Effect of urbanization and its environmental stressors on the intraspecific variation of flight functional traits in two bumblebee species.

The way urbanization shapes the intraspecific variation of pollinator functional traits is little understood. However, this topic is relevant for investigating ecosystem services and pollinator health. Here, we studied how urbanization affects the functional traits of workers in two bumblebee species (Bombus terrestris and B. pascuorum) sampled in 37 sites along a gradient of urbanization in North Italy (an area of 1800 km2 including the metropolitan context of Milan and other surrounding capital districts). Namely, we investigated the effect of land use composition, configuration, air temperature, flower resource abundance, and air pollutants on the variation of traits related to flight performance and of stress during insect development (i.e., wing size, wing shape and size fluctuating asymmetry). The functional traits of the two bumblebees responded idiosyncratically to urbanization. Urban temperatures were associated with smaller wing sizes in B. pascuorum and with more accentuated fluctuating asymmetry of wing size in B. terrestris. Moreover, flower abundance correlated with bigger wings in B. terrestris and with less asymmetric wing size in B. pascuorum. Other traits did not vary significantly, and other urban variables played minor effects. These species-specific variation patterns highlight that environmental stressor linked to urbanization negatively impact the traits related to flight performance and development stability of these syntopic bumblebees, with possible consequences on the pollination service they provide.

Extracellular RNAs: A Secret Arm of Immune System Regulation.

The immune system has evolved to protect multicellular organisms from the attack of a variety of pathogens. To exert this function efficiently, the system has developed the capacity to coordinate the function of different cell types and the ability to down-modulate the response when the foreign attack is over. For decades, immunologists believed that these two characteristics were primarily related to cytokine/chemokine-based communication and cell-to-cell direct contact. More recently, it has been shown that immune cells also communicate by transferring regulatory RNAs, microRNAs in particular, from one cell to the other. Several studies have suggested a functional role of extracellular regulatory RNAs in cell-to-cell communication in different cellular contexts. This minireview focuses on the potential role of extracellular RNA transfer in the regulation of adaptive immune response, also contextualizing it in a broader field of what is known of cell-free RNAs in communication among different organisms in the evolutionary scale.

DNA Barcoding Meets Nanotechnology: Development of a Universal Colorimetric Test for Food Authentication.

Food trade globalization and the growing demand for selected food varieties have led to the intensification of adulteration cases, especially in the form of species substitution and mixing with cheaper taxa. This phenomenon has huge economic impact and sometimes even public health implications. DNA barcoding represents a well-proven molecular approach to assess the authenticity of food items, although its use is hampered by analytical constraints and timeframes that are often prohibitive for the food market. To address such issues, we have introduced a new technology, named NanoTracer, that allows for rapid and naked-eye molecular traceability of any food and requires limited instrumentation and cost-effective reagents. Moreover, unlike sequencing, this method can be used to identify not only the substitution of a fine ingredient, but also its dilution with cheaper ones.

Exploration and Retrieval of Virus-Related Molecular Data Using ExTaxsI: The Monkeypox Use Case.

Retrieval and visualization of biological data are essential for understanding complex systems. With the increasing volume of data generated from high-throughput sequencing technologies, effective and optimized data visualization tools have become indispensable. This is particularly relevant in the COVID-19 postpandemic period, where understanding the diversity and interactions of microbial communities (i.e., viral and bacterial) constitutes an important asset to develop and plan suitable interventions.In this chapter, we show the usage and the potentials of ExTaxsI (Exploring Taxonomy Information) tool to retrieve viral biodiversity data stored in National Center for Biotechnology Information (NCBI) databases and create the related visualization. In addition, by integrating different functions and modules, the tool generates relevant types of visualization plots to facilitate the exploration of microbial biodiversity communities useful to deep dive into ecological and taxonomic relationships among different species and identify potential significant targets.Using the Monkeypox virus as a case study, this work points out significant perspectives on biological data visualization, which can be used to gain insights into the ecology, evolution, and pathogenesis of viruses. Accordingly, we show the potentiality of ExTaxsI to organize and describe the available/downloaded data in an easy, simple, and interpretable way allowing the user to interact dynamically with the visualization plots through specific filters, zoom, and explore functions.

