Parasite and virus dynamics in the honeybee Apis mellifera unicolor on a tropical island recently invaded by Varroa destructor.

In La Réunion, the established honeybee subspecies Apis mellifera unicolor, an endemic subspecies of African lineage, is facing considerable challenges. Since the introduction of the Varroa destructor mite in 2017 high colony losses have been recorded. We investigated the dynamics of V. destructor and two viruses, the Deformed Wing Virus (DWV), known to be transmitted by the mite, and the Chronic Bee Paralysis Virus (CBPV), in A. m. unicolor. Colonies from two apiaries located at 300 and 900 m a.s.l were monitored twice for one year without any acaricide treatment. The brood area, V. destructor infestation rates, DWV and CBPV prevalence and load were recorded monthly. A. m. unicolor maintained brood rearing throughout the year. Varroa destructor infestation resulted in high colony mortality (up to 85 %) and high phoretic mite rates (up to 52 mites per hundred bees). The establishment of DWV in colonies occurred after that of V. destructor and the mite infestation rate had a significant effect on the virus prevalence and load. CBPV appeared only transiently throughout the surveys. The data showed that, in tropical colonies with permanent brood rearing, V. destructor and DWV can reach high levels, but are still subject to seasonal variations that appear to be influenced by environmental conditions. This suggests that beekeeping practices could be adapted by favouring sites and periods for transhumance or acaricide treatment.

Geographic population structure of the honeybee microsporidian parasite Vairimorpha (Nosema) ceranae in the South West Indian Ocean.

The microsporidian Vairimorpha (Nosema) ceranae is one of the most common parasites of the honeybee. A single honeybee carries many parasites and therefore multiple alleles of V. ceranae genes that seem to be ubiquitous. As a consequence, nucleotide diversity analyses have not allowed discriminating genetic structure of parasite populations. We performed deep loci-targeted sequencing to monitor the haplotype frequencies of genome markers in isolates from discontinuous territories, namely the tropical islands of the South West Indian Ocean. The haplotype frequency distribution corroborated the suspected tetraploidy of the parasite. Most major haplotypes were ubiquitous in the area but with variable frequency. While oceanic isolates differed from European and Asian outgroups, parasite populations from distinct archipelagoes also differed in their haplotype distribution. Interestingly an original and very divergent Malagasy isolate was detected. The observed population structure allowed formulating hypotheses upon the natural history of V. ceranae in this oceanic area. We also discussed the usefulness of allelic distribution assessment, using multiple informative loci or genome-wide analyses, when parasite population is not clonal within a single host.

Evolutionarily diverse origins of deformed wing viruses in western honey bees.

Novel transmission routes can allow infectious diseases to spread, often with devastating consequences. Ectoparasitic varroa mites vector a diversity of RNA viruses, having switched hosts from the eastern to western honey bees (Apis cerana to Apis mellifera). They provide an opportunity to explore how novel transmission routes shape disease epidemiology. As the principal driver of the spread of deformed wing viruses (mainly DWV-A and DWV-B), varroa infestation has also driven global honey bee health declines. The more virulent DWV-B strain has been replacing the original DWV-A strain in many regions over the past two decades. Yet, how these viruses originated and spread remains poorly understood. Here, we use a phylogeographic analysis based on whole-genome data to reconstruct the origins and demography of DWV spread. We found that, rather than reemerging in western honey bees after varroa switched hosts, as suggested by previous work, DWV-A most likely originated in East Asia and spread in the mid-20th century. It also showed a massive population size expansion following the varroa host switch. By contrast, DWV-B was most likely acquired more recently from a source outside East Asia and appears absent from the original varroa host. These results highlight the dynamic nature of viral adaptation, whereby a vector's host switch can give rise to competing and increasingly virulent disease pandemics. The evolutionary novelty and rapid global spread of these host-virus interactions, together with observed spillover into other species, illustrate how increasing globalization poses urgent threats to biodiversity and food security.

Spatio-temporal modelling of Culicoides Latreille (Diptera: Ceratopogonidae) populations on Reunion Island (Indian Ocean).

