Potential of Ilhéus virus to emerge.

Ilhéus virus (ILHV)(Flaviviridae:Orthoflavivirus) is an arthropod-borne virus (arbovirus) endemic to Central and South America and the Caribbean. First isolated in 1944, most of our knowledge derives from surveillance and seroprevalence studies. These efforts have detected ILHV in a broad range of mosquito and vertebrate species, including humans, but laboratory investigations of pathogenesis and vector competence have been lacking. Here, we develop an immune intact murine model with several ages and routes of administration. Our model closely recapitulates human neuroinvasive disease with ILHV strain- and mouse age-specific virulence, as well as a uniformly lethal Ifnar-/- A129 immunocompromised model. Replication kinetics in several vertebrate and invertebrate cell lines demonstrate that ILHV is capable of replicating to high titers in a wide variety of potential host and vector species. Lastly, vector competence studies provide strong evidence for efficient infection of and potential transmission by Aedes species mosquitoes, despite ILHV's phylogenetically clustering with Culex vectored flaviviruses, suggesting ILHV is poised for emergence in the neotropics.

Consequences of polyploidy and divergence as revealed by cytogenetic mapping of tandem repeats in African clawed frogs (Xenopus, Pipidae).

Repetitive elements have been identified in several amphibian genomes using whole genome sequencing, but few studies have used cytogenetic mapping to visualize these elements in this vertebrate group. Here, we used fluorescence in situ hybridization and genomic data to map the U1 and U2 small nuclear RNAs and histone H3 in six species of African clawed frog (genus Xenopus), including, from subgenus Silurana, the diploid Xenopus tropicalis and its close allotetraploid relative X. calcaratus and, from subgenus Xenopus, the allotetraploid species X. pygmaeus, X. allofraseri, X. laevis, and X. muelleri. Results allowed us to qualitatively evaluate the relative roles of polyploidization and divergence in the evolution of repetitive elements because our focal species include allotetraploid species derived from two independent polyploidization events - one that is relatively young that gave rise to X. calcaratus and another that is older that gave rise to the other (older) allotetraploids. Our results demonstrated conserved loci number and position of signals in the species from subgenus Silurana; allotetraploid X. calcaratus has twice as many signals as diploid X. tropicalis. However, the content of repeats varied among the other allotetraploid species. We detected almost same number of signals in X. muelleri as in X. calcaratus and same number of signals in X. pygmaeus, X. allofraseri, X. laevis as in the diploid X. tropicalis. Overall, these results are consistent with the proposal that allopolyploidization duplicated these tandem repeats and that variation in their copy number was accumulated over time through reduction and expansion in a subset of the older allopolyploids.

Divergent subgenome evolution in the allotetraploid frog Xenopus calcaratus.

Allopolyploid genomes are divided into compartments called subgenomes that are derived from lower ploidy ancestors. In African clawed frogs of the subgenus Xenopus (genus Xenopus), allotetraploid species have two subgenomes (L and S) with morphologically distinct homoeologous chromosomes. In allotetraploid species of the sister subgenus Silurana, independently evolved subgenomes also exist, but their cytogenetics has not been investigated in detail. We used a diverse suite of cytogenetic and molecular FISH techniques on an allotetraploid species in Silurana-Xenopus calcaratus-to explore evolutionary dynamics of chromosome morphology and rearrangements. We find that the subgenomes of X. calcaratus have distinctive characteristics, with a more conserved a-subgenome resembling the closely related genome of the diploid species X. tropicalis, and a more rapidly evolving b-subgenome having more pronounced changes in chromosome structure, including diverged heterochromatic blocks, repetitive sequences, and deletion of a nucleolar secondary constriction. Based on these cytogenetic differences, we propose a chromosome nomenclature for X. calcaratus that may apply to other allotetraploids in subgenus Silurana, depending on as yet unresolved details of their evolutionary origins. These findings highlight the potential for large-scale asymmetry in subgenome evolution following allopolyploidization.

