A Series of 69 COVID-related Dermatoses With Biopsy, Immunohistochemistry With Anti-spike 3, in situ Hybridization and PCR: A Critical Reappraisal of Viral Involvement in COVID-19 Skin Lesions. / [Articulo traducido] Serie de 69 dermatosis relacionadas con la COVID-19 con biopsia, estudio inmunohistoquímico con anti-spike 3, hibridación in situ y PCR: Una revisión crítica de la afectación cutánea relacionada con la COVID-19.

BACKGROUND: Despite the large number of articles published on skin lesions related to COVID-19, clinicopathological correlation has not been performed consistently and immunohistochemistry to demonstrate spike 3 protein expression has not been validated through RT-PCR. MATERIAL AND METHODS: We compiled 69 cases of patients with confirmed COVID-19, where skin lesions were clinically and histopathologically studied. Immunohistochemistry (IHC) and RT-PCR was performed in skin biopsies. RESULTS: After a careful review of the cases, 15 were found to be dermatosis not related to COVID-19, while the rest of the lesions could be classified according to their clinical characteristics as vesicular (4), maculopapular eruptions (41), urticariform (9), livedo and necrosis (10) and pernio-like (5). Although histopathological features were similar to previously reported results, we found two previously unreported findings, maculopapular eruptions with squamous eccrine syringometaplasia and neutrophilic epitheliotropism. IHC showed in some cases endothelial and epidermal staining but RT-PCR was negative in all the tested cases. Thus, direct viral involvement could not be demonstrated. CONCLUSIONS: Despite presenting the largest series of confirmed COVID-19 patients with histopathologically studied skin manifestations, direct viral involvement was difficult to establish. Vasculopathic and urticariform lesions seem to be those more clearly related to the viral infection, despite IHC or RT-PCR negative results failed to demonstrate viral presence. These findings, as in other dermatological areas, highlight the need of a clinico-pathological correlation to increase knowledge about viral involvement in COVID-19 skin-related lesions.

A Series of 69 COVID-related Dermatoses With Biopsy, Immunohistochemistry With Anti-spike 3, in situ Hybridization and PCR: A Critical Reappraisal of Viral Involvement in COVID-19 Skin Lesions.

BACKGROUND: Despite the large number of articles published on skin lesions related to COVID-19, clinicopathological correlation has not been performed consistently and immunohistochemistry to demonstrate spike 3 protein expression has not been validated through RT-PCR. MATERIAL AND METHODS: We compiled 69 cases of patients with confirmed COVID-19, where skin lesions were clinically and histopathologically studied. Immunohistochemistry (IHC) and RT-PCR was performed in skin biopsies. RESULTS: After a careful review of the cases, 15 were found to be dermatosis not related to COVID-19, while the rest of the lesions could be classified according to their clinical characteristics as vesicular (4), maculopapular eruptions (41), urticariform (9), livedo and necrosis (10) and pernio-like (5). Although histopathological features were similar to previously reported results, we found two previously unreported findings, maculopapular eruptions with squamous eccrine syringometaplasia and neutrophilic epitheliotropism. IHC showed in some cases endothelial and epidermal staining but RT-PCR was negative in all the tested cases. Thus, direct viral involvement could not be demonstrated. CONCLUSIONS: Despite presenting the largest series of confirmed COVID-19 patients with histopathologically studied skin manifestations, direct viral involvement was difficult to establish. Vasculopathic and urticariform lesions seem to be those more clearly related to the viral infection, despite IHC or RT-PCR negative results failed to demonstrate viral presence. These findings, as in other dermatological areas, highlight the need of a clinico-pathological correlation to increase knowledge about viral involvement in COVID-19 skin-related lesions.

MicroBooNE and the ν_{e} Interpretation of the MiniBooNE Low-Energy Excess.

A new generation of neutrino experiments is testing the 4.7σ anomalous excess of electronlike events observed in MiniBooNE. This is of huge importance for particle physics, astrophysics, and cosmology, not only because of the potential discovery of physics beyond the standard model, but also because the lessons we will learn about neutrino-nucleus interactions will be crucial for the worldwide neutrino program. MicroBooNE has recently released results that appear to disfavor several explanations of the MiniBooNE anomaly. Here, we show quantitatively that MicroBooNE results, while a promising start, unquestionably do not probe the full parameter space of sterile neutrino models hinted at by MiniBooNE and other data, nor do they probe the ν_{e} interpretation of the MiniBooNE excess in a model-independent way.

