Secretory Cells in Halla parthenopeia (Oenonidae): Potential Implications for the Feeding and Defence Strategies of a Carnivorous Burrowing Polychaete.

Carnivorous polychaetes are known to bear diversified and often unique anatomical and behavioural adaptations for predation and defence. Halla parthenopeia, a species known to be a specialized predator of clams, thrives in the soft bottoms of the Mediterranean Sea, holding potential for polyculture and biotechnology due to the secretion of bioactive compounds. Our objective was to provide a comprehensive description of H. parthenopeia's anatomy and microanatomy, shedding light on the relation between morphology and habitat, chemical defences, and feeding behaviour. The pharynx, housing maxillae and mandibles connected to an extensive mucus gland, occupies a considerable portion of the worm's length, reaching beyond the oesophagus. This unique gland is responsible for secreting the feeding mucus, which immobilizes and aids in the digestion of clams probably acting as a vehicle of bioactive compounds synthesized by specialized serous cells in the mouth. Moreover, H. parthenopeia combines behavioural tactics, such as burrowing, and anatomical defences to evade predators. Examination of its epidermis revealed a thick cuticle layer and abundant mucocytes secreting locomotion mucus, both of which save the worm from mechanical harm during movement. When it is preyed upon, the worm can release a substantial amount of Hallachrome, a toxic anthraquinone produced by specific cells in its distal region. This pigment, with its known antimicrobial properties, likely acts as a chemical shield in case of injury. The results suggest that the ability of H. parthenopeia to prey on bivalves and to provide mechanical protection plus defence against pathogens rely on its ability to secrete distinct types of mucus. The interplay between highly specialized microanatomical features and complex behaviours underscores its adaptation as a predator in marine benthic environments.

Exploring the Antimicrobial Potential of Hallachrome, a Defensive Anthraquinone from the Marine Worm Halla parthenopeia (Polychaeta).

Antimicrobial resistance is a critical global health issue, with rising resistance among bacteria and fungi. Marine organisms have emerged as promising, but underexplored, sources of new antimicrobial agents. Among them, marine polychaetes, such as Halla parthenopeia, which possess chemical defenses, could attract significant research interest. This study explores the antimicrobial properties of hallachrome, a unique anthraquinone found in the purple mucus of H. parthenopeia, against Gram-negative bacteria (Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 9027), Gram-positive bacteria (Enterococcus faecalis ATCC 29212, Staphylococcus aureus ATCC 6538, Staphylococcus epidermidis ATCC 12228), and the most common human fungal pathogen Candida albicans ATCC 10231. Antibacterial susceptibility testing revealed that Gram-negative bacteria were not inhibited by hallachrome at concentrations ≤2 mM. However, Gram-positive bacteria showed significant growth inhibition at 0.12-0.25 mM, while C. albicans was inhibited at 0.06 mM. Time-kill studies demonstrated dose-dependent growth inhibition of susceptible strains by hallachrome, which exerted its effect by altering the membrane permeability of C. albicans, E. faecalis, and S. epidermidis after 6 h and S. aureus after 24 h. Additionally, hallachrome significantly reduced biofilm formation and mature biofilm in S. aureus, E. faecalis, and C. albicans. Additionally, it inhibited hyphal growth in C. albicans. These findings highlight hallachrome's potential as a novel antimicrobial agent, deserving further exploration for clinical experimentation.

Can We Apply Snyder's Arthroscopic Classification to Ultrasound for Evaluating Rotator Cuff Tears? A Comparative Study with MR Arthrography.

