Peri-operative management of endoscopic endonasal cerebrospinal fluid leak repair: an international survey.

PURPOSE: Peri-operative management of nasal cerebrospinal fluid (CSF) leaks is not consensual due to limited evidence. The main aim of this study was to identify key factors in peri-operative management of endoscopic endonasal CSF leak repair among international experts. METHODS: A 60-item survey questionnaire collected opinions of members of international learned societies of ENT surgeons and neurosurgeons on nasal packing, post-operative instructions, antibiotic prophylaxis, and CSF volume depletion. RESULTS: The survey had 153 respondents (124 otorhinolaryngologists and 29 neurosurgeons). A resting position was recommended by 85% (130/151) of respondents for extended CSF leak of the anterior skull base, mainly in Fowler's position (72% (110/153)). Nasal packing was used by 85% (130/153) of respondents; 33.3% (51/153) used it to stabilize the reconstruction, and 22.2% (34/153) to prevent bleeding. It was usually removed after 48 h in 44.4% of cases (68/153). CSF depletion was considered by 47.1% (72/153) of respondents in case of CSF leak recurrence and by 34.6% (53/153) in cases of increased intracranial pressure. All respondents gave specific postoperative instructions to patients including driving, running, swimming, diving restrictions and flighting restrictions. In subgroup analysis, ENT surgeons more often recommended a resting position than neurosurgeons (71% vs. 37.9% ; p = 0.0008) and prescribed more antibiotics (82.3% vs. 21.4% ; p < 0.0001). CONCLUSION: Although postoperative management after CSF closure remains challenging and not codified, this international survey revealed some points of consensus concerning resting position and restriction of post-operative activities. Prospective clinical studies must be undertaken to evaluate their efficiency.

Extended sphenoidotomy combined with transpterygoid approach for sphenoidal sinus inverted papilloma.

PURPOSE: Inverted papilloma of the sphenoid sinus (IPSS) is a rare tumor with debated surgical management due to its proximity to vital structures. The aim of this manuscript is to highlight the role of a transpterygoid approach (TPA) and pedicle-orientated strategy in case of involvement of critical structures in IPSS and compare it with data from the literature. METHODS: Patients with primary IPSS between January 2000 and June 2021 were included. Pre-operative CT/MRI were analyzed to classify the pneumatization of the SS and predict the insertion point of the inverted papilloma. All patients were treated with a trans-sphenoidal approach which was combined with a TPA in case of lateral insertion point. A systematic search was also performed to summarize the available literature. RESULTS: Twenty-two patients were treated for IPSS. By CT, the SS was categorized with type III pneumatization in 72.8% of cases. Eleven patients (50%) were treated with a TPA with a statistical association with the insertion point on the SS lateral wall (p = 0.01), rather than a SS pneumatization (p = 0.63). The overall success was 95.5% after a mean follow-up of 35.9 months. For the literature, 26 publications were included on 97 patients and described a trans-sphenoidal approach with a success rate of 84.6% after a mean follow-up of 24.5 months. CONCLUSION: IPSS is generally treated with a sphenoidotomy approach, although in selected cases, a TPA should be preferred to expose the whole SS lateral wall though allowing a complete pedicled oriented resection of the tumor.

Venous sinus stenting after repair of skull base spontaneous cerebrospinal fluid leaks: A single-center retrospective cohort series with case-control analysis.

