Oral health of people with emotional disorders: A systematic review and meta-analysis.

OBJECTIVES: This study aims to determine the association between severe mental disorders and oral health among individuals over 18 years of age. METHODS: An electronic search was conducted in six electronic databases and gray literature. Qualitative and quantitative analyses were performed on studies that met the inclusion criteria. The methodology of the included studies was assessed using the Joanna Briggs Institute Critical Appraisal tool. A meta-analysis of proportions with a random effect was carried out. The certainty of evidence was evaluated using the GRADE tool. RESULTS: After searching the databases, 5,734 references were retrieved, and twenty articles were selected for synthesis. Considering the DMFT index between the groups with mental disorders and the control group, the values of the DMFT index were higher among individuals with schizophrenia [MD = 5.27; 95% CI = 4.13 - 6.42; I2 = 35%] and bipolar disorder [MD = 1.90; 95% CI = 0.87 - 2.93]. Values were lower among individuals with obsessive-compulsive disorder [MD = -0.85; 95% CI = -1.46-0.24]. The risk of bias was considered low for 16 studies, and four were classified with a moderate risk of bias. The certainty of evidence was very low. CONCLUSION: Patients with schizophrenia and bipolar disorder exhibit increased frequency in the number of decayed, missing, or filled teeth. There was no effect in relation to periodontal probing depth, plaque index, and TMD, but the evidence is still uncertain for this outcome. CLINICAL RELEVANCE: These findings underscore the need for a comprehensive health approach.

[Oral Anticoagulation and the Incidence of Stroke Associated with Atrial Fibrillation in Mainland Portugal: A Modelling Study]. / Anticoagulação Oral e Incidência de Acidente Vascular Cerebral Associado a Fibrilhação Auricular em Portugal Continental: Um Estudo de Modelação.

INTRODUCTION: Atrial fibrillation is the most prevalent persistent dysrhythmia, contributing to a significant social and economic burden. The main objective of this study was to evaluate the association between oral anticoagulant use and the incidence of stroke associated with atrial fibrillation, in mainland Portugal. METHODS: The number of episodes of inpatient care with a main diagnosis of stroke and an additional diagnosis of atrial fibrillation, occurring monthly between January 2012 and December 2018, in individuals aged 18 years or over, was extracted from the hospital morbidity database. The number of patients with an atrial fibrillation code documented in this database was used as a proxy for the prevalence of known atrial fibrillation. The number of anticoagulated patients was estimated from total medicine sales of vitamin K antagonists and novel oral anticoagulants (apixaban, dabigatran, edoxaban and rivaroxaban) in mainland Portugal. Descriptive analyses were performed, and seasonal autoregressive integrated moving average (SARIMA) models were built using the R software. RESULTS: The mean number of episodes of stroke per month was 522 (± 57). The number of anticoagulated patients increased gradually from 68 943 to 180 389 per month. The decreasing trend in the number of episodes has been observed since 2016, along with the increased use of new oral anticoagulants compared to vitamin K antagonists. The final model indicated that the increase in oral anticoagulation use between 2012 and 2018, in mainland Portugal, was associated with a decrease in the number of episodes of stroke associated with atrial fibrillation. It was estimated that the shift in the type of anticoagulation used, between 2016 and 2018, was associated with a reduction of 833 episodes of stroke in patients with atrial fibrillation (4.2%). CONCLUSION: The use of oral anticoagulation was associated with a reduced incidence of stroke in patients with atrial fibrillation in mainland Portugal. This reduction was more relevant in the period between 2016 and 2018, and is probably related with the introduction of the novel oral anticoagulants.