Errors in ribosomal sequence datasets generated using PCR-coupled 'panbacterial' pyrosequencing, and the establishment of an improved approach.

Universal bacterial primers are often used in PCR-coupled sequencing approaches to investigate environmental and host-associated bacterial communities. Some of these primers can also amplify eukaryotic DNA. This is leading to the submission of datasets to public databases which are erroneously annotated as prokaryotic sequences. The present note sends a message about the risk of submitting incorrectly annotated sequence data and suggests a reliable approach for the sequencing of 16S rRNA genes and identification of bacteria within complex communities.

Urban habitat fragmentation and floral resources shape the occurrence of gut parasites in two bumblebee species.

Urbanization and the expansion of human activities foster radical ecosystem changes with cascading effects also involving host-pathogen interactions. Urban pollinator insects face several stressors related to landscape and local scale features such as green habitat loss, fragmentation and availability reduction of floral resources with unpredictable effects on parasite transmission. Furthermore, beekeeping may contribute to the spread of parasites to wild pollinators by increasing the number of parasite hosts. Here we used DNA-based diagnostics tools to evaluate how the occurrence of parasites, namely microsporidians (Nosema spp.), trypanosomatids (Crithidia spp.) and neogregarines (Apicystis bombi), is shaped by the above-mentioned stressors in two bumblebee species (i.e. Bombus terrestris and Bombus pascuorum). Infection rates of the two species were different and generally higher in B. terrestris. Moreover, they showed different responses towards the same ecological variables, possibly due to differences in body size and foraging habits supposed to affect their susceptibility to parasite infection. The probability of infection was found to be reduced in B. pascuorum by green habitat fragmentation, while increased along with floral resource availability. Unexpectedly, B. terrestris had a lower parasite richness nearby apiaries maybe due to the fact that parasites are prone to be transmitted among the most abundant species. Our finding supports the need to design proper conservation measures based on species-specific knowledge, as suggested by the variation in the parasite occurrence of the two species. Moreover, conservation policies aiming at safeguarding pollinators through flower planting should consider the indirect effects of these measures for parasite transmission together with pollinator biodiversity issues.

Red mark syndrome: Is the aquaculture water microbiome a keystone for understanding the disease aetiology?

Aquaculture significantly contributes to the growing demand for food worldwide. However, diseases associated with intensive aquaculture conditions, especially the skin related syndromes, may have significant implications on fish health and industry. In farmed rainbow trout, red mark syndrome (RMS), which consists of multiple skin lesions, currently lacks recognized aetiological agents, and increased efforts are needed to elucidate the onset of these conditions. Most of the past studies were focused on analyzing skin lesions, but no study focused on water, a medium constantly interacting with fish. Indeed, water tanks are environmental niches colonized by microbial communities, which may be implicated in the onset of the disease. Here, we present the results of water and sediment microbiome analyses performed in an RMS-affected aquaculture facility, bringing new knowledge about the environmental microbiomes harbored under these conditions. On the whole, no significant differences in the bacterial community structure were reported in RMS-affected tanks compared to the RMS-free ones. However, we highlighted significant differences in microbiome composition when analyzing different samples source (i.e., water and sediments). Looking at the finer scale, we measured significant changes in the relative abundances of specific taxa in RMS-affected tanks, especially when analyzing water samples. Our results provide worthwhile insight into a mostly uncharacterized ecological scenario, aiding future studies on the aquaculture built environment for disease prevention and monitoring.

DNA barcoding: a six-question tour to improve users' awareness about the method.

