Association of Root Biofilm Bacteriome with Root Caries Lesion Severity and Activity.

INTRODUCTION: This research aimed to assess the association of root biofilm bacteriome with root caries lesion severity and activity in institutionalised Colombian elderlies and was conducted to gather data on the root caries bacteriome in this population. METHODS: A bacteriome evaluation of biofilm samples from sound and carious root surfaces was performed. Root caries was categorised (ICDAS Root criteria) based on severity (sound surfaces, initial: non-cavitated, moderate/extensive combined: cavitated) and activity status (active and inactive). DNA was extracted and the V4 region of the 16S rRNA gene was sequenced; afterwards the classification of features was conducted employing amplicon sequence variants and taxonomic assignment via the Human Oral Microbiome Database (HOMD). Bacterial richness, diversity (Simpson's and Shannon's indices), and relative abundance estimation were assessed and compared based on root caries severity and activity status (including Sound surfaces). RESULTS: A total of 130 biofilm samples were examined: sound (n = 45) and with root caries lesions (n = 85; by severity: initial: n = 41; moderate/extensive: n = 44; by activity: active: n = 60; inactive: n = 25). Species richness was significantly lower in biofilms from moderate/extensive and active groups compared to sound sites. There was a higher relative abundance of species like Lechtotricia wadei, Capnocytophaga granulosa, Cardiobacterium valvarum, Porphyromonas pasteri - in sound sites; Dialister invisus, Streptococcus mutans, Pseudoramibacter alactolyticus and Bacteroidetes (G-5) bacterium 511 - in moderate/extensive lesions, and Fusobacterium nucleatum subsp. animalis, Prevotella denticola, Lactobacillus fermentum, Saccharibacteria (TM7) (G-5)bacterium HMT 356 - in active lesions. CONCLUSION: Root caries bacteriome exhibited differences in species proportions between the compared groups. Specifically, cavitated caries lesions and active caries lesions showed higher relative abundance of acidogenic bacteria.

Effect of chitosan irrigant solutions on the release of bioactive proteins from root dentin.

OBJECTIVE: To identify the effect of two chitosan solutions on the release of root dentin matrix proteins and to describe the chemical changes observed following conditioning with chelating agents. MATERIALS AND METHODS: The release of dentin sialoprotein (DSP), transforming growth factor-beta 1 (TGF-ß1), vascular endothelial growth factor (VEGF), and platelet-derived growth factor-BB (PDGF-BB) with different chelating agents, including ethylenediaminetetraacetic acid (EDTA), chitosan solution (CS), and nanoparticulate chitosan (CSnp), was investigated. DSP was quantified using an enzyme-linked immunosorbent assay (ELISA). TGF-ß1, VEGF, and PDGF-BB were quantified using a cytokine bead panel (CBA). Raman spectroscopy was performed to identify surface chemical changes. Statistical analysis was performed using Kruskal-Wallis test with Mann-Whitney-Wilcoxon rank-sum test (p < 0.05). RESULTS: TGF-ß1, VEGF, and DSP solubilized in all irrigants tested. CSnp showed the highest concentration of DSP. PDGF-BB did not exceed the detection limits. Raman spectroscopy revealed a decrease in the phosphate and carbonate peaks, representing the chelating effect of EDTA, CS, and CSnp. Additionally, CSnp showed the greatest preservation of the amide I and III content. CONCLUSION: Proteins can be released from dentin via EDTA, CS, and CSnp conditioning. Raman spectroscopic revealed changes in the inorganic content of the root dentin after chelation. Furthermore, use of CSnp facilitated a preservation of the organic content. CLINICAL RELEVANCE: Chelation allows the release of proteins, justifying the use of chelating agents in regenerative endodontics. The chitosan-dentin matrix interaction also promotes the protection of the organic content as an additional benefit to its protein releasing effect.

Tracing ΦX174 bacteriophage spreading during aerosol-generating procedures in a dental clinic.