Introdução: A fibrilhação auricular é a disritmia persistente mais prevalente, tendo um importante impacto social e económico. O objetivo principal deste estudo foi avaliar a associação entre a utilização de anticoagulantes orais e a incidência de acidente vascular cerebral associado a fibrilhação auricular, em Portugal continental. Métodos: A base de dados de morbilidade hospitalar foi utilizada para a contabilização dos episódios de internamento com um diagnóstico principal de acidente vascular cerebral e um diagnóstico adicional de fibrilhação auricular, ocorridos durante cada mês do período em análise (janeiro de 2012 a dezembro de 2018), em indivíduos com idade igual ou superior a 18 anos. O número de doentes com registo de fibrilhação auricular presentes nesta base de dados foi utilizado como um proxy da prevalência de fibrilhação auricular conhecida. O número de doentes anticoagulados foi estimado a partir das estatísticas das vendas de antagonistas da vitamina K e novos anticoagulantes orais (apixabano, dabigatrano, edoxabano e rivaroxabano) em Portugal continental. Foi realizada uma análise descritiva das variáveis, construindo-se depois modelos auto-regressivos integrados de médias móveis sazonais (seasonal autoregressive integrated moving average, SARIMA), com recurso ao software R. Resultados: Ocorreram, em média, 522 (± 57) episódios de acidente vascular cerebral por mês. Verificou-se um aumento gradual do número de doentes anticoagulados, passando de 68 943 para 180 389, por mês. A tendência decrescente no número de episódios verificou-se a partir de 2016, a par da maior utilização dos novos anticoagulantes orais, comparativamente aos antagonistas da vitamina K. O modelo final estimado indicou que o aumento do consumo de anticoagulação oral entre 2012 e 2018 em Portugal continental foi associado a um decréscimo do número de acidentes vasculares cerebrais associados a fibrilhação auricular. Estimou-se que, entre 2016 e 2018, a mudança no tipo de anticoagulação se associou a uma redução de 833 episódios de acidentes vascular cerebrais em doentes com fibrilhação auricular (4,2%). Conclusão: A anticoagulação oral associou-se à redução da incidência de acidente vascular cerebral em doentes com fibrilhação auricular, em Portugal continental. Esta redução foi mais relevante no período 2016 a 2018, em provável relação com a introdução dos novos anticoagulantes orais.

Glioblastoma Factors Increase the Migration of Human Brain Endothelial Cells In Vitro by Increasing MMP-9/CXCR4 Levels.

BACKGROUND/AIM: Glioblastoma (GB) is the most aggressive type of tumor in the central nervous system and is characterized by resistance to therapy and abundant vasculature. Tumor vessels contribute to the growth of GB, and the tumor microenvironment is thought to influence tumor vessels. We evaluated the molecular communication between human GB cells and human brain microvascular endothelial cells (HBMEC) in vitro. MATERIALS AND METHODS: We investigated whether GB-conditioned media (GB-CM) influenced HBMEC proliferation and migration, as well as the levels of MMP-9, CXCL12, CXCR4, CXCR7, VEGFs, VEGFR-2, and WNT5a in HBMEC. RESULTS: Although HBMEC proliferation was not modified, increased HBMEC migration was detected after GB-CM treatment. Furthermore, treatment of HBMEC with GB-CM resulted in increased levels of MMP-9 and CXCR4. The levels of WNT5a, VEGFs and VEGFR-2 were not affected. CONCLUSION: GB-secreted factors lead to increased endothelial cell migration and to increased levels of MMP-9 and CXCR4.

Biofilm formation of Brazilian meticillin-resistant Staphylococcus aureus strains: prevalence of biofilm determinants and clonal profiles.