Loperamide exerts a direct bactericidal effect against M. tuberculosis, M. bovis, M. terrae and M. smegmatis.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis. TB is highly prevalent, characterized by the constant occurrence of drug-resistant cases, and confounded by the incidence of respiratory disease caused by non-tuberculous mycobacteria (NTB). Expanding the spectrum of drugs for the treatment of TB is indispensable. Loperamide, an antidiarrhoeal drug, enhances immune-driven antimycobacterial activity, and we aimed to evaluate its bactericidal activity against M. tuberculosis, Mycobacterium bovis BCG, Mycobacterium terrae and Mycobacterium smegmatis. Loperamide exhibited an inhibitory effect against all mycobacterial species tested, with MICs of 100 and 150 µg ml-1 . Thus, loperamide is a mycobactericidal drug with potential as adjunctive therapy for TB and NTB infections.

Short Disordered Epitope of CRTAM Ig-Like V Domain as a Potential Target for Blocking Antibodies.

Class-I Restricted T Cell-Associated Molecule (CRTAM) is a protein that is expressed after T cell activation. The interaction of CRTAM with its ligand, nectin-like 2 (Necl2), is required for the efficient production of IL-17, IL-22, and IFNγ by murine CD4 T cells, and it plays a role in optimal CD8 T and NK cell cytotoxicity. CRTAM promotes the pro-inflammatory cytokine profile; therefore, it may take part in the immunopathology of autoimmune diseases such as diabetes type 1 or colitis. Thus, antibodies that block the interaction between CRTAM and Necl2 would be useful for controlling the production of these inflammatory cytokines. In this work, using bioinformatics predictions, we identified three short disordered epitopes (sDE1-3) that are located in the Ig-like domains of murine CRTAM and are conserved in mammalian species. We performed a structural analysis by molecular dynamics simulations of sDE1 (QHPALKSSKY, Ig-like V), sDE2 (QRNGEKSVVK, Ig-like C1), and sDE3 (CSTERSKKPPPQI, Ig-like C1). sDE1, which is located within a loop of the contact interface of the heterotypic interaction with Nectl2, undergoes an order-disorder transition. On the contrary, even though sDE2 and sDE3 are flexible and also located within loops, they do not undergo order-disorder transitions. We evaluated the immunogenicity of sDE1 and sDE3 through the expression of these epitopes in chimeric L1 virus-like particles. We confirmed that sDE1 induces polyclonal antibodies that recognize the native folding of CRTAM expressed in activated murine CD4 T cells. In contrast, sDE3 induces polyclonal antibodies that recognize the recombinant protein hCRTAM-Fc, but not the native CRTAM. Thus, in this study, an exposed disordered epitope in the Ig-like V domain of CRTAM was identified as a potential site for therapeutic antibodies.

Identification and biofilm development by a new fungal keratitis aetiologic agent.

BACKGROUND: In recent years, human keratitis caused by fungal plant pathogens has become more common. Biofilm is a structure that confers adaptations and virulence to fungi in keratitis. Neoscytalidium spp. are phytopathogenic and recently have been recognised as a human pathogen, using biofilm formation as a virulence factor. OBJECTIVES: The aim of this study was isolation, identification (at the species level) and characterisation of a new fungal keratitis agent. PATIENTS/METHODS: The fungus was isolated from a 67-year-old male patient with a corneal ulcer. Biofilm formation and structure were evaluated by colorimetric methods and microscopy. To identify the fungus, morphological characteristics were examined and a phylogenetic analysis was performed. RESULTS AND CONCLUSIONS: We report the identification of a fungus, a member of the genus Neoscytalidium which is associated with human keratitis. Phylogenetic analysis and morphological observations on conidiogenous cells, which occur only in arthric chains in aerial mycelium and the coelomycetous synasexual morph is absent, identified a new species, Neoscytalidium oculus sp. nov. The fungus formed biofilm at a concentration of 1 × 106  conidia/mL, during 96 hours of incubation at 37°C, and also manifested haemolysis and melanin production. This is the first report in Latin America of a new species of Neoscytalidium from a clinical isolate has been identified.

