Saliva Research Articles

Salivary effector DcE1 suppresses plant defense and facilitates the successful feeding of Asian citrus psyllid, Diaphorina citri.

Piercing-sucking insects secrete diverse repertoires of effectors into their hosts to weaken host defenses and promote infestation. The Asian citrus psyllid, Diaphorina citri Kuwayama, is the most destructive insect pest in citrus orchards because of its role as a vector for the huanglongbing pathogen, Candidatus Liberibacter asiaticus (CLas). However, specific effector proteins and their functions in D. citri remain unclear. We demonstrate that DcE1, a salivary protein gene from D. citri, is predominantly expressed in the salivary gland tissues and is delivered into host plants during feeding. Transient expression in tobacco leaves revealed that DcE1 was subcellularly localized in the cytoplasm and plasma membrane, where it inhibited BAX- and INF1-induced cell death, suppressed callose deposition, and activated the salicylic acid pathway by upregulating the expression of endo-β-1,3-glucanase NtBGL2 and regulatory protein NtNPR1. Further, DcE1 knockdown by double-stranded RNA (dsRNA) injection decreased the survival rates of D. citri and interrupted D. citri phloem-feeding on host plants. These results indicate that DcE1 is a novel effector that promotes plant susceptibility and enables D. citri feeding. These findings enhance our understanding of D. citri-plant interactions and offer a potential new target gene for the development of citrus protection strategies. © 2024 Society of Chemical Industry.

Disinfection effect of ozonated water on SARS-CoV-2 in the presence of salivary proteins.

Ozonated water is expected to be an effective disinfectant for SARS-CoV-2 present on environmental fomites; however, ozone is consumed by organic substances, resulting in attenuation of its effect. SARS-CoV-2 present in saliva can contaminate environmental surfaces; therefore, it is essential to understand the effect of organic substances in saliva on the disinfectant properties of ozonated water. To assess organic factors in saliva and the extent to which they diminish the effect of ozonated water on SARS-CoV-2. Ozonated water was exposed to salivary organic factors and residual ozone concentrations were measured. SARS-CoV-2 was exposed to a salivary factor and virus inactivation by ozonated water was measured. Amylase and mucin consumed ozone in a concentration-dependent manner. Urea did not. Ozonated water appeared to inactivate SARS-CoV-2 within 30 sec. The amount of inactivated SARS-CoV-2 decreased as the protein concentration increased. Virus inactivation was stronger by 1.5 mg/L ozonated water than by 0.5 mg/L ozonated water. This study suggests that the salivary amylase and mucin decay ozone in a concentration-dependent manner, thereby attenuating the disinfection properties of ozonated water for SARS-CoV-2. An increase of the initial amount of ozone can ameliorate the disinfection effect of ozonated water on SARS-CoV-2. Ozone consumption should be taken into consideration for virus infection control. These results provide fundamental information about the effect of ozonated water when used to decontaminate surfaces harbouring SARS-CoV-2 in saliva.

Reduced Salivary Gustin and Statherin in Long-COVID Cohort with Impaired Bitter Taste

Background/Objectives: Taste dysfunction is a frequent symptom of acute coronavirus disease (COVID)-19 caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). While the majority of those affected reported recovery over time, emerging data suggest that 20–25% of individuals experience persistent taste dysfunction, constituting a common symptom of long COVID. Gustation is mediated by continuously renewing taste bud cells. A balance between the counteracting processes of cell generation and cell death maintains the homeostatic turnover. Sonic hedgehog (SHH) is a morphogenic protein that promotes taste cell proliferation and differentiation. Enzymatic proteins such as gustin modulate the environment around the taste receptors and influence taste perception. Hence, we hypothesized that increased taste cell turnover and reduced taste-related salivary proteins contribute to the taste dysfunction in long COVID. Methods: Unstimulated whole saliva (UWS) was collected from individuals with long COVID experiencing taste dysfunction after obtaining informed consent. The normal control included archived saliva samples catalogued prior to 2019. Taste perception was objectively determined by the waterless empirical taste test. The SHH, gustin, and inflammatory cytokines in UWS were determined with ELISA. The expressions of epithelial and taste-cell-specific markers in cellular saliva were assessed by immunoflurorescence. Results: Impaired bitter taste was the most common dysfunction in the long-COVID cohort. Salivary gustin was significantly lower in those with long COVID and correlated with lower bitter taste score. Cellular saliva showed keratin-10- and small-proline-rich protein-positive epithelial cells as well as SHH-, occluding- and KCNQ1-positive taste cells. Conclusions: Salivary gustin could be a marker for impaired bitter taste in long COVID.

