Impact of intraspecific variation in insect microbiomes on host phenotype and evolution.

Microbes can be an important source of phenotypic plasticity in insects. Insect physiology, behaviour, and ecology are influenced by individual variation in the microbial communities held within the insect gut, reproductive organs, bacteriome, and other tissues. It is becoming increasingly clear how important the insect microbiome is for insect fitness, expansion into novel ecological niches, and novel environments. These investigations have garnered heightened interest recently, yet a comprehensive understanding of how intraspecific variation in the assembly and function of these insect-associated microbial communities can shape the plasticity of insects is still lacking. Most research focuses on the core microbiome associated with a species of interest and ignores intraspecific variation. We argue that microbiome variation among insects can be an important driver of evolution, and we provide examples showing how such variation can influence fitness and health of insects, insect invasions, their persistence in new environments, and their responses to global environmental changes. A and B are two stages of an individual or a population of the same species. The drivers lead to a shift in the insect associated microbial community, which has consequences for the host. The complex interplay of those consequences affects insect adaptation and evolution and influences insect population resilience or invasion.

Changes in Bacterial Community Structure Across the Different Life Stages of Black Soldier Fly (Hermetia illucens).

The digestive capacity of organic compounds by the black soldier fly (BSF, Hermetia illucens, Diptera: Stratiomyidae, Linnaeus, 1758) is known to rely on complex larva-microbiota interactions. Although insect development is known to be a driver of changes of bacterial communities, the fluctuations along BSF life cycle in terms of composition and diversity of bacterial communities are still unknown. In this work, we used a metabarcoding approach to explore the differences in bacterial diversity along all four BSF developmental stages: eggs, larvae, pupae, and adult. We detected not only significant differences in bacterial community composition and species richness along the development of BSF, but also nine prevalent amplicon single variants (ASVs) forming the core microbiota. Out of the 2010 ASVs identified, 160 were significantly more abundant in one of the life stages. Moreover, using PICRUSt2, we inferred 27 potential metabolic pathways differentially used among the BSF life cycle. This distribution of metabolic pathways was congruent with the bacterial taxonomic distribution among life stages, demonstrating that the functional requirements of each phase of development are drivers of bacterial composition and diversity. This study provides a better understanding of the different metabolic processes occurring during BSF development and their links to changes in bacterial taxa. This information has important implications for improving bio-waste processing in such an economically important insect species.

Development of a multiprimer metabarcoding approach to understanding trophic interactions in agroecosystems.

To understand trophic interactions and the precise ecological role of each predatory species, it is important to know which arthropod and plant resources are used by generalist predators in agroecosystems. Molecular approaches, such as the use of high-throughput sequencing (HTS), play a key role in identifying these resources. This study develops a multiprimer metabarcoding approach for screening the most common trophic interactions of two predatory arthropods with contrasting morphologies, Rhagonycha fulva (Coleoptera: Cantharidae) and Anthocoris nemoralis (Hemiptera: Anthocoridae) collected from a peach crop. To reduce the time and cost of this metabarcoding approach, we first evaluated the effect of using two different predator-pools of different size (10 and 23 individuals of the same species). We also used our system to analyze the performance of one and two primer pairs in the same library. Our results show that the analysis of 23 individuals together with the use of two primer pairs in the same library optimize the HTS analysis. Using these best-performing conditions, we then analyzed the entire bodies of field-collected predators as well as the washing solutions used to clean the insect bodies. We were able to identify both gut content (i.e., diet) and external pollen load (i.e., on the insects' bodies). This study also demonstrates the importance of washing predatory insects' bodies prior to HTS analysis when the target species have a considerable size (>10 mm) and hairy structures. This metabarcoding approach has significant potential for the study of trophic links in agriculture, revealing expected and unexpected trophic relationships.

Viral clearance capacity by continuous Protein A chromatography step using Sequential MultiColumn Chromatography.