BACKGROUND: Management of idiopathic intracranial hypertension (IIH) is recommended after surgical repair of spontaneous cerebrospinal fluid leaks (sCSF-leaks) of the skull base for prevention of recurrence. PURPOSE: To assess the feasibility of venous sinus stenting, a treatment commonly used for the treatment of IIH associated with intracranial venous sinus stenosis (VSS), after sCSF-leaks closure. MATERIALS AND METHODS: A single-center cohort series of consecutive patients who underwent sCSF-leak closure was retrospectively analyzed. Stenting was considered either for leak recurrence or in prophylactic manner after repair in patients with VSS as confirmed by cerebral venous imaging. Leak recurrence, need for new repair or adjunctive treatment of IIH, meningitis, and stenting complications were determined at the last follow-up. Cases who had prophylactic stenting were compared to historical controls before stenting option. RESULTS: Twenty-two patients had intracranial venous stenting after sCSF-leak closure. Their median age was 58 years (Q1=45; Q3=68), BMI=31 kg.m-2 (Q1=27; Q3=36), and female rate=85%. The overall rate of successful repair after stenting was 95% (95% CI = 87-100%) at a median follow-up of 2.4 years (Q1=1.2; Q3=3.3). Adjunctive treatment for IIH was needed in 4 patients (4/22, 18%) including 2 patients without leak recurrence. No meningitis, permanent morbidity or mortality was observed after stenting. Compared to 18 controls, cases had significantly less recurrence (P = 0.03), and a trend for less adjunctive treatment for IIH (P = 0.06). CONCLUSIONS: Our study suggests that stenting might be a valid option for prevention of sCSF-leak recurrences after repair in patients with intracranial venous sinus stenosis.

Strategy to Develop and Evaluate a Multiplex RT-ddPCR in Response to SARS-CoV-2 Genomic Evolution.

The worldwide emergence and spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) since 2019 has highlighted the importance of rapid and reliable diagnostic testing to prevent and control the viral transmission. However, inaccurate results may occur due to false negatives (FN) caused by polymorphisms or point mutations related to the virus evolution and compromise the accuracy of the diagnostic tests. Therefore, PCR-based SARS-CoV-2 diagnostics should be evaluated and evolve together with the rapidly increasing number of new variants appearing around the world. However, even by using a large collection of samples, laboratories are not able to test a representative collection of samples that deals with the same level of diversity that is continuously evolving worldwide. In the present study, we proposed a methodology based on an in silico and in vitro analysis. First, we used all information offered by available whole-genome sequencing data for SARS-CoV-2 for the selection of the two PCR assays targeting two different regions in the genome, and to monitor the possible impact of virus evolution on the specificity of the primers and probes of the PCR assays during and after the development of the assays. Besides this first essential in silico evaluation, a minimal set of testing was proposed to generate experimental evidence on the method performance, such as specificity, sensitivity and applicability. Therefore, a duplex reverse-transcription droplet digital PCR (RT-ddPCR) method was evaluated in silico by using 154 489 whole-genome sequences of SARS-CoV-2 strains that were representative for the circulating strains around the world. The RT-ddPCR platform was selected as it presented several advantages to detect and quantify SARS-CoV-2 RNA in clinical samples and wastewater. Next, the assays were successfully experimentally evaluated for their sensitivity and specificity. A preliminary evaluation of the applicability of the developed method was performed using both clinical and wastewater samples.

Germline mutations in the new E1' cryptic exon of the VHL gene in patients with tumours of von Hippel-Lindau disease spectrum or with paraganglioma.

BACKGROUNDS: The incidence of germline mutations in the newly discovered cryptic exon (E1') of VHL gene in patients with von Hippel-Lindau (VHL) disease and in patients with paraganglioma or pheochromocytoma (PPGL) is not currently known. METHODS: We studied a large international multicentre cohort of 1167 patients with a previous negative genetic testing. Germline DNA from 75 patients with a single tumour of the VHL spectrum ('Single VHL tumour' cohort), 70 patients with multiple tumours of the VHL spectrum ('Multiple VHL tumours' cohort), 76 patients with a VHL disease as described in the literature ('VHL-like' cohort) and 946 patients with a PPGL were screened for E1' genetic variants. RESULTS: Six different genetic variants in E1' were detected in 12 patients. Two were classified as pathogenic, 3 as variants of unknown significance and 1 as benign. The rs139622356 was found in seven unrelated patients but described in only 16 patients out of the 31 390 of the Genome Aggregation Database (p<0.0001) suggesting that this variant might be either a recurrent mutation or a modifier mutation conferring a risk for the development of tumours and cancers of the VHL spectrum. CONCLUSIONS: VHL E1' cryptic exon mutations contribute to 1.32% (1/76) of 'VHL-like' cohort and to 0.11% (1/946) of PPGL cohort and should be screened in patients with clinical suspicion of VHL, and added to panels for Next Generation Sequencing (NGS) diagnostic testing of hereditary PPGL. Our data highlight the importance of studying variants identified in deep intronic sequences, which would have been missed by examining only coding sequences of genes/exomes. These variants will likely be more frequently detected and studied with the upcoming implementation of whole-genome sequencing into clinical practice.