We aimed to demonstrate the applicability of Snyder's arthroscopic classification of rotator cuff tears (RCT) in shoulder ultrasound (US) and to compare it with MR arthrography (MRA). Forty-six patients (34 males; mean age:34 ± 14 years) underwent shoulder US and MRA. Two radiologists (R1 = 25 years of experience; R2 = 2 years of experience) assigned A1-4, B1-4, or C1-4 values depending on the extent of RCT in both US and MRA. Inter-reader intra-modality and intra-reader inter-modality agreement were calculated using Cohen's kappa coefficient. US sensitivity and specificity of both readers were calculated using MRA as the gold standard. Patients were divided into intact cuff vs. tears, mild (A1/B1) vs. moderate (A2-3/B2-3) tears, mild-moderate (A2/B2) vs. high-moderate (A3/B3) cuff tears, moderate (A2-3/B2-3) vs. advanced (A4/B4) and full-thickness (C) tears. The highest agreement values in inter-reader US evaluation were observed for mild-moderate vs. high-moderate RCT (K = 0.745), in inter-reader MRA evaluation for mild vs. moderate RCT (K = 0.821), in R1 inter-modality (US-MRA) for mild-moderate vs. high-moderate and moderate vs. advanced/full-thickness RCT (K = 1.000), in R2 inter-modality (US-MRA) for moderate vs. advanced/full-thickness RCT (K = 1.000). US sensitivity ranged from 88.89%(R1)-84.62%(R2) to 100% (both readers), while specificity from 77.78%(R1)-90.00%(R2) to 100% (both readers). Snyder's classification can be used in US to ensure the correct detection and characterization of RCT.

Radiomics in PI-RADS 3 Multiparametric MRI for Prostate Cancer Identification: Literature Models Re-Implementation and Proposal of a Clinical-Radiological Model.

PI-RADS 3 prostate lesions clinical management is still debated, with high variability among different centers. Identifying clinically significant tumors among PI-RADS 3 is crucial. Radiomics applied to multiparametric MR (mpMR) seems promising. Nevertheless, reproducibility assessment by external validation is required. We retrospectively included all patients with at least one PI-RADS 3 lesion (PI-RADS v2.1) detected on a 3T prostate MRI scan at our Institution (June 2016-March 2021). An MRI-targeted biopsy was used as ground truth. We assessed reproducible mpMRI radiomic features found in the literature. Then, we proposed a new model combining PSA density and two radiomic features (texture regularity (T2) and size zone heterogeneity (ADC)). All models were trained/assessed through 100-repetitions 5-fold cross-validation. Eighty patients were included (26 with GS ≥ 7). In total, 9/20 T2 features (Hector's model) and 1 T2 feature (Jin's model) significantly correlated to biopsy on our dataset. PSA density alone predicted clinically significant tumors (sensitivity: 66%; specificity: 71%). Our model obtained a sensitivity of 80% and a specificity of 76%. Standard-compliant works with detailed methodologies achieve comparable radiomic feature sets. Therefore, efforts to facilitate reproducibility are needed, while complex models and imaging protocols seem not, since our model combining PSA density and two radiomic features from routinely performed sequences appeared to differentiate clinically significant cancers.

Novel Natural Compounds and Their Anatomical Distribution in the Stinging Fireworm Hermodice carunculata (Annelida).

Increasing evidence in the field of bioprospection fosters the necessity of studying poorly investigated poisonous marine invertebrates to expand knowledge on animal venom biology. Among marine annelids, amphinomid fireworms are notorious for their bearded trunk equipped with a powerful stinging capacity. Here, a methodological workflow based on analytical chemistry techniques (compound isolation followed by mass spectrometry and spectroscopy analyses) was applied to gain new insights, leading to the identification and structural elucidation of an array of natural products from Mediterranean specimens of Hermodice carunculata. Eight betaine-derived unprecedented compounds, named "carunculines", were detected, bearing two terminal ammonium groups tri-and disubstituted at the Cα (A, B) and a series of different alkyl chains (I-VIII). The mixture of chemicals was found in all the body parts of H. carunculata, supporting a mechanism of action triggered by their vehiculation inside the dorsal chaetae, and subsequent injection when chaetae break off on contact. Preliminary investigations to understand adaptive features were also performed, showing a trend in carunculine abundance that fits into the evolutionary history of these worms. These findings shed light on the chemical ecology of amphinomids, giving reasons for the success of H. carunculata in benthic environments and providing promising novel metabolites for biotechnological implications.

Superb Microvascular Imaging (SMI) Compared with Color Doppler Ultrasound for the Assessment of Hepatic Artery in Pediatric Liver Transplants: A Feasibility Study.