Multi-omics approaches define novel aphid effector candidates associated with virulence and avirulence phenotypes.

Compatibility between plant parasites and their hosts is genetically determined {Citation}both interacting organisms. For example, plants may carry resistance (R) genes or deploy chemical defences. Aphid saliva contains many proteins that are secreted into host tissues. Subsets of these proteins are predicted to act as effectors, either subverting or triggering host immunity. However, associating particular effectors with virulence or avirulence outcomes presents challenges due to the combinatorial complexity. Here we use defined aphid and host genetics to test for co-segregation of expressed aphid transcripts and proteins with virulent or avirulent phenotypes. We compared virulent and avirulent pea aphid parental genotypes, and their bulk segregant F1 progeny on Medicago truncatula genotypes carrying or lacking the RAP1 (Resistance to Acyrthosiphon pisum 1) resistance quantitative trait locus. Differential gene expression analysis of whole body and head samples, in combination with proteomics of saliva and salivary glands, enabled us to pinpoint proteins associated with virulence or avirulence phenotypes. There was relatively little impact of host genotype, whereas large numbers of transcripts and proteins were differentially expressed between parental aphids, likely a reflection of their classification as divergent biotypes within the pea aphid species complex. Many fewer transcripts intersected with the equivalent differential expression patterns in the bulked F1 progeny, providing an effective filter for removing genomic background effects. Overall, there were more upregulated genes detected in the F1 avirulent dataset compared with the virulent one. Some genes were differentially expressed both in the transcriptome and in the proteome datasets, with aminopeptidase N proteins being the most frequent differentially expressed family. In addition, a substantial proportion (27%) of salivary proteins lack annotations, suggesting that many novel functions remain to be discovered. Especially when combined with tightly controlled genetics of both insect and host plant, multi-omics approaches are powerful tools for revealing and filtering candidate lists down to plausible genes for further functional analysis as putative aphid effectors.

Infiltration of salivary proteins into dentin during erosive processes

ObjectiveUltrastructural analyses showed that during erosion under oral cavity conditions, dentin is infiltrated by a substrate morphologically similar to salivary proteins. This in-situ study aimed to investigate the presence of salivary proteins in demineralized dentin. MethodsBovine dentin specimens were attached to individual maxillary splints (n = 1 per subject and condition) and worn intraorally by four subjects for 1 min. During intraoral exposure, the subjects rinsed with 10 ml of either sterile water or 1% citric acid. Additional specimens were eroded in-vitro as controls (n = 2). The specimens were then fixed, embedded in acrylic resin, and ultrathin sections were prepared. The salivary protein lysozyme was labelled with immunogold staining and examined using transmission electron microscopy. ResultsLysozyme was absent in the in-vitro controls but was detected on the dentin surface after rinsing with water, indicating its role in the initial pellicle formation on dentin. After rinsing with citric acid, lysozyme infiltrated deeper layers of the dentin. ConclusionsThese findings support the initial morphological observations that lysozyme, representing other salivary proteins, is deposited in demineralized dentin during erosion with citric acid. Clinical significanceInfiltration of salivary proteins into dentin during erosive processes can be a factor to consider in remineralization approaches under oral cavity conditions.

Characterization of potential salivary biomarkers in Behçet's disease.

Diagnosis of Behçet's disease (BD) has always been a challenging task due to the heterogeneity and complexity of the disease, while available literature on BD biomarkers is largely inconclusive. The aim of this study was to evaluate the expression of a panel of salivary biomarkers in a group of BD patients and test their association with BD. The study population comprised 23 BD patients (12 females and 11 males) and a control group of age and gender-matched healthy individuals. Enzyme-linked immunosorbent assays were used to test the expression of 5 salivary proteins including: interleukin 1β (IL-1β), epidermal growth factor (EGF), endothelin, vascular endothelial growth factor (VEGF) and transferrin (TrF). BD activity was evaluated using the International Society for Behçet's disease (ISBD) current activity form. Salivary endothelin, EGF and VEGF were significantly higher in the BD group compared to controls (16.3 Vs. 7.7pg/ml, 194.5 Vs. 163.4pg/ml and 262 Vs. 150.2pg/ml, p<0.05, respectively) whereas TrF was significantly lower in BD group compared to controls (497 Vs. 2206pg/ml, p<0.001). The correlation analysis showed a trend of a positive linear association between endothelin expression and both BD activity (r=0.41) and BD activity index score (r=0.4), without reaching statistical significance (p>0.05). In conclusion, salivary TrF, endothelin, VEGF and EGF may represent potential biomarkers for BD. These findings require further validation in larger samples and different activity states of BD.