Functional treatment of temporal bone chondroblastoma: retrospective analysis of 3 cases.

PURPOSE: To describe and analyse functional treatment of temporal bone chondroblastoma (TBCh). METHODS: From January 2000 to June 2019, at the Department of Otorhinolaryngology, Hôpital Lariboisière, Paris, France, patients with TBCh were enrolled in this study. All cases routinely performed pre-operatory work-up including evaluation of performance status, audio-vestibular function test, ear endoscopy, contrasted CT scan and MRI of head and neck region; in one case we also performed an angio-CT scan. All patients underwent resection of the tumour with a "functional" approach RESULTS: Three male patients (mean age 46,6 years)-two primary tumours and one recurrence-were treated. In all three cases the tumour invaded the middle ear with a variable degree of hearing loss and infiltration of temporal bone structures. All surgeries were performed with a microscopic approach associated with open/endoscopic approach when necessary. Inner ear and facial nerve were speared in all cases and the TMJ was partially resected in 2 cases due to its moderate involvement. At present, after a mean follow-up of 103 months (range 40-225 months), only one case presented recurrence which has been successfully treated with radiotherapy. CONCLUSIONS: Our results of treatment suggest that functional surgery can be relevant in the management of TBCh since it is focused on both treatment of this pathology and maintenance of a good quality of life.

MRI contribution for the detection of endolymphatic hydrops in patients with superior canal dehiscence syndrome.

PURPOSE: Patients with superior semicircular canal dehiscence syndrome, which can only be treated by surgery, present cochleo-vestibular symptoms related to a third-mobile window but also endolymphatic hydrops. Since cVEMP and oVEMP are disturbed by the presence of the dehiscence, the aim of the study is to assess the value of MRI for the diagnosis of endolymphatic hydrops in patients with superior semicircular canal dehiscence syndrome in comparison with cVEMP and oVEMP. METHODS: In this retrospective cohort study we enrolled 33 ears in 24 patients with superior semicircular dehiscence syndrome who underwent a 4-h delayed intravenous Gd-enhanced 3D-FLAIR MRI and pure tone audiometry, cVEMP and oVEMP. For each patient MRI images were evaluated by two radiologists who used a compartmental endolymphatic hydrops grading system in comparison with cVEMP and oVEMP. RESULTS: Endolymphatic hydrops was found on MRI in 9 out of 33 SCDS ears (27.3%). We found no significant correlation between the presence of endolymphatic hydrops on MRI and cVEMP and oVEMP (p = 0.36 and p = 0.7, respectively). However, there was a significant correlation between the presence of endolymphatic hydrops on MRI and the degree of sensorineural hearing loss, Air Conduction-Pure Tone Average level (p = 0.012) and Bone Conduction-Pure Tone Average level (p = 0.09), respectively. CONCLUSION: We demonstrated that EH might be observed in 27.3% of superior semicircular dehiscence syndrome ears. The role of inner ear MRI is important to detect endolymphatic hydrops, since cVEMP and oVEMP are disturbed by the presence of the dehiscence, because these patients could benefit from a medical treatment. LEVEL OF EVIDENCE: Level 3.

Electrophysiological and inner ear MRI findings in patients with bilateral vestibulopathy.

