Localization of nitric oxide-producing hemocytes in Aedes and Culex mosquitoes infected with bacteria.

Mosquitoes are significant vectors of various pathogens. Unlike vertebrates, insects rely solely on innate immunity. Hemocytes play a crucial role in the cellular part of the innate immune system. The gaseous radical nitric oxide (NO) produced by hemocytes acts against pathogens and also functions as a versatile transmitter in both the immune and nervous systems, utilizing cyclic guanosine monophosphate (cGMP) as a second messenger. This study conducted a parallel comparison of NO synthase (NOS) expression and NO production in hemocytes during Escherichia coli K12 infection in four vector species: Aedes aegypti, Aedes albopictus, Culex pipiens molestus, and Culex pipiens quinquefasciatus. Increased NOS expression by NADPH diaphorase (NADPHd) staining and NO production by immunofluorescence against the by-product L-citrulline were observed in infected mosquito hemocytes distributed throughout the abdomens. NADPHd activity and citrulline labeling were particularly found in periostial hemocytes near the heart, but also on the ventral nerve chord (VNC). Pericardial cells of Ae. aegypti and Cx. p. molestus showed increased citrulline immunofluorescence, suggesting their involvement in the immune response. Oenocytes displayed strong NADPHd and citrulline labeling independent of infection status. This comparative study, consistent with findings in other species, suggests a widespread phenomenon of NO's role in hemocyte responses during E. coli infection. Found differences within and between genera highlight the importance of species-specific investigations.

West Nile and Usutu viruses are efficiently inactivated in platelet concentrates by UVC light using the THERAFLEX UV-Platelets system.

BACKGROUND AND OBJECTIVES: West Nile virus (WNV) and Usutu virus (USUV) are mosquito-borne flaviviruses (Flaviviridae) that originated in Africa, have expanded their geographical range during the last decades and caused documented infections in Europe in the last years. Acute WNV and USUV infections have been detected in asymptomatic blood donors by nucleic acid testing. Thus, inactivation of both viral pathogens before blood transfusion is necessary to ensure blood product safety. This study aimed to investigate the efficacy of the THERAFLEX UV-Platelets system to inactivate WNV and USUV in platelet concentrates (PCs). MATERIALS AND METHODS: Plasma-reduced PCs were spiked with the virus suspension. Spiked PC samples were taken after spiking (load and hold sample) and after UVC illumination on the Macotronic UV illumination machine with different light doses (0.05, 0.1, 0.15 and 0.2 (standard) J/cm2). Virus loads of WNV and USUV before and after illumination were measured by titration. RESULTS: Infectivity assays showed that UVC illumination inactivated WNV and USUV in a dose-dependent manner. At a UVC dose of 0.2 J/cm2, the WNV titre was reduced by a log10 factor of 3.59 ± 0.43 for NY99 (lineage 1) and 4.40 ± 0.29 for strain ED-I-33/18 (lineage 2). USUV titres were reduced at the same UVC dose by a log10 factor of 5.20 ± 0.70. CONCLUSIONS: Our results demonstrate that the THERAFLEX UV-Platelets procedure is an effective technology to inactivate WNV and USUV in contaminated PCs.

Attentional prioritisation and facilitation for similar stimuli in visual working memory.

Visual working memory (VWM) allows for the brief retention of approximately three to four items. Interestingly, when these items are similar to each other in a feature domain, memory recall performance is elevated compared to when they are dissimilar. This similarity benefit is currently not accounted for by models of VWM. Previous research has suggested that this similarity benefit may arise from selective attentional prioritisation in the maintenance phase. However, the similarity effect has not been contrasted under circumstances where dissimilar item types can adequately compete for memory resources. In Experiment 1, similarity benefits were seen for all-similar over all-dissimilar displays. This was also seen in mixed displays, change detection performance was higher when one of the two similar items changed, compared to when the dissimilar item changed. Surprisingly, the similarity effect was stronger in these mixed displays then when comparing the all-similar and all-dissimilar. Experiment 2 investigated this further by examining how attention was allocated in the memory encoding phase via eye movements. Results revealed that attention prioritised similar over dissimilar items in the mixed displays. Similar items were more likely to receive the first fixation and were fixated more often than dissimilar items. Furthermore, dwell times were elongated for dissimilar items, suggesting that encoding was less efficient. These results suggest that there is an attentional strategy towards prioritising similar items over dissimilar items, and that this strategy's influence can be observed in the memory encoding phase.

