Targeting cathepsin S promotes activation of OLF1-BDNF/TrkB axis to enhance cognitive function.

BACKGROUND: Cathepsin S (CTSS) is a cysteine protease that played diverse roles in immunity, tumor metastasis, aging and other pathological alterations. At the cellular level, increased CTSS levels have been associated with the secretion of pro-inflammatory cytokines and disrupted the homeostasis of Ca2+ flux. Once CTSS was suppressed, elevated levels of anti-inflammatory cytokines and changes of Ca2+ influx were observed. These findings have inspired us to explore the potential role of CTSS on cognitive functions. METHODS: We conducted classic Y-maze and Barnes Maze tests to assess the spatial and working memory of Ctss-/- mice, Ctss+/+ mice and Ctss+/+ mice injected with the CTSS inhibitor (RJW-58). Ex vivo analyses including long-term potentiation (LTP), Golgi staining, immunofluorescence staining of sectioned whole brain tissues obtained from experimental animals were conducted. Furthermore, molecular studies were carried out using cultured HT-22 cell line and primary cortical neurons that treated with RJW-58 to comprehensively assess the gene and protein expressions. RESULTS: Our findings reported that targeting cathepsin S (CTSS) yields improvements in cognitive function, enhancing both working and spatial memory in behavior models. Ex vivo studies showed elevated levels of long-term potentiation levels and increased synaptic complexity. Microarray analysis demonstrated that brain-derived neurotrophic factor (BDNF) was upregulated when CTSS was knocked down by using siRNA. Moreover, the pharmacological blockade of the CTSS enzymatic activity promoted BDNF expression in a dose- and time-dependent manner. Notably, the inhibition of CTSS was associated with increased neurogenesis in the murine dentate gyrus. These results suggested a promising role of CTSS modulation in cognitive enhancement and neurogenesis. CONCLUSION: Our findings suggest a critical role of CTSS in the regulation of cognitive function by modulating the Ca2+ influx, leading to enhanced activation of the BDNF/TrkB axis. Our study may provide a novel strategy for improving cognitive function by targeting CTSS.

Examining Human-Smartphone Interaction as a Proxy for Circadian Rhythm in Patients With Insomnia: Cross-Sectional Study.

BACKGROUND: The sleep and circadian rhythm patterns associated with smartphone use, which are influenced by mental activities, might be closely linked to sleep quality and depressive symptoms, similar to the conventional actigraphy-based assessments of physical activity. OBJECTIVE: The primary objective of this study was to develop app-defined circadian rhythm and sleep indicators and compare them with actigraphy-derived measures. Additionally, we aimed to explore the clinical correlations of these indicators in individuals with insomnia and healthy controls. METHODS: The mobile app "Rhythm" was developed to record smartphone use time stamps and calculate circadian rhythms in 33 patients with insomnia and 33 age- and gender-matched healthy controls, totaling 2097 person-days. Simultaneously, we used standard actigraphy to quantify participants' sleep-wake cycles. Sleep indicators included sleep onset, wake time (WT), wake after sleep onset (WASO), and the number of awakenings (NAWK). Circadian rhythm metrics quantified the relative amplitude, interdaily stability, and intradaily variability based on either smartphone use or physical activity data. RESULTS: Comparisons between app-defined and actigraphy-defined sleep onsets, WTs, total sleep times, and NAWK did not reveal any significant differences (all P>.05). Both app-defined and actigraphy-defined sleep indicators successfully captured clinical features of insomnia, indicating prolonged WASO, increased NAWK, and delayed sleep onset and WT in patients with insomnia compared with healthy controls. The Pittsburgh Sleep Quality Index scores were positively correlated with WASO and NAWK, regardless of whether they were measured by the app or actigraphy. Depressive symptom scores were positively correlated with app-defined intradaily variability (ß=9.786, SD 3.756; P=.01) and negatively correlated with actigraphy-based relative amplitude (ß=-21.693, SD 8.214; P=.01), indicating disrupted circadian rhythmicity in individuals with depression. However, depressive symptom scores were negatively correlated with actigraphy-based intradaily variability (ß=-7.877, SD 3.110; P=.01) and not significantly correlated with app-defined relative amplitude (ß=-3.859, SD 12.352; P=.76). CONCLUSIONS: This study highlights the potential of smartphone-derived sleep and circadian rhythms as digital biomarkers, complementing standard actigraphy indicators. Although significant correlations with clinical manifestations of insomnia were observed, limitations in the evidence and the need for further research on predictive utility should be considered. Nonetheless, smartphone data hold promise for enhancing sleep monitoring and mental health assessments in digital health research.