DNA barcoding is a recent and widely used molecular-based identification system that aims to identify biological specimens, and to assign them to a given species. However, DNA barcoding is even more than this, and besides many practical uses, it can be considered the core of an integrated taxonomic system, where bioinformatics plays a key role. DNA barcoding data could be interpreted in different ways depending on the examined taxa but the technique relies on standardized approaches, methods and analyses. The existing reference towards a common way to treat DNA barcoding data, analyses and results is the Barcode of Life Data Systems. However, the scientific community has produced in the recent years a number of alternative methods to manage barcoding data. The present work starts from this point, because users should be aware of the consequences their choices produce on the results. Despite the fact that a strict standardization is the essence of DNA barcoding, we propose a tour of six questions to improve the users' awareness about the method, the correct use of concepts and alternative tools provided by scientific community.

SKIOME Project: a curated collection of skin microbiome datasets enriched with study-related metadata.

Large amounts of data from microbiome-related studies have been (and are currently being) deposited on international public databases. These datasets represent a valuable resource for the microbiome research community and could serve future researchers interested in integrating multiple datasets into powerful meta-analyses. However, this huge amount of data lacks harmonization and it is far from being completely exploited in its full potential to build a foundation that places microbiome research at the nexus of many subdisciplines within and beyond biology. Thus, it urges the need for data accessibility and reusability, according to findable, accessible, interoperable and reusable (FAIR) principles, as supported by National Microbiome Data Collaborative and FAIR Microbiome. To tackle the challenge of accelerating discovery and advances in skin microbiome research, we collected, integrated and organized existing microbiome data resources from human skin 16S rRNA amplicon-sequencing experiments. We generated a comprehensive collection of datasets, enriched in metadata, and organized this information into data frames ready to be integrated into microbiome research projects and advanced post-processing analyses, such as data science applications (e.g. machine learning). Furthermore, we have created a data retrieval and curation framework built on three different stages to maximize the retrieval of datasets and metadata associated with them. Lastly, we highlighted some caveats regarding metadata retrieval and suggested ways to improve future metadata submissions. Overall, our work resulted in a curated skin microbiome datasets collection accompanied by a state-of-the-art analysis of the last 10 years of the skin microbiome field. Database URL:  https://github.com/giuliaago/SKIOMEMetadataRetrieval.

ExTaxsI: an exploration tool of biodiversity molecular data.

BACKGROUND: The increasing availability of multi-omics data is leading to regularly revised estimates of existing biodiversity data. In particular, the molecular data enable novel species to be characterized and the information linked to those already observed to be increased with new genomics data. For this reason, the management and visualization of existing molecular data, and their related metadata, through the implementation of easy-to-use IT tools have become a key point to design future research. The more users are able to access biodiversity-related information, the greater the ability of the scientific community to expand its knowledge in this area. RESULTS: In this article we focus on the development of ExTaxsI (Exploring Taxonomy Information), an IT tool that can retrieve biodiversity data stored in NCBI databases and provide a simple and explorable visualization. We use 3 case studies to show how an efficient organization of the available data can lead to obtaining new information that is fundamental as a starting point for new research. Using this approach highlights the limits in the distribution of data availability, a key factor to consider in the experimental design phase of broad-spectrum studies such as metagenomics. CONCLUSIONS: ExTaxsI can easily retrieve molecular data and its metadata with an explorable visualization, with the aim of helping researchers to improve experimental designs and highlight the main gaps in the coverage of available data.

Human intraocular filariasis caused by Dirofilaria sp. nematode, Brazil.

A case of human intraocular dirofilariasis is reported from northern Brazil. The nematode was morphologically and phylogenetically related to Dirofilaria immitis but distinct from reference sequences, including those of D. immitis infesting dogs in the same area. A zoonotic Dirofilaria species infesting wild mammals in Brazil and its implications are discussed.

Exploring Changes in the Microbiota of Aedes albopictus: Comparison Among Breeding Site Water, Larvae, and Adults.

