Connectomic reconstruction predicts the functional organization of visual inputs to the navigation center of the Drosophila brain.

Many animals, including humans, navigate their surroundings by visual input, yet we understand little about how visual information is transformed and integrated by the navigation system. In Drosophila melanogaster, compass neurons in the donut-shaped ellipsoid body of the central complex generate a sense of direction by integrating visual input from ring neurons, a part of the anterior visual pathway (AVP). Here, we densely reconstruct all neurons in the AVP using FlyWire, an AI-assisted tool for analyzing electron-microscopy data. The AVP comprises four neuropils, sequentially linked by three major classes of neurons: MeTu neurons, which connect the medulla in the optic lobe to the small unit of anterior optic tubercle (AOTUsu) in the central brain; TuBu neurons, which connect the anterior optic tubercle to the bulb neuropil; and ring neurons, which connect the bulb to the ellipsoid body. Based on neuronal morphologies, connectivity between different neural classes, and the locations of synapses, we identified non-overlapping channels originating from four types of MeTu neurons, which we further divided into ten subtypes based on the presynaptic connections in medulla and postsynaptic connections in AOTUsu. To gain an objective measure of the natural variation within the pathway, we quantified the differences between anterior visual pathways from both hemispheres and between two electron-microscopy datasets. Furthermore, we infer potential visual features and the visual area from which any given ring neuron receives input by combining the connectivity of the entire AVP, the MeTu neurons' dendritic fields, and presynaptic connectivity in the optic lobes. These results provide a strong foundation for understanding how distinct visual features are extracted and transformed across multiple processing stages to provide critical information for computing the fly's sense of direction.

Non-celestial polarization vision in arthropods.

Most insects can detect the pattern of polarized light in the sky with the dorsal rim area in their compound eyes and use this visual information to navigate in their environment by means of 'celestial' polarization vision. 'Non-celestial polarization vision', in contrast, refers to the ability of arthropods to analyze polarized light by means of the 'main' retina, excluding the dorsal rim area. The ability of using the main retina for polarization vision has been attracting sporadic, but steady attention during the last decade. This special issue of the Journal of Comparative Physiology A presents recent developments with a collection of seven original research articles, addressing different aspects of non-celestial polarization vision in crustaceans and insects. The contributions cover different sources of linearly polarized light in nature, the underlying retinal and neural mechanisms of object detection using polarization vision and the behavioral responses of arthropods to polarized reflections from water.

Behavioral responses of free-flying Drosophila melanogaster to shiny, reflecting surfaces.

Active locomotion plays an important role in the life of many animals, permitting them to explore the environment, find vital resources, and escape predators. Most insect species rely on a combination of visual cues such as celestial bodies, landmarks, or linearly polarized light to navigate or orient themselves in their surroundings. In nature, linearly polarized light can arise either from atmospheric scattering or from reflections off shiny non-metallic surfaces like water. Multiple reports have described different behavioral responses of various insects to such shiny surfaces. Our goal was to test whether free-flying Drosophila melanogaster, a molecular genetic model organism and behavioral generalist, also manifests specific behavioral responses when confronted with such polarized reflections. Fruit flies were placed in a custom-built arena with controlled environmental parameters (temperature, humidity, and light intensity). Flight detections and landings were quantified for three different stimuli: a diffusely reflecting matt plate, a small patch of shiny acetate film, and real water. We compared hydrated and dehydrated fly populations, since the state of hydration may change the motivation of flies to seek or avoid water. Our analysis reveals for the first time that flying fruit flies indeed use vision to avoid flying over shiny surfaces.

Oncolytic Measles Virus Encoding MicroRNA for Targeted RNA Interference.

Virotherapy is a promising, novel form of cancer immunotherapy currently being investigated in pre-clinical and clinical settings. While generally well-tolerated, the anti-tumor potency of oncolytic virus-based monotherapies needs to be improved further. One of the major factors limiting the replication efficiency of oncolytic viruses are the antiviral defense pathways activated by tumor cells. In this study, we have designed and validated a universal expression cassette for artificial microRNAs that can now be adapted to suppress genes of interest, including potential resistance factors. Transcripts are encoded as a primary microRNA for processing via the predominantly nuclear RNase III Drosha. We have engineered an oncolytic measles virus encoding this universal expression cassette for artificial microRNAs. Virally encoded microRNA was expressed in the range of endogenous microRNA transcripts and successfully mediated target protein suppression. However, absolute expression levels of mature microRNAs were limited when delivered by an oncolytic measles virus. We demonstrate that measles virus, in contrast to other cytosolic viruses, does not induce translocation of Drosha from the nucleus into the cytoplasm, potentially resulting in a limited processing efficiency of virus-derived, cytosolically delivered artificial microRNAs. To our knowledge, this is the first report demonstrating functional expression of microRNA from oncolytic measles viruses potentially enabling future targeted knockdown, for instance of antiviral factors specifically in tumor cells.