Biofilms plays an important role in medical-device-related infections. This study aimed to determine the factors that influence adherence and biofilm production, as well as the relationship between strong biofilm production and genetic determinants in clinical isolates of meticillin-resistant Staphylococcus aureus (MRSA). Fifteen strains carrying different chromosomal cassettes recovered from hospitalized patients were selected; five SCCmecII, five SCCmecIII and five SCCmecIV. The SCCmec type, agr group and the presence of the virulence genes (bbp, clfA, icaA, icaD, fnbB, bap, sasC and IS256) were assessed by PCR. PFGE and multilocus sequence typing (MLST) techniques were also performed. The initial adhesion and biofilm formation were examined by quantitative assays. The surface tension and hydrophobicity of the strains were measured by the contact angle technique to evaluate the association between these parameters and adhesion ability. SCCmecIII and IV strains were less hydrophilic, with a high value for the electron acceptor parameter and higher adhesion in comparison with SCCmecII strains. Only SCCmecIII strains could be characterized as strong biofilm producers. The PFGE showed five major pulsotypes (A-E); however, biofilm production was related to the dissemination of one specific PFGE clone (C) belonging to MLST ST239 (Brazilian epidemic clonal complex). The genes agrI, fnbB and IS256 in SCCmecIII strains were considered as genetic determinants associated with strong biofilm-formation by an ica-independent biofilm pathway. This study contributes to the understanding of biofilm production as an aggravating factor potentially involved in the persistence and severity of infections caused by multidrug-resistant MRSA belonging to this genotype.

NaOCl effect on biofilm produced by Staphylococcus aureus isolated from the milking environment and mastitis infected cows / Efeito do hipoclorito de sódio em biofilmes produzidos por Staphylococcus aureus isolados do ambiente de ordenha e de vacas com mastite

Biofilms constitute a physical barrier, protecting the encased bacteria from detergents and sanitizers. The objective of this work was to analyze the effectiveness of sodium hypochlorite (NaOCl) against strains of Staphylococcus aureus isolated from raw milk of cows with subclinical mastitis and Staphylococcus aureus isolated from the milking environment (blowers and milk conducting tubes). The results revealed that, in the presence of NaOCl (150ppm), the number of adhered cells of the twelve S. aureus strains was significantly reduced. When the same strains were evaluated in biofilm condition, different results were obtained. It was found that, after a contact period of five minutes with NaOCl (150ppm), four strains (two strains from milk , one from the blowers and one from a conductive rubber) were still able to grow. Although with the increasing contact time between the bacteria and the NaOCl (150ppm), no growth was detected for any of the strains. Concerning the efficiency of NaOCl on total biofilm biomass formation by each S. aureus strain, a decrease was observed when these strains were in contact with 150 ppm NaOCl for a total period of 10 minutes. This study highlights the importance of a correct sanitation protocol of all the milk processing units which can indeed significantly reduce the presence of microorganisms, leading to a decrease of cow´s mastitis and milk contamination.

Biofilmes são constituídos de bactérias aderidas a uma superfície e aderidas entre si envolvidas por um polissacarídeo de constituição proteica, lipídica e glicídica que conferem uma barreira física às bactérias dentro deste microambiente. O objetivo deste trabalho foi analisar a eficácia do hipoclorito de sódio (NaOCl) contra estirpes de Staphylococcus aureus isoladas de leite cru de vacas com mastite subclínica e Staphylococcus aureus isolados do ambiente de ordenha (borrachas de ordenhadeiras e mangueiras condutoras de leite). Os resultados revelaram que, na presença de hipoclorito de sódio (150ppm), o número de células aderidas das 12 estirpes de S. aureus analisadas foi significativamente reduzido. Quando as mesmas estirpes foram avaliadas em condições de biofilme, diferentes resultados foram obtidos. Verificou-se que, após um período de contato de cinco minutos com NaOCl (150ppm), quatro estirpes (duas estirpes de leite, uma estirpe das borrachas das ordenhadeiras e uma estirpe de uma mangueira condutora de leite) ainda eram capazes de crescer. Com o aumento do tempo de contato do hipoclorito e as bactérias, cada vez maior, na concentração de 150ppm, não foi detectado o crescimento das estirpes. Em relação à eficácia do NaOCl na formação total da biomassa do biofilme por cada uma das estirpes de S. aureus, observou-se decréscimo da biomassa dos biofilmes quando estas estirpes estavam em contato com o NaOCl na concentração de 150ppm durante um tempo total de 10 minutos. O estudo demonstra a importância de um protocolo de saneamento correto de todas as unidades de processamento de leite, que pode, efetivamente, reduzir a presença de microrganismos de forma significativa, conduzindo a uma diminuição da mastite e da contaminação do leite.

