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The molecular factors and gene regulation involved in sex determination and gonad differentiation in bivalve molluscs are unknown. It has been suggested that doubly uniparental inheritance (DUI) of mitochondria may be involved in these processes in species such as the ubiquitous and commercially relevant Manila clam, Ruditapes philippinarum. We present the first long-read-based de novo genome assembly of a Manila clam, and a RNA-Seq multi-tissue analysis of 15 females and 15 males. The highly contiguous genome assembly was used as reference to investigate gene expression, alternative splicing, sequence evolution, tissue-specific co-expression networks, and sexual contrasting SNPs. Differential expression (DE) and differential splicing (DS) analyses revealed sex-specific transcriptional regulation in gonads, but not in somatic tissues. Co-expression networks revealed complex gene regulation in gonads, and genes in gonad-associated modules showed high tissue specificity. However, male gonad-associated modules showed contrasting patterns of sequence evolution and tissue specificity. One gene set was related to the structural organization of male gametes and presented slow sequence evolution but high pleiotropy, whereas another gene set was enriched in reproduction-related processes and characterized by fast sequence evolution and tissue specificity. Sexual contrasting SNPs were found in genes overrepresented in mitochondrial-related functions, providing new candidates for investigating the relationship between mitochondria and sex in DUI species. Together, these results increase our understanding of the role of DE, DS, and sequence evolution of sex-specific genes in an understudied taxon. We also provide resourceful genomic data for studies regarding sex diagnosis and breeding in bivalves.

Mitonuclear coevolution is an important prerequisite for efficient energy production in eukaryotes. However, many bivalve taxa experience doubly uniparental inheritance (DUI) and have sex-specific mitochondrial (mt) genomes, providing a challenge for mitonuclear coevolution. We examined possible mechanisms to reconcile mitonuclear coevolution with DUI. No nuclear-encoded, sex-specific OXPHOS paralogs were found in the DUI clam Ruditapes philippinarum, refuting OXPHOS paralogy as a solution in this species. It is also unlikely that mt changes causing disruption of nuclear interactions are strongly selected against because sex-specific mt-residues or those under positive selection in M mt genes were not depleted for contacting nuclear-encoded residues. However, M genomes showed consistently higher d(N)/d(S) ratios compared to putatively ancestral F genomes in all mt OXPHOS genes and across all DUI species. Further analyses indicated that this was consistently due to relaxed, not positive selection on M vs. F mt OXPHOS genes. Similarly, selection was relaxed on the F genome of DUI species compared to species with strict maternal inheritance. Coupled with recent physiological and molecular evolution studies, we suggest that relaxed selection on M mt function limits the need to maintain mitonuclear interactions in M genomes compared to F genomes. We discuss our findings with regard to OXPHOS function and the origin of DUI.

Ripristino del network uditivo dopo impianto cocleare in soggetti affetti da sordità prelinguale: studio del potenziale P300 attraverso l’utilizzo di LORETA.La letteratura è concorde su come, in pazienti affetti da ipoacusia perlinguale, il ripristino uditivo secondario ad impianto cocleare (IC) sia paragonabile ad un modello di network sinaptico attivato dall’esperienza uditiva. Scopo di questo studio è la valutazione di quali networks cerebrali si attivino secondariamente ad impianto cocleare in una popolazione di pazienti ipoacusici preverbali. In particolare, verrà valutato come alcuni fattori, quali l’età all’attivazione ed il tempo di utilizzo dell’IC, incidano nel ripristino della percezione uditiva. Sono stati valutati 20 pazienti suddivisi in 3 gruppi: impianto precoce e lungo utilizzo dell’IC (gruppo A); impianto tardivo e lungo utilizzo dell’IC (gruppo B); impianto tardivo e breve utilizzo dell’IC (gruppo C). Tutti i pazienti sono stati comparati con soggetti normoacusici di pari età. La valutazione si è avvalsa dell’utilizzo di potenziali uditivi evento correlati (ERPs); in particolare sono state valutate le latenze dei potenziali (N200 e P300) e le sorgenti corticali tramite utilizzo del software LORETA. Le latenze del potenziale P300 sono risultate aumentate nei pazienti rispetto ai controlli. Dall’analisi delle sorgenti corticali è stato riscontrato che l’attivazione corticale nei pazienti è direttamente correlata con il tempo di utilizzo del dispositivo ed inversamente correlata con l’età d’impianto. Dal confronto dei pazienti suddivisi in gruppi rispetto ai controlli, un’attivazione corticale similare nelle aree frontali e cingolata, è stata riscontrata solo nel gruppo A, in minor misura nel gruppo B e notevolmente ridotta nel gruppo C. In conclusione, l’IC apporta una reale attivazione delle aree cerebrali implicate nelle funzioni sensoriali e cognitive ma, per ottenere risposte simili ai normoudenti, necessita di un tempo di utilizzo prolungato. I risultati di questo studio evidenziano come la precocità di impianto ed il tempo di utilizzo del dispositivo siano fattori cruciali per l’attivazione dei newtorks cerebrali nei soggetti ipoacusici preverbali.The concept of auditory restoration after cochlear implant (CI) in prelingual deafness is well described by a synaptic network model, whose development depends on sensory experience. The aim of this work was to study the associative networks activated by the CI in a population of prelingually deaf patients. In particular, the impact of age at time of first CI fitting and duration of CI use was evaluated. Twenty patients were tested and divided into three groups: early implanted and lengthy CI use (group A); late implanted and lengthy CI use (group B); late implanted and short CI use (group C). Each patient group was compared with a normal hearing age matched control group. All subjects underwent to auditory event-related potentials (ERPs) registration. ERP latencies and their cortical sources were investigated. Cortical source analysis was performed using LORETA (Low Resolution Electromagnetic Tomography) software. P300 latencies were significantly longer in patients than in controls. The amount of cortical activation was found to be significantly directly correlated with duration of implant use and significantly correlated inversely with age at implant. When comparing patients and controls, comparable cortical activation was only found in A patient group, and to a lesser extent in group B, while significantly lower activation was found in patient group C in the frontal and cingulate areas. CI adds a sensory modality in deafness patients, i.e. the auditory one. This involves areas implicated in sensory and cognitive functions, and needs some time to form. The duration of CI use is crucial: our results demonstrate the importance of long term use of the device in addition to an early time of implant.

Mitochondrial DNA (mtDNA) is present in multiple copies within an organism. Since these copies are not identical, a single individual carries a heterogeneous population of mtDNAs, a condition known as heteroplasmy. Several factors play a role in the dynamics of the within-organism mtDNA population: among them, genetic bottlenecks, selection, and strictly maternal inheritance are known to shape the levels of heteroplasmy across mtDNAs. In Metazoa, the only evolutionarily stable exception to the strictly maternal inheritance of mitochondria is the doubly uniparental inheritance (DUI), reported in 100+ bivalve species. In DUI species, there are two highly divergent mtDNA lineages, one inherited through oocyte mitochondria (F-type) and the other through sperm mitochondria (M-type). Having both parents contributing to the mtDNA pool of the progeny makes DUI a unique system to study the dynamics of mtDNA populations. Since, in bivalves, the spermatozoon has few mitochondria (4–5), M-type mtDNA faces a tight bottleneck during embryo segregation, one of the narrowest mitochondrial bottlenecks investigated so far. Here, we analyzed the F- and M-type mtDNA variability within individuals of the DUI species Ruditapes philippinarum and investigated for the first time the effects of such a narrow bottleneck affecting mtDNA populations. As a potential consequence of this narrow bottleneck, the M-type mtDNA shows a large variability in different tissues, a condition so pronounced that it leads to genotypes from different tissues of the same individual not to cluster together. We believe that such results may help understanding the effect of low population size on mtDNA bottleneck.

Extending the interval between the end of neoadjuvant chemoradiotherapy (CRT) and surgery may enhance tumor response in patients with locally advanced rectal cancer. However, data on the association of delaying surgery with long-term outcome in patients who had a minor or poor response are lacking.To assess a large series of patients who had minor or no tumor response to CRT and the association of shorter or longer waiting times between CRT and surgery with short- and long-term outcomes.This is a multicenter retrospective cohort study. Data from 1701 consecutive patients with rectal cancer treated in 12 Italian referral centers were analyzed for colorectal surgery between January 2000 and December 2014. Patients with a minor or null tumor response (ypT stage of 2 to 3 or ypN positive) stage greater than 0 to neoadjuvant CRT were selected for the study. The data were analyzed between March and July 2020.Patients who had a minor or null tumor response were divided into 2 groups according to the wait time between neoadjuvant therapy end and surgery. Differences in surgical and oncological outcomes between these 2 groups were explored.The primary outcomes were overall and disease-free survival between the 2 groups.Of a total of 1064 patients, 654 (61.5%) were male, and the median (IQR) age was 64 (55-71) years. A total of 579 patients (54.4%) had a shorter wait time (8 weeks or less) 485 patients (45.6%) had a longer wait time (greater than 8 weeks). A longer waiting time before surgery was associated with worse 5- and 10-year overall survival rates (67.6% [95% CI, 63.1%-71.7%] vs 80.3% [95% CI, 76.5%-83.6%] at 5 years; 40.1% [95% CI, 33.5%-46.5%] vs 57.8% [95% CI, 52.1%-63.0%] at 10 years; P .001). Also, delayed surgery was associated with worse 5- and 10-year disease-free survival (59.6% [95% CI, 54.9%-63.9%] vs 72.0% [95% CI, 67.9%-75.7%] at 5 years; 36.2% [95% CI, 29.9%-42.4%] vs 53.9% [95% CI, 48.5%-59.1%] at 10 years; P .001). At multivariate analysis, a longer waiting time was associated with an augmented risk of death (hazard ratio, 1.84; 95% CI, 1.50-2.26; P .001) and death/recurrence (hazard ratio, 1.69; 95% CI, 1.39-2.04; P .001).In this cohort study, a longer interval before surgery after completing neoadjuvant CRT was associated with worse overall and disease-free survival in tumors with a poor pathological response to preoperative CRT. Based on these findings, patients who do not respond well to CRT should be identified early after the end of CRT and undergo surgery without delay.