BACKGROUND: Reunion Island regularly faces outbreaks of bluetongue and epizootic hemorrhagic diseases, two insect-borne orbiviral diseases of ruminants. Hematophagous midges of the genus Culicoides (Diptera: Ceratopogonidae) are the vectors of bluetongue (BTV) and epizootic hemorrhagic disease (EHDV) viruses. In a previous study, statistical models based on environmental and meteorological data were developed for the five Culicoides species present in the island to provide a better understanding of their ecology and predict their presence and abundance. The purpose of this study was to couple these statistical models with a Geographic Information System (GIS) to produce dynamic maps of the distribution of Culicoides throughout the island. METHODS: Based on meteorological data from ground weather stations and satellite-derived environmental data, the abundance of each of the five Culicoides species was estimated for the 2214 husbandry locations on the island for the period ranging from February 2016 to June 2018. A large-scale Culicoides sampling campaign including 100 farms was carried out in March 2018 to validate the model. RESULTS: According to the model predictions, no husbandry location was free of Culicoides throughout the study period. The five Culicoides species were present on average in 57.0% of the husbandry locations for C. bolitinos Meiswinkel, 40.7% for C. enderleini Cornet & Brunhes, 26.5% for C. grahamii Austen, 87.1% for C. imicola Kieffer and 91.8% for C. kibatiensis Goetghebuer. The models also showed high seasonal variations in their distribution. During the validation process, predictions were acceptable for C. bolitinos, C. enderleini and C. kibatiensis, with normalized root mean square errors (NRMSE) of 15.4%, 13.6% and 16.5%, respectively. The NRMSE was 27.4% for C. grahamii. For C. imicola, the NRMSE was acceptable (11.9%) considering all husbandry locations except in two specific areas, the Cirque de Salazie-an inner mountainous part of the island-and the sea edge, where the model overestimated its abundance. CONCLUSIONS: Our model provides, for the first time to our knowledge, an operational tool to better understand and predict the distribution of Culicoides in Reunion Island. As it predicts a wide spatial distribution of the five Culicoides species throughout the year and taking into consideration their vector competence, our results suggest that BTV and EHDV can circulate continuously on the island. As further actions, our model could be coupled with an epidemiological model of BTV and EHDV transmission to improve risk assessment of Culicoides-borne diseases on the island.

[Automatic superimpositions of 3-dimensional craniofacial imaging: illustration with a mandibular distraction osteogenesis case]. / Superpositions automatisées d'imageries tridimensionnelles cranio-faciales : illustration par un cas clinique de distraction mandibulaire chirurgicale.

When performed several times over the course of an orthodontic or surgical orthodontic treatment, three-dimensional (3D) imaging like CBCT or CT-Scans can be superimposed. The purpose of this article is to illustrate the implementation of voxel-based 3D superimpositions with a clinical case of mandibular distraction osteogenesis A 13-year old patient underwent a surgical orthodontic treatment with a « 3D ¼ mandibular distractor. He presented a bilateral atrophy of his mandibular condyles due to a staphylococcus aureus bacteremia at birth. 3D general and mandibular regional superimpositions were performed using CT-Scan and CBCT images acquired respectively before and after completion of the distraction osteogenesis. Two superimposition methods were used, one relying on commercially-available software and the other one relying on a series of « open-source ¼ softwares. Using commercially-available software, 3D superimpositions were automatically performed in a few minutes by a sparsely-trained operator. The method relying on « open-source ¼ software asked for more training and time. Results of the superimpositions were presented under various formats. Evaluation of 3D superimposition results is still challenging, as only a qualitative evaluation can be easily performed. In years to come, this interpretation should however become more straightforward for clinicians.

Modelling temporal dynamics of Culicoides Latreille (Diptera: Ceratopogonidae) populations on Reunion Island (Indian Ocean), vectors of viruses of veterinary importance.