Patterns of helminth infections in Rattus rattus and Mus musculus from two Mayan communities in Mexico.

The black rat Rattus rattus and the house mouse Mus musculus are two commensal rodent species that harbour and shed zoonotic pathogens, including helminths. The aim of this survey was to study the helminth community and the patterns of infections in R. rattus and M. musculus from two Mayan communities in Mexico. Gastrointestinal helminths were isolated from 322 M. musculus and 124 R. rattus, including Gongylonema neoplasticum, Hassalstrongylus aduncus, Hassalstrongylus musculi, Hydatigera taeniaeformis metacestode, Hymenolepis diminuta, Nippostrongylus brasiliensis, Oligacanthorhynchidae gen. sp., Syphacia muris, Syphacia obvelata, Rodentolepis microstoma and Trichuris muris. The overall richness of helminths was seven in R. rattus and six in M. musculus. The results of generalized linear models showed that juvenile rodents had lower probabilities of being infected with G. neoplasticum, H. taeniaeformis and H. musculi than adult rodents. A positive association between the prevalence of S. muris and rat abundance was found. The intensity of infection with S. muris was higher in the rainy season than in the dry season; the opposite result was found for H. musculi infection. Male R. rattus harboured more S. muris specimens. The intensity of infection with T. muris was inversely associated with mouse abundance. The presence of the zoonotic H. diminuta, as well as H. taeniaeformis and R. microstoma in rodent populations indicates that there is risk of transmission, and that their entire life cycle occurs in the study area.

Effect of point mutations on the ultrafast photo-isomerization of Anabaena sensory rhodopsin.

Anabaena sensory rhodopsin (ASR) is a particular microbial retinal protein for which light-adaptation leads to the ability to bind both the all-trans, 15-anti (AT) and the 13-cis, 15-syn (13C) isomers of the protonated Schiff base of retinal (PSBR). In the context of obtaining insight into the mechanisms by which retinal proteins catalyse the PSBR photo-isomerization reaction, ASR is a model system allowing to study, within the same protein, the protein-PSBR interactions for two different PSBR conformers at the same time. A detailed analysis of the vibrational spectra of AT and 13C, and their photo-products in wild-type ASR obtained through femtosecond (pump-) four-wave-mixing is reported for the first time, and compared to bacterio- and channelrhodopsin. As part of an extensive study of ASR mutants with blue-shifted absorption spectra, we present here a detailed computational analysis of the origin of the mutation-induced blue-shift of the absorption spectra, and identify electrostatic interactions as dominating steric effects that would entail a red-shift. The excited state lifetimes and isomerization reaction times (IRT) for the three mutants V112N, W76F, and L83Q are studied experimentally by femtosecond broadband transient absorption spectroscopy. Interestingly, in all three mutants, isomerization is accelerated for AT with respect to wild-type ASR, and this the more, the shorter the wavelength of maximum absorption. On the contrary, the 13C photo-reaction is slightly slowed down, leading to an inversion of the ESLs of AT and 13C, with respect to wt-ASR, in the blue-most absorbing mutant L83Q. Possible mechanisms for these mutation effects, and their steric and electrostatic origins are discussed.

Flavonoids from Boldoa purpurascens inhibit proinflammatory cytokines (TNF-α and IL-6) and the expression of COX-2.

The flavonoids comprise a large class of plant metabolites distributed in food plants. These compounds have antioxidant, antitumor, antiallergic, and anti­inflammatory effects. The molecular mechanisms of their biological activities remain to be clearly understood. We investigated the in vitro anti­inflammatory potential of a flavonoid mixture and isolated compounds from the leaves of Boldoa purpurascens. Our results provide direct evidence of the anti­inflammatory effects of the mixture, which are mediated by the inhibition of the proinflammatory cytokines tumor necrosis factor α and interleukin 6 as well as the modulation of the expression of cyclooxygenase 2.

Jaw closing movement and sex differences in temporomandibular joint energy densities.