Attentional selection is a sufficient cause for visual working memory interference.

Visual attention and visual working memory (VWM) are intertwined processes that allow navigation of the visual world. These systems can compete for highly limited cognitive resources, creating interference effects when both operate in tandem. Performing an attentional task while maintaining a VWM load often leads to a loss of memory information. These losses are seen even with very simple visual search tasks. Previous research has argued that this may be due to the attentional selection process, of choosing the target item out of surrounding nontarget items. Over two experiments, the current study disentangles the roles of search and selection in visual search and their influence on a retained VWM load. Experiment 1 revealed that, when search stimuli were relatively simple, target-absent searches (which did not require attentional selection) did not provoke memory interference, whereas target-present search did. In Experiment 2, the number of potential targets was varied in the search displays. In one condition, participants were required to select any one of the items displayed, requiring an attentional selection but no need to search for a specific item. Importantly, this condition led to memory interference to the same extent as a condition where a single target was presented among nontargets. Together, these results show that the process of attentional selection is a sufficient cause for interference with a concurrently maintained VWM load.

Microbiota-derived tryptophan metabolites in vascular inflammation and cardiovascular disease.

The essential amino acid tryptophan (Trp) is metabolized by gut commensals, yielding in compounds that affect innate immune cell functions directly, but also acting on the aryl hydrocarbon receptor (AHR), thus regulating the maintenance of group 3 innate lymphoid cells (ILCs), promoting T helper 17 (TH17) cell differentiation, and interleukin-22 production. In addition, microbiota-derived Trp metabolites have direct effects on the vascular endothelium, thus influencing the development of vascular inflammatory phenotypes. Indoxyl sulfate was demonstrated to promote vascular inflammation, whereas indole-3-propionic acid and indole-3-aldehyde had protective roles. Furthermore, there is increasing evidence for a contributory role of microbiota-derived indole-derivatives in blood pressure regulation and hypertension. Interestingly, there are indications for a role of the kynurenine pathway in atherosclerotic lesion development. Here, we provide an overview on the emerging role of gut commensals in the modulation of Trp metabolism and its influence in cardiovascular disease development.

Histochemical staining techniques in Culex pipiens and Drosophila melanogaster (Diptera) with a comparison to mammals.

Insects play an important role in ecosystems. Changes in their abundance and biodiversity are of paramount interest, as there has not only been an alarming decline of insects important for ecosystem health throughout the past decades, but also an increase in insects detrimental for biomes. Furthermore, insects pose a threat to modern society as arbovirus-transmitting vectors. Therefore, detailed knowledge of insect staining characteristics could be beneficial as a basis for further studies, whether in the context of species conservation or control of insect pests. Thus, this study compared 14 histochemical stains for their usefulness in insects regarding nervous tissue, connective tissue components, mucins and polysaccharides, mineralization, and microorganisms. The study used formalin-fixed paraffin-embedded tissue sections of mammals (Equus caballus) and 2 dipterans (Culex pipiens biotype molestus, Drosophila melanogaster). Several histochemical stains were suitable for tissue assessment in insects and mammals, in particular for nervous tissue (Bielschowsky silver stain, luxol fast blue-cresyl violet) and polysaccharides (alcian blue, periodic acid-Schiff with and without diastase treatment, toluidine blue). Other stains proved useful for visualization of insect-specific organ characteristics such as Gomori's reticulin stain for tracheoles in both dipteran species, Heidenhain's azan for midgut-associated connective tissue, and von Kossa for mineral deposition in Malpighian tubules of C. pipiens biotype molestus. In summary, this study provides comparable insights into histochemical procedures in mammals and insects and their usefulness for histological assessment of C. pipiens biotype molestus and D. melanogaster.

Inhibition continues to guide search under concurrent visual working memory load.

