Proso Millet (Panicum miliaceum L.) as Alternative Source of Starch and Phenolic Compounds: A Study on Twenty-Five Worldwide Accessions.

Proso millet has been proposed as an effective anti-diabetic food thanks to the combined action of polyphenols and starch. This study aimed to characterize the chemical composition of twenty-five accessions, in order to enhance this cereal as an alternative to available starch for food applications or to propose new food ingredients with health benefits. Proso millet contained a high percentage of starch, reaching values of 58.51%. The amylose content showed high variability, with values ranging from 1.36 to 42.70%, and significantly higher values were recorded for the white accessions than for the colored ones. High-resistant starch content (13.41-26.07%) was also found. The HPLC-MS analysis showed the same phenolic pattern in all the samples. Cinnamic acids are the most abundant compounds and significant differences in their total content were found (0.69 to 1.35 mg/g DW), while flavonoids were only detected in trace amounts. Statistical results showed significantly higher antiradical activity in the colored accessions than in the white ones.

Challenges and a call to action for protecting European red wood ants.

Red wood ants (RWAs) are a group of keystone species widespread in temperate and boreal forests of the Northern Hemisphere. Despite this, there is increasing evidence of local declines and extinctions. We reviewed the current protection status of RWAs throughout Europe and their International Union for the Conservation of Nature (IUCN) threat classification. Only some RWA species have been assessed at a global scale, and not all national red lists of the countries where RWAs are present include these species. Different assessment criteria, inventory approaches, and risk categories are used in different countries, and data deficiency is frequent. Legislative protection is even more complex, with some countries protecting RWAs implicitly together with the wildlife fauna and others explicitly protecting the whole group or particular species. This complexity often occurs within countries, for example, in Italy, where, outside of the Alps, only the introduced species are protected, whereas the native species, which are in decline, are not. Therefore, an international, coordinated framework is needed for the protection of RWAs. This first requires that the conservation target should be defined. Due to the similar morphology, complex taxonomy, and frequent hybridization, protecting the entire RWA group seems a more efficient strategy than protecting single species, although with a distinction between autochthonous and introduced species. Second, an update of the current distribution of RWA species is needed throughout Europe. Third, a protection law cannot be effective without the collaboration of forest managers, whose activity influences RWA habitat. Finally, RWA mounds offer a peculiar microhabitat, hosting a multitude of taxa, some of which are obligate myrmecophilous species on the IUCN Red List. Therefore, RWAs' role as umbrella species could facilitate their protection if they are considered not only as target species but also as providers of species-rich microhabitats.

Las hormigas rojas de la madera (HRM) conforman un grupo de especies clave con amplia distribución en los bosques templados y boreales del Hemisferio Norte. A pesar de lo anterior, cada vez hay más evidencia de su declinación y extinción local. Revisamos el estado actual de protección de las HRM en toda Europa y su clasificación en la Lista Roja de la Unión Internacional para la Conservación de la Naturaleza (UICN). Sólo se han evaluado algunas especies de HRM a escala mundial y no todas las listas rojas nacionales de los países con presencia de HRM incluyen a estas especies. Los diferentes países usan criterios de evaluación, estrategias de inventario y categorías de riesgo distintos, además de que la información deficiente es habitual. La protección legislativa es todavía más compleja pues algunos países protegen implícitamente a las HRM junto con la fauna silvestre y otros protegen explícitamente a todo el grupo o a una especie particular. Esta complejidad ocurre a menudo en los países (por ejemplo: Italia) en donde, fuera de los Alpes, sólo se protege a las especies introducidas, mientras a las especies nativas, que están declinando, no se les protege. Por lo tanto, se requiere un marco de trabajo internacional y coordinado para proteger a las HRM. Esto necesita primero que se defina el objetivo de conservación. Ya que las HRM tienen similitudes morfológicas, una taxonomía compleja e hibridación frecuente, la protección del grupo completo, con la distinción entre las especies autóctonas y las introducidas, parece ser una estrategia más eficiente que la protección de una sola especie. Segundo, se debe actualizar la distribución actual de las HRM en Europa. Tercero, una ley de protección no puede ser efectiva sin la colaboración de los gestores forestales, cuya actividad influye sobre el hábitat de las HRM Finalmente, los montículos de las HRM ofrecen un microhábitat peculiar pues hospedan a una multitud de taxones, algunos de los cuales son especies mirmecófilas obligadas presentes en la Lista Roja de la UICN. Así, el papel de las HRM como especie paraguas podría facilitar su protección si se les considera no sólo como especies diana sino también como proveedoras de microhábitats con riqueza de especies.