Gene flow in phylogenomics: Sequence capture resolves species limits and biogeography of Afromontane forest endemic frogs from the Cameroon Highlands.

Puddle frogs of the Phrynobatrachus steindachneri species complex are a useful group for investigating speciation and phylogeography in Afromontane forests of the Cameroon Volcanic Line, western Central Africa. The species complex is represented by six morphologically relatively cryptic mitochondrial DNA lineages, only two of which are distinguished at the species level - southern P. jimzimkusi and Lake Oku endemic P. njiomock, leaving the remaining four lineages identified as 'P. steindachneri'. In this study, the six mtDNA lineages are subjected to genomic sequence capture analyses and morphological examination to delimit species and to study biogeography. The nuclear DNA data (387 loci; 571,936 aligned base pairs) distinguished all six mtDNA lineages, but the topological pattern and divergence depths supported only four main clades: P. jimzimkusi, P. njiomock, and only two divergent evolutionary lineages within the four 'P. steindachneri' mtDNA lineages. One of the two lineages is herein described as a new species, P. amieti sp. nov. Reticulate evolution (hybridization) was detected within the species complex with morphologically intermediate hybrid individuals placed between the parental species in phylogenomic analyses, forming a ladder-like phylogenetic pattern. The presence of hybrids is undesirable in standard phylogenetic analyses but is essential and beneficial in the network multispecies coalescent. This latter approach provided insight into the reticulate evolutionary history of these endemic frogs. Introgressions likely occurred during the Middle and Late Pleistocene climatic oscillations, due to the cyclic connections (likely dominating during cold glacials) and separations (during warm interglacials) of montane forests. The genomic phylogeographic pattern supports the separation of the southern (Mt. Manengouba to Mt. Oku) and northern mountains at the onset of the Pleistocene. Further subdivisions occurred in the Early Pleistocene, separating populations from the northernmost (Tchabal Mbabo, Gotel Mts.) and middle mountains (Mt. Mbam, Mt. Oku, Mambilla Plateau), as well as the microendemic lineage restricted to Lake Oku (Mt. Oku). This unique model system is highly threatened as all the species within the complex have exhibited severe population declines in the past decade, placing them on the brink of extinction. In addition, Mount Oku is identified to be of particular conservation importance because it harbors three species of this complex. We, therefore, urge for conservation actions in the Cameroon Highlands to preserve their diversity before it is too late.

Conserving the forgotten: New insights from a Central African biodiversity hotspot on the anthropogenic perception of nocturnal primates (Mammalia: Strepsirrhini).

The growing dependence of villagers on local forests (food, wood, etc.) makes the comparative assessment of the perceptions they have of the forest and its wildlife increasingly important for setting conservation priorities. While hunting and habitat loss are important threats to primates' existence worldwide, more attention has been focused on diurnal species, while little is known about their nocturnal counterparts. Strepsirrhini is a group of nocturnal primates with galago and potto as the only representatives on mainland Africa. To assess the perception of locals and their impacts on the conservation of these primates, questionnaires were administered to 79 household heads in four villages located in community forests around Mount Cameroon National Park. Amongst the respondents, over 90% admitted that these animals are eaten in their communities. Nocturnal primates were not only hunted for food, but also used in medicine and rituals and to make drums. However, the habit of eating nocturnal primates seems to be uncommon, as most respondents had not consumed any primate bushmeat in the last 5 years. The knowledge and support of wildlife conservation manifested by the villagers did not reflect the reality on the ground, as forest clearing for agriculture takes place regularly across the villages. Our findings suggest that bushmeat is not the main threat to nocturnal primates in this area, with habitat loss potentially representing a bigger problem for their existence in the near future.

Effects of disturbances by forest elephants on diversity of trees and insects in tropical rainforests on Mount Cameroon.