Comparative genomics has become a central tool for evolutionary biology, and a better knowledge of understudied taxa represents the foundation for future work. In this study, we characterized the transcriptome of male and female mature gonads in the European clam Ruditapes decussatus, compared with that in the Manila clam Ruditapes philippinarum providing, for the first time in bivalves, information about transcription dynamics and sequence evolution of sex-biased genes. In both the species, we found a relatively low number of sex-biased genes (1,284, corresponding to 41.3% of the orthologous genes between the two species), probably due to the absence of sexual dimorphism, and the transcriptional bias is maintained in only 33% of the orthologs. The dN/dS is generally low, indicating purifying selection, with genes where the female-biased transcription is maintained between the two species showing a significantly higher dN/dS. Genes involved in embryo development, cell proliferation, and maintenance of genome stability show a faster sequence evolution. Finally, we report a lack of clear correlation between transcription level and evolutionary rate in these species, in contrast with studies that reported a negative correlation. We discuss such discrepancy and call into question some methodological approaches and rationales generally used in this type of comparative studies.

Many bivalve species have two types of mitochondrial DNA passed independently through the female line (F genome) and male line (M genome). Here we study the cytochrome oxidase I protein in such bivalve species and provide evidence for differences between the F and M proteins in amino acid property values, particularly relating to hydrophobicity and helicity. The magnitude of these differences varies between different regions of the protein and the change from the ancestor is most marked in the M protein. The observed changes occur in parallel and in the same direction in the different species studied. Two possible causes are considered, first relaxation of purifying selection with drift and second positive selection. These may operate in different ways in different regions of the protein. Many different amino acid substitutions contribute in a small way to the observed variation, but substitutions involving alanine and serine have a quantitatively large effect. Some of these substitutions are potential targets for phosphorylation and some are close to residues of functional importance in the catalytic mechanism. We propose that the observed changes in the F and M proteins might contribute to functional differences between them relating to ATP production and mitochondrial membrane potential with implications for sperm function. Ministerio de Economía y Competitividad | Ref. AGL2014-52062-R Xunta de Galicia | Ref. ED431C 2016-037

Introduction Permanent hearing impairment is the most common sensory disorder in newborns. The Universal Newborn Hearing Screening (UNHS) is widely adopted as a cost-effective procedure to achieve early identification and treatment of congenital hearing impairment, with the final goal of an improved linguistic and cognitive outcome for hearing impaired children. The Italian Ministry of Health has recently comprised UNHS in the Essential Level of Health Assistance. Nevertheless, programs still vary both across and within Italian Regions in terms of coverage, testing, referral and tracking protocols. In Friuli-Venezia Giulia region the program for the early identification of newborn and childhood hearing impairment is operative since 2012. In order to minimize the lost to follow-up cases, UNHS and childhood hearing surveillance activities have been organized in close collaboration among birth centres, paediatric audiology services, territorial Family Paediatricians and the sole regional centre for paediatric hearing loss management. Material and methods We performed a five years’ retrospective analysis of the UNHS experience in Friuli-Venezia Giulia comparing the UNHS activity of year 2013 and year 2017. The focus of the study concerns the “missing” cases. Three different typologies of “miss” cases (“documentation-miss”, “access-miss” and “pathway-miss”) have been defined in correspondence with main reasons for their occurrence. Results Births in Friuli-Venezia Giulia were 9465 and 8432, respectively in 2013 and 2017. International quality indicators improved with a gain of efficiency in 5 years’ experience. However, “missing” cases were 486 in 2013 and 321 in 2017, mainly due to the lack of an efficient documentation system. Conclusion UNHS programs have proven to be valuable and cost-effective in Friuli-Venezia Giulia and other Italian regions. New resources and efforts are required to achieve a complete standardization and informatisation of the UNHS data to avoid documentation gaps. A possible strategy would point to the opportunity to unify data management systems for all the ongoing newborn screening programs (metabolic, hearing and visual), linking the integrated IT system with the regional repository of current datasets.

Advanced MRI findings in two patients with probable chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS) are presented. Diffusion tensor imaging indices (fractional anisotropy and mean diffusivity), evaluated in both patients at baseline MRI examination before treatment and during follow-up, indicated white matter structural changes not only affecting the brainstem, which represents the primary site of inflammatory damage, but also projection (corona radiata) and associative tracts in both patients, while alterations within the corpus callosum were detected in patient 1# at follow-up. Susceptibility weighted imaging (SWI) revealed hypointense lesions in both patients, MRI spectroscopy (MRS) indicated a mildly increased Cho/NAA ratio with no evidence of lipids/lactate peaks, indicating that it may be used as a non-invasive marker to identify CLIPPERS cases suspected for progression to lymphoma.

Background Morgagni hernias present technical challenges. The laparoscopic approach for repair was first described in 1992; however, as these hernias are uncommon in adult life, few data exist on the optimal method for surgical management. The purpose of this study was to analyze a method for laparoscopic repair of Morgagni giant hernias using laparoscopic primary closure with V lock (Medtronic, Covidien). Methods This case series describes a method of laparoscopic Morgagni hernia repair using primary closure. In all patients, a laparoscopic transabdominal approach was used. The content of the hernia was reduced into the abdomen, and the diaphragmatic defect was closed with a running laparoscopic suture using a self-fixating suture. Clips were placed at the edges of the suture to secure the pledged sutures to both the anterior and posterior fascia. Demographic data such as BMI and operative and postoperative data were collected. Results Retrospectively collected data for 9 patients were analyzed. There were 1 (11.1%) males and 8 (88.8%) females. The median BMI was 29.14±52 kg/m2. The median operative time was 80±25 minutes. There were no intraoperative complications or conversions to open surgery. Patients began a fluid diet on the first postoperative day and were discharged after a median hospital stay of 3±1.87 days. In a median follow-up of 36 months, we did not observe any recurrences. Conclusions Transabdominal laparoscopic approach with primary closure of the diaphragmatic defect is a viable approach for the repair of Morgagni hernia. In our experience, the use of laparoscopic transabdominal suture fixed to the fascia allowed the closure of the defect laparoscopically with minimal tension on the repairs.

BACKGROUND Transanal endoscopic microsurgery (TEM) was introduced to combine the curativeness of full thickness excision with minimum morbidity, while traditional rectal surgery is burdened by high morbidity and mortality rates. However, while it is still a matter of considerable debate whether local excision is an adequate approach for curative resection of rectal cancer, new minimally invasive operative techniques have been introduced. The purpose of this paper was to show the indications, the tips and long term results of this technique through the review of the largest single-center database available to date. The showed results derived from the single center experience of the Clinica Chirurgica of Polytechnic University of Marche. METHODS We retrospectively reviewed a 25-year database from May 1992 to May 2017. We divided the patients into three different groups of patients according to the preoperative diagnosis: rectal cancers, adenomas and other rectal lesions. Rectal adenomas were divided into two groups according to their diameter (> or

The tissue inflammatory response can influence the outcome of anastomotic healing. Anastomotic leakage represents a dreadful complication after gastrointestinal surgery, in particular sepsis and intra-abdominal infections impair the restorative process of colic anastomoses. It has been debated whether the administration of non-steroidal anti-inflammatory drugs (NSAIDs) is a risk factor for dehiscence, since many patients receive NSAIDs in the early postoperative period. Our aim was, for the first time, to analyze the morpho-functional effects of postoperative administration of two commonly used NSAIDs, Diclofenac and Ketorolac, on the healing process of colo-colic anastomoses constructed under condition of fecal peritonitis in a rat model. Sixty adult male rats underwent two surgical procedures: peritonitis induction and colo-colic anastomosis, and were divided into three groups: 20 rats received saline; 20 rats 4 mg/kg Diclofenac and 20 rats 5 mg/kg Ketorolac. We assessed anastomosis strength, morphological features of tissue wound healing, immunohistochemical metalloproteinase 9 (MMP9) expression and collagen deposition and content by Sirius red staining and hydroxyproline level. We found no significant difference in bursting pressure, collagen content and organization and morphological features between the groups, except a significantly reduced presence of inflammatory cells and MMP9 expression in the groups treated with NSAIDs. Our findings showed that Diclofenac and Ketorolac administration did not affect post-surgical healing and did not increase the leakage risk of colo-colic anastomoses during peritonitis.