BACKGROUND: Reunion Island regularly faces outbreaks of epizootic haemorrhagic disease (EHD) and bluetongue (BT), two viral diseases transmitted by haematophagous midges of the genus Culicoides (Diptera: Ceratopogonidae) to ruminants. To date, five species of Culicoides are recorded in Reunion Island in which the first two are proven vector species: Culicoides bolitinos, C. imicola, C. enderleini, C. grahamii and C. kibatiensis. Meteorological and environmental factors can severely constrain Culicoides populations and activities and thereby affect dispersion and intensity of transmission of Culicoides-borne viruses. The aim of this study was to describe and predict the temporal dynamics of all Culicoides species present in Reunion Island. METHODS: Between 2016 and 2018, 55 biweekly Culicoides catches using Onderstepoort Veterinary Institute traps were set up in 11 sites. A hurdle model (i.e. a presence/absence model combined with an abundance model) was developed for each species in order to determine meteorological and environmental drivers of presence and abundance of Culicoides. RESULTS: Abundance displayed very strong heterogeneity between sites. Average Culicoides catch per site per night ranged from 4 to 45,875 individuals. Culicoides imicola was dominant at low altitude and C. kibatiensis at high altitude. A marked seasonality was observed for the three other species with annual variations. Twelve groups of variables were tested. It was found that presence and/or abundance of all five Culicoides species were driven by common parameters: rain, temperature, vegetation index, forested environment and host density. Other parameters such as wind speed and farm building opening size governed abundance level of some species. In addition, Culicoides populations were also affected by meteorological parameters and/or vegetation index with different lags of time, suggesting an impact on immature stages. Taking into account all the parameters for the final hurdle model, the error rate by Normalized Root mean Square Error ranged from 4.4 to 8.5%. CONCLUSIONS: To our knowledge, this is the first study to model Culicoides population dynamics in Reunion Island. In the absence of vaccination and vector control strategies, determining periods of high abundance of Culicoides is a crucial first step towards identifying periods at high risk of transmission for the two economically important viruses they transmit.

[Front Block distraction]. / Distraction sagittale du bloc incisivo-canin mandibulaire.

The contribution of the segmental osteotomies in the ortho-surgical protocols is no longer to demonstrate and found a new lease of life thanks to the combination with the bone distraction techniques. The osteotomy of Köle, initially described to close infraclusies, and then used to level very marked curves of Spee has more recently been used to correct anterior crowding. This support is therefore aimed at patients with an incisor and canine Class 2 but molar Class 1 with an isolated mandibular footprint. With minimal orthodontic preparation we can create in two weeks bilateral diastemas that will then be used to align the incisivocanin crowding without stripping or bicuspid extractions. Dental orthodontic movements can be resumed one month after the end of the distraction. This technique is therefore likely to avoid bicuspid extraction and replace some sagittal osteotomy advancement by correction of the overjet. It also helps to correct a incisors labial or lingual tipping playing on differential activation of the cylinders and the distractor. This segmental surgery can be combined with Le Fort 1 surgeries with correction of the transverse and associated meanings, but in a second time, to a mandibular advancement and/or a genioplasty.

Emergence of Coxiella burnetii in ruminants on Reunion Island? Prevalence and risk factors.

Q fever is a widespread zoonosis that is caused by Coxiella burnetii (C. burnetii), and ruminants are identified as the main sources of human infections. Some human cases have been described, but very limited information was available about Q fever in ruminants on Reunion Island, a tropical island in the Indian Ocean. A cross-sectional study was undertaken from March 2011 to August 2012 to assess the Q fever prevalence and to identify the major risk factors of C. burnetii infection in ruminants. A total of 516 ruminants (245 cattle, 137 sheep and 134 goats) belonging to 71 farms and localized in different ecosystems of the island were randomly selected. Samples of blood, vaginal mucus and milk were concomitantly collected from females, and a questionnaire was submitted to the farmers. Ticks from positively detected farms were also collected. The overall seropositivity was 11.8% in cattle, 1.4% in sheep and 13.4% in goats. C. burnetii DNA was detected by PCR in 0.81%, 4.4% and 20.1% in cow, sheep and goat vaginal swabs, respectively. C. burnetii shedding in milk was observed in 1% of cows, 0% in sheep and 4.7% in goats. None of the ticks were detected to be positive for C. burnetii. C. burnetii infection increased when the farm was exposed to prevailing winds and when there were no specific precautions for a visitor before entering the farm, and they decreased when a proper quarantine was set up for any introduction of a new ruminant and when the animals returned to the farm at night. MLVA genotyping confirmed the role of these risk factors in infection.