The mosquito body hosts highly diverse microbes, which influence different physiological traits of both larvae and adults. The composition of adult mosquito microbiota is tightly linked to that of larvae, which are aquatic and feed on organic detritus, algae and prokaryotic microorganisms present in their breeding sites. Unraveling the ecological features of larval habitats that shape the structure of bacterial communities and their interactions with the mosquito host is still a poorly investigated topic in the Asian tiger mosquito Aedes albopictus, a highly invasive species that is vector of numerous arboviruses, including Dengue, Chikungunya, and Zika viruses. In this study, we investigated the composition of the bacterial community present in the water from a natural larval breeding site in which we separately reared wild-collected larvae and hatched eggs of the Foshan reference laboratory strain. Using sequence analysis of bacterial 16S rRNA gene amplicons, we comparatively analyzed the microbiota of the larvae and that of adult mosquitoes, deriving information about the relative impact of the breeding site water on shaping mosquito microbiota. We observed a higher bacterial diversity in breeding site water than in larvae or adults, irrespective of the origin of the sample. Moreover, larvae displayed a significantly different and most diversified microbial community than newly emerged adults, which appeared to be dominated by Proteobacteria. The microbiota of breeding site water significantly increased its diversity over time, suggesting the presence of a dynamic interaction among bacterial communities, breeding sites and mosquito hosts. The analysis of Wolbachia prevalence in adults from Foshan and five additional strains with different geographic origins confirmed the described pattern of dual wAlbA and wAlbB strain infection. However, differences in Wolbachia prevalence were detected, with one strain from La Reunion Island showing up to 18% uninfected individuals. These findings contribute in further understanding the dynamic interactions between the ecology of larval habitats and the structure of host microbiota, as well as providing additional information relative to the patterns of Wolbachia infection.

Fermented food products in the era of globalization: tradition meets biotechnology innovations.

Omics tools offer the opportunity to characterize and trace traditional and industrial fermented foods. Bioinformatics, through machine learning, and other advanced statistical approaches, are able to disentangle fermentation processes and to predict the evolution and metabolic outcomes of a food microbial ecosystem. By assembling microbial artificial consortia, the biotechnological advances will also be able to enhance the nutritional value and organoleptics characteristics of fermented food, preserving, at the same time, the potential of autochthonous microbial consortia and metabolic pathways, which are difficult to reproduce. Preserving the traditional methods contributes to protecting the hidden value of local biodiversity, and exploits its potential in industrial processes with the final aim of guaranteeing food security and safety, even in developing countries.

Plant biodiversity assessment through pollen DNA metabarcoding in Natura 2000 habitats (Italian Alps).

Monitoring biodiversity is of increasing importance in natural ecosystems. Metabarcoding can be used as a powerful molecular tool to complement traditional biodiversity monitoring, as total environmental DNA can be analyzed from complex samples containing DNA of different origin. The aim of this research was to demonstrate the potential of pollen DNA metabarcoding using the chloroplast trnL partial gene sequencing to characterize plant biodiversity. Collecting airborne biological particles with gravimetric Tauber traps in four Natura 2000 habitats within the Natural Park of Paneveggio Pale di San Martino (Italian Alps), at three-time intervals in 1 year, metabarcoding identified 68 taxa belonging to 32 local plant families. Metabarcoding could identify with finer taxonomic resolution almost all non-rare families found by conventional light microscopy concurrently applied. However, compared to microscopy quantitative results, Poaceae, Betulaceae, and Oleaceae were found to contribute to a lesser extent to the plant biodiversity and Pinaceae were more represented. Temporal changes detected by metabarcoding matched the features of each pollen season, as defined by aerobiological studies running in parallel, and spatial heterogeneity was revealed between sites. Our results showcase that pollen metabarcoding is a promising approach in detecting plant species composition which could provide support to continuous monitoring required in Natura 2000 habitats for biodiversity conservation.