Two mosquito salivary antigens demonstrate promise as biomarkers of recent exposure to P. falciparum infected mosquito bites.

Measuring malaria transmission intensity using the traditional entomological inoculation rate is difficult. Antibody responses to mosquito salivary proteins like SG6 have been used as biomarkers of exposure to Anopheles mosquito bites. Here, we investigate four mosquito salivary proteins as potential biomarkers of human exposure to mosquitoes infected with P. falciparum: mosGILT, SAMSP1, AgSAP, and AgTRIO. We tested population-level human immune responses in longitudinal and cross-sectional plasma from individuals with known P. falciparum infection from low and moderate transmission areas in Senegal using a multiplexed magnetic bead-based assay. AgSAP and AgTRIO were the best indicators of recent exposure to infected mosquitoes. Antibody responses to AgSAP, in a moderate endemic area, and to AgTRIO in both low and moderate endemic areas, were significantly higher than responses in a healthy non-endemic control cohort (p-values = 0.0245, 0.0064, and <0.0001 respectively). No antibody responses significantly differed between the low and moderate transmission area, or between equivalent groups during and outside the malaria transmission seasons. For AgSAP and AgTRIO, reactivity peaked 2-4 weeks after clinical P. falciparum infection and declined 3 months after infection. Reactivity to AgSAP and AgTRIO peaked after infection, with no differences between transmission seasons within region or between low and moderate transmission regions. This suggests that reactivity reflects exposure to infectious mosquitoes or recent bites rather than general mosquito exposure. Kinetics suggest reactivity is relatively short-lived. AgSAP and AgTRIO are promising candidates to incorporate into multiplexed assays for serosurveillance of population-level changes in P. falciparum-infected mosquito exposure.

Effects of combined action of proline-rich peptides of human saliva with antimicrobial proteins

BACKGROUND: The substances that make up mixed saliva exhibit activity against bacteria, viruses and fungi. Among them are various cationic antimicrobial peptides: alpha- and beta-defensins, cathelicidins, histatins, but their concentration is relatively low. On the other hand, saliva widely contains a fraction of proline-rich proteins and products of their proteolysis — peptides, the functions of which currently remain poorly studied and unclear. The study of the antimicrobial activity of proline-rich proteins, in particular, when they act together with antimicrobial peptides, and the deciphering of the molecular and cellular basis for the implementation of the protective functions of proline-rich peptides is an urgent task in medicine and biology. The data obtained will help to understand the mechanisms of anti-infective protection of the oral cavity with the participation of the innate immune system, and in the future will allow the creation of drugs based on proline-rich proteins in humans and animals. AIM: Study of the individual antibacterial action of proline-rich salivary proteins, their combined action with the antimicrobial proteins lactoferrin and lysozyme, as well as with one of the most significant endogenous protective peptides — cathelicidin LL-37 on the formation of bacterial biofilms. MATERIALS AND METHODS: Human lactoferrin and egg lysozyme were used in the work. The minimum inhibitory concentrations of peptides against gram-negative and gram-positive bacteria were estimated using serial dilutions in a liquid nutrient medium. Using serial dilutions using the “chessboard” method, it was shown that proline-rich proteins can increase the antibacterial activity of innate immunity peptides present in the oral cavity. The ability of combinations of cationic proline-rich peptides with cathelicidin LL-37 to prevent the formation of bacterial biofilms was studied. RESULTS: In the presence of proline-rich peptides, the antimicrobial activity of some antimicrobial peptides and proteins present in saliva against planktonic bacteria increases. The combined action of proline-rich proteins with LL-37 increases the inhibition of bacterial biofilm formation. CONCLUSIONS: As a result of the study, it was shown that the analyzed cationic proline-rich peptides of human saliva exhibit anti-inflammatory effects, which suggests their significant role in the regulation of inflammatory processes in the oral cavity, as well as their participation in recovery processes.