Immunogenicity of a spike protein subunit-based COVID-19 vaccine with broad protection against various SARS-CoV-2 variants in animal studies.

SARS-CoV-2 pandemic has profound impacts on human life and global economy since the outbreak in 2019. With the new variants continue to emerge with greater immune escaping capability, the protectivity of the available vaccines is compromised. Therefore, development a vaccine that is capable of inducing immunity against variants including omicron strains is in urgent need. In this study, we developed a protein-based vaccine BCVax that is consisted of antigen delta strain spike protein and QS21-based adjuvant AB801 in nanoparticle immune stimulation complex format (AB801-ISCOM). Results from animal studies showed that high level of anti-S protein IgG was induced after two doses of BCVax and the IgG was capable of neutralizing multiple variants of pseudovirus including omicron BA.1 or BA.2 strains. In addition, strong Th1 response was stimulated after BCVax immunization. Furthermore, BCvax with AB801-ISCOM as the adjuvant showed significant stronger immunity compared with the vaccine using aluminum hydroxide plus CpG 1018 as the adjuvant. BCVax was also evaluated as a booster after two prior vaccinations, the IgG titers and pseudovirus neutralization activities against BA.2 or BA.4/BA.5 were further enhanced suggesting BCVax is a promising candidate as booster. Taken together, the pre-clinical data warrant BCVax for further development in clinic.

A tale of two lineages: how the strains of the earliest divergent symbiotic Frankia clade spread over the world.

It is currently assumed that around 100 million years ago, the common ancestor to the Fabales, Fagales, Rosales and Cucurbitales in Gondwana, developed a root nodule symbiosis with a nitrogen-fixing bacterium. The symbiotic trait evolved first in Frankia cluster-2; thus, strains belonging to this cluster are the best extant representatives of this original symbiont. Most cluster-2 strains could not be cultured to date, except for Frankia coriariae, and therefore many aspects of the symbiosis are still elusive. Based on phylogenetics of cluster-2 metagenome-assembled genomes (MAGs), it has been shown that the genomes of strains originating in Eurasia are highly conserved. These MAGs are more closely related to Frankia cluster-2 in North America than to the single genome available thus far from the southern hemisphere, i.e., from Papua New Guinea.To unravel more biodiversity within Frankia cluster-2 and predict routes of dispersal from Gondwana, we sequenced and analysed the MAGs of Frankia cluster-2 from Coriaria japonica and Coriaria intermedia growing in Japan, Taiwan and the Philippines. Phylogenetic analyses indicate there is a clear split within Frankia cluster-2, separating a continental from an island lineage. Presumably, these lineages already diverged in Gondwana.Based on fossil data on the host plants, we propose that these two lineages dispersed via at least two routes. While the continental lineage reached Eurasia together with their host plants via the Indian subcontinent, the island lineage spread towards Japan with an unknown host plant.

Role of Recipient Susceptibility Factors During Contact-Dependent Interbacterial Competition.