It is well known that attention can be automatically attracted to salient items. However, recent studies show that it is possible to avoid distraction by a salient item (with a known feature), leading to facilitated search. This article tests a proposed mechanism for distractor inhibition: that a mental representation of the distractor feature held in visual working memory (VWM) allows attention to be guided away from the distractor. We tested this explanation by examining color-based inhibition in visual search for a shape target with and without VWM load. In Experiment 1 the presence of a distractor facilitated visual search under low and high VWM loads, as reflected in faster response times when the distractor was present (compared to absent), and in fewer eye movements to the salient distractor than the non-target items. However, the eye movement inhibition effect was noticeably weakened in the load conditions. Experiment 2 explored further, to distinguish between inhibition of the distractor color and activation of the (irrelevant) target color. Intermittently presenting single-color search trials that contained only either a target, distractor or a neutral-colored singleton revealed that the distractor color attracted attention less than the neutral color with and without VWM load. The target color, however, only attracted attention more than neutral colors under no load, whereas a VWM load completely eliminated this effect. This suggests that although VWM plays a role in guiding attention to the (irrelevant) target color, distractor-feature inhibition can operate independently.

Vector and Host C-Type Lectin Receptor (CLR)-Fc Fusion Proteins as a Cross-Species Comparative Approach to Screen for CLR-Rift Valley Fever Virus Interactions.

Rift Valley fever virus (RVFV) is a mosquito-borne bunyavirus endemic to Africa and the Arabian Peninsula, which causes diseases in humans and livestock. C-type lectin receptors (CLRs) represent a superfamily of pattern recognition receptors that were reported to interact with diverse viruses and contribute to antiviral immune responses but may also act as attachment factors or entry receptors in diverse species. Human DC-SIGN and L-SIGN are known to interact with RVFV and to facilitate viral host cell entry, but the roles of further host and vector CLRs are still unknown. In this study, we present a CLR-Fc fusion protein library to screen RVFV-CLR interaction in a cross-species approach and identified novel murine, ovine, and Aedes aegypti RVFV candidate receptors. Furthermore, cross-species CLR binding studies enabled observations of the differences and similarities in binding preferences of RVFV between mammalian CLR homologues, as well as more distant vector/host CLRs.

Intact Type I Interferon Receptor Signaling Prevents Hepatocellular Necrosis but Not Encephalitis in a Dose-Dependent Manner in Rift Valley Fever Virus Infected Mice.

Rift Valley fever (RVF) is a zoonotic and emerging disease, caused by the RVF virus (RVFV). In ruminants, it leads to "abortion storms" and enhanced mortality rates in young animals, whereas in humans it can cause symptoms like severe hemorrhagic fever or encephalitis. The role of the innate and adaptive immune response in disease initiation and progression is still poorly defined. The present study used the attenuated RVFV strain clone 13 to investigate viral spread, tissue tropism, and histopathological lesions after intranasal infection in C57BL/6 wild type (WT) and type I interferon (IFN-I) receptor I knockout (IFNAR-/-) mice. In WT mice, 104 PFU RVFV (high dose) resulted in a fatal encephalitis, but no hepatitis 7-11 days post infection (dpi), whereas 103 PFU RVFV (low dose) did not cause clinical disease or significant histopathological lesions in liver and the central nervous system (CNS). In contrast, IFNAR-/- mice infected with 103 PFU RVFV developed hepatocellular necrosis resulting in death at 2-5 dpi and lacked encephalitis. These results show that IFNAR signaling prevents systemic spread of the attenuated RVFV strain clone 13, but not the dissemination to the CNS and subsequent fatal disease. Consequently, neurotropic viruses may be able to evade antiviral IFN-I signaling pathways by using the transneuronal instead of the hematogenous route.

The Role of Frontoparietal Cortex across the Functional Stages of Visual Search.

