Non-invasive approaches to morphological and molecular identification of insects from museum, archaeo-funerary and forensic contexts

Species identification is the starting point for the formulation of any further ipothesis in insect sciences and they are particularly important in forensic entomology and funerary arcaheoentomology. Two main approaches are currently used for identifying insects, the morphological one, preferred by old-school taxonomists and the more recent molecular one, favoured by non-specialists. Both approaches have respectively some limitations: lack of morphological identification keys, especially for the immature stages and the destruction of the specimens during the DNA extraction, as examples. The development of easy-access identification keys and of non-invasive DNA extraction methods permitting the repeatability of the analyses have been the main target of this thesis. To achieve this aims morphological analysis and DNA based methods were integrated and used on samples belonging to different contexts –forensic, museum and archaeological –and different timeframe –0 to 1000 years old-. The attention was paid to adults and the poor investigate puparia of flies from each context and the different timeframes. For the DNA analysis commercial kits and homemade digestion buffers were tested on modern and old specimens but applying operational procedure to reserve the morphology of the specimens. More then 10 puparia belonging to species in the family Heleopmyzidae, Sphaeroceridae, Piophilidae, Muscidae and Calliphoridae have been described and illustrated. Forty new DNA sequences, mainly from poor investigated taxa, were generated and 37 deposited in GenBank despite some problems arised, mainly due to presence of PCR inibithors in and on the cuticle. The results, reported in this work, will impact in the anlysis of forensic and archaeological material but as well in the study of museum material, that especialy for the species typusis particularly precius and can not be destroid or altered during the identification process. In addition specific question as the separation and identification of Hydrotaea puparia belonging to different species, the identification of poorly investigated species, the systematic of Sarcophaga species in the subgenus Sarcophaga and the evaluation of pre-Columbian funerary cerimonies have been answered. In addition the pictorial material produced will help entomologists working with fragmented material especially from old cases or archaeological sites. Several questions are still open and they request further investigations to be done in the future

Effect of aminergic signaling on the humoral innate immunity response of Drosophila

Biogenic amines are crucial signaling molecules that modulate various physiological life functions both in vertebrates and invertebrates. In humans, these neurotransmitters influence the innate and adaptive immunity systems. In this work, we analyzed whether the aminergic neurotransmission of dopamine, serotonin, and octopamine could have an impact on the humoral innate immune response of Drosophila melanogaster. This is a powerful model system widely used to uncover the insect innate immunity mechanisms which are also conserved in mammals. We found that the neurotransmission of all these amines positively modulates the Toll-responsive antimicrobial peptide (AMP) drosomycin (drs) gene in adult flies infected with the Micrococcus luteus bacterium. Indeed, we showed that either blocking the neurotransmission in their specific aminergic neurons by expressing shibire(ts) (Shi(ts)) or silencing the vesicular monoamine transporter gene (dVMAT) by RNAi caused a significantly reduced expression of the Toll-responsive drs gene. However, upon M. luteus infection, the block of aminergic transmission did not alter the expression of AMP attacin genes responding to the immune deficiency (Imd) and Toll pathways. Overall, our results not only reveal a neuroimmune function for biogenic amines in humoral immunity but also further highlight the complexity of the network controlling AMP gene regulation

Sleep-disordered breathing and pregnancy outcomes: The impact of maternal oxygen saturation

ObjectiveTo investigate pathological associations between sleep-disordered breathing (SDB) and pregnancy outcomes. MethodsFrom May 2016 to September 2019, obese women during their uncomplicated singleton pregnancies underwent screening sleep questionnaires, oxygen saturation monitoring, and, in proper cases, complete overnight polysomnography. Their medical records were also recorded. ResultsIn all, 112 pregnant women were included in the study cohort; 44 showed an oxygen desaturation index & GE;10, and their newborns had a significantly higher rate of congenital abnormalities and respiratory distress syndrome compared with the women with normal pulse oximetry. Stepwise multivariate regression analysis showed that basal oxygen saturation was independently associated with the occurrence of fetal growth restriction. ConclusionAmong obese pregnant women, the rate of congenital abnormalities is higher in the ones with altered pulse oximetry. Maternal basal oxygen saturation in the first trimester of pregnancy predicts fetal growth restriction independently of maternal age, ethnicity, body mass index, gravidity, and hypertensive disorders of pregnancy

A combined protocol for identification of maggots of forensic interest

In Forensic Entomology the estimation of the age of insects is used for the estimation of the minimum postmortem interval. As insect development is temperature dependent and species specific, a correct species identification is therefore fundamental. In the majority of cases the molecular identification is based on a destructive approach. In this paper a working protocol for molecular identification of fly larvae without affecting the anatomical characters used for morphological identification is presented. The suggested technique allows the preservation of the larval exoskeleton and of the unused soft tissues in the same vial allowing a repetition of both the morphological and molecular identification and reducing the risk of loss of the evidence. This method also allows the possibility of measuring the size of the specimens before their morphological and biomolecular characterization. In order to demonstrate that this technique can be applied on maggots of a large spectrum of dimensions it has been tested and validated using larvae of different size from ~1.7–1.3 cm [Calliphora vomitoria and Lucilia sericata (Diptera: Calliphoridae)] to ~10–6.5 mm [Musca domestica (Diptera: Muscidae) and Megaselia scalaris (Diptera: Phoridae)]. The importance of a unique identifier and of a complete database with all the specimen information (origin, sample size, identification, etc.) is also discussed