Bacteria evolved multiple strategies to survive and develop optimal fitness in their ecological niche. They deployed protein secretion systems for robust and efficient delivery of antibacterial toxins into their target cells, therefore inhibiting their growth or killing them. To maximize antagonism, recipient factors on target cells can be recognized or hijacked to enhance the entry or toxicity of these toxins. To date, knowledge regarding recipient susceptibility (RS) factors and their mode of action is mostly originating from studies on the type Vb secretion system that is also known as the contact-dependent inhibition (CDI) system. Yet, recent studies on the type VI secretion system (T6SS), and the CDI by glycine-zipper protein (Cdz) system, also reported the emerging roles of RS factors in interbacterial competition. Here, we review these RS factors and their mechanistic impact in increasing susceptibility of recipient cells in response to CDI, T6SS, and Cdz. Past and future strategies for identifying novel RS factors are also discussed, which will help in understanding the interplay between attacker and prey upon secretion system-dependent competition. Understanding these mechanisms would also provide insights for developing novel antibacterial strategies to antagonize aggressive bacteria-killing pathogens.

A quantitative luciferase-based cell-cell fusion assay to measure four-serotype dengue virus E protein-triggered membrane fusion.

The class II membrane fusion induced by flavivirus E proteins is a unique pH-dependent membrane fusion process differently from the class I or III membrane fusion by other enveloped virus proteins. The fusion peptide on the DII of the flavivirus E proteins can insert into the cell membrane as a cell entry process besides the receptor bindings. A traditional assay using C6/36 mosquito cells infected by dengue viruses has been previously reported but did not provide efficient quantitation to measure the virus-triggered membrane fusion. Here we reported the development of a quantitative cell fusion assay for four serotypes of dengue viruses and the recently emerged Zika viruses. We used a pCI-neo vector encoding the prME genes of dengue and Zika viruses and investigated the cell fusion in transfected 293, Vero and CHO cells. Donor cells were co-transfection of the prME genes of dengue and Zika prME gene and T7 RNA polymerase to react with the indicator cells transfected with the luciferase gene under the control of the T7 promoter. Quantification of the virus-induced cell fusion was determined by the luciferase expression levels under a switch of pH from 7.4 to 5.4 in the co-cultured donor and indicator cells. The quantitative luciferase-based assay was applied to measure the anti-fusion activity by two monoclonal antibodies mAb 4G2 and mAb DB42 against dengue virus infections. This assay could quality as a quantitative bioassay for testing the potency of anti-fusion monoclonal antibodies.

A novel method for assessing oral mixing ability based on the spatial clusters quantified by variogram.

BACKGROUND: The two-colour chewing test (TCCT) has been widely used for assessing oral mixing ability, a critical component of masticatory performance. Most studies focused on quantifying the evenness of colour distribution. It remained unknown if the variation of colour clustering was a valid index of oral mixing ability. OBJECTIVE: The study aims to investigate the oral mixing ability based on the spatial clusters quantified by variogram. METHODS: Fifty older people (15 male/35 female, age: 66.0 ± 7.8 years) were assessed for the TCCT and the colour-changeable chewing gum test (CCGT). For the CCGT, we quantified the degree of colour change (ΔE). For the TCCT, the highest peak in colour histogram (HP), the standard deviation of colour values (SDC) and the range of variogram from colour spatial distribution (VARG) were quantified. The participants were grouped according to the contacts of posterior teeth, as assessed by Eichner Index (EI). RESULTS: Highest peak, SDC and VARG showed statistically significant differences between the EI groups (two-tailed independent t test P < .05). Higher VARG (ie a lower degree of clustering) was significantly negatively correlated with ΔE (r = -.36, one-tailed P < .01). The binary logistic regression revealed that among the spatial indices (HP, SDC and VARG), only VARG achieved statistical significance in prediction to the EI group. Eliminating other indices was insignificant to the model performance. CONCLUSIONS: Our results show that the averaged cluster sizes, quantified by variogram, are a valid index for quantifying the TCCT. Compared with other spatial indices, it had the best predictability to the condition of posterior contact.

Association between regional brain volume and masticatory performance differed in cognitively impaired and non-impaired older people.