Minimum information about a biofilm experiment (MIABiE): standards for reporting experiments and data on sessile microbial communities living at interfaces.

The minimum information about a biofilm experiment (MIABiE) initiative has arisen from the need to find an adequate and scientifically sound way to control the quality of the documentation accompanying the public deposition of biofilm-related data, particularly those obtained using high-throughput devices and techniques. Thereby, the MIABiE consortium has initiated the identification and organization of a set of modules containing the minimum information that needs to be reported to guarantee the interpretability and independent verification of experimental results and their integration with knowledge coming from other fields. MIABiE does not intend to propose specific standards on how biofilms experiments should be performed, because it is acknowledged that specific research questions require specific conditions which may deviate from any standardization. Instead, MIABiE presents guidelines about the data to be recorded and published in order for the procedure and results to be easily and unequivocally interpreted and reproduced. Overall, MIABiE opens up the discussion about a number of particular areas of interest and attempts to achieve a broad consensus about which biofilm data and metadata should be reported in scientific journals in a systematic, rigorous and understandable manner.

Adhesion of Candida biofilm cells to human epithelial cells and polystyrene after treatment with silver nanoparticles.

This study investigated the adhesion to human epithelial cells and polystyrene surface of viable yeasts recovered from Candida biofilms treated with silver nanoparticles (SN). Biofilm resuspended Candida cells were added to HeLa cells or to empty wells of microtiter plates and the adhesion was verified using crystal violet staining. The adhesion of Candida cells was significantly reduced, mainly when biofilms were pretreated with 54 µg/mL SN. These new findings allow to conclude that SN may induce changes in viable yeasts, which can decrease the dissemination of Candida infections, mainly in susceptible patients.

Feed optimization in fed-batch culture.

Fed-batch processes are a current preference for the production of recombinant proteins in mammalian cells. The use of nutrient feeding prevents the depletion of important medium components and results in improved culture longevity and high cell and product yields. To take maximum advantage of these effects, it is important to optimize the fed-batch process for each application. In this chapter, a simple strategy for fed-batch optimization is described, consisting of the development of a feed medium based on spent media analysis and the establishment of a feeding strategy that consists of adding variable volumes of feed media at specific intervals, after off-line measurement of the concentration of a reference nutrient.

Evaluation of solid and porous microcarriers for cell growth and production of recombinant proteins.

Microcarrier technology opened new perspectives for anchorage-dependent cell culture, by providing increased surface areas for cell adhesion and proliferation, and therefore improving both cell and product yields obtained in these cultures. The establishment of a successful microcarrier culture depends on many factors, such as the type of microcarrier, the cells, and the culture conditions. In this chapter, the basic steps required for the evaluation and optimization of a microcarrier culture for the purpose of production of recombinant proteins are described, for both solid and porous microcarriers.

Glycosylation: impact, control and improvement during therapeutic protein production.

The emergence of the biopharmaceutical industry represented a major revolution for modern medicine, through the development of recombinant therapeutic proteins that brought new hope for many patients with previously untreatable diseases. There is a ever-growing demand for these therapeutics that forces a constant technological evolution to increase product yields while simultaneously reducing costs. However, the process changes made for this purpose may also affect the quality of the product, a factor that was initially overlooked but which is now a major focus of concern. Of the many properties determining product quality, glycosylation is regarded as one of the most important, influencing, for example, the biological activity, serum half-life and immunogenicity of the protein. Consequently, monitoring and control of glycosylation is now critical in biopharmaceutical manufacturing and a requirement of regulatory agencies. A rapid evolution is being observed in this context, concerning the influence of glycosylation in the efficacy of different therapeutic proteins, the impact on glycosylation of a diversity of parameters/processes involved in therapeutic protein production, the analytical methodologies employed for glycosylation monitoring and control, as well as strategies that are being explored to use this property to improve therapeutic protein efficacy (glycoengineering). This work reviews the main findings on these subjects, providing an up-to-date source of information to support further studies.