The Remineralization of Enamel from Saliva: A Chemical Perspective

The natural remineralization of enamel is of major importance for oral health. In principle, early erosions (demineralization) induced by acidic beverages and foods as well as initial caries lesions can be covered and remineralized by the deposition of calcium phosphate, i.e., tooth mineral. This remineralization effect is characterized by the presence of calcium and phosphate ions in saliva that form hydroxyapatite on the enamel surface. Although it is apparently a simple crystallization, it turns out that remineralization under in vivo conditions is actually a very complex process. Calcium phosphate can form a number of solid phases of which hydroxyapatite is only one. Precipitation involves the formation of metastable phases like amorphous calcium phosphate that convert into biological apatite in a number of steps. Nanoscopic clusters of calcium phosphate that can attach on the enamel surface are also present in saliva. Thus, remineralization under strictly controlled in vitro conditions (e.g., pH, ion concentrations, no additives) is already complex, but it becomes even more complicated under the actual conditions in the oral cavity. Here, biomolecules are present in saliva, which interact with the forming calcium phosphate mineral. For instance, there are salivary proteins which have the function of inhibiting crystallization to avoid overshooting remineralization. Finally, the presence of bacteria and an extracellular matrix in plaque and the presence of proteins in the pellicle have strong influences on the precipitation on the enamel surface. The current knowledge on the remineralization of the enamel is reviewed from a chemical perspective with a special focus on the underlying crystallization phenomena and the effects of biological compounds that are present in saliva, pellicle, and plaque. Basically, the remineralization of enamel follows the same principles as calculus formation. Notably, both processes are far too complex to be understood on a microscopic basis under in vivo conditions, given the complicated process of mineral formation in the presence of a plethora of foreign ions and biomolecules.

Prediction of Oral Cancer Biomarkers by Salivary Proteomics Data.

Oral cancer, representing 2-4% of all cancer cases, predominantly consists of Oral Squamous Cell Carcinoma (OSCC), which makes up 90% of oral malignancies. Early detection of OSCC is crucial, and identifying specific proteins in saliva as biomarkers could greatly improve early diagnosis. Here, we proposed a strategy to pinpoint candidate biomarkers. Starting from a list of salivary proteins detected in 10 OSCC patients and 20 healthy controls, we combined a univariate approach and a multivariate approach to select candidates. To reduce the number of proteins selected, a Protein-Protein Interaction network was built to consider only connected proteins. Then, an over-representation analysis (ORA) determined the enriched pathways. The network from 172 differentially abundant proteins highlighted 50 physically connected proteins, selecting relevant candidates for targeted experimental validations. Notably, proteins like Heat shock 70 kDa protein 1A/1B, Pyruvate kinase PKM, and Phosphoglycerate kinase 1 were suggested to be differentially regulated in OSCC patients, with implications for oral carcinogenesis and tumor growth. Additionally, the ORA revealed enrichment in immune system, complement, and coagulation pathways, all known to play roles in tumorigenesis and cancer progression. The employed method has successfully identified potential biomarkers for early diagnosis of OSCC using an accessible body fluid.

Ultrasound and its coupled oak blocks treatment based on absorbed energy density for comprehensive insight and scale-up consideration of phenolic and astringency profiles in red wines

Ultrasound and ultrasound-coupled oak blocks techniques on the phenolic and astringency profiles of Petit Verdot wines were discussed in this study. The relationship between techniques at varying absorbed energy density (AED) gradients and astringency was revealed, elucidating potential molecular mechanisms regarding compound interactions. Ultrasound was found to promote phenolics degradation and condensed tannins maturation, while oak blocks improved ellagitannins release into wines, facilitating polycondensation reactions to form larger complexes. Phenolics binding to salivary proteins decreased at low AED values (0–33.07 J/mL), enhanced at high AED values (66.14–165.34 J/mL), and reduced salivary proteins' precipitation ability. Treated samples significantly altered astringency's global terminology and sensory attributes across oral regions, with oak blocks enriching the astringency hierarchy. Multivariate analysis identified an optimal ultrasound AED of 0 to 33.07 J/mL, and an effective PLS-DA model was developed for industrial scale-up considerations, aiming to cost-effectively enhance the organoleptic quality of wines.