PURPOSE: Bilateral vestibulopathy (BV) is an uncommon disorder and the etiology remained idiopathic in most cases. Delayed 3D-FLAIR sequences have provided new insights into various inner ear diseases, allowing the evaluation of the endolymphatic space and the permeability of the blood-labyrinthine barrier (BLB). The aim of this study was to assess both the morphology of the endolymphatic space and the permeability of the BLB in patients with BV as evaluated by delayed 3D-FLAIR sequences. METHODS: In this retrospective study, we performed 3D-FLAIR sequences 4 h after administering contrast media to 42 patients with BV. Two radiologists independently evaluated the morphology of the endolymphatic space (either vestibular atelectasis or endolymphatic hydrops) and the permeability of the BLB. RESULTS: Morphologic anomalies of the endolymphatic space and vestibular blood-labyrinthine barrier impairment were observed in 59.6% of patients with BV. Bilateral vestibular atelectasis (VA) was found in 21 patients (50%), involving only the utricle and all three ampullas while the saccule was always observed with no sign of collapse: idiopathic BV (n = 19), aminoglycoside administration (n = 1) and few days following abdominal surgery (n = 1). One patient had bilateral vestibular malformation. BLB impairment was observed in five patients (11.9%): paraneoplastic (n = 1), lymphoma (n = 1), autoimmune (n = 1), and vestibular "neuritis" (n = 2). Seventeen patients (40.4%) had normal MRI with no endolymphatic space anomaly or BLB impairment. CONCLUSION: Patients with BV presented with morphologic anomalies of the endolymphatic space or BLB impairment in 59.6% of patients.

Use of Whole Genome Sequencing Data for a First in Silico Specificity Evaluation of the RT-qPCR Assays Used for SARS-CoV-2 Detection.

The current COronaVIrus Disease 2019 (COVID-19) pandemic started in December 2019. COVID-19 cases are confirmed by the detection of SARS-CoV-2 RNA in biological samples by RT-qPCR. However, limited numbers of SARS-CoV-2 genomes were available when the first RT-qPCR methods were developed in January 2020 for initial in silico specificity evaluation and to verify whether the targeted loci are highly conserved. Now that more whole genome data have become available, we used the bioinformatics tool SCREENED and a total of 4755 publicly available SARS-CoV-2 genomes, downloaded at two different time points, to evaluate the specificity of 12 RT-qPCR tests (consisting of a total of 30 primers and probe sets) used for SARS-CoV-2 detection and the impact of the virus' genetic evolution on four of them. The exclusivity of these methods was also assessed using the human reference genome and 2624 closely related other respiratory viral genomes. The specificity of the assays was generally good and stable over time. An exception is the first method developed by the China Center for Disease Control and prevention (CDC), which exhibits three primer mismatches present in 358 SARS-CoV-2 genomes sequenced mainly in Europe from February 2020 onwards. The best results were obtained for the assay of Chan et al. (2020) targeting the gene coding for the spiking protein (S). This demonstrates that our user-friendly strategy can be used for a first in silico specificity evaluation of future RT-qPCR tests, as well as verifying that the former methods are still capable of detecting circulating SARS-CoV-2 variants.

Evaluating the prognostic potential of the Ki67 proliferation index and tumour-infiltrating lymphocytes in olfactory neuroblastoma.

AIMS: Olfactory neuroblastomas (ONBs) are rare malignant tumours that arise in the nasal vault. To date, the Hyams grade remains the only widely used histological grading system. However, it is based only on morphological criteria, and has not been updated since 1988. The objective of this study was to explore the prognostic potential of the Ki67 proliferation index (PI) and tumour-infiltrating lymphocytes (TILs) in ONB. METHODS AND RESULTS: A retrospective study was conducted on a bicentric series of 45 cases. The Ki67 PI was determined by counting at least 1000 nuclei on whole slides. TILs were evaluated with CD20, CD4 and CD8 immunohistochemical markers on whole slides. In this series, Hyams grades I, II, III and IV accounted for 13.4%, 44.4%, 20% and 22.2% of all cases, respectively. The Ki67 PI ranged from 1 to 93; the Ki67 PI was significantly higher in Hyams grade III-IV ONBs than in Hyams grade I-II ONBs (P < 0.0001). A Ki67 PI of ≥25 was associated with poorer survival (P = 0.02). TILs were present in both stromal and intratumoral compartments, but were located predominantly in the stromal component of the tumour. The numbers of intratumoral CD8+ cells/mm2 and CD4+ cells/mm2 were greater in high-grade ONBs than in low-grade ONBs (P = 0.0015 and P = 0.043, respectively). The numbers of T cells/mm2 and B cells/mm2 were not associated with survival, but a CD4/CD8 ratio of >2 was significantly associated with shorter survival (P = 0.04). CONCLUSION: Our findings suggest that the Ki67 PI and TILs could be used as prognostic markers, as a potential alternative to the Hyams grade.

