Computational Tools for the Reliability Assessment and the Engineering Design of Procedures and Devices in Bariatric Surgery.

Obesity is one of the main health concerns worldwide. Bariatric Surgery (BS) is the gold standard treatment for severe obesity. Nevertheless, unsatisfactory weight loss and complications can occur. The efficacy of BS is mainly defined on experiential bases; therefore, a more rational approach is required. The here reported activities aim to show the strength of experimental and computational biomechanics in evaluating stomach functionality depending on bariatric procedure. The experimental activities consisted in insufflation tests on samples of swine stomach to assess the pressure-volume behaviour both in pre- and post-surgical configurations. The investigation pertained to two main bariatric procedures: adjustable gastric banding (AGB) and laparoscopic sleeve gastrectomy (LSG). Subsequently, a computational model of the stomach was exploited to validate and to integrate results from experimental activities, as well as to broad the investigation to a wider scenario of surgical procedures and techniques. Furthermore, the computational approach allowed analysing stress and strain fields within stomach tissues because of food ingestion. Such fields elicit mechanical stimulation of gastric receptors, contributing to release satiety signals. Pressure-volume curves assessed stomach capacity and stiffness according to the surgical procedure. Both AGB and LSG proved to reduce stomach capacity and to increase stiffness, with markedly greater effect for LSG. At an internal pressure of 5 kPa, outcomes showed that in pre-surgical configuration the inflated volume was about 1000 mL, after AGB the inflated volume was slightly lower, while after LSG it fell significantly, reaching 100 mL. Computational modelling techniques showed the influence of bariatric intervention on mechanical stimulation of gastric receptors due to food ingestion. AGB markedly enhanced the mechanical stimulation within the fundus region, while LSG significantly reduced stress and strain intensities. Further computational investigations revealed the potentialities of hybrid endoscopic procedures to induce both reduction of stomach capacity and enhancement of gastric receptors mechanical stimulation. In conclusion, biomechanics proved to be useful for the investigation of BS effects. Future exploitations of the biomechanical methods may largely improve BS reliability, efficacy and penetration rate.

First isolation of Yersinia entomophaga in human urinary tract.

Yersinia entomophaga is an insect pathogen first isolated from larvae of Coleoptera in New Zealand in 2011. We report here the first isolation of Y. entomophaga from human urine. Using whole-genome sequencing, we confirmed the presence of specific chromosomal virulence genes and identified a plasmid harbouring a quinolone resistance gene.

Detection, seroprevalence and antimicrobial resistance of Yersinia enterocolitica and Yersinia pseudotuberculosis in pig tonsils in Northern Italy.

Yersiniosis is the third most common reported zoonoses in Europe, with Y. enterocolitica and Y. pseudotuberculosis responsible for 98.66% and 0.94% of the confirmed human cases in 2013. From June 2013 to October 2014, 201 pigs at slaughter belonging to 67 batches were tested for Y. enterocolitica and Y. pseudotuberculosis in tonsils. Diaphragm muscle samples were tested for antibodies against Yersinia by a commercially available ELISA test. Y. enterocolitica 4/O:3 was detected in 55/201 pig tonsils (27.4%; 95% CI 23.1-37.1). The positive pigs came from 38/67 batches (56.7%) and were reared in 36/61 farms (59.0%). There was no statistical difference between farrow-to-finish and finishing farms. The mean count of Y. enterocolitica was 3.56±0.85log10CFU/g with a minimum of 2.0log10CFU/g and a maximum of 4.78log10CFU/g. Y. pseudotuberculosis was isolated from 4/201 pig tonsils (2.0%; 95% CI 0.0-4.5). Three isolates belonged to serotype O:3 and one to serotype O:1. The positive pigs belonged to 4/67 batches (6.0%) and came from finishing farms only. Y. pseudotuberculosis could be enumerated in one sample only (4.27log10CFU/g). The ELISA test demonstrated that 56.1% of the meat juice samples were positive for Yersinia antibodies. Serological positivity was found in 67.9% (36/53) of the Y. enterocolitica- and 75.0% (3/4) of the Y. pseudotuberculosis positive pigs. A significant association was found between serological results and the presence of Y. enterocolitica in tonsils (OR=1.97, p=0.044). All the Y. enterocolitica 4/O:3 isolates were susceptible to amoxicillin-clavulanic acid, gentamicin, ceftazidime, ertapenem and meropenem, 94.5% to cefotaxime, 89.1% to kanamycin and 78.2% to tetracycline. The highest resistance rates were observed for ampicillin (100%), sulphonamides (98.2%) and streptomycin (78.2%). Y. pseudotuberculosis strains were sensitive to all the antimicrobials tested, i.e. amoxicillin, amoxicillin/clavulanic acid, azithromycin, cephalothin, cefoxitin, ceftriaxone, ciprofloxacin, nalidixic acid, sulphonamide, tetracycline and ticarcillin. The study shows that Italian fattening pigs are frequently infected with human pathogenic Y. enterocolitica 4/O:3. Although the isolation rate is slightly lower than in other European countries, the serological test demonstrates that the infection is widespread among pig population. In fact, seroprevalence is similar to other EU countries. The detection of Y. pseudotuberculosis serotypes O:1 and O:3 in pig tonsils is of concern. Since tonsils may represent a contamination source for pig meat at slaughter, further studies regarding human infections by both microbial species are strongly recommended.