BACKGROUND: Trastuzumab (TZ) therapy requires careful monitoring of left ventricular (LV) ejection fraction (LVEF) because it can be potentially cardiotoxic. However, LVEF is an imperfect parameter and there is a need to find other variables to predict cardiac dysfunction early. Left atrium (LA) enlargement has proven to be a powerful predictor of adverse outcomes in several disease entities. HYPOTHESIS: Baseline LA volume enlargement might predict TZ‐related LV dysfunction. METHODS: HER2‐positive breast cancer patients receiving TZ and undergoing transthoracic echocardiography at baseline and at follow‐up every 3 months were retrospectively recruited. One‐hundred sixty‐two patients formed the study population. RESULTS: Baseline LAVI was dilated in 14 patients (8.6%). Mean follow‐up was 14 ± 4 months. Cardiotoxicity occurred in 24 patients (14.8%). LAVI was an independent predictor of TZ‐induced LV dysfunction in a clinical model, after adjustment for age and hypertension (odds ratio per 5‐mL/m(2) LAVI increase: 1.34, 95% confidence interval: 1.03‐1.82, P = 0.03); and in a hemodynamic model, including ventricular sizes and systolic blood pressure level (odds ratio per 5‐mL/m(2) LAVI increase: 1.34, 95% confidence interval: 1.01‐1.81, P = 0.04). The predicted probability of developing cardiotoxicity increased progressively, in parallel with LAVI values. CONCLUSIONS: Baseline LA dilatation emerges as a condition associated with the development of cardiotoxicity in HER2‐positive breast cancer patients treated with TZ.

Background The role of endoscopic ultrasound (EUS) in the management of pancreatobiliary and digestive diseases is well established in adults, but it remains limited in children. The aim of this study was to evaluate the feasibility, safety, and clinical impact of EUS use in children. Methods This is a retrospective analysis of a prospectively acquired database of consecutive pediatric (< 18 years) patients presenting an indication for EUS for pancreatobiliary and gastrointestinal disorders. Results Between January 2010 and January 2016, 47 procedures were performed in 40 children (mean age of 15.1 ± 4.7 years; range 3–18). The majority of EUS (n = 32; 68.1%) were performed for pancreatobiliary and upper gastrointestinal pathologies, including suspected common bile duct stones (CBDs), acute biliary pancreatitis, recurrent/chronic pancreatitis, cystic pancreatic mass, recurrent hypoglycemia, duodenal polyp, gastric submucosal lesion, and perigastric abscess. In only 2 out of 18 children with suspected CBDs or acute biliary pancreatitis, EUS confirmed CBDs. EUS-guided fine needle aspiration was performed in 3 (6.4%) patients. Fifteen (31.9%) procedures were performed for lower gastrointestinal tract disorders, including suspected anal Crohn’s disease, fecal incontinence, and encopresis. Overall, EUS had a significant impact on the subsequent clinical management in 87.2% of patients. Conclusion The present findings were consistent with results observed in the current relevant literature and support EUS as a safe and feasible diagnostic and therapeutic tool, which yields a significant clinical impact in children with pancreatobiliary and gastrointestinal disorders.

Bivalves exhibit an astonishing diversity of sexual systems and sex-determining mechanisms. They can be gonochoric, hermaphroditic or androgenetic, with both genetic and environmental factors known to determine or influence sex. One unique sex-determining system involving the mitochondrial genome has also been hypothesized to exist in bivalves with doubly uniparental inheritance (DUI) of mtDNA. However, the link between DUI and sex determination remains obscure. In this study, we performed a comparative gonad transcriptomics analysis for two DUI-possessing freshwater mussel species to better understand the mechanisms underlying sex determination and DUI in these bivalves. We used a BLAST reciprocal analysis to identify orthologs between Venustaconcha ellipsiformis and Utterbackia peninsularis and compared our results with previously published sex-specific bivalve transcriptomes to identify conserved sex-determining genes. We also compared our data with other DUI species to identify candidate genes possibly involved in the regulation of DUI. A total of ∼12,000 orthologous relationships were found, with 2,583 genes differentially expressed in both species. Among these genes, key sex-determining factors previously reported in vertebrates and in bivalves (e.g., Sry, Dmrt1, Foxl2) were identified, suggesting that some steps of the sex-determination pathway may be deeply conserved in metazoans. Our results also support the hypothesis that a modified ubiquitination mechanism could be responsible for the retention of the paternal mtDNA in male bivalves, and revealed that DNA methylation could also be involved in the regulation of DUI. Globally, our results suggest that sets of genes associated with sex determination and DUI are similar in distantly-related DUI species.

Feeding and swallowing impairment are present in up to 80% of oculo-auriculo-vertebral spectrum (OAVS) patients. Salivary gland abnormalities have been reported in OAVS patients but their rate, features, and relationship with phenotype severity have yet to be defined.Parotid and submandibular salivary gland hypo/aplasia was evaluated on head MRI of 25 OAVS patients (16 with severe phenotype, Goldenhar syndrome) and 11 controls.All controls disclosed normal salivary glands. Abnormal parotid glands were found exclusively ipsilateral to facial microsomia in 21/25 OAVS patients (84%, aplasia in six patients) and showed no association with phenotype severity (14/16 patients with Goldenhar phenotype vs 7/9 patients with milder phenotype, p = 0.6). Submandibular salivary gland hypoplasia was detected in six OAVS patients, all with concomitant ipsilateral severe involvement of the parotid gland (p 0.001). Submandibular salivary gland hypoplasia was associated to Goldenhar phenotype (p 0.05). Parotid gland abnormalities were associated with ipsilateral fifth (p 0.001) and seventh cranial nerve (p = 0.001) abnormalities. No association was found between parotid gland anomaly and ipsilateral internal carotid artery, inner ear, brain, eye, or spine abnormalities (p 0.6).Salivary gland abnormalities are strikingly common in OAVS. Their detection might help the management of OAVS-associated swallowing and feeding impairment.

The latest robotic bipolar vessel sealing tools have been described to be effective allowing to perform procedures with reduced blood loss and shorter operative times. The aim of this study was to assess the efficacy and reliability of these devices applied in different robotic procedures.All robotic operations, between 2014 and 2016, were performed using the EndoWrist One VesselSealer (EWO, Intuitive Surgical, Sunnyvale, CA), a bipolar fully wristed device. Data, including age, gender, body mass index (BMI), were collected. Robot docking time, intraoperative blood loss, robot malfunctioning and overall operative time were analyzed. A meta-analysis of the literature was carried out to point the attention to three different parameters (mean blood loss, operating time and hospital stay) trying to identify how different coagulation devices may affect them.In 73 robotic procedures, the mean operative time was 118.2 minutes (75-125 minutes). Mean hospital stay was four days (2-10 days). There were two post-operative complications (2.74%).The bipolar vessel sealer offers the efficacy of bipolar diathermy and the advantages of a fully wristed instrument. It does not require any change of instruments for coagulation or involvement of the bedside assistant surgeon. These characteristics lead to a reduction in operative time.