Natural disturbances are essential for tropical forests biodiversity. In the Afrotropics, megaherbivores have played a key role before their recent decline. Contrastingly to savanna elephants, forest elephants' impact on ecosystems remains poorly studied. Few decades ago, forests on Mount Cameroon were divided by lava flows, not being crossed by a local population of forest elephants until now. We assessed communities of trees, butterflies and two guilds of moths in the disturbed and undisturbed forests split by the longest lava flow. We surveyed 32 plots, recording 2025 trees of 97 species, and 7853 insects of 437 species. The disturbed forests differed in reduced tree density, height, and high canopy cover, and in increased DBH. Forest elephants' selective browsing and foraging also decreased tree species richness and altered their composition. The elephant disturbance increased butterfly species richness and had various effects on species richness and composition of the insect groups. These changes were likely caused by disturbance-driven alterations of habitats and species composition of trees. Moreover, the abandonment of forests by elephants led to local declines of range-restricted butterflies. The recent declines of forest elephants across the Afrotropics probably caused similar changes in forest biodiversity and should be reflected by conservation actions.

Precipitation and vegetation shape patterns of genomic and craniometric variation in the central African rodent Praomys misonnei.

Predicting species' capacity to respond to climate change is an essential first step in developing effective conservation strategies. However, conservation prioritization schemes rarely take evolutionary potential into account. Ecotones provide important opportunities for diversifying selection and may thus constitute reservoirs of standing variation, increasing the capacity for future adaptation. Here, we map patterns of environmentally associated genomic and craniometric variation in the central African rodent Praomys misonnei to identify areas with the greatest turnover in genomic composition. We also project patterns of environmentally associated genomic variation under future climate change scenarios to determine where populations may be under the greatest pressure to adapt. While precipitation gradients influence both genomic and craniometric variation, vegetation structure is also an important determinant of craniometric variation. Areas of elevated environmentally associated genomic and craniometric variation overlap with zones of rapid ecological transition underlining their importance as reservoirs of evolutionary potential. We also find that populations in the Sanaga river basin, central Cameroon and coastal Gabon are likely to be under the greatest pressure from climate change. Lastly, we make specific conservation recommendations on how to protect zones of high evolutionary potential and identify areas where populations may be the most susceptible to climate change.

First records of 31 species of butterflies and moths (Lepidoptera) in Cameroon, with remarks on their elevational ranges.

BACKGROUND: The biodiversity of West and Central Africa is understudied, including butterflies and moths (Lepidoptera). Cameroon, through its position in between few biogeographic regions and diversity of habitats, is an important hotspot of lepidopteran diversity. However, the country also ranks low when it comes to local biodiversity knowledge. During our long-term ecological projects in the Cameroonian part of the Gulf of Guinea Highlands, we collected rich material of butterflies and moths, including a number of interesting faunistic records. NEW INFORMATION: In this study, we report 31 species of butterflies and moths which have not yet been recorded in Cameroon. These species comprised eight new genera records for the country. In many cases, our records represented an important extension of the species' known distribution, including ten species whose distribution ranges extended into the Guinean biogeographic region. We also comment on the species' elevational distribution ranges on Mount Cameroon where most of our records originated. Additionally, we confirm the presence of a butterfly Telchinia encedena, after more than a century since its first and so far its only record in Cameroon.

Evolutionary history of the Cameroon radiation of puddle frogs (Phrynobatrachidae: Phrynobatrachus), with descriptions of two critically endangered new species from the northern Cameroon Volcanic Line.

