Evaluation of identification methods for cryptic Bactrocera dorsalis (Diptera: Tephritidae) specimens: combining morphological and molecular techniques.

The potential for population genomics to elucidate invasion pathways of a species is limited by taxonomic identification issues. The Oriental fruit fly pest, Bactrocera dorsalis (Hendel) belongs to a complex in which several sympatric species are attracted to the same lure used in trapping and are morphologically cryptic and/or reported to hybridize. In this study, we evaluated the taxonomic ambiguity between B. dorsalis and 2 major cryptic species, based on morphological expertise and 289 target specimens sampled across the whole distribution range. Specimens were then subjected to DNA sequence analyses of the COI mitochondrial barcode and the EIF3L nuclear marker to evaluate the potential for molecular identification, in particular for specimens for which morphological identification was inconclusive. To this aim, we produced reference datasets with DNA sequences from target specimens whose morphological identification was unambiguous, which we complemented with 56 new DNA sequences from closest relatives and 76 published and curated DNA sequences of different species in the complex. After the necessary morphological observation, about 3.5% of the target dataset and 47.6% of the specimens from Southeast Asian islands displayed ambiguous character states shared with B. carambolae and/or B. occipitalis. Critical interpretation of DNA sequence data solved morphological ambiguities only when combining both mitochondrial and nuclear markers. COI discriminated B. dorsalis from 5 species; EIF3L and ITS from another species. We recommend this procedure to ensure correct identification of B. dorsalis specimens in population genetics studies and surveillance programs.

Evaluation of 96-well high-throughput DNA extraction methods for 16S rRNA gene metabarcoding.

Gaining meaningful insights into bacterial communities associated with animal hosts requires the provision of high-quality nucleic acids. Although many studies have compared DNA extraction methods for samples with low bacterial biomass (e.g. water) or specific PCR inhibitors (e.g. plants), DNA extraction bias in samples without inherent technical constraint (e.g. animal samples) has received little attention. Furthermore, there is an urgent need to identify a DNA extraction methods in a high-throughput format that decreases the cost and time for processing large numbers of samples. We here evaluated five DNA extraction protocols, using silica membrane-based spin columns and a 96-well microplate format and based on either mechanical or enzymatic lysis or a combination of both, using three bacterial mock communities and Illumina sequencing of the V4 region of the 16SrRNA gene. Our results showed that none of the DNA extraction methods fully eliminated bias associated with unequal lysis efficiencies. However, we identified a DNA extraction method with a lower bias for each mock community standard. Of these methods, those including an enzymatic lysis showed biases specific to some bacteria. Altogether, these results again demonstrate the importance of DNA extraction standardization to be able to compare the microbiome results of different samples. In this attempt, we advise for the use of the 96-well DNeasy Blood and Tissue kit (Qiagen) with a zirconia bead-beating procedure, which optimizes altogether the cost, handling time and bacteria-specific effects associated with enzymatic lysis.

Does Host Plant Drive Variation in Microbial Gut Communities in a Recently Shifted Pest?

Biotic interactions can modulate the responses of organisms to environmental stresses, including diet changes. Gut microbes have substantial effects on diverse ecological and evolutionary traits of their hosts, and microbial communities can be highly dynamic within and between individuals in space and time. Modulations of the gut microbiome composition and their potential role in the success of a species to maintain itself in a new environment have been poorly studied to date. Here we examine this question in a large wood-boring beetle Cacosceles newmannii (Cerambycidae), that was recently found thriving on a newly colonized host plant. Using 16S metabarcoding, we assessed the gut bacterial community composition of larvae collected in an infested field and in "common garden" conditions, fed under laboratory-controlled conditions on four either suspected or known hosts (sugarcane, tea tree, wattle, and eucalyptus). We analysed microbiome variation (i.e. diversity and differentiation), measured fitness-related larval growth, and studied host plant lignin and cellulose contents, since their degradation is especially challenging for wood-boring insects. We show that sugarcane seems to be a much more favourable host for larval growth. Bacterial diversity level was the highest in field-collected larvae, whereas lab-reared larvae fed on sugarcane showed a relatively low level of diversity but very specific bacterial variants. Bacterial communities were mainly dominated by Proteobacteria, but were significantly different between sugarcane-fed lab-reared larvae and any other hosts or field-collected larvae. We identified changes in the gut microbiome associated with different hosts over a short time frame, which support the hypothesis of a role of the microbiome in host switches.

