Enterotoxigenic profiles and submerged and interface biofilms in Bacillus cereus group isolates from foods.

Biofilm formation by Bacillus cereus strains is now recognized as a systematic contamination mechanism in foods; the aim of this study was to evaluate the production of submerged and interface biofilms in strains of B. cereus group in different materials, the effect of dextrose, motility, the presence of genes related to biofilms and the enterotoxigenic profile of the strains. We determine biofilm production by safranin assay, motility on semi-solid medium, toxin gene profiling and genes related to biofilm production by PCR in B. cereus group isolated from food. In this study, we observe strains used a higher production of biofilms in PVC; in the BHI broth, no submerged biofilms were found compared to phenol red broth and phenol red broth supplemented with dextrose; no strains with the ces gene were found, the enterotoxin profile was the most common the profile that includes genes for the three enterotoxins. We observed a different distribution of tasA and sipW with the origin of isolation of the strain, being more frequent in the strains isolated from eggshell. The production and type of biofilms are differential according to the type of material and culture medium used.

Biofilm-forming capacity of blood-borne Candida albicans strains and effects of antifungal agents.

Infections related to Candida albicans biofilms and subsequent antifungal resistance have become more common with the increased use of indwelling medical devices. Regimens for preventing fungal biofilm formation are needed, particularly in high-risk patients. In this study, we investigated the biofilm formation rate of multiple strains of Candida albicans (n=162 clinical isolates), their antifungal susceptibility patterns, and the efficacy of certain antifungals for preventing biofilm formation. Biofilm formation was graded using a modified Christensen's 96-well plate method. We further analyzed 30 randomly chosen intense biofilm-forming isolates using the XTT method. Minimum biofilm inhibition concentrations (MBIC) of caspofungin, micafungin, anidulafungin, fluconazole, voriconazole, posaconazole, itraconazole, and amphotericin B were determined using the modified Calgary biofilm method. In addition, the inhibitory effects of antifungal agents on biofilm formation were investigated. Our study showed weak, moderate, and extensive biofilm formation in 29% (n=47), 38% (n=61), and 23% (n=37) of the isolates, respectively. We found that echinocandins had the lowest MBIC values and that itraconazole inhibited biofilm formation in more isolates (26/32; 81.3%) than other tested agents. In conclusion, echinocandins were most effective against formed biofilms, while itraconazole was most effective for preventing biofilm formation. Standardized methods are needed for biofilm antifungal sensitivity tests when determining the treatment and prophylaxis of C. albicans infections.

[Networks involving quorum sensing, cyclic-di-GMP and nitric oxide on biofilm production in bacteria]. / Redes de señalización en la producción de biopelículas en bacterias: quorum sensing, di-GMPc y óxido nítrico.

Bacterial biofilms are ubiquitous in nature, and their flexibility is derived in part from a complex extracellular matrix that can be made-to-order to cope with environmental demand. Although common developmental stages leading to biofilm formation have been described, an in-depth knowledge of genetic and signaling is required to understand biofilm formation. Bacteria detect changes in population density by quorum sensing and particular environmental conditions, using signals such as cyclic di-GMP or nitric oxide. The significance of understanding these signaling pathways lies in that they control a broad variety of functions such as biofilm formation, and motility, providing benefits to bacteria as regards host colonization, defense against competitors, and adaptation to changing environments. Due to the importance of these features, we here review the signaling network and regulatory connections among quorum sensing, c-di-GMP and nitric oxide involving biofilm formation.

Addressing catheter lock therapy: Does heparin reduce the bioactivity of dalbavancin when together in solution during freezing?

INTRODUCTION: The possible use of dalbavancin as a catheter lock solution was previously demonstrated by our study group. However, it was needed to assess whether heparin could affect dalbavancin bioactivity during freezing storage. METHODS: We tested the bioactivity of a dalbavancin+heparin (DH) vs. dalbavancin (D) against Staphylococcal biofilms comparing DH median value of cfu counts and metabolic activity with that obtained for D before and during storage under freezing up to 6 months. RESULTS: Despite there was a slight decrease in the median percentage reduction of metabolic activity at month 3 in Staphylococcus epidermidis between DH and D (97.6 vs. 100, p=0.037), considering the clinical criteria, no significant reduction in any of the variables tested was observed at the end of the experiment between D and DH solutions. CONCLUSION: The addition of heparin to a dalbavancin lock solution did not affect its bioactivity against staphylococcal biofilms irrespective of its preservation time under freezing.

Characterization of Candida albicans and Staphylococcus aureus polymicrobial biofilm on different surfaces.