Epizootic haemorrhagic disease virus in Reunion Island: evidence for the circulation of a new serotype and associated risk factors.

Bluetongue virus (BTV) and epizootic haemorrhagic disease virus (EHDV) are members of the Orbivirus genus of the Reoviridae family transmitted between ruminants by the bites of Culicoides midges. BTV went undetected in Reunion Island between its first documented emergence in 1979 and two other serious outbreaks with both BTV-3 and EHDV-6 in 2003, and both EHDV-6 and BTV-2 in 2009. In these outbreaks, infected animals developed symptoms including hyperthermia, anorexia, congestion, prostration and nasal discharge. Samples were collected in 2011 to assess the prevalence of BT and EHD in ruminants native to Reunion Island by serological analysis. A cross-sectional study was undertaken on 67 farms, including a total of 276 cattle, 142 sheep and 71 goats. The prevalence rates of BT and EHD were 58% (95% CI [54.03-62.94]) and 38% (95% CI [33.85-42.63], respectively. Two further suspected outbreaks were confirmed to involve EHDV and BTV/EHDV. A new circulating EHDV serotype 1 of unknown origin was isolated. Our results confirm that the prevalence of both BT and EHD is high and that both are likely currently circulating. A high risk of BTV and EHDV infections was associated with the introduction of ruminants from neighbouring farms without quarantine, the presence of organic and other waste on the farm, and treatment against ectoparasites and insects.

High-intensity focused ultrasound ablation of thyroid nodules: first human feasibility study.

BACKGROUND: Thyroid surgery is common, but complications may occur. High-intensity focused ultrasound (HIFU) is a minimally invasive alternative to surgery. We hypothesized that an optimized HIFU device could be safe and effective for ablating benign thyroid nodules without affecting neighboring structures. METHODS: In this open, single-center feasibility study, 25 patients were treated with HIFU with real-time ultrasound imaging 2 weeks before a scheduled thyroidectomy for multinodular goiter. Thyroid ultrasonography imaging, thyroid function, were evaluated before and after treatment. Adverse events were carefully recorded. Each patient received HIFU for one thyroid nodule, solid or mixed, with mean diameter ≥8 mm, and no suspicion of malignancy. The HIFU device was progressively adjusted with stepwise testing. The energy level for ablation ranged from 35 to 94 J/pulse for different groups of patients. One pathologist examined all removed thyroids. RESULTS: Three patients discontinued treatment due to pain or skin microblister. Among the remaining 22 patients, 16 showed significant changes by ultrasound. Macroscopic and histological examinations showed that all lesions were confined to the targeted nodule without affecting neighboring structures. At pathological analysis, the extent of nodule destruction ranged from 2% to 80%. Five out of 22 patients had over 20% pathological lesions unmistakably attributed to HIFU. Seventeen cases had putative lesions including nonspecific necrosis, hemorrhage, nodule detachment, cavitations, and cysts. Among these 17 cases, 12 had both ultrasound changes and cavitation at histology that may be expected for an HIFU effect. In the last three patients ablated at the highest energy level, significant ultrasound changes and complete coagulative necrosis were observed in 80%, 78%, and 58% of the targeted area, respectively. There were no major complications of ablation. CONCLUSION: This study showed the potential efficacy of HIFU for human thyroid nodule ablation. Lesions were clearly visible by histology and ultrasound after high energy treatments, and safety and tolerability were good. We identified a power threshold for optimal necrosis of the target thyroid tissue. Further studies are ongoing to assess nodule changes at longer follow-up times.