Bladder tissue biomechanical behavior: Experimental tests and constitutive formulation.

A procedure for the constitutive analysis of bladder tissues mechanical behavior is provided, by using a coupled experimental and computational approach. The first step pertains to the design and development of mechanical tests on specimens from porcine bladders. The bladders have been harvested, and the specimens have been subjected to uniaxial cyclic tests at different strain rates along preferential directions, considering the distribution of tissue fibrous components. Experimental results showed the anisotropic, non-linear and time-dependent stress-strain behavior, due to tissue conformation with fibers distributed along preferential directions and their interaction phenomena with ground substance. In detail, experimental data showed a greater tissue stiffness along transversal direction. Viscous behavior was assessed by strain rate dependence of stress-strain curves and hysteretic phenomena. The second step pertains the development of a specific fiber-reinforced visco-hyperelastic constitutive model, in the light of bladder tissues structural conformation and experimental results. Constitutive parameters have been identified by minimizing the discrepancy between model and experimental data. The agreement between experimental and model results represent a term for evaluating the reliability of the constitutive models by means of the proposed operational procedure.

Typing and clustering of Yersinia pseudotuberculosis isolates by restriction fragment length polymorphism analysis using insertion sequences.

Yersinia pseudotuberculosis is an enteropathogen that has an animal reservoir and causes human infections, mostly in temperate and cold countries. Most of the methods previously used to subdivide Y. pseudotuberculosis were performed on small numbers of isolates from a specific geographical area. One aim of this study was to evaluate the typing efficiency of restriction fragment length polymorphism of insertion sequence hybridization patterns (IS-RFLP) compared to other typing methods, such as serotyping, ribotyping, and multilocus sequence typing (MLST), on the same set of 80 strains of Y. pseudotuberculosis of global origin. We found that IS100 was not adequate for IS-RFLP but that both IS285 and IS1541 efficiently subtyped Y. pseudotuberculosis. The discriminatory index (DI) of IS1541-RFLP (0.980) was superior to those of IS285-RFLP (0.939), ribotyping (0.944), MLST (0.861), and serotyping (0.857). The combination of the two IS (2IS-RFLP) further increased the DI to 0.998. Thus, IS-RFLP is a powerful tool for the molecular typing of Y. pseudotuberculosis and has the advantage of exhibiting well-resolved banding patterns that allow for a reliable comparison of strains of worldwide origin. The other aim of this study was to assess the clustering power of IS-RFLP. We found that 2IS-RFLP had a remarkable capacity to group strains with similar genotypic and phenotypic markers, thus identifying robust populations within Y. pseudotuberculosis. Our study thus demonstrates that 2IS- and even IS1541-RFLP alone might be valuable tools for the molecular typing of global isolates of Y. pseudotuberculosis and for the analysis of the population structure of this species.

