault - MDRD - CKD-EPI – débit de filtration glomérulaire (DFG) – diabète

Phylogeography of Rift Valley Fever Virus in Africa Reveals Multiple Introductions in Senegal and Mauritania

Rift Valley Fever (RVF) virus (Family Bunyaviridae) is an arthropod-borne RNA virus that infects primarily domestic ruminants and occasionally humans. RVF epizootics are characterized by numerous abortions and mortality among young animals. In humans, the illness is usually characterized by a mild self-limited febrile illness, which could progress to more serious complications. RVF virus is widespread and endemic in many regions of Africa. In Western Africa, several outbreaks have been reported since 1987 when the first major one occurred at the frontier of Senegal and Mauritania. Aiming to evaluate the spreading and molecular epidemiology in these countries, RVFV isolates from 1944 to 2008 obtained from 18 localities in Senegal and Mauritania and 15 other countries were investigated. Our results suggest that a more intense viral activity possibly took place during the last century compared to the recent past and that at least 5 introductions of RVFV took place in Senegal and Mauritania from distant African regions. Moreover, Barkedji in Senegal was possibly a hub associated with the three distinct entries of RVFV in West Africa

Letter to the Editor : Sylvatic Dengue Viruses Share the Pathogenic Potential of Urban/Endemic Dengue Viruses

Dengue virus (DENV) exists in both sylvatic and urban/endemic ecotypes (15), and the potential for emergence of sylvatic strains has become a focus of research. Recently Mota and Rico-Hesse (10) attempted to evaluate the pathogenic potential of viruses belonging to different genetic subgroups of DENV serotype 2 (DENV-2). Based on the viremia levels and erythema index profiles of one sylvatic genotype and three (Asian, American, and Indian) urban/endemic genotypes evaluated using the NOD-scid IL2rγnull humanized mouse model, the authors concluded that sylvatic DENV-2 viruses possess a reduced pathogenic potential compared to strains belonging to urban/endemic DENV-2 genotypes. However, these conclusions ignore both patterns in their own data and a wealth of published ex vivo, in vivo, and epidemiological evidence collected over the past 40 years

Development of a Usutu virus specific real-time reverse transcription PCR assay based on sequenced strains from Africa and Europe

Usutu virus (USUV) has been isolated in several African and European countries mainly from mosquitoes and birds. However, previous benign and two recent severe cases of human infections point out the need of a tool for the identification of USUV in human samples. A published real-time reverse transcription (RT) PCR assay for the detection of USUV in human blood or cerebrospinal fluid does not take into account the genetic variability of USUV in different geographic regions. Therefore, this article presents a quantitative real-time RT-PCR assay based on sequences from Europe and Africa. Primers and probe were designed in conserved regions among USUV strains that differed from closely related flaviviruses. The specificity of the assay was investigated by testing 16 other flaviviruses circulating in Africa. The sensitivity was determined by testing serial dilutions of virus and RNA standard. Intra- and inter-assay coefficients of variation were evaluated by 10 reactions in a same and in different assays, respectively. The assay provides high analytical specificity for USUV and detection limits of 1.2pfu/reaction for virus dilutions in L-15 medium or human serum and 60 copies/reaction for the RNA standard. The assay needs to be evaluated in a clinical context and integrated in standard diagnosis of flaviviral diseases