BACKGROUND: Staphylococcus aureus and Candida albicans have been co-isolated from biofilm-associated diseases such as denture stomatitis, periodontitis, and burn wound infections, as well as from medical devices. However, the polymicrobial biofilm of both microorganisms has not been fully characterized. AIMS: To characterize the polymicrobial biofilm of C. albicans and S. aureus in terms of microbial density, synergy, composition, structure, and stability against antimicrobials and chemical agents. METHODS: Crystal violet assay was used to measure the biofilm formation. Scanning electron microscopy and confocal microscopy were used to analyze the structure and chemical composition of the biofilms, respectively. RESULTS: Supplemented media with fetal bovine serum (FBS) decreased the biofilm formation of S. aureus and the polymicrobial biofilm. For C. albicans, depending on the culture media, the addition of glucose or FBS had a positive effect in biofilm formation. FBS decreased the adhesion to polystyrene wells for both microorganisms. Supplementing the media with glucose and FBS enhanced the growth of C. albicans and S. aureus, respectively. It seems that C. albicans contributes the most to the adhesion process and to the general structure of the biofilms on all the surfaces tested, including a catheter model. Interestingly, S. aureus showed a great adhesion capacity to the surface of C. albicans in the biofilms. Proteins and ß-1,6-linked polysaccharides seem to be the most important molecules in the polymicrobial biofilm. CONCLUSIONS: The polymicrobial biofilm had a complex structure, with C. albicans serving as a scaffold where S. aureus adheres, preferentially to the hyphal form of the fungus. Detection of polymicrobial infections and characterization of biofilms will be necessary in the future to provide a better treatment.

Bacterial profile of the supragingival dental biofilm in children with deciduous and early mixed dentition using next generation sequencing (HOMINGS) technique. / Perfil bacteriano del biofilm dental supragingival en niños con dentición temporal y mixta temprana utilizando la técnica de secuenciación de próxima generación (HOMINGS).

OBJECTIVE: Tdescribe the bacterial profile of the supragingival biofilm of children with temporary dentition (CTD) and early mixed dentition (CEMD), with the next-generation sequencing (HOMINGS) technique. METHOD: A comparative descriptive study was carried out with 30 systemically healthy children aged between 5 and 7 years old from public schools in Cartagena-Colombia. All participants were caries-free applying the criteria of the International Caries Detection and Assessment System (ICDAS II) and had no caries experience according to the Decayed, Missing and Filled Teeth (DMFT) index. Supragingival biofilm samples were collected. Bacterial DNA was extracted and used for analysis using HOMINGS (Human Oral Microbe Identification using Next-Generation Sequencing) based on the sequencing of the V3-V4 region of the 16S rRNA gene using the Illumina MiSeq platform (V3-V4 primers). RESULTS: A total of 360 species-specific and 65 genus-specific probes were identified. The bacterial genus most predominant in CTD were Streptococcus, Actinomyces, Veillonella and Fusobacterium (29.2% of all bacterial DNA present), while in CEMD the most predominant were Streptococcus, Leptotrichia, TM7 and Porphyromonas (24.5% of all bacterial DNA present). The bacterial species with the highest relative abundance in the oral biofilm microbiome from CTD were Streptococcus sanguinis, Rothia aeria, Gemella haemolysans, while in CEMD they were S. sanguinis, Leptotrichia spp. HOT-417 and Leptotrichia spp. HOT-498. The Shannon diversity index was 2.77 (SD=0.26) for CTD and 3.01 (SD=0.39) for CEMD (P=0.06). CONCLUSIONS: The analysis of the bacterial profile of the supragingival dental biofilm in children with DMFT, by means of HOMINGS showed low microbiological diversity both in presence and in relative abundance in terms of genus as well as bacterial species.

Anti-Candidaalbicans germ tube antibodies reduce in vitro growth and biofilm formation of C. albicans.

BACKGROUND: Invasive candidiasis by Candida albicans is associated with high morbidity and mortality, due in part to the late implementation of an appropriate antifungal therapy hindered by the lack of an early diagnosis. AIMS: We aimed to evaluate the in vitro antifungal activity of the antibodies against C. albicans germ tubes (CAGTA) raised in a rabbit model of candidemia. METHODS: We measured the effect of CAGTA activity by colorimetric XTT and crystal violet assays, and colony forming units count, both on C. albicans planktonic cells and during the course of biofilm formation and maturation. Viability and cell morphology were assessed by optical, fluorescent or scanning electron microscopy. RESULTS: CAGTA ≥50µg/ml caused a strong inhibition of C. albicans blastospores growth, and DiBAC fluorescent staining evidenced a fungicidal activity. Moreover, electron microscopy images revealed that CAGTA induced morphological alterations of the surface of C. albicans germ tubes grown free as well as in biofilm. Interestingly, CAGTA ≥80µg/ml reduced the amount of C. albicans biofilm, and this effect started at the initial adhesion stage of the biofilm formation, during the first 90min. CONCLUSIONS: This is the first report showing that CAGTA reduce C. albicans growth, and impair its metabolic activity and ability to form biofilm in vitro. The antigens recognized by CAGTA could be the basis for the development of immunization protocols that might protect against Candida infections.