Characterization of the anisotropic mechanical behaviour of colonic tissues: experimental activity and constitutive formulation.

The aim was to investigate the biomechanical behaviour of colonic tissues by a coupled experimental and numerical approach. The wall of the colon is composed of different tissue layers. Within each layer, different fibre families are distributed according to specific spatial orientations, which lead to a strongly anisotropic configuration. Accounting for the complex histology of the tissues, mechanical tests must be planned and designed to evaluate the behaviour of the colonic wall in different directions. Uni-axial tensile tests were performed on tissue specimens from 15 fresh pig colons, accounting for six different loading directions (five specimens for each loading direction). The next step of the investigation was to define an appropriate constitutive framework and develop a procedure for identification of the constitutive parameters. A specific hyperelastic formulation was developed that accounted for the multilayered conformation of the colonic wall and the fibre-reinforced configuration of the tissues. The parameters were identified by inverse analyses of the mechanical tests. The comparison of model results with experimental data, together with the evaluation of satisfaction of material thermomechanics principles, confirmed the reliability of the analysis developed. This work forms the basis for more comprehensive activities that aim to provide computational tools for the interpretation of surgical procedures that involve the gastrointestinal tract, considering the specific biomedical devices adopted.

Investigation of the mechanical behaviour of the foot skin.

BACKGROUND/PURPOSE: The aim of this work was to provide computational tools for the characterization of the actual mechanical behaviour of foot skin, accounting for results from experimental testing and histological investigation. Such results show the typical features of skin mechanics, such as anisotropic configuration, almost incompressible behaviour, material and geometrical non linearity. The anisotropic behaviour is mainly determined by the distribution of collagen fibres along specific directions, usually identified as cleavage lines. METHODS: To evaluate the biomechanical response of foot skin, a refined numerical model of the foot is developed. The overall mechanical behaviour of the skin is interpreted by a fibre-reinforced hyperelastic constitutive model and the orientation of the cleavage lines is implemented by a specific procedure. Numerical analyses that interpret typical loading conditions of the foot are performed. The influence of fibres orientation and distribution on skin mechanics is outlined also by a comparison with results using an isotropic scheme. RESULTS: A specific constitutive formulation is provided to characterize the mechanical behaviour of foot skin. The formulation is applied within a numerical model of the foot to investigate the skin functionality during typical foot movements. Numerical analyses developed accounting for the actual anisotropic configuration of the skin show lower maximum principal stress fields than results from isotropic analyses. CONCLUSION: The developed computational models provide reliable tools for the investigation of foot tissues functionality. Furthermore, the comparison between numerical results from anisotropic and isotropic models shows the optimal configuration of foot skin.

Biomechanical behaviour of ankle ligaments: constitutive formulation and numerical modelling.

This study was aimed at the definition of a constitutive formulation of ankle ligaments and of a procedure for the constitutive parameters evaluation, for the biomechanical analysis by means of numerical models. To interpret the typical features of ligaments mechanical response, as anisotropic configuration, geometric non-linearity, non-linear elasticity and time-dependent behaviour, a specific fibre-reinforced visco-hyperelastic model is provided. The identification of constitutive parameters is performed by a stochastic-deterministic procedure that minimises the discrepancy between experimental and computational results. A preliminary evaluation of parameters is performed by analytical models in order to define reference values. Afterwards, solid models are developed to consider the complex histo-morphometric configuration of samples as a basis for the definition of numerical models. The results obtained are adopted for upgrading parameter values by comparison with specific mechanical tests. Assuming the new parameters set, the final numerical results are compared with the overall set of experimental data, to assess the reliability and efficacy of the analysis developed for the interpretation of the mechanical response of ankle ligaments.

Analysis of the biomechanical behaviour of gastrointestinal regions adopting an experimental and computational approach.