Development and validation of an integrated DNA walking strategy to detect GMO expressing cry genes.

BACKGROUND: Recently, an integrated DNA walking strategy has been proposed to prove the presence of GMO via the characterisation of sequences of interest, including their transgene flanking regions and the unnatural associations of elements in their transgenic cassettes. To this end, the p35S, tNOS and t35S pCAMBIA elements have been selected as key targets, allowing the coverage of most of GMO, EU authorized or not. In the present study, a bidirectional DNA walking method anchored on the CryAb/c genes is proposed with the aim to cover additional GMO and additional sequences of interest. RESULTS: The performance of the proposed bidirectional DNA walking method anchored on the CryAb/c genes has been evaluated in a first time for its feasibility using several GM events possessing these CryAb/c genes. Afterwards, its sensitivity has been investigated through low concentrations of targets (as low as 20 HGE). In addition, to illustrate its applicability, the entire workflow has been tested on a sample mimicking food/feed matrices analysed in GMO routine analysis. CONCLUSION: Given the successful assessment of its performance, the present bidirectional DNA walking method anchored on the CryAb/c genes can easily be implemented in GMO routine analysis by the enforcement laboratories and allows completing the entire DNA walking strategy in targeting an additional transgenic element frequently found in GMO.

Risk assessment of maternally inherited SDHD paraganglioma and phaeochromocytoma.

BACKGROUND: Germline mutations in the SDHD tumour suppressor gene (11q23.1) predispose to phaeochromocytomas and paragangliomas (PPGL) mainly on a paternal transmission. However, PPGL have been recently reported in three carriers of a maternally inherited SDHD mutation. OBJECTIVE: To assess the risk of PPGL occurrence on maternal transmission of SDHD mutation. METHODS: Pedigrees of 80 SDHD-related families have been reviewed. 35 asymptomatic subjects carrying a maternally transmitted SDHD mutation were identified. 20 of them accepted to benefit from a PPGL imaging screening. RESULTS: A unique histologically proven biochemically negative phaeochromocytoma has been diagnosed in a 35-year-old woman. Molecular investigations carried out on tumour tissue revealed that the loss of heterozygosity encompassed the paternally derived q arm and the maternally derived p arm of chromosome 11. CONCLUSIONS: This study demonstrates that the risk of developing PPGL for a subject carrying a germline SDHD mutation on the maternal allele remains a rare scenario but does exist. Our data suggest an adjustment of current genetic counselling and clinical care recommendations for at-risk subjects. A targeted familial genetic test should be proposed from the age of 18 years to every subject having a mother carrying a germline SDHD mutation and a first medical workup, including imaging, should be recommended to SDHD-positive mutation carriers.

Author's reply to "Letter to the editor on the article: Saccular measurements in routine MRI can predict hydrops in Menière's disease by Simon F et al."