Energy densities (ED, mJ/mm3 ) quantify mechanical work imposed on articular cartilages during function. This cross-sectional study examined differences in temporomandibular joint (TMJ) ED during asymmetric versus symmetric jaw closing in healthy females versus males. ED component variables were tested for differences between and within sexes for two types of jaw closing. Seventeen female and 17 male subjects gave informed consent to participate. Diagnostic criteria for temporomandibular disorders and images (magnetic resonance (MR), computed tomography) were used to confirm healthy TMJ status. Numerical modelling predicted TMJ loads (Fnormal ) consequent to unilateral canine biting. Dynamic stereometry combined MR imaging and jaw-tracking data to measure ED component variables during 10 trials of each type of jaw closing in each subject's TMJs. These data were then used to calculate TMJ ED during jaw closing asymmetrically and symmetrically. Paired and Student's t tests assessed ED between jaw closing movements and sexes, respectively. Multivariate data analyses assessed ED component variable differences between jaw closing movements and sexes (α = 0.05). Contralateral TMJ ED were 3.6-fold and significantly larger (P < .0001) during asymmetric versus symmetric jaw closing, due to significantly larger (P ≤ .001) distances of TMJ stress-field translation in asymmetric versus symmetric movement. During asymmetric jaw closing, contralateral TMJ ED were twofold and significantly larger (P = .036) in females versus males, due to 1.5-fold and significantly smaller (P ≤ .010) TMJ disc cartilage volumes under stress fields in females versus males. These results suggest that in healthy individuals, asymmetric compared to symmetric jaw closure in females compared to males has higher TMJ mechanical fatigue liabilities.

Comparative proteomic profiles reveal characteristic Mycobacterium tuberculosis proteins induced by cholesterol during dormancy conditions.

Cholesterol has been reported to play an important role during Mycobacterium tuberculosis infection and during its dormant state inside the host. We present the determination of proteomic profiles of M. tuberculosis H37Rv in the presence of cholesterol as the sole carbon source under exponential growth and in two in vitro dormancy phases (NRP1 and NRP2). Using 2D-PAGE, we detected that M. tuberculosis expressed a high diversity of proteins in both exponential and non-replicative phases. We also found that cholesterol was involved in the overexpression of some proteins related to sulfur metabolism (CysA2), electron transport (FixB), cell wall synthesis (Ald), iron storage (BfrB), protein synthesis (Tig and EF-Tu) and dormancy maintenance (HspX and TB 31.7). According to our results we propose that proteins Ald, BfrB, FadA5 and TB31.7 are likely to play a fundamental role during in vitro dormancy of M. tuberculosis in the presence of cholesterol, helping to counteract its intracellular hostile microenvironment.

TMJ energy densities in healthy men and women.

OBJECTIVE: Cartilage fatigue, due to mechanical work, may account for the early development of degenerative joint disease (DJD) in the temporomandibular joint (TMJ), and why women are three times more likely to be afflicted. This study tested for gender differences in mechanical energy densities in women and men with healthy TMJs. DESIGN: Eighteen women and eighteen men gave informed consent. Research diagnostic criteria including imaging were used to ensure that subjects' TMJs were normal, without disc displacement or signs of DJD. Numerical modeling determined TMJ loads (Fnormal). Jaw tracking and three-dimensional dynamic stereometry characterized individual-specific data of stress-field dynamic mechanics during 10 symmetrical jaw closing cycles. These data were used to estimate tractional forces (Ftraction). Energy densities were then calculated, where: Energy Density = W/Q (W = work done or mechanical energy input = Ftraction*distance of stress-field translation, Q = volume of cartilage). Two-way analysis of variance (ANOVA) and follow-up two-group comparisons tested mean energy densities for ipsilateral and contralateral TMJs in women vs men. RESULTS: Mean energy densities ± standard deviations in ipsilateral and contralateral TMJs in women were 9.0 ± 9.7 and 8.4 ± 5.5 mJ/mm3, respectively, and were significantly larger (P = 0.004 and 0.001, respectively) compared to ipsilateral and contralateral TMJs in men, which were 5.6 ± 4.2 and 6.3 ± 4.2 mJ/mm3, respectively. CONCLUSIONS: Energy densities were significantly larger in healthy TMJs of women than men. Larger TMJ energy densities during normal jaw functions could predispose earlier mechanical fatigue of the TMJ disc.

A survey of zoonotic pathogens carried by house mouse and black rat populations in Yucatan, Mexico.

