Annamaria Giorgi

Effects of the Diet on the Microbiota of the Red Palm Weevil (Coleoptera: Dryophthoridae)

Rhynchophorus ferrugineus, also known as the red palm weevil, is regarded as the major pest of palm trees. Although studies of the microbiota associated with this species have been performed in recent years, little attention has been dedicated to the influence of the diet in shaping the host bacterial community. Here, we investigated the influence of food sources (i.e. palm tissues vs apple based substrate) on the microbial diversity associated with RPW, which was compared with the microbiota associated with wild individuals of the sister species Rhynchophorus vulneratus. The bacterial characterization was performed using a culture independent approach, i.e. the 16S rRNA pyrotag, and a culture dependent approach for a subset of the samples, in order to obtain bacterial isolates from RPW tissues. The bacterial community appeared significantly influenced by diet. Proteobacteria resulted to be the most abundant clade and was present in all the specimens of the three examined weevil groups. Within Proteobacteria, Enterobacteriaceae were identified in all the organs analysed, including hemolymph and reproductive organs. The apple-fed RPWs and the wild R. vulneratus showed a second dominant taxon within Firmicutes that was scarcely present in the microbiota associated with palm-fed RPWs. A comparative analysis on the bacteria associated with the palm tissues highlighted that 12 bacterial genera out of the 13 identified in the plant tissues were also present in weevils, thus indicating that palm tissues may present a source for bacterial acquisition.

Influence of environmental factors on composition of phenolic antioxidants of Achillea collina Becker ex Rchb.

Effects of environmental growth conditions on the antioxidant capacity, total phenolic content and composition of Achillea collina Becker ex Rchb. were investigated. Methanol extracts and infusions obtained from leaves and inflorescences of plants cultivated in the Italian Alps at two different altitudes (600 and 1050 m a.s.l.) were evaluated. Infusions exhibited the highest antioxidant capacity (1/IC50 values from 4.35 ± 0.72 to 8.90 ± 0.74), total phenolic content (from 31.39 ± 4.92 to 49.36 ± 5.70 mg gallic acid equivalents (GAE) g−1 DW), chlorogenic acid (from 9.21 ± 1.52 to 31.27 ± 6.88 mg g−1 DW), 3,5-di-O-caffeoylquinic acid (from 12.28 ± 3.25 to 25.13 ± 1.99 mg g−1 DW) and 4,5-di-O-caffeoylquinic acid (from 7.38 ± 1.01 to 12.78 ± 2.61 mg g−1 DW) content. Climate (as influenced by altitude) was shown to be the main environmental factor influencing yarrow composition and properties. Leaf extracts from the higher experimental site showed a 2–4-fold increase of chlorogenic acid level. Achillea collina can be considered as a very good source of bioactive phenolic compounds, and growing it at high altitude may constitute an effective way to significantly enhance its quality for both medicinal and nutritional uses.

Antioxidant and cytoprotective properties of infusions from leaves and inflorescences of Achillea collina Becker ex Rchb.

Plants are the main source of molecules with antioxidant and radical scavenging properties that aid the natural defence systems of cells and may be involved in the preservation of human health, particularly preventing all the physiopathological conditions where oxidative damage is a hallmark. Achillea collina Becker ex Rchb. is a medicinal plant of the Achillea millefolium aggregate (yarrow) traditionally used, particularly in mountain areas, as an infusion or alcohol extract for its digestive, antiinflammatory, analgesic, antipyretic and wound healing properties. The aim of this study was to investigate the antioxidant capacity and cytoprotective activity against oxidative stress of infusions obtained from the leaves and inflorescences of Achillea collina Becker ex Rchb., assessed by chemical (free radical scavenging activity by DPPH and Folin Ciocalteu assay) and biological assays (in vitro model of cytotoxicity and lipid peroxidation in PC12 cells line). Infusions of leaves had the highest antioxidant properties and cytoprotective activity. The antioxidant capacity was significantly correlated with the total phenolic content but not with the cytoprotective profile. Achillea collina Becker ex Rchb. has good antioxidant and cytoprotective properties, suggesting further investigations on its chemical composition and potential health value, particularly for traditionally prepared infusions of leaves. Copyright

Metamicrobiomics in herbivore beetles of the genus Cryptocephalus (Chrysomelidae): toward the understanding of ecological determinants in insect symbiosis.