Genome wide association study of agronomic and seed traits in a world collection of proso millet (Panicum miliaceum L.).

BACKGROUND: The climate crisis threatens sustainability of crop production worldwide. Crop diversification may enhance food security while reducing the negative impacts of climate change. Proso millet (Panicum milaceum L.) is a minor cereal crop which holds potential for diversification and adaptation to different environmental conditions. In this study, we assembled a world collection of proso millet consisting of 88 varieties and landraces to investigate its genomic and phenotypic diversity for seed traits, and to identify marker-trait associations (MTA). RESULTS: Sequencing of restriction-site associated DNA fragments yielded 494 million reads and 2,412 high quality single nucleotide polymorphisms (SNPs). SNPs were used to study the diversity in the collection and perform a genome wide association study (GWAS). A genotypic diversity analysis separated accessions originating in Western Europe, Eastern Asia and Americas from accessions sampled in Southern Asia, Western Asia, and Africa. A Bayesian structure analysis reported four cryptic genetic groups, showing that landraces accessions had a significant level of admixture and that most of the improved proso millet materials clustered separately from landraces. The collection was highly diverse for seed traits, with color varying from white to dark brown and width spanning from 1.8 to 2.6 mm. A GWAS study for seed morphology traits identified 10 MTAs. In addition, we identified three MTAs for agronomic traits that were previously measured on the collection. CONCLUSION: Using genomics and automated seed phenotyping, we elucidated phylogenetic relationships and seed diversity in a global millet collection. Overall, we identified 13 MTAs for key agronomic and seed traits indicating the presence of alleles with potential for application in proso breeding programs.

Effectiveness of a Khorasan Wheat-Based Replacement on Pain Symptoms and Quality of Life in Patients with Fibromyalgia.

OBJECTIVE: To investigate the effects of a replacement diet with Khorasan wheat products in patients with fibromyalgia, in comparison with a similar replacement diet with control products made from organic semi-whole-grain modern wheat. DESIGN: Randomized, double-blinded crossover trial. SETTING: Outpatient clinic. SUBJECTS: Twenty subjects (19 female and one male, mean age = 48.9 ± 12.3 years) with fibromyalgia. METHODS: Participants were randomly assigned to consume either Khorasan or control wheat products (pasta, bread, crackers, biscuits) for eight weeks and then crossed. Validated self-administered questionnaires were collected from each subject at the beginning and end of each intervention period. RESULTS: A general linear model for repeated measurement, adjusted for potential confounders, showed that the overall score reported from each questionnaire improved after both intervention and control periods, but the effect was more evident after the intervention with Khorasan. In particular, a statistically significant difference in Widespread Pain Index (WPI) + Severity Scale (SS) and Functional Outcome of Sleep Questionnaire (FOSQ) was observed, which decreased significantly by 21.5% and 11.7% respectively, only after the Khorasan period, while no statistically significant variations were reported after the control period. Similarly, FM Impact Questionnaire scores decreased significantly only after the Khorasan period, with a reduction that was significantly different between the intervention and control periods (-22.5% vs -0.3%, P = 0.037). The improvement was even greater in people with higher symptom severity. CONCLUSIONS: A dietary intervention with Khorasan wheat products seems to benefit patients with fibromyalgia, especially those with greater symptom severity.

Sub-lethal doses of imidacloprid alter food selection in the invasive garden ant Lasius neglectus.