In response to the strong demand of biological protein therapeutics, such as monoclonal antibodies (MAbs), continuous downstream process was developed to deliver these molecules while maintaining desired product consistency and quality attributes, and providing manufacturing efficiency and flexibility. Viral safety is a critical quality attribute for biopharmaceuticals, such as MAbs. Evaluation of the viral clearance by the downstream process is a key component of risk mitigation. Protein A chromatography is typically used as an initial capture step for MAbs and efficient for the removal of process-related impurities like Host Cell Proteins (HCP). This step can also contribute to the clearance of potential viral contaminants. Murine Minute Virus (MMV)-spiking experiments were performed at small scale to investigate the impact on the viral clearance efficiency of the way the Protein A chromatography step is carried out, whether in batch or multicolumn mode. Protein A chromatography step using Novasep Sequential MultiColumn Chromatography (SMCC) technology demonstrated no statistical difference in the viral reduction with reduction factor (RF) of 3.7 log10 (vs. RF of 3.8 log10 for batch). The experiments showed also similar viral distribution over the purification cycles and columns. Data confirmed that the viral clearance capacity by the continuous Protein A chromatography step using SMCC technology is maintained and efficient.

Towards robust and repeatable sampling methods in eDNA-based studies.

DNA-based techniques are increasingly used for measuring the biodiversity (species presence, identity, abundance and community composition) of terrestrial and aquatic ecosystems. While there are numerous reviews of molecular methods and bioinformatic steps, there has been little consideration of the methods used to collect samples upon which these later steps are based. This represents a critical knowledge gap, as methodologically sound field sampling is the foundation for subsequent analyses. We reviewed field sampling methods used for metabarcoding studies of both terrestrial and freshwater ecosystem biodiversity over a nearly three-year period (n = 75). We found that 95% (n = 71) of these studies used subjective sampling methods and inappropriate field methods and/or failed to provide critical methodological information. It would be possible for researchers to replicate only 5% of the metabarcoding studies in our sample, a poorer level of reproducibility than for ecological studies in general. Our findings suggest greater attention to field sampling methods, and reporting is necessary in eDNA-based studies of biodiversity to ensure robust outcomes and future reproducibility. Methods must be fully and accurately reported, and protocols developed that minimize subjectivity. Standardization of sampling protocols would be one way to help to improve reproducibility and have additional benefits in allowing compilation and comparison of data from across studies.

Agroecological management of a soil-dwelling orthopteran pest in vineyards.

The efficacy of different combinations of undervine and inter-row treatments for managing a soil-dwelling orthopteran pest, weta (Hemiandrus sp.), in vineyards was investigated over 2 seasons. This insect damages vine buds, thus reducing subsequent grape yield. The undervine treatments comprised pea straw mulch, mussel shells, tick beans [Vicia faba Linn. var minor (Fab)], plastic sleeves on vine trunks (treated control) and control (no intervention), while inter-rows contained either the existing vegetation or tick beans. Treatments were arranged in a randomized complete block design with 10 replicates. Data were collected on weta densities, damage to beans and components of yield. The latter were numbers of bud laid down per vine, shoots per bud, clusters per shoot, grape bunches per vine, bunch weight and yield. The undervine treatments significantly affected all variables except the number of shoots per bud. In contrast, none of the variables was significantly affected by the inter-row treatments or their interaction with undervine treatments, apart from weta density. At the end of the experiment, weta density in the shell treatment was about 58% lower than in the control. As a result, there was about 39% significant yield increase in that treatment compared to the control. Although the undervine beans and sleeves treatments increased yield, there were no reductions in weta density. With undervine beans, the insect fed on the bean plants instead of vine buds. Thus, yield in that treatment was approximately 28% higher than in the control. These results demonstrate that simple agroecological management approaches can reduce above-ground damage by soil-dwelling insects.

Diet Composition and Variability of Wild Octopus vulgaris and Alloteuthis media (Cephalopoda) Paralarvae: a Metagenomic Approach.