Older patients with cognitive impairment exhibit worse masticatory performance (MP), which may be attributed to poorer abilities of control, learning, and adaptation of oral sensorimotor functions. This study tested the hypothesis that the association between the structural signature of the right premotor cortex (which relates to sensorimotor integration) and masticatory performance would differ between cognitively impaired (CI) and healthy older people. Thirty-one CI and 31 non-CI older participants (aged 60-84 years; male: female = 28: 34) were recruited and between-group matched for the average age and sex ratio. All the participants received T1-weighted magnetic resonance imaging (MRI) and assessments of MP and the number of missing teeth (NMT). Voxel-based morphometry (VBM) was performed to quantify the gray matter volume (GMV). VBM analyses revealed that in the non-CI but not the CI group, MP was positively correlated with regional GMV at the right premotor cortex (PMC) (p = 0.036, corrected for familywise error under small volume correction), controlling for the participants' sex, age, NMT, and total intracranial volume. Multiple linear regression models revealed that in the non-CI group, the right PMC (beta = 0.3, p = 0.049) was a significant predictor of individual MP. In the CI group, only NMT (beta = -0.7, p = 0.001) was a significant predictor of MP. In non-CI older people, both the NMT and the regional GMV of the right PMC contributed to individual MP. In contrast, in CI patients, tooth loss was the dominant factor in MP. An altered association of the brain-stomatognathic system could be linked to cognitive impairment.

Association between tooth loss and gray matter volume in cognitive impairment.

Previous studies have reported an association between tooth loss and gray matter volume (GMV) in healthy adults. The study aims to elucidate the link between tooth loss, brain volume differences, and cognitive impairment by investigating the total and regional differences in GMV associated with tooth loss in older people with and without cognitive impairment. Forty older participants with mild cognitive impairment or Alzheimer's disease [the cognitive impairment (CI) group] and 30 age- and sex-matched healthy participants [the control (CON) group] received T1-weighted magnetic resonance imaging scans and assessments of oral functions, including masticatory performance (MP) and the number of missing teeth (NMT). Voxel-based morphometry was used to assess the total and regional GMV, including that of the medial temporal lobe and motor-related areas. (A) When the total intracranial volume and age were controlled for, an increased MP was associated with a larger GMV in the premotor cortex in the CON group. (B) In the CI group, an increased NMT was significantly correlated with smaller regional GMV of the bilateral primary motor cortex and the premotor cortex. (C) In the CI group, but not the CON group, an increased NMT was associated with both smaller total GMV and regional GMV of the left medial temporal lobe, including the left hippocampus and parahippocampus. Tooth loss may be preferentially related to the structural differences in the medial temporal lobe in cognitively impaired older people. Further research is required to understand the mechanisms of the relationships.

A High-throughput Interbacterial Competition Platform.

Contact-dependent interbacterial competition is a common strategy used by bacteria to fight for their ecological niches. Interbacterial competition is monitored by a competition assay involving co-culturing the attacker and the recipient bacterial cells on agar, followed by recovery of the surviving recipient cells. Conventional interbacterial competition assays rely on serial dilution, plate spreading, and colony counting experiments for the readout. The high demand for time and labor in a competition assay limits its use for large-scale screening. However, a high-throughput interbacterial competition screening method is required to screen genetic factors involved in an interbacterial competition. Here, using Agrobacterium tumefaciens as an attacker and Escherichia coli as a recipient, we developed a robust, fast, efficient, and high-throughput type VI secretion system-dependent interbacterial competition screening platform. This system allows for 96 simultaneous competition assays without the need for serial dilution and plate spreading. Data analysis of this system relies on only direct and straightforward colony counting. This platform may be easily adapted to identify novel factors involved in any contact-dependent interbacterial competition systems.

Dengue and Zika Virus Domain III-Flagellin Fusion and Glycan-Masking E Antigen for Prime-Boost Immunization.