This is an answer to the Letter to the Editor regarding our previously published article « Saccular measurements in routine MRI can predict hydrops in Menière's disease ¼. We thank the authors for their interest in our article and their insightful comments. We would like to emphasise that our article was a preliminary study and to our knowledge the first published series establishing a correlation between measurements of saccular morphology in T2-weighed 3D CISS images and clinical definite Menière's disease. Therefore, we agree with the authors of the Letter that verifying reproducibility is paramount for the technique to be widely used. Further studies should be conducted to investigate the risk of artefacts reducing the reliability of saccular width measurements and to confirm the clinical implications. We recommend the use of saccular height measurements which have higher reliability. Also, the goal of the study was to present a possible radiological alternative to the more established methods of endolymphatic hydrops visualisation. If accessible, we agree that the latter techniques should be preferred, but we find that they are unfortunately too often unavailable in routine clinical practice to ENT physicians.

Saccular measurements in routine MRI can predict hydrops in Menière's disease.

Most patients with suspicion of hydrops do not have access to MRI with 3D reconstruction of the endolymphatic space. Our main objective was to show that measurements of the saccule on a non-enhanced 3D-T2 MRI could show hydrops and help diagnose Menière disease. We conducted a prospective study from 2015 to 2016 to compare consecutive patients consulting for Menière's disease to a control group (patients with unilateral non-hydrops disorders and contralateral healthy ears). They all received full auditory and vestibular testing. They also underwent a 3-Tesla 3D-T2 MRI using CISS sequence (0.4 mm thick slices), which were blindly evaluated by two independent neuroradiologists. The saccular height and width were measured in a coronal plane and Menière's disease patients' symptomatic ears were compared to asymptomatic and control ears. 36 patients with definite Menière's disease and 36 control patients were studied, including 42 symptomatic Menière, 30 asymptomatic Menière and 72 control ears. Saccular measurements were significantly different between symptomatic Menière ears compared to healthy ears (1.59 vs 1.32 mm, p < 0.001 for height; 1.13 vs 0.90 mm, p < 0.001 for width). Symptomatic and asymptomatic Menière ears' measurements were not significantly different (p = 0.307 and p = 0.109). Using ROC curve, we found cut-off values for saccular height 1.51 mm, Se = 63%, Sp = 95% and width 1.05 mm, Se = 41%, Sp = 95%. Routine 3D-T2 MRI, which patients must undergo for differential diagnosis, could help diagnose hydrops with high specificity using saccular measurements.

How I do it. The pedicled temporoparietal fascia flap for skull base reconstruction after endonasal endoscopic approaches.

BACKGROUND: Endoscopic endonasal approaches (EEA) are an alternative for removing challenging nasopharyngeal or skull base lesions. In some cases, a nasoseptal flap (NSF) is not always available and such complex procedures may lead to carotid arteries exposition and/or dura mater (DM) wide opening. Meticulous carotid coverage and DM reconstruction are crucial for preventing early and delayed complications. METHOD: We propose a step-by-step description of the pedicled temporoparietal fascia flap (TPFF) technique, with a focus on its pitfalls, advantages and limits. CONCLUSION: The TPFF is a reliable flap for skull base reconstruction when other pedicled flaps are not available. KEY POINTS: 1. Reliable flap even for irradiated patients 2. CT angiography with 3D reconstruction as pre-operative imaging 3. Doppler ultrasound probe to draw the artery's trajectory on skin 4. TPFF elevation concomitantly to the endoscopic procedure 5. Hemicoronal incision sufficient to harvest the TPFF 6. Superficial dissection in a plane just beneath the hair follicles 7. Dissection plane deep to the fad pad to preserve the frontal branch of the facial nerve 8. Surgical corridor wide enough to avoid any compression of the pedicle 9. Double visualization to avoid any twist and Doppler control of the STA patency 10. Close follow-up, toilet in clinics.

Value of systematic analysis of the olfactory cleft in case of cerebrospinal rhinorrhea: incidence of olfactory arachnoid dilatation.

