Assessing synergistic interaction between Gardnerella vaginalis and other urogenital pathogens

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)

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)

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

Long‐term outcomes and molecular analysis of a large cohort of patients with 46,XY disorder of sex development due to partial gonadal dysgenesis

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/145419/1/cen13717_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/145419/2/cen13717.pd

Convalescent plasma for COVID-19 in hospitalised patients : an open-label, randomised clinical trial

Background: The effects of convalescent plasma (CP) therapy in hospitalised patients with coronavirus disease 2019 (COVID-19) remain uncertain. This study investigates the effect of CP on clinical improvement in these patients. Methods: This is an investigator-initiated, randomised, parallel arm, open-label, superiority clinical trial. Patients were randomly (1:1) assigned to two infusions of CP plus standard of care (SOC) or SOC alone. The primary outcome was the proportion of patients with clinical improvement 28 days after enrolment. Results: A total of 160 (80 in each arm) patients (66.3% critically ill, 33.7% severely ill) completed the trial. The median (interquartile range (IQR)) age was 60.5 (48–68) years; 58.1% were male and the median (IQR) time from symptom onset to randomisation was 10 (8–12) days. Neutralising antibody titres >1:80 were present in 133 (83.1%) patients at baseline. The proportion of patients with clinical improvement on day 28 was 61.3% in the CP+SOC group and 65.0% in the SOC group (difference −3.7%, 95% CI −18.8–11.3%). The results were similar in the severe and critically ill subgroups. There was no significant difference between CP+SOC and SOC groups in pre-specified secondary outcomes, including 28-day mortality, days alive and free of respiratory support and duration of invasive ventilatory support. Inflammatory and other laboratory marker values on days 3, 7 and 14 were similar between groups. Conclusions: CP+SOC did not result in a higher proportion of clinical improvement on day 28 in hospitalised patients with COVID-19 compared to SOC alone

Mortality from gastrointestinal congenital anomalies at 264 hospitals in 74 low-income, middle-income, and high-income countries: a multicentre, international, prospective cohort study

Summary Background Congenital anomalies are the fifth leading cause of mortality in children younger than 5 years globally. Many gastrointestinal congenital anomalies are fatal without timely access to neonatal surgical care, but few studies have been done on these conditions in low-income and middle-income countries (LMICs). We compared outcomes of the seven most common gastrointestinal congenital anomalies in low-income, middle-income, and high-income countries globally, and identified factors associated with mortality. Methods We did a multicentre, international prospective cohort study of patients younger than 16 years, presenting to hospital for the first time with oesophageal atresia, congenital diaphragmatic hernia, intestinal atresia, gastroschisis, exomphalos, anorectal malformation, and Hirschsprung’s disease. Recruitment was of consecutive patients for a minimum of 1 month between October, 2018, and April, 2019. We collected data on patient demographics, clinical status, interventions, and outcomes using the REDCap platform. Patients were followed up for 30 days after primary intervention, or 30 days after admission if they did not receive an intervention. The primary outcome was all-cause, in-hospital mortality for all conditions combined and each condition individually, stratified by country income status. We did a complete case analysis. Findings We included 3849 patients with 3975 study conditions (560 with oesophageal atresia, 448 with congenital diaphragmatic hernia, 681 with intestinal atresia, 453 with gastroschisis, 325 with exomphalos, 991 with anorectal malformation, and 517 with Hirschsprung’s disease) from 264 hospitals (89 in high-income countries, 166 in middleincome countries, and nine in low-income countries) in 74 countries. Of the 3849 patients, 2231 (58·0%) were male. Median gestational age at birth was 38 weeks (IQR 36–39) and median bodyweight at presentation was 2·8 kg (2·3–3·3). Mortality among all patients was 37 (39·8%) of 93 in low-income countries, 583 (20·4%) of 2860 in middle-income countries, and 50 (5·6%) of 896 in high-income countries (p<0·0001 between all country income groups). Gastroschisis had the greatest difference in mortality between country income strata (nine [90·0%] of ten in lowincome countries, 97 [31·9%] of 304 in middle-income countries, and two [1·4%] of 139 in high-income countries; p≤0·0001 between all country income groups). Factors significantly associated with higher mortality for all patients combined included country income status (low-income vs high-income countries, risk ratio 2·78 [95% CI 1·88–4·11], p<0·0001; middle-income vs high-income countries, 2·11 [1·59–2·79], p<0·0001), sepsis at presentation (1·20 [1·04–1·40], p=0·016), higher American Society of Anesthesiologists (ASA) score at primary intervention (ASA 4–5 vs ASA 1–2, 1·82 [1·40–2·35], p<0·0001; ASA 3 vs ASA 1–2, 1·58, [1·30–1·92], p<0·0001]), surgical safety checklist not used (1·39 [1·02–1·90], p=0·035), and ventilation or parenteral nutrition unavailable when needed (ventilation 1·96, [1·41–2·71], p=0·0001; parenteral nutrition 1·35, [1·05–1·74], p=0·018). Administration of parenteral nutrition (0·61, [0·47–0·79], p=0·0002) and use of a peripherally inserted central catheter (0·65 [0·50–0·86], p=0·0024) or percutaneous central line (0·69 [0·48–1·00], p=0·049) were associated with lower mortality. Interpretation Unacceptable differences in mortality exist for gastrointestinal congenital anomalies between lowincome, middle-income, and high-income countries. Improving access to quality neonatal surgical care in LMICs will be vital to achieve Sustainable Development Goal 3.2 of ending preventable deaths in neonates and children younger than 5 years by 2030