Is sonographic measurement of head-perineum distance useful to predict obstetrical anal sphincter injury in case of vacuum delivery?

OBJECTIVE: Determine if head-perineum distance (HPD) measurement before vacuum extraction (VE) was predictive of an obstetric anal sphincter injury (OASIS) occurrence. METHODS: Retrospective, bicentric (Lille and Poissy, France) cohort study conducted from January 2019 to June 2020. All VE in singleton pregnancies of ≥34 weeks were included. HPD measurement was performed without compression of the tissues before each VE. The judgment criterion was the occurrence of an OASIS. RESULTS: Of 12 568 deliveries, VE was performed in 1093 (8.6%). Among these 1093 women undergoing VE, 675 (61.7%) with HPD measurement were included. OASIS was found in 6.5% of women (n = 44; 95% CI 4.5-8.7). HPD was not associated with OASIS (38.5 ± 12.6 mm in women with OASIS vs 37.4 ± 12.0 mm in women without; adjusted OR [aOR] per 5 mm increase = 0.92; 95% CI 0.79-1.06). Increased HPD was associated with higher risk of sequential extraction (aOR = 1.19; 95% CI 1.06-1.32), extraction duration >10 min (aOR = 1.12; 95% CI 1.02-1.23) and shoulder dystocia (aOR = 1.20; 95% CI 1.03-1.40). CONCLUSION: Ultrasound-measured head-perineum distance does not predict the occurrence of obstetric anal sphincter injury during a VE. The interest of HPD is more about predicting the success or difficulty of VE rather its specific complications.

Barcoding pest species in a biodiversity hot-spot: the South African polyphagous broad-nosed weevils (Coleoptera, Curculionidae, Entiminae).

Polyphagous broad nosed weevils (Curculionidae: Entiminae) constitute a large and taxonomically challenging subfamily that contains economically significant agricultural pests worldwide. South Africa is a hot-spot for biodiversity and several species of indigenous and endemic genera of Entiminae have shifted on to cultivated plants, with some being phytosanitary pests. The sporadic pest status of many species (where the species has an occasional economic impact on the agricultural industry, but is not encountered often enough that is is readily recognisable by researchers and agricultural extension workers) and the presence of pest complexes and cryptic species represent an identification challenge to non-specialists. Furthermore, no comprehensive identification tools exist to identify immature stages that may be found in crops/soil. In this paper, a curated barcoding database with 70 COI sequences from 41 species (39 Entiminae, 2 Cyclominae) is initiated, to assist with the complexity of identification of species in this group.

DNA barcoding for bio-surveillance of emerging pests and species identification in Afrotropical Prioninae (Coleoptera, Cerambycidae).

DNA barcoding has been succesfully used for bio-surveillance of forest and agricultural pests in temperate areas, but has few applications in the tropics and particulary in Africa. Cacosceles newmannii (Coleoptera: Cerambycidae) is a Prioninae species that is locally causing extensive damage in commercially-grown sugarcane in the KwaZulu-Natal Province in South Africa. Due to the risk of spread of this species to the rest of southern Africa and to other sugarcane growing regions, clear and easy identification of this pest is critical for monitoring and for phytosanitary services. The genus Cacosceles Newman, 1838 includes four species, most being very similar in morphology. The damaging stage of the species is the larva, which is inherently difficult to distinguish morphologically from other Cerambycidae species. A tool for rapid and reliable identification of this species was needed by plant protection and quarantine agencies to monitor its potential abundance and spread. Here, we provide newly-generated barcodes for C. newmannii that can be used to reliably identify any life stage, even by non-trained taxonomists. In addition, we compiled a curated DNA barcoding reference library for 70 specimens of 20 named species of Afrotropical Prioninae to evaluate DNA barcoding as a valid tool to identify them. We also assessed the level of deeply conspecific mitochondrial lineages. Sequences were assigned to 42 different Barcode Index Numbers (BINs), 28 of which were new to BOLD. Out of the 20 named species barcoded, 11 (52.4%) had their own unique Barcode Index Number (BIN). Eight species (38.1%) showed multiple BINs with no morphological differentiation. Amongst them, C. newmannii showed two highly divergent genetic clusters which co-occur sympatrically, but further investigation is required to test whether they could represent new cryptic species.