Italian odonates in the Pandora's box: A comprehensive DNA barcoding inventory shows taxonomic warnings at the Holarctic scale.

The Odonata are considered among the most endangered freshwater faunal taxa. Their DNA-based monitoring relies on validated reference data sets that are often lacking or do not cover important biogeographical centres of diversification. This study presents the results of a DNA barcoding campaign on Odonata, based on the standard 658-bp 5' end region of the mitochondrial COI gene, involving the collection of 812 specimens (409 of which barcoded) from peninsular Italy and its main islands (328 localities), belonging to all the 88 species (31 Zygoptera and 57 Anisoptera) known from the country. Additional BOLD and GenBank data from Holarctic samples expanded the data set to 1,294 DNA barcodes. A multi-approach species delimitation analysis involving two distance (OT and ABGD) and four tree-based (PTP, MPTP, GMYC and bGMYC) methods was used to explore these data. Of the 88 investigated morphospecies, 75 (85%) unequivocally corresponded to distinct molecular operational units, whereas the remaining ones were classified as 'warnings' (i.e. showing a mismatch between morphospecies assignment and DNA-based species delimitation). These results are in contrast with other DNA barcoding studies on Odonata showing up to 95% of identification success. The species causing warnings were grouped into three categories depending on if they showed low, high or mixed genetic divergence patterns. The analysis of haplotype networks revealed unexpected intraspecific complexity at the Italian, Palearctic and Holarctic scale, possibly indicating the occurrence of cryptic species. Overall, this study provides new insights into the taxonomy of odonates and a valuable basis for future DNA and eDNA-based monitoring studies.

Toward a resolution of a taxonomic enigma: first genetic analyses of Paradoxornis webbianus and Paradoxornis alphonsianus (Aves: Paradoxornithidae) from China and Italy.

Paradoxornis webbianus and Paradoxornis alphonsianus naturally occur in South-East Asia. Due to a recent introduction, a mixed population currently occurs in northern Italy. A preliminary phylogeographic analysis using samples from Italy and China found little genetic differentiation between the two taxa and revealed the existence of two molecular lineages, sympatric in some part of China, that do not correspond to the morphological classification. Possible taxonomic changes and preliminary inferences on the relationships between Chinese and the Italian populations and on the likely provenance of the founders introduced in Italy are also discussed.

Identification of poisonous plants by DNA barcoding approach.

The plant exposures are one of the most frequent poisonings reported to poison control centres. The diagnosis of intoxicated patients is usually based on the morphological analysis of ingested plant portions; this procedure requires experience in systematic botany, because the plant identification is based on few evident traits. The objective of this research is to test DNA barcoding approach as a new universal tool to identify toxic plants univocally and rapidly. Five DNA barcode regions were evaluated: three cpDNA sequences (trnH-psbA, rpoB and matK) and two nuclear regions (At103 and sqd1). The performance of these markers was evaluated in three plant groups: (1) a large collection of angiosperms containing different toxic substances, (2) congeneric species showing different degrees of toxicity and (3) congeneric edible and poisonous plants. Based on assessments of PCR, sequence quality and resolution power in species discrimination, we recommend the combination of plastidial and nuclear markers to identify toxic plants. Concerning plastidial markers, matK and trnH-psbA showed consistent genetic variability. However, in agreement with CBOL Plant Working Group, we selected matK as the best marker, because trnH-psbA showed some problems in sequences sizes and alignments. As a final and relevant observation, we also propose the combination of matK with a nuclear marker such as At103 to distinguish toxic hybrids form parental species. In conclusion, our data support the claim that DNA barcoding is a powerful tool for poisonous plant identifications.

The hidden 'plant side' of insect novel foods: A DNA-based assessment.

