A holistic approach for the investigation of bee health

Despite current evidence is in strong disagreement with an emergency for the conservation of Apis mellifera, great concern is related to profitability of beekeeping operations. A growing involvement of veterinary science in addressing bee health topics will therefore be fundamental to preserve and protect the entire sector. The experiments in this thesis focused on two different and interdependent levels related to bee health: the biochemical level and the parasitological level. At the biochemical level the impact of plant protection products on bee physiology and survival was studied, elucidating synergistic interactions between poor nutrition and pesticide exposure in A. mellifera and between an insecticide and a fungicide in Osmia bicornis. Moreover, an innovative fingerprinting approach on honey bee haemolymph was applied to detect population imbalances in the hive. The control of Varroa infestations was studied both at the biochemical and parasitological level. A panel of biomarkers in honey bee haemolymph was applied to compare different mite control protocols. This resulted in relevant indications for beekeeping operations pursuing the least impact on nutritional status of the colonies. To guide the decision making of beekeepers, a new formic acid evaporator was tested in comparison with a more established one. Considering its widespread distribution in the country, efforts were directed also towards N. ceranae. In particular, the pivotal aspect of diagnosis was studied, proposing a new qPCR method to overcome some limits of the existing ones. In conclusion, this works fills some of the knowledge gaps of the beekeeping sector. However, many of them still need to be addressed and the upcoming menaces of climate change and dispersal of pathogens via globalization should be targeted by research efforts in the near future. Therefore, a multifaceted vision of bee health is of capital importance, aware of the complementarity of reductionist and holistic approaches.

Applied genomics to monitor the genetic diversity and integrity of honey bee (Apis mellifera) populations and define sustainable strategies to benefit the beekeeping sector

Honey bees are considered keystone species in ecosystem, the effect of harmful pesticides for the honey bees, the action of extreme climatic waves and their consequence on honey bees health can cause the loss of many colonies which could contribute to the reduction of the effective population size and incentive the use of non-autochthonous queens to replace dead colonies. Over the last decades, the use of non-ligustica bee subspecies in Italy has increased and together with the mentioned phenomena exposed native honey bees to hybridization, laeding to a dramatic loss of genetic erosion and admixture. Healthy genetic diversity within honey bee populations is critical to provide tolerance and resistance to current and future threatening. Nowadays it is urgent to design strategies for the conservation of local subspecies and their valorisation on a productive scale. In this Thesis we applied genomics tool for the analysis of the genetic diversity and the genomic integrity of honey bee populations in Italy are described. In this work mtDNA based methods are presented using honey bee DNA or honey eDNA as source of information of the genetic diversity of A. mellifera at different level. Taken together, the results derived from these studies should enlarge the knowledge of the genetic diversity and integrity of the honey bee populations in Italy, filling the gap of information necessary to design efficient conservation programmes. Furthermore, the methods presented in these works will provide a tool for the honey authentication to sustain and valorise beekeeping products and sector against frauds.

New functional and biomechanical assessments for the improvement of the surgical navigation systems for joint replacement in the human lower limb