The Cameroon Volcanic Line, a mountain chain located between West and Central Africa, is a region of numerous endemic diversifications, including of puddle frogs (Phrynobatrachus). This study reviews the phylogeny and taxonomy of puddle frogs of the "Cameroon radiation," which is a clade containing mainly montane but also at least three lowland species. Molecular data revealed a novel evolutionary lineage from high altitudes in the northern part of the mountains. Puddle frogs from the new, minute-sized (SVL < 20 mm) lineage are identified using molecular, morphological and acoustic data and described as two new species, Phrynobatrachus arcanus sp. nov. (Gotel Mountains, Cameroon-Nigeria) and P. mbabo sp. nov. (Tchabal Mbabo, Cameroon). The tadpole of the first species is also described. Phylogenetic analyses placed the new lineage to the proximity of the recently described lowland small-sized taxa (P. horsti, P. ruthbeateae). Based on the inferred phylogeny, we propose five species groups within the Cameroon radiation: P. arcanus, P. chukuchuku, P. ruthbeateae, P. steindachneri, and P. werneri. The taxonomically enigmatic P. hylaios is proposed to be a member of the P. ruthbeateae species group. The basal radiation evolved during the late Miocene with subsequent diversifications occurring during the Pliocene, while closely related terminal taxa originated during the Pleistocene. We recommend that the newly described species are categorized as Critically Endangered due to their limited ranges and because recent surveys did not identify any individuals at the type localities. This further supports the need for conservation interventions in the mountains of Cameroon and Nigeria.

The role of ultraviolet reflectance and pattern in the pollination system of Hypoxis camerooniana (Hypoxidaceae).

Apart from floral morphology and colours perceived by the human eye, ultraviolet (UV) reflectance acts as an important visual advertisement of numerous flowering plant species for pollinators. However, the effect of UV signalling on attracting pollinators of particular plant species is still insufficiently studied, especially in the Afrotropics. Therefore, we studied the pollination system of Hypoxis camerooniana in montane grasslands of Mount Cameroon, West/Central Africa. We focused mainly on the effects of the flowers' UV reflectance on its visitors. We experimentally removed UV reflection from petals either completely or partially. Thereafter, flower visitors were recorded and pistils were collected post-flowering to quantify germinated pollen tubes per treatments. The most important visitors were bees, followed by flies. Due to their contacts with reproductive organs bees are considered as the primary pollinators. Visitation rates were lower when UV reflectance was completely removed, whereas the decrease of frequency on half-treated flowers did not differ significantly from control treatments. The complete removal of UV also affected bees' landing behaviour, but not that of flies. We showed that the presence of UV reflectance is more important than UV pattern for bees visiting flowers of H. camerooniana. We hypothesize that exploiting all flowers irrespective of their pattern can be more efficient for pollinators in the open grasslands of high altitudes to spot these relatively scarce flowers by their UV reflectance. Furthermore, we highlight the necessity of both experimental and natural controls in similar studies to control for additional effects of the used UV manipulations. Many plants advertise their flowers with UV reflectance visible to their insect visitors. By manipulating the UV reflectance and pattern of Hypoxis camerooniana in the Afromontane grasslands of Mount Cameroon, we have shown how crucial it is for the predominant visitor, bees. Both bees' preferences for flowers and their behaviour during visits are influenced by changes in UV reflectance. However, the presence of some UV signal is more important than the specific pattern. Especially in montane grasslands with higher UV irradiation, the UV floral colours are important for recognition of flowers by potential pollinators.

Distribution modeling and lineage diversity of the chytrid fungus Batrachochytrium dendrobatidis (Bd) in a central African amphibian hotspot.