Cancer during Pregnancy: A Review of Preclinical and Clinical Transplacental Transfer of Anticancer Agents.

The occurrence of cancer during pregnancy is observed in 1 in 1000 pregnancies and is expected to increase given the trend of delaying childbearing. While breast cancer is the most common, the incidence of other cancers, such as cervical, ovarian, and lung cancers as well as hemopathies and melanomas, is also increasing. Thus, cancer occurrence in pregnant women raises questions of management during pregnancy and, especially, assessment of the treatment benefit-risk ratio to ensure optimal management for the mother while ensuring the safety of the fetus. Chemotherapy remains a cornerstone of cancer management. If the use of anticancer agents appears possible during pregnancy, while avoiding the first trimester, the extent of placental transfer of different anticancer agents varies considerably thereafter. Furthermore, the significant physiological pharmacokinetic variations observed in pregnant women may have an impact on the placental transfer of anticancer agents. Given the complexity of predicting placental transfer of anticancer agents, preclinical studies are therefore mandatory. The aim of this review was to provide updated data on in vivo and ex vivo transplacental transfer of anticancer agents used in the management of the most common pregnancy-associated cancers to better manage these highly complex cases.

How Bank Vole-PUUV Interactions Influence the Eco-Evolutionary Processes Driving Nephropathia Epidemica Epidemiology-An Experimental and Genomic Approach.

In Europe, Puumala virus (PUUV) is responsible for nephropathia epidemica (NE), a mild form of hemorrhagic fever with renal syndrome (HFRS). Despite the presence of its reservoir, the bank vole, on most of French territory, the geographic distribution of NE cases is heterogeneous and NE endemic and non-endemic areas have been reported. In this study we analyzed whether bank vole-PUUV interactions could partly shape these epidemiological differences. We performed crossed-experimental infections using wild bank voles from French endemic (Ardennes) and non-endemic (Loiret) areas and two French PUUV strains isolated from these areas. The serological response and dynamics of PUUV infection were compared between the four cross-infection combinations. Due to logistical constraints, this study was based on a small number of animals. Based on this experimental design, we saw a stronger serological response and presence of PUUV in excretory organs (bladder) in bank voles infected with the PUUV endemic strain. Moreover, the within-host viral diversity in excretory organs seemed to be higher than in other non-excretory organs for the NE endemic cross-infection but not for the NE non-endemic cross-infection. Despite the small number of rodents included, our results showed that genetically different PUUV strains and in a lesser extent their interaction with sympatric bank voles, could affect virus replication and diversity. This could impact PUUV excretion/transmission between rodents and to humans and in turn at least partly shape NE epidemiology in France.

Detection and Genetic Characterization of Puumala Orthohantavirus S-Segment in Areas of France Non-Endemic for Nephropathia Epidemica.

Puumala virus (PUUV) in Europe causes nephropathia epidemica (NE), a mild form of hemorrhagic fever with renal syndrome (HFRS). The incidence of NE is highly heterogeneous spatially, whereas the geographic distribution of the wild reservoir of PUUV, the bank vole, is essentially homogeneous. Our understanding of the processes driving this heterogeneity remains incomplete due to gaps in knowledge. Little is known about the current distribution and genetic variation of PUUV in the areas outside the well-identified zones of NE endemicity. We trapped bank voles in four forests in French regions in which NE is considered non-endemic, but sporadic NE cases have been reported recently. We tested bank voles for anti-PUUV IgG and characterized the S segment sequences of PUUV from seropositive animals. Phylogenetic analyses revealed specific amino-acid signatures and genetic differences between PUUV circulating in non-endemic and nearby NE-endemic areas. We also showed, in temporal surveys, that the amino-acid sequences of PUUV had undergone fewer recent changes in areas non-endemic for NE than in endemic areas. The evolutionary history of the current French PUUV clusters was investigated by phylogeographic approaches, and the results were considered in the context of the history of French forests. Our findings highlight the need to monitor the circulation and genetics of PUUV in a larger array of bank vole populations, to improve our understanding of the risk of NE.