Areas in frontoparietal cortex have been shown to be active in a range of cognitive tasks and have been proposed to play a key role in goal-driven activities (Dosenbach, N. U. F., Fair, D. A., Miezin, F. M., Cohen, A. L., Wenger, K. K., Dosenbach, R. A. T., et al. Distinct brain networks for adaptive and stable task control in humans. Proceedings of the National Academy of Sciences, U.S.A., 104, 11073-11078, 2007; Duncan, J. The multiple-demand (MD) system of the primate brain: Mental programs for intelligent behavior. Trends in Cognitive Sciences, 14, 172-179, 2010). Here, we examine the role this frontoparietal system plays in visual search. Visual search, like many complex tasks, consists of a sequence of operations: target selection, stimulus-response (SR) mapping, and response execution. We independently manipulated the difficulty of target selection and SR mapping in a novel visual search task that involved identical stimulus displays. Enhanced activity was observed in areas of frontal and parietal cortex during both difficult target selection and SR mapping. In addition, anterior insula and ACC showed preferential representation of SR-stage information, whereas the medial frontal gyrus, precuneus, and inferior parietal sulcus showed preferential representation of target selection-stage information. A connectivity analysis revealed dissociable neural circuits underlying visual search. We hypothesize that these circuits regulate distinct mental operations associated with the allocation of spatial attention, stimulus decisions, shifts of task set from selection to SR mapping, and SR mapping. Taken together, the results show frontoparietal involvement in all stages of visual search and a specialization with respect to cognitive operations.

Binding of HasA by its transmembrane receptor HasR follows a conformational funnel mechanism.

HasR in the outer membrane of Serratia marcescens binds secreted, heme-loaded HasA and translocates the heme to the periplasm to satisfy the cell's demand for iron. The previously published crystal structure of the wild-type complex showed HasA in a very specific binding arrangement with HasR, apt to relax the grasp on the heme and assure its directed transfer to the HasR-binding site. Here, we present a new crystal structure of the heme-loaded HasA arranged with a mutant of HasR, called double mutant (DM) in the following that seemed to mimic a precursor stage of the abovementioned final arrangement before heme transfer. To test this, we performed first molecular dynamics (MD) simulations starting at the crystal structure of the complex of HasA with the DM mutant and then targeted MD simulations of the entire binding process beginning with heme-loaded HasA in solution. When the simulation starts with the former complex, the two proteins in most simulations do not dissociate. When the mutations are reverted to the wild-type sequence, dissociation and development toward the wild-type complex occur in most simulations. This indicates that the mutations create or enhance a local energy minimum. In the targeted MD simulations, the first protein contacts depend upon the chosen starting position of HasA in solution. Subsequently, heme-loaded HasA slides on the external surface of HasR on paths that converge toward the specific arrangement apt for heme transfer. The targeted simulations end when HasR starts to relax the grasp on the heme, the subsequent events being in a time regime inaccessible to the available computing power. Interestingly, none of the ten independent simulation paths visits exactly the arrangement of HasA with HasR seen in the crystal structure of the mutant. Two factors which do not exclude each other could explain these observations: the double mutation creates a non-physiologic potential energy minimum between the two proteins and /or the target potential in the simulation pushes the system along paths deviating from the low-energy paths of the native binding processes. Our results support the former view, but do not exclude the latter possibility.

Top-down modulation of gaze capture: Feature similarity, optimal tuning, or tuning to relative features?

It is well-known that we can tune attention to specific features (e.g., colors). Originally, it was believed that attention would always be tuned to the exact feature value of the sought-after target (e.g., orange). However, subsequent studies showed that selection is often geared towards target-dissimilar items, which was variably attributed to (1) tuning attention to the relative target feature that distinguishes the target from other items in the surround (e.g., reddest item; relational tuning), (2) tuning attention to a shifted target feature that allows more optimal target selection (e.g., reddish orange; optimal tuning), or (3) broad attentional tuning and selection of the most salient item that is still similar to the target (combined similarity/saliency). The present study used a color search task and assessed gaze capture by differently coloured distractors to distinguish between the three accounts. The results of the first experiment showed that a very target-dissimilar distractor that matched the relative color of the target but was outside of the area of optimal tuning still captured very strongly. As shown by a control condition and a control experiment, bottom-up saliency modulated capture only weakly, ruling out a combined similarity-saliency account. With this, the results support the relational account that attention is tuned to the relative target feature (e.g., reddest), not an optimal feature value or the target feature.

Gene expression profiles in neurological tissues during West Nile virus infection: a critical meta-analysis.