Pain provocation tests and clinical entities in male football players with longstanding groin pain are associated with pain intensity and disability.

BACKGROUND: Clinical examination of male football players with longstanding groin pain can be considered difficult. Pain provocation tests are used to examine and classify longstanding groin pain into clinical entities as adductor-, iliopsoas-, inguinal-, and pubic-related. It is unknown if pain provocation tests and clinical entities are associated with pain intensity and disability. OBJECTIVES: To investigate if the number of positive pain provocation tests and clinical entities are associated with pain intensity and disability, measured by the Copenhagen 5-Second Squeeze Test (5SST) and the Copenhagen Hip and Groin Outcome Score (HAGOS), respectively. DESIGN: Cross-sectional. METHOD: Forty male football players (age: mean 24 years [SD: 3.2]; height: mean 182 cm [SD: 5.7]; weight: mean 78 Kg [SD: 6.6]) with longstanding groin pain for a median of 8.5 months (IQR: 4-36) were included. The players underwent a bilateral groin examination with 33 pain provocation tests and were classified with clinical entities (0-7) based on the test findings. RESULTS: The number of positive pain provocation tests (median 10, range 2-23) correlated with pain intensity (5SST: rs = 0.70 [95% CI: 0.50, 0.83]) and disability (HAGOS subscales Sport: rs =-0.62 [95% CI: -0.81, -0.36], Pain: rs = -0.38 [95% CI: -0.69, -0.06], Symptoms: rs = 0.52 [95% CI: -0.73, -0.24], ADL: rs = -0.48 [95% CI: -0.71, -0.18]). The number of clinical entities (median 3, range: 1-7) showed similar but weaker correlations to pain intensity and disability. CONCLUSIONS: In male football players with longstanding groin pain, the number of positive pain provocation tests and clinical entities shows weak to strong correlations with pain intensity and disability. Consequently, when pain intensity and disability are severe, a higher number of pain provocation tests may be positive, and more clinical entities may be present.

Cytokine release syndrome-like serum responses after COVID-19 vaccination are frequent and clinically inapparent under cancer immunotherapy.

Patients with cancer frequently receive immune-checkpoint inhibitors (ICIs), which may modulate immune responses to COVID-19 vaccines. Recently, cytokine release syndrome (CRS) was observed in a patient with cancer who received BTN162b2 vaccination under ICI treatment. Here, we analyzed adverse events and serum cytokines in patients with 23 different tumors undergoing (n = 64) or not undergoing (n = 26) COVID-19 vaccination under ICI therapy in a prospectively planned German single-center cohort study (n = 220). We did not observe clinically relevant CRS (≥grade 2) after vaccination (95% CI 0-5.6%; Common Terminology of Adverse Events v.5.0) in this small cohort. Within 4 weeks after vaccination, serious adverse events occurred in eight patients (12.5% 95% CI 5.6-23%): six patients were hospitalized due to events common under cancer therapy including immune related adverse events and two patients died due to conditions present before vaccination. Despite absence of CRS symptoms, a set of pairwise-correlated CRS-associated cytokines, including CXCL8 and interleukin-6 was >1.5-fold upregulated in 40% (95% CI 23.9-57.9%) of patients after vaccination. Hence, elevated cytokine levels are common and not sufficient to establish CRS diagnosis.

Rapid Spike in Hip Adduction Strength in Early Adolescent Footballers: A Study of 125 Elite Male Players From Youth to Senior.