DNA Metabarcoding as a Tool for Disentangling Food Webs in Agroecosystems.

Better knowledge of food webs and related ecological processes is fundamental to understanding the functional role of biodiversity in ecosystems. This is particularly true for pest regulation by natural enemies in agroecosystems. However, it is generally difficult to decipher the impact of predators, as they often leave no direct evidence of their activity. Metabarcoding via high-throughput sequencing (HTS) offers new opportunities for unraveling trophic linkages between generalist predators and their prey, and ultimately identifying key ecological drivers of natural pest regulation. Here, this approach proved effective in deciphering the diet composition of key predatory arthropods (nine species.; 27 prey taxa), insectivorous birds (one species, 13 prey taxa) and bats (one species; 103 prey taxa) sampled in a millet-based agroecosystem in Senegal. Such information makes it possible to identify the diet breadth and preferences of predators (e.g., mainly moths for bats), to design a qualitative trophic network, and to identify patterns of intraguild predation across arthropod predators, insectivorous vertebrates and parasitoids. Appropriateness and limitations of the proposed molecular-based approach for assessing the diet of crop pest predators and trophic linkages are discussed.

Multilocus phylogeography of the world populations of Elaeidobius kamerunicus (Coleoptera, Curculionidae), pollinator of the palm Elaeis guineensis.

Elaeidobius kamerunicus Faust (Coleoptera, Curculionidae) is one of the specific pollinators on inflorescences of the African oil palm Elaeis guineensis Jacquin. This derelomine weevil is native to tropical Africa. During the late 20th century, it was introduced into all tropical regions where E. guineensis is grown, in order to improve its pollination and fruit set. Despite an overall success, a decline in pollination efficiency has been documented in several regions. In this study, we reconstructed a multilocus phylogeography of the world populations of E. kamerunicus, in order to explore its genetic diversity in its native and introduced ranges. Our results showed that African populations of E. kamerunicus are forming two differentiated mitochondrial clusters in West and central Africa, forming a contact zone along the Cameroon Volcanic Line. The existence of this sharp contact zone along this weak altitudinal barrier suggests that other parameters, such as climate, may be driving the distribution of populations. A differential genetic structure between mitochondrial and nuclear genes, and the strong level of genetic structure of the mitochondrial gene, also suggest sex-biased dispersal in this species, with males dispersing more than females. The genetic structure inferred from Asian and South American populations suggests that they originate from populations of both western and central tropical Africa and that a bottleneck has probably been experienced by these populations.

Renal colic with ureterohydronephrosis due to menstrual cup.

We report a rare case of ureterohydronephrosis due to an improperly positioned menstrual cup.

Deciphering host-parasitoid interactions and parasitism rates of crop pests using DNA metabarcoding.

An accurate estimation of parasitism rates and diversity of parasitoids of crop insect pests is a prerequisite for exploring processes leading to efficient natural biocontrol. Traditional methods such as rearing have been often limited by taxonomic identification, insect mortality and intensive work, but the advent of high-throughput sequencing (HTS) techniques, such as DNA metabarcoding, is increasingly seen as a reliable and powerful alternative approach. Little has been done to explore the benefits of such an approach for estimating parasitism rates and parasitoid diversity in an agricultural context. In this study, we compared the composition of parasitoid species and parasitism rates between rearing and DNA metabarcoding of host eggs and larvae of the millet head miner, Heliocheilus albipunctella De Joannis (Lepidoptera, Noctuidae), collected from millet fields in Senegal. We first assessed the detection threshold for the main ten endoparasitoids, by sequencing PCR products obtained from artificial dilution gradients of the parasitoid DNAs in the host moth. We then assessed the potential of DNA metabarcoding for diagnosing parasitism rates in samples collected from the field. Under controlled conditions, our results showed that relatively small quantities of parasitoid DNA (0.07 ng) were successfully detected within an eight-fold larger quantity of host DNA. Parasitoid diversity and parasitism rate estimates were always higher for DNA metabarcoding than for host rearing. Furthermore, metabarcoding detected multi-parasitism, cryptic parasitoid species and differences in parasitism rates between two different sampling sites. Metabarcoding shows promise for gaining a clearer understanding of the importance and complexity of host-parasitoid interactions in agro-ecosystems, with a view to improving pest biocontrol strategies.