The aim of this study was to analyse audiometric and speech perception outcomes after cochlear implantation (CI) in adult and elderly patients in the first year post-CI activation. We evaluated 42 subjects who underwent CI at the Otorhinolaryngological Clinic of Padua Hospital. The subjects enrolled were post-lingually deafened patients who were unilaterally implanted for bilateral, severe-to-profound hearing loss. The overall sample was divided into three groups according to the age at the time of implantation: group A (35-49 years), group B (50-64 years) and group C (≥ 65 years). The subjects were assessed, both before and after surgery (at months 1, 3, 6 and 12), using pure tone audiometry, speech audiometry and speech perception tests and the CAP questionnaire. Statistical analysis of outcomes was using a Student's t-test for paired data. In all study groups a significant improvement was demonstrated in auditory performance examinations post-CI compared to the pre-operative scores. All subjects in all age groups obtained significant improvements in PTA scores before surgery and post-CI activation. Comparison of PTA values among the three age groups did not reveal any significant difference. Considerable improvement was obtained even in the speech audiometry thresholds in all groups at follow-up, with no significant differences between groups. The speech perception examination and CAP questionnaire showed good progress in all study groups, although younger patients tended to achieve more complex categories than older ones. In conclusion, CI is an effective treatment for severe-to-profound hearing loss with no significant differences in auditory performances between older and younger CI recipients. Even if somewhat slower, subjects older than 65 reached good performance and therefore are good candidates for a cochlear implant.Questo studio è volto alla valutazione degli outcomes audiometrici e logopedici dei pazienti anziani portatori di impianto cocleare durante il primo anno di utilizzo del dispositivo. Sono stati valutati 42 pazienti impiantati tra marzo 2010 e settembre 2014 presso l’UO ORL dell’Azienda Ospedaliera Universitaria di Padova. Sono stati inclusi nello studio pazienti affetti da sordità bilaterale postlinguale di grado severo-profondo impiantati unilateralmente. I soggetti sono stati divisi in tre gruppi in base all’epoca della chirurgia: 14 soggetti con impianto fra i 35 e i 49 anni, 14 fra i 50 e i 64 anni e 14 impiantati a un’età superiore di 65 anni. Tutti i pazienti sono stati valutati prima e dopo la chirurgia (a 1, 3, 6 e 12 mesi di follow-up) attraverso l’esecuzione di: audiometria tonale, audiometria vocale, test logopedici e somministrazione del questionario delle categorie percettive (CAP). L’analisi statistica è stata effettuata attraverso il Student’s t-test. La totalità dei soggetti nei tre gruppi hanno dimostrato significativi miglioramenti all’audiometria tonale e vocale ai controlli post chirurgici rispetto alle performance ottenute precedentemente all’impianto. In particolare si sono verificati miglioramenti della soglia audiometrica media (PTA) senza differenze statisticamente significative tra i tre gruppi. risultati ottenuti nei test logopedici e dalla somministrazione del CAP hanno dimostrato evidenti miglioramenti in tutti i tre gruppi in studio. Abbiamo riscontrato, però, che i soggetti più giovani hanno raggiunto maggiori punteggi ai controlli post impianto rispetto a quelli più anziani. Concludendo, possiamo affermare che l’impianto cocleare è un trattamento efficace per soggetti affetti da ipoacusia severa-profonda senza differenze significative nelle performance audiologiche e logopediche in relazione all’età di impianto. Anche se più lentamente, i pazienti impiantati dopo i 65 anni di età raggiungono performance ottimali e possono essere ritenuti dei candidati ottimali all’intervento.

We investigated the efficacy of colistin combined with teicoplanin or daptomycin in an experimental mouse model of multiresistant Acinetobacter baumannii infection. Animal received intraperitoneally 1ml saline containing 2×1010CFU of A. baumannii. Colistin, daptomycin, teicoplanin, and colistin plus daptomycin or teicoplanin were given by intraperitoneal administration 2h after bacterial challenge. A control group received sodium chloride solution. In the in vitro study A. baumannii showed to be susceptible only to colistin with MIC of 2mg/l. In the in vivo study, colistin alone showed a good antimicrobial efficacy. When combined with teicoplanin or daptomycin, colistin produced the lowest bacterial and the best survival rates. In immunological studies, when colistin was associated to daptomycin or teicoplanin, both the number and the cytotoxic activity of NK cells increased. In conclusion, colistin combined with teicoplanin or daptomycin may improve the therapy of multiresistant A. baumannii infection.

Introduction: The combination of Cushing’s syndrome and obesity, one of the most common manifestations of the syndrome itself, may be associated with the worsening of post-operative outcomes in cas...

Aims Exercise performance is known to predict outcome in hypertrophic cardiomyopathy (HCM), but whether sex-related differences exist is unresolved. We explored whether functional impairment, assessed by exercise echocardiography, has comparable predictive accuracy in females and males with HCM. Methods We retrospectively evaluated 292 HCM patients (46 ± 16 years, 72% males), consecutively referred for exercise echocardiography; 242 were followed for 5.9 ± 4.2 years. Results Peak exercise capacity was 6.5 ± 1.6 metabolic equivalents (METs). Sixty patients (21%) showed impaired exercise capacity (≤5 METs). Exercise performance was reduced in females, compared with males (5.6 ± 1.6 vs 6.9 ± 1.5 METs, p 50 years of age. At multivariable analysis, female sex was independently associated with impaired exercise capacity (odds ratio: 4.67; 95% confidence interval (CI): 1.83–11.90; p = 0.001). During follow-up, 24 patients (10%) met the primary endpoint (a combination of cardiac death, heart failure requiring hospitalization, sustained ventricular tachycardia, appropriate implantable cardioverter defibrillator discharge, resuscitated sudden cardiac death and cardioembolic stroke). Event-free survival was reduced in females (p = 0.035 vs males). Peak METs were inversely related to outcome in males (hazard ratio (HR) per unit increase: 0.57; 95% CI: 0.39–0.84; p = 0.004) but not in females (HR: 1.22; 95% CI: 0.66–2.24; p = 0.53). Conclusions Female patients with HCM showed significant age-related impairment in functional capacity compared with males, particularly evident in post-menopausal age groups. While women were at greater risk of HCM-related complications and death, impaired exercise capacity predicted adverse outcome only in men. These findings suggest the need for sex-specific management strategies in HCM.

Background: Celiac disease is an auto-immune disorder characterized by clinical manifestations that appear in genetically predisposed subjects after gluten ingestion. In the last years, there has been a progressive change in clinical manifestations. Our aim was to evaluate the nutritional status of children with celiac disease at diagnosis and how the gluten-free diet (GFD) influences their growth. Methods: A search on PubMed/Medline was performed using “celiac disease”, “body mass index” and “children” as key words. Medline, Scopus, PubMed publisher and Google Scholar were searched as well. We selected clinical studies describing the nutritional status of patients before and after GFD using indicators like height, weight, BMI, skeletal age. We excluded papers referred to adult population or in which other diseases were related to celiac disease. Also literature-reviews were excluded. Results: From 1999 to 2018, 10 studies were found. Overall, 1383 patients in pediatric age were evaluated for their nutritional status at diagnosis of celiac disease and after a variable period from 1 to 17 years of GFD. Indicators considered were height, weight, BMI and skeletal age. Conclusions: the nutritional status of celiac patients at diagnosis is variable including an increasing number of overweight and obese. GFD has a beneficial impact on growth changes determining a correction of BMI distribution towards a Gaussian shape. (www.actabiomedica.it)

BACKGROUND AND PURPOSE: Cranial nerve abnormalities might be observed in hemifacial microsomia and microtia (oculo-auriculo-vertebral spectrum), but the rate, features, and relationship with functional impairment or phenotype severity have not yet been defined. This study aimed at investigating absence/asymmetry, abnormal origin, morphology and course of cranial nerves, and presence/asymmetry of the foramen ovale and inferior alveolar nerve canal in a cohort of oculo-auriculo-vertebral spectrum patients. MATERIALS AND METHODS: Twenty-nine patients with oculo-auriculo-vertebral spectrum (mean age, 7 years; age range, 0.2–31 years; 12 females) underwent brain MR imaging, CT, and neurologic evaluation; 19 patients had a more severe phenotype (Goldenhar syndrome). RESULTS: Cranial nerve abnormalities were detected only in patients with Goldenhar syndrome (17/19, bilaterally in 8) and were involved the second (4/19), third (1/18), fifth (11/19), sixth (8/16), seventh (11/18), and eighth (8/18) cranial nerves. Multiple cranial nerve abnormalities were common (11/17). Eleven patients showed bone foramina abnormalities. Trigeminal and facial nerve dysfunctions were common (44% and 58%, respectively), especially in patients with Goldenhar syndrome. Trigeminal abnormalities showed a good correlation with ipsilateral dysfunction (P = .018), which further increased when bone foramina abnormalities were included. The facial nerve showed a trend toward correlation with ipsilateral dysfunction (P = .081). Diplopia was found only in patients with Goldenhar syndrome and was associated with third and sixth cranial nerve abnormalities (P = .006). CONCLUSIONS: Among patients with oculo-auriculo-vertebral spectrum, cranial nerve morphologic abnormalities are common, correlate with phenotype severity, and often entail a functional impairment. The spectrum of cranial nerve abnormalities appears wider than simple hypo-/aplasia and includes an anomalous cisternal course and partial/complete fusion of diverse cranial nerves.

Bacterial meningitis may cause inner ear fibrosis and progressive cochlear ossification with irreversible profound hearing loss (HL). Recognition of potential ossification is essential for effective management. We present a clinical case of a 4 year old boy who developed a progressive HL starting 3 weeks after meningitis. For the prospective risk of cochlear ossification, bilateral cochlear implantation (CI) was performed. Unexpectedly, unaided hearing threshold began to show improvement on the left ear, starting 4 months after meningitis and continuing for years post CI surgery. In order to explore the residual cochlear function, a trial of exclusively acoustic amplification was performed on the improved left side 5 years post implantation, providing good results. A certain degree of hearing recovery may be expected after meningitis related deafness. This case encourages surgeons to always adopt atraumatic surgical techniques that can enable the preservation of cochlear structure and residual function after CI surgery.

Recent data from mitochondrial genomics and proteomics research demonstrate the existence of several atypical mitochondrial protein-coding genes (other than the standard set of 13) and the involvement of mtDNA-encoded proteins in functions other than energy production in several animal species including humans. These results are of considerable importance for evolutionary and cellular biology because they indicate that animal mtDNAs have a larger functional repertoire than previously believed. This review summarizes recent studies on animal species with a non-standard mitochondrial functional repertoire and discusses how these genetic novelties represent promising candidates for studying the role of the mitochondrial genome in speciation.