Co-spray drying whey protein isolate with polysaccharides provides additional lubrication impacting the sensory profile of model beverages

For older adults, whey protein is associated with negative sensory attributes, including undesirable mouthfeel qualities, that limit consumption and acceptance. Previous work suggests that increased lubrication may have the ability to reduce whey protein-associated mouthdrying, a driver for disliking, by limiting interactions of whey protein with salivary proteins and mucin. This was investigated in the current study by co-spray drying whey protein with combinations of maltodextrin, xanthan gum and/or guar gum; the resulting powders were used to make 10% suspensions. The particle size, zeta potential, rheological and tribological profiles of these suspensions were measured. It was shown that co-spray drying whey protein with guar gum led to a reduction in instrumental friction, irrespective of changes in viscosity. These samples were perceived as significantly more mouthcoating and smooth when assessed by a trained sensory panel. Contrastingly, suspensions containing xanthan gum showed increased viscosity and enhanced shear thinning compared with whey protein, but no change in instrumental friction at higher sliding speeds. This may be a result of a larger particle size, representing increased aggregation in samples containing xanthan gum. There was no significant difference in mouthdrying or slipperiness perception between the suspensions. These findings suggest that the incorporation of guar gum has the capacity to reduce oral friction and impact mouthfeel in whey protein model beverages. This should be taken forward into temporal sensory trials to further investigate the effects of additional lubrication.

Explorative study of stimulated saliva proteome in head and neck cancer patients pre- and post-treatment

Objectivesto compare saliva proteome of patients before treatment of head and neck cancer and six months post-treatment with controls. DesignFive dentate patients and five age and gender-matched controls were included. The stimulated salivary secretion rate was determined, and saliva was stored at −80 °C. After thawing, 30 mg of each sample and a reference (aliqouts of all samples) was trypsin digested. The digested peptides were analyzed by mass spectrometry. The relative abundances were transformed to log2 and significant differences determined. Relative abundances of mucins were compared with patient's problems with dry mouth, sticky saliva and swallowing. Data are available via ProteomeXchange with identifier PXD047500. Results966 proteins with ≥2 unique peptides were found. Compared with controls, 30 proteins were found in significantly lower relative abundances and 65 in higher at pre-treatment and 38 proteins in significantly lower relative abundances and 34 proteins in higher post-treatment. Regarding proteins from the salivary glands, a significantly lower relative abundance of Cystatins was detected pre-treatment and significantly lower relative abundances of Cystatin, Cysteine-rich secretory protein 3, Lactoperoxidase, Prolactin-inducible protein and Proline-rich protein 4 post-treatment. No clear relation between relative abundance of mucins and dry mouth, sticky saliva and problems with swallowing was detected. ConclusionDecreases in several salivary gland proteins post cancer treatment might lead to a reduced defense against oral disorders. Knowledge about changes in saliva proteins in connection with oral cancer treatment is important for planning dental care for these patients.

Salivary cysteine levels as a potential biochemical indicator of oral cancer risk in tobacco consumers.

Aim: Oral cancer is the leading cause of mortality, with a survival rate of less than 5 years, and is predominantly influenced by tobacco mutagens. Invasive diagnostic methods hinder early detection of oral cancer biomarkers. The present study performed salivary biochemical analysis for early oral cancer screening in tobacco consumers.Materials & methods: Three study groups included healthy controls (n=25), tobacco users (n=25) and oral cancer patients (n=25). Salivary total protein, amylase, TNF-α and amino acid levels were evaluated using enzymatic tests, Enzyme linked Immunosorbent Assay (ELISA) and High-Performance Liquid Chromatography (HPLC).Results: Compared with healthy controls, salivary total protein and TNF-αlevels were significantly (p=0.04) higher in oral cancer patients. Salivary amylase levels were significantly lower in tobacco smokers (p=0.02) and higher in oral cancer patients (p=0.01). Interestingly, the amino acid cysteine concentration was significantly higher (p=0.02) in tobacco consumers (62.5±10) than in healthy controls (116.1±28).Conclusion: In high-risk populations, such as tobacco users, salivary biochemical analysis can serve as a promising noninvasive diagnostic method for early oral cancer screening. As a salivary biomarker, the amino acid cysteine exhibits potential as a means of detecting the progression of oral cancer in individuals who consume tobacco.

Exploring the Relationship Between Stress, Salivary C-Reactive Protein, and Embodied Physiological Responses in a Nigerian Population.

