High diversity in species, reproductive modes and distribution within the Paramacrobiotus richtersi complex (Eutardigrada, Macrobiotidae)

For many years, Paramacrobiotus richtersi was reported to consist of populations with different chromosome numbers and reproductive modes. To clarify the relationships among different populations, the type locality of the species (Clare Island, Ireland) and several Italian localities were sampled. Populations were investigated with an integrated approach, using morphological (LM, CLSM, SEM), morphometric, karyological, and molecular (18S rRNA, cox1 genes) data. Paramacrobiotus richtersi was redescribed and a neotype designed from the Irish bisexual population. Animals of all populations had very similar qualitative and quantitative characters, apart from the absence of males and the presence of triploidy in some of them, whereas some differences were recorded in the egg shell. All populations examined had the same 18S haplotype, while 21 haplotypes were found in the cox1 gene. In four cases, those qualitative characters were correlated with clear molecular (cox1) differences (genetic distance 14.6\u201321.8%). The integrative approach, which considered the morphological differences in the eggs, the reproductive biology and the wide genetic distances among putative species, led to the description of four new species (Paramacrobiotus arduus sp. n., Paramacrobiotus celsus sp. n., Paramacrobiotus depressus sp. n., Paramacrobiotus spatialis sp. n.) and two Unconfirmed Candidate Species (UCS) within the P. richtersi complex. Paramacrobiotus fairbanksi, the only ascertained parthenogenetic, triploid species, was redescribed and showed a wide distribution (Italy, Spain, Poland, Alaska), while the amphimictic species showed limited distributions. The difference in distribution between apomictic and amphimictic populations can be explained by the difference in the dispersal potentials associated with these two types of reproduction

Life-history traits and description of the new gonochoric amphimictic Mesobiotus joenssoni (Eutardigrada: Macrobiotidae) from the island of Elba, Italy

Comparative analyses of life-history theory studies are based on the characteristics of the life cycles of different species. For tardigrades, life-history traits are available only from laboratory cultures, most of which have involved parthenogenetic species. The discovery of a new gonochoristic bisexual Mesobiotus species in a moss collected on the island of Elba (Italy) provides us with the opportunity to describe Mesobiotus joenssoni sp. nov. and to collect data on the life-history traits of cultured specimens to increase our knowledge of the life-history strategies present in tardigrades. This new species is differentiated from all other species of the genus by the presence of granules (~1 µm in diameter) on the dorsal cuticle of the last two body segments, two large bulges (gibbosities) on the hindlegs and long, conical egg processes. The species exhibits sexual dimorphism in body length, with females being longer than males of the same age. The mean lifespan of specimens was 86 days, with a maximum of 150 days. The mean age at first oviposition was 19.8 days and the mean egg hatching time 15.4 days. The life-cycle traits correspond to those collected for the only other two macrobiotid species with gonochoric amphimictic reproduction examined so far

Cardiac involvement in Erdheim- Chester disease: MRI findings and literature revision

: Erdheim-Chester disease (ECD) is a rare form of non-Langerhans cell histiocytosis, characterized by the involvement of several organs. The lesions may be skeletal or extra-skeletal: in particular, long bones, skin, lungs, and the cardiovascular and the central nervous systems can be affected. In this report, we describe a case of a 34-year-old man, who came to our observation with symptomatic ECD, for a correct assessment of the degree of cardiac involvement through magnetic resonance imaging (MRI)

Resistance to Extreme Stresses by a Newly Discovered Japanese Tardigrade Species, Macrobiotus kyoukenus (Eutardigrada, Macrobiotidae)

Tardigrades are small micrometazoans able to resist several environmental stresses in any stage of their life cycle. An integrated analysis of tardigrade specimens collected in Tsukuba (Japan) revealed a peculiar morphology and a new sensory field in the cloaca. Molecular taxonomy and phylogenetic analysis on different genes (COI, ITS2, 18S and 28S) confirmed that this population is a new species, Macrobiotus kyoukenus sp. nov., belonging to the widespread Macrobiotus hufelandi group. The stress resistance capabilities of M. kyoukenus sp. nov. have been tested by submitting animals to extreme desiccation, rapid freezing, and high levels of ultraviolet radiations (UVB and UVC). Animals were able to survive desiccation (survivorship 95.71 ± 7.07%) and freezing up to -80 °C (82.33 ± 17.11%). Both hydrated and desiccated animals showed a high tolerance to increasing UV radiations: hydrated animals survived to doses up to 152.22 kJ m−2 (UVB) and up to 15.00 kJ m−2 (UVC), while desiccated specimens persisted to radiations up to 165.12 kJ m−2 (UVB) and up to 35.00 kJ m−2 (UVC). Present data contribute to the discovery of a larger tardigrade biodiversity in Japan, and the tolerance capabilities of M