The amphibian disease chytridiomycosis in amphibians is caused by the chytrid fungus Batrachochytrium dendrobatidis (Bd) and has resulted in dramatic declines and extinctions of amphibian populations worldwide. A hypervirulent, globally-dispersed pandemic lineage (Bd-GPL) is thought to be largely responsible for population declines and extinctions, although numerous endemic lineages have also been found. Recent reports of amphibian declines have been linked to the emergence of Bd in Cameroon, a major hotspot of African amphibian diversity. However, it is not known whether Bd-GPL or other lineages have been found in this region. This study therefore aims to examine Bd lineage diversity in the region and predict the distribution of this pathogen under current and future climate conditions using data from this study and from historical records. Almost 15% (52/360) of individuals tested positive for Bd using a standard quantitative PCR diagnostic. Infected amphibians were found at all eight sites sampled in this study. Species distribution models generated in BIOMOD2 indicate that areas with highest predicted environmental suitability occur in the Cameroon highlands and several protected areas throughout the country. These areas of high environmental suitability for Bd are projected to shift or decrease in size under future climate change. However, montane regions with high amphibian diversity are predicted to remain highly suitable. Phylogenetic analysis of the ITS sequences obtained from a set of positive Bd samples indicate that most fall within the Bd-GPL lineage while the remainder group with isolates from either Brazil or South Korea. Although more in depth phylogenetic analyses are needed, identification of Bd-GPL lineages in areas of high amphibian diversity emphasizes the need to continue to monitor for Bd and develop appropriate conservation strategies to prevent its further spread.

Flying between raindrops: Strong seasonal turnover of several Lepidoptera groups in lowland rainforests of Mount Cameroon.

Although seasonality in the tropics is often less pronounced than in temperate areas, tropical ecosystems show seasonal dynamics as well. Nevertheless, individual tropical insects' phenological patterns are still poorly understood, especially in the Afrotropics. To fill this gap, we investigated biodiversity patterns of Lepidoptera communities at three rainforest localities in the foothills of Mount Cameroon, West Africa, one of the wettest places in the world. Our multitaxa approach covered six lepidopteran groups (fruit-feeding butterflies and moths, the families Sphingidae, Saturniidae, and Eupterotidae, and the subfamily Arctiinae of Erebidae) with diverse life strategies. We sampled adults of the focal groups in three distinct seasons. Our sampling included standardized bait trapping (80 traps exposed for 10 days per locality and season) and attraction by light (six full nights per locality and season). Altogether, our dataset comprised 20,576 specimens belonging to 559 (morpho)species of the focal groups. The biodiversity of Lepidoptera generally increased in the high-dry season, and either increased (fruit-feeding moths, Arctiinae, Saturniidae) or decreased (butterflies, Sphingidae) in the transition to the wet season in particular groups. Simultaneously, we revealed a strong species turnover of fruit-feeding Lepidoptera and Arctiinae among the seasons, indicating relatively high specialization of these communities for particular seasons. Such temporal specialization can make the local communities of butterflies and moths especially sensitive to the expected seasonal perturbations caused by the global change. Because of the key role of Lepidoptera across trophic levels, such changes in their communities could strengthen this impact on entire tropical ecosystems.

Determination of the predictive factors of long-lasting insecticide-treated net ownership and utilisation in the Bamenda Health District of Cameroon.

BACKGROUND: Malaria is a serious health concern in Africa. In Cameroon, an endemic country where malaria remains a major public health problem, several control measures have been put in place among which the use of insecticide-treated bednets (LLINs/ITNs) is considered one of the core vector control strategies. However, the greatest challenges include ownership and utilisation by individuals and households. Factors such as age, marital status, gender, education and occupation of the household head, household size, knowledge of bednets, socioeconomic status, and environmental factors have been suggested to have an impact on bednet ownership and utilisation in different settings. The present study sought to determine bednet ownership and utilisation rates and to assess the impact of predictive factors on bednet ownership and use in the Bamenda Health District (BHD) of Cameroon. METHODS: A cross-sectional study involving 384 households was conducted in six health areas in the BHD. A structured and semi-structured questionnaire was used to collect data on demographic and household characteristics as well as information on their bednet ownership and utilisation. Descriptive statistics, bivariate and multivariate logistic regression analysis were performed. RESULTS: Frequency of bednet ownership was relatively high (63.5%) with LLINs being most abundant (91.9%); the majority of households (87.7%) obtained their bednets during the 2011 free distribution campaign. Utilisation was relatively high (69.3%), with negligence (29.3%) and heat discomfort (26.7%) accounting most for non-usage of bednets. Children less than 5 years (63%) and pregnant women (60%) most often used these nets. Households headed by a married couple, those with older household heads, household with smaller size (5-12 persons), and knowledge of bednets (good knowledge) had positive impacts on bednet ownership (p < 0.05). The gender of the household head (males), their educational level, environmental conditions (presence of suitable mosquito breeding sites), bednet number in households (greater number of bednets) and the prioritised groups (children < 5 and pregnant women) had positive impacts on bednet utilisation in households (p < 0.05). There was a negative association between bednet ownership and utilisation by households as bednet ownership was high and utilisation of these nets was low. Marital status and age of household head, household size, and knowledge of bednets had impacts on bednet ownership while gender and educational level of the household head, environmental suitability, the number of bednets and the two prioritised groups had an impact on bednet usage. CONCLUSION: These factors may be relevant for policy makers and in decision making for the intensification of campaign strategies to ensure more effective subsequent distribution campaigns in the BHD and beyond.