An integrated experimental and computational procedure is provided for the evaluation of the biomechanical behaviour that characterizes the pressure-volume response of gastrointestinal regions. The experimental activity pertains to inflation tests performed on specific gastrointestinal conduct segments. Different inflation processes are performed according to progressively increasing volumes. Each inflation test is performed by a rapid liquid in-flaw, up to a prescribed volume, which is held constant for about 300s to allow the development of relaxation processes. The different tests are interspersed by 600s of rest to allow the recovery of the specimen mechanical condition. A physio-mechanical model is developed to interpret both the elastic behaviour of the sample, as the pressure-volume trend during the rapid liquid in-flaw, and the time-dependent response, as the pressure drop during the relaxation processes. The minimization of discrepancy between experimental data and model results entails the identification of the parameters that characterize the viscoelastic model adopted for the definition of the behaviour of the gastrointestinal regions. The reliability of the procedure is assessed by the characterization of the response of samples from rat small intestine.

Analysis of heel pad tissues mechanics at the heel strike in bare and shod conditions.

A combined experimental and numerical approach is used to investigate the interaction phenomena occurring between foot and footwear during the heel strike phase of the gait. Two force platforms are utilised to evaluate the ground reaction forces of a subject in bare and shod walking. The reaction forces obtained from the experimental tests are assumed as loading conditions for the numerical analyses using three dimensional models of the heel region and of the running shoe. The heel pad region, as fat and skin tissues, is described by visco-hyperelastic and fibre-reinforced hyperelastic formulations respectively and bone region by a linear orthotropic formulation. Different elastomeric foams are considered with regard to the outsole, the midsole and the insole layers. The mechanical properties are described by a hyperfoam formulation. The evaluation of the mechanical behaviour of the heel pad tissues at the heel strike in bare and shod conditions is performed considering different combinations of materials for midsole and insole layers. Results allow for the definition of the influence of different material characteristics on the mechanical response of the heel pad region, in particular showing the compressive stress differentiation in the bare and shod conditions.

Resistance to plague of Mus spretus SEG/Pas mice requires the combined action of at least four genetic factors.

We have previously described SEG/Pas as the first mouse inbred strain able to survive subcutaneous injection of virulent Yersinia pestis, the agent of plague, and we identified Yprl1, Yprl2 and Yprl3 as three quantitative trait loci (QTLs) controlling this exceptional phenotype in females from a backcross between SEG/Pas and C57BL/6 strains. We have now developed congenic strains to further characterize the extent and effect of these genomic regions. In this study, we confirm the importance of two of these regions, both in males and females, while the third one may well be a spurious association. We show that no genomic region alone is able to increase the survival of C57BL/6 mice, but that C57BL/6 mice carrying both Yprl2 and Yprl3 exhibit intermediate resistance. Each of these two QTLs contains at least two subregions, which are required to increase survival. Finally, through the analysis of congenic strains in an F1 background, we establish the mode of inheritance of the SEG-derived resistance alleles. Altogether, this study has clarified and enhanced our understanding of the genetic architecture of resistance to plague in SEG/Pas mice.

Investigation on the load-displacement curves of a human healthy heel pad: In vivo compression data compared to numerical results.

The aims of the present work were to build a 3D subject-specific heel pad model based on the anatomy revealed by MR imaging of a subject's heel pad, and to compare the load-displacement responses obtained from this model with those obtained from a compression device used on the subject's heel pad. A 30 year-old European healthy female (mass=54kg, height=165cm) was enrolled in this study. Her left foot underwent both MRI and compression tests. A numerical model of the heel region was developed based on a 3D CAD solid model obtained by MR images. The calcaneal fat pad tissue was described with a visco-hyperelastic model, while a fiber-reinforced hyperelastic model was formulated for the skin. Numerical analyses were performed to interpret the mechanical response of heel tissues. Different loading conditions were assumed according to experimental tests. The heel tissues showed a non-linear visco-elastic behavior and the load-displacement curves followed a characteristic hysteresis form. The energy dissipation ratios measured by experimental tests (0.25±0.02 at low strain rate and 0.26±0.03 at high strain rate) were comparable with those evaluated by finite element analyses (0.23±0.01 at low strain rate and 0.25±0.01 at high strain rate). The validity and efficacy of the investigation performed was confirmed by the interpretation of the mechanical response of the heel tissues under different strain rates. The mean absolute percentage error between experimental data and model results was 0.39% at low strain rate and 0.28% at high strain rate.