The Cryptocephalus marginellus (Coleoptera: Chrysomelidae) complex is composed by six species that are supposed to have originated by events of allo‐ or parapatric speciation. In the present study we investigated the alternative hypotheses that the bacterial communities associated with six populations of this species complex are shaped by environmental factors, or reflect the proposed pattern of speciation. The microbiota associated with the six populations, from five species of the complex, have been characterized through 16S rRNA pyrotag sequencing. Based on a 97% sequence similarity threshold, data were clustered into 381 OTUs, which were analyzed using a variety of diversity indices. The microbiota of C. acquitanus and C. marginellus (Calanques) were the most diverse (over 100 OTUs), while that from C. zoiai yielded less bacterial diversity (45 OTUs). Taxonomic assignment revealed Proteobacteria, Tenericutes and Firmicutes as the dominant components of these beetles’ microbiota. The most abundant genera were Ralstonia, Sphingomonas, Rickettsia, and Pseudomonas. Different strains of Rickettsia were detected in C. eridani and C. renatae. The analysis of β‐diversity revealed high OTU turnover among the populations of C. marginellus complex, with only few shared species. Hierarchical clustering taking into account relative abundances of OTUs does not match the phylogeny of the beetles, therefore we hypothesize that factors other than phylogenetic constraints play a role in shaping the insects’ microbiota. Environmental factors that could potentially affect the composition of bacterial communities were tested by fitting them on the results of a multi‐dimensional scaling analysis. No significant correlations were observed towards the geographic distances or the host plants, while the composition of the microbiota appeared associated with altitude. The metabolic profiles of the microbiotas associated with each population were inferred from bacterial taxonomy, and interestingly, the obtained clustering pattern was consistent with the host phylogeny.

The relationships between phenolic content, pollen diversity, physicochemical information and radical scavenging activity in honey.

Honey is rich in different secondary plant metabolites acting as natural antioxidants and contributing to human health. Radical scavenging activity (RSA) is related to antioxidant activity, while the correlation between the phenolic content and RSA is often weak. Consequently, exclusive information on phenolics is often insufficient to qualify the RSA and the health promoting effects of honey. The paper deals with a case study of honey samples originating from the alpine areas of Italy’s Lombardia and Veneto regions and realized by standard physicochemical and statistical analytical methods. In pure honey, the total phenolic content and the RSA were measured in spectrophotometric tests with the 2,2-diphenyl-1-picrylhydrazyl (DPPH·) free radical and Folin-Ciocalteu assays, respectively. Melissopalynological data was used to qualify pollen diversity through rank-frequency curves separating the samples into two groups. On the basis of physicochemical data, the samples were analyzed through multivariate classification and ranking procedures resulting in the identification of an outlier. Elimination of the outlier produced a high correlation between the total phenolic content and RSA in the two pollen diversity groups. The case study suggests that, after disregarding outliers, the RSA activity can be satisfactorily qualified on the basis of phenolics with pollen diversity as a covariate.

Melissopalynological and Volatile Compounds Analysis of Buckwheat Honey from Different Geographical Origins and Their Role in Botanical Determination

Volatile organic compounds (VOCs) have been proposed as one of the main factors for differentiating honeys from different botanical/floral origins. In this work, we investigated the volatile profile of honeys, commercially labeled as buckwheat honeys, from the Alps and its relationship with melissopalynological investigation. The results showed that buckwheat honey samples that contained, to different extents, buckwheat pollen grains on melissopalynological analyses showed similar VOCs profiles, distinguishing them from the other honey floral types analyzed. Among VOCs identified, 3-methylbutanal, butanoic acid, pentanoic acid, and isovaleric acid were considerably greater in the buckwheat honey samples from the Alps. Other compounds were identified only in the honeys containing buckwheat pollen grains such as 3-methyl-2-buten-1-ol, 2-butanone, 2-hydroxy-3-pentanone, 4-methylpentanoic acid, 4-pentanoic acid, butanal, 2-methylbutanal, pentanal, dihydro-2-methyl-3(2H)-furanone, 5-methylfurfural, and cis-linalool oxide. These compounds give to buckwheat honey its characteristic aromatic and organoleptic properties and may be considered interesting as potential “variety markers” for botanical determination.

HS-SPME-GC/MS analysis of the volatile compounds of Achillea collina : Evaluation of the emissions fingerprint induced by Myzus persicae infestation

A Headspace Solid-phase Microextraction (HS-SPME) method combined with Gas Chromatography-Mass Spectrometry (GC/MS) was developed and optimized to extrat and analyze the volatile compounds of aerial parts of Achillea collina Becker ex Rchb. and to investigate the effect of the phlem feeding aphid Myzus persicae Sulzer on the Volatile Organic Compounds (VOCs) emitted by the infested plants. The extraction of 1 g of powdered freeze dried plant samples for 120 min at 30°C using divinylbenzene-carbowax-polydimethylsiloxane (DVB/CAR/PDMS) fiber showed the highest area counts for the majority of the volatile compounds. Overall, 62 and 80 volatile compounds were detected in control and infested plant samples respectively. In A. collina infested plants, we observed a great increase in both monoterpenes and sesquiterpenes fractions. Several changes among alcohols also occurred, particularly regarding Z-3-hexen-1-ol, E-3-hexen-1-ol and E-2-hexen-1-ol proposing these compounds as herbivore-induces plant volatiles (HIPVs). New perspective for agricultural practice may derive from the opportunity to identify novel herbivores-induced plant VOCs active as plant protection agents.