PURPOSE: To investigate the differences in hip adductor and abductor muscle strength in elite male footballers from youth to senior level. METHODS: We tested 125 players from the under-13-years (U'13) to senior squads of a Danish male professional football club in this cross-sectional design study. Hip adductor and abductor force (in newtons), torque (in newton meters), normalized torque (in newton meters per body mass), and adduction-to-abduction ratio were measured using handheld dynamometry. RESULTS: Between U'13 and senior level, adductor force increased by 104%, torque by 127%, and normalized torque by 21%. Abductor force increased by 78%, torque by 126%, and normalized torque by 17%. For incremental differences between age groups, significant increases were observed between the ages of U'13 to U'14 (18%-39%) and U'14 to U'15 (19%-33%) for all strength measures (P ≤ .021). No incremental difference was observed for adductor-to-abductor ratio. CONCLUSIONS: The large increases in hip adductor and abductor strength occurring between the ages of U'13 and U'15 offer insight into the strength capabilities and stress demands in these players, which may relate to injury vulnerability, and facilitate clinicians in selecting best-suited exercise interventions.

Hip adduction strength and provoked groin pain: A comparison of long-lever squeeze testing using the ForceFrame and the Copenhagen 5-Second-Squeeze test.

OBJECTIVES: To compare long-lever squeeze testing using the ForceFrame and the Copenhagen 5-Second-Squeeze test (5SST) for assessment of hip adduction strength and provoked groin pain in elite male soccer players. DESIGN: Cross-sectional study. SETTING: Pre-season testing at facilities of a Danish professional 1st tier soccer club and academy. PARTICIPANTS: Elite male soccer players (n = 83, mean age; 16 ± 2.7 years) from U13, U14, U15, U17, U19 and senior teams cleared for full training and match participation. MAIN OUTCOME MEASURES: Maximum isometric hip adduction strength (Nm/kg) and provoked groin pain (NRS 0-10). RESULTS: Hip adduction strength was 16% lower in the ForceFrame. A Bland-Altman plot showed a systematic bias (-0.47 Nm/kg, 95% CI [-0.57; -0.38]) and lack of agreement (95% limits of agreement: -1.31; 0.39 Nm/kg). In the ForceFrame, provoked groin pain was less intense (median NRS 0 [IQR: 0-1] vs. 5SST: 1 [IQR: 0-3], p < 0.001) and reported by fewer players (NRS >0) (27% [n = 22] vs. 5SST: 61.4% [n = 51], p < 0.001). CONCLUSIONS: The ForceFrame and the 5SST lack agreement and are not interchangeable methods. This may have implications when selecting a method for screening and detecting early groin problems in male soccer players.

Biodiversity and succession of lactic microbiota involved in Brazilian buffalo mozzarella cheese production.

The biodiversity and succession of lactic acid bacteria (LAB) involved in the production and storage of Brazilian buffalo mozzarella cheese were evaluated. The isolates were characterized by Gram staining and catalase test, by the ability to grow at different conditions: temperatures, pH, concentrations of NaCl, and production of CO2 from glucose. The biodiversity and succession of 152 LAB isolated during cheese production were evaluated by 16S rRNA gene sequencing, Random Amplified Polymorphic DNA (RAPD-PCR), and Restriction Fragment Length Polymorphism (RFLP-PCR) techniques. Most of the strains grow well at 30 °C and are tolerant to 6.5% of NaCl, and in general, the best pH for growing was 9.6. Leuconostoc mesenteroides, Lacticaseibacillus casei, Limosilactobacillus fermentum, and Enterococcus sp. were prevalent and present in almost all steps of production. The LAB strains are typically found in the traditional Italian cheese, except the Leuconostoc citreum species. Sixty clusters were obtained by RAPD-PCR with 85% of similarity (114 isolates) while most of the LAB was clustered with 100% of similarity by the RFLP-PCR technique. The applied techniques enabled a valuable elucidation of the LAB biodiversity and succession, contributing to a better understanding of the specific microbial cultures with a technological aptitude of this cheese.

Synaptic targets of photoreceptors specialized to detect color and skylight polarization in Drosophila.