The high mortality of cephalopod early stages is the main bottleneck to grow them from paralarvae to adults in culture conditions, probably because the inadequacy of the diet that results in malnutrition. Since visual analysis of digestive tract contents of paralarvae provides little evidence of diet composition, the use of molecular tools, particularly next generation sequencing (NGS) platforms, offers an alternative to understand prey preferences and nutrient requirements of wild paralarvae. In this work, we aimed to determine the diet of paralarvae of the loliginid squid Alloteuthis media and to enhance the knowledge of the diet of recently hatched Octopus vulgaris paralarvae collected in different areas and seasons in an upwelling area (NW Spain). DNA from the dissected digestive glands of 32 A. media and 64 O. vulgaris paralarvae was amplified with universal primers for the mitochondrial gene COI, and specific primers targeting the mitochondrial gene 16S gene of arthropods and the mitochondrial gene 16S of Chordata. Following high-throughput DNA sequencing with the MiSeq run (Illumina), up to 4,124,464 reads were obtained and 234,090 reads of prey were successfully identified in 96.87 and 81.25% of octopus and squid paralarvae, respectively. Overall, we identified 122 Molecular Taxonomic Units (MOTUs) belonging to several taxa of decapods, copepods, euphausiids, amphipods, echinoderms, molluscs, and hydroids. Redundancy analysis (RDA) showed seasonal and spatial variability in the diet of O. vulgaris and spatial variability in A. media diet. General Additive Models (GAM) of the most frequently detected prey families of O. vulgaris revealed seasonal variability of the presence of copepods (family Paracalanidae) and ophiuroids (family Euryalidae), spatial variability in presence of crabs (family Pilumnidae) and preference in small individual octopus paralarvae for cladocerans (family Sididae) and ophiuroids. No statistically significant variation in the occurrences of the most frequently identified families was revealed in A. media. Overall, these results provide new clues about dietary preferences of wild cephalopod paralarvae, thus opening up new scenarios for research on trophic ecology and digestive physiology under controlled conditions.

Ecological and pest-management implications of sex differences in scarab landing patterns on grape vines.

BACKGROUND: Melolonthinae beetles, comprising different white grub species, are a globally-distributed pest group. Their larvae feed on roots of several crop and forestry species, and adults can cause severe defoliation. In New Zealand, the endemic scarab pest Costelytra zealandica (White) causes severe defoliation on different horticultural crops, including grape vines (Vitis vinifera). Understanding flight and landing behaviours of this pest can help inform pest management decisions. METHODS: Adult beetles were counted and then removed from 96 grape vine plants from 21:30 until 23:00 h, every day from October 26 until December 2, during 2014 and 2015. Also, adults were removed from the grape vine foliage at dusk 5, 10, 15, 20 and 25 min after flight started on 2015. Statistical analyses were performed using generalised linear models with a beta-binomial distribution to analyse proportions and with a negative binomial distribution for beetle abundance. RESULTS: By analysing C. zealandica sex ratios during its entire flight season, it is clear that the proportion of males is higher at the beginning of the season, gradually declining towards its end. When adults were successively removed from the grape vines at 5-min intervals after flight activity begun, the mean proportion of males ranged from 6-28%. The male proportion suggests males were attracted to females that had already landed on grape vines, probably through pheromone release. DISCUSSION: The seasonal and daily changes in adult C. zealandica sex ratio throughout its flight season are presented for the first time. Although seasonal changes in sex ratio have been reported for other melolonthines, changes during their daily flight activity have not been analysed so far. Sex-ratio changes can have important consequences for the management of this pest species, and possibly for other melolonthines, as it has been previously suggested that C. zealandica females land on plants that produce a silhouette against the sky. Therefore, long-term management might evaluate the effect of different plant heights and architecture on female melolonthine landing patterns, with consequences for male distribution, and subsequently overall damage within horticultural areas.

Impacts of Endemic Earthworms (Megascolecidae) in Biosolids-Amended Soil.

Biosolids can be a valuable fertilizer for agriculture and in ecological restoration, although there are concerns about contaminants. Earthworm activity, including vermicomposting of biosolids, may influence the efficacy of their use. We investigated how two New Zealand endemic anecic species of (cf. ) responded to biosolids amendment and affected the mobility of nutrients and trace elements as well as greenhouse gas emissions in biosolids-amended soil. Earthworms were incubated with mixtures of biosolids-amended soil (0, 6.25, 12.5, 25, and 50% biosolids by volume) for 21 d. All species survived in the soil-biosolids mixtures but not in 100% biosolids. The native earthworms, and sp.2, increased KCl-extractable NH and NO by up to 29%, substantially more than . All species significantly increased microbial biomass carbon and Ca(NO)-extractable Cu but significantly decreased dehydrogenase enzymes activity in biosolids-amended soil. Concentrations of Ca(NO)-extractable Mg, S, Fe, Mn, Cd, Co, and Zn varied between earthworm species and with biosolids addition rates. New Zealand native earthworms exacerbated NO emissions from soil, whereas did not. is clearly a preferred species for vermicomposting biosolids and is more tolerant of high concentrations of biosolids. However, New Zealand native earthworms may be more suitable for improving the fertility of soil amended with biosolids.

