Lactobacillus crispatus as the etiological agent in cytolytic vaginosis

Introduction: Lactobacillus spp. dominate the vaginal niche but can also be involved in other vaginal dysbiosis, such as cytolytic vaginosis (CV), which remains poorly studied. It is characterized by a cryptic symptomatology, that often confounds the clinic. Goals: The aim of this work was to search for the etiological agent of CV, by studying the vaginal microbiome and metabolomics of women afflicted with this disease and compare it with women with other clinical diagnostic. Methods: Twenty-one vaginal washes have been collected from women attending a gynaecology consultation of a private clinic. The samples were categorized according with clinical diagnosis at the time of sampling (CV, 11; vulvovaginal candidosis, 8; Healthy, 2). The distribution of bacterial species, and their prevalence was assessed by next-generation sequencing of the 16S V4 region. In addition, total lactate D-lactic acid and L-lactic acid was quantified in all washes by a commercial kit, as well as lactate dehydrogenase (LDH) activity. Results: L. crispatus was dominant (>70%) in all CV samples. Lactate was increased in CV in comparison with other cases. The presence of D-lactic acid isomer was associated with presence of L. crispatus. LDH activity was increased in vaginal washes that tested positive for the presence of L. crispatus, however no direct association was found with CV cases. Discussion/Conclusions: The microbiome of women afflicted with CV was dominated in all cases by L. crispatus, contrarily with the results obtained for women diagnosed with other clinical symptomatology. In addition, the finding that an increase in D-lactic acid is associated with CV patients can be related to the role of L. crispatus in CV. The determination of LDH activity did not correlate exclusively with CV cases. On the other hand, D-lactic acid and total lactate quantification could be used as a valuable biomarker to diagnose this cryptic vaginal infection.info:eu-repo/semantics/publishedVersio

Assessment of Lactobacillus crispatus´ role in vaginal infections: friend or foe?

Lactobacillus spp. are the most prevalent microorganisms in the vaginal milleu. Several probiotic mechanisms have been associated with Lactobacillus, but the most relevant one is lactate production – resulting in a low pH value, typical of the healthy vagina. However, its pathogenic state is still unknown. We aim to elucidate the role of a commensal vaginal microorganism, Lactobacillus crispatus, in vaginal infections. Twenty-four vaginal washes have been collected from women attending a gynaecology consultation of a private clinic. The samples were categorized according with clinical diagnosis at the time of sampling. The distribution of bacterial species, and their prevalence was assessed by Next-Generation Sequencing of the 16S rRNA V4 region. In addition, L- and D-lactate concentration, and LDH enzymatic activity was quantified in all washes by a commercial kit. Detection of Candida spp. was performed by PCR. L. crispatus was dominant (>70%) in 11 out of 24 samples – diagnosed for vaginal atrophy (VA, 3), cytolytic vaginosis (CV, 2) and lactobacillosis (LB, 2). Lactate was increased in CV, LB and VA cases only. The remaining samples, diagnosed for vulvovaginal candidosis in its majority, had lower prevalence of L. crispatus; and lower to moderate lactate metabolite. There was not a direct relationship between LDH enzymatic activity and clinical diagnosis. L. crispatus dominance, associated with increased lactate production, was observed in CV, LB and VA cases. These results indicate that this microorganism might have a role in dysbiosis of the vagina associated with these specific pathologies.info:eu-repo/semantics/publishedVersio

Role of Lactobacillus crispatus in vaginal infections: insights from metagenomics and metabolomics studies

Objective: Lactobacillus spp. constitute the most prevalent bacterial microorganism in the vaginal milleu. Several probiotic mechanisms have been associated with Lactobacillus, but the most relevant one is lactate production – resulting in a low pH value, typical of the healthy vagina. We aim to elucidate the role of a native vaginal microorganism, Lactobacillus crispatus, in vaginal infections. Methods: Twenty-four vaginal washes have been collected from women (mean 38±14 years of age) attending a gynecology consultation of a private clinic. The samples were categorized according with clinical diagnosis at the time of sampling. The distribution of bacterial species, and their prevalence was assessed by next-generation sequencing of the 16S V4 region. In addition, lactate was quantified in all washes by a commercial kit. Results: L. crispatus was dominant (>70%) in 11 out of 24 samples – diagnosed for vaginal atrophy (VA, 3), cytolytic vaginosis (CV, 2), lactobacillosis (LB, 2) and vulvovaginal candidosis (VVC, 1). For three samples, diagnostic was inconclusive. Lactate was increased in CV, LB and VA cases only. One sample, diagnosed with VVC had 23% dominance of L. crispatus; and lower lactate metabolite. L. crispatus was not detected in the remaining 12 cases (6 VVC, 4 VA, 1 healthy and 1 inconclusive). Of these, only 5 samples had moderate lactate metabolite. Conclusions: L. crispatus dominance, associated with increased lactate production, was observed in CV, LB and VA cases. These results indicate that this microorganism might have a role in dysbiosis of the vagina associated with these specific pathologies.info:eu-repo/semantics/publishedVersio