Despite its functional conservation, mtDNA presents strikingly different features among eukaryotes, such as size, rearrangement frequency, and amount of intergenic regions. The reasons beyond this diversity have been object of extensive studies that have investigated the correlation between different mtDNA evolutionary patterns and body mass, metabolic rate, reactive oxygen species production, lifespan, etc. The fundamental role of non-adaptive processes such as random genetic drift and mutation rate in genome evolution has been recently highlighted by the work of several Authors. Under this light, mitochondrial bottlenecks and number of germ line replications are critical factors for mtDNA evolution, and different patterns of germ line differentiation could be responsible for the mtDNA diversity observed across eukaryotic lineages. Among metazoans, bivalve mollusc mtDNAs show unusual features, like hypervariable gene arrangement, high mutation rate, large amount of intergenic regions and, in some species, an unique inheritance system, the Doubly Uniparental Inheritance (DUI). In DUI species, two mitochondrial lineages are present: one is transmitted maternally (F-type), the other paternally (M-type). We exploited the DUI species Ruditapes philippinarumto study intergenic mtDNA functions, and to assess mitochondrial transcription and polymorphism in gonads. The coupling of high mutation rate with quite large genomes (i.e., large proportion of intergenic DNA) would be in contrast with the Mutation Pressure theory, but this is not true if most of that intergenic DNA has a function. We observed the presence of conserved functional elements and novel ORFs that could explain the evolutionary persistence of such regions and possibly be involved in DUI-specific aspects. Accordingly, the RNA-Seq analysis showed that mtDNA transcription is lineage-specific and it is independent from the nuclear background. This is consistent with an involvement in transcriptional control of the lineage- specific structures found inside the intergenic regions. We also found that male-transmitted and female- transmitted mtDNAs have a similar amount of polymorphism, but of a different kind. In contrast with organisms that show male-driven evolution, in this species male and female germ cells undergo a similar number of replication events due to gonad physiology, and this is reflected by the number of SNPs in M and F mtDNAs. The different kind of polymorphism is instead an effect of a different population size (different bottlenecks) and of a different efficiency of selection on the two mitochondrial lineages. Our results are consistent with the hypothesis that mtDNA evolution is strongly dependent on the dynamics of germ line formation. The DUI system offers the possibility to study the evolutionary dynamics of mtDNAs that, despite being in the same organism, experience different genetic drifts (due to different mitochondrial population size in gametes), but also different selective pressures. Actually, DUI animals are the only known organisms in which the mtDNA (i.e. the M-type) is subject to selection for male function. In a broadcast spawning organism the performance of spermatozoa is extremely important and selection for viable sperm, as well as sperm competition, is more efficient on a mtDNA which is transmitted through males. For this reason, we think that even if DUI probably arose by chance through completely stochastic events, its evolutionary persistence may be adaptive, because an increased male fitness would be beneficial for the whole species.

We investigated the efficacy of colistin combined with pexiganan in experimental mouse models of Acinetobacter baumannii infection.Adult male BALB/c mice received intraperitoneally 1 mL saline containing 2 × 10 CFU of susceptible and multiresistant A. baumannii. Two hours after bacterial challenge, animals received 1 mg/kg of colistin, 1 mg/kg of pexiganan, or 1 mg/kg of colistin plus 1 mg/kg of pexiganan.Blood culture positivity, the quantities of bacteria in the intra-abdominal fluid, the rate of lethality and immunological studies, such as immunophenotyping and NK cytotoxicity, were evaluated.In the in vitro study, A. baumannii showed susceptibility to colistin and pexiganan and a strong synergy was observed by testing colistin combined with pexiganan with fractionary inhibitory concentration index of 0.312 for both strains.In the in vivo study colistin or pexiganan alone showed a good antimicrobial efficacy. When colistin was combined with pexiganan, the positive interaction produced low bacterial counts that were statistically significant versus singly treated groups. For both strains the highest rate of survival was observed in combined-treated groups (90%).Pexiganan increased NK cytotoxic activity over the levels of infected and colistin-treated animals.In conclusion, pexiganan combined with colistin was found to be efficacious against A. baumannii infection.

Mitochondrial ORFans (Open Reading Frames having no detectable homology and with unknown function) were discovered in bivalve molluscs with Doubly Uniparental Inheritance (DUI) of mitochondria. In these animals two mitochondrial lineages are present, one transmitted through eggs (F-type), the other through sperm (M-type), each showing a specific ORFan. In the present study, we used in situ hybridization and immunocytochemistry to provide evidence for the expression of Ruditapes philippinarum male-specific ORFan (orf21): both the transcript and the protein (RPHM21) were localized in spermatogenetic cells and mature spermatozoa; the protein was localized in sperm mitochondria and nuclei, and in early embryos. Also, in silico analyses of orf21 flanking region and RPHM21 structure supported its derivation from viral sequence endogenization. We propose that RPHM21 prevents the recognition of M-type mitochondria by the degradation machinery, allowing their survival in the zygote. The process might involve a mechanism similar to that of Modulators of Immune Recognition, viral proteins involved in the immune recognition pathway, to which RPHM21 showed structural similarities. A viral origin of RPHM21 may also support a developmental role, since some integrated viral elements are involved in development and sperm differentiation of their host. Mitochondrial ORFans could be responsible for or participate in the DUI mechanism and their viral origin could explain the acquired capability of M-type mitochondria to avoid degradation and invade the germ line, that is what viruses do best: to elude host immune system and proliferate.

Background The retention of a genome in mitochondria (mtDNA) has several consequences, among which the problem of ensuring a faithful transmission of its genetic information through generations despite the accumulation of oxidative damage by reactive oxygen species (ROS) predicted by the free radical theory of ageing. A division of labour between male and female germ line mitochondria was proposed: since mtDNA is maternally inherited, female gametes would prevent damages by repressing oxidative phosphorylation, thus being quiescent genetic templates. We assessed mitochondrial activity in gametes of an unusual biological system (doubly uniparental inheritance of mitochondria, DUI), in which also sperm mtDNA is transmitted to the progeny, thus having to overcome the problem of maintaining genetic information viability while producing ATP for swimming. Results Ultrastructural analysis shows no difference in the conformation of mitochondrial cristae in male and female mature gametes, while mitochondria in immature oocytes exhibit a simpler internal structure. Our data on transcriptional activity in germ line mitochondria show variability between sexes and different developmental stages, but we do not find evidence for transcriptional quiescence of mitochondria. Our observations on mitochondrial membrane potential are consistent with mitochondria being active in both male and female gametes. Conclusions Our findings and the literature we discussed may be consistent with the hypothesis that template mitochondria are not functionally silenced, on the contrary their activity might be fundamental for the inheritance mechanism. We think that during gametogenesis, fertilization and embryo development, mitochondria undergo selection for different traits (e.g. replication, membrane potential), increasing the probability of the transmission of functional organelles. In these phases of life cycle, the great reduction in mtDNA copy number per organelle/cell and the stochastic segregation of mtDNA variants would greatly improve the efficiency of selection. When a higher mtDNA copy number per organelle/cell is present, selection on mtDNA deleterious mutants is less effective, due to the buffering effect of wild-type variants. In our opinion, a combination of drift and selection on germ line mtDNA population, might be responsible for the maintenance of viable mitochondrial genetic information through generations, and a mitochondrial activity would be necessary for the selective process. Reviewers This article was reviewed by Nick Lane, Fedor S Severin and Fyodor Kondrashov.

Complicated skin and soft tissue infections constitute a heterogeneous group of severe disorders, with surgical site infections being the most common hospital-acquired ones. The aim of our study was to investigate the synergistic and bactericidal activities of antimicrobial combinations of fosfomycin with rifampicin and tigecycline against Enterococcus faecalis, Enterococcus faecium and methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates, and also to evaluate their in vivo effects in a mouse wound infection model. In in vitro studies, the combinations of fosfomycin with rifampicin and tigecycline were both synergistic. These synergies were confirmed in in vivo studies: the drug combinations showed the highest antimicrobial effects compared to monotherapy. In conclusion, the efficacy of fosfomycin combinations, also confirmed in our in vivo model, may suggest new directions in the treatment of infected skin and a possible alternative way to control bacterial skin infection.

Mitochondrial homoplasmy, which is maintained by strictly maternal inheritance and a series of bottlenecks, is thought to be an adaptive condition for metazoans. Doubly uniparental inheritance (DUI) is a unique mode of mitochondrial transmission found in bivalve species, in which two distinct mitochondrial genome (mtDNA) lines are present, one inherited through eggs (F) and one through sperm (M). During development, the two lines segregate in a sex- and tissue-specific manner: females lose M during embryogenesis, whereas males actively segregate it in the germ line. These two pivotal events are still poorly characterized. Here we investigated mtDNA replication dynamics during embryogenesis and pre-adulthood of the venerid Ruditapes philippinarum using real-time quantitative PCR. We found that both mtDNAs do not detectably replicate during early embryogenesis, and that the M line might be lost from females around 24 h of age. A rise in mtDNA copy number was observed before the first reproductive season in both sexes, with the M mitochondrial genome replicating more than the F in males, and we associate these boosts to the early phase of gonad production. As evidence indicates that DUI relies on the same molecular machine of mitochondrial maternal inheritance that is common in most animals, our data are relevant not only to DUI but also to shed light on how differential segregations of mtDNA variants, in the same nuclear background, may be controlled during development.

