Genetic tracing of the illegal trade of the white-bellied pangolin (Phataginus tricuspis) in western Central Africa.

The white-bellied pangolin is subject to intense trafficking, feeding both local and international trade networks. In order to assess its population genetics and trace its domestic trade, we genotyped 562 pangolins from local to large bushmeat markets in western central Africa. We show that the two lineages described from the study region (WCA and Gab) were overlapping in ranges, with limited introgression in southern Cameroon. There was a lack of genetic differentiation across WCA and a significant signature of isolation-by-distance possibly due to unsuspected dispersal capacities involving a Wahlund effect. We detected a c. 74.1-82.5% decline in the effective population size of WCA during the Middle Holocene. Private allele frequency tracing approach indicated up to 600 km sourcing distance by large urban markets from Cameroon, including Equatorial Guinea. The 20 species-specific microsatellite loci provided individual-level genotyping resolution and should be considered as valuable resources for future forensic applications. Because admixture was detected between lineages, we recommend a multi-locus approach for tracing the pangolin trade. The Yaoundé market was the main hub of the trade in the region, and thus should receive specific monitoring to mitigate pangolins' domestic trafficking. Our study also highlighted the weak implementation of CITES regulations at European borders.

Mutualistic interactions between ants and fungi: A review.

The large amount of dead plant biomass caused by the final extinction events triggered a fungi proliferation that mostly differentiated into saprophytes degrading organic matter; others became parasites, predators, likely commensals, and mutualists. Among the last, many have relationships with ants, the most emblematic seen in the Neotropical myrmicine Attina that cultivate Basidiomycota for food. Among them, leaf-cutting, fungus-growing species illustrate an ecological innovation because they grow fungal gardens from fresh plant material rather than arthropod frass and plant debris. Myrmecophytes shelter "plant-ants" in hollow structures, the domatia, whose inner walls are lined with thin-walled Ascomycota hyphae that, in certain cases, are eaten by the ants, showing a form of convergence. Typically, these Ascomycota have antibacterial properties illustrating cases of farming for protection. Ant gardens, or mutualistic associations between certain ant species and epiphytes, shelter endophytic fungi that promote the growth of the epiphytes. Because the cell walls of certain Ascomycota hyphae remain sturdy after the death of the mycelium, they form resistant fibers used by ants to reinforce their constructions (e.g., galleries, shelters for tended hemipterans, and carton nests). Thus, we saw cases of "true" fungal agriculture involving planting, cultivating, and harvesting Basidiomycota for food with Attina. A convergence with "plant-ants" feeding on Ascomycota whose antibacterial activity is generally exploited (i.e., farming for protection). The growth of epiphytes was promoted by endophytic fungi in ant gardens. Finally, farming for structural materials occurred with, in one case, a leaf-cutting, fungus-growing ant using Ascomycota fibers to reinforce its nests.

Venomics survey of six myrmicine ants provides insights into the molecular and structural diversity of their peptide toxins.

Among ants, Myrmicinae represents the most speciose subfamily. The venom composition previously described for these social insects is extremely variable, with alkaloids predominant in some genera while, conversely, proteomics studies have revealed that some myrmicine ant venoms are peptide-rich. Using integrated transcriptomic and proteomic approaches, we characterized the venom peptidomes of six ants belonging to the different tribes of Myrmicinae. We identified a total of 79 myrmicitoxins precursors which can be classified into 38 peptide families according to their mature sequences. Myrmicine ant venom peptidomes showed heterogeneous compositions, with linear and disulfide-bonded monomers as well as dimeric toxins. Several peptide families were exclusive to a single venom whereas some were retrieved in multiple species. A hierarchical clustering analysis of precursor signal sequences led us to divide the myrmicitoxins precursors into eight families, including some that have already been described in other aculeate hymenoptera such as secapin-like peptides and voltage-gated sodium channel (NaV) toxins. Evolutionary and structural analyses of two representatives of these families highlighted variation and conserved patterns that might be crucial to explain myrmicine venom peptide functional adaptations to biological targets.