Combined effect of linezolid and N-acetylcysteine against Staphylococcus epidermidis biofilms

Introduction: Staphylococcus epidermidis is an organism commonly associated with infections caused by biofilms. Biofilms are less sensible to antibiotics and therefore are more difficult to eradicate. Linezolid and N-acetylcysteine (NAC), have demonstrated to be active against gram-positive microorganisms. Therefore and since linezolid and NAC have different modes of action, the main objective of this work was to investigate the single and synergistic effect of linezolid and NAC against S. epidermidis biofilms. Methods: This work reports the in vitro effect of linezolid and NAC against S. epidermidis biofilms, treated with MIC (4 mgml−1) and 10×MIC of NAC, and MIC (1 μgml−1) and peak serum concentration (PS = 18μgml−1) of linezolid alone and in combination. After exposure of S. epidermidis biofilms to linezolid and/or NAC for 24 h, several biofilm parameters were evaluated, namely the number of cultivable cells [colony forming unit (CFU) enumeration], total biofilm biomass and cellular activity. Results: When tested alone, NAC at 10 × MIC was the most effective agent against S. epidermidis biofilms. However, the combination linezolid (MIC) + NAC (10×MIC) showed a synergistic effect and was the most biocidal treatment tested, promoting a 5 log reduction in the number of biofilm viable cells. Conclusion: This combination seems to be a potential candidate to combat infections caused by S. epidermidis biofilms, namely as a catheter lock solution therapy

Introducción: Staphylococcus epidermidis es un organismo comúnmente asociado con infecciones causadas por biofilms. Los biofilms son menos sensibles a los antibióticos y, por lo tanto, más difíciles de erradicar. El linezolid y la N-acetilcisteína (NAC) han demostrado ser activos contra los microorganismos grampositivos. Por lo tanto, y puesto que el linezolid y la NAC tienen diferentes modos de acción, el objetivo principal de este trabajo fue investigar el efecto individual y sinérgico del linezolid y la NAC frente a biofilms de S. epidermidis. Métodos: Este trabajo reporta el efecto in vitro del linezolid y la NAC solos y en combinación contra biofilms de S. epidermidis, tratados con NAC en las concentraciones de MIC (4mg·ml–1) y 10×MIC, y linezolid en las concentraciones MIC (1mg·ml–1) y concentración sérica máxima (PS = 18μg ml–1). Después de la exposición de los biofilms de S. epidermidis al linezolid y/o la NAC durante 24h, fueron evaluados varios parámetros del biofilm, a saber, el número de células cultivables (recuento de unidades formadoras de colonias [UFC]), la biomasa total del biofilm y la actividad celular. Resultados: Durante el ensayo solo, la NAC en la concentración de 10×MIC fue el agente más eficaz contra biofilms de S. epidermidis. Sin embargo, la combinación linezolid (MIC)+NAC (10×MIC) mostró un efecto sinérgico y fue el tratamiento con mayor efecto biocida, promoviendo una reducción logarítmica de 5 en el número de células viables del biofilm. Conclusión: Esta combinación parece ser un potencial candidato en el combate de las infecciones causadas por biofilms de S. epidermidis, es decir, como una terapia de solución de bloqueo del catéter

Update on the challenging role of biofilms in peritoneal dialysis.