Optimization of in vitro model for growth of Gardnerella vaginalis biofilm

Bacterial vaginosis (BV) is the most common vaginal disorder in women of reproductive age. Despite its high prevalence, BV aetiology remains elusive. Recently, it was described that BV occurrence involves the presence of an adherent biofilm on vaginal epithelium, being Gardnerella vaginalis the predominant bacterial species. Furthermore, this bacterial biofilm persists on the vaginal epithelium after therapy with metronidazole, suggesting that G. vaginalis biofilm plays a key role in treatment failure and recurrence of BV. Despite the relevance of G. vaginalis biofilms in the BV pathogenesis, there has not been detailed information addressing the environmental conditions influence in G. vaginalis biofilm formation. In this sense, we designed a series of in vitro assays in order to evaluate the influence of some common culture conditions on biofilm formation of this bacterial species. Thus, we used four G. vaginalis strains isolated from women with BV and compared their biofilm-forming ability using distinct culture conditions, namely inoculum concentration, incubation period, feeding conditions and culture medium composition. Our results showed that growth conditions strongly influenced G. vaginalis biofilm formation and that biofilm formation was enhanced when starting the culture with a higher inoculum, supplemented the medium with glucose and using a fed-batch system. To conclude, this study provides new insights about optimized culture conditions for G. vaginalis biofilm formation, which is extremely important for future fundamental studies involving this bacterial community. This work was co-funded by FCT project RECI/BBB-EBI/0179/2012 (FCOMP-01-0124-FEDER-027462), FCT Strategic Project PEst-C/SAU/UI0709/2011 and by QREN, FEDER, ON2 project (NORTE-07-0124-FEDER-000027)

Opioids, sex and gender

JUSTIFICATIVA E OBJETIVOS: Sexo é um fator importante na modulação da experiência dolorosa. Evidências significativas têm demonstrado que a experiência à dor difere entre homens e mulheres, bem como na resposta à ativação do sistema opioide e seus efeitos analgésicos. Há evidências que as mulheres têm menor limiar que os homens para alguns estímulos álgicos. Os neurotransmissores opioides e seus receptores estão centralmente envolvidos na resposta ao estresse, na supressão à dor e na ação dos analgésicos opioides. O objetivo deste estudo foi analisar a relação entre sexo, gênero e sistema opioide e discutir a relevância de um dos aspectos mais intrigantes da fisiologia da dor: a presença da diferença entre sexo e gênero, sistema opioide e as respostas da analgesia por opioides. CONTEÚDO: Uma revisão da literatura sobre opioides, sexo e gênero, cujo objetivo foi mostrar dados atuais sobre a experiência dolorosa entre homens e mulheres, a ativação opioide central e a resposta aos analgésicos opioides. CONCLUSÃO: Os dados disponíveis na literatura, e os trabalhos em andamento indicam que o sexo provavelmente seja responsável pelas diferenças à analgesia opioide em homens e mulheres, mas a direção e a magnitude destas diferenças dependem de variáveis que se interagem. Como fatores importantes que interagem na percepção dolorosa e na resposta analgésica opioide, devem ser levados em consideração os fatores socioculturais e biológicos, incluindo as variações hormonais em mulheres e a presença do hormônio masculino nos homens.BACKGROUND AND OBJECTIVES: Sex is a major factor for pain modulation. Significant evidences have shown that pain experience is different between men and women, as well as the response to opioid system activation and its analgesic effects. There are evidences that women have lower pain threshold compared to men. Opioid neurotransmitters and their receptors are centrally involved with stress response, pain suppression and opioids analgesic action. This study aimed to illustrate the relationship between sex, gender and the opioid system and to discuss the relevance of one of the most intriguing aspects of pain physiology: differences between sex and gender, opioid system and opioids analgesic response. CONTENTS: A literature review on opioids, sex and gender, aiming to show current data on pain experience between men and women, central opioid activation and response to opioids. CONCLUSION: Data in the literature and ongoing studies indicate that sex is probably responsible for differences in opioid analgesia between men and women, but the direction and magnitude of such differences depend on interacting variables. Socio-cultural and biological factors, including hormonal variations in women and the presence of male hormone in men are important factors interacting with pain perception and opioid analgesic response