The impacts of stress on inflammation, although hypothesized, have not been thoroughly examined, especially in relation to social and environmental factors and particularly within Black populations. This study aims to explore the biological mechanisms of embodiment linking stress and health to understand physiological changes in the body's response to psychological stress in a Nigerian population. Through a multidisciplinary approach, this study queries the relationship between stress, cortisol, and salivary C-reactive protein (sCRP), a biomarker of inflammation, while also validating the use of sCRP as a potential and accurate stress indicator in the field. In this cross-sectional study, 138 passive drool saliva samples (nfemale = 89 nmale = 49) were collected and assessed for sCRP and cortisol levels in adults. Participants also completed a short demographic survey and, to measure psychological stress, the General Health Questionnaire 12 (GHQ-12). Relationships between sCRP and stress-related variables (i.e., cortisol, GHQ-12, and demographic data) were assessed using Spearman's correlations, simple regression, multivariable linear regression, and exploratory factor analysis. sCRP levels ranged from 20.57 to 6879.41 pg/mL across all samples, with significant differences between female and male participants. The GHQ-12 was not a significant predictor of sCRP variability. However, socio-demographic factors such as body mass index (BMI), age, self-reported sex, ethnic identity, and cortisol were significant predictors, collectively explaining 24%-27% of the variation in sCRP. Socio-demographic predictors like BMI, age, sex, and particularly ethnic group experience in Nigeria encapsulate aspects of embodied stress, that significantly affect sCRP variability.

Oral SARS-CoV-2 host responses predict the early COVID-19 disease course

Oral fluids provide ready detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and host responses. This study sought to evaluate relationships between oral virus, oral and systemic anti-SARS-CoV-2-specific antibodies, and symptoms. Oral fluids (saliva/throat wash (saliva/TW)) and serum were collected from asymptomatic and symptomatic, nasopharyngeal (NP) SARS-CoV-2 RT-qPCR+ human participants (n = 45). SARS-CoV-2 RT-qPCR and N-antigen detection by immunoblot and lateral flow assay (LFA) were performed. RT-qPCR for subgenomic RNA (sgRNA) was sequence confirmed. SARS-CoV-2-anti-S protein RBD LFA and ELISA assessed IgM and IgG responses. Structural analysis identified host salivary molecules analogous to SARS-CoV-2-N-antigen. At time of enrollment (baseline, BL), LFA-detected N-antigen in 86% of TW and was immunoblot-confirmed. However, only 3/17 were saliva/TW qPCR+ . Sixty percent of saliva and 83% of TW demonstrated persistent N-antigen at 4 weeks. N-antigen LFA signal in three anti-spike sero-negative participants suggested potential cross-detection of 4 structurally analogous salivary RNA binding proteins (alignment 19–29aa, RMSD 1–1.5 Angstroms). At enrollment, symptomatic participants demonstrated replication-associated sgRNA junctions, were IgG+ (94%/100% in saliva/TW), and IgM+ (63%/54%). At 4 weeks, SARS-CoV-2 IgG (100%/83%) and IgM (80%/67%) persisted. Oral and serum IgG correlated 100% with NP+ PCR status. Cough and fatigue severity (p = 0.010 and 0.018 respectively), and presence of weakness, nausea, and composite upper respiratory symptoms (p = 0.037, 0.005, and 0.017, respectively) were negatively associated with saliva IgM but not TW or serum IgM. Throat wash IgM levels were higher in women compared to men, although the association did not reach statistical significance (median: 290 (female) versus 0.697, p = 0.056). Important to transmission and disease course, oral viral replication and persistence showed clear relationships with select symptoms and early oral IgM responses during early infection. N-antigen cross-reactivity may reflect mimicry of structurally analogous host proteins.

Mosquito salivary apyrase regulates blood meal hemostasis and facilitates malaria parasite transmission