COVID-19 symptoms at hospital admission vary with age and sex: results from the ISARIC prospective multinational observational study

Background: The ISARIC prospective multinational observational study is the largest cohort of hospitalized patients with COVID-19. We present relationships of age, sex, and nationality to presenting symptoms. Methods: International, prospective observational study of 60 109 hospitalized symptomatic patients with laboratory-confirmed COVID-19 recruited from 43 countries between 30 January and 3 August 2020. Logistic regression was performed to evaluate relationships of age and sex to published COVID-19 case definitions and the most commonly reported symptoms. Results: ‘Typical’ symptoms of fever (69%), cough (68%) and shortness of breath (66%) were the most commonly reported. 92% of patients experienced at least one of these. Prevalence of typical symptoms was greatest in 30- to 60-year-olds (respectively 80, 79, 69%; at least one 95%). They were reported less frequently in children (≤ 18 years: 69, 48, 23; 85%), older adults (≥ 70 years: 61, 62, 65; 90%), and women (66, 66, 64; 90%; vs. men 71, 70, 67; 93%, each P &lt; 0.001). The most common atypical presentations under 60 years of age were nausea and vomiting and abdominal pain, and over 60 years was confusion. Regression models showed significant differences in symptoms with sex, age and country. Interpretation: This international collaboration has allowed us to report reliable symptom data from the largest cohort of patients admitted to hospital with COVID-19. Adults over 60 and children admitted to hospital with COVID-19 are less likely to present with typical symptoms. Nausea and vomiting are common atypical presentations under 30 years. Confusion is a frequent atypical presentation of COVID-19 in adults over 60 years. Women are less likely to experience typical symptoms than men

Prevalência da Gardnerella vaginalis em mulheres grávidas portuguesas. Novas estratégias terapêuticas para a bacteriose vaginal