BACKGROUND: Infections with the West Nile virus (WNV) can attack neurological tissues in the host and alter gene expression levels therein. Several individual studies have analyzed these changes in the transcriptome based on measurements with DNA microarrays. Individual microarray studies produce a high-dimensional data structure with the number of studied genes exceeding the available sample size by far. Therefore, the level of scientific evidence of these studies is rather low and results can remain uncertain. Furthermore, the individual studies concentrate on different types of tissues or different time points after infection. A general statement regarding the transcriptional changes through WNV infection in neurological tissues is therefore hard to make. We screened public databases for transcriptome expression studies related to WNV infections and used different analysis pipelines to perform meta-analyses of these data with the goal of obtaining more stable results and increasing the level of evidence. RESULTS: We generated new lists of genes differentially expressed between WNV infected neurological tissues and control samples. A comparison with these genes to findings of a meta-analysis of immunological tissues is performed to figure out tissue-specific differences. While 5.879 genes were identified exclusively in the neurological tissues, 15 genes were found exclusively in the immunological tissues, and 44 genes were commonly detected in both tissues. Most findings of the original studies could be confirmed by the meta-analysis with a higher statistical power, but some genes and GO terms related to WNV were newly detected, too. In addition, we identified gene ontology terms related to certain infection processes, which are significantly enriched among the differentially expressed genes. In the neurological tissues, 17 gene ontology terms were found significantly different, and 2 terms in the immunological tissues. CONCLUSIONS: A critical discussion of our findings shows benefits but also limitations of the meta-analytic approach. In summary, the produced gene lists, identified gene ontology terms and network reconstructions appear to be more reliable than the results from the individual studies. Our meta-analysis provides a basis for further research on the transcriptional mechanisms by WNV infections in neurological tissues.

Association of Batai Virus Infection and Encephalitis in Harbor Seals, Germany, 2016.

We isolated Batai virus from the brain of a euthanized, 26-year-old, captive harbor seal with meningoencephalomyelitis in Germany. We provide evidence that this orthobunyavirus can naturally infect the central nervous system of a mammal. The full-genome sequence showed differences from a previously reported virus isolate from a mosquito in Germany.

Detection, infection dynamics and small RNA response against Culex Y virus in mosquito-derived cells.

Many insect cell lines are persistently infected with insect-specific viruses (ISV) often unrecognized by the scientific community. Considering recent findings showing the possibility of interference between arbovirus and ISV infections, it is important to pay attention to ISV-infected cell lines. One example is the Entomobirnavirus, Culex Y virus (CYV). Here we describe the detection of CYV using a combination of small RNA sequencing, electron microscopy and PCR in mosquito cell lines Aag2, U4.4 and C7-10. We found CYV-specific small RNAs in all three cell lines. Interestingly, the magnitude of the detected viral RNA genome is variable among cell passages and leads to irregular detection via electron microscopy. Gaining insights into the presence of persistent ISV infection in commonly used mosquito cells and their interactions with the host immune system is beneficial for evaluating the outcome of co-infections with arboviruses of public health concern.

Visual search for emotional expressions: Effect of stimulus set on anger and happiness superiority.

Prior reports of preferential detection of emotional expressions in visual search have yielded inconsistent results, even for face stimuli that avoid obvious expression-related perceptual confounds. The current study investigated inconsistent reports of anger and happiness superiority effects using face stimuli drawn from the same database. Experiment 1 excluded procedural differences as a potential factor, replicating a happiness superiority effect in a procedure that previously yielded an anger superiority effect. Experiments 2a and 2b confirmed that image colour or poser gender did not account for prior inconsistent findings. Experiments 3a and 3b identified stimulus set as the critical variable, revealing happiness or anger superiority effects for two partially overlapping sets of face stimuli. The current results highlight the critical role of stimulus selection for the observation of happiness or anger superiority effects in visual search even for face stimuli that avoid obvious expression related perceptual confounds and are drawn from a single database.

Co-circulation of a novel phlebovirus and Massilia virus in sandflies, Portugal.