The house mouse (Mus musculus) and the black rat (Rattus rattus) are reservoir hosts for zoonotic pathogens, several of which cause neglected tropical diseases (NTDs). Studies of the prevalence of these NTD-causing zoonotic pathogens, in house mice and black rats from tropical residential areas are scarce. Three hundred and two house mice and 161 black rats were trapped in 2013 from two urban neighbourhoods and a rural village in Yucatan, Mexico, and subsequently tested for Trypanosoma cruzi, Hymenolepis diminuta and Leptospira interrogans. Using the polymerase chain reaction we detected T. cruzi DNA in the hearts of 4·9% (8/165) and 6·2% (7/113) of house mice and black rats, respectively. We applied the sedimentation technique to detect eggs of H. diminuta in 0·5% (1/182) and 14·2% (15/106) of house mice and black rats, respectively. Through the immunofluorescent imprint method, L. interrogans was identified in 0·9% (1/106) of rat kidney impressions. Our results suggest that the black rat could be an important reservoir for T. cruzi and H. diminuta in the studied sites. Further studies examining seasonal and geographical patterns could increase our knowledge on the epidemiology of these pathogens in Mexico and the risk to public health posed by rodents.

The sigma factor SigD of Mycobacterium tuberculosis putatively enhances gene expression of the septum site determining protein under stressful environments.

This work examined the expression of the septum site determining gene (ssd) of Mycobacterium tuberculosis CDC1551 and its ∆sigD mutant under different growing conditions. The results showed an up-regulation of ssd during stationary phase and starvation conditions, but not during in vitro dormancy, suggesting a putative role for SigD in the control of ssd expression mainly under lack-of-nutrients environments. Furthermore, we elucidated a putative link between ssd expression and cell elongation of bacilli at stationary phase. In addition, a -35 sigD consensus sequence was found for the ssd promoter region, reinforcing the putative regulation of ssd by SigD, and in turn, supporting this protein role during the adaptation of M. tuberculosis to some stressful environments.

A pilot study of nocturnal temporalis muscle activity in TMD diagnostic groups of women.

Temporomandibular disorder (TMD) incidences are believed to be related to parafunctional behaviours like teeth clenching. This pilot study aimed to (i) develop an automated clench-detection algorithm, and (ii) apply the algorithm to test for differences in nocturnal clenching in women with and without TMD. Subjects gave informed consent to participate. Adult women were categorised using Diagnostic Criteria for TMD according to presence/absence (+/-) of both TM joint disc placement (DD) and chronic pain (P) into two groups (+DD+P, -DD-P) with 12 subjects each. Surface temporalis electromyography was recorded during oral tasks performed by subjects at two laboratory sessions. The data were used to characterise muscle activity per N of bite force (µV/N) for each subject, develop the clench-detection algorithm and test its accuracy. Ambulatory surface temporalis electromyography was self-recorded by each subject over three nights and analysed using the algorithm and bite force (N) versus muscle activity µV/N calibrations. Bonferroni-adjusted homoscedastic t-tests assessed for significant between-group differences in clenching (P < 0·05). Sensitivity, specificity and accuracy of algorithm-detected laboratory clenches were all ≥96%. During self-recordings 95% of clenches had durations of <4 s and peak forces of <10 N in both groups. Mean clench durations were significantly longer (P = 0·042) in +DD+P (1·9 ± 0·8 s) than -DD-P subjects (1·4 ± 0·4 s). Mean temporalis duty factors (%clench time/total recording time) were significantly larger (P = 0·041) in +DD+P (0·47 ± 0·34%) than -DD-P (0·26 ±0·22%) subjects. Nocturnal temporalis muscle activities detected by a validated algorithm were longer per clench and recording time in +DD+P compared to -DD-P women.

Optimizing CIGB-300 intralesional delivery in locally advanced cervical cancer.