Constitutive formulation and numerical analysis of the heel pad region.

The aim of this work is to provide a numerical approach for the investigation of the mechanical behaviour of the heel pad region. A visco-hyperelastic model is formulated with regard to fat pad tissue, while a fibre-reinforced hyperelastic model is considered for the heel skin tissue. Bone components are defined by means of an orthotropic linear elastic model. Particular attention is paid to the evaluation of constitutive parameters within different models adopted in consideration of experimental tests data. Preliminarily, indentation tests on a skinless cadaveric foot are considered with regard to fat pad tissue. Indentation tests on an intact heel pad of a cadaveric foot are subsequently adopted for the final identification of constitutive parameters of fat pad and skin tissues. A numerical model of the rear foot is defined and different loading conditions are assumed according to experimental data. A comparison between experimental and numerical data leads to the evaluation of the real capability of the procedure to interpret the actual response of the rear foot.

A numerical model for investigating the mechanics of calcaneal fat pad region.

The present paper pertains to the definition of a numerical model of the calcaneal fat pad region, considering a structure composed of adipose and connective tissues organized in fibrous septae and adipose chambers. The mechanical response is strongly influenced by the structural conformation, as the dimension of adipose chambers, the thickness of connective septae walls and the mechanical properties of the different soft tissues. In order to define the constitutive formulation of adipose tissues, experimental data from pig specimens are considered, according to the functional similarity, while the mechanical response of connective tissue septae is assumed with regard to the mechanical behaviour that characterize ligaments. Different numerical models are provided accounting for the variation of chambers dimensions, septae wall thickness and tissues characteristics. The spiral angles of collagen fibres within the septae influence the capability of the structure to withstand the bulging of chambers. The analysis considers different orientation of the fibres. The response of calcaneal fat pad region is evaluated in comparison with experimental data from unconfined compression tests. The present work provides a preliminary approach to enhance the correlation between the structural conformation and tissues mechanical properties towards the biomechanical response of overall heel pad region.

Biomechanical behaviour of heel pad tissue: experimental testing, constitutive formulation, and numerical modelling.

This paper deals with the constitutive formulation of heel pad tissue and presents a procedure for identifying constitutive parameters using experimental data, with the aim of developing a computational approach for investigating the actual biomechanical response. The preliminary definition of constitutive parameters was developed using a visco-hyperelastic formulation, considering experimental data from in vitro compression tests on specimens of fat pad tissue and data from in vivo tests to identify the actual trend of tissue stiffness. The discrepancy between model results and experimental data was evaluated on the basis of a specific cost function, adopting a stochastic/deterministic procedure. The parameter evaluation was upgraded by considering experimental tests performed on the fat pad tissues of a cadaveric foot using in situ indentation tests at 0.01 and 350 mm/s strain rates. The constitutive formulation was implemented in a numerical model. The comparison of data from in situ tests and numerical results led to an optimal domain of parameters based on an admissible discrepancy criterion. Numerical results evaluated for different sets of parameters inside the domain are reported and compared with experimental data for a reliability evaluation of the proposed procedure.

Mus spretus SEG/Pas mice resist virulent Yersinia pestis, under multigenic control.

Laboratory mice are well known to be highly susceptible to virulent strains of Yersinia pestis in experimental models of bubonic plague. We have found that Mus spretus-derived SEG/Pas (SEG) mice are exceptionally resistant to virulent CO92 and 6/69 wild type strains. Upon subcutaneous injection of 10(2) colony-forming units (CFU), 90% of females and 68% of males survived, compared with only an 8% survival rate for both male and female C57BL/6 mice. Furthermore, half of the SEG mice survived a challenge of up to 10(7) CFU. The time required for mortality was similar between B6 and SEG, suggesting that survival is dependent on early rather than late processes. The analysis of 322 backcross mice identified three significant quantitative trait loci (QTLs) on chromosomes 3, 4 and 6, with dominant SEG protective alleles. Each QTL increased the survival rate by approximately 20%. The three QTLs function additively, thereby accounting for 67% of the difference between the parental phenotypes. Mice heterozygous for the three QTLs were just as resistant as SEG mice to Y. pestis challenge. The SEG strain therefore offers an invaluable opportunity to unravel mechanisms and underlying genetic factors of resistance against Y. pestis infection.

