Molecular methods for diagnosis of monkeypox: A mini-review.

BACKGROUND: Monkeypox is a global public health issue caused by the monkeypox virus (MPXV). As of October 28, 2022, a total of 77,115 laboratory-confirmed cases and 3,610 probable cases, including 36 deaths, were reported, with 9,070 cases reported in Brazil, the second most affected country. The need to develop national technologies for the rapid diagnosis of emerging diseases for mass testing of the population is evident, as observed in the SARS-CoV-2 pandemic. OBJECTIVE: With that in mind, this article provides an overview of current methods, techniques, and their applications in the molecular detection of monkeypox, focusing the search on real-time polymerase chain reaction (qPCR), polymerase chain reaction (PCR), and polymerase chain reaction-enzyme linked immunosorbent assay (PCR-ELISA). METHODS: The relevant documents or papers covered in this study were selected by a search in international bibliographic databases. The search terms used in the databases were aimed at summarizing existing knowledge on molecular diagnostic methods, such as monkeypox; MPX, MPXV, qPCR, PCR, PCR-ELISA, diagnosis and detection searched separately or together using the Boolean operator "AND" either in the title or abstract. The searches took place in September 2022, and the corresponding articles were selected between 2012 and 2022. RESULTS: We found 256 documents in total and twelve studies addressing the molecular diagnosis of monkeypox were classified as possible sources for this review. CONCLUSION: It is evident there is a pressing need to develop national technologies for rapid diagnosis of emerging diseases for mass testing of the population. It is also extremely important to have national detection kits with greater diagnostic capacity to assist in developing effective public policies in countries affected by this disease.

Aerobic degradation of 2,4,6-trinitrophenol by Proteus sp. strain OSES2 obtained from an explosive contaminated tropical soil.

A 2,4,6-trinitrophenol (TNP) degrading bacterial strain isolated from a site polluted with explosives was identified as Proteus sp. strain OSES2 via 16S rRNA gene sequencing. Metabolic investigation showed that the organism grew exponentially on 100 mg l-1 of TNP as a source of carbon, nitrogen, and energy. In addition, the growth of the organism was sustainable on 3-nitrotoluene, 2,4-dinitrotoluene, 2,4,6-trinitrotoluene, 4-nitrophenol, methyl-3-nitrobenzoate, 4-nitroaniline, aniline and nitrobenzene. Strain OSES2 was able to utilize TNP within a concentration range of 100 mg l-1 to 500 mg l-1. The specific growth rate and degradation rates on TNP were 0.01043 h-1 and 0.01766 mg l-1 h-1 respectively. Effective degradation of TNP in a chemically defined medium was evident with a gradual reduction in the concentration of TNP concomitant with an increase in cell density as well as the substantial release of ammonium (NH4+), nitrite (NO2-), and nitrate (NO3-) as metabolites in 96 h. Degradation competence of the organism was enhanced in the presence of starch and acetate. On starch-supplemented TNP, the highest specific growth rate and degradation rates were 0.02634 h-1 and 0.04458 mg l-1 h-1, respectively, while the corresponding values on acetate were 0.02341 h-1 and 0.02811 mg l-1 h-1. However, amendment with nitrogen sources yielded no substantial improvement in degradation. TNP was utilized optimally at pH 7 to 9 and within the temperature range of 30 °C to 37 °C. The enzyme hydride transferase II [HTII], encoded by the npdI gene which is the first step involved in the TNP degradation pathway, was readily expressed by the isolate thus suggesting that substrate was utilized through the classical metabolic pathway.