Biofilms are commonly associated with an increased risk of patient infection. In peritoneal dialysis (PD), catheter associated infection, especially peritonitis, remains a clinically relevant problem. Although the presence of a biofilm is recognized in relapsing, repeat, and catheter-related peritonitis, it remains poorly characterized. In this review, an update on the role of biofilms in PD infections is presented. The emerging concept that host cells and tissue associated biofilms, in addition to the biofilms on the catheters themselves, contribute to the recalcitrance of infections is discussed. Furthermore, the evidence of biofilms on PD catheters, their developmental stages, and the possible influence of the PD environment are reviewed. The focus is given to ex vivo and in vitro studies that contribute to the elucidation of the interplay between host, microbial, and dialysis factors. The key issues that are still to be answered and the challenges to clinical practice are discussed.

Antifungal activity of silver nanoparticles in combination with nystatin and chlorhexidine digluconate against Candida albicans and Candida glabrata biofilms.

Although silver nanoparticles (SN) have been investigated as an alternative to conventional antifungal drugs in the control of Candida-associated denture stomatitis, the antifungal activity of SN in combination with antifungal drugs against Candida biofilms remains unknown. Therefore, the aim of this study was to evaluate the antifungal efficacy of SN in combination with nystatin (NYT) or chlorhexidine digluconate (CHG) against Candida albicans and Candida glabrata biofilms. The drugs alone or combined with SN were applied on mature Candida biofilms (48 h), and after 24 h of treatment their antibiofilm activities were assessed by total biomass quantification (by crystal violet staining) and colony forming units enumeration. The structure of Candida biofilms was analysed by scanning electron microscopy (SEM) images. The data indicated that SN combined with either NYT or CHG demonstrated synergistic antibiofilm activity, and this activity was dependent on the species and on the drug concentrations used. SEM images showed that some drug combinations were able to disrupt Candida biofilms. The results of this study suggest that the combination of SN with NYT or CHG may have clinical implications in the treatment of denture stomatitis. However, further studies are needed before recommending the use of these drugs safely in clinical situations.

N-acetylcysteine and vancomycin alone and in combination against staphylococci biofilm

INTRODUCTION: The ability of staphylococci to produce biofilm is an important virulence mechanism that allows bacteria both to adhere and to live on artificial surfaces and to resist to the host immune factors and antibiotics. Staphylococcal infections have become increasingly difficult to treat due their antibiotic resistance. Therefore, there is a continuous need for new and effective treatment alternatives against staphylococcal infections. The main goal of this study was to test N-acetylcysteine (NAC) and vancomycin alone and in combination against S. epidermidis and S. aureus biofilms. METHODS: Biofilms were treated with NAC at minimum inhibitory concentration (MIC) and 10 × MIC concentrations and vancomycin at MIC and peak serum concentrations. RESULTS: The use of NAC 10 × MIC alone showed a significant antibactericidal effect, promoting a 4-5 log10 CFU/ mL reduction in biofilm cells. The combination of NAC 10 × MIC with vancomycin (independently of the concentration used) reduced significantly the number of biofilm cells for all strains evaluated (5-6 log10). CONCLUSION: N-acetylcysteine associated to vancomycin can be a potential therapeutic strategy in the treatment of infections associated to biofilms of S. epidermidis or S. aureus.

Combined effect of linezolid and N-acetylcysteine against Staphylococcus epidermidis biofilms.

INTRODUCTION: Staphylococcus epidermidis is an organism commonly associated with infections caused by biofilms. Biofilms are less sensible to antibiotics and therefore are more difficult to eradicate. Linezolid and N-acetylcysteine (NAC), have demonstrated to be active against gram-positive microorganisms. Therefore and since linezolid and NAC have different modes of action, the main objective of this work was to investigate the single and synergistic effect of linezolid and NAC against S. epidermidis biofilms. METHODS: This work reports the in vitro effect of linezolid and NAC against S. epidermidis biofilms, treated with MIC (4mgml(-1)) and 10×MIC of NAC, and MIC (1µgml(-1)) and peak serum concentration (PS=18µgml(-1)) of linezolid alone and in combination. After exposure of S. epidermidis biofilms to linezolid and/or NAC for 24h, several biofilm parameters were evaluated, namely the number of cultivable cells [colony forming unit (CFU) enumeration], total biofilm biomass and cellular activity. RESULTS: When tested alone, NAC at 10×MIC was the most effective agent against S. epidermidis biofilms. However, the combination linezolid (MIC)+NAC (10×MIC) showed a synergistic effect and was the most biocidal treatment tested, promoting a 5log reduction in the number of biofilm viable cells. CONCLUSION: This combination seems to be a potential candidate to combat infections caused by S. epidermidis biofilms, namely as a catheter lock solution therapy.