Physical integrity and residual bio-efficacy of used LLINs in three cities of the South-West region of Cameroon 4 years after the first national mass-distribution campaign.

BACKGROUND: Long-lasting insecticidal nets (LLINs) are effective tools at reducing malaria transmission. In Cameroon, following the first national mass distribution campaign in 2011, there has been no follow up on the efficacy of LLINs distributed. The aim of this report is to assess the physical integrity and insecticidal potency of LLINs distributed in three cities with contrasting socio economic status (SES) and to evaluate the use and care for bed nets owned by individuals. METHODS: The study was conducted in Limbe, Tiko and Buea three localities in the Fako division of the South West Region of Cameroon. Tiko had the highest SES based on the type of building materials used, followed respectively by Limbe and Buea. The use and care for bed nets was assessed using a questionnaire, the physical state of bed nets evaluated following WHOPES recommendation for determining size of holes and the residual insecticidal activity of retrieved bed nets determined through a cone bioassay using susceptible strains of mosquitoes. RESULTS: Of the 241 households visited in Limbe (n = 81), Tiko (n = 80) and Buea (n = 80), 186 (77.2%) had LLINs, with no significant variations from the selected study locations. However, bed net coverage was not meeting World Health Organization standards (p < 0.0001). Six different brands of LLINs were recorded, and the majority were those provided by the NMCP through the 2011 national mass campaign or antenatal care programme. Based on the calculation of the Proportionate Hole Index (PHI) as indicator of physical integrity of nets, the proportion of nets classified as serviceable (versus too torn) differed statistically according to locations (p value = 0.04), with 63.8% in Tiko, 50% in Limbe and 47% in Buea. Of the 20 nets tested for their efficacy against susceptible strains of mosquito, 42.6% (3 nets) were optimally effective in Tiko, 57.4% (4), 16.7% (1) and 14.3% (1) were minimally effective in Tiko, Buea and Limbe respectively. Finally; 85.7% (6) and 83.3% (5) were not effective in Limbe and Buea. CONCLUSIONS: These findings pinpoint the need for more frequent replacement of LLINs especially for people with low SES and also the need for the promotion of good practices on the maintenance and washing of nets.

The Effect of Long Lasting Insecticide Bed Net Use on Malaria Prevalence in the Tombel Health District, South West Region-Cameroon.