Acute toxicity of plant extracts towards Daphnia magna

The demand for natural based products for the cosmetics industry is increasing sharply and therefore the search for new alternatives to the traditionally used plants is growing. These alternative plants can be an important source of bioactive compounds under a circular economy approach. Within the framework of Inovep project, several plant species, some of which autochthonous from Portugal, were identified as potential sources of bioactive compounds, namely: gum rockrose (Cistus ladanifer), curry plant (Helichrysum italicum), hop (Humulus lupulus), Mediterranean thyme (Thymbra capitata) and basil (Ocimum basilicum). Considering the potential future use of these plant extracts by the industry, it is necessary to assess the risk associated with their introduction into the ecosystem. This work aims to evaluate the toxic effects of different extracts of these five species. Acute toxicity tests using the model organism Daphnia magna were performed and the immobilization after 24 and 48 hours of exposure was evaluated. The EC50 (the concentration estimated to immobilize 50 per cent of the Daphnia) varied from 201.8±3.28 x10-5 mg.L-1 at 24 hours and 199.5±5.9410x10-5 mg.L-1 at 48 hours for Cistus ladanifer extract and 11.2±0.2403 mg.L- 1 at 24 hours and 11.2±0.4095 mg.L-1 at 48 hours for Thymbra capitata essential oil. For Humulus lupulus, Helichrysum italicum, Thymbra capitata in the form of hidrolate, and Ocimum basilicum no immobilization was observed until the highest concentrations tested for various types of extracts, suggesting these extracts present low to no risk towards D. magna

Preliminary studies for the aplication of Thymbra capitata essential oil as potential antimicrobial agent in Bacterial Vaginosis

Bacterial vaginosis (BV) is the most common gynecological clinical condition in women in reproductive age, and has been associated with an increased risk of development of preterm labor, spontaneous abortion, and several sexually transmitted diseases such as HIV [1]. BV is not only a condition that involves excessive growth of anaerobic microorganisms, but also involves the presence of an adherent biofilm [2]. Despite its high prevalence, the aetiology of BV remains unknown, but frequently Gardnerella vaginalis is the main microorganism associated with this disorder [3], that is often the main species in biofilms presented in BV [4]. These results led to the theory that the microorganisms that form biofilms may be relevant for the aetiology of BV [5]. Traditionally, the treatment of BV involves the antibiotics metronidazole, clindamicin or tinidazole [6]. However, the recurrence rate remains high and this can be related to the biofilm-forming ability of G. vaginalis [5]. In this context, recently it was reported that natural compounds, such as Thymbra capitata essential oil, can reduce the activity of biofilms [7]. To test if T. capitata could be used in the treatment of BV, we determined the biofilm-forming ability and the effect of essential oil in seven strains of G. vaginalis isolated from women with BV. Thus, the biofilm-forming ability was assessed under anaerobic conditions for 48 hours, allowing each strain to form biofilm. These assays revealed that in fact, G. vaginalis strains produced moderate to tenacious biofilm. We then determined the sensibility of the tested bacteria to the common antibiotics used to treat BV. Not surprisingly, most of strains were resistant of metronidazole and tinidazole which are more commonly administered. However half of strains were sensitive to clindamicin. We then evaluated the antibacterial activity of essential oil upon G. vaginalis strains, determining minimum inhibitory concentration (MIC) and minimum lethal concentration (MLC). The T. capitata essential oil exhibited a potent anti-G. vaginalis effect, which was confirmed by the low values of MIC (0.16 µL.mL-1) and MLC (0.16 - 0.31 µL.mL-1). To conclude, these results showed that T. capitata essential oil presents a strong antibacterial activity upon antibiotic resistant and biofilm forming of G. vaginalis strains, reinforcing the potential interest of this essential oil as antimicrobial agent in the treatment of BV

Anti-inflammatory potential of Portuguese thermal waters

In light of Medical Hydrology, thermal waters (TW) are all-natural mineral waters that emerge inside a thermal resort and have therapeutic applications. Their beneficial effect has been empirically recognized for centuries, being indicated for symptom alleviation and/or treatment of several diseases, almost all associated with inflammation. Indeed, an anti-inflammatory effect has been attributed to many different Portuguese TW but there is no scientific validation supporting this empiric knowledge. In the present study, we aimed to investigate the anti-inflammatory properties of 14 TW pertaining to thermal centers located in the Central Region of Portugal, and grouped according to their ionic profile. Mouse macrophage cells stimulated with lipopolysaccharide (LPS), a Toll-like receptor 4 agonist, were exposed to culture medium prepared in TW. Metabolism, nitric oxide (NO) production, inducible nitric oxide synthase (iNOS) expression levels and the scavenging capacity of TW, were investigated in vitro. 11 out of 14 TW reduced NO production and/or iNOS expression, and/or scavenging activity, in macrophages exposed to LPS. The sulphated/calcic TW did not show any effect on at least one of the inflammatory parameters evaluated. Two sulphurous/bicarbonate/sodic TW and the sulphurous/chlorinated/sodic TW promoted an increase in NO production and/or iNOS expression. Our results validate, for the first time, the anti-inflammatory properties of Portuguese TW, supporting their therapeutic use in the treatment of inflammation-related diseases and promoting their putative application in cosmetic products and medical devices