Hearing loss is considered the most common congenital disease and the prevalence of neonatal deafness can be estimated between 1 and 2 cases per 1000 live births. Infant deafness must be diagnosed as early as possible and an effective therapeutic intervention needs to be carried out in order to avoid the serious consequences of hearing deprivation during the evolutionary period: alterations in the development of central auditory pathways and lack of language acquisition. The cochlear implant (CI) has proved to be the best instrument to solve the problem of auditory deprivation. In particular, the bilateral CI gives the patient access to binaural hearing which results in benefits in terms of sound localisation and discrimination. The optimal age of application of the CI is a widely discussed topic in the scientific community and the current guidelines indicate a period between 12 and 24 months of age, even though the supporters of the application before 12 months of age are nowadays increasing.The study is observational, retrospective, monocentric. 49 paediatric patients (18 years) with simultaneous bilateral CIs were included. The audiometric threshold and speech tests were carried out during the follow-up 3, 6 and 12 months after the CIs activation and when the patient reached 2 years of age.The statistical analysis showed that undergoing bilateral implantation surgery before 2 years of age allows a satisfactory audiometric performance, while there are no particular benefits in performing the surgery before 1 year of age. As far as the speech outcome is concerned, the statistical analysis didn't show significant correlation between the earlier age of implantation and better speech performance if the operation is carried out before 2.5 years of age.The results of the study indicate that the optimal age to perform the simultaneous bilateral CIs surgery is between 12 and 24 months, without demonstrating any particular benefit in carrying out the procedure before 1 year of age. This may be clinically relevant in terms of avoiding the risks of diagnostic mistakes and reducing the related surgical risk in children under 1 year of age.

Prolonged hospitalization and antibiotic therapy are risk factors for the development of methicillin-resistant Staphylococcus aureus (MRSA) infections in thermal burn patients. We used a rat model to study the in vivo efficacy of daptomycin in the treatment of burn wound infections by S. aureus , and we evaluated the wound healing process through morphological and immunohistochemical analysis. A copper bar heated in boiling water was applied on a paraspinal site of each rat, resulting in two full-thickness burns. A small gauze was placed over each burn and inoculated with 5 × 10 7 CFU of S. aureus ATCC 43300. The study included two uninfected control groups with and without daptomycin treatment, an infected control group that did not receive any treatment, and two infected groups treated, respectively, with intraperitoneal daptomycin and teicoplanin. The main outcome measures were quantitative culture, histological evaluation of tissue repair, and immunohistochemical expression of wound healing markers: epidermal growth factor receptor (EGFR) and fibroblast growth factor 2 (FGF-2). The highest inhibition of infection was achieved in the group that received daptomycin, which reduced the bacterial load from 10 7 CFU/ml to about 10 3 CFU/g ( P < 0.01). The groups treated with daptomycin showed better overall healing with epithelialization and significantly higher collagen scores than the other groups, and these findings were also confirmed by immunohistochemical data. In conclusion, our results support the hypothesis that daptomycin is an important modulator of wound repair by possibly reducing hypertrophic burn scar formation.

The success of transanal endoscopic microsurgery (TEM) for early rectal cancer depends on proper indications and strict patient selection. When unfavorable pathologic features are identified after TEM operation, total mesorectal excision is recommended to minimize the risk of recurrence. In this study, data were collected in a retrospective series of patients to determine the results of laparoscopic reoperation after TEM. All patients underwent an accurate rectal-digital examination and clinical tumor staging by transanal endosonography, CT, and/or MRI. The histologic examination included an evaluation of the free margins, depth of tumor infiltration according to International Union Against Cancer guidelines, degree of tumor differentiation, and the presence of lymphovascular and perineural invasion. When a high-risk tumor was identified, reoperation was performed within 6 weeks from TEM. The patients were divided into two groups according to the procedure performed: laparoscopic anterior resection (LAR) or laparoscopic abdominal perineal amputation (LAPR). Sixty-eight patients (5.3%) underwent reoperation: 38 underwent LAR and 30 underwent LAPR. The mean operative time was 148.24 min (± 35.8, p = 0.62). Meanwhile, the mean distance of the TEM scar from the anal verge differed statistically between the two groups (p = 0.003) and was statistically correlated with abdominal perineal amputation (p = 0.0001) in multivariate analysis. Conversion to open surgery was required in 6 patients (15.7%) in the LAR group and 3 patients (10%) in the LAPR group (p = 0.38). The histologic examination revealed residual cancer cells in 3 cases (3 pT2N0) and 1 case (1 pT3N0), respectively, and lymph node metastases in 4 cases. No residual neoplasms were detected in the remaining 60 cases (88.3%). After a mean follow-up of 108 months, the overall disease-free survival was 98% (95% CI 88–99%). In our experience, reoperation after TEM using a laparoscopic approach is feasible and safe, with low conversion rates and optimal postoperative results.

Intracranial vascular, bone, and brain abnormalities might be observed in oculo-auriculo-vertebral spectrum (OAVS) patients even though these structures do not derive embryologically from the first and second pharyngeal arches. This neuroimaging study investigated their type, frequency and phenotype correlations among OAVS patients. Thirty-five OAVS patients (mean-age 4.8 ± 6.0 years; age range 0.3–30, 14 females) underwent head CT (all patients) and brain MRI (32 patients); 19 patients had a more severe phenotype (Goldenhar syndrome). Internal carotid artery hypoplasia/agenesia and abnormal course, brain abnormalities, internal acoustic canal stenosis/aplasia, cochlear-vestibular malformations, facial nerve bony canal anomalies, and oval window atresia were recorded. Nine of 35 (26%) OAVS patients showed internal carotid artery anomalies; 18/32 (56%) had protean brain MRI abnormalities, ranging from tegmental cap anomaly to mild ventriculomegaly, which were associated with Goldenhar syndrome (p

Objectives Congenital Aural Atresia (CAA) is a deformity of the external ear and it is commonly associated with malformations of middle and inner ear and, in some cases, with other facial deformities. Very few assessment measures exist for evaluating the functional impairment in children with CAA. Purpose of this study is to introduce and describe an assessment Checklist, (nominated FOS Checklist) that covers feeding abilities (F), oralmotor skills (O), communication/language development (S) in children with CAA. FOS wants to offer a range of assessment providing a profile of the child in comparison to hearing peers and it aims to make clinicians able to identify additional problems and areas of difficulties as well as specific abilities and skills. Secondary, we want to investigate the presence of correlations between disorders and side of CAA. Methods a new Checklist (FOS Checklist) was administered to 68 children with CAA. Results Feeding abilities are age-adequate in 94,3% of all patients. 54,4% of all patients are in need for further assessment of their oral-motor skills; delays in language development were found in 44,1% of cases. Orofacial development delays have been observed in 57.2% of subjects among the bilateral CAA group, in 53.9% among the right CAA group and in 53.4% among the left CAA group. Patients referred for further language evaluation were 42,9% in the bilateral CAA group, 33.3% in the right CAA group and 33.3% in the left CAA group. According to the χ2 analysis, referral for further assessment is independent from side of aural atresia. Conclusions Subjects with bilateral CAA are more likely to be referred for further assessment, both for oral motor aspects and for speech perception and language development. However, there is not a significant statistical difference between the performances of children with bilateral or unilateral CAA. FOS Checklist is simple, reliable and time effective and can be used in everyday clinical practice. FOS enable clinicians to identify additional problems and areas of difficulties as well as specific abilities and skills; moreover, FOS allows to determine appropriate referrals and intervention strategies.

AIM: To describe our experience in treating rectal cancer by transanal endoscopic microsurgery (TEM), report morbidity and mortality and oncological outcome. METHODS: A total of 425 patients with rectal cancer (120 T1, 185 T2, 120 T3 lesions) were staged by digital rectal examination, rectoscopy, transanal endosonography, magnetic resonance imaging and/or computed tomography. Patients with T1-N0 lesions and favourable histological features underwent TEM immediately. Patients with preoperative stage T2-T3-N0 underwent preoperative high-dose radiotherapy; from 1997 those aged less than 70 years and in good general health also underwent preoperative chemotherapy. Patients with T2-T3-N0 lesions were restaged 30 d after radiotherapy and were then operated on 40-50 d after neoadjuvant therapy. The instrumentation designed by Buess was used for all procedures. RESULTS: There were neither perioperative mortality nor intraoperative complications. Conversion to other surgical procedures was never required. Major complications (urethral lesions, perianal or retroperitoneal phlegmon and rectovaginal fistula) occurred in six (1.4%) patients and minor complications (partial suture line dehiscence, stool incontinence and rectal haemorrhage) in 42 (9.9%). Postoperative pain was minimal. Definitive histological examination of the 425 malignant lesions showed 80 (18.8%) pT0, 153 (36%) pT1, 151 (35.5%) pT2, and 41 (9.6%) pT3 lesions. Eighteen (4.2%) patients (ten pT2 and eight pT3) had a local recurrence and 16 (3.8%) had distant metastasis. Cancer-specific survival rates at the end of follow-up were 100% for pT1 patients (253 mo), 93% for pT2 patients (255 mo) and 89% for pT3 patients (239 mo). CONCLUSION: TEM is a safe and effective procedure to treat rectal cancer in selected patients without evidence of nodal involvement. T2-T3 lesions require preoperative neoadjuvant therapy.