Despite several restrictions to their use, neonicotinoid insecticides are still widely employed worldwide. Residual sub-lethal amounts of these chemicals can have detrimental effects on the behavior of non-target insects. Toxic effects on economically important species such as bees have been widely documented, but less is known about their toxic action on other social insects, such as ants. In this study, we assessed the effect of different sub-lethal doses of the neonicotinoid imidacloprid on the ability of colonies of the invasive ant Lasius neglectus to select the most profitable resource. We used Y-shaped mazes having an imidacloprid-polluted or an unpolluted sucrose solution on the two branches. Two sucrose (0.1 M, 0.5 M) and two imidacloprid (1 µg/ml, 10 µg/ml) concentrations were used. In parallel, we evaluated the marking activity of foragers who fed on the same solutions. We found that the 0.1 M sugar solution polluted with 1 µg/ml imidacloprid was significantly more frequently selected in binary choices experiments than the unpolluted resource. Moreover, the ingestion of the same combination of sugar and imidacloprid significantly increased the marking rate of foragers. The higher concentration of the pollutant had lower effects, probably because of the hormesis phenomenon. Results suggest that the lower sub-lethal dose of imidacloprid can lead ants to select again the polluted resource. This "active" selection of the pollutant may magnify the negative effects on the colonies. Due to their ecological role, any impairment of ant survival or behavior may have detrimental cascade effects on the whole ecosystem.

Keep Your Eggs Away: Ant Presence Reduces Ceratitis capitata Oviposition Behaviour through Trait-Mediated Indirect Interactions.

Alternative methods to achieve sustainable agricultural production while reducing the use of chemical pesticides, such as biological control, are increasingly needed. The exploitation of trait-mediated indirect interactions (TMIIs), in which pests modify their behavior in response to some cues (e.g., pheromones and other semiochemicals) to avoid predation risk, may be a possible strategy. In this study, we tested the effect of TMIIs of two Mediterranean ant species, Crematogaster scutellaris and Tapinoma nigerrimum, on the oviposition behaviour of Ceratitis capitata (Diptera: Tephritidae), one of the world's most economically damaging pests, which attacks fruits. For each ant species, we performed choice experiments using ant-scented and control plums, counting the time spent by medflies on fruits and the number of pupae emerging from them. Results of both ant species tests showed a significantly shorter time spent by ovipositing medflies on ant-exposed plums and a lower number of pupae, when compared to the control group. Our findings highlighted that the semiochemicals released by ants on plums triggered an avoidance behaviour by medfly females, leading to lower oviposition rates. This study contributes to the understanding of indirect ant-pest interactions in Mediterranean agricultural settings and points out the potential of utilising ant-borne semiochemicals in sustainable IPM strategies.

Comparative Analysis of Facial Coloration between Introduced and Source Populations of the Red Wood Ant Formica paralugubris.

The variation in the typical black-reddish color of red wood ants (Formica rufa group) has been recently suggested as a good indicator of habitat quality, being dependent on environmental conditions. However, the relative contribution of external factors and heritability in shaping this trait is poorly investigated. In this study, we compared the facial coloration of workers from four introduced populations of Formica paralugubris with those of the two Alpine populations from which they had been taken. We used a Relative Warp Analysis to describe the variations in the shape of this trait. We expected each introduced population to be more similar to its population of origin if the color pattern was predominantly genetically determined. On the contrary, due to the considerable differences in habitat type and climate between the Alps and the Apennines, we expected to observe differences between the introduced population and their origin population if the coloration was mostly environmentally determined. With one exception that we discuss, the results showed that ants from the two source populations had different phenotypes, and that the introduced populations had a shape similar to the population of origin, suggesting a stable genetic background. Surprisingly, the habitat type seems to have a less clear effect, even if within-population differences suggest the influence of very localized environmental factors. Finally, we found that the facial coloration shape is affected by the ant's size, a result in line with previous studies.

Acute and chronic ingestion of polyethylene (PE) microplastics has mild effects on honey bee health and cognition.