OBJECTIVE: The aim of this study was to test the plausibility of using the ΦX174 bacteriophage as a tracer of viral aerosols spreading in a dental aerosol-generating procedure (AGP) model. METHODS: ΦX174 bacteriophage (~ 108 plaque-forming units (PFU)/mL) was added into instrument irrigation reservoirs and aerosolized during class-IV cavity preparations followed by composite fillings on natural upper-anterior teeth (n = 3) in a phantom head. Droplets/aerosols were sampled through a passive approach that consisted of Escherichia coli strain C600 cultures immersed in a LB top agar layer in Petri dishes (PDs) in a double-layer technique. In addition, an active approach consisted of E coli C600 on PDs sets mounted in a six-stage cascade Andersen impactor (AI) (simulating human inhalation). The AI was located at 30 cm from the mannequin during AGP and afterwards at 1.5 m. After collection PDs were incubated overnight (18 h at 37 °C) and bacterial lysis was quantified. RESULTS: The passive approach disclosed PFUs mainly concentrated over the dental practitioner, on the mannequin's chest and shoulder and up to 90 cm apart, facing the opposite side of the AGP's source (around the spittoon). The maximum aerosol spreading distance was 1.5 m in front of the mannequin's mouth. The active approach disclosed collection of PFUs corresponding to stages (and aerodynamic diameters) 5 (1.1-2.1 µm) and 6 (0.65-1.1 µm), mimicking access to the lower respiratory airways. CONCLUSION: The ΦX174 bacteriophage can be used as a traceable viral surrogate in simulated studies contributing to understand dental bioaerosol's behavior, its spreading, and its potential threat for upper and lower respiratory tract. CLINICAL RELEVANCE: The probability to find infectious virus during AGPs is high. This suggests the need to continue characterizing the spreading viral agents in different clinical settings through combination of passive and active approaches. In addition, subsequent identification and implementation of virus-related mitigation strategies is relevant to avoid occupational virus infections.

Purification of RgpA from external outer membrane vesicles of Porphyromonas gingivalis.

INTRODUCTION: Purification of native gingipains is challenging because these proteases are frequently associated with the cell surface, which affects yield. This study aimed to purify native Arg-gingipain (RgpA) from Porphyromonas gingivalis Outer Membrane Vesicles (OMV). METHODS: Native RgpA was purified from P. gingivalis strain ATCC33277 OMV using a strategy including ultracentrifugation, sonication, and successive anionic and cationic fast protein liquid chromatography (FPLC). The presence and purity of the protease were confirmed by SDS-PAGE and detection of protease activity using fluorogenic substrates. Rat antibodies produced against the unique adhesin hemagglutinin (H1) domain of RgpA (amino acids 719-865) were titrated by ELISA at a 1:100 dilution using whole P. gingivalis lysate as an antigen and western blotting to detect a 75 kDa band corresponding to RgpA. RESULTS: Double anionic-cationic FLPC yielded prominent peaks with evident amidolytic gingipain activity of the appropriate molecular weight, as confirmed by western blotting. The final RgpA yield from 1 L of bacterial culture with colony forming unit (CFU) (Log10) 7.4 ± 0.08/mL was of 12.6% (2 mg/mL), with 3.2 FU/µg of amidolytic activity. CONCLUSIONS: This protocol allows purification of native RgpA from OMV that retains protease activity.

Human Metapneumovirus: Laboratory Methods for Isolation, Propagation, and Plaque Titration.

The human metapneumovirus (hMPV) is an important viral agent associated with severe infections of the upper and lower airways, especially in young children and immunosuppressed subjects. Nevertheless, in vitro studies of hMPV are very difficult due to the little knowledge we have on its laboratory manipulation. OBJECTIVE: The aim of this study was to isolate and propagate hMPV from patients, and to establish a method to quantify the virus by plaque assay. METHOD: As part of a Latin American respiratory virus surveillance study, 12 nasal secretion samples - hMPV-positive by direct fluorescence - were inoculated on LLC-MK2 cells to isolate the virus. The supernatants were re-inoculated and the cytopathic effect and syncytium formation were evaluated daily; the infection was confirmed by immunofluorescence and RT-PCR. A protocol to titrate the harvested virus was established inoculating serial dilutions on LLC-MK2 cells, and agarose was then added as an overlay. After different time periods, the monolayers were fixed and stained with Naphthol blue/black or crystal violet and finally the viral titer was obtained. RESULTS: Eight out of 12 hMPV-positive respiratory samples were positive for the isolation and confirmed by RT-PCR and immunofluorescence, but the cytopathic effect and syncytium formation were observed only in 5 cultures. One out of 8 viral isolates was used for propagation and plaque assay standardization. We found that incubation for 7 days in the semisolid overlay yielded plaques with appropriate size and shape to be counted, although crystal violet staining showed slightly larger plaques than those seen with Naphthol blue/black staining. CONCLUSIONS: The isolation and propagation from patient-derived hMPV and the standardization of a practical, reliable, and inexpensive method of detection and quantification of hMPV were carried out, without the additional use of antibodies that had not been reported previously. These results offer some important insights for future studies of cellular and molecular biology of hMPV.