Percentage-based Author Contribution Index: a universal measure of author contribution to scientific articles.

BACKGROUND: Deciphering the amount of work provided by different co-authors of a scientific paper has been a recurrent problem in science. Despite the myriad of metrics available, the scientific community still largely relies on the position in the list of authors to evaluate contributions, a metric that attributes subjective and unfounded credit to co-authors. We propose an easy to apply, universally comparable and fair metric to measure and report co-authors contribution in the scientific literature. METHODS: The proposed Author Contribution Index (ACI) is based on contribution percentages provided by the authors, preferably at the time of submission. Researchers can use ACI to compare the contributions of different authors, describe the contribution profile of a particular researcher or analyse how contribution changes through time. We provide such an analysis based on contribution percentages provided by 97 scientists from the field of ecology who voluntarily responded to an online anonymous survey. RESULTS: ACI is simple to understand and to implement because it is based solely on percentage contributions and the number of co-authors. It provides a continuous score that reflects the contribution of one author as compared to the average contribution of all other authors. For example, ACI(i) = 3, means that author i contributed three times more than what the other authors contributed on average. Our analysis comprised 836 papers published in 2014-2016 and revealed patterns of ACI values that relate to career advancement. CONCLUSION: There are many examples of author contribution indices that have been proposed but none has really been adopted by scientific journals. Many of the proposed solutions are either too complicated, not accurate enough or not comparable across articles, authors and disciplines. The author contribution index presented here addresses these three major issues and has the potential to contribute to more transparency in the science literature. If adopted by scientific journals, it could provide job seekers, recruiters and evaluating bodies with a tool to gather information that is essential to them and cannot be easily and accurately obtained otherwise. We also suggest that scientists use the index regardless of whether it is implemented by journals or not.

Assessing pollinators' use of floral resource subsidies in agri-environment schemes: An illustration using Phacelia tanacetifolia and honeybees.

BACKGROUND: Honeybees (Apis mellifera L.) are frequently used in agriculture for pollination services because of their abundance, generalist floral preferences, ease of management and hive transport. However, their populations are declining in many countries. Agri-Environment Schemes (AES) are being implemented in agricultural systems to combat the decline in populations of pollinators and other insects. Despite AES being increasingly embedded in policy and budgets, scientific assessments of many of these schemes still are lacking, and only a few studies have examined the extent to which insect pollinators use the floral enhancements that are part of AES and on which floral components they feed (i.e., pollen and/or nectar). METHODS: In the present work, we used a combination of observations on honeybee foraging for nectar/pollen from the Californian annual plant Phacelia tanacetifolia in the field, collection of pollen pellets from hives, and pollen identification, to assess the value of adding phacelia to an agro-ecosystem to benefit honeybees. RESULTS: It was found that phacelia pollen was almost never taken by honeybees. The work here demonstrates that honeybees may not use the floral enhancements added to a landscape as expected and points to the need for more careful assessments of what resources are used by honeybees in AES and understanding the role, if any, which AES play in enhancing pollinator fitness. DISCUSSION: We recommend using the methodology in this paper to explore the efficacy of AES before particular flowering species are adopted more widely to give a more complete illustration of the actual efficacy of AES.

Scarcity of ecosystem services: an experimental manipulation of declining pollination rates and its economic consequences for agriculture.

Ecosystem services (ES) such as pollination are vital for the continuous supply of food to a growing human population, but the decline in populations of insect pollinators worldwide poses a threat to food and nutritional security. Using a pollinator (honeybee) exclusion approach, we evaluated the impact of pollinator scarcity on production in four brassica fields, two producing hybrid seeds and two producing open-pollinated ones. There was a clear reduction in seed yield as pollination rates declined. Open-pollinated crops produced significantly higher yields than did the hybrid ones at all pollination rates. The hybrid crops required at least 0.50 of background pollination rates to achieve maximum yield, whereas in open-pollinated crops, 0.25 pollination rates were necessary for maximum yield. The total estimated economic value of pollination services provided by honeybees to the agricultural industry in New Zealand is NZD $1.96 billion annually. This study indicates that loss of pollination services can result in significant declines in production and have serious implications for the market economy in New Zealand. Depending on the extent of honeybee population decline, and assuming that results in declining pollination services, the estimated economic loss to New Zealand agriculture could be in the range of NZD $295-728 million annually.