Evaluation of macroporous and microporous carriers for CHO-K1 cell growth and monoclonal antibody production.

The emergence of microcarrier technology has brought a renewed interest in anchorage-dependent cell culture for high-yield processes. Well-known in vaccine production, microcarrier culture also has potential for application in other fields. In this work, two types of microcarriers were evaluated for small-scale monoclonal antibody (mAb) production by CHOK1 cells. Cultures (5 ml) of microporous Cytodex 3 and macroporous CultiSpher-S carriers were performed in vented conical tubes and subsequently scaled-up (20 ml) to shake-flasks, testing combinations of different culture conditions (cell concentration, microcarrier concentration, rocking methodology, rocking speed, and initial culture volume). Culture performance was evaluated by considering the mAb production and cell growth at the phases of initial adhesion and proliferation. The best culture performances were obtained with Cytodex 3, regarding cell proliferation (average 1.85 ± 0.11 × 10(6) cells/ml against 0.60 ± 0.08 × 10(6) cells/ ml for CultiSpher-S), mAb production (2.04 ± 0.41 µg/ml against 0.99 ± 0.35 µg/ml for CultiSpher-S), and culture longevity (30 days against 10-15 days for CultiSpher-S), probably due to the collagen-coated dextran matrix that potentiates adhesion and prevents detachment. The culture conditions of greater influence were rocking mechanism (Cytodex 3, pulse followed by continuous) and initial cell concentration (CultiSpher-S, 4 × 10(5) cells/ml). Microcarriers proved to be a viable and favorable alternative to standard adherent and suspended cultures for mAb production by CHO-K1 cells, with simple operation, easy scale-up, and significantly higher levels of mAb production. However, variations of microcarrier culture performance in different vessels reiterate the need for optimization at each step of the scale-up process.

Corrosion behaviour of titanium in the presence of Streptococcus mutans.

OBJECTIVE: The main aim of this in vitro study was to evaluate the influence of Streptococcus mutans on the corrosion of titanium. METHODS: S. mutans biofilms were formed on commercially pure titanium (CP-Ti) square samples (10mm×10mm×1mm) using a culture medium enriched with sucrose. Open circuit potential (OCP) and electrochemical impedance spectroscopy (EIS) measurements were used to evaluate the corrosion behaviour of CP-Ti in the presence of S. mutans in Fusayama's artificial saliva. The corrosion of biofilm-free CP-Ti samples was also evaluated in artificial saliva. Biofilms biomass was measured by spectrophotometry, using crystal violet staining, after 1, 2 and 7 days. RESULTS: The OCP values recorded on CP-Ti in the presence of S. mutans (-0.3±0.02V vs. SCE) was lower than those on biofilm-free CP-Ti (-0.1±0.01V vs. SCE) after 2h of immersion in artificial saliva (p<0.05). That reveals a high reactivity of titanium in presence of S. mutans. Impedance spectra revealed the formation of a compact passive film on titanium in artificial saliva or in the presence of a 2 days old S. mutans biofilm even though the corrosion resistance of CP-Ti has decreased in presence of a S. mutans biofilm. CONCLUSION: The presence of bacterial colonies, such as S. mutans, negatively affected the corrosion resistance of the titanium.