A prospective cohort study to assess seroprevalence, incidence, knowledge, attitudes and practices, willingness to pay for vaccine and related risk factors in dengue in a high incidence setting.

BACKGROUND: Dengue is one of the most important vector-borne diseases in the world, causing significant morbidity and economic impact. In Colombia, dengue is a major public health problem. Departments of La Guajira, Cesar and Magdalena are dengue endemic areas. The objective of this research is to determine the seroprevalence and the incidence of dengue virus infection in the participating municipalities from these Departments, and also establish the association between individual and housing factors and vector indices with seroprevalence and incidence. We will also assess knowledge, attitudes and practices, and willingness-to-pay for dengue vaccine. METHODS: A cohort study will be assembled with a clustered multistage sampling in 11 endemic municipalities. Approximately 1000 homes will be visited to enroll people older than one year who living in these areas, who will be followed for 1 year. Dengue virus infections will be evaluated using IgG indirect ELISA and IgM and IgG capture ELISA. Additionally, vector indices will be measured, and adult mosquitoes will be captured with aspirators. Ovitraps will be used for continuous estimation of vector density. DISCUSSION: This research will generate necessary knowledge to design and implement strategies with a multidimensional approach that reduce dengue morbidity and mortality in La Guajira and other departments from Colombian Caribbean.

Detection of all four dengue serotypes in Aedes aegypti female mosquitoes collected in a rural area in Colombia.

The Aedes aegypti vector for dengue virus (DENV) has been reported in urban and periurban areas. The information about DENV circulation in mosquitoes in Colombian rural areas is limited, so we aimed to evaluate the presence of DENV in Ae. aegypti females caught in rural locations of two Colombian municipalities, Anapoima and La Mesa. Mosquitoes from 497 rural households in 44 different rural settlements were collected. Pools of about 20 Ae. aegypti females were processed for DENV serotype detection. DENV in mosquitoes was detected in 74% of the analysed settlements with a pool positivity rate of 62%. The estimated individual mosquito infection rate was 4.12% and the minimum infection rate was 33.3/1,000 mosquitoes. All four serotypes were detected; the most frequent being DENV-2 (50%) and DENV-1 (35%). Two-three serotypes were detected simultaneously in separate pools. This is the first report on the co-occurrence of natural DENV infection of mosquitoes in Colombian rural areas. The findings are important for understanding dengue transmission and planning control strategies. A potential latent virus reservoir in rural areas could spill over to urban areas during population movements. Detecting DENV in wild-caught adult mosquitoes should be included in the development of dengue epidemic forecasting models.

Identification of proteins from human permanent erupted enamel.

Proteins from the extracellular matrix of enamel are highly specific and necessary for proper enamel formation. Most proteins are removed from the matrix by enamel proteases before complete mineralization is achieved; however, some residual protein fragments persist in the mineralized matrix of erupted enamel. So far, only amelogenin peptides obtained by traditional bottom-up proteomics have been recovered and identified in human permanent erupted enamel. In this study, we hypothesize that other enamel-specific proteins are also found in human permanent enamel, by analysing human erupted third molars. Pulverized enamel was used to extract proteins, and the protein extract was subjected directly to liquid-chromatography coupled to tandem mass spectrometry (LC-MS/MS) without a previous trypsin-digestion step. Amelogenin and non-amelogenin proteins (ameloblastin and enamelin) were succesfully identified. The sequences of the naturally occurring peptides of these proteins are reported, finding in particular that most of the peptides from the amelogenin X-isoform come from the tyrosine-rich amelogenin peptide (TRAP) and that some were identified in all specimens. In conclusion, our LC-MS/MS method without trypsin digestion increased the coverage of identification of the enamel proteome from a few amelogenin peptides to a higher number of peptides from three enamel-specific proteins.