Novel Eudragit® -based polymeric nanoparticles for sustained release of simvastatin

This paper reports on the development of nanoparticles aiming at the in vitro controlled release of simvastatin (SVT). The nanoparticles were prepared by the nanoprecipitation method with polymers Eudragit® FS30D (EDGFS) or Eudragit® NE30D (EDGNE). EDGFS+SVT nanoparticles showed mean size of 148.8 nm and entrapment efficiency of 76.4%, whereas EDGNE+SVT nanoparticles showed mean size of 105.0 nm and entrapment efficiency of 103.2%. Infrared absorption spectra demonstrated that SVT incorporated into the polymer matrix, especially EDGNE. Similarly, the differential scanning calorimeter (DSC) curve presented no endothermic peak of fusion, indicating that the system is amorphous and the drug is not in the crystalline state. The maintenance of SVT in the amorphous state may favors its solubilization in the target release sites. In the in vitro dissolution assay, the SVT incorporated into the EDGFS+SVT nanoparticles showed a rapid initial release, which may be related to the pH of the dissolution medium used. Regarding the EDGNE+SVT nanoparticles, the in vitro release occurred in a bimodal behavior, i.e., an initial "burst" followed by a sustained delivery, with the kinetics of drug release following Baker-Lonsdale's mathematical model. All these features suggest the nanoparticulate system's potential to modulate SVT delivery and enhance its bioavailability.

Smoking History Decreases Survival in Patients with Squamous Cell Carcinoma of the Mouth: A Retrospective Study with 15 Years of Follow-up.

Background: The purpose of this study was to evaluate the influence of smoking history on the clinical-pathological, sociodemographic and prognostic characteristics of patients with oral squamous cell carcinoma (SCC). Materials and Methods: A retrospective cohort study was carried out with the records of 136 smokers with SCC and 68 nonsmokers with oral SCC who were diagnosed and treated at Haroldo Juaçaba Hospital (2000-2014). Data on patient sex, age, race, education level, tumor location, tumor size, lymph node involvement, distant metastasis, treatment type, marital status, method of health care access (public or private health systems) and overall survival (15 years) were analyzed by the X² test, Mantel-Cox tests and multinomial and Cox logistic regression models (SPSS 20.0, p <0.05). Results: Smoking history was directly associated with male sex (p <0.001), low levels of education (p = 0.001), tumors of the mouth and palate (p = 0.001), stage T3/4 tumors (p = 0.014), lymph node metastasis (N+) (p = 0.024), palliative treatment (p = 0.024) and receiving health care through the public health system (p = 0.006), with education level being the only independently associated factor (p = 0.039). Lower survival was observed in patients who were smokers (p = 0,002), with low levels of education (p = 0.001), who had stage T3/4 tumors (p = 0.004), with N+ (p = 0.021), and had received palliative treatment (p = 0.002). Age (>65 years old, p = 0.015) and T staging (T3/4, p = 0.033) decreased the survival of SCC patients regardless of the other factors. Conclusions: Smoking history had an independent association with low education level and a history of alcoholism, and survival was negatively associated with older age and larger tumor size, which were more prevalent in smokers.

Differences in respiratory syncytial virus and influenza infection in a house-dust-mite-induced asthma mouse model: consequences for steroid sensitivity.

A significant number of clinical asthma exacerbations are triggered by viral infection. We aimed to characterize the effect of virus infection in an HDM (house dust mite) mouse model of asthma and assess the effect of oral corticosteroids. HDM alone significantly increased eosinophils, lymphocytes, neutrophils, macrophages and a number of cytokines in BAL (bronchoalveolar lavage), all of which were sensitive to treatment with prednisolone (with the exception of neutrophils). Virus infection also induced cell infiltration and cytokines. RSV (respiratory syncytial virus) infection in HDM-treated animals further increased all cell types in BAL (except eosinophils, which declined), but induced no further increase in HDM-elicited cytokines. However, while HDM-elicited TNF-α (tumour necrosis factor-α), IFN-γ (interferon-γ), IL (interleukin)-2, IL-5 and IL-10 were sensitive to prednisolone treatment, concomitant infection with RSV blocked the sensitivity towards steroid. In contrast, influenza infection in HDM- challenged animals resulted in increased BAL lymphocytes, neutrophils, IFN-γ, IL-1ß, IL-4, IL-5, IL-10 and IL-12, but all were attenuated by prednisolone treatment. HDM also increased eNO (exhaled NO), which was further increased by concomitant virus infection. This increase was only partially attenuated by prednisolone. RSV infection alone increased BAL mucin. However, BAL mucin was increased in HDM animals with virus infection. Chronic HDM challenge in mice elicits a broad inflammatory response that shares many characteristics with clinical asthma. Concomitant influenza or RSV infection elicits differing inflammatory profiles that differ in their sensitivity towards steroids. This model may be suitable for the assessment of novel pharmacological interventions for asthmatic exacerbation.