Color and polarization provide complementary information about the world and are detected by specialized photoreceptors. However, the downstream neural circuits that process these distinct modalities are incompletely understood in any animal. Using electron microscopy, we have systematically reconstructed the synaptic targets of the photoreceptors specialized to detect color and skylight polarization in Drosophila, and we have used light microscopy to confirm many of our findings. We identified known and novel downstream targets that are selective for different wavelengths or polarized light, and followed their projections to other areas in the optic lobes and the central brain. Our results revealed many synapses along the photoreceptor axons between brain regions, new pathways in the optic lobes, and spatially segregated projections to central brain regions. Strikingly, photoreceptors in the polarization-sensitive dorsal rim area target fewer cell types, and lack strong connections to the lobula, a neuropil involved in color processing. Our reconstruction identifies shared wiring and modality-specific specializations for color and polarization vision, and provides a comprehensive view of the first steps of the pathways processing color and polarized light inputs.

Virotherapy in Germany-Recent Activities in Virus Engineering, Preclinical Development, and Clinical Studies.

Virotherapy research involves the development, exploration, and application of oncolytic viruses that combine direct killing of cancer cells by viral infection, replication, and spread (oncolysis) with indirect killing by induction of anti-tumor immune responses. Oncolytic viruses can also be engineered to genetically deliver therapeutic proteins for direct or indirect cancer cell killing. In this review-as part of the special edition on "State-of-the-Art Viral Vector Gene Therapy in Germany"-the German community of virotherapists provides an overview of their recent research activities that cover endeavors from screening and engineering viruses as oncolytic cancer therapeutics to their clinical translation in investigator-initiated and sponsored multi-center trials. Preclinical research explores multiple viral platforms, including new isolates, serotypes, or fitness mutants, and pursues unique approaches to engineer them towards increased safety, shielded or targeted delivery, selective or enhanced replication, improved immune activation, delivery of therapeutic proteins or RNA, and redirecting antiviral immunity for cancer cell killing. Moreover, several oncolytic virus-based combination therapies are under investigation. Clinical trials in Germany explore the safety and potency of virotherapeutics based on parvo-, vaccinia, herpes, measles, reo-, adeno-, vesicular stomatitis, and coxsackie viruses, including viruses encoding therapeutic proteins or combinations with immune checkpoint inhibitors. These research advances represent exciting vantage points for future endeavors of the German virotherapy community collectively aimed at the implementation of effective virotherapeutics in clinical oncology.

Vision: Space and colour meet in the fly optic lobes.

Colour vision involves colour-opponent cells, which are excited and inhibited by different wavelengths. Synaptic interconnections between Drosophila Dm8 cells are required for forming spatio-chromatic receptive fields with a center and surround of opposing polarity which can invert, depending on the stimulus.

Colour opponency: Chromatic and achromatic circuits in the mix.

The molecular genetic dissection of Drosophila colour vision circuitry reveals converging pathways previously categorized as being chromatic versus achromatic. Amacrine-like Dm8 cells receive direct and indirect inputs with different spectral sensitivity tuning, thereby forming the second stage of colour-opponent processing.

Hamstring and Quadriceps Muscle Strength in Youth to Senior Elite Soccer: A Cross-Sectional Study Including 125 Players.

PURPOSE: Increasing age, high quadriceps strength, and low hamstring muscle strength are associated with hamstring strain injury in soccer. The authors investigated the age-related variation in maximal hamstring and quadriceps strength in male elite soccer players from under-13 (U-13) to the senior level. METHODS: A total of 125 elite soccer players were included from a Danish professional soccer club and associated youth academy (first tier; U-13, n = 19; U-14, n = 16; U-15, n = 19; U-17, n = 24; U-19, n = 17; and senior, n = 30). Maximal voluntary isometric force was assessed for the hamstrings at 15° knee joint angle and for the quadriceps at 60° knee joint angle (0° = full extension) using an external-fixated handheld dynamometer. Hamstring-to-quadriceps strength (H:Q) ratio and hamstring and quadriceps maximal voluntary isometric force levels were compared across age groups (U-13 to senior). RESULTS: Senior players showed 18% to 26% lower H:Q ratio compared with all younger age groups (P ≤ .026). Specific H:Q ratios (mean [95% confidence interval]) were as follows: senior, 0.45 (0.42-0.48); U-19, 0.61 (0.55-0.66); U-17, 0.56 (0.51-0.60); U-15, 0.59 (0.54-0.64); U-14, 0.54 (0.50-0.59); and U-13, 0.57 (0.51-0.62). Hamstring strength increased from U-13 to U-19 with a significant drop from U-19 to the senior level (P = .048), whereas quadriceps strength increased gradually from U-13 to senior level. CONCLUSION: Elite senior soccer players demonstrate lower H:Q ratio compared with youth players, which is driven by lower hamstring strength at the senior level compared with the U-19 level combined with a higher quadriceps strength. This discrepancy in hamstring and quadriceps strength capacity may place senior-level players at increased risk of hamstring muscle strain injuries.