Formación de biopelícula bacteriana sobre las superficies de pilares de cicatrización de PEEK y titanio

Objetivos. Explorar el efecto de las características de superficie sobre el volumen total y la viabilidad de la biopelícula formada sobre pilares de cicatrización de PEEK y titanio. Métodos. Los parámetros de rugosidad (S a y S k) y la energía superficial de pilares de cicatrización de PEEK y titanio (n=3) fueron determinados mediante microscopía confocal láser de barrido (CLSM) y ángulo de contacto, respectivamente. Se determinó luego el volumen total y la viabilidad de una biopelícula bacteriana multiespecie cultivada por 30 días, mediante CLSM y el reactivo LIVE/DEAD Kit BacLight. El tamaño del efecto se determinó mediante d de Cohen. Resultados. Los pilares de PEEK mostraron una mayor rugosidad que los de titanio (S a 0,41 µm vs 0,17 µm), pero no se observaron diferencias en la energía superficial. Si bien el volumen total de biopelícula fue mayor en titanio que en PEEK (696 µm3 vs 419 µm3), no hubo diferencias en la proporción de bacterias vivas entre ambos materiales. Conclusiones. La viabilidad de la biopelícula bacteriana formada no guarda relación directa con las características superficiales de pilares de cicatrización de PEEK y titanio.

Objetivo. Explorar o efeito das características da superfície no volume total e viabilidade do biofilme formado em PEEK e pilares de cicatrização de titânio. Métodos. Parâmetros de rugosidade (S a e S k) e energia de superfície de PEEK e pilares de titânio (n = 3) foram determinados por microscopia confocal de varredura a laser (CLSM) e ângulo de contato, respectivamente. O volume total e a viabilidade de um biofilme bacteriano multiespécie cultivado por 30 dias foram então determinados usando CLSM e o reagente LIVE/DEAD Kit BacLight. O tamanho do efeito foi determinado usando o d de Cohen. Resultados. Os pilares de PEEK mostraram maior rugosidade do que os de titânio (S a 0,41 µm vs 0,17 µm), mas não foram observadas diferenças na energia de superfície. Embora o volume total de biofilme tenha sido maior no titânio do que no PEEK (696 µm3 vs 419 µm3), não houve diferenças na proporção de bactérias vivas entre os dois materiais. Conclusões. A viabilidade do biofilme bacteriano formado não está diretamente relacionada às características da superfície dos pilares de cicatrização de PEEK e titânio.

Objectives . To explore the effect of surface characteristics on the total volume and viability of a bacterial biofilm developed on the surface of PEEK and titanium healing abutments. Methods. Surface parameters S a and S k, as well as the surface energy of PEEK and titanium healing abutments (n=3) were determined using confocal laser scanning microscopy (CLSM) and contact angle, respectively. The total volume and viability of a multispecies bacterial biofilm cultivated for 30 days were determined using CLSM and the LIVE/DEAD BacLight reactive kit. Effect size was determined using Cohen's d. Results. PEEK healing abutments displayed a higher surface roughness than titanium (S a 0.41 µm vs 0.17 µm), although no differences in surface energy were observed. Despite the higher total volume of the biofilm measured on titanium abutments compared to PEEK (696 µm3 vs 419 µm3), no differences in the live/dead bacterial ratio were observed. Conclusions. Bacterial viability of the biofilm did not show a direct relation to the surface characteristics of PEEK and titanium healing abutments.

Influencia de la higiene bucal en el estado de salud de los tejidos periimplantarios / Influence of oral hygiene on peri-implant tissue health status

Introducción: el estado de salud de los tejidos periimplantarios es de vital importancia en el éxito de la rehabilitación implantosoportada, por esta razón, es necesario observar todos aquellos factores que contribuyen a mantener este estado y dentro de ellos, principalmente: la higiene bucal. Objetivo: determinar la influencia de la higiene bucal en el estado de salud de los tejidos periimplantarios. Métodos: se realizó un estudio descriptivo, observacional y transversal en el servicio de Prótesis de la Facultad de Estomatología de Villa Clara, en el período comprendido entre los años 2017 y 2019. El universo de estudio estuvo constituido por 45 pacientes portadores de rehabilitaciones implantosoportadas; las unidades de análisis fueron los implantes y los tejidos que rodean a las 85 prótesis fijas realizadas a dichos pacientes que cumplieron con los criterios de inclusión. Se emplearon la observación clínica y radiográfica, y se elaboró un formulario como instrumento. Se evaluó la higiene bucal y el estado de los tejidos periimplantarios como principales variables. La información obtenida se recopiló en una base de datos, se procesó y se sometió a pruebas de independencia (el estadígrafo Ji cuadrado y su posibilidad asociada) para mostrar la relación entre las variables. Resultados: las variables analizadas evidenciaron una relación significativa de la higiene bucal con el estado de salud de los tejidos periimplantarios a favor de la buena higiene y los tejidos sanos. Conclusiones: la buena higiene bucal evidenciada contribuyó a que los tejidos periimplantarios se mantuvieran sanos.