Description of Heteracris lecoqi Yetchom-Fondjo amp; Kekeunou sp. nov. and Heteracris hannai Wandji amp; Kekeunou sp. nov. and redescription of Heteracris guineensis (Krauss, 1890) (Orthoptera: Acrididae) with comments on its ecology in the Southern part of Cameroon.

The specimens studied were collected with sweep net and pitfall in the forests, agro-forests, herbaceous fallows, and crop fields of 14 localities in the southern part of Cameroon, from August 2015 to February 2018. The results show that Heteracris hannai sp. nov. and Heteracris lecoqi sp. nov. differ from previously known species of the same genus by the pattern of coloration and the details or shape of the phallic complex. H. lecoqi sp. nov. is characterized by light brownish body; male cercus with flattened, downcurved and obtuse apex; lophus strongly curved; interlophal space with V shape; apodeme of cingulum bow, convergent, with V-shape; valve of cingulum in lateral view longer than apical valve of penis; ramus in ventral view joined. H. hannai sp. nov. is distinct to other species by brown to grey body; male cercus with apex rounded, curved inside; lophus curved; interlophal space with U-shape; apodeme of cingulum thick, slightly parallel, with U shape; valve of cingulum in lateral view hardly longer than apical valve of penis; ramus in ventral view opened. The characteristics of H. guineensis are closer to H. hannai sp. nov. than to H. lecoqi sp. nov. H. lecoqi sp. nov. was collected only in the fallows while H. hannai sp. nov. was collected in the forests, agro-forests, fallows, and crop fields. Compared to H. guineensis, both new species are scarce in the natural vegetation and their distribution area is limited to two and four localities respectively for H. lecoqi sp. nov. and H. hannai sp. nov. All these three grasshopper's species were recorded as accidental species in all types of vegetation.

Development and characterization of 20 polymorphic microsatellite markers for the white-bellied pangolin Phataginus tricuspis (Mammalia, Pholidota).

Pangolins, or scaly anteaters, have recently been flagshiped as one of the most illegally traded mammals, and as a corollary, as potential intermediate hosts at the origin of the COVID-19 pandemic. In order to improve the traceability of their trade, we developed 20 polymorphic microsatellite loci for the white-bellied pangolin (Phataginus tricuspis), the species most frequently found on African bushmeat markets. We genotyped 24 white-bellied pangolins from the Douala market, Cameroon, originating from the Ebo forest c. 75 km north-east of Douala. The number of alleles per locus ranged from 4 to 12 (mean = 6.95), and mean observed and expected heterozygosities were 0.592 (0.208-0.875) and 0.671 (0.469-0.836), respectively. Genetic diversity was higher than that cross-estimated from microsatellite loci developed for other species of pangolins. Two loci deviated from Hardy-Weinberg equilibrium and two loci showed linkage disequilibrium. Genetic variance (PCoA) was increased with the addition of 13 pangolins of unknown origin, possibly suggesting that the Douala market is fed from differentiated source populations of white-bellied pangolins. Each of the 37 individuals had a unique multilocus genotype. The unbiased probability of identity (uPI) and the probability of identity among siblings (PIsibs) were both very low (uPI = 8.443 e-21; PIsibs = 1.011 e-07). Only five microsatellite loci were needed to reach the conservative value of PIsibs < 0.01, overall indicating a powerful discriminating power of our combined loci. These 20 newly developed microsatellite loci might prove useful in tracing the local-to-global trade of the white-bellied pangolin, and will hopefully contribute to the DNA-assisted implementation of future conservation strategies at reasonable costs.

Geographic distribution and abundance of the Afrotropical subterranean scale insect Stictococcus vayssierei (Hemiptera: Stictococcidae), a pest of root and tuber crops in the Congo basin.