(1) Background: Despite progression in surgical techniques and immunological treatments, hepatic artery (HA) thrombosis and stenosis still develop as an early or late liver transplant (LT) complication. We aimed to compare superb microvascular imaging (SMI) with conventional Doppler imaging (CDI) in the assessment of HA in a cohort of pediatric patients undergoing follow-up ultrasound (US) for LT. (2) Methods: This prospective, observational study included 73 pediatric LT recipients (median age, 7 years; IQR, 5.8 years; 35 females) who underwent US during LT follow-up from March to December 2019. For each examination, CDI and SMI were separately assessed in terms of HA visibility and spectral waveform morphology (SWM). The former was scored based on HA discrimination from the blooming signal of the surrounding vessels, as follows: 0, not visible; 1, majority course hardly distinguishable; and 2, majority course clearly distinguishable. The latter was scored on a two-point scale: 0, combined venous and arterial SWM, and 1, pure arterial SWM. The patient's overall score was finally calculated by adding the two individual scores. (3) Results: Both the absolute scores and frequency of overall scores equal to 3 (maximum global score) were higher using SMI compared with CDI. The median overall score was 3 for SMI and 2 for CDI (p = 0.011; IQR = 1). An overall score equal to 3 was obtained in 74% and 49.3% of the study population using SMI and CDI, respectively (p = 0.002). This was attributable to a better score in HA visibility (p = 0.007). (4) Conclusions: SMI has shown promise for assessing HA in pediatric LT recipients, possibly serving as a complementary non-invasive tool of CDI in everyday practice.

Accuracy of Inflow Inversion Recovery (IFIR) for Upper Abdominal Arteries Evaluation: Comparison with Contrast-Enhanced MR and CTA.

Background: Inflow-sensitive inversion recovery (IFIR) is a recently introduced technique to perform unenhanced magnetic resonance angiography (MRA). The purpose of our study is to determine the accuracy of IFIR-MRA in the evaluation of upper abdominal arteries, compared to standard MRA and computed tomography angiography (CTA). Materials and Methods: Seventy patients undergoing upper abdomen Magnetic Resonance Imaging (MRI) in different clinical settings were enrolled. The MRI protocol included an IFIR-MRA sequence that was intra-individually compared by using a qualitative 4-point scale in the same patients who underwent concomitant or close MRA (n = 65) and/or CTA (n = 44). Celiac trunk (CA), common-proper-left-right hepatic artery (C-P-L-R-HA), left gastric artery (LGA), gastroduodenal artery (GDA), splenic artery (SA), renal arteries (RA) and superior mesenteric artery (SMA) were assessed. Results: IFIR-MRA images were better rated in comparison with MRA. Particularly, all arteries obtained a statistically significant higher qualitative rating value (all p < 0.05). IFIR-MRA and MRA exhibited acceptable intraclass correlation coefficients (ICC) values for CA, C-L-R-HA, and SMA (ICC 0.507, 0.591, 0.615, 0.570, 0.525). IFIR-MRA and CTA showed significant correlations in C-P-L-R-HA (τ = 0.362, 0.261, 0.308, 0.307, respectively; p < 0.05), and in RA (τ = 0.279, p < 0.05). Conclusions: Compared to MRA, IFIR-MRA demonstrated a higher image quality in the majority of upper abdomen arterial vessels assessment. LHA and RHA branches could be better visualized with IFIR sequences, when visualizable. Based on these findings, we suggest to routinely integrate IFIR sequences in upper abdomen MRI studies.

Response to Tilic and Bartolomaeus's Commentary on the original Research Paper "Unravelling the ultrastructure and mineralogical composition of fireworm stinging bristles" (Zoology, 144).

In their Commentary to our paper recently published in Zoology (Righi et al., 2021a), Tilic and Bartolomaeus question our findings that the chaetae of Hermodice carunculata (Annelida) are hollow and able to store and deliver venoms. They sustain the idea that inflammatory chemicals are secreted through epidermal glands and possibly exposed to predator trough wounds caused by the brittle chaetae. We provide evidence-based arguments in support of our considerations. The sample preparation procedures did not affect the native inner structure of unfixed fireworm chaetae, which is clearly hollow as supported by both ultrastructure observation and crystal-chemical analysis of constituents. Furthermore, our previous and more recent feeding bioassays and chemical analysis indicate both that chaetae retain strong deterrent capacities even when isolated from the body of H. carunculata, and that they contain venoms. The cellular mechanisms involved in fireworm chaeta storage and deliver of chemicals are still unstudied. We strongly believe that this lack of knowledge should draw further attention on H. carunculata biology, pursuing new hypotheses and studies based on the noteworthy information which has been obtained so far.