Fertility specialists’ views, behavior, and attitudes towards the use of endometrial scratching in Italy

Background: Endometrial scratching (ES) or injury is intentional damage to the endometrium performed to improve reproductive outcomes for infertile women desiring pregnancy. Moreover, recent systematic reviews with meta-analyses and randomized controlled trials demonstrated that ES is not effective, data on the safety are limited, and it should not be recommended in clinical practice. The aim of the current study was to assess the view and behavior towards ES among fertility specialists throughout infertility centers in Italy, and the relationship between these views and the attitudes towards the use of ES as an add-on in their commercial setting. Methods: Online survey among infertility centers, affiliated to Italian Society of Human Reproduction (SIRU), was performed using a detailed questionnaire including 45 questions with the possibility to give "closed" multi-choice answers for 41 items and "open" answers for 4 items. Online data from the websites of the infertility centers resulting in affiliation with the specialists were also recorded and analyzed. The quality of information about ES given on infertility centers websites was assessed using a scoring matrix including 10 specific questions (scored from 0 to 2 points), and the possible scores ranged from 0 to 13 points ('excellent' if the score was 9 points or more, 'moderate' if the score was between 5 and 8, and 'poor' if it was 4 points or less). Results: The response rate was of 60.6% (43 questionnaires / 71 infertility SIRU-affiliated centers). All included questionnaires were completed in their entirety. Most physicians (~ 70%) reported to offer ES to less than 10% of their patients. The procedure is mainly performed in the secretory phase (69.2%) using pipelle (61.5%), and usually in medical ambulatory (56.4%) before IVF cycles to improve implantation (71.8%) without drugs administration (e.g., pain drugs, antibiotics, anti-hemorrhagics, or others) before (76.8%) or after (64.1%) the procedure. Only a little proportion of infertility centers included in the analysis proposes formally the ES as an add-on procedure (9.3%), even if, when proposed, the full description of the indications, efficacy, safety, and costs is never addressed. However, the overall information quality of the websites was generally "poor" ranging from 3 to 8 and having a low total score (4.7 ± 1.6; mean ± standard deviation). Conclusions: In Italy, ES is a procedure still performed among fertility specialists for improving the implantation rate in IVF patients. Moreover, they have a poor attitude in proposing ES as an add-on in the commercial setting

The Use of Ceus Software with No Contrast Media Administration in the Diagnosis of Pneumoperitoneum

Background: Pneumoperitoneum is defined by the presence of free air in the abdominal cavity; gastrointestinal perforation is an important cause of this pathological condition. In emergency situations, radiology is considered vital in the early detection and identification of the site and cause of the perforation, which is critical for proper surgical planning. Aim: The aim of our study was to evaluate a new diagnostic US tool, based on the US contrast-specific software generally used during contrast-enhanced US examination (CEUS), without the administration of sonographic contrast media, and to describe the specific imaging features in the detection of free intra-peritoneal air. Subjects and Methods: One hundred and fifty-seven consecutive and hemodynamically stable patients, who arrived in our E.D. with an acute abdomen between April 2018 and October 2019, underwent US and CT examination, performed by three radiologists (with 5, 5, and 25 years of experience). The US was performed first and divided into two steps, using B-mode US and both B-mode and contrast-specific software US, with no contrast media administration. All the patients underwent CT examination. Results: In 32 out of 157 patients, the surgery confirmed GI perforation. CT correctly detected 31 out of 32 patients; the contrast-specific software US identified 30 perforated patients. CT reached a sensitivity value of 97% and specificity value of 100%; contrast-specific software US demonstrated higher values than B-mode US in sensitivity (93% vs. 70%, respectively) and specificity (98% vs. 88%, respectively). Conclusion: the use of contrast-specific software in emergencies improves image quality, and reaches higher levels of sensitivity and specificity with no time delay compared to standard US examination, helping radiologists expedite diagnoses

Identification of the trehalose-6-phosphate synthase (tps) gene in desiccation tolerant and intolerant tardigrades

The presence of the gene coding for trehalose-6-phosphate synthase (tps) was determined for the first time in Tardigrada in twelve species belonging to both Heterotardigrada and Eutardigrada. The tps gene was found in all semiterrestrial and anhydrobiotic species examined, correlating trehalose presence with anhydrobiotic ability. In contrast, tps was not detected in two limnic and desiccation intolerant species. The surprising presence of tps in another limnic and desiccation intolerant species, Dactylobiotus parthenogeneticus, allowed us to infer that in tardigrades trehalose could be produced and involved, not just in anhydrobiosis, but also in the regulation of other biological functions, such as encystment. Our findings, integrated with literature data, indicate that anhydrobiotic ability in tardigrades can depend on the combined action of numerous different molecules, only partially identified. With nucleotide tps sequences, it is impossible to infer phylogenetic relationships inside the phylum Tardigrada, while amino acid sequences further support the relationship between Tardigrada and Pancrustacea