In the context of novel foods, a category for which the market demand is increasing worldwide, the consumption of edible insects and related insect-based products is expected to grow in the next years. Insects represent an important source of energy for the human diet but there is a lack of scientific knowledge about their processing to ensure safe food items to the consumer. In this study we adopted a combined DNA-based approach to verify the identity of the declared species in five categories of commercial insect-based products (mt COI DNA barcoding) and to characterize plant declared ingredients or contaminants (nu ITS2 DNA metabarcoding) with particular attention to putative elements of allergenic concern belonging, for example to the insect rearing substrate. Moreover, the same approach has been used to assess its sensitivity to cases of contamination and counterfeits to insect flour with low cost (and potentially allergenic) vegetable flours like wheat and soybean. Results show the success of insect DNA barcoding authentication even for highly processed products. Furthermore, the DNA metabarcoding analysis revealed a high efficacy as a screening method to identify both plant ingredients and vegetal traces belonging to insect farming or possible adulteration events, also acting as an early warning strategy for the occurrence of allergens of human concern. This approach could support the development of new risk assessment procedures for novel foods by regulatory authorities to ensure their quality, safety, and acceptance which will become more required in order to face the challenge of feeding the world population in the next decades.

Rethinking Urban and Food Policies to Improve Citizens Safety After COVID-19 Pandemic.

The ongoing pandemic caused by the coronavirus disease 2019 (COVID-19) is literally changing the world. From December 2019 to date, more than 22 million cases have been reported worldwide and global health institutions are acting to slow down the virus transmission and are looking for possible prevention strategies in case of a new outbreak. As in other endemic or pandemic phenomena, the issues mostly covered by scientific and media attention are related to the diagnostic and therapeutic approach of COVID-19. However, a still neglected issue regards the adoption of a more systemic approach considering the close connection among the infection, the environment, and human behaviors, including the role of diet and urban management. To shed light on this issue, we brought together a faculty group involving experts in environment and biodiversity, food safety, human nutrition, and behavior, bioprospecting, as well as medical doctors having a deep knowledge of the complex historical relationship between humanity and vector-borne infections. Two main aspects emerged from the integrative overview of the current COVID-19 pandemic: (i) the scientific community should start sharing social actions and policy advocacy based on the assumption that human health strongly depends upon a sustainable exploitation of natural resources in populated areas; (ii) the specific strategic role of the cities in developing sustainable food systems and promoting healthy dietary patterns. Definitely, some priority issues should be addressed to achieve these goals, such as global efforts to increase food safety and security, which would benefit from urban and peri-urban agriculture enhancement, smallholder food producers support, and ecosystem services and local biodiversity maintenance.

Integrated taxonomy: traditional approach and DNA barcoding for the identification of filarioid worms and related parasites (Nematoda).

BACKGROUND: We compared here the suitability and efficacy of traditional morphological approach and DNA barcoding to distinguish filarioid nematodes species (Nematoda, Spirurida). A reliable and rapid taxonomic identification of these parasites is the basis for a correct diagnosis of important and widespread parasitic diseases. The performance of DNA barcoding with different parameters was compared measuring the strength of correlation between morphological and molecular identification approaches. Molecular distance estimation was performed with two different mitochondrial markers (coxI and 12S rDNA) and different combinations of data handling were compared in order to provide a stronger tool for easy identification of filarioid worms. RESULTS: DNA barcoding and morphology based identification of filarioid nematodes revealed high coherence. Despite both coxI and 12S rDNA allow to reach high-quality performances, only coxI revealed to be manageable. Both alignment algorithm, gaps treatment, and the criteria used to define the threshold value were found to affect the performance of DNA barcoding with 12S rDNA marker. Using coxI and a defined level of nucleotide divergence to delimit species boundaries, DNA barcoding can also be used to infer potential new species. CONCLUSION: An integrated approach allows to reach a higher discrimination power. The results clearly show where DNA-based and morphological identifications are consistent, and where they are not. The coherence between DNA-based and morphological identification for almost all the species examined in our work is very strong. We propose DNA barcoding as a reliable, consistent, and democratic tool for species discrimination in routine identification of parasitic nematodes.