Unravelling the ultrastructure and mineralogical composition of fireworm stinging bristles.

Amphinomid fireworms are notorious for their stinging dorsal bristles (notochaetae), but it is still unclear whether the irritation they cause is merely mechanical or if the notochaetae contain toxins. Furthermore, although fireworm chaetae have always been described as calcareous, their composition has never been investigated to date and strong debates are ongoing on their internal structure. Unravelling the native ultrastructure and composition of fireworm chaetae is the first crucial step to assess whether the hypothesis of toxin vehiculation could be fully considered. We examined for the first time the chemical and mineralogical composition, the ultrastructure and the external structure of the dorsal and ventral chaetae of the large species Hermodice carunculata. All the measurements were carried out on samples prepared without the use of chemical reagents, except for those targeted to investigate if decalcification altered the ultrastructure of the chaetae. A crystal-chemical strategy, combining chemical, diffraction and thermal analyses clearly showed the occurrence of crystalline calcium carbonate and clusters of phosphatic amorphous material. Scanning electron micrographs and energy dispersive X-ray measurements showed that the dorsal chaetae have an extremely shallow insertion point in the body respect to the ventral chaetae, that could facilitate the release of the notochaetae in the environment. Their proximal part is characterized by canals with a hexagonal pattern rich in Ca and P, followed by a large cavity upwards. The harpoon-shaped ends and the central canals of the notochaetae completely disappeared after exposure to EDTA. The notochaetae are hollow and may be able to vehicle toxins. The absence of the honeycomb pattern in the distal part of the notochaetae and their slenderness probably contribute to their brittleness and high sensitivity to breakage on contact. These observations constitute keystone understandings to shed light on fireworm defensive and offensive capacities and their ecological success.

The Immune Response of the Invasive Golden Apple Snail to a Nematode-Based Molluscicide Involves Different Organs.

The spreading of alien and invasive species poses new challenges for the ecosystem services, the sustainable production of food, and human well-being. Unveiling and targeting the immune system of invasive species can prove helpful for basic and applied research. Here, we present evidence that a nematode (Phasmarhabditis hermaphrodita)-based molluscicide exerts dose-dependent lethal effects on the golden apple snail, Pomacea canaliculata. When used at 1.7 g/L, this biopesticide kills about 30% of snails within one week and promotes a change in the expression of Pc-bpi, an orthologue of mammalian bactericidal/permeability increasing protein (BPI). Changes in Pc-bpi expression, as monitored by quantitative PCR (qPCR), occurred in two immune-related organs, namely the anterior kidney and the gills, after exposure at 18 and 25 °C, respectively. Histological analyses revealed the presence of the nematode in the snail anterior kidney and the gills at both 18 and 25 °C. The mantle and the central nervous system had a stable Pc-bpi expression and seemed not affected by the nematodes. Fluorescence in situ hybridization (FISH) experiments demonstrated the expression of Pc-bpi in circulating hemocytes, nurturing the possibility that increased Pc-bpi expression in the anterior kidney and gills may be due to the hemocytes patrolling the organs. While suggesting that P. hermaphrodita-based biopesticides enable the sustainable control of P. canaliculata spread, our experiments also unveiled an organ-specific and temperature-dependent response in the snails exposed to the nematodes. Overall, our data indicate that, after exposure to a pathogen, the snail P. canaliculata can mount a complex, multi-organ innate immune response.

Trans-generational plasticity in physiological thermal tolerance is modulated by maternal pre-reproductive environment in the polychaete Ophryotrocha labronica.