Stictococcus vayssierei is a major pest of root and tuber crops in central Africa. However, data on its ecology are lacking. Here we provide an updated estimate of its distribution with the aim of facilitating the sustainable control of its populations. Surveys conducted in nine countries encompassing 13 ecological regions around the Congo basin showed that African root and tuber scale was present in Cameroon, Central African Republic, Congo, Democratic Republic of Congo, Equatorial Guinea, Gabon and Uganda. It was not found on the sites surveyed in Chad and Nigeria. The pest occurred in the forest and the forest-savannah mosaic as well as in the savannah where it was never recorded before. However, prevalence was higher in the forest (43.1%) where cassava was the most infested crop, compared to the savannah (9.2%) where aroids (cocoyam and taro) were the most infested crops. In the forest habitat, the pest was prevalent in all but two ecological regions: the Congolian swamp forests and the Southern Congolian forest-savanna mosaic. In the savannah habitat, it was restricted to the moist savannah highlands and absent from dry savannahs. The scale was not observed below 277 m asl. Where present, the scale was frequently (87.1% of the sites) attended by the ant Anoplolepis tenella. High densities (>1000 scales per plant) were recorded along the Cameroon-Gabon border. Good regulatory measures within and between countries are required to control the exchange of plant materials and limit its spread. The study provides information for niche modeling and risk mapping.

The Peptide Venom Composition of the Fierce Stinging Ant Tetraponera aethiops (Formicidae: Pseudomyrmecinae).

In the mutualisms involving certain pseudomyrmicine ants and different myrmecophytes (i.e., plants sheltering colonies of specialized "plant-ant" species in hollow structures), the ant venom contributes to the host plant biotic defenses by inducing the rapid paralysis of defoliating insects and causing intense pain to browsing mammals. Using integrated transcriptomic and proteomic approaches, we identified the venom peptidome of the plant-ant Tetraponera aethiops (Pseudomyrmecinae). The transcriptomic analysis of its venom glands revealed that 40% of the expressed contigs encoded only seven peptide precursors related to the ant venom peptides from the A-superfamily. Among the 12 peptide masses detected by liquid chromatography-mass spectrometry (LC-MS), nine mature peptide sequences were characterized and confirmed through proteomic analysis. These venom peptides, called pseudomyrmecitoxins (PSDTX), share amino acid sequence identities with myrmeciitoxins known for their dual offensive and defensive functions on both insects and mammals. Furthermore, we demonstrated through reduction/alkylation of the crude venom that four PSDTXs were homo- and heterodimeric. Thus, we provide the first insights into the defensive venom composition of the ant genus Tetraponera indicative of a streamlined peptidome.

Correction: Spodoptera frugiperda Smith (Lepidoptera: Noctuidae) in Cameroon: Case study on its distribution, damage, pesticide use, genetic differentiation and host plants.

[This corrects the article DOI: 10.1371/journal.pone.0215749.].

Spodoptera frugiperda Smith (Lepidoptera: Noctuidae) in Cameroon: Case study on its distribution, damage, pesticide use, genetic differentiation and host plants.