Ocorrência de perda dentária entre os usuários da estratégia de saúde da família do município de Campina Grande - PB / Occurrence of tooth loss among users of the family health strategy in Campina Grande - PB

OBJETIVO: Avaliar a ocorrência de perda dentária em usuários da Estratégia de Saúde da Família do município de Campina Grande-PB. MATERIAL E MÉTODOS: A amostra, do tipo não probabilística, foi composta por 204 usuários das UBSFs, com idade igual ou superior a 18 anos. Utilizou-se como instrumento de coleta de dados um formulário específico, com questões abertas e fechadas, dicotômicas e de múltipla escolha. As variáveis estudadas foram: sexo, idade, perda dentária, tipo de elemento dentário ausente e motivo da perda. O exame bucal foi realizado por um único pesquisador, sob luz natural, no próprio ambiente da UBSF, usando apenas espátulas de madeira descartáveis. Os dados foram analisados através do SPSS (Statistical Package for Social Sciences) versão 13.0, e apresentados por meio da estatística descritiva (distribuições absolutas e percentuais). RESULTADOS: Constatou-se que 87,7% da amostra apresentavam perda dentária, predominando os molares (69,3%). Indivíduos com idade igual ou superior a 56 anos apresentaram o maior percentual de elementos dentários perdidos (53,1%). Os homens apresentaram um maior percentual de perda dentária (34,4%) quando comparados às mulheres. A cárie dentária se constituiu no principal motivo para a exodontia (47,3%). CONCLUSÃO: Verificou-se ser elevada a prevalência de edentulismo, acometendo, principalmente, indivíduos do sexo masculino e acima de 56 anos de idade. Os dentes posteriores foram os mais frequentemente perdidos, sendo a cárie dentária a principal causa

OBJECTIVE: To evaluate the occurrence of tooth loss among users of the Family Health Strategy in Campina Grande-PB. METHOD: The non-probabilistic sample was composed by 204 users of FHUs, aged 18 years or more. Data collection instrument used was a specific form, containing open, closed, multiple choice and dichotomous questions. The variables studied were sex, age, tooth loss, type of missing tooth and reason for losing. The oral examination was performed by a single examiner, under natural light at the FHU environment, using only disposable wooden spatulas. Data were analyzed using the SPSS (Statistical Package for Social Sciences) version 13.0, and presented by means of descriptive statistics (absolute and percentage distributions). RESULTS: 87.7% of the sample was found to present teeth loss, and molars predominated (69.3%). Individuals aged over 56 years had the highest percentage of teeth lost (53.1%). Men had a higher percentage of tooth loss (34.4%) compared to women. Decay was the main reason for tooth extraction (47.3%). CONCLUSION: Tooth loss prevalence was proven to be high, affecting mainly males and above 56 years of age. Posterior teeth were the most frequently lost, being tooth decays the main cause

Therapy with TLR7 agonists induces lymphopenia: correlating pharmacology to mechanism in a mouse model.

BACKGROUND AND OBJECTIVE: Synthetic TLR7 agonists have been proposed as oral replacements for interferonα (IFNα) therapy in the treatment of hepatitis C virus infection. However, adverse effects, such as lymphopenia and cardiovascular irregularities, have been observed in the clinical following treatment with TLR7 agonists. We wished to understand and characterise the relationship between TLR7 agonism and adverse effects. METHODS: We compared responses to two prototypic TLR7 agonists (Resiquimod: R-848; and PF-04878691) in a mouse model and compared the responses to treatment with IFNα. We measured clinically relevant adverse effects such as lymphopenia and cardiovascular irregularities and related them to plasma drug levels and clinically relevant efficacy biomarkers such as the pro-inflammatory cytokine IP-10, 2'5'OAS and TLR7 receptor expression. RESULTS: By 2 h post dose all agents had induced a dose-dependent transient lymphopenia. IFNα increased heart rate immediately following dosing, persisting for 5 h, whilst PF-04878691 induced significant reductions in blood pressure. Lymphopenia co-incided with maximum plasma drug levels, raised levels of IP-10 and the auto-induction of TLR7 expression in the blood and lymph nodes. Peak levels of 2'5'OAS occurred at 24 h post-dose and only at doses which also induced lymphopenia. CONCLUSIONS: We conclude that systemic delivery of TLR7 agonists or IFNα induces similar exaggerated pharmacology, consistent with there being a narrow therapeutic window between efficacy and safety. This clinically validated mouse model will help to investigate whether more potent agonists or optimised dosing schedules, will be successful strategies for targeting TLR7 in patients.