Farnesol induces cell detachment from established S. epidermidis biofilms.

Antibiotic resistance is a serious problem in Staphylococcus epidermidis infections as many clinical isolates of this organism are resistant to up to eight different antibiotics. The increased resistance to conventional antibiotic therapy has lead to the search for new antimicrobial therapeutic agents. Farnesol, an essential oil found in many plants, has been shown to be active against S. epidermidis. Using a type control strain we recently described that although farnesol was not efficient at killing biofilm bacteria, a strong reduction on biofilm biomass was detected, and we hypothesize that farnesol could, somehow, induce biofilm detachment. In this report, to test our hypothesis we used 36 representative clinical strains of S. epidermidis from different geographic locations and characterized them in terms of genetic variability by multilocus sequence typing and staphylococcal chromosome cassette mec. Strains were tested for biofilm formation, and the presence of ica, bhp and aap genes was determined. Stronger biofilms had always the presence of ica operon but often co-harbored bhp and aap genes. Farnesol was then used in biofilm-forming strains, and biofilm detachment was detected in half of the strains tested. Furthermore, we also showed that farnesol inability to kill biofilm bacteria was not the result of the biofilm structure but was related to high cell density. Our results demonstrate, for the first time, that the biomass reduction previously found by us, and many other groups, is the result not of cell killing but instead is the result of biofilm detachment.

The impact of microcarrier culture optimization on the glycosylation profile of a monoclonal antibody.

Microcarriers are widely used for the large-scale culture of attachment-dependent cells with increased cell densities and, ultimately, higher product yield. In these processes, the specific culture conditions can affect the quality of the product, which is closely related to its glycosylation pattern. Furthermore, the lack of studies in the area reinforces the need to better understand the effects of microcarrier culture in product glycosylation. Consequently, in this work, the glycosylation profile of a monoclonal antibody (mAb) produced by adherent CHO-K1 cells grown in Cytodex 3 was evaluated under different conditions, and compared to that obtained of typical adherent cultures. It was found that microcarrier cultures result in a glycosylation profile with different characteristics from T-flask cultures, with a general increase in galactosylation and decrease in fucosylation levels, both with a potentially positive impact on mAb activity. Sialylation also varied but without a general tendency. This study then showed that the specific culture conditions used in microcarrier culture influence the mAb glycan profile, and each functional element (galactose, core fucose, sialic acid) is independently affected by these conditions. In particular, great reductions of fucosylation (from 79 to 55%) were obtained when using half volume at inoculation, and notable decreases in sialylation (from 23 to 2%) and glycoform heterogeneity (from 20 to 11 glycoforms) were observed for shake flask culture, potentially associated with the improved cell densities achieved in these culture vessels.

Advances and drawbacks of the adaptation to serum-free culture of CHO-K1 cells for monoclonal antibody production.

Currently, mammalian cell technology has become the focus of biopharmaceutical production, with strict regulatory scrutiny of the techniques employed. Major concerns about the presence of animal-derived components in the culture media led to the development of serum-free (SF) culture processes. However, cell adaptation to SF conditions is still a major challenge and limiting step of process development. Thus, this study aims to assess the impact of SF adaptation on monoclonal antibody (mAb) production, identify the most critical steps of cell adaptation to the SF EX-CELL medium, and create basic process guidelines. The success of SF adaptation was dependent on critical steps that included accentuated cell sensitivity to common culture procedures (centrifugation, trypsinization), initial cell concentration, time given at each step of serum reduction, and, most importantly, medium supplements used to support adaptation. Indeed, only one of the five supplement combinations assessed (rhinsulin, ammonium metavanadate, nickel chloride, and stannous chloride) succeeded for the Chinese hamster ovary-K1 cell line used. This work also revealed that the chemically defined EX-CELL medium benefits mAb production in comparison with the general purpose Dulbecco's Modified Eagle's Medium, but the complete removal of serum attenuates these positive effects.