Localized ablation of thyroid tissue by high-intensity focused ultrasound: improvement of noninvasive tissue necrosis methods.

BACKGROUND: Although thyroid nodules are frequently detected in patients during routine examinations, such nodules are rarely malignant. Surgical treatment of nodules is controversial because of the possible complications associated with surgery, and there is an unmet need for a minimally invasive alternative. We previously reported on a high-intensity focused ultrasound (HIFU) device that induced necrosis in ewe thyroids. This complementary study on 27 ewes evaluated the use of the device to produce thyroid lesions, characterized the HIFU-induced lesions on the thyroid and surrounding structures, and evaluated the safety and reproducibility of the method. METHODS: A spherical 3-MHz transducer that was coupled to a 5-MHz linear array ultrasound imaging probe was used to generate powerful acoustic waves to destroy thyroid tissue. Three series of experiments were conducted: thyroid lesion experiments (10 ewes), safety experiments (4 ewes), and reproducibility experiments (13 ewes). After fixation of the ewe's neck, tissue lesions were examined both macroscopically and histologically. RESULTS: First, individual pulsed acoustical waves were used to induce lesions in 19 thyroid lobes. In most lesions, there was coagulative necrosis that was replaced later by fibrosis. Macroscopic examination of adjacent organs revealed skin lesions and muscle injuries. A second series of experiments evaluated the consequences of HIFU pulsed waves on structures surrounding the thyroid to better characterize possible side effects of HIFU. Firings at the periphery of eight lobes revealed macroscopic lesions in the trachea of one ewe and superficial esophagus lesions in three ewes. The recurrent nerves were damaged bilaterally in one ewe that died from dysphagia 3 days after HIFU. Four ewes were found to have muscle injuries, but no skin lesions were observed. A third series of experiments evaluated the reproducibility of a HIFU prototype designed specifically for human use. Thyroid lesions were obtained in 25 of the 26 treated lobes. No damage to the nerves, trachea, esophagus, or muscles was observed. About 3 of the 13 ewes had superficial skin burns. CONCLUSION: The results obtained in the ewe model show that thyroid lesions with a defined volume can be induced safely and suggest that the HIFU device is now ready for evaluation in humans.

High-intensity focused ultrasound for localized thyroid-tissue ablation: preliminary experimental animal study.

OBJECTIVE: Thyroid surgery is common and complications are not rare. High-intensity focused ultrasound (HIFU) could be a possible minimally invasive alternative to surgery. The aim of this study was to assess the feasibility of using HIFU to obtain localized ablation of thyroid tissue without affecting neighboring structures. METHODS: The ewe was chosen as the model because its thyroid is easily accessible, with a size comparable to the human gland. An HIFU device designed to treat human prostate cancer was used. Eight ewes were anesthetised and their thyroids were ablated with ultrasound-guided HIFU. HIFU was generated by a 3-MHz spherical piezocomposite transducer that delivered an average of 24 (range, 9-44) ultrasound pulses per lobe covering a mean volume of 0.7 cm(3) (range, 0.2-1.8). RESULTS: Ewes were sacrificed 6-13 days after HIFU treatment and the anterior part of the neck was fixed in formalin before macroscopic and microscopic examinations. Adverse events occurred mainly for the first treated ewes before the energy delivered to the thyroid tissues was fully mastered: one animal died 3 days after HIFU, most probably because of inhalation pneumonia; the ultrasound beam hit adjacent organs in three animals. As expected, typical histologic lesions of the thyroid were obtained: central coagulative necrosis with ghost vesicular structures, disappearance of the nuclei, and cytoplasmic flocculation. At the periphery of the necrotic zone, a cell reaction was observed with fibroblastic granulation tissue, mononuclear cell infiltrate and regenerating thyroid tissue. CONCLUSION: The results of this preliminary study confirm the possibility of using HIFU in order to destroy a defined area in thyroid tissue. Future experiments in ewes will be focused on the safety of the method by refining the HIFU parameters and by developing a new equipment specially built for the thyroid.