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this study evidenced the presence of parasites in a cesspit of an aristocratic palace of nineteenth century in Sardinia (Italy) by the use of classical paleoparasitological techniques coupled with nextgeneration sequencing. Parasite eggs identified by microscopy included helminth genera pathogenic for humans and animals: the whipworm Trichuris sp., the roundworm Ascaris sp., the flatworm Dicrocoelium sp. and the fish tapeworm Diphyllobothrium sp. In addition, 18S rRNA metabarcoding and metagenomic sequencing analysis allowed the first description in Sardinia of aDNA of the human specific T. trichiura species and Ascaris genus. Their presence is important for understanding the health conditions, hygiene habits, agricultural practices and the diet of the local inhabitants in the period under study. Paleoparasitology, the study of ancient parasites recovered from archaeological sites, is a branch of paleopathology important to understand the health conditions and lifestyle of past populations 1-5. Classical paleoparasitology consist on the rehydration and microscopic analysis of coprolites, latrine sediments, pelvic soil of skeletons or intestines of mummified bodies, followed by identification of recovered parasite eggs basing on morphometry and other characteristics (e.g. opercula, caps and surface structures) 6. With this approach, since the first description in 1910 of Schistosoma haematobium eggs in a renal tissue of an Egyptian mummy 7 , helminth eggs have been identified in coprolites, latrines, mummified bodies, and archaeological contests all over the world 4,8-15. Despite microscopy is still a method of choice for paleoparasitological studies, it allows to identify the parasites mostly to genus level, as the eggs of related species are often indistinguishable 5,16,17. For a better taxonomic identification, immunological, hybridization and molecular techniques were developed and used in combination with classical methods 18-21. Molecular paleoparasitological studies are mainly based on PCR amplification and Sanger sequencing of short barcoding loci as 18S rDNA, mostly using primers for specific parasite taxa 21-24. The recent developed next generation sequencing (NGS) allows to identify multiple taxonomic groups at the same time by direct shotgun sequencing of DNA extracted from the samples (metagenomics) or by PCR-based metabarcoding of target genes 25-29 , but the application of this technique to paleoparasitology is so far limited to few studies 22,30,31. In Europe, human intestinal parasites were described from Palaeolithic until the middle of 1900 17 , but in Sardinia

Diagnosis of cystic echinococcosis (CE) in sheep is essentially based on necropsy findings. Clinical symptoms can be easily overlooked, while the use of immunological tests is still not recommended for an intra vitam diagnosis. This study assessed the performances of three post-mortem laboratory methods in the diagnosis of ovine CE. In the absence of a single and accurate test as a gold standard, the results of multiple analytical tests can be combined to estimate diagnostic performance based on a Bayesian statistical approach. For this purpose, livers (n = 77), and lungs (n = 79) were sampled from adult sheep and examined using gross pathology, histopathology and molecular analyses. Data from the three diagnostic methods were analyzed using a Bayesian latent class analysis model to evaluate their diagnostic accuracy in terms of sensitivity (Se), specificity (Sp), positive predictive value (PPV) and negative predictive value (NPV). The gross pathology examination revealed excellent diagnostic capabilities in diagnosing ovine CE with an Se of 99.7 (96.7–99.8), Sp of 97.5 (90.3–99.8), PPV of 97.6 (90.5–100), and NPV of 99.7 (96.5–100). The experimental design used in this work could be implemented as a validation protocol in a quality assurance system.

The Sardinian coloured donkey Equus asinus (Perissodactyla: Equidae) and its albino colour morph represent the wildlife species most typical of the island of Asinara. This Mediterranean island represents a favourable context for ticks and tick-borne diseases; however, knowledge of the tick fauna on Asinara is scarce. A total of 106 Sardinian donkeys were inspected for tick infestation from June to November 2015. All ticks found were collected, classified by stage and sex, and identified to species level. The level of infestation of each donkey was determined; both the overall tick infestation and infestations of each detected species were classified on a scale of 1-3 to give an infestation score (IS). Overall, 256 hard ticks were collected from 60 of 106 donkeys (56.6%). Rhipicephalus bursa, Haemaphysalis punctata and Hyalomma marginatum (all: Ixodida: Ixodidae) infested 26.4%, 28.3% and 6.6% of donkeys, respectively. Different variables affected the IS. With reference to overall tick infestation, a higher IS was observed in donkeys grazing on grassland and Mediterranean shrubland and in albino donkeys compared with coloured donkeys. The collected ticks included species involved in the transmission of pathogens to humans, which highlights the risks for public health in a tourist destination such as Asinara National Park.