Alemtuzumab treatment in Denmark: A national study based on the Danish Multiple Sclerosis Registry.

OBJECTIVE: To investigate clinical outcomes in a real-world setting in the complete population-based cohort of alemtuzumab-treated MS patients in Denmark. METHODS: Data were retrieved from The Danish Multiple Sclerosis Registry between 2009 and 2019. Demographic and disease-specific patient parameters related to treatment history, efficacy, and safety outcomes were assessed at baseline and during follow-up visits. RESULTS: A total of 209 patients (78% female) started treatment with alemtuzumab during the study period with 3.1 ± 1.4 years follow-up. After 2 years, 75% of patients were relapse-free compared to 48% the year before alemtuzumab (p < 0.001). The annual number of relapses was reduced by 69% in year 4 compared with the year prior alemtuzumab. More active disease before alemtuzumab increased the annual hazard rate for relapse (HR: 2.88, p < 0.001). The Expanded Disability Status Scale (EDSS) score remained stable or improved in 81% of patients after 2 years. The need for an additional treatment course was associated with higher number of relapses in the year before alemtuzumab (odds ratio (OR) = 1.95, p = 0.001). CONCLUSION: In a country with primarily escalation strategy, relapse rate reduction was maintained for 5 years, and EDSS stabilized/improved in majority of patients. Higher relapse rate 1 year before alemtuzumab increased the odds for additional courses. Novel serious AEs were not observed.

The development and function of neuronal subtypes processing color and skylight polarization in the optic lobes of Drosophila melanogaster.

The retinal mosaics of many insects contain different ommatidial subtypes harboring photoreceptors that are both molecularly and morphologically specialized for comparing between different wavelengths versus detecting the orientation of skylight polarization. The neural circuits underlying these different inputs and the characterization of their specific cellular elements are the subject of intense research. Here we review recent progress on the description of both assembly and function of color and skylight polarization circuitry, by focusing on two cell types located in the distal portion of the medulla neuropil of the fruit fly Drosophila melanogaster's optic lobes, called Dm8 and Dm9. In the main part of the retina, Dm8 cells fall into two molecularly distinct subtypes whose center becomes specifically connected to either one of randomly distributed 'pale' or 'yellow' R7 photoreceptor fates during development. Only in the 'dorsal rim area' (DRA), both polarization-sensitive R7 and R8 photoreceptors are connected to different Dm8-like cell types, called Dm-DRA1 and Dm-DRA2, respectively. An additional layer of interommatidial integration is introduced by Dm9 cells, which receive input from multiple neighboring R7 and R8 cells, as well as providing feedback synapses back into these photoreceptors. As a result, the response properties of color-sensitive photoreceptor terminals are sculpted towards being both maximally decorrelated, as well as harboring several levels of opponency (both columnar as well as intercolumnar). In the DRA, individual Dm9 cells appear to mix both polarization and color signals, thereby potentially serving as the first level of integration of different celestial stimuli. The molecular mechanisms underlying the establishment of these synaptic connections are beginning to be revealed, by using a combination of live imaging, developmental genetic studies, and cell type-specific transcriptomics.

Parallel Visual Pathways with Topographic versus Nontopographic Organization Connect the Drosophila Eyes to the Central Brain.

One hallmark of the visual system is a strict retinotopic organization from the periphery toward the central brain, where functional imaging in Drosophila revealed a spatially accurate representation of visual cues in the central complex. This raised the question how, on a circuit level, the topographic features are implemented, as the majority of visual neurons enter the central brain converge in optic glomeruli. We discovered a spatial segregation of topographic versus nontopographic projections of distinct classes of medullo-tubercular (MeTu) neurons into a specific visual glomerulus, the anterior optic tubercle (AOTU). These parallel channels synapse onto different tubercular-bulbar (TuBu) neurons, which in turn relay visual information onto specific central complex ring neurons in the bulb neuropil. Hence, our results provide the circuit basis for spatially accurate representation of visual information and highlight the AOTU's role as a prominent relay station for spatial information from the retina to the central brain.