BACKGROUND: In Portugal, entomological surveys to detect phleboviruses in their natural vectors have not been performed so far. Thus, the aims of the present study were to detect, isolate and characterize phleboviruses in sandfly populations of Portugal. FINDINGS: From May to October 2007-2008, 896 female sandflies were trapped in Arrábida region, located on the southwest coast of Portugal. Phlebovirus RNA was detected by using a pan-phlebovirus RT-PCR in 4 out of 34 Phlebotomus perniciosus pools. Direct sequencing of the amplicons showed that 2 samples exhibited 72 % nucleotide identity with Arbia virus, and two showed 96 % nucleotide identity with Massilia virus. The Arbia-like virus (named Alcube virus) was isolated in cell culture and complete genomic sequences of one Alcube and two Massila viruses were determined using next-generation sequencing technology. Phylogenetic analysis demonstrated that Alcube virus clustered with members of the Salehabad virus species complex. Within this clade, Alcube virus forms a monophyletic lineage with the Arbia, Salehabad and Adana viruses sharing a common ancestor. Arbia virus has been identified as the most closely related virus with 20-28 % nucleotide and 10-27 % amino acid divergences depending on the analysed segment. CONCLUSIONS: We have provided genetic evidence for the circulation of a novel phlebovirus species named Alcube virus in Ph. perniciosus and co-circulation of Massilia virus, in Arrábida region, southwest of Portugal. Further epidemiological investigations and surveillance for sandfly-borne phleboviruses in Portugal are needed to elucidate their medical importance.

Physico-Chemical Characteristics of Culex pipiens sensu lato and Culex torrentium (Diptera: Culicidae) Breeding Sites in Germany.

Culex pipiens sensu lato (L.) and Culex torrentium Martini, 1925 are two widely distributed mosquito species in Europe. Due to difficulties in morphological discrimination, the current knowledge on differences in the breeding ecology is fragmentary. Therefore, this study evaluated the relation between the presence-absence of both species at various types of breeding habitats in response to physico-chemical parameters, using a recently developed molecular assay that allows reliable species-specific typing of larvae and pupae. The results revealed that the two species often occur in sympatry at the studied breeding sites, and there were no substantial differences concerning presence-absence of the two species with regard to the various environmental parameters investigated.

Distinct neural networks for target feature versus dimension changes in visual search, as revealed by EEG and fMRI.

In visual search, responses are slowed, from one trial to the next, both when the target dimension changes (e.g., from a color target to a size target) and when the target feature changes (e.g., from a red target to a green target) relative to being repeated across trials. The present study examined whether such feature and dimension switch costs can be attributed to the same underlying mechanism(s). Contrary to this contention, an EEG study showed that feature changes influenced visual selection of the target (i.e., delayed N2pc onset), whereas dimension changes influenced the later process of response selection (i.e., delayed s-LRP onset). An fMRI study provided convergent evidence for the two-system view: Compared with repetitions, feature changes led to increased activation in the occipital cortex, and superior and inferior parietal lobules, which have been implicated in spatial attention. By contrast, dimension changes led to activation of a fronto-posterior network that is primarily linked with response selection (i.e., pre-motor cortex, supplementary motor area and frontal areas). Taken together, the results suggest that feature and dimension switch costs are based on different processes. Specifically, whereas target feature changes delay attention shifts to the target, target dimension changes interfere with later response selection operations.

Implicit false-belief processing in the human brain.

Eye-movement patterns in 'Sally-Anne' tasks reflect humans' ability to implicitly process the mental states of others, particularly false-beliefs - a key theory of mind (ToM) operation. It has recently been proposed that an efficient ToM system, which operates in the absence of awareness (implicit ToM, iToM), subserves the analysis of belief-like states. This contrasts to consciously available belief processing, performed by the explicit ToM system (eToM). The frontal, temporal and parietal cortices are engaged when humans explicitly 'mentalize' about others' beliefs. However, the neural underpinnings of implicit false-belief processing and the extent to which they draw on networks involved in explicit general-belief processing are unknown. Here, participants watched 'Sally-Anne' movies while fMRI and eye-tracking measures were acquired simultaneously. Participants displayed eye-movements consistent with implicit false-belief processing. After independently localizing the brain areas involved in explicit general-belief processing, only the left anterior superior temporal sulcus and precuneus revealed greater blood-oxygen-level-dependent activity for false- relative to true-belief trials in our iToM paradigm. No such difference was found for the right temporal-parietal junction despite significant activity in this area. These findings fractionate brain regions that are associated with explicit general ToM reasoning and false-belief processing in the absence of awareness.