Introduction: peri-implant tissue health state is of vital importance in the success of implant-supported rehabilitation; for this reason, it is necessary to observe all those factors that contribute to maintaining this state, mainly oral hygiene. Objective: to determine the influence of oral hygiene on peri-implant tissue health status. Methods: a descriptive, observational and cross-sectional study was carried out in the Prosthesis service at the Dental Faculty of Villa Clara between 2017 and 2019. The universe of study consisted of 45 patients with implant-supported rehabilitations; the units of analysis were the implants and the tissues surrounding the 85 fixed prostheses performed on those patients who met the inclusion criteria. Clinical and radiographic observations were used, and a form was developed as an instrument. Oral hygiene and peri-implant tissue state were evaluated as the main variables. The information obtained was compiled in a database as well as processed and subjected to independence tests (the Chi-square statistic and its associated possibility) to show the relationship among the variables. Results: the analyzed variables showed a significant relationship between oral hygiene and the peri-implant tissue health status in favour of good hygiene and healthy tissues. Conclusions: the evidenced good oral hygiene contributed to the maintenance of healthy peri-implant tissues.

Tipificación bioquímica y evaluación de la patogenicidad de aislamientos vulvovaginales del complejo Candida albicans / Biochemical typing and evaluation of pathogenicity in vulvovaginal isolates of Candida albicans complex

Introducción. Candida albicans, C. dubliniensis y C. africana forman el complejo Candida albicans. Objetivo. Identificar las características fenotípicas y patogénicas de aislamientos del complejo C. albicans conservados en una colección. Materiales y métodos. Se evaluaron 300 aislamientos identificados presuntivamente como del complejo C. albicans, utilizando CHROMagarTM Candida. Se determinó la producción del tubo germinal mediante tres métodos, se evaluó la producción de clamidosporas, se caracterizaron las colonias en agares artesanales (Rosmarinus officinalis y Nicotiana tabacum) y se utilizó MALDI-TOF como prueba de referencia para la identificación. Para detectar factores de patogenicidad, se evaluó la actividad hemolítica de los aislamientos independientes y en cocultivo con Staphylococcus aureus, la producción de enzima coagulasa y la formación de biopelículas. Resultados. El 43,7 % de los aislamientos produjo tubo germinal en caldo de medio infusión de cerebro-corazón y el 47 % generó clamidosporas. En los medios artesanales, en el 6 % de los aislamientos se obtuvieron colonias de color café en agar romero y, en el 5 %, en agar tabaco. Ninguna de las cepas hemolizó el agar sangre comercial (ni en presencia o ausencia de S. aureus), mientras que el 50 % hemolizó el agar papa dextrosa suplementado con sangre. Todos los aislamientos produjeron enzima coagulasa y la producción de biopelículas fue variable. Para la producción de tubo germinal, el método de suero humano mostró igual positividad que el de caldo de leche. Todos los aislamientos fueron identificados como C. albicans por MALDITOF. Conclusiones. Se requieren herramientas de proteómica y pruebas moleculares, o la combinación de métodos, para poder discriminar entre especies.

Introduction. Candida albicans, C. dubliniensis, and C. africana form the Candida albicans complex. Objective. To identify the phenotypic and pathogenic characteristics of isolates of the C. albicans complex preserved in a collection. Materials and methods. Three hundred presumptive strains of the C. albicans complex were evaluated using CHROMagarTM Candida. Germ tube production was determined by three methods, chlamydospores formation was assessed and colonies were characterized in artisanal agars (Rosmarinus officinalis and Nicotiana tabacum). MALDI-TOF was used as the gold standard identification test. To detect pathogenicity factors, we evaluated the hemolytic activity of each isolate and cocultured with Staphylococcus aureus, coagulase enzyme production, and biofilm formation. Results. Out of the 300 isolates, 43.7% produced germ tube in the heart-brain infusion broth and 47% of the isolates produced chlamydospores. In the artisan media, 6% of the isolates produced brown colonies on rosemary agar and 5% did so on tobacco agar. None of the strains hemolyzed the blood agar alone or cocultured with S. aureus. However, 50% of the isolates hemolyzed the potato dextrose agar supplemented with blood. All strains were coagulase producers, and biofilm production was variable. For germ tube production, the human serum method showed the same positivity as the milk broth method. All isolates were identified as C. albicans by MALDI-TOF. Conclusions. The use of proteomics, molecular tests or a combination of methods is required for species identification.

Microbiological diagnosis of biofilm-related infections. / Diagnóstico microbiológico de las infecciones relacionadas con la formación de biopelículas.

Biofilm-related infections represent a serious health problem, accounting for 65- 80% of all infections. The infections are generally chronic and characterized by the persistence of the microorganism, due to the increased resistance of biofilms to both the immune system and antimicrobials. Biofilms can be located to almost every human body tissue and on exogenous devices such as catheters, pacemakers, prosthetic material, implants, urinary catheters, etc. Traditional antimicrobial susceptibility studies in clinical microbiology laboratories have lied on the study of planktonic form of microorganisms. However, this approach might lead to miss the biofilm characteristics and to a treatment failure. Microbiological diagnosis and antimicrobial susceptibility studies of biofilm-related infections are complex and, nowadays, represent a challenge that clinicians and microbiologists have to address as a team in the absence of consensus or standardized protocols.