Maize farmers in sub-Saharan Africa recently experienced unusual damage in their farms, attributed to the fall armyworm (FAW) Spodoptera frugiperda (J. E. Smith). This pest was first recorded in Africa in 2016, but detailed information on its distribution and damage and farmer's response in invaded areas are largely lacking. In this study, we determined FAW distribution, genetic diversity, host plants, crop damage, and farmers' responses. S. frugiperda was recorded in the 10 regions of Cameroon. Average percentage of infested plants and damage severity (on a scale of 1 to 5) were lowest-20.7 ± 7.4% and 2.1 ± 0.1 respectively-in the Sahelian regions and greatest-69.0 ± 4.3% and 3.1 ± 0.1 respectively-in the Western Highlands. Altitude did not influence FAW incidence and severity and its larvae infrequently co-occurred with maize stemborers on the same plants, suggesting possible direct and/or indirect competition between the two groups of maize pests. In response to this new threat to maize production, farmers have opted for the application of synthetic pesticides. Although our experiments were not designed to determine pesticide efficacy, as parameters such as time since application were not considered, our observations suggest lack of a drastic effect on S. frugiperda infestations on maize. There were two haplotypes of FAW co-occurring in Cameroon corresponding to the rice and corn strains and separated by 1.7% sequence divergence, which does not support the existence of cryptic species. S. frugiperda larvae were also recorded on Sorghum bicolor (L.) Moench (10.6%), Solanum tuberosum L. (2.8%), Ipomoea batatas (L.) Lam. (1.9%), Saccharum officinarum L (0.8%), Phaseolus vulgaris L. (0.4%) and Gossypium hirsutum L. (1.9%). This study show that two strains are present in all agroecological zones in Cameroon, and probably in neighboring countries of central Africa sharing the same agroecologies. Management options should therefore consider the use of specific natural enemies and an informed decision of intervention based on strain capture and damage threshold, to avoid pesticide resistance that may arise from inadequate use of chemicals. Further studies should also be undertaken to assess the response of the two S. frugiperda strains to biopesticides and botanical insecticides.

Thermotolerance adaptation to human-modified habitats occurs in the native range of the invasive ant Wasmannia auropunctata before long-distance dispersal.

Key evolutionary events associated with invasion success are traditionally thought to occur in the introduced, rather than the native range of species. In the invasive ant Wasmannia auropunctata, however, a shift in reproductive system has been demonstrated within the native range, from the sexual non-dominant populations of natural habitats to the clonal dominant populations of human-modified habitats. Because abiotic conditions of human- modified habitats are hotter and dryer, we performed lab experiments on workers from a set of native and introduced populations, to investigate whether these ecological and genetic transitions were accompanied by a change in thermotolerance and whether such changes occurred before establishment in the introduced range. Thermotolerance levels were higher in native populations from human-modified habitats than in native populations from natural habitats, but were similar in native and introduced populations from human-modified habitats. Differences in thermotolerance could not be accounted for by differences in body size. A scenario based on local adaptation in the native range before introduction in remote areas represents the most parsimonious hypothesis to account for the observed phenotypic pattern. These findings highlight the importance of human land use in explaining major contemporary evolutionary changes.

Genetic structure, nestmate recognition and behaviour of two cryptic species of the invasive big-headed ant Pheidole megacephala.

BACKGROUND: Biological invasions are recognized as a major cause of biodiversity decline and have considerable impact on the economy and human health. The African big-headed ant Pheidole megacephala is considered one of the world's most harmful invasive species. METHODOLOGY/PRINCIPAL FINDINGS: To better understand its ecological and demographic features, we combined behavioural (aggression tests), chemical (quantitative and qualitative analyses of cuticular lipids) and genetic (mitochondrial divergence and polymorphism of DNA microsatellite markers) data obtained for eight populations in Cameroon. Molecular data revealed two cryptic species of P. megacephala, one inhabiting urban areas and the other rainforests. Urban populations belong to the same phylogenetic group than those introduced in Australia and in other parts of the world. Behavioural analyses show that the eight populations sampled make up four mutually aggressive supercolonies. The maximum distance between nests from the same supercolony was 49 km and the closest distance between two nests belonging to two different supercolonies was 46 m. The genetic data and chemical analyses confirmed the behavioural tests as all of the nests were correctly assigned to their supercolony. Genetic diversity appears significantly greater in Africa than in introduced populations in Australia; by contrast, urban and Australian populations are characterized by a higher chemical diversity than rainforest ones. CONCLUSIONS/SIGNIFICANCE: Overall, our study shows that populations of P. megacephala in Cameroon adopt a unicolonial social structure, like invasive populations in Australia. However, the size of the supercolonies appears several orders of magnitude smaller in Africa. This implies competition between African supercolonies and explains why they persist over evolutionary time scales.

Initial behavior in colony fragments of an introduced population of the invasive ant Wasmannia auropunctata.