The massive use of plastic has contributed to huge quantities of hazardous refuse at a global scale and represents one of the most prominent issues of the Anthropocene. Microplastics (MPs) have been detected in almost all environments and pose a potential threat to a variety of plant and animal species. Many studies have reported a variety of effects, from negligible to detrimental, of MPs to aquatic organisms. Conversely, much less is known about their effect on terrestrial biota, and particularly on animal behavior and cognition. We assessed the oral toxicity of polyethylene (PE) MPs at three different concentrations (0.5, 5, and 50 mg L-1), and at different timescales (1 day and 7 days of exposure) and tested for their effects on survival, food intake, sucrose responsiveness, habituation to sucrose and appetitive olfactory learning and memory in the honey bee Apis mellifera. We found that workers were not completely unaffected by acute and prolonged ingestion of this polymer. A significant effect of PE on bee mortality was found for the highest concentration but not for lower ones. PE affected feeding behavior in a concentration-dependent manner, with bees consuming more food than controls when exposed to low concentration PE. Regarding our behavioral and cognitive experiments, the high concentration PE was found to affect only bees' ability to respond consistently to sucrose but not sucrose sensitivity, habituation to sucrose or learning and memory abilities, even for prolonged exposure to PE. While these last results may look somewhat encouraging, we discussed why caution is warranted before ruling out the possibility that PE particles at environmental concentrations are harmful to honey bees.

Intraspecific Relationships and Nest Mound Shape Are Affected by Habitat Features in Introduced Populations of the Red Wood Ant Formica paralugubris.

Ants belonging to the Formica rufa group build large nest mounds, which aid their survival during severe winters. We investigated whether different environmental features of the habitats affected the nest mound shape and the population structure. We assessed the shape of all the nest mounds and mapped inter-nest trails connecting mounds for three imported populations of Formica paralugubris in three forest habitats: fir-dominated, beech-dominated, and a mixture of fir and beech. Single-nest mounds were averagely smaller and flatter in the beech-dominated forest, probably because of lighter building materials. Nonetheless, by summing the volumes of all interconnected nests, the size was similar among all three sites. In fir- and beech-dominated forests, large nests were also central in the networks, suggesting a central place foraging model with these nests as reference. We finally performed aggression tests, and found that aggressiveness was significantly higher among nests belonging to the same population than between populations. The results highlight the plasticity of the species to adapt nest and colony structure to different environments. Additionally, it appears that none of these populations is unicolonial, as observed in various alpine sites, there and the observed patterns of aggression are coherent with the 'nasty neighbor' effect.

Assessing Molecular Diversity in Native and Introduced Populations of Red Wood Ant Formica paralugubris.

The Formica rufa group comprises several ant species which are collectively referred to as "red wood ants" and play key roles in boreal forest ecosystems, where they are ecologically dominant and greatly influence habitat dynamics. Owing to their intense predatory activity, some of these species are used as biocontrol agents against several forest insect pests and for this aim in Italy, nearly 6000 ant nests were introduced from their native areas in the Alps to several Appeninic sites during the last century. In this work, we assessed and compared the genetic variability and structure of native and introduced populations of F. paralugubris, thus evaluating the extent of genetic drift that may have occurred since the time of introduction, using amplified fragment length polymorphism (AFLP) markers. PCR amplification with a fam_EcoRI-TAC/MseI-ATG primers combination produced a total of 147 scorable bands, with 17 identified as outlier loci. The genetic variation was higher in the introduced population compared to the native ones that, on the other hand, showed a higher diversity between nests. AMOVA results clearly pointed out that the overall genetic structure was dominated by among-worker variation, considering all populations, the Alpine vs. Apennine groups and the comparison among native and related introduced populations (all ranging between 77.84% and 79.84%). Genetic analyses unveiled the existence of six main different groups that do not entirely mirror their geographic subdivision, pointing towards a wide admixture between populations, but, at the same time, rapid diversification of some Apennine populations. Future studies based on high-throughput genomic methods are needed to obtain a thorough understanding of the effects of environmental pressure on the genetic structure and mating system of these populations.

The pivotal role of cultivar affinity to arbuscular mycorrhizal fungi in determining mycorrhizal responsiveness to water deficit.

Arbuscular mycorrhizal fungi (AMF) have gained remarkable importance, having been proved to alleviate drought stress-induced damage in wheat due to their ability to ameliorate plant water use efficiency and antioxidant enzyme activity. However, despite the current relevance of the topic, the molecular and physiological processes at the base of this symbiosis never consider the single cultivar affinity to mycorrhization as an influencing factor for the metabolic response in the AMF-colonized plant. In the present study, the mycorrhizal affinity of two durum wheat species (T. turgidum subsp. durum (Desf.)) varieties, Iride and Ramirez, were investigated. Successively, an untargeted metabolomics approach has been used to study the fungal contribution to mitigating water deficit in both varieties. Iride and Ramirez exhibited a high and low level of mycorrhizal symbiosis, respectively; resulting in a more remarkable alteration of metabolic pathways in the most colonised variety under water deficit conditions. However, the analysis highlighted the contribution of AMF to mitigating water deficiency in both varieties, resulting in the up- and down-regulation of many amino acids, alkaloids, phenylpropanoids, lipids, and hormones.