Enterotoxigenic profiles and submerged and interface biofilms in Bacillus cereus group isolates from foods / Perfiles enterotoxigénicos y biopelículas de superficie y sumergidas en aislados del grupo Bacillus cereus de alimentos

Abstract Biofilm formation by Bacillus cereus strains is now recognized as a systematic contaminaron mechanism in foods; the aim of this study was to evaluate the production of submerged and interface biofilms in strains of B. cereus group in different materials, the effect of dex-trose, motility, the presence of genes related to biofilms and the enterotoxigenic profile of the strains. We determine biofilm production by safranin assay, motility on semi-solid medium, toxin gene profiling and genes related to biofilm production by PCR in B. cereus group iso-lated from food. In this study, we observe strains used a higher production of biofilms in PVC; in the BHI broth, no submerged biofilms were found compared to phenol red broth and phenol red broth supplemented with dextrose; no strains with the ces gene were found, the enterotoxin profile was the most common the profile that includes genes for the three enterotoxins. We observed a different distribution of tasA and sipW with the origin of isolation of the strain, being more frequent in the strains isolated from eggshell. The production and type of biofilms are differential according to the type of material and culture medium used.

Resumen La formación de biopelículas por cepas de Bacillus cereus es reconocida como un mecanismo de contaminación sistemática en alimentos; el objetivo del estudio fue evaluar la producción de biopelículas sumergidas y de superficie en cepas del grupo de Bacillus cereus en diferentes materiales, el efecto de la dextrosa, la motilidad, la presencia de genes relacionados a biopelículas y el perfil enterotoxigénico de las cepas. Determinamos la producción de biopelículas por el ensayo de safranina, motilidad en medio sólido, perfil enterotoxigénico y genes relacionados a producción de biopelículas por PCR en aislados del grupo de Bacillus cereus de alimentos. En este estudio, observamos en las cepas utilizadas una alta producción de biopelículas en PVC; en caldo BHI, no se encontraron biopelículas sumergidas en comparación con el caldo rojo de fenol y caldo rojo de fenol suplementando con dextrosa; no se encontraron cepas con el gen ces, el perfil de enterotoxinas más común fue el perfil que incluía los genes de las tres enterotoxinas, también observamos una distribución diferente de tasA y sipW con relación al origen de la cepa, siendo más frecuente estos genes en las cepas aisladas de huevos. La producción y el tipo de biopelículas es diferente de acuerdo con el tipo de material y el medio de cultivo utilizado.

Las biopelículas y su interrelación con la patología crónica del oído medio / Biofilms and their relationship with chronic middle ear pathology

El concepto de biopelículas ha surgido de forma paulatina durante un largo período; se presentan como estructuras tridimensionales compuestas por células sésiles de microorganismos que crecen y se adhieren irreversiblemente a superficies, tanto vivas como inertes. Su capacidad de desarrollarse, tanto en superficies bióticas como abióticas, es una característica que los relaciona directamente con la salud humana. Distintas infecciones óticas se han inculpado a la presencia de biopelículas en las mucosas como en la otitis media con efusión, de igual forma se manifiestan en la aparición y persistencia de la otitis media crónica. Las biopelículas afines con otitis media, generalmente, contienen uno o múltiples especies de bacterias otopatógenas primarias. La comprensión de la biopelicula auxiliará el progreso de nuevas terapias y estrategias de control, al evitar enfermedades infecciosas ya que las bacterias formadoras de biopelículas son una seria amenaza para la salud pública debido a su alta resistencia a los antimicrobianos.

The concept of biofilms has emerged gradually over a long period; they appear as three-dimensional structures composed of sessile cells of microorganisms that grow and adhere irreversibly to surfaces, both living and inert. Their ability to develop, both on biotic and abiotic surfaces, is a characteristic that directly relates them to human health. Different ear infections have been blamed on the presence of biofilms on the mucous membranes, such as otitis media with effusion, in the same way they manifest themselves in the appearance and persistence of chronic otitis media. Otitis media-related biofilms generally contain one or multiple species of primary otopathogenic bacteria. The understanding of the biofilm will help us refine new therapies and control strategies, by avoiding infectious diseases since biofilm-forming bacteria are a serious threat to public health due to their high resistance to antimicrobials.

Microbial biofilms and their impact on medical areas: physiopathology, diagnosis and treatment. / Biopelículas microbianas y su impacto en áreas médicas: fisiopatología, diagnóstico y tratamiento

Biofilms are communities of microorganisms that grow aggregated and surrounded by an extracellular matrix, which they produce and favors them to adhere covalently to inert and living surfaces; it also helps them to develop high tolerance to molecules with antimicrobial activity. Moreover, biofilms are associated with chronic and persistent infections, which negatively impact different medical areas since they generate high costs to health care systems and patients every year because they are difficult to treat with conventional antimicrobial drugs. Additionally, they generate high rates of morbidity and mortality. The objective of this review was to present extensive and up-to-date information on the origin, biosynthesis, and pathophysiology of biofilms. Also, its relationship with chronic infections, diagnosis, current antimicrobial treatments with antibiotic activity, and perspectives on the search for new treatments, since the latter still represent an important area of research.