We investigated in the laboratory the initial behavior of propagules of the invasive ant Wasmannia auropunctata in Cameroon where it has been introduced. Both workers and queens at first feigned death (thanatosis), and then the workers slowly moved around the experimental arena; the queens did the same about 10seconds later. Each queen antennated selected workers that then aggregated together by grasping the hind leg of another ant with their mandibles. When encountering the queen again, the lead worker climbed up the queen's hind leg and onto her back, followed by some other individuals. The remaining workers followed the queen to a location in the experimental arena. When brood was present, the workers transferred it to this location. Orphaned workers did not aggregate, but gathered the brood together and took care of it. By permitting propagules to survive, these behaviors likely contribute to the success of W. auropunctata as an invader.

Bactrocera invadens (Diptera: Tephritidae), a new invasive fruit fly pest for the Afrotropical region: host plant range and distribution in West and Central Africa.

In 2003, the invasive fruit fly Bactrocera invadens Drew, Tsuruta & White (Diptera: Tephritidae) (Drew et al. 2005), of possible Sri Lankan origin, has been detected in the East and about 1 yr later in West Africa. In regular surveys in Benin and Cameroon covering 4 yr, samples from 117 plant species across 43 families have been obtained. Incubation of field-collected fruits demonstrate that in West and Central Africa (WCA) B. invadens is highly polyphagous, infesting wild and cultivated fruits of at least 46 species from 23 plant families with guava (Psidium spp.), mango (Mangifera spp.), and citrus (spp.), and the wild hosts tropical almond (Terminalia catappa L.), African wild mango (Irvingia gabonensis (Aubry-Lecomte) Baill.), and sheanut (Vitellaria paradoxa C.F.Gaertn.) showing the highest infestation index. B. invadens occurs in 22 countries of WCA with new records for Angola, Central African Republic, the Congo, DR Congo, Equatorial Guinea, Gabon, Gambia, Guinea Bissau, Mali, Mauritania, Niger, and Sierra Leone. Overall, the pest has spread across a North-South distance of ≍5,000 km representing a contiguous area of >8.3 million km(2) within WCA. B. invadens has adapted to a wide range of ecological and climatic conditions extending from low land rainforest to dry savanna. Because of its highly destructive and invasive potential, B. invadens poses a serious threat to horticulture in Africa if left uncontrolled. Moreover, the presence of this quarantine pest causes considerable restrictions on international trade of affected crops.

Worldwide invasion by the little fire ant: routes of introduction and eco-evolutionary pathways.

Biological invasions are generally thought to occur after human aided migration to a new range. However, human activities prior to migration may also play a role. We studied here the evolutionary genetics of introduced populations of the invasive ant Wasmannia auropunctata at a worldwide scale. Using microsatellite markers, we reconstructed the main routes of introduction of the species. We found three main routes of introduction, each of them strongly associated to human history and trading routes. We also demonstrate the overwhelming occurrence of male and female clonality in introduced populations of W. auropunctata, and suggest that this particular reproduction system is under selection in human-modified habitats. Together with previous researches focused on native populations, our results suggest that invasive clonal populations may have evolved within human modified habitats in the native range, and spread further from there. The evolutionarily most parsimonious scenario for the emergence of invasive populations of the little fire ant might thus be a two-step process. The W. auropunctata case illustrates the central role of humans in biological change, not only due to changes in migration patterns, but also in selective pressures over species.

The predatory behaviour of a tramp ant species in its native range.

Workers of the pest ant Paratrechina longicornis participate in a type of group hunting. Each individual forages with its long antennae wide open and moves quickly (6.3 cm/s) along an erratic path surrounded by nestmates behaving in the same way and within range of a recruiting pheromone. They detect prey by contact with successful workers singly capturing and retrieving small prey and seizing larger ones by an appendage. Then they recruit nestmates at short-range; all together they spread-eagle the prey and retrieve them whole.