Can ecosystem-scale translocations mitigate the impact of climate change on terrestrial biodiversity? Promises, pitfalls, and possibilities: Ecosystem-scale translocations.

Because ecological interactions are the first components of the ecosystem to be impacted by climate change, future forms of threatened-species and ecosystem management should aim at conserving complete, functioning communities rather than single charismatic species. A possible way forward is the deployment of ecosystem-scale translocation (EST), where above- and below-ground elements of a functioning terrestrial ecosystem (including vegetation and topsoil) are carefully collected and moved together. Small-scale attempts at such practice have been made for the purpose of ecological restoration. By moving larger subsets of functioning ecosystems from climatically unstable regions to more stable ones, EST could provide a practical means to conserve mature and complex ecosystems threatened by climate change. However, there are a number of challenges associated with EST in the context of climate change mitigation, in particular the choice of donor and receptor sites. With the aim of fostering discussion and debate about the EST concept, we  1) outline the possible promises and pitfalls of EST in mitigating the impact of climate change on terrestrial biodiversity and 2) use a GIS-based approach to illustrate how  potential source and receptor sites, where EST could be trialed and evaluated globally, could be identified.

Preference of a native beetle for "exoticism," characteristics that contribute to invasive success of Costelytra zealandica (Scarabaeidae: Melolonthinae).

Widespread replacement of native ecosystems by productive land sometimes results in the outbreak of a native species. In New Zealand, the introduction of exotic pastoral plants has resulted in diet alteration of the native coleopteran species, Costelytra zealandica (White) (Scarabaeidae) such that this insect has reached the status of pest. In contrast, C. brunneum (Broun), a congeneric species, has not developed such a relationship with these 'novel' host plants. This study investigated the feeding preferences and fitness performance of these two closely related scarab beetles to increase fundamental knowledge about the mechanisms responsible for the development of invasive characteristics in native insects. To this end, the feeding preference of third instar larvae of both Costelytra species was investigated using an olfactometer device, and the survival and larval growth of the invasive species C. zealandica were compared on native and exotic host plants. Costelytra zealandica, when sampled from exotic pastures, was unable to fully utilise its ancestral native host and showed higher feeding preference and performance on exotic plants. In contrast, C. zealandica sampled from native grasslands did not perform significantly better on either host and showed similar feeding preferences to C. brunneum, which exhibited no feeding preference. This study suggests the possibility of strong intraspecific variation in the ability of C. zealandica to exploit native or exotic plants, supporting the hypothesis that such ability underpins the existence of distinct host-races in this species.

The PGI enzyme system and fitness response to temperature as a measure of environmental tolerance in an invasive species.

In the field of invasion ecology, the determination of a species' environmental tolerance, is a key parameter in the prediction of its potential distribution, particularly in the context of global warming. In poikilothermic species such as insects, temperature is often considered the most important abiotic factor that affects numerous life-history and fitness traits through its effect on metabolic rate. Therefore the response of an insect to challenging temperatures may provide key information as to its climatic and therefore spatial distribution. Variation in the phosphoglucose-6-isomerase (PGI) metabolic enzyme-system has been proposed in some insects to underlie their relative fitness, and is recognised as a key enzyme in their thermal adaptation. However, in this context it has not been considered as a potential mechanism contributing to a species invasive cability. The present study aimed to compare the thermal tolerance of an invasive scarabaeid beetle, Costelytra zealandica (White) with that of the closely related, and in part sympatrically occurring, congeneric non-invasive species C. brunneum (Broun), and to consider whether any correlation with particular PGI genotypes was apparent. Third instar larvae of each species were exposed to one of three different temperatures (10, 15 and 20 °C) over six weeks and their fitness (survival and growth rate) measured and PGI phenotyping performed via cellulose acetate electrophoresis. No consistent relationship between PGI genotypes and fitness was detected, suggesting that PGI may not be contributing to the invasion success and pest status of C. zealandica.