Genetic variability assessment of 127 Triticum turgidum L. accessions for mycorrhizal susceptibility-related traits detection.

Positive effects of arbuscular mycorrhizal fungi (AMF)-wheat plant symbiosis have been well discussed by research, while the actual role of the single wheat genotype in establishing this type of association is still poorly investigated. In this work, the genetic diversity of Triticum turgidum wheats was exploited to detect roots susceptibility to AMF and to identify genetic markers in linkage with chromosome regions involved in this symbiosis. A tetraploid wheat collection of 127 accessions was genotyped using 35K single-nucleotide polymorphism (SNP) array and inoculated with the AMF species Funneliformis mosseae (F. mosseae) and Rhizoglomus irregulare (R. irregulare), and a genome-wide association study (GWAS) was conducted. Six clusters of genetically related accessions were identified, showing a different mycorrhizal colonization among them. GWAS revealed four significant quantitative trait nucleotides (QTNs) involved in mycorrhizal symbiosis, located on chromosomes 1A, 2A, 2B and 6A. The results of this work enrich future breeding activities aimed at developing new grains on the basis of genetic diversity on low or high susceptibility to mycorrhization, and, possibly, maximizing the symbiotic effects.

Combining metal and stable isotope analyses to disentangle contaminant transfer in a freshwater community dominated by alien species.

Freshwater ecosystems are negatively impacted by various pollutants, from agricultural, urban and industrial wastewater, with metals being one of the largest concerns. Moreover, freshwater ecosystems are often affected by alien species introductions that can modify habitats and trophic relationships. Accordingly, the threat posed by metals interacts with those by alien species, since the latter can accumulate and transfer these substances across the food web to higher trophic levels. How metals transfer within such communities is little studied. We analysed the concentration of 14 metals/metalloids (Al, As, Cd, Co, Cr, Cu, Fe, Hg, Mg, Mn, Ni, Pb, Se, Zn, hereafter 'metal(s)') of eight fish and three crustacean species co-existing in the Arno River (Central Italy), most of which were alien. To assess the pathway of contaminants within the community, we coupled metal analysis with carbon and nitrogen stable isotope analysis derived from the same specimens. Crustaceans showed higher metal concentration than fish, except for Cd, Hg and Se that were higher in fish. We found evidence of trophic transfer for six metals (Cd, Cr, Hg, Mg, Se, Zn). Additionally, ontogenetic differences and differences among various fish tissues (muscle, liver, and gills) were found in metals concentration. Considerable biomagnification along the trophic chain was found for Hg, while other metals were found to biodilute. Using stable isotopes and Hg as a third diet tracer, we refined the estimations of consumed preys in the diet previously reconstructed with stable isotope mixing models. Alien species reach high biomass and can both survive to and accumulate high pollutants concentrations, potentially posing a risk for their predators and humans. A combined effect of environmental filtering and increased competition may potentially contribute to the disappearance of native species with lower tolerances.

Small-scale drivers on plant and ant diversity in a grassland habitat through a multifaceted approach.

Semi-natural grasslands are characterized by high biodiversity and require multifaceted approaches to monitor their biodiversity. Moreover, grasslands comprise a multitude of microhabitats, making the scale of investigation of fundamental importance. Despite their wide distribution, grasslands are highly threatened and are considered of high conservation priority by Directive no. 92/43/EEC. Here, we investigate the effects of small-scale ecological differences between two ecosites present within the EU habitat of Community Interest of semi-natural dry grasslands on calcareous substrates (6210 according to Dir. 92/43/EEC) occurring on a Mediterranean mountain. We measured taxonomic and functional diversity of plant and ant communities, evaluating the differences among the two ecosites, how these differences are influenced by the environment and whether vegetation affects composition of the ant community. Our results show that taxonomic and functional diversity of plant and ant communities are influenced by the environment. While vegetation has no effect on ant communities, we found plant and ant community composition differed across the two ecosites, filtering ant and plant species according to their functional traits, even at a small spatial scale. Our findings imply that small-scale monitoring is needed to effectively conserve priority habitats, especially for those that comprise multiple microhabitats.