Maternal temperature is known to affect many aspects of offspring phenotype, but its effect on offspring physiological thermal tolerance has received less attention, despite the importance of physiological traits in defining organismal ability to cope with temperature changes. To fill this gap, we used the marine polychaete Ophryotrocha labronica to investigate the influence of maternal temperature on offspring upper and lower thermal tolerance limits, and assess whether maternal influence changed according to the stage of offspring pre-zygotic development at which a thermal cue was provided. Measurements were taken on adult offspring acclimated to 18 or 30°C, produced by mothers previously reared at 24°C and then exposed to 18 or 30°C at an early and late stage of oogenesis. When the shift from 24°C was provided early during oogenesis, mothers produced offspring with greater cold and heat tolerance whenever mother-offspring temperatures did not match, with respect to when they matched, suggesting the presence of an anticipatory maternal effect triggered by the thermal variation. Conversely, when the cue was provided later during oogenesis, more tolerant offspring were observed when temperatures persisted across generations. In this case, maternal exposure to 18 or 30°C may have benefited offspring performance, while limitations in the transmission of the thermal cue may account for the lack of correlation between maternal experiences and offspring performance when mother-offspring environments did not match. Our results provided evidence for a trans-generational effect of temperature on physiological performance characterised by a high context dependency, and are discussed in the light of maternal pre-reproductive experiences.

Novel antifouling agent--zinc pyrithione: short- and long-term effects on survival and reproduction of the marine polychaete Dinophilus gyrociliatus.

The recent ban on TBT in boat antifouling paints has resulted in a large employment of the biocide zinc pyrithione (ZnPT) as substitute. Despite concerns of its environmental toxicity at ultra-trace concentrations, ZnPT has received little attention, as it was assumed to photo-degrade easily. However, recent evidence has suggested that ZnPT degrades only partially, and persists in the marine environment, especially where the influence of light is limited, such as in harbours. Short-term acute toxicity tests and life table response experiments (LTREs) were performed to evaluate the effects of ZnPT on the polychaete Dinophilus gyrociliatus at both individual and population level. The 96h-LC(50) values for ZnPT on D. gyrociliatus were 7.8 and 11.5nM under dark and 12h light/12h dark conditions, respectively. In LTREs, laboratory cultured cohorts of D. gyrociliatus were exposed to sublethal concentrations of ZnPT (0.5 and 1nM) and compared to a control cohort. The survival among individuals exposed to the highest concentration decreased already during the 2nd week of life with respect to the control. The effects of the biocide on fecundity were even more evident: ZnPT caused a considerable reduction in both the exposed groups. The demographic approach applied here succeeded in identifying ZnPT effects both on the biological cycle and on the growth potential of polychaete D. gyrociliatus. The net growth rate (R(0)) appeared to be the demographic parameter most sensitive to ZnPT, as the biocide exposure was associated with a sharp decline of R(0) in both the 0.5 and the 1nM groups (-41% and -63%, respectively) in comparison to the control cohort. The population growth rate lambda (often used as an index of population fitness) and the life expectancy e(0) were also significantly reduced, while ZnPT exposure did not affect the generation time T. These results highlight the potential ecological threat posed by the biocide ZnPT, even at the very low tested concentrations, which are already detectable in some European marinas.

Responses of a Northern Adriatic Ampelisca-Corbula community to seasonality and short-term hydrological changes in the Po river.

During the 20th century, increased nutrient inflow has increased the frequency and intensity of eutrophication events in the North-western Adriatic Sea. More recently, evidence of a reduction in the Po river flow and a recovery of the benthic environment were reported. We examined the role played by Po river regime and seasonality in determining the temporal pattern of macrozoobenthos variation: samples were collected from a site located 10-12km off Cesenatico during two years from October 2004 to September 2006, when the mean Po discharge remained exceptionally low. 88 taxa were found, and the community was dominated by few species: Ampelisca diadema, Lumbrineris latreillii, Corbula gibba, Aricidea claudiae, Levinsenia gracilis and Nucula nucleus account for about 80% of total abundances. The density of these species exhibited a marked seasonal variability. Moreover, the total abundance and the density of the sensitive species, A. diadema, were negatively affected by several disturbance events (hypoxic conditions, frequent storms and an anomalous winter flood) that occurred between November 2005 and March 2006. The reduction of C. gibba and the increase of A. diadema densities observed in the investigated period, and verified recently by other authors could be related to the reduction in river inputs as a consequence of climatic changes.