Modulation of TRPV1 on Odontoblast-like Cells Using Capsazepine-Loaded Nanogels.

The modulation of TRPV1 emerges as a promising strategy for dental pain management. This study aimed to assess TRPV1 modulation in a human odontoblast-like cell model using Capsazepine (CZP) loaded in a nanogel delivery system. Gelatin nanogels, synthesized via the emulsification-gelation technique, were characterized and loaded with the TRPV1 antagonist, CZP. HPLC determined a remarkable 67.5 ± 0.04% CZP loading efficiency, with 71.7% of nanogels falling within the 300-950 nm size range, as evidenced by light microscopy. Moreover, CZP-loaded nanogels had a low cytotoxicity. An FTIR analysis showed no adverse chemical interactions, ensuring stability and active release. When examining biological responses, TRPV1 expression and channel activity were assessed in odontoblast-like cells. On the fifth day post-treatment, cells treated with CZP-loaded nanogels exhibited an increased TRPV1 expression and a reduction in calcium fluxes after agonist stimulus (F/F0 ratio 1.18 ± 0.18), resembling the response in free CZP-treated cells (1.28 ± 0.15). A two-way analysis of variance and the Tukey's test were used to determine statistical significance (p < 0.05). This delivery system, proven to be economical and straightforward, holds promise for dental pain management and potential local use. Local administration minimizes systemic adverse effects, making it a practical solution for releasing molecules in the oral cavity.

Zika Virus: A Neurotropic Warrior against High-Grade Gliomas-Unveiling Its Potential for Oncolytic Virotherapy.

Gliomas account for approximately 75-80% of all malignant primary tumors in the central nervous system (CNS), with glioblastoma multiforme (GBM) considered the deadliest. Despite aggressive treatment involving a combination of chemotherapy, radiotherapy, and surgical intervention, patients with GBM have limited survival rates of 2 to 5 years, accompanied by a significant decline in their quality of life. In recent years, novel management strategies have emerged, such as immunotherapy, which includes the development of vaccines or T cells with chimeric antigen receptors, and oncolytic virotherapy (OVT), wherein wild type (WT) or genetically modified viruses are utilized to selectively lyse tumor cells. In vitro and in vivo studies have shown that the Zika virus (ZIKV) can infect glioma cells and induce a robust oncolytic activity. Consequently, interest in exploring this virus as a potential oncolytic virus (OV) for high-grade gliomas has surged. Given that ZIKV actively circulates in Colombia, evaluating its neurotropic and oncolytic capabilities holds considerable national and international importance, as it may emerge as an alternative for treating highly complex gliomas. Therefore, this literature review outlines the generalities of GBM, the factors determining ZIKV's specific tropism for nervous tissue, and its oncolytic capacity. Additionally, we briefly present the progress in preclinical studies supporting the use of ZIKV as an OVT for gliomas.

Development and Optimization of a Multiplex Real-Time RT-PCR to Detect SARS-CoV-2 in Human Samples.

PCR and its variants (RT-PCR and qRT-PCR) are valuable and innovative molecular techniques for studying nucleic acids. qPCR has proven to be highly sensitive, efficient, and reproducible, generating reliable results that are easy to analyze. During the COVID-19 pandemic, qPCR became the gold standard technique for detecting the SARS-CoV-2 virus that allowed to confirm the infection event, and those asymptomatic ones, and thus save millions of lives. In-house multiplex qPCR tests were developed worldwide to detect different viral targets and ensure results, follow the infections, and favor the containment of a pandemic. Here, we present the detailed fundamentals of the qPCR technique based on fluorogenic probes and processes to develop and optimize a successful multiplex RT-qPCR test for detecting SARS-CoV-2 that could be used to diagnose COVID-19 accurately.

In vivo differential susceptibility of sensory neurons to rabies virus infection.