Tese de Doutoramento em Engenharia Química e BiológicaA nível mundial, a bacteriose vaginal (BV) é a infeção vaginal mais comum em mulheres em idade reprodutiva, associando-se a complicações na gravidez. Esta infeção carateriza-se pela diminuição da população microbiana vaginal normal (predominantemente lactobacilar) e a simultânea proliferação de bacterias anaeróbias. Devido à sua natureza polimicrobiana, a etiologia da BV permanece por esclarecer. No entanto, o desenvolvimento da BV está fortemente associado ao desenvolvimento de biofilmes polimicrobianos, compostos maioritariamente por Gardnerella vaginalis. Atualmente, os tratamentos disponíveis para a BV são insatisfatórios. Os principais objetivos deste trabalho foram determinar a prevalência da G. vaginalis e da BV em mulheres grávidas portuguesas e explorar novas estratégias terapêuticas para esta infeção. Para tal, foi desenvolvido um estudo transversal com mulheres grávidas da região norte de Portugal. Verificouse uma baixa prevalência da BV (3.88%) mas uma elevada colonização vaginal por G. vaginalis (67.48%). Um elevado risco de colonização por G. vaginalis foi encontrado em mulheres com BV, com nível educacional básico, fumadoras e no segundo trimestre de gravidez. A história de doença crónica, parto prematuro e a colonização por G. vaginalis foram identificados como factores de risco para a BV. Foram, também, optimizadas as condições de formação de biofilmes in vitro por G. vaginalis. Demonstramos que os biofilmes de G. vaginalis apresentam maior tolerância aos antibióticos e a outros agentes antimicrobianos que culturas planctónicas. Por fim, avaliámos o potential terapêutico do óleo essencial de Thymbra capitata e o seu componente maioritário (carvacrol) contra biofilmes de G. vaginalis. Ambos exibiram uma potente atividade antimicrobiana contra culturas planctónicas de G. vaginalis. Contudo, apenas o óleo foi capaz de reduzir significativamente os biofilmes de G. vaginalis e, adicionalmente, apresentou baixa atividade antimicrobiana sob os lactobacilos vaginais. Assim, o óleo essencial T. capitata destaca-se como um promissor agente terapêutico para a BV.Worldwide, bacterial vaginosis (BV) is the most common gynaecological infection in women in reproductive age, being related with pregnancy complications. This vaginal infection is characterized by the replacement of beneficial microflora (often lactobacilli) and concurrent overgrowth of anaerobic bacteria. Being polymicrobial in nature, BV etiology remains unclear. Nevertheless, it is commonly accepted that BV involves the presence of multispecies biofilm, mainly composed by Gardnerella vaginalis. A major issue related to BV is the recurrent failure of conventionally antibiotic therapies. So, the main aims of this work were to determine the prevalence of G. vaginalis and BV in Portuguese pregnant women and to search for novel strategies to treat this vaginal infection. To achieve this, we first conducted a cross-sectional study among pregnant women in the Northern region of Portugal. Our epidemiological findings showed that BV frequency was low (3.88%) but G. vaginalis colonization was high (67.48%) among pregnant women. Furthermore, a higher risk of G. vaginalis colonization was found in women with BV, basic educational level, smokers and during the second trimester of pregnancy. In contrast, past history of chronic disease, preterm delivery and colonization by G. vaginalis were identified as BV risk factors during pregnancy. Next, we performed a series of in vitro assays in order to determine the influence of environmental conditions in G. vaginalis biofilm formation. Using optimized in vitro conditions, we demonstrated that G. vaginalis biofilms display a higher tolerance to antibiotics and other antimicrobial agents than planktonic cultures. Lastly, we evaluated the potential therapeutic of Thymbra capitata essential oil and its main component (carvacrol) against G. vaginalis biofilms. We found that both T. capitata essential oil and carvacrol displayed a potent antibacterial activity against G. vaginalis planktonic cultures. However, only the intact oil was able to reduce significantly G. vaginalis biofilms. Interestingly, it presented a lower antimicrobial activity on vaginal lactobacilli. Thus, T. capitata essential oil stands up as a promising therapeutic agent for BV.The work presented in this thesis was funded by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684), the project RECI/BBB-EBI/0179/2012 (FCOMP-01-0124-FEDER-027462) and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. DM acknowledges the FCT individual grant with reference SFRH/BD/87569/2012

Resumos em andamento - Direito

Resumos em andamento - Direit