Discontinuities in quinoa biodiversity in the dry Andes: An 18-century perspective based on allelic genotyping.

History and environment shape crop biodiversity, particularly in areas with vulnerable human communities and ecosystems. Tracing crop biodiversity over time helps understand how rural societies cope with anthropogenic or climatic changes. Exceptionally well preserved ancient DNA of quinoa (Chenopodium quinoa Willd.) from the cold and arid Andes of Argentina has allowed us to track changes and continuities in quinoa diversity over 18 centuries, by coupling genotyping of 157 ancient and modern seeds by 24 SSR markers with cluster and coalescence analyses. Cluster analyses revealed clear population patterns separating modern and ancient quinoas. Coalescence-based analyses revealed that genetic drift within a single population cannot explain genetic differentiation among ancient and modern quinoas. The hypothesis of a genetic bottleneck related to the Spanish Conquest also does not seem to apply at a local scale. Instead, the most likely scenario is the replacement of preexisting quinoa gene pools with new ones of lower genetic diversity. This process occurred at least twice in the last 18 centuries: first, between the 6th and 12th centuries-a time of agricultural intensification well before the Inka and Spanish conquests-and then between the 13th century and today-a period marked by farming marginalization in the late 19th century likely due to a severe multidecadal drought. While these processes of local gene pool replacement do not imply losses of genetic diversity at the metapopulation scale, they support the view that gene pool replacement linked to social and environmental changes can result from opposite agricultural trajectories.

Evolution of almond genetic diversity and farmer practices in Lebanon: impacts of the diffusion of a graft-propagated cultivar in a traditional system based on seed-propagation.

BACKGROUND: Under cultivation, many outcrossing fruit tree species have switched from sexual reproduction to vegetative propagation. Traditional production systems have persisted, where cultivar propagation is based on a mixed reproductive system. For millenia, almond, Prunus dulcis, has been propagated by seeds. Almond grafting remained of little importance until recently. In Lebanon, both sexual and clonal reproductions are used for almond propagation. We used 15 microsatellite markers to investigate the effect of introducing graft-propagated cultivars and associated practices, on the structure of the genetic diversity among and within the two main Lebanese cultivars. RESULTS: As expected, the sexually propagated cultivar Khachabi exhibited more genotypic and genetic diversity than the vegetatively propagated cultivar Halwani. It also exhibited lower differentiation among populations. The distribution of clones showed that propagation modes were not exclusive: farmers have introduced clonal propagation in the seed-propagated cultivar while they have maintained a diversity of genotypes within populations that were mostly graft-propagated. These practices are also important to avoid mate limitations that hamper fruit production in a self-incompatible species. 'Khachabi' is structured into two gene pools separated by the Lebanese mountains. As to 'Halwani', two different gene pools were introduced. The most ancient one shares the same geographic range as 'Khachabi'; longtime coexistence and sexual reproduction have resulted in admixture with 'Khachabi'. In contrast, the more recent introduction of the second gene pool in the Bekaa region followed an evolution towards more extensive clonal propagation of 'Halwani' limiting hybridizations. Furthermore, some pairs of geographically distant 'Halwani' orchards, exhibited low genetic distances, suggesting that a network of exchanges between farmers was effective on a large scale and/or that farmers brought clonal plant material from a common source. CONCLUSIONS: Almond diversification in Lebanon is clearly related to the evolution of propagation practices adapted to self-incompatible cultivars. The comparison between both cultivars demonstrated the genetic effects of the introduction of a new cultivar and the associated grafting propagation practices. Our study provided information to develop a strategy for in situ conservation of cultivars and to limit gene flow from introduced material to ancient orchards.

Mind the cell: Seasonal variation in telomere length mirrors changes in leucocyte profile.