Las biopelículas son comunidades de microorganismos que crecen agregados y rodeados por una matriz extracelular que ellos mismos producen, la cual favorece la adhesión covalente sobre superficies inertes y vivas; además, les ayuda a desarrollar alta tolerancia a las moléculas con actividad antimicrobiana. Por otra parte, las biopelículas se asocian con infecciones crónicas y persistentes que impactan de manera negativa en distintas áreas médicas. Además, generan altos costos a los sistemas de salud y a los pacientes cada año, porque son difíciles de tratar con antimicrobianos convencionales; adicionalmente, generan altas tasas de morbilidad y mortalidad. El objetivo de esta revisión es presentar información extensa y actualizada sobre el origen, la biosíntesis y la fisiopatología de las biopelículas, así como sobre su relación con infecciones crónicas, el diagnóstico, los tratamientos antimicrobianos actuales con actividad antibiopelícula y las perspectivas sobre la búsqueda de nuevos tratamientos. Estos últimos aún representan una importante área de investigación.

Candida parapsilosis endocarditis without vegetations at Echocardiography: a case report / Endocarditis por Candida parapsilosis sin vegetaciones ecocardiográficas: Un reporte de caso

ABSTRACT Candida endocarditis is a very serious manifestation of candida infections, and it has increased in incidence over the past years. Of these, C. parapsilosis has been described as a cause of endocarditis in native valves of intravenous drug users and prosthetic valves. We report the case of a female that developed a cerebrovascular accident secondary to emboli from aortic prosthetic valve C. parapsilosis endocarditis, despite apparently normal echocardiography. She received antifungal therapy without surgical intervention.

RESUMEN La endocarditis por Candida spp. es una manifestación muy grave de las infecciones por este patógeno y su incidencia ha ido aumentando en los últimos años. La Candida parapsilosis se ha descrito como causa de endocarditis en válvulas nativas de usuarios de drogas intravenosas y prótesis valvulares. Presentamos el caso de una mujer que desarrolló un accidente cerebrovascular secundario a émbolos fúngicos procedentes de una prótesis valvular aórtica infectada por C. parapsilosis, a pesar de un ecocardiograma sin vegetaciones. La paciente recibió tratamiento antifúngico sin necesidad de intervención quirúrgica.

Evaluación de la formación de biopelículas en aislamientos bacterianos y fúngicos por el método semicuantitativo de microtitulación con cristal violeta y el cualitativo de agar con rojo Congo / Assessment of biofilms formation of bacterial and fungal isolates using qualitative Congo red agar and semiquantitative crystal violet microtiter methods

Introducción. El 65 % de las infecciones humanas son producidas por bacterias o levaduras, cuya capacidad de formar biopelículas las hace más resistentes a los antimicrobianos y antifúngicos. Objetivo. Determinar la capacidad de formación de biopelículas en aislamientos bacterianos y fúngicos por medio de los métodos cuantitativo de microtitulación con cristal violeta y cualitativo de cultivo en agar con rojo Congo. Materiales y métodos. Con el método cuantitativo, se utilizaron los medios de cultivo infusión cerebro-corazón, tripticasa de soya y Müeller-Hinton para aislamientos bacterianos; para levaduras, se usaron caldo infusión cerebro-corazón y Sabouraud dextrosa. Para el método cualitativo de cultivo en agar, se utilizaron los mismos medios de cultivo más una solución con 3 % de rojo Congo y 10 % de dextrosa. Cómo método de referencia, se utilizó la propuesta de Stepanovic et al. Resultados. Se evaluaron 103 aislamientos bacterianos y 108 de levaduras. No es recomendable sustituir el caldo infusión cerebro-corazón por los caldos tripticasa de soya y Müeller-Hinton en el método cuantitativo, para evaluar la formación de biopelículas en los aislamientos bacterianos. El medio Sabouraud dextrosa, en caldo y agar, puede sustituir al de infusión de cerebro-corazón para evaluar la formación de biopelículas en levaduras, tanto por el método cuantitativo como por el cualitativo. Conclusión. El estudio de las biopelículas en el laboratorio de microbiología, a partir del método cualitativo de cultivo en agar con rojo Congo, es un procedimiento sencillo, rápido y de bajo costo, que proporciona información útil para el diagnóstico y la terapéutica de infecciones persistentes causadas por bacterias y levaduras.