The viral E proteins of dengue virus (DENV) and Zika virus (ZIKV) are the major viral proteins involved in receptor binding and fusion, and for the induction of protective antibodies against viral infections. DIII of the E proteins is an independent domain and stretches out on the virion surface that can elicit type-specific neutralizing antibodies. For recombinant DIII vaccine development, prime-boost immunizations can provide an advantage of eliciting more type-specific neutralizing antibodies by recalling DIII antigens after DIII booster to improve protection. Methods: The DIII of the E genes of DENV and ZIKV were fused with bacterial fliC gene for the expression of flagellin-DIII (FliC-DIII) fusion proteins. Prime-boost immunization strategies by the second-dose booster of four DENV serotype or ZIKV FliC-DIII fusion proteins were used to investigate the induction of neutralizing antibodies and protection against viral infections. Cross-reactive non-neutralizing antibodies in each group of antisera were also examined using in vitro antibody-dependent enhancement (ADE) assay. A series of glycan-masking E antigens were finally constructed for prime-boost immunizations to abolish the elicitation of cross-reactive non-neutralizing antibodies for ADE activity. Results: We showed that inclusion of a bivalent live-attenuated vaccine with a FliC-DIII booster is superior in eliciting neutralization titers and protection in vivo against all four-serotype DENVs. We also demonstrated that recombinant adenovirus vectors encoding four-serotype DENV prMEs with a FliC-DIII prime-boost scheme is capable of eliciting good antibody responses. In contract, recombinant adenovirus vector of ZIKV prME gene priming, followed by ZIKV FliC-DIII booster did not improve vaccine efficacy. The glycan-masking mutation on the ZIKV E protein ij loop (E-248NHT), but not on DENV2 E protein ij loop (E-242NHT), resulted in abolishing the elicitation of cross-reactive antibodies for DENV and ZIKV infection enhancements. Conclusions: Our findings can provide useful information for designing novel immunogens and vaccination strategies in an attempt to develop a safe and efficacious DENV or ZIKV vaccine.

Lysosomal cysteine protease cathepsin S is involved in cancer cell motility by regulating store-operated Ca2+ entry.

Cathepsin S (CTSS), a lysosomal cysteine protease, has been reported to be associated with extracellular matrix (ECM) degradation, thus promoting cell migration and invasion, but whether CTSS regulates other intracellular mechanisms during metastasis remains unknown. The expression of CTSS was knocked down using siRNA transfection, and enzymatic activity was inhibited by the highly-selective CTSS inhibitor RJW-58. The results of in vitro functional assays, western blot analysis, and an in vivo colonization model demonstrated that CTSS was positively related to cellular adhesive ability. Moreover, both CTSS knockdown and inhibition significantly decreased Ca2+ influx via store-operated Ca2+ entry (SOCE) without changing STIM1 and Orai1 expression levels, while RJW-58 dose-dependently reduced the activation of the Ca2+-dependent downstream effectors, NFAT1 and Rac1. The results of immunoprecipitation assays demonstrated that CTSS could bind to STIM1, which was reversed by CTSS inhibition. In addition, confocal microscopy and super-resolution imaging showed that CTSS inhibition led to STIM1 puncta accumulation in the endoplasmic reticulum and reduced the interaction between active STIM1 and EB1. In conclusion, we have demonstrated for the first time that the lysosomal cysteine protease, CTSS, plays an important role in mediating Ca2+ homeostasis by regulating STIM1 trafficking, which leads to the suppression of cell migration and invasion.

Temporal Stability of Smartphone Use Data: Determining Fundamental Time Unit and Independent Cycle.