DNA barcoding e strategie riproduttive nel genere Ramazzottius (Tardigrada, Eutardigrada)

Nell’ambito del progetto MoDNA (morfologia e DNA) è stato utilizzato l’approccio del DNA barcoding, associato a comparazioni morfologiche e cariologiche. E’ stata valutata la variabilità inter- ed intraspecifica nel genere Ramazzottius, caratterizzato dalla presenza di specie criptiche e da diverse modalità riproduttive e talvolta diverso grado di ploidia. Mediante approccio morfologico (LM, SEM) e molecolare (mtDNA cox1) sono stati analizzati esemplari di Ramazzottius provenienti da 15 località europee. Per correlare morfologia e sequenze sono stati allestiti voucher specimens, compresi gusci di uova (molto importanti per la diagnosi specifica) dalle quali sono sgusciati neonati utilizzati per l’analisi molecolare. Tre campioni contengono rispettivamente Ramazzottius varieornatus, Ramazzottius tribulosus ed una nuova specie di Ramazzottius, distinguibili sia su base morfologica che molecolare. Negli altri campioni gli esemplari (animali e uova) sono tutti morfologicamente attribuibili a Ramazzottius oberhaeuseri, ma si distinguono tra loro per sex ratio, ploidia e per notevoli differenze molecolari. I dati evidenziano la validità dell’approccio integrato nello studio della diversità nei tardigradi e dell’utilizzo del DNA barcoding. Nel genere Ramazzottius, ed in particolare nell’ambito della morfospecie R. oberhaeuseri, è confermata la presenza di specie criptiche, alcune delle quali con ampia diffusione e con strategie riproduttive diverse

Survival and DNA degradation in anhydrobiotic tardigrades

Anhydrobiosis is a highly stable state of suspended animation in an organism due to its desiccation, which is followed by recovery after rehydration. Changes occurring during drying could damage molecules, including DNA. Using as a model organism the anhydrobiotic tardigrade Paramacrobiotus richtersi, we have evaluated the effects of environmental factors, such as temperature and air humidity level (RH), on the survival of desiccated animals and on the degradation of their DNA. Tardigrades naturally desiccated in leaf litter and tardigrades experimentally desiccated on blot paper were considered. Replicates were kept at 37°C and at different levels of RH for 21 days. RH values and temperature, as well as time of exposure to these environmental factors, have a negative effect on tardigrade survival and on the time required by animals to recover active life after desiccation. DNA damages (revealed as single strand breaks) occurred only in desiccated tardigrades kept for a long time at high RH values. These results indicate that during the anhydrobiotic state, damages take place and accumulate with time. Two hypotheses can be formulated to explain the results: i. oxidative damages occur in desiccated specimens of P. richtersi, ii. high temperature and high RH values change the state of the disaccharide trehalose, reducing its protective role

Approccio molecolare, modalità riproduttive e specie criptiche nei tardigradi.

Gli studi molecolari hanno portato ad attribuire i tardigradi agli Ecdysozoa e confermato sia la loro appartenenza ai Panarthropoda, che la loro suddivisione in ordini. Ancora assai limitato è invece l’utilizzo della biologia molecolare per discriminare le loro specie. Numerosi sono, infatti, i ritrovamenti di tardigradi attribuibili alla medesima morfospecie, ma distinti in popolazioni unisessuate e bisessuate, spesso diverse anche per grado di ploidia. Le indagini condotte finora non hanno però consentito di stabilire con certezza se tra queste popolazioni vi fosse o meno flusso genico. Utilizzando come marcatore una sequenza di 640 bp del gene mitocondriale COI, sono state confrontate popolazioni riferibili all’eutardigrado Macrobiotus richtersi con diversa provenienza geografica (Modena e Trento) e verificate le loro modalità riproduttive. In entrambe le province sono state rinvenute, in località distinte, sia popolazioni anfimittiche (diploidi) che partenogenetiche telitoche (triploidi e ameiotiche). L’analisi delle sequenze ha rilevato notevole somiglianza nell’aplotipo delle popolazioni partenogenetiche, pur se geograficamente distanti, e forti differenze tra queste e le anfimittiche. Queste sono coerenti con quanto da noi già riscontrato tra popolazioni anfimittiche e partenogenetiche di un’altra morfospecie (Richtersius coronifer) utilizzando allozimi. La costanza e l’ampiezza delle differenze nell’aplotipo evidenziano che non c’è flusso genico tra le popolazioni e che siamo in presenza di specie criptiche. Data l’esistenza di numerose morfospecie di tardigradi con più modalità riproduttive, il loro studio a livello molecolare, associato a quello della loro distribuzione geografica, dovrebbero portare nuova luce sulla microevoluzione all’interno del phylum e, più in generale, sui microinvertebrati caratterizzati da forte dispersione passiva