Introduction. Sixty-five percent of human infections are caused by bacteria or yeasts able to form biofilms. This feature makes them more resistant to antimicrobials and antifungals. Objective. To determine biofilm formation capacity of bacterial and fungal isolates by quantitative crystal violet microtiter and qualitative Congo red agar methods. Materials and methods. Brain-heart infusion, trypticase soy broth and Müeller-Hinton culture media were used in bacterial isolates for the quantitative method; brain-heart infusion broth and Sabouraud dextrose were used for yeasts. The same culture media plus 3% Congo red and 10% dextrose were used to apply the qualitative method in agar. The proposal by Stepanovic, et al. was used as a reference method. Results. We evaluated 103 bacterial isolates and 108 yeasts isolates. We did not recommend substitute brain-heart infusion broth for trypticase soy and Müeller-Hinton broths for biofilm formation assessment in bacterial isolates using the quantitative method. Sabouraud dextrose medium, both broth and agar, can replace brain-heart infusion to assess biofilm formation in yeasts, quantitatively and qualitatively. Conclusion. The study of biofilms in the microbiology laboratory, using Congo red agar qualitative method, is a simple, fast, and inexpensive procedure that provides precise information for the diagnosis and treatment of persistent infections caused by bacteria and yeasts.

Biofilme em parafusos ortopédicos prontos para uso adquiridos por meio de sistema de consiganação/comodato / Biopelícula en tornillos ortopédicos listos para uso adquiridos por medio del sistema de consignación/comodato / Biofilm on patient-ready orthopaedic screws acquired through the loaner system

Resumo Objetivo Avaliar a integridade da superfície e as condições microbiológicas de parafusos prontos para uso em bandejas ortopédicas após múltiplos processamentos. Métodos Após o processamento completo, as bandejas utilizadas em cirurgias de pequenos fragmentos, fornecidas por meio de sistema de consignação/comodato em um hospital brasileiro, foram selecionadas aleatoriamente durante quatro meses. Os parafusos mais utilizados (números 14, 16 e 18 - Grupo 1) e menos utilizados (números 10 e 38 - Grupo 2), portanto, os mais e menos expostos a agentes biológicos, químicos e físicos, foram aleatoriamente removidos e submetidos a inspeção visual (n=126), seguido de cultura bacteriana (n=6 parafusos/bandeja, 9 bandejas), teste de proteínas (n=6 parafusos/bandeja, 9 bandejas) e Microscopia Eletrônica de Varredura (MEV) (n=2 parafusos/bandeja, 9 bandejas). As culturas positivas foram submetidas a métodos automatizados de identificação bacteriana e suscetibilidade antimicrobiana. Resultados Foram detectadas ranhuras em 8,7% dos parafusos, predominantemente no Grupo 2 (8/11). Proteína residual foi detectada em 96,3%, e não houve diferença estatisticamente significativa na quantidade de proteína entre os grupos (P=0,07). Crescimento bacteriano foi identificado em 3/54 parafusos. Danos na superfície e presença de sujidade foram visualizados em todos os parafusos submetidos a MEV. Formação de biofilmes extensos foi detectada em oito parafusos, três do Grupo 1 e cinco do Grupo 2. Conclusão Recuperação de bactérias viáveis, acúmulo de biofilme e danos na superfície foram detectados nos parafusos prontos para uso. Os parafusos costumam permanecer nas bandejas cirúrgicas e serem submetidos a múltiplos processamento, sendo expostos a contaminação e danos repetidas vezes. Esses achados apontam para a necessidade de discutir e repensar a forma como esses implantes de uso único são atualmente disponibilizados para cirurgias.

Resumen Objetivo Evaluar la integridad de la superficie y las condiciones microbiológicas de tornillos listos para uso en bandejas ortopédicas después de múltiples procesamientos. Métodos Después del procesamiento completo, fueron seleccionadas aleatoriamente durante cuatro meses las bandejas utilizadas en cirugías de pequeños fragmentos, proporcionadas mediante el sistema de consignación/comodato en un hospital brasileño. Los tornillos más utilizados (números 14, 16 y 18 - Grupo 1) y menos utilizados (números 10 y 38 - Grupo 2), por lo tanto, los más y menos expuestos a agentes biológicos, químicos y físicos, fueron quitados aleatoriamente y sometidos a inspección visual (n=126), seguido de cultivo bacteriano (n=6 tornillos/bandeja, 9 bandejas), prueba de proteínas (n=6 tornillos/bandeja, 9 bandejas) y microscopía electrónica de barrido (MEB) (n=2 tornillos/bandeja, 9 bandejas). Los cultivos positivos fueron sometidos a métodos automatizados de identificación bacteriana y susceptibilidad antimicrobiana. Resultados Se detectaron ranuras en el 8,7 % de los tornillos, predominantemente en el Grupo 2 (8/11). Se detectó proteína residual en el 96,3 % y no se encontró diferencia estadísticamente significativa en la cantidad de proteína entre los grupos (P=0,07). En 3/54 tornillos se identificó crecimiento bacteriano. Se visualizaron daños en la superficie y presencia de suciedad en todos los tornillos sometidos a MEB. En ocho tornillos se detectó la formación de biopelículas, tres del Grupo 1 y cinco del Grupo 2. Conclusión Se detectó recuperación de bacterias viables, acumulación de biopelícula y daños en la superficie en los tornillos listos para uso. Los tornillos suelen permanecer en las bandejas quirúrgicas y son sometidos a múltiples procesamientos, donde están expuestos a contaminación y daños repetidas veces. Estos descubrimientos señalan la necesidad de discutir y repensar la forma como estos implantes de uso único se ponen a disposición para cirugía actualmente.