Leucocytes are typically considered as a whole in studies examining telomere dynamics in mammals. Such an approach may be precarious, as leucocytes represent the only nucleated blood cells in mammals, their composition varies temporally, and telomere length differs between leucocyte types. To highlight this limitation, we examined here whether seasonal variation in leucocyte composition was related to variation in telomere length in free-ranging mandrills (Mandrilllus sphinx). We found that the leucocyte profile of mandrills varied seasonally, with lower lymphocyte proportion being observed during the long dry season presumably because of the combined effects of high nematode infection and stress at that time of the year. Interestingly, this low lymphocyte proportion during the long dry season was associated with shorter telomeres. Accordingly, based on longitudinal data, we found that seasonal changes in lymphocyte proportion were reflected by corresponding seasonal variation in telomere length. Overall, these results suggest that variation in lymphocyte proportion in blood can significantly affect telomere measurements in mammals. However, lymphocyte proportion did not entirely explain variation in telomere length. For instance, a lower lymphocyte proportion with age could not fully explain shorter telomeres in older individuals. Overall, our results show that telomere length and leucocyte profile are strongly although imperfectly intertwined, which may obscure the relationship between telomere dynamics and ageing processes in mammals.

Experimental infections of wild bank voles (Myodes glareolus) from nephropatia epidemica endemic and non-endemic regions revealed slight differences in Puumala virological course and immunological responses.

In Europe, the occurrence of nephropathia epidemica (NE), a human disease caused by Puumala virus (PUUV), exhibits considerable geographical heterogeneity despite the continuous distribution of its reservoir, the bank vole Myodes glareolus. To better understand the causes of this heterogeneity, wild voles sampled in two adjacent NE endemic and non-endemic regions of France were infected experimentally with PUUV. The responses of bank voles to PUUV infection, based on the levels of anti-PUUV IgG and viral RNA, were compared. Slight regional differences were highlighted despite the high inter-individual variability. Voles from the NE non-endemic region showed greater immune responsiveness to PUUV infection, but lower levels of RNA in their organs than voles from the endemic region. These results suggest the existence of regional variations in the sensitivity of bank voles that could contribute to the apparent absence of PUUV circulation among voles and the absence of NE in the non-endemic region.

Social organization and space use of a wild mandrill (Mandrillus sphinx) group.

Mandrills (Mandrillus sphinx) are enigmatic Old World primates whose social organization and ecology remain poorly known. Previous studies indicated, for example, that groups are composed of only adult females and their young or that several units composed of one adult male and several females make up larger permanent social units. Here, we present the first data on group composition and male ranging patterns from the only habituated wild mandrill group and examine how home range size and daily path length varied with environmental and demographic factors over a 15-month period. Our study site is located in southern Gabon where we followed the group on a daily basis, collecting data on presence, ranging, behavior, and parasite load of its individual members. Throughout the study, the group was made up of about 120 individuals, including several non-natal and natal adult and sub-adult males. One-male units were never observed. The mandrills traveled an estimated 0.44-6.50 km/day in a home range area of 866.7 ha. Exploratory analyses revealed that precipitation, the number of adult males present, and the richness of protozoan parasites were all positively correlated with daily path length. These results clarify the social system of mandrills and provide first insights into the factors that shape their ranging patterns.

Repeated evolution of fungal cultivar specificity in independently evolved ant-plant-fungus symbioses.

Some tropical plant species possess hollow structures (domatia) occupied by ants that protect the plant and in some cases also provide it with nutrients. Most plant-ants tend patches of chaetothyrialean fungi within domatia. In a few systems it has been shown that the ants manure the fungal patches and use them as a food source, indicating agricultural practices. However, the identity of these fungi has been investigated only in a few samples. To examine the specificity and constancy of ant-plant-fungus interactions we characterised the content of fungal patches in an extensive sampling of three ant-plant symbioses (Petalomyrmex phylax/Leonardoxa africana subsp. africana, Aphomomyrmex afer/Leonardoxa africana subsp. letouzeyi and Tetraponera aethiops/Barteria fistulosa) by sequencing the Internal Transcribed Spacers of ribosomal DNA. For each system the content of fungal patches was constant over individuals and populations. Each symbiosis was associated with a specific, dominant, primary fungal taxon, and to a lesser extent, with one or two specific secondary taxa, all of the order Chaetothyriales. A single fungal patch sometimes contained both a primary and a secondary taxon. In one system, two founding queens were found with the primary fungal taxon only, one that was shown in a previous study to be consumed preferentially. Because the different ant-plant symbioses studied have evolved independently, the high specificity and constancy we observed in the composition of the fungal patches have evolved repeatedly. Specificity and constancy also characterize other cases of agriculture by insects.