BACKGROUND: We conducted a phase 1 trial in patients with locally advanced cervical cancer by injecting 0.5 ml of the CK2-antagonist CIGB-300 in two different sites on tumours to assess tumour uptake, safety, pharmacodynamic activity and identify the recommended dose. METHODS: Fourteen patients were treated with intralesional injections containing 35 or 70 mg of CIGB-300 in three alternate cycles of three consecutive days each before standard chemoradiotherapy. Tumour uptake was determined using (99)Tc-radiolabelled peptide. In situ B23/nucleophosmin was determined by immunohistochemistry. RESULTS: Maximum tumour uptake for CIGB-300 70-mg dose was significantly higher than the one observed for 35 mg: 16.1 ± 8.9 vs 31.3 ± 12.9 mg (P = 0.01). Both, AUC24h and biological half-life were also significantly higher using 70 mg of CIGB-300 (P < 0.001). Unincorporated CIGB-300 diffused rapidly to blood and was mainly distributed towards kidneys, and marginally in liver, lungs, heart and spleen. There was no DLT and moderate allergic-like reactions were the most common systemic side effect with strong correlation between unincorporated CIGB-300 and histamine levels in blood. CIGB-300, 70 mg, downregulated B23/nucleophosmin (P = 0.03) in tumour specimens. CONCLUSION: Intralesional injections of 70 mg CIGB-300 in two sites (0.5 ml per injection) and this treatment plan are recommended to be evaluated in phase 2 studies.

A pilot study of ambulatory masticatory muscle activities in temporomandibular joint disorders diagnostic groups.

OBJECTIVE: To determine differences in masticatory muscle usage between temporomandibular joint disorders diagnostic groups. SETTING AND SAMPLE POPULATION: Seventy-one informed and consented subjects (27 men; 44 women) participated at the University at Buffalo. MATERIAL AND METHODS: Research diagnostic criteria and imaging data were used to categorize subjects according to the presence/absence +/- of TMJ disc placement (DD) and chronic pain (P) (+DD+P, n=18; +DD-P, n=14; -DD-P, n=39). Electromyographic (EMG)/bite-force calibrations determined subject-specific masseter and temporalis muscle activities per 20 N bite-force (T20N , µV). Over 3 days and nights, subjects collected EMG recordings. Duty factors (DFs, % of recording time) were determined based on threshold intervals (5-9, 10-24, 25-49, 50-79, ≥80% T20N ). anova and Tukey-Kramer post hoc tests identified 1) diagnostic group differences in T20N and 2) the effects of diagnostic group, gender, time and interval on muscle DFs. RESULTS: Mean (±SE) temporalis T20N in +DD+P subjects was significantly higher (71.4±8.8 µV) than masseter T20N in these subjects (19.6±8.8 µV; p=0.001) and in -DD-P subjects (25.3±6.0 µV, p=0.0007). Masseter DFs at 5-9% T20N were significantly higher in +DD-P women (3.48%) than +DD-P men (0.85%) and women and men in both other diagnostic groups (all p<0.03), and in +DD+P women (2.00%) compared to -DD-P men (0.83%; p=0.029). Night-time DFs at 5-9% T20N in +DD-P women (1.97%) were significantly higher than in -DD-P men (0.47%) and women (0.24%; all p<0.01). CONCLUSIONS: Between-group differences were found in masticatory muscle activities in both laboratory and natural environmental settings.

Infield masticatory muscle activity in subjects with pain-related temporomandibular disorders diagnoses.

OBJECTIVES: Pain-related temporomandibular disorders (TMDs) are the most prevalent conditions among TMDs. There is contrasting evidence available for association of pain-related TMD and masticatory muscle activity (MMA). The present investigation assesses the associations between MMA levels of masseter and temporalis muscles during awake and sleep among pain-related TMD diagnostic groups. SETTING AND SAMPLE POPULATION: The department of Oral Diagnostic Sciences, University at Buffalo. Twenty females and six males participated in this study. MATERIAL AND METHODS: Using the diagnostic criteria for temporomandibular disorders (DC-TMDs), participants were diagnostically categorized. Subjects used a custom monitoring system, which recorded infield muscle activities. A factorial model tested for association between independent variable (muscle, time period, MMA level, and diagnostic group) effects and the logarithm of MMA. Greenhouse-Geisser test was used to determine any statistically significant associations (p≤0.003). RESULTS: No statistically significant association was found between four-way, three-way, and two-way analyses. However, among the main effects, range of magnitudes was the only variable to be statistically significant. Although the data suggest a trend of increased masseter MMA in the pain-related TMD diagnoses group both during awake and sleep time periods, such observation is not maintained for the temporalis muscle. In addition, temporalis MMA was found to be higher in the pain-related TMD diagnoses group only at extreme activity levels (<25 and ≥80% ranges). CONCLUSION: This data support the association between masticatory muscle hyperactivity and painful TMD conditions.