An imidazopiperidine series of CCR5 antagonists for the treatment of HIV: the discovery of N-{(1S)-1-(3-fluorophenyl)-3-[(3-endo)-3-(5-isobutyryl-2-methyl-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-1-yl)-8-azabicyclo[3.2.1]oct-8-yl]propyl}acetamide (PF-232798).

Preventing entry of HIV into human host cells has emerged as an attractive approach to controlling viral replication. Maraviroc 1 is an approved antagonist of the human CCR5 receptor which prevents the entry of HIV. Herein, we report the design and discovery of a series of imidazopiperidine CCR5 antagonists which retain the attractive antiviral profile and window over hERG activity of maraviroc 1, combined with improved absorption profiles in rat and dog. Furthermore, this series of compounds has been shown to retain activity against a laboratory generated maraviroc-resistant HIV-1 strain, which indicates an alternative resistance profile to that of maraviroc 1. Compound 41f (PF-232798) was selected as a clinical candidate from the imidazopiperidine series and is currently in phase II clinical trials.

Use of a highly sensitive strand-specific quantitative PCR to identify abortive replication in the mouse model of respiratory syncytial virus disease.

BACKGROUND: The BALB/c mouse is commonly used to study RSV infection and disease. However, despite the many advantages of this well-characterised model, the inoculum is large, viral replication is restricted and only a very small amount of virus can be recovered from infected animals. A key question in this model is the fate of the administered virus. Is replication really being measured or is the model measuring the survival of the virus over time? To answer these questions we developed a highly sensitive strand-specific quantitative PCR (QPCR) able to accurately quantify the amount of RSV replication in the BALB/c mouse lung, allowing characterisation of RSV negative and positive strand RNA dynamics. RESULTS: In the mouse lung, no increase in RSV genome was seen above the background of the original inoculum whilst only a limited transient increase (< 1 log) in positive strand, replicative intermediate (RI) RNA occurred. This RNA did however persist at detectable levels for 59 days post infection. As expected, ribavirin therapy reduced levels of infectious virus and RI RNA in the mouse lung. However, whilst Palivizumab therapy was also able to reduce levels of infectious virus, it failed to prevent production of intracellular RI RNA. A comparison of RSV RNA kinetics in human (A549) and mouse (KLN205) cell lines demonstrated that RSV replication was also severely delayed and impaired in vitro in the mouse cells. CONCLUSIONS: This is the first time that such a sensitive strand-specific QPCR technique has been to the RSV mouse system. We have accurately quantified the restricted and abortive nature of RSV replication in the mouse. Further in vitro studies in human and mouse cells suggest this restricted replication is due at least in part to species-specific host cell-viral interactions.

Avaliação ecotoxicológica preliminar das águas das bacias hidrográficas dos rios Tarumã, São Raimundo e Educandos

The Tarumã, São Raimundo and Educandos basins, and their tributaries, are impacted by antropic influence. This study tried to evaluate the water toxicity of the hydrographic basins, through acute toxicity bioassay using Daphnia similis as test-organism and physical and chemical analysis. The results showed that, of the three basins, the water from Tarumã caused the most deleterious effect on D. similis.

As Bacias hidrográficas dos riosTarumã, São Raimundo e Educandos e seus afluentes estão sendo impactados, devido as influências antrópicas. Neste estudo foi realizada uma avaliação da toxicidade das águas destes ecossistemas, através de testes de toxicidade aguda com Daphnia similis e análises físico-químicas. Os resultados obtidos demostraram um maior efeito deletério sobre D. similis, nas amostras da microbacia Tarumã.