Aptamer-Functionalized Gold Nanoparticles for Drug Delivery to Gynecological Carcinoma Cells

Cervical cancer is one of the most common cancers and is one of the major cause of deaths in women, especially in underdeveloped countries. The patients are usually treated with surgery, radiotherapy, and chemotherapy. However, these treatments can cause several side effects and may lead to infertility. Another concerning gynecologic cancer is endometrial cancer, in which a high number of patients present a poor prognosis with low survival rates. AS1411, a DNA aptamer, increases anticancer therapeutic selectivity, and through its conjugation with gold nanoparticles (AS1411-AuNPs) it is possible to improve the anticancer effects. Therefore, AS1411-AuNPs are potential drug carriers for selectively delivering therapeutic drugs to cervical cancer. In this work, we used AS1411-AuNPs as a carrier for an acridine orange derivative (C8) or Imiquimod (IQ). The AS1411 aptamer was covalently bound to AuNPs, and each drug was associated via supramolecular assembly. The final nanoparticles presented suitable properties for pharmaceutical applications, such as small size, negative charge, and favorable drug release properties. Cellular uptake was characterized by confocal microscopy and flow cytometry, and effects on cellular viability were determined by MTT assay. The nanoparticles were then incorporated into a gel formulation of polyethylene glycol, suitable for topical application in the female genital tract. This gel showed promising tissue retention properties in Franz cells studies in the porcine vaginal epithelia. These findings suggest that the tested nanoparticles are promising drug carriers for cervical cancer therapy

Levantamento sorológico de anticorpos anti-Neospora caninum e anti-Toxoplasma gondii em ovinos no interior de São Paulo, SP: Serological survey of anti-Neospora caninum and anti-Toxoplasma gondii antibodies in sheep in the interior of São Paulo, SP

A ovinocultura no Brasil é uma atividade lucrativa, produzindo no ano de 2019 em torno de 19,7 milhões de cabeças. No entanto, algumas doenças podem trazer prejuízo ao produtor, como por exemplo, a neosporose e a toxoplasmose, que são responsáveis por perdas econômicas significativas especialmente devido ao abortamento. A neosporose é uma importante enfermidade causada pelo protozoário Neospora caninum. A toxoplasmose é uma zoonose de distribuição mundial, causada pelo Toxoplasma gondii, protozoário unicelular. Nesse contexto, o presente estudo teve como objetivo realizar a pesquisa de anticorpos anti-Neospora caninum e anti-Toxoplasma gondii em ovinos da região de Indaiatuba, interior do estado de São Paulo. Para tanto, foram colhidas amostras de sangue de 100 animais de diferentes idade e sexo. Para a detecção de anticorpos foram empregadas a reação de imunofluorescência indireta (RIFI). Para a detecção de IgG anti-Neospora caninum foi utilizado como antígeno taquizoítos da cepa NC-1. Como anticorpo secundário foi utilizado o conjugado comercial anti-ovino IgG (Sigma, USA, F7887) marcado com isotiocianato de fluoresceína. Para a detecção de anticorpos IgG anti-Toxoplasma gondii foi adotado o protocolo preconizado por Camargo (1964), utilizando-se anticorpos anti-IgG-ovino (Sigma®) conjugado ao isotiocianato de fluoresceína, com ponto de corte 64, utilizando-se como antígeno, taquizoítos da cepa RH. Em todas as reações foram incluídos controle positivo e negativo, previamente conhecidos. A RIFI foi realizada no Departamento de Medicina Veterinária (DMV), da Universidade Federal Rural de Pernambuco (UFRPE). Dos 100 animais testados para Neospora caninum, os resultados do teste de sorodiagnóstico indicaram 12% (12/100) de animais positivos, sendo 25% (3/12) filhotes, 25% (3/12) jovens e 50% (6/12) adultos. Em relação ao sexo, 16,70% (2/12) eram machos e 83,30% (10/12) fêmeas. Os resultados para Toxoplasma gondii indicaram 37% (37/100) de animais soropositivos. Em relação a idade, observou-se 16,22% (6/37) filhotes, 2,70% (1/37) jovens e 81,08% (30/37) adultos. Em relação ao sexo verificou-se que dos animais positivos, 27,02% (10/37) eram machos e 72,97% (27/37) fêmeas. Com base nos resultados do presente estudo concluiu-se que a ocorrência de animais com neoporose e toxoplasmose é expressiva na propriedade pesquisada. Novos estudos devem ser conduzidos para verificação dos fatores de riscos a fim de adotarem medidas de prevenção e controle