Malaria remains a major public health problem in Africa, and its prevalence in Cameroon stands at 29%. Long Lasting Insecticide Nets (LLINs) were distributed in 2011 to reduce malaria mortality and morbidity; however, assessment of this intervention is scanty. The present study in the Tombel health district (THD) investigated the impact of this distribution on malaria prevalence. A total of 31,657 hospital records from 3 health facilities in 3 health areas for 2010-2013 were examined. Records for 2010 and 2011 provided predistribution baseline data, while those of 2012 and 2013 represented postdistribution data. 8,679 (27.4%) patients were positive for malaria. Children below 5 years had the highest prevalence (40.7%). The number of confirmed cases was highest from June to August (peak rainy season). Malaria prevalence was higher in males (25.3%) than in females (23.2%). Malaria prevalence increased in THD from 26.7% in 2010 to 30.7% in 2011 but dropped to 22.7% in 2012 and then increased in 2013 to 29.5%. There was an overall drop in the total number of confirmed malaria cases in 2012; this decrease was significant in Ebonji (p < 0.001) and Nyasoso (p < 0.015) health areas. The distribution of LLINs led to a short lived reduction in malaria prevalence in THD. LLIN distribution and other control activities should be reinforced to keep malaria prevalence low especially among the 0-5-year group.

A newly isolated reovirus has the simplest genomic and structural organization of any reovirus.

UNLABELLED: A total of 2,691 mosquitoes representing 17 species was collected from eight locations in southwest Cameroon and screened for pathogenic viruses. Ten isolates of a novel reovirus (genus Dinovernavirus) were detected by culturing mosquito pools on Aedes albopictus (C6/36) cell cultures. A virus that caused overt cytopathic effects was isolated, but it did not infect vertebrate cells or produce detectable disease in infant mice after intracerebral inoculation. The virus, tentatively designated Fako virus (FAKV), represents the first 9-segment, double-stranded RNA (dsRNA) virus to be isolated in nature. FAKV appears to have a broad mosquito host range, and its detection in male specimens suggests mosquito-to-mosquito transmission in nature. The structure of the T=1 FAKV virion, determined to subnanometer resolution by cryoelectron microscopy (cryo-EM), showed only four proteins per icosahedral asymmetric unit: a dimer of the major capsid protein, one turret protein, and one clamp protein. While all other turreted reoviruses of known structures have at least two copies of the clamp protein per asymmetric unit, FAKV's clamp protein bound at only one conformer of the major capsid protein. The FAKV capsid architecture and genome organization represent the most simplified reovirus described to date, and phylogenetic analysis suggests that it arose from a more complex ancestor by serial loss-of-function events. IMPORTANCE: We describe the detection, genetic, phenotypic, and structural characteristics of a novel Dinovernavirus species isolated from mosquitoes collected in Cameroon. The virus, tentatively designated Fako virus (FAKV), is related to both single-shelled and partially double-shelled viruses. The only other described virus in this genus was isolated from cultured mosquito cells. It was previously unclear whether the phenotypic characteristics of that virus were reflective of this genus in nature or were altered during serial passaging in the chronically infected cell line. FAKV is a naturally occurring single-shelled reovirus with a unique virion architecture that lacks several key structural elements thought to stabilize a single-shelled reovirus virion, suggesting what may be the minimal number of proteins needed to form a viable reovirus particle. FAKV evolved from more complex ancestors by losing a genome segment and several virion proteins.

Individual variation in thermal performance curves: swimming burst speed and jumping endurance in wild-caught tropical clawed frogs.

The importance of studying individual variation in locomotor performance has long been recognized as it may determine the ability of an organism to escape from predators, catch prey or disperse. In ectotherms, locomotor performance is highly influenced by ambient temperature (Ta), yet several studies have showed that individual differences are usually retained across a Ta gradient. Less is known, however, about individual differences in thermal sensitivity of performance, despite the fact that it could represent adaptive sources of phenotypic variation and/or additional substrate for selection to act upon. We quantified swimming and jumping performance in 18 wild-caught tropical clawed frogs (Xenopus tropicalis) across a Ta gradient. Maximum swimming velocity and acceleration were not repeatable and individuals did not differ in how their swimming performance varied across Ta. By contrast, time and distance jumped until exhaustion were repeatable across the Ta gradient, indicating that individuals that perform best at a given Ta also perform best at another Ta. Moreover, thermal sensitivity of jumping endurance significantly differed among individuals, with individuals of high performance at low Ta displaying the highest sensitivity to Ta. Individual differences in terrestrial performance increased with decreasing Ta, which is opposite to results obtained in lizards at the inter-specific and among-individual levels. To verify the generality of these patterns, we need more studies on individual variation in thermal reaction norms for locomotor performance in lizards and frogs.