Gonochorism vs. hermaphroditism: relationship between life history and fitness in three species of Ophryotrocha (Polychaeta: Dorvilleidae) with different forms of sexuality.

1. The relationships between life history, fitness and sexuality, together with their ecological and evolutionary significance, has been analysed comparing the main life-history traits and demography in three closely related species belonging to the genus Ophryotrocha. The species are: the gonochoristic O. labronica, the simultaneous hermaphrodite O. diadema and the protandrous hermaphrodite O. puerilis. 2. Survivorship and reproductive data were collected weekly and were used to construct life tables and population projection matrices for each species and compare life-history characteristics. Elasticity, life-table response and decomposition analyses were performed to examine the relative contribution of fecundity and survivorship to differences in lamda between species. 3. The gonochoristic and hermaphroditic species differ in all the main life-history parameters and also in demographic characteristics. In particular the value of lamda, used commonly to express fitness, is markedly higher in the gonochoristic species while in terms of fitness simultaneous and sequential hermaphroditism are very similar. In the genus Ophryotrocha gonochorism currently represents the most widespread condition, being characteristic of the majority of the known species in the genus. 4. Given the demographic advantage ensured by gonochorism, it remains be understood why some species have retained simultaneous hermaphroditism and one has evolved a sequential type hermaphroditism; the most probable hypothesis is correlated with the density of the species in natural habitats.

Effects of zinc exposure on the polychaete Dinophilus gyrociliatus: a life-table response experiment.

A life-table response experiment (LTRE) was performed in order to study the effects of enhanced zinc on the survival, fecundity and demographic indices of Dinophilus gyrociliatus (Polychaeta). A control group and three treated cohorts kept in artificial seawater enriched with nominal concentrations of zinc at 0.25, 0.50 and 1.00 microg Zn/ml, respectively, were considered. Zinc marginally affected survival but had a strong impact on the reproduction of D. gyrociliatus. Zinc enrichment had no effect on age at maturity, whereas it induced a slight increase in fecundity during the early reproductive phases at 0.50 and 1.00 microg Zn/ml, and a sharp decrease in fecundity from the 20th day of life at all levels of metal enrichment tested. Treated groups were characterised by a marked reduction of the net growth rate (R0) and a decrease in generation time (T) with respect to the control groups, but no effect on life expectancy (e0) or on population growth rates (lambda) was observed. R0 and lambda, which are the most commonly used indices of fitness, were not equally susceptible to zinc. Lambda appears less sensitive to stress induced by metal enrichment with respect to R0 and other life history traits like age-specific fecundity. D. gyrociliatus appears to be an appropriate bioindicator species in LTRE to assess the consequences of environmental heavy metals on individual life-history and population dynamics.

Demographic responses of the polychaete Dinophilus gyrociliatus to chromium exposure.

The effects of chromium on the life history and demography of Dinophilus gyrociliatus (Polychaeta) were studied on a control and three treated cohorts kept in artificial seawater enriched with nominal concentrations of chromium at 1.0, 1.4, and 1.6 microg/ml. Survival and fecundity were significantly reduced by chromium. Fecundity was higher in the chromium treatments than in the control group at the beginning of the reproductive period; however, overall, the highest fecundity was observed in the control. Life expectancy, net growth rate, and generation time were also significantly reduced by chromium. The population growth rate (lambda) was higher in the treated animals than in the controls. The differences in lambda between the treated animals and the controls (deltalambda) were almost completely due to the differences in the life tables observed during the first three weeks of life. The differences in survival between the treated and control animals did not contribute to deltalambda. The contribution to deltalambda of the slight increase in age-specific fecundity in the early reproductive events compensated for the marked decrease in overall fecundity in the treated animals. The complex effects of chromium enrichment on D. gyrociliatus underline the importance of analyzing both individual and population responses when assessing the effects of pollutants on organisms. Our results confirm that D. gyrociliatus might serve as an indicator species in confined areas of polluted seawater such as lagoons and harbors.