Investigation of foot plantar pressure: experimental and numerical analysis.

The analysis of interaction phenomena occurring between the plantar region of the foot and insole was investigated using a combined experimental-numerical approach. Experimental data on the plantar pressure for treadmill walking of a subject were obtained using the Pedar(®) system. The plantar pressure resultant was monitored during walking and adopted to define the loading conditions for a subsequent static numerical analysis. Geometrical configuration of the foot model is provided on the basis of biomedical images. Because the mechanical behaviour of adipose tissues and plantar fascia is the determinant factor in affecting the paths of the plantar pressure, specific attention was paid to define an appropriate constitutive model for these tissues. The numerical model included sole and insole, providing for friction contact conditions between foot-insole and insole-sole pairs as well. Two different numerical analyses were performed with regards to different loading conditions during the gait cycle. The plantar pressure peaks predicted by the numerical model for the two loading conditions are 0.16 and 0.12 MPa, and 0.09 and 0.12 MPa in the posterior and anterior regions of the foot, respectively. These values are in agreement with experimental evidence, showing the suitability of the model proposed.

Constitutive formulation and analysis of heel pad tissues mechanics.

This paper presents a visco-hyperelastic constitutive model developed to describe the biomechanical response of heel pad tissues. The model takes into account the typical features of the mechanical response such as large displacement, strain phenomena, and non-linear elasticity together with time-dependent effects. The constitutive model was formulated, starting from the analysis of the complex structural and micro-structural configuration of the tissues, to evaluate the relationship between tissue histology and mechanical properties. To define the constitutive model, experimental data from mechanical tests were analyzed. To obtain information about the mechanical response of the tissue so that the constitutive parameters could be established, data from both in vitro and in vivo tests were investigated. Specifically, the first evaluation of the constitutive parameters was performed by a coupled deterministic and stochastic optimization method, accounting for data from in vitro tests. The comparison of constitutive model results and experimental data confirmed the model's capability to describe the compression behaviour of the heel pad tissues, regarding both constant strain rate and stress relaxation tests. Based on the data from additional experimental tests, some of the constitutive parameters were modified in order to interpret the in vivo mechanical response of the heel pad tissues. This approach made it possible to interpret the actual mechanical function of the tissues.

Immunogenicity and safety of combined intradermal recombinant Hepatitis B with BCG vaccines at birth.

This randomized, prospective, non-inferiority study aimed to quantify anti-HBs titers induced by recombinant Hepatitis B vaccine from healthy infants vaccinated with combined Hepatitis B and Bacillus Calmette-Guérin (BCG) vaccines (HbsAg 10 microg plus BCG suspension 0.1mg) and compare them to titers obtained with separated vaccines. Infants were immunized at birth either with combined intradermal (ID) BCG and Hepatitis B or ID BCG alone and intramuscular (IM) Hepatitis B. Both groups received IM Hepatitis B at 1 and 6 months of age. After the third dose anti-HBs titers > or =10 IU/mL were observed in 99% of vaccinees and > or =1000 IU/mL in 71%. There were no adverse events in both groups. Combination of HbsAg with BCG as first dose did not modify the profile of the humoral immune response for Hepatitis B indicating safety and immunogenicity of this vaccine in newborn.

3 IS-RFLP: a powerful tool for geographical clustering of global isolates of Yersinia pestis.

Multiple copies of several classes of insertion sequences (IS) are found in the genome of Yersinia pestis, the causative agent of bubonic and pneumonic plague. We used the genetic instability generated by these IS to develop a method (designated 3IS-RFLP) based on the restriction fragment length polymorphism of the IS100, IS285 and IS1541 elements for studying Y. pestis strains of worldwide origin. We show that 31S-RFLP is a powerful tool to group Y. pestis isolates according to their geographical origin, and therefore that this method may be valuable for investigating the origin of new or re-emerging plague foci or for addressing forensic issues.