BACKGROUND: Assessing human behaviors via smartphone for monitoring the pattern of daily behaviors has become a crucial issue in this century. Thus, a more accurate and structured methodology is needed for smartphone use research. OBJECTIVE: The study aimed to investigate the duration of data collection needed to establish a reliable pattern of use, how long a smartphone use cycle could perpetuate by assessing maximum time intervals between 2 smartphone periods, and to validate smartphone use and use/nonuse reciprocity parameters. METHODS: Using the Know Addiction database, we selected 33 participants and passively recorded their smartphone usage patterns for at least 8 weeks. We generated 4 parameters on the basis of smartphone use episodes, including total use frequency, total use duration, proactive use frequency, and proactive use duration. A total of 3 additional parameters (root mean square of successive differences, Control Index, and Similarity Index) were calculated to reflect impaired control and compulsive use. RESULTS: Our findings included (1) proactive use duration correlated with subjective smartphone addiction scores, (2) a 2-week period of data collection is required to infer a 2-month period of smartphone use, and (3) smartphone use cycles with a time gap of 4 weeks between them are highly likely independent cycles. CONCLUSIONS: This study validated temporal stability for smartphone use patterns recorded by a mobile app. The results may provide researchers an opportunity to investigate human behaviors with more structured methods.

Brain signatures associated with swallowing efficiency in older people.

Swallowing is a complex movement consisting of the sequential and orderly activation of the swallowing muscles. Neuroimaging evidence has revealed a complex cortical and subcortical representation of voluntary swallowing. The repetitive saliva swallowing test (RSST) is a convenient and simple method for assessing swallowing performance in older people. It remains unclear whether individual differences in swallowing performance are associated with variations in structural brain signatures. We aimed to investigate the association between swallowing efficiency (SWE, measured by the RSST) and gray matter volume (GMV) in healthy older adults. Forty healthy older adults (52-82 years old, 28 female) underwent T1-weighted magnetic resonance imaging (MRI) and clinical assessments of SWE, stimulated/unstimulated salivary flow rate, masticatory cycle, and walking speed. GMV was quantified using a voxel-based morphometry (VBM) approach based on the MRI data. SWE was significantly negatively correlated with age. The association between SWE and hand grip strength, but not the other clinical metrics, was statistically significant. The GMV of the left posterior cerebellum (from cerebellum crus to lobule VIII) was significantly positively correlated with SWE, as evidenced by the results of whole-brain and cerebellum-specific VBM analyses. SWE was significantly positively correlated with the cerebellar volume in the region-of-interest analyses based on automated segmentation. In healthy older adults, swallowing efficiency was positively correlated with cerebellar GMV. The findings suggested that in older people, structural variations of the brain may play a key role in individual differences in swallowing performance.

A High-Throughput Interbacterial Competition Screen Identifies ClpAP in Enhancing Recipient Susceptibility to Type VI Secretion System-Mediated Attack by Agrobacterium tumefaciens.

The type VI secretion system (T6SS) is an effector delivery system used by Gram-negative bacteria to kill other bacteria or eukaryotic hosts to gain fitness. The plant pathogen Agrobacterium tumefaciens utilizes its T6SS to kill other bacteria, such as Escherichia coli. We observed that the A. tumefaciens T6SS-dependent killing outcome differs when using different T6SS-lacking, K-12 E. coli strains as a recipient cell. Thus, we hypothesized that the A. tumefaciens T6SS killing outcome not only relies on the T6SS activity of the attacker cells but also depends on the recipient cells. Here, we developed a high-throughput interbacterial competition platform to test the hypothesis by screening for mutants with reduced killing outcomes caused by A. tumefaciens strain C58. Among the 3,909 strains in the E. coli Keio library screened, 16 mutants with less susceptibility to A. tumefaciens C58 T6SS-dependent killing were identified, and four of them were validated by complementation test. Among the four, the clpP encoding ClpP protease, which is universal and highly conserved in both prokaryotes and eukaryotic organelles, was selected for further characterizations. We demonstrated that ClpP is responsible for enhancing susceptibility to the T6SS killing. Because ClpP protease depends on other adapter proteins such as ClpA and ClpX for substrate recognition, further mutant studies followed by complementation tests were carried out to reveal that ClpP-associated AAA+ ATPase ClpA, but not ClpX, is involved in enhancing susceptibility to A. tumefaciens T6SS killing. Moreover, functional and biochemical studies of various ClpP amino acid substitution variants provided evidence that ClpA-ClpP interaction is critical in enhancing susceptibility to the T6SS killing. This study highlights the importance of recipient factors in determining the outcome of the T6SS killing and shows the universal ClpP protease as a novel recipient factor hijacked by the T6SS of A. tumefaciens.