The evolution of hematophagy involves a series of adaptations that allow blood-feeding insects to access and consume blood efficiently while managing and circumventing the host’s hemostatic and immune responses. Mosquito, and other insects, utilize salivary proteins to regulate these responses at the bite site during and after blood feeding. We investigated the function of Anopheles gambiae salivary apyrase (AgApyrase) in regulating hemostasis in the mosquito blood meal and in Plasmodium transmission. Our results demonstrate that salivary apyrase, a known inhibitor of platelet aggregation, interacts with and activates tissue plasminogen activator, facilitating the conversion of plasminogen to plasmin, a human protease that degrades fibrin and facilitates Plasmodium transmission. We show that mosquitoes ingest a substantial amount of apyrase during blood feeding, which reduces coagulation in the blood meal by enhancing fibrin degradation and inhibiting platelet aggregation. AgApyrase significantly enhanced Plasmodium infection in the mosquito midgut, whereas AgApyrase immunization inhibited Plasmodium mosquito infection and sporozoite transmission. This study highlights a pivotal role for mosquito salivary apyrase for regulation of hemostasis in the mosquito blood meal and for Plasmodium transmission to mosquitoes and to the mammalian host, underscoring the potential for strategies to prevent malaria transmission.

First Insight into Lignin Valorization as a Promising Biopolymer for the Modulation of the Physicochemical Properties of Port Wine.

Lignosulfonate (LS), kraft lignin (KL), and organosolv lignin (OL) were evaluated as potential modulating agents of the physicochemical properties of Port wine at two different concentrations for 7 and 30 days. KL and LS demonstrated the ability to remove proteins and potentiate the anthocyanin concentration. LS reduced the tannin content and the interaction of salivary acidic proline-rich proteins with wine phenolic compounds. None of the lignin promoted a perceptible color change; however, the yellowish color of KL and OL at 100 g/hL contributed to an increase in the yellow tones of wines. Lignin improved wine aroma by reducing the amount of unwanted volatiles by 30% and increasing the content of ethyl esters associated with fruity aromas by up to 60%. The results suggest that lignin, especially LS, can be employed as a modulating agent, positively impacting wine's physicochemical properties. This valorization of a byproduct opens up new opportunities for the wine industry.

Searching for Protein Off-Targets of Prostate-Specific Membrane Antigen-Targeting Radioligands in the Salivary Glands.

Background: Prostate specific membrane antigen (PSMA)-targeted radioligand therapies represent a highly effective treatment for metastatic prostate cancer. However, high and sustain uptake of PSMA-ligands in the salivary glands led to dose limiting dry mouth (xerostomia), especially with α-emitters. The expression of PSMA and histologic analysis couldn't directly explain the toxicity, suggesting a potential off-target mediator for uptake. In this study, we set out to search for possible off-target non-PSMA protein(s) in the salivary glands. Methods: A machine-learning based quantitative structure activity relationship (QSAR) model was built for seeking the possible off-target(s). The resulting target candidates from the model prediction were subjected to further analysis for salivary protein expression and structural homology at key regions required for PSMA-ligand binding. Furthermore, cellular binding assays were performed utilizing multiple cell lines with high expression of the candidate proteins and low expression of PSMA. Finally, PSMA knockout (PSMA-/-) mice were scanned by small animal PET/MR using [68Ga]Ga-PSMA-11 for in-vivo validation. Results: The screening of the trained QSAR model did not yield a solid off-target protein, which was corroborated in part by cellular binding assays. Imaging using PSMA-/- mice further demonstrated markedly reduced PSMA-radioligand uptake in the salivary glands. Conclusion: Uptake of the PSMA-targeted radioligands in the salivary glands remains primarily PSMA-mediated. Further investigations are needed to illustrate a seemingly different process of uptake and retention in the salivary glands than that in prostate cancer.

De Novo Transcriptome Profiling of Mustard Aphid (Lipaphis erysimi) and Differential Expression of Transcripts Associated with Feeding and Non-Feeding Conditions and Developmental Stages.

Lipaphis erysimi is a specialist aphid of the Indian subcontinent that causes significant yield losses in oilseed Brassicas. Several aphid genes have been used as preferred targets in RNAi-based transgenic plants for aphid resistance. In order to enhance the repertoire of potential target genes for aphid control and to identify the genes associated with aphid feeding and development, we performed a two-way comparative study of differential gene expression profiles between (i) feeding and non-feeding adults and (ii) adult and nymph developmental stages of L. erysimi. De novo RNA-seq of aphids using Illumina technology generated a final transcriptome comprising 52,652 transcripts. Potential transcripts for host selection, detoxification, salivary proteins and effectors, molecular chaperones and developmental genes were identified. Differential gene expression studies identified variations in the expression of 1502 transcripts between feeding and non-feeding adults and 906 transcripts between nymphs and adults. These data were used to identify novel target genes for RNAi-based aphid control and facilitate further studies on the molecular basis of aphid feeding and development.