There is controversy with regard to the entry pathway of the rabies virus (RABV) into the central nervous system (CNS). Some authors have suggested that the virus inoculated at the periphery is captured and transported to CNS only by motor neurons; however, it has been reported that dorsal root ganglia (DRG) sensory neurons capture and transport the virus to the spinal cord (SC) and then to the brain. It is probable that preferences for one pathway or another depend on the site of inoculation and the post-infection time. Therefore, in the present study, we evaluated different vertebral segments and post-infection times, along with the location, number, and subpopulation of sensory neurons susceptible to infection after inoculating RABV in the footpads of adult mice. It was noted that the virus inoculated in the footpad preferentially entered the CNS through the large-sized DRG sensory neurons, while infection of the motor neurons occurred later. Further, it was found that the virus was dispersed in spinal cord trans-synaptically through the interneurons, arriving at both sensory neurons and contralateral motor neurons. In conclusion, we observed that RABV inoculated in the plantar footpad is captured preferentially by large sensory neurons and is transported to the DRG, where it replicates and is spread to the SC using transynaptic jumps, infecting sensory and motor neurons at the same level before ascending to the brain.

Deubiquitinating Enzyme Inhibitors Block Chikungunya Virus Replication.

Ubiquitination and deubiquitination processes are widely involved in modulating the function, activity, localization, and stability of multiple cellular proteins regulating almost every aspect of cellular function. Several virus families have been shown to exploit the cellular ubiquitin-conjugating system to achieve a productive infection: enter the cell, promote genome replication, or assemble and release viral progeny. In this study, we analyzed the role of deubiquitinating enzymes (DUBs) during chikungunya virus (CHIKV) infection. HEK293T, Vero-E6, and Huh-7 cells were treated with two DUB inhibitors (PR619 or WP1130). Then, infected cells were evaluated by flow cytometry, and viral progeny was quantified using the plaque assay method. The changes in viral proteins and viral RNA were analyzed using Western blotting and RT-qPCR, respectively. Results indicate that treatment with DUB inhibitors impairs CHIKV replication due to significant protein and viral RNA synthesis deregulation. Therefore, DUB activity may be a pharmacological target for blocking CHIKV infection.

Capsazepine antagonizes TRPV1 activation induced by thermal and osmotic stimuli in human odontoblast-like cells.

Objectives: Dental pain, which is the main reason for patients consulting dentists, is classified as a public health concern. The study of cellular and molecular mechanisms contributing to pain is a fundamental element for developing new analgesics. By using a selective antagonist in an in vitro model, this study aimed to establish the role of TRPV-1 in human odontoblast-like cells (OLCs) as a therapeutic target for dental pain mediated by noxious thermal and osmotic stimuli. Methods: OLCs were differentiated from dental pulp mesenchymal cells and TRPV1 expression was evaluated. Activation of TRPV-1 was determined by evaluating changes in calcium concentration after stimulation with mannitol and xylitol hyperosmotic solutions or DMEM heated at 45 °C, using the fluorescent calcium probe Fluo-4 AM. In addition, changes in fluorescence (F/F0) due to calcium flux were evaluated using fluorometry and flow cytometry. Simultaneously, the cells were co-stimulated with the selective antagonist capsazepine (CZP). Results: OLCs expressed DSPP and DMP-1, confirming their cellular phenotype. TRPV1 was expressed, and its activation by different stimuli produced an increase in cytosolic Ca2+ which was reduced by the antagonist. Both methods used to evaluate TRPV1 activation through the measurement of calcium probe fluorescence showed similar patterns. Conclusions: These results suggest that TRPV-1 modulation using an antagonist can be implemented as a pharmacological strategy for managing dental pain mediated by hyperosmotic and thermal stimuli.

Relaxin treatment stimulates the differentiation of mesenchymal stem cells into osteoblasts.

Background/purpose: Several studies have determined that relaxin stimulates differentiation and regulates the activity of mature osteoclasts, but little is known about its effect on the differentiation of mesenchymal cells towards the osteogenic lineage. Therefore, this study aimed to determine the effect of relaxin on the proliferation and differentiation of the osteoblastic lineage of mesenchymal cells derived from human dental pulp (hDPSC). Materials and methods: In this in vitro study, hDPSC were characterized and treated with relaxin at different doses (10-80 ng/ml) and times (1-21 days). Morphology was assessed by microscopy, and proliferation was assessed using a resazurin assay. Osteoblastic differentiation was evaluated by Alizarin Red staining, alkaline phosphatase (ALP) labeling, and changes in the expression of the osteoblastic differentiation genes RUNX2 and BMP2. Results: Relaxin treatment did not induce changes in the proliferation or viability of hDPSCs; however, larger cells and increased cytoplasmic prolongation were observed. Relaxin treatment (20 and 80 ng/ml) significantly increased calcified nodule formation on days 14 and 21. The cytochemical signals for ALP, RUNX2, and BMP2 gene expression were significantly (P < 0.05) increased by the relaxin treatment. Conclusion: Relaxin treatment does not induce changes in hDPSC proliferation but induces morphological changes, increases ALP detection, calcified nodule formation, and increases expression of RUNX2 and BMP2, suggesting the induction of osteoblastic differentiation of hDPSC.