Secondary metabolites and antioxidant capacities of Waldheimia glabra (Decne.) Regel from Nepal.

BACKGROUND The phenolic content, volatile compound fingerprint and antioxidant capacity of Waldheimia glabra (Decne.) Regel (Asteraceae), a wild plant from the Himalayan mountains used in Sherpa religious rituals and in traditional medicine, were determined for the first time to investigate its suitability as a source of natural antioxidants. RESULTS The total phenolic content and antioxidant capacity of dried and powdered plants from two different altitudes (5000 and 5200 m above sea level) in Khumbu Valley, Nepal were estimated by the Folin-Ciocalteu and 2,2-diphenyl-1-picrylhydrazyl radical methods respectively. The volatile organic compound (VOC) fingerprints from leafing plants (in vivo and in situ VOC sampling) and from dried plant samples were evaluated by headspace solid phase microextraction. The results indicated that W. glabra was a good source of phenolics with antioxidant properties. A correlation between total phenolic content and antioxidant capacity was found (r² = 0.9111). Seychellene, a volatile compound that characterises patchouli oil, was detected for the first time in a species of the Asteraceae family. CONCLUSION Waldheimia glabra was proved to be a good source of antioxidants, partially supporting its traditional uses. Further studies aimed at deepening knowledge on W. glabra bioactivity and its possible uses will be of interest, considering the lack of proper scientific awareness regarding these aspects.

Secondary Metabolite Profile, Antioxidant Capacity, and Mosquito Repellent Activity of Bixa orellana from Brazilian Amazon Region

The Brazilian flora was widely used as source of food and natural remedies to treat various diseases. Bixa orellana L. (Bixaceae), also known as annatto, urucu, or achiote, is a symbol for the Amazonian tribes that traditionally use its seeds as coloured ink to paint their bodies for religious ceremonies. The aim of this study was to investigate the volatile organic compounds (VOCs) profile of B. orellana fresh fruits (in vivo sampled), dried seeds, wood, bark, and leaves analyzed with Headspace solid-phase microextraction coupled with gas chromatography and mass spectrometry. A screening on phenolic content (the Folin-Ciocalteu assay) and antiradical activity (DPPH assay) of seeds was also conducted. In addition, the repellent properties of seed extracts against Aedes aegypti L. were investigated. Volatile compounds detected in B. orellana samples consisted mainly of sesquiterpenes, monoterpenes, and arenes: α-humulene is the major volatile compound present in seed extracts followed by D-germacrene, γ-elemene, and caryophyllene. B. orellana proved to be a good source of antioxidants. Preliminary data on repellency against A. aegypti of three different dried seed extracts (hexane, ethanol, and ethanol/water) indicated a significant skin protection activity. A protection of 90% and 73% for hexane and ethanol/water extracts was recorded.

Volatile Fingerprint of Italian Populations of Orchids Using Solid Phase Microextraction and Gas Chromatography Coupled with Mass Spectrometry

The volatile fraction of Ophrys sphegodes Mill. subsp. sphegodes, Ophrys bertolonii subsp. benacensis (Reisigl) O. Danesch, E. Danasch & Ehrend. and Neotinea tridentata (Scop.) R.M. Bateman, Pridgeon & M.W. Case, three orchid species with different pollinator attraction strategies, sampled in vivo and in situ, were evaluated by headspace solid phase microextraction coupled with gas-chromatography and mass spectrometry. The results were compared with the volatile compounds emitted by flowering plant samples picked from the same populations of orchid species. Hydrocarbons, aldehydes, alcohols and terpenes were the major constituents of “in vivo” orchid scents and some distinctive differences in volatile metabolite composition were observed between Ophrys and Neotinea species. Moreover, the odour bouquets of the picked flowering plant samples were different from the in vivo ones and in particular different proportions of the various terpenes and an increase of α-pinene were observed. In conclusion HS/SPME GCMS proved to be a suitable technique for analyzing and distinguishing the volatile fingerprint of different orchid species, sampled in vivo and in situ in a non-disruptive way, with potentially great advantages for ecophysiological studies of rare and endangered species.