In case of severe osteoarthritis at the knee causing pain, deformity, and loss of stability and mobility, the clinicians consider that the substitution of these surfaces by means of joint prostheses. The objectives to be pursued by this surgery are: complete pain elimination, restoration of the normal physiological mobility and joint stability, correction of all deformities and, thus, of limping. The knee surgical navigation systems have bee developed in computer-aided surgery in order to improve the surgical final outcome in total knee arthroplasty. These systems provide the surgeon with quantitative and real-time information about each surgical action, like bone cut executions and prosthesis component alignment, by mean of tracking tools rigidly fixed onto the femur and the tibia. Nevertheless, there is still a margin of error due to the incorrect surgical procedures and to the still limited number of kinematic information provided by the current systems. Particularly, patello-femoral joint kinematics is not considered in knee surgical navigation. It is also unclear and, thus, a source of misunderstanding, what the most appropriate methodology is to study the patellar motion. In addition, also the knee ligamentous apparatus is superficially considered in navigated total knee arthroplasty, without taking into account how their physiological behavior is altered by this surgery. The aim of the present research work was to provide new functional and biomechanical assessments for the improvement of the surgical navigation systems for joint replacement in the human lower limb. This was mainly realized by means of the identification and development of new techniques that allow a thorough comprehension of the functioning of the knee joint, with particular attention to the patello-femoral joint and to the main knee soft tissues. A knee surgical navigation system with active markers was used in all research activities presented in this research work. Particularly, preliminary test were performed in order to assess the system accuracy and the robustness of a number of navigation procedures. Four studies were performed in-vivo on patients requiring total knee arthroplasty and randomly implanted by means of traditional and navigated procedures in order to check for the real efficacy of the latter with respect to the former. In order to cope with assessment of patello-femoral joint kinematics in the intact and replaced knees, twenty in-vitro tests were performed by using a prototypal tracking tool also for the patella. In addition to standard anatomical and articular recommendations, original proposals for defining the patellar anatomical-based reference frame and for studying the patello-femoral joint kinematics were reported and used in these tests. These definitions were applied to two further in-vitro tests in which, for the first time, also the implant of patellar component insert was fully navigated. In addition, an original technique to analyze the main knee soft tissues by means of anatomical-based fiber mappings was also reported and used in the same tests. The preliminary instrumental tests revealed a system accuracy within the millimeter and a good inter- and intra-observer repeatability in defining all anatomical reference frames. In in-vivo studies, the general alignments of femoral and tibial prosthesis components and of the lower limb mechanical axis, as measured on radiographs, was more satisfactory, i.e. within ±3°, in those patient in which total knee arthroplasty was performed by navigated procedures. As for in-vitro tests, consistent patello-femoral joint kinematic patterns were observed over specimens throughout the knee flexion arc. Generally, the physiological intact knee patellar motion was not restored after the implant. This restoration was successfully achieved in the two further tests where all component implants, included the patellar insert, were fully navigated, i.e. by means of intra-operative assessment of also patellar component positioning and general tibio-femoral and patello-femoral joint assessment. The tests for assessing the behavior of the main knee ligaments revealed the complexity of the latter and the different functional roles played by the several sub-bundles compounding each ligament. Also in this case, total knee arthroplasty altered the physiological behavior of these knee soft tissues. These results reveal in-vitro the relevance and the feasibility of the applications of new techniques for accurate knee soft tissues monitoring, patellar tracking assessment and navigated patellar resurfacing intra-operatively in the contest of the most modern operative techniques. This present research work gives a contribution to the much controversial knowledge on the normal and replaced of knee kinematics by testing the reported new methodologies. The consistence of these results provides fundamental information for the comprehension and improvements of knee orthopedic treatments. In the future, the reported new techniques can be safely applied in-vivo and also adopted in other joint replacements.

The Effect of Preservative Interventions on the Chemical- Physical and Structural Characteristics of Panel Painting

This work studies the impact of two traditional Romanian treatments, Red Petroleum and Propolis, in terms of real efficiency and consequence on the wooden artifacts. The application of these solutions is still a widely adopted and popular technique in preservative conservation but the impact of these solutions is not well known. It is important to know the effect of treatments on chemical-physical and structural characteristics of the artifacts, not only for understanding the influence on present conditions but also for foreseeing the future behavior. These treatments with Romanian traditional products are compared with a commercial antifungal product, Biotin R, which is utilized as reference to control the effectiveness of Red Petroleum and Propolis. Red Petroleum and Propolis are not active against mould while Biotin R is very active. Mould attack is mostly concentrated in the painted layer, where the tempera, containing glue and egg, enhance nutrition availability for moulds. Biotin R, even if is not a real insecticide but a fungicide, was the most active product against insect attack of the three products, followed by Red Petroleum, Propolis and untreated reference. As for colour, it did not change so much after the application of Red Petroleum and Biotin R and the colour difference was almost not perceptible. On the contrary, Propolis affected the colour a lot. During the exposure at different RH, the colour changes significantly at 100% RH at equilibrium and this is mainly due to the mould attack. Red Petroleum penetrates deeply into wood, while Propolis does not penetrate and remains only on the surface. However, Red Petroleum does not interact chemically with wood substance and it is easy volatilized in oven-dry condition. On the contrary Propolis interacts chemically with wood substance and hardly volatilized, even in oven-dry condition and consequently Propolis remains where it penetrated, mostly on the surface. Treatment by immersion has impact on wood physical parameters while treatment by brushing does not have significant impact. Especially Red Petroleum has an apparent impact on moisture content (MC) due to the penetration of solution, while Propolis does not penetrate so much and remains only on surface therefore Propolis does not have so much impact as Red Petroleum. However, if the weight of the solution penetrated in wood is eliminated, there is not significant difference in MC between treated and untreated samples. Considering physical parameters, dimensional stability is an important parameter. The variation of wood moisture content causes shrinkages/swelling of the wood that polychrome layer can only partially follow. The dimension of wooden supports varied under different moisture conditioning; the painted layer cannot completely follow this deformation, and consequently a degradation and deterioration caused by detachment, occurs. That detachment affects the polychrome stratification of the panel painting and eventually the connections between the different layer compositions of the panel painting.