Brain activations associated with fearful experience show common and distinct patterns between younger and older adults in the hippocampus and the amygdala.

Revisiting threat-related scenes elicits fear and activates a brain network related to cognitive-affective processing. Prior experience may contribute to the present fearful experience. We aimed to investigate (a) patterns of brain activation associated with individual differences in past fearful experiences (pFear) and the present fear elicited by watching videos (eFear) and (b) age-related differences in the activation patterns. Forty healthy adults, including 20 younger adults (YA) and 20 older adults (OA), underwent functional magnetic resonance imaging while watching videos containing high- and low-threat scenes of medical treatment. Both age subgroups showed positive correlations between pFear and bilateral hippocampal activation. Only YA showed threat-related activation in the bilateral anterior insula and activation positively correlated with pFear in the bilateral S1 and the amygdala. The evidence suggests that the hippocampus, amygdala and S1 may play key roles in bridging past fearful experiences and the present fear elicited by revisiting visual scenes and that the interaction between memory and emotional processing may be age dependent.

Functional Exploration of the Bacterial Type VI Secretion System in Mutualism: Azorhizobium caulinodans ORS571-Sesbania rostrata as a Research Model.

The bacterial type VI secretion system (T6SS) has been considered the armed force of bacteria because it can deliver toxin effectors to prokaryotic or eukaryotic cells for survival and fitness. Although many legume symbiotic rhizobacteria encode T6SS in their genome, the biological function of T6SS in these bacteria is still unclear. To elucidate this issue, we used Azorhizobium caulinodans ORS571 and its symbiotic host Sesbania rostrata as our research model. By using T6SS gene deletion mutants, we found that T6SS provides A. caulinodans with better symbiotic competitiveness when coinfected with a T6SS-lacking strain, as demonstrated by two independent T6SS-deficient mutants. Meanwhile, the symbiotic effectiveness was not affected by T6SS because the nodule phenotype, nodule size, and nodule nitrogen-fixation ability did not differ between the T6SS mutants and the wild type when infected alone. Our data also suggest that under several lab culture conditions tested, A. caulinodans showed no T6SS-dependent interbacterial competition activity. Therefore, instead of being an antihost or antibacterial weapon of the bacterium, the T6SS in A. caulinodans ORS571 seems to participate specifically in symbiosis by increasing its symbiotic competitiveness.

Zika virus structural biology and progress in vaccine development.

The growing number of zika virus (ZIKV) infections plus a 20-fold increase in neonatal microcephaly in newborns in Brazil have raised alarms in many countries regarding the threat to pregnant women. Instances of microcephaly and central nervous system malformations continue to increase in ZIKV outbreak regions. ZIKV is a small enveloped positive-strand RNA virus belonging to the Flavivirus genus of the Flaviviridae family. High-resolution ZIKV structures recently identified by cryo-electron microscopy indicate that the overall ZIKV structure is similar to those of other flaviviruses. With its compact surface, ZIKV is more thermally stable than the dengue virus (DENV). ZIKV E proteins have a characteristic "herringbone" structure with a single glycosylation site. The ZIKV E protein, the major protein involved in receptor binding and fusion, is formed as a head-to-tail dimer on the surfaces of viral particles. The E monomer consists of three distinct domains: DI, DII, and DIII. The finger-like DII contains a fusion loop (FL) that is inserted into the host cell endosomal membrane during pH-dependent conformational changes that drive fusion. Quaternary E:E dimer epitopes located at the interaction site of prM and E dimers can be further divided into two dimer epitopes. To date, more than 50 ZIKV vaccine candidates are now in various stages of research and development. Candidate ZIKV vaccines that are currently in phase I/II clinical trials include inactivated whole viruses, recombinant measles viral vector-based vaccines, DNA and mRNA vaccines, and a mosquito salivary peptide vaccine. Stabilized forms of ZIKV E:E dimer proteins have been successfully obtained either by introducing additional inter-subunit disulfide bond(s) in DII or via the direct assembly of E:E dimer proteins by immobilization with monomeric E proteins. The VLP-based approach is another alternative method for presenting native E:E dimer antigens among the vaccine components. Several forms of ZIKV VLPs have been reported featuring the co-expression of the prM-E, prM-E-NS1, C-prM-E, and NS2B/NS3 viral genes in human cells. To minimize the effect of the cross-reactive ADE-facilitating antibodies between ZIKV and DENV, several novel mutations have been reported either in or near the FL of DII or DIII to dampen the production of cross-reactive antibodies. Future ZIKV vaccine design efforts should be focused on eliciting improved neutralizing antibodies with a reduced level of cross-reactivity to confer sterilizing immunity.