Children with Peutz-Jeghers syndrome (PJS) have increased risk of polyp-related complications and emergency laparotomies. The aim of the present study was to assess the efficacy and the safety of endoscopic therapy of small bowel polyps using single-balloon enteroscopy (SBE) in children affected by PJS.Between January 2010 and December 2011, prospectively consecutive PJS children with polyps15 mm or polyps actively bleeding previously identified using video capsule endoscopy and magnetic resonance imaging underwent therapeutic SBE. The main outcome measurements were the feasibility, the technical performance, and the safety.A total of 10 children (6 boys; median age 13.7 years, range 5.6-15.6) underwent 23 SBE procedures. Four patients had a history of abdominal surgery. A total of 53 polyps were removed, and 23 of them were15 mm. The majority of polyps were found in jejunum (85%). The mean insertion depths for antegrade and retrograde approach were 200 ± 80 and 100 ± 50 cm beyond the ileal valve, respectively. The mean procedure time was 75 ± 25 minutes. Mild abdominal pain was reported after 3 procedures. In 1 patient a postpolypectomy perforation occurred.In conclusion, SBE is an effective endoscopic tool for treating small bowel polyps in children with PJS, and well-timed polypectomy may optimize patients' care, preventing polyp-related complications and emergency laparotomy. Further larger multicenter studies are warranted to accurately determine the safety of therapeutic SBE in children.

Transanal endoscopic microsurgery (TEM) is a minimally invasive technique allowing to excise large rectal adenomas by local approach. This study shows the feasibility and results of a long term experience of TEM applied in this field.A retrospective investigation was performed in patients affected by rectal adenomas (5-11 cm in diameter) who had undergone transanal endoscopic microsurgery from 1992 to 2015. A series of 207 patients met the inclusion criteria and were analyzed for characteristics, intraoperative and postoperative results and long-term follow-up.Median age was 71 years (52-83 years). Of the 207 total patients, 77 were women (37.2%) and 130 men (62.8%). Median distance from the anal verge was 8 cm (4-20 cm). Median size was 6 cm, with lesion ranging from 5 to 11 cm in diameter. Median operative time was 57 minutes (42-90 minutes). We observed 17 (8.2%) minor complications and 2 (0.9%) major complication according to the Clavien-Dindo classification. We did not observe any intraoperative or post-operative mortality. Histological examination showed preoperative undiagnosed cancer in 8 (3.8%) cases in which the pathologist revealed adenocarcinoma staged as T1 Sm1 according to the Kikuchi classification. The median follow-up was 115 months (12-288 months). Six recurrences (2.9%), we observed with a median time for recurrence of 14 months.TEM provides excellent oncological outcomes in the treatment of large benign rectal lesions, ensuring a minimal risk of resection margin infiltration at pathological examination, and is associated with low risk of complications.

Background and aim A complex relationship between immune system and metabolic pathway exists and can induce oxidative stress. The objective of this study was to design a new methodology allowing the measurement of oxidative status of leukocytes. Methods and results We developed a flow cytometry technique, based on C11-BODIPY 581/591 staining, to evaluate peroxidation in leukocytes. We defined the Peroxidation of Leukocytes Index Ratio (PLIR) as the ratio between the damage after AAPH-induced and PMA-induced peroxidation, using Trolox as standard antioxidant. Sensitivity of the method was assessed by correlating results with plasma antioxidant capacity (TRAP and FRAP), levels of endogenous antioxidants (uric acid and sulfhydryls) and markers of metabolic status (cholesterol, triglycerides, glucose and insulin). PLIR measures the ratio between the resistance to exogenous and endogenous ROS injury, independently from baseline level of oxidation, which was directly correlated with plasma cholesterol on lymphocytes (0.738, p = 0.029), monocytes (0.691, p = 0.047) and neutrophils (0.690, p = 0.047). PLIR of lymphocytes was inversely correlated with uric acid (− 0.810, p = 0.009) and FRAP (− 0.738, p = 0.029) levels. On the other hand, PLIR of monocytes was directly correlated with the total scavenger antioxidant capacity attributable to nutritional antioxidants (0.738, p = 0.029), calculated as the difference between TRAP and the contribution of uric acid and sulfhydryls to its value. Conclusions This study reports a feasible and reproducible new flow cytometry assay for assessing the leukocytes redox status. PLIR discriminates between reducing and scavenger activities and is able to appreciate the potentially dangerous effect of uric acid on innate immune response.

STUDY DESIGN: A cross-sectional study. OBJECTIVE: To assess using postoperative magnetic resonance imaging whether the posterior longitudinal ligament (PLL) caused residual cord compression after anterior cervical decompression and fusion (ACDF) in a series of patients in whom the PLL was retained. SUMMARY OF BACKGROUND DATA: There is a lack of data evaluating the postoperative compressive effects of the PLL in patients undergoing ACDF providing guidance as to whether to remove or retain the PLL during discectomy to facilitate adequate decompression. METHODS: Postoperative gadolinium enhanced magnetic resonance images were reviewed in a series of 33 patients who underwent ACDF for cervical radiculomyelopathy and who had persistent or recurrent postoperative symptoms. Patients with ossification of the posterior longitudinal ligament or with a herniated disc behind the PLL were excluded from this study. RESULTS: There were no cases of discernible compression by the retained PLL identified on the magnetic resonance image (P < 0.001) as assessed by 2 independent reviewers. Four patients underwent subsequent revision surgery unrelated to the PLL. CONCLUSION: We were unable to demonstrate magnetic resonance imaging evidence to suggest that the retained PLL caused compression after ACDF in this patient cohort. Therefore we suggest that removing the PLL should be considered for reasons other than concern about residual compression.

Restaurant foods have been shown to be high in sodium and limited sodium content information provided through menu nutrition information (MNI) is available at the point of purchase. Dining out and use of MNI are behaviors that can be altered by consumers who are trying to decrease their sodium intake.The aim of this study was to determine the relationship between reported consumer actions to decrease sodium intake and dining out frequency and awareness and use/or intended use of MNI.A secondary analysis was conducted using responses from 5,588 US adults aged 20 years or older who participated in the 2013-2014 cross-sectional National Health and Nutrition Examination Survey household interview.The main outcomes were dining out frequency and seeing MNI, using MNI if seen, or would use MNI if provided.Linear and logistic regression models were used to assess the relationship of consumers reporting and not reporting action to decrease sodium intake and the outcome measures.Reported consumer action to decrease sodium intake compared to no action was associated with an overall decreased dining out frequency of approximately one meal per week (mean±standard error=3.12±0.10 compared to 4.11±0.14; P0.01). When separated by type of restaurant, the relationship was significant for fast-food or pizza establishments (mean±standard error=1.35±0.05 meals compared to 2.00±0.07 meals; P0.001), but not other types of foodservice operations. The odds of seeing MNI, using MNI when seen, or would use MNI if provided were higher for consumers reporting actions to decrease their sodium intake compared to those who were not for both fast-food or pizza establishments and restaurants with wait staff (odds ratio ranged from 1.17 [95% CI 1.04 to 1.32] to 2.24 [95% CI 1.82 to 2.76]; P values ranged from0.05 to0.001).Compared to consumers reporting no actions to decrease sodium intake, consumers reporting actions indicate they dine out less frequently, specifically at fast-food or pizza restaurants and report they are more likely to use MNI. These results may inform the restaurant industry of the actions of a potentially growing consumer group and provide insights for future public health initiatives targeting population sodium reduction.

Musculista senhousia is a marine mussel with Doubly Uniparental Inheritance (DUI) of mitochondria. In this study we analysed the largest unassigned region (LUR) of its female- and male- transmitted mitochondrial genomes, described their fine characteristics and searched for shared features. Our results suggest that both LURs contain the control region of their respective mitochondrial genomes. The female-transmitted control region is duplicated in tandem, with the two copies evolving in concert. This makes the F-mtDNA of M. senhousia the first Bivalve mitochondrial genome with this feature. We also compared M. senhousia control regions to that of other Mytilidae, and demonstrate that signals for basic mtDNA functions are retained over evolutionary times even among the fast-evolving mitochondrial genomes of DUI species. Finally, we discussed on how similarities between female and male LURs may be explained in the context of DUI evolution and if the duplicated female control region might have influenced the DUI system in this species.