A tangled threesome: understanding arbovirus infection in Aedes spp. and the effect of the mosquito microbiota.

Arboviral infections transmitted by Aedes spp. mosquitoes are a major threat to human health, particularly in tropical regions but are expanding to temperate regions. The ability of Aedes aegypti and Aedes albopictus to transmit multiple arboviruses involves a complex relationship between mosquitoes and the virus, with recent discoveries shedding light on it. Furthermore, this relationship is not solely between mosquitoes and arboviruses, but also involves the mosquito microbiome. Here, we aimed to construct a comprehensive review of the latest information about the arbovirus infection process in A. aegypti and A. albopictus, the source of mosquito microbiota, and its interaction with the arbovirus infection process, in terms of its implications for vectorial competence. First, we summarized studies showing a new mechanism for arbovirus infection at the cellular level, recently described innate immunological pathways, and the mechanism of adaptive response in mosquitoes. Second, we addressed the general sources of the Aedes mosquito microbiota (bacteria, fungi, and viruses) during their life cycle, and the geographical reports of the most common microbiota in adults mosquitoes. How the microbiota interacts directly or indirectly with arbovirus transmission, thereby modifying vectorial competence. We highlight the complexity of this tripartite relationship, influenced by intrinsic and extrinsic conditions at different geographical scales, with many gaps to fill and promising directions for developing strategies to control arbovirus transmission and to gain a better understanding of vectorial competence. The interactions between mosquitoes, arboviruses and their associated microbiota are yet to be investigated in depth.

A strategy for entomo-virological surveillance of yellow fever, dengue, zika, and chikungunya viruses in field-collected mosquitoes.

Arboviruses transmitted by Culicidae insects are significant threats to human health, presenting dynamic transmission cycles and involving different vectors and hosts. The surveillance and characterization of the vectors involved in these cycles are crucial for understanding and preventing potential outbreaks. Therefore, we propose a strategy that we used for entomological surveillance of urban, rural, and sylvatic mosquitoes and to characterize natural infection by four major arboviruses.•Immature and adult mosquitoes were collected intra, peri and extradomicilie of urban and rural households, using different collection methodologies.•Mosquitoes were pooled or separated in head-thorax and abdomen, according to the species.•A multiplex nested RT-PCR (Reverse transcription polymerase chain reaction) method was used for the simultaneous detection of dengue virus (DENV), zika virus (ZIKV), chikungunya virus (CHIKV), and yellow fever virus (YFV).Overall, this strategy proved helpful for vectors surveillance at different ecosystems, as well as for implementing a low-cost molecular surveillance system that allows the early detection of potential outbreaks, and identify other potential vectors involved in viral transmission.

Seroprevalence, infection, and personal protective equipment use among Colombian healthcare workers during the COVID-19 pandemic.