Genetic and phenotypic characterization of sylvatic dengue virus type 2 strains.

The four serotypes of endemic dengue viruses (DENV) circulate between humans and peridomestic Aedes mosquitoes. At present endemic DENV infect 100 million people per year, and a third of the global population is at risk. In contrast, sylvatic DENV strains are maintained in a transmission cycle between nonhuman primates and sylvatic Aedes species, and are evolutionarily and ecologically distinct from endemic DENV strains. Phylogenetic analyses place sylvatic strains basal to each of the endemic serotypes, supporting the hypothesis that each of the endemic DENV serotypes emerged independently from sylvatic ancestors. We utilized complete genome analyses of both sylvatic and endemic DENV serotype 2 (DENV-2) to expand our understanding of their genetic relationships. A high degree of conservation was observed in both the 5'- and 3'-untranslated genome regions, whereas considerable differences at the nucleotide and amino acid levels were observed within the open reading frame. Additionally, replication of the two genotypes was compared in cultured cells, where endemic DENV strains produced a significantly higher output of progeny in human liver cells, but not in monkey kidney or mosquito cells. Understanding the genetic relationships and phenotypic differences between endemic and sylvatic DENV genotypes may provide valuable insight into DENV emergence and guide monitoring of future outbreaks.

Evolutionary processes among sylvatic dengue type 2 viruses.

Sylvatic dengue viruses (DENV) are transmitted in an enzootic cycle between nonhuman primates and arboreal Aedes mosquitoes in Southeast Asia and West Africa. Although previous analyses have revealed the evolutionary processes among endemic (human) DENV, little is known about viral evolution in the sylvatic cycle. Through an analysis of 14 complete coding regions of sylvatic Dengue type 2 virus sampled over a 33-year period, we show that both the rate of evolutionary change and the pattern of natural selection are similar among endemic and sylvatic DENV, although the latter have a uniquely high frequency of positive selection in the NS4B protein gene. Our findings support a recent cross-species transmission event and suggest the possibility of future DENV reemergence from the sylvatic cycle.

Potential of ancestral sylvatic dengue-2 viruses to re-emerge.

Dengue viruses (DENV) are the most important arboviral pathogens in tropical and subtropical regions throughout the world. DENV transmission includes both a sylvatic, enzootic cycle between nonhuman primates and arboreal mosquitoes of the genus Aedes, and an urban, endemic/epidemic cycle between Aedes aegypti, a mosquito with larval development in peridomestic water containers, and human reservoir hosts. All 4 serotypes of endemic DENV evolved independently from ancestral sylvatic viruses and have become both ecologically and evolutionarily distinct; this process may have involved adaptation to (i) peridomestic mosquito vectors and/or (ii) human reservoir hosts. To test the latter hypothesis, we assessed the ability of sylvatic and endemic DENV-2 strains, representing major genotypes from Southeast Asia, West Africa and the Americas, to replicate in two surrogate human model hosts: monocyte-derived, human dendritic cells (moDCs), and mice engrafted with human hepatoma cells. Although the various DENV-2 strains showed significant inter-strain variation in mean replication titers in both models, no overall difference between sylvatic and endemic strains was detected in either model. Our findings suggest that emergence of endemic DENV strains from ancestral sylvatic strains may not have required adaptation to replicate more efficiently in human reservoir hosts, implying that the potential for re-emergence of sylvatic dengue strains into the endemic cycle is high. The shared replication profiles of the American endemic and sylvatic strains suggest that American strains have maintained or regained the ancestral phenotype.