In species with Doubly Uniparental Inheritance (DUI), males are heteroplasmic for two sex-linked mitochondrial genomes (M- and F-mtDNA). While a role of M-mtDNA in male gametogenesis and sperm function is evident, there is an ongoing debate on whether it is transcribed or not in male soma. In this work we report a qPCR analysis in the DUI species Ruditapes philippinarum, showing that M-mtDNA is transcribed in somatic tissues. We observed a correlation between DNA copy numbers of the two analyzed genes, cytochrome b and a novel male-specific mitochondrial gene thought to be involved in DUI (orf21), and between their transcription levels. No correlation between a transcript and its DNA copy number was found, supporting the existence of complex regulatory mechanisms of mitochondrial transcription. We found the highest amount of mtDNA and mtRNA in gonads, likely due to the intense cell proliferation and high energy request for gametogenesis, while the observed variation among specimens is probably related to their different stage of gonad development. Finally, orf21 showed a highly variable transcription in advanced stages of gametogenesis. We hypothesize a differential storage of orf21 transcripts in spermatozoa, representing different paternal contributions to progeny, possibly leading to different developmental outcomes. A transcriptional activity does not necessarily imply the translation of M- mtDNA genes, and studies on mitochondrial proteins and their localization are needed to definitively assess the functioning of male-transmitted mitochondria in male soma. All that considered, the male soma of DUI species may represent an intriguing experimental model to test for cytoplasmatic genome conflicts.

Background Hepatitis B vaccination has proven to be very safe and highly effective. This study assessed the proportion of successfully vaccinated individuals among cases with acute hepatitis B, the proportion of preventable cases if individuals were vaccinated as recommended, and the reasons for failures. Methods We analyzed data reported to the Italian Surveillance System for Acute Viral Hepatitis from 1993 to 2014. Results A total of 362 of 11 311 (3.2%) cases with acute hepatitis B were vaccinated. Of the 277 cases for whom immunization data were available, 50 (18%) received a complete vaccination course according to the correct schedule and before exposure to hepatitis B virus. Molecular characterization of 17 of these cases showed that 6 were infected with S-gene mutants. Among the 10 949 unvaccinated cases, 213 (1.9%) escaped mandatory vaccination and 2821 (25.8%) were not vaccinated despite being at increased risk of infection. Among the latter, the most common risk factors were cohabitation with hepatitis B surface antigen (HBsAg) carriers, intravenous drug use, and homosexual/bisexual practices. Thirty-seven percent of the unvaccinated households with HBsAg carriers were aware of their risk. Lack of trust in the vaccination, negative attitude, and inaccurate beliefs followed by lack of or poor communication and low perceived severity of the disease were the most frequent reasons for vaccine hesitancy. Conclusions Development of acute disease in successfully vaccinated individuals is a rare event. Further efforts are needed to enhance the vaccine coverage rate in individuals at increased risk of infection.

The aim of this work was to determine the in vitro activity of tigecycline and its bactericidal effect for a large number of Gram-positive cocci, as well as to investigate its in vitro interaction with six clinically used antibiotics. In vivo, a wound model was established through the panniculus carnosus of BALB/c mice, and then inoculated with 5 × 10(7) colony-forming units (CFU) of Staphylococcus aureus or Enterococcus faecalis. For each bacterial strain, the study included an infected or non-infected group that did not receive any treatment, three groups singly treated with tigecycline, rifampin, and daptomycin, and two groups that received tigecycline treatment plus rifampin or daptomycin. In the in vitro studies, tigecycline, daptomycin, and teicoplanin were active against all of the 48 Gram-positive isolates. The combination of tigecycline with rifampicin and daptomycin was synergistic against S. aureus and Enterococcus spp. In the in vivo studies, all groups treated with single drugs showed statistically significant results compared to the control group. The two groups treated with a combination of drugs showed the highest antimicrobial efficacy. In conclusion, our results suggested a strong activity of tigecycline alone and in combination with other antimicrobial agents against multi-resistant Gram-positive organisms isolated from wound infections.

Background Gram-negative sepsis ranks as the leading cause of death in intensive care units, and its incidence is increasing steadily and mortality rates has not changed much over recent decades. Materials and Methods We investigated the efficacy of the amphibian peptide, citropin 1.1 alone and in combination with tazobactam-piperacillin (TZP) in two experimental mice models of gram-negative sepsis. Animals were given an intraperitoneal injection of (1) 1 mg Escherichia coli 0111:B4 LPS, and (2) 2 × 10 10 CFU of E. coli ATCC 25922. For each model, all animals were randomized to receive intraperitoneally isotonic sodium chloride solution, 1 mg/Kg citropin 1.1 and 120 mg/Kg of TZP, and finally 1 mg/Kg citropin 1.1 plus 60 mg/Kg of TZP. Lethality, bacterial growth in blood and peritoneum, and oxidative stress indices in plasma were evaluated. Results All compounds reduced the lethality compared with controls. Treatment with citropin 1.1 resulted in significant decrease in plasma endotoxin and cytokine levels, while TZP exerted opposed effect. The combination between citropin 1.1 and TZP proved to be the most effective treatment in reducing all variables measured. Conclusion Due to its multifunctional properties, citropin 1.1 may become an important future consideration to treat conditions in which oxidative organ failure may be present.

An experimental study was performed to evaluate the interaction between s-thanatin and colistin both in vitro and in vivo, using two Pseudomonas aeruginosa strains with different patterns of susceptibilities. We evaluated whether selecting for colistin-resistant P. aeruginosa could be prevented in vitro by combining colistin with s-thanatin. The strains were serially exposed in broth to twofold stepwise increasing concentrations of colistin alone or in combination with a fixed concentration [0.25× minimum inhibitory concentration (MIC)] of s-thanatin. We also performed an in vitro synergy study. For in vivo studies, a mouse model of Pseudomonas sepsis has been used. Main outcome measures were lethality and quantitative blood cultures. Exposure to colistin alone gradually selected for Pseudomonas strains with an increased MIC. In vitro studies, s-thanatin showed a positive interaction with colistin, and was able to prevent its resistance. In vivo studies, s-thanatin combined with colistin exhibited the highest efficacy on all main outcome measurements. These results highlight the potential usefulness of this combination and provide a future therapeutic alternative in severe Pseudomonas infections.

We investigated the efficacy of the peptide s-thanatin alone and in combination with tigecycline in an animal model of sepsis induced by cecal ligation and puncture. Adult male Wistar rats were randomized to receive intravenously isotonic sodium chloride solution, 5 mg/kg s-thanatin, 2 mg/kg tigecycline, 5 mg/kg s-thanatin combined with 2 mg/kg tigecycline. The experiment was also performed with administration of the drugs 360 min after the surgical procedure to better investigate the clinical situation where there is an interval between the onset of sepsis and the initiation of therapy. Lethality, bacterial growth in blood, peritoneum, spleen and liver, and NO indices were evaluated. All compounds reduced the lethality when compared to control. In all experiments, the compounds reduced significantly bacterial growth and lethality compared with saline treatment. Treatment with s-thanatin resulted in significant decrease in plasma NO levels compared to tigecycline and control group. The combination between s-thanatin and tigecycline proved to be the most effective treatment in reducing all variables measured. S-thanatin may have potential therapeutic usefulness alone and when associated to tigecycline in polymicrobial peritonitis.

The aim of this study was to evaluate the influence of short-term dietary intake of bread obtained by a selected variety of old grain grown in Tuscany, Italy on some parameters related to the atherosclerotic process. Twenty healthy subjects (median age, 39.5 years) followed for 10 weeks a diet containing bread (150 g/day) made from the test grain (test period) and for the same period a diet containing commercially available bread of the same quantity (control period). Lipid, inflammatory, and hemorheological profiles before and after dietary intervention were evaluated. The test period showed a significant (P < .05) improvement of total cholesterol (pre-intervention, 211.2 +/- 10.8 mg/dL; post-intervention, 196.5 +/- 9.8 mg/dL) and low-density lipoprotein-cholesterol levels (pre-intervention, 137.5 +/- 8.1 mg/dL; post-intervention, 119.5 +/- 7.5 mg/dL), whereas no significant changes during the control period were observed. With regard to inflammatory and hemorheological parameters, the test period showed a significant decrease in some of the parameters investigated (interleukin-8 [pre-intervention vs. post-intervention, 67.4 +/- 10.7 vs. 43.9 +/- 4.1 pg/mL], whole blood viscosity at high [4.36 +/- 0.03 vs. 4.32 +/- 0.03 mPa x s, respectively] and low [26.1 +/- 0.4 vs. 24.8 +/- 0.5 mPa x s, respectively] shear rates, and erythrocyte filtration [8.4 +/- 0.7% vs. 9.1 +/- 0.6%, respectively]) relative to the control period, which showed no significant changes. Short-term dietary intake of whole grain bread obtained from an old grain variety seems to impose a favorable status with regard to lower circulating levels of markers of atherosclerosis.