Exploring the Gut Microbiome Alteration of the European Hare (Lepus europaeus) after Short-Term Diet Modifications.

This study aimed to characterise the gut microbiome composition of European hares (Lepus europaeus) and its potential changes after a short-term diet modification. The high sensitivity of European hare to habitat changes makes this species a good model to analyse possible alterations in gut microbiome after the introduction of additional nourishment into the diet. In total, 20 pairs were chosen for the experiments; 10 pairs formed the control group and were fed with standard fodder. The other 10 pairs represented the experimental group, whose diet was integrated with apples and carrots. The DNA from fresh faecal pellets collected after 4 days from the start of the experiment was extracted and the V3-V4 hypervariable regions were amplified and sequenced using the Illumina MiSeq® platform. The obtained amplicon sequence variants were classified into 735 bacterial genera belonging to 285 families and 36 phyla. The control and the experimental groups appeared to have a homogenous dispersion for the two taxonomic levels analysed with the most abundant phyla represented by Bacteroidetes and Firmicutes. No difference between control and experimental samples was detected, suggesting that the short-term variation in food availability did not alter the hares' gut microbiome. Further research is needed to estimate significant time threshold.

Palatability of glyphosate in ants: a field experiment reveals broad acceptance of highly polluted solutions in a Mediterranean ant.

Glyphosate is a systemic herbicide still used in many countries, though there are several known detrimental effects on animals. Previous studies concerning its effects on social insects are available, but they are primarily focused on honeybees; little is known about the interactions of this compound with ants. Here, we assessed whether different concentrations of glyphosate can be perceived by ant workers and to what extent. As a model species, we used the Mediterranean ant Crematogaster scutellaris, commonly found in agroecosystems. We performed 3000 individual tests of acceptance using ten different solutions of various concentrations of the herbicide. Half of the solutions contained added sucrose in order to test the possible masking effect of the sugar taste on glyphosate. We used comparable glyphosate concentrations to those previously used in other studies on social insects or suggested by the producer. We found that the acceptance of the solutions decreased as the concentration of the herbicide increased. However, a significant percentage of ants drank the solutions with concentrations up to dozens of times higher than those inducing toxic effects in bees. In light of these results, we urge further assessment of the effects of glyphosate on ants, particularly because the food ingested by workers is transferred to the brood and queens, posing a potential threat to the health of the entire colony. Surprisingly, we did not record any difference in acceptance between solutions with and without sugar; this point is discussed regarding drought stress.

Serpentine soils affect heavy metal tolerance but not genetic diversity in a common Mediterranean ant.

Natural habitats with serpentine soils are rich in heavy metal ions, which may significantly affect ecological communities. Exposure to metal pollutants results, for instance, in a reduction of population genetic diversity and a diffused higher tolerance towards heavy metals. In this study, we investigated whether chronic exposure to metals in serpentine soils affect accumulation patterns, tolerance towards metal pollutants, and genetic diversity in ants. In particular, we studied colonies of the common Mediterranean ant, Crematogaster scutellaris, along a contamination gradient consisting of two differently contaminated forests and a reference soil with no geogenic contamination. We first evaluated the metal content in both soil and ants' body. Then, we tested for tolerance towards metal pollutants by evaluating the mortality of ants fed with nickel (Ni) solutions of increasing concentrations. Finally, differences in genetic diversity among ants from different areas were assessed using eight microsatellite loci. Interestingly, a higher tolerance to nickel solutions was found in ants sampled in sites with intermediate levels of heavy metals. This may occur, because ants inhabiting strongly contaminated areas tend to accumulate higher amounts of contaminants. Additional ingestion of toxicants beyond the saturation threshold would lead to death. There was no difference in the genetic diversity among ant colonies sampled in different sites. This was probably the result of queen mediated gene flow during nuptial flights across uncontaminated and contaminated areas of limited geographical extent.