To report on the presence of olfactory arachnoid dilatations (OAD), a previously undescribed radiologic feature of spontaneous cerebrospinal fluid (CSF) rhinorrhea originating from the cribriform plate of the ethmoid bone. The medical records of all patients treated between 2001 and 2011 at a tertiary care center for a spontaneous CSF rhinorrhea originating from the cribriform plate were retrospectively reviewed. The radiological work-up included high-resolution computed tomography and magnetic resonance imaging with at least the following sequences: T1, T2, and T2 with fast imaging employing steady state acquisition (FIESTA). Thirty cases were identified. The mean age at diagnosis was 49. Fourteen patients (47 %) had a body mass index (BMI) of 30 or more and 3 patients (10 %) had a BMI between 25 and 29.9. Five patients had a history of meningitis. The imaging work-up revealed a bone defect of the cribriform plate in 6 cases (20 %), associated to a typical meningocele in 14 cases (47 %). In ten patients (33 %), there was no defect of the cribriform plate, but ultrathin coronal T2-FIESTA sequences revealed an OAD, i.e. a dilatation of the arachnoid sheath of the olfactory fibers, in nine cases (30 %), or a "pseudo-polyp" outlined by a thin layer of arachnoid (1 patient, 3 %). Preoperative imaging should be carefully analyzed for the presence of OAD or "pseudo-polyp" in patients presenting with a CSF rhinorrhea without bony defect of the cribriform plate.

The binding force of the staphylococcal adhesin SdrG is remarkably strong.

SdrG is a cell surface adhesin from Staphylococcus epidermidis which binds to the blood plasma protein fibrinogen (Fg). Ligand binding follows a 'dock, lock and latch' model involving dynamic conformational changes of the adhesin that result in a greatly stabilized adhesin-ligand complex. To date, the force and dynamics of this multistep interaction are poorly understood. Here we use atomic force microscopy (AFM) to unravel the binding strength and cell surface localization of SdrG at molecular resolution. Single-cell force spectroscopy shows that SdrG mediates time-dependent attachment to Fg-coated surfaces. Single-molecule force spectroscopy with Fg-coated AFM tips demonstrates that the adhesin forms nanoscale domains on the cell surface, which we believe contribute to strengthen cell adhesion. Notably, we find that the rupture force of single SdrG-Fg bonds is very large, ∼ 2 nN, equivalent to the strength of a covalent bond, and shows a low dissociation rate, suggesting that the bond is very stable. The strong binding force, slow dissociation and clustering of SdrG provide a molecular foundation for the ability of S. epidermidis to colonize implanted biomaterials and to withstand physiological shear forces.

Integrated DNA walking system to characterize a broad spectrum of GMOs in food/feed matrices.

BACKGROUND: In order to provide a system fully integrated with qPCR screening, usually used in GMO routine analysis, as well as being able to detect, characterize and identify a broad spectrum of GMOs in food/feed matrices, two bidirectional DNA walking methods targeting p35S or tNOS, the most common transgenic elements found in GM crops, were developed. These newly developed DNA walking methods are completing the previously implemented DNA walking method targeting the t35S pCAMBIA element. METHODS: Food/feed matrices containing transgenic crops (Bt rice or MON863 maize) were analysed using the integrated DNA walking system. RESULTS: First, the newly developed DNA walking methods, anchored on the sequences used for the p35S or tNOS qPCR screening, were tested on Bt rice that contains these two transgenic elements. Second, the methods were assessed on a maize sample containing a low amount of the GM MON863 event, representing a more complex matrix in terms of genome size and sensitivity. Finally, to illustrate its applicability in GMO routine analysis by enforcement laboratories, the entire workflow of the integrated strategy, including qPCR screening to detect the potential presence of GMOs and the subsequent DNA walking methods to characterize and identify the detected GMOs, was applied on a GeMMA Scheme Proficiency Test matrix. Via the characterization of the transgene flanking region between the transgenic cassette and the plant genome as well as of a part of the transgenic cassette, the presence of GMOs was properly confirmed or infirmed in all tested samples. CONCLUSION: Due to their simple procedure and their short time-frame to get results, the developed DNA walking methods proposed here can be easily implemented in GMO routine analysis by the enforcement laboratories. In providing crucial information about the transgene flanking regions and/or the transgenic cassettes, this DNA walking strategy is a key molecular tool to prove the presence of GMOs in any given food/feed matrix.