Invasion success of a scarab beetle within its native range: host range expansion versus host-shift.

Only recently has it been formally acknowledged that native species can occasionally reach the status of 'pest' or 'invasive species' within their own native range. The study of such species has potential to help unravel fundamental aspects of biological invasions. A good model for such a study is the New Zealand native scarab beetle, Costelytra zealandica (White), which even in the presence of its natural enemies has become invasive in exotic pastures throughout the country. Because C. zealandica still occurs widely within its native habitat, we hypothesised that this species has only undergone a host range expansion (ability to use equally both an ancestral and new host) onto exotic hosts rather than a host shift (loss of fitness on the ancestral host in comparison to the new host). Moreover, this host range expansion could be one of the main drivers of its invasion success. In this study, we investigated the fitness response of populations of C. zealandica from native and exotic flora, to several feeding treatments comprising its main exotic host plant as well as one of its ancestral hosts. Our results suggest that our initial hypothesis was incorrect and that C. zealandica populations occurring in exotic pastures have experienced a host-shift rather than simply a host-range expansion. This finding suggests that an exotic plant introduction can facilitate the evolution of a distinct native host-race, a phenomenon often used as evidence for speciation in phytophagous insects and which may have been instrumental to the invasion success of C. zealandica.

[Cytological and virological medium performance and stability assessment using the Cobas 4800 HPV test (Roche Diagnostics) used in France]. / Étude des performances et de la stabilité des principaux milieux de conservation et/ou de transport de cellules du col de l'utérus utilisés en France par le test Cobas 4800 HPV (Roche Diagnostics).

Analytical and stability performances of ten media were compared to PreservCyt medium using the Cobas 4800 HPV test: Easyfix (Labonord SAS, France), Qualicyt (Qualicyt, France), NovaPrep HQ+ (NovaCyt, France), CMDH (SARL Alphapath France), Cyt-All (Cytomega, France), Digene Cervical Sampler (Qiagen, USA), Aptima (Gen-Probe, USA), Multi-Collect (Abbott, Allemagne), M4RT Micro Test (Remel, USA), et PCR-Media (Roche, Suisse). Most of media show a good correlation for all the performance characteristics studied. Cyt-All and NovaPrep HQ+ media are perfectly concordant with PreservCyt all parameters and genotypes considered. CMDH and M4RT have a reduced stability at +25°C (3 and 2 days respectively) and would not be conformed to current shipping practices. Most of media tested show analytical and stability performances equivalent with the reference medium. The prospects of such study are interesting because in the near future, providers would make available media adapted to the problem of cervical smear but also to the conservation, transport of virus or bacteria for performing simultaneous searches of HPV, Chlamydia trachomatis or Neisseria gonorrhea.

Pyrosequencing of prey DNA in faeces of carnivorous land snails to facilitate ecological restoration and relocation programmes.

Identifying and understanding predator diets is of high importance in biological conservation. This is particularly true for the introduction, establishment and maintenance of predator populations in newly created or modified ecological communities, such as translocation sites or restored habitats. Conservation status of predators may not permit captive feeding trials or intrusive gut-content methods, so non-intrusive diet assessment is required, such as faecal analysis. However, prey such as earthworms leave no morphological clues suitable for accurately discriminating between species consumed through visual faecal analysis. This study uses non-intrusive molecular methods on earthworm DNA extracted from the faeces of the carnivorous land snail Powelliphanta patrickensis to identify its earthworm diet and any seasonal trends. Data from 454-pyrosequencing revealed earthworm DNA in all samples (n = 60). Sequences were compared to a DNA library created from published and unpublished studies of New Zealand's endemic earthworms and online databases. Unidentified earthworm sequences were clustered into molecular operational taxonomic units (MOTUs). Twenty-six MOTUs were identified, 17 of which matched the library, whereas nine did not. Similarity indices indicate that there were seasonal differences (P < 0.05) in the earthworm communities represented in the summer and the winter diets. This study highlights the importance of utilising the vast body of data generated by pyrosequencing to investigate potential temporal diet shifts in protected species. The method described here is widely applicable to a wide range of predatory species of conservation interest and can further inform habitat restoration and relocation programmes to optimize the long-term survival of the target species.