A review of morphological characters for the identification of three common European species of Sarcophaga s. str. (Diptera: Sarcophagidae), with an emphasis on female terminalia

FIGURE 3. Female terminalia of Sarcophaga (s. str.), sternites 6–8. A. S. subvicina (SMNS_Dip_007081): length (L) of sternite 7 (ST7) equal to or longer than width (W). B. S. carnaria (SMNS_Dip_007077): ST7 wider (W) than long (L); anterior, lateral and posterior margins of ST7 straight. C. S. carnaria (SMNS_Dip_007079): anterior margin of ST7 with an indentation in the middle, lateral margins bending inwards on basal half, and posterior margin straight. D. S. variegata (SMNS_Dip_007041): lateral margins of ST7 bending inwards on basal 1/5. E. S. variegata (SMNS_Dip_007040): all margins of ST7 straight. F. S. variegata (SMNS_Dip_007042): lateral margins of ST7 bending inwards on basal half and posterior margin sinuous.Published as part of Schönberger, Daniel, Giordani, Giorgia, Vanin, Stefano & Whitmore, Daniel, 2022, A review of morphological characters for the identification of three common European species of Sarcophaga s. str. (Diptera: Sarcophagidae), with an emphasis on female terminalia, pp. 463-480 in Zootaxa 5205 (5) on page 471, DOI: 10.11646/zootaxa.5205.5.4, http://zenodo.org/record/731853

Retrotransposons Down- and Up-Regulation in Aging Somatic Tissues

none5noThe transposon theory of aging hypothesizes the activation of transposable elements (TEs) in somatic tissues with age, leading to a shortening of the lifespan. It is thought that TE activation in aging produces an increase in DNA double-strand breaks, contributing to genome instability and promoting the activation of inflammatory responses. To investigate how TE regulation changes in somatic tissues during aging, we analyzed the expression of some TEs, as well as a source of small RNAs that specifically silence the analyzed TEs; the Drosophila cluster named flamenco. We found significant variations in the expression levels of all the analyzed TEs during aging, with a trend toward reduction in middle-aged adults and reactivation in older individuals that suggests dynamic regulation during the lifespan.openGiordani, Giorgia; Cavaliere, Valeria; Gargiulo, Giuseppe; Lattanzi, Giovanna; Andrenacci, DavideGiordani, Giorgia; Cavaliere, Valeria; Gargiulo, Giuseppe; Lattanzi, Giovanna; Andrenacci, David

The human Smoothened inhibitor PF-04449913 induces exit from quiescence and loss of multipotent Drosophila hematopoietic progenitor cells

International audienceThe efficient treatment of hematological malignancies as Acute Myeloid Leukemia, myelofibrosis and Chronic Myeloid Leukemia, requires the elimination of cancer-initiating cells and the prevention of disease relapse through targeting pathways that stimulate generation and maintenance of these cells. In mammals, inhibition of Smoothened, the key mediator of the Hedgehog signaling pathway, reduces Chronic Myeloid Leukemia progression and propagation. These findings make Smo a candidate target to inhibit maintenance of leukemia-initiating cells. In Drosophila melanogaster the same pathway maintains the hematopoietic precursor cells of the lymph gland, the hematopoietic organ that develops in the larva. Using Drosophila as an in vivo model, we investigated the mode of action of PF-04449913, a small-molecule inhibitor of the human Smo protein. Drosophila larvae fed with PF-04449913 showed traits of altered hematopoietic homeostasis. These include the development of melanotic nodules, increase of circulating hemocytes, the size increase of the lymph gland and accelerated differentiation of blood cells likely due to the exit of multi-potent precursors from quiescence. Importantly, the Smo inhibition can lead to the complete loss of hematopoietic precursors. We conclude that PF-04449913 inhibits Drosophila Smo blocking the Hh signaling pathway and causing the loss of hematopoietic precursor cells. Interestingly, this is the effect expected in patients treated with PF-04449913: number decrease of cancer initiating cells in the bone marrow to reduce the risk of leukemia relapse. Altogether our results indicate that Drosophila comprises a model system for the in vivo study of molecules that target evolutionary conserved pathways implicated in human hematological malignancies

Record of Leptometopa latipes (Diptera: Milichiidae) from a human cadaver in the Mediterranean area

In forensic entomology, insects are used mainly to obtain information about the time since death. This information is obtained studying the developmental rate of the first colonizers, principally species in the families Calliphoridae, Sarcophagidae, Muscidae, Stratiomyidae and Phoridae. However, species belonging to other families can provide information about body transfer or the season of the death. Among them Milichiidae are flies rarely reported from human cases despite the larvae of some species are known as saprophagous feeding on plant and animal decomposing matter. A potential cause of the lack of records of these species from forensic cases can be related with the paucity of descriptions and illustrations of the immature stages. In this article, the entomological samples collected from a human body found inside an apartment in a Maghreb country, in Northern Africa, is reported and Leptometopa latipes (Diptera: Milichiidae) is described in detail. Molecular analysis is also reported to confirm the morphological analysis