Age- and sex-related differences in masseter size and its role in oral functions.

BACKGROUND: The masseter muscle plays a key structural and functional role in the stomatognathic system. Researchers' cumulative evidence has suggested that the variation in the size of a person's masseter muscle may be a critical factor related to individual differences in oral functions. However, researchers have not yet investigated systematically the effect of a person's age and sex on masseter muscle size and the association of masseter muscle size with other clinical metrics, including masticatory performance (MP) and salivary flow rate (SFR). Using T1-weighted magnetic resonance imaging (MRI) data provides a noninvasive method for assessing masseter muscle volume (MMV). METHODS: Using T1-weighted MRI data, the authors developed a voxel-based method to assess MMV and investigated the associations among MMV, MP, and SFR. RESULTS: The authors acquired T1-weighted MRI data from scans of the heads of 62 healthy adults and assessed MMV by means of using a voxel-based approach. The authors' assessment results had acceptable rates of inter-rater and intrarater reliability. MMV was significantly lower in the older subgroup and in the female subgroup. In addition, the correlation for MMV was significantly positive with MP and stimulated SFR. CONCLUSIONS: The study results revealed evidence that the authors' voxel-based approach, which they designed on the basis of T1-weighted MRI data, would be a reliable method for quantifying MMV. PRACTICAL IMPLICATIONS: The findings suggest that the variation in masseter muscle size may be a critical factor to assess individual differences in oral functions.

Age-Related Difference in Functional Brain Connectivity of Mastication.

The age-related decline in motor function is associated with changes in intrinsic brain signatures. Here, we investigated the functional connectivity (FC) associated with masticatory performance, a clinical index evaluating general masticatory function. Twenty-six older adults (OA) and 26 younger (YA) healthy adults were recruited and assessed using the masticatory performance index (MPI) and resting-state functional magnetic resonance imaging (rs-fMRI). We analyzed the rs-fMRI FC network related to mastication, which was constructed based on 12 bilateral mastication-related brain regions according to the literature. For the OA and the YA group, we identified the mastication-related hubs, i.e., the nodes for which the degree centrality (DC) was positively correlated with the MPI. For each pair of nodes, we identified the inter-nodal link for which the FC was positively correlated with the MPI. The network analysis revealed that, in the YA group, the FC between the sensorimotor cortex, the thalamus (THA) and the cerebellum was positively correlated with the MPI. Consistently, the cerebellum nodes were defined as the mastication-related hubs. In contrast, in the OA group, we found a sparser connection within the sensorimotor regions and cerebellum and a denser connection across distributed regions, including the FC between the superior parietal lobe (SPL), the anterior insula (aINS) and the dorsal anterior cingulate cortex (dACC). Compared to the YA group, the network of the OA group also comprised more mastication-related hubs, which were spatially distributed outside the sensorimotor regions, including the right SPL, the right aINS, and the bilateral dACC. In general, the findings supported the hypothesis that in OA, higher masticatory performance is associated with a widespread pattern of mastication-related hubs. Such a widespread engagement of multiple brain regions associated with the MPI may reflect an increased demand in sensorimotor integration, attentional control and monitoring for OA to maintain good mastication.