Introduction: Healthcare workers (HCWs) are at the forefront of the COVID-19 response and frequently come into close contact with patients and their virus-contaminated body fluids. Recent studies have identified differential risks of infection and the use of personal protective equipment (PPE) among HCWs. However, available data might be interpreted with caution because of differences in the national health systems, local implementation issues, and adherence limitations to guidelines. A comprehensive description of infection, exposure at work, and biosafety habits during the COVID-19 pandemic has not been conducted among the HCW groups in Latin American populations. Objective: To describe SARS-CoV-2 seroprevalence, infections, and extent of PPE use during the COVID-19 pandemic among HCWs at three different times, including dental practitioners (DP), nursing assistants (NA), physicians (P), and respiratory therapists (RT), from Bogotá, Colombia. Methods: After IRB approval, this cross-sectional study included 307 HCWs. Participants provided nasopharyngeal swabs and blood samples to detect viral RNA (RT-qPCR) and IgM/IgG anti-SARS-CoV-2 (ELFA-ELISA) at baseline (BL) and two follow-ups. Infection prevalence was defined as the number of positive-tested participants (RT-qPCR and/or IgM). Data on clinical status and biosafety habits were collected each time. Results: Differential infection prevalence was found among HCWs through the study timeline (BL: RT-qPCR = 2.6%, IgM = 1.6%; follow-up 1 (45 days after BL): RT-qPCR = 4.5%, IgM = 3.9%; follow-up 2 (60 days after BL): RT-qPCR = 3.58%, IgM = 1.3%. Dental practitioners showed a higher infection frequency in BL and follow-up 1. IgG-positive tested HCWs percentage progressively increased from BL to follow-ups among the whole sample while index values decreased. Limitations in N95 availability and a high perception of occupational risk were reported. Conclusion: A low prevalence of active SARS-CoV-2 infections among HCWs groups was found. Over time, there was an increase in participants showing IgG antibodies, although the levels of these antibodies in the blood decreased. Additionally, HCWs reported limitations in the availability of PPE as well as a variation in their safety practices.

The Interaction Effect of Anti-RgpA and Anti-PPAD Antibody Titers: An Indicator for Rheumatoid Arthritis Diagnosis.

Porphyromonas gingivalis secretes virulence factors like Arg-gingipains and peptidyl arginine deiminase (PPAD), that are associated with rheumatoid arthritis (RA) pathogenesis. However, there is no information regarding the antibody titers for these bacterial enzymes as systemic indicators or biomarkers in RA. In this cross-sectional study, 255 individuals were evaluated: 143 were diagnosed with RA, and 112 were without RA. Logistic regression models adjusted for age, sex, basal metabolic index, smoking, and periodontitis severity were used to evaluate the association of RA with rheumatoid factor (RF), anti-citrullinated protein antibodies (ACPAs), erythrocyte sedimentation rate, high sensitivity C-reactive protein, anti-RgpA, anti-PPAD, and double positive anti-RgpA/anti-PPAD. It was found that RF (odds ratio [OR] 10.6; 95% confidence interval [CI] 4.4-25), ACPAs (OR 13.7; 95% CI 5.1-35), and anti-RgpA/anti-PPAD double positivity (OR 6.63; 95% CI 1.61-27) were associated with RA diagnoses. Anti-RgpA was also associated with RA (OR 4.09; 95% CI 1.2-13.9). The combination of anti-RgpA/anti-PPAD showed a high specificity of 93.7% and 82.5% PPV in identifying individuals with RA. RgpA antibodies were associated with the periodontal inflammatory index in RA individuals (p < 0.05). The double positivity of the anti-RgpA/anti-PPAD antibodies enhanced the diagnosis of RA. Therefore, RgpA antibodies and anti-RgpA/anti-PPAD may be biomarkers for RA.

In vivo infection by a neuroinvasive neurovirulent dengue virus.

Although neurological manifestations associated with dengue infections have been reported in endemic countries, the viral or host characteristics determining the infection or alteration of nervous function have not been described. In order to investigate neurobiological conditions related to central nervous system dengue virus (DENV) infection, we established a mouse model of neuroinfection. A DENV-4 isolate was first adapted to neuroblastoma cells, later inoculated in suckling mice brain, and finally, this D4MB-6 viral variant was inoculated intraperitoneally in Balb/c mice at different postnatal days (pnd). Virus-induced fatal encephalitis in 2 and 7 pnd mice but infected at 14 and 21 pnd mice survived. The younger mice presented encephalitis at the sixth day postinfection with limb paralysis and postural instability concomitant with efficient viral replication in brain. In this mice model, we found activated microglial cells positive to viral antigen. Neurons, oligodendrocytes, and endothelial cells were also infected by the D4MB-6 virus in neonatal mice, which showed generalized and local plasma leakage with blood-brain barrier (BBB) severe damage. These results suggest that there was a viral fitness change which led to neuroinfection only in immune or neurological immature mice. Infection of neurons, endothelial, and microglial cells may be related to detrimental function or architecture found in susceptible mice. This experimental neuroinfection model could help to have a better understanding of neurological manifestations occurring during severe cases of dengue infection.