Abstract Objective Assess the surface integrity and microbiological conditions of patient-ready screws in orthopaedic trays that had been multiply reprocessed. Methods After full reprocessing, clinical trays used for small fragment surgery provided through a loaner system to a Brazilian hospital were randomly selected during four months. The most (numbers 14, 16 and 18 - Group 1) and least (numbers 10 and 38 - Group 2) frequently implanted screws, therefore, the ones estimated to be the most and least exposed to biological, chemical and physical agents, were randomly removed and subjected to visual inspection (n=126), followed by bacterial culture (n=6 screws/tray, 9 trays), protein test (n=6 screws/tray, 9 trays) and Scanning Electron Microscopy (SEM) (n=2 screws/tray, 9 trays). Positive cultures were subjected to automated bacterial identification and antimicrobial susceptibility tests. Results Grooves were detected on 8.7% screws, predominantly in Group 2 (8/11). Residual protein was detected on 96,3%, and there was no statistically significant difference in the amount of protein between the groups (P=0.07). Bacterial growth was identified in 3/54 screws. Surface damage and soil were visualized on all screws subjected to SEM. Extensive biofilms were detected on eight screws, three from Group 1 and five from Group 2. Conclusion Recovery of bacteria, biofilm accumulation and surface damage were detected on patient-ready screws. Screws frequently remain in surgical trays for multiple reprocessing; thus they are repeatedly exposed to contamination and damage. These findings point to the need to discuss and review the way these single-use implants are currently made available for surgeries.

Estandarización de un ensayo de adhesión de Escherichia coli para el cribado de agentes antibiopelículas / Standardization of an Escherichia coli adhesion assay for anti-biofilm agents screening

Introducción: Las biopelículas constituyen un factor clave en el desarrollo de enfermedades infecciosas y la resistencia a fármacos para su control. Nuevas estrategias terapéuticas incluyen productos naturales como agentes antibiopelículas. Sin embargo, la comparación de los resultados suele ser difícil debido a la falta de homogeneidad y estandarización en los métodos empleados para estudiar la formación de biopelículas in vitro. Objetivo: Estandarizar un ensayo de adhesión en microplaca de Escherichia coli para su uso en el cribado de potenciales agentes antibiopelículas. Métodos: Se utilizó el método de adherencia en microplaca y la tinción con violeta cristal. Se evaluó la influencia de condiciones experimentales como la concentración bacteriana, el medio de cultivo y el tiempo de incubación. Resultados: Se identificaron como condiciones óptimas para la formación de biopelículas: el medio Luria Bertani (LB), la concentración bacteriana a 105 UFC/mL y un tiempo de incubación de 24 h. Conclusiones: Los resultados mostraron que las condiciones de cultivo influyen en la formación de biopelículas. Se determinaron las condiciones de cultivo óptimas para la formación de biopelículas de E. coli, que podrían emplearse en futuros estudios acerca del efecto de productos naturales sobre la inhibición o destrucción de biopelículas.

Introduction: Biofilms are a key factor in the development of infectious diseases and drug resistance for their control. New therapeutic strategies include natural products as anti-biofilm agents. However, comparing results is often difficult due to the lack of homogeneity and standardization of the methods used to study biofilm formation in vitro. Objective: To standardize an Escherichia coli microplate adhesion assay for potential anti-biofilm agents screening. Methods: The microplate adhesion method and crystal violet staining were used. The influence of experimental conditions such as bacterial concentration, culture medium, and incubation time were evaluated. Results: Optimal conditions for biofilm formation included: Luria Bertani (LB) medium, bacterial concentration at 105 CFU/mL, and an incubation time of 24 hours. Conclusions: The results showed that culture conditions influence biofilm formation. Optimal culture conditions for the formation of E. coli biofilms were determined, which could be used in further studies on the effect of natural products on the inhibition or destruction of biofilms.

Antibacterial potential of essential oils against planktonic and sessile cells of Escherichia coli isolated from diarrhea cases in swine / Potencial antibacteriano de aceites esenciales contra células de Escherichia coli planctónicas y sésiles aisladas de diarrea porcina

Escherichia coli is a pathogen associated with infections in piglets in the post-weaning phase, its pathogenicity is related to the animal's susceptibility to bacterial enterotoxins. The objective of the present study was to determine the EOs activity against E. colistrain, in the form planktonic and sessile. Although the Disc-Diffusion tests to determine the Minimum Inhibitory Concentration, do not fully corroborate with the other analyzes of this study, it was noticed bacteria inhibition. The EOs were prepared at 0.4%, 0.8% and 1.0% for tests. The tested EOs were effective against E. coliplanktonic cells (p<0.05). As for the sessile cells, the most significant result was inhibition and 100% sessile cells at the concentration of 1.0% of Cymbopogon citratusEO. Although there was resistance in some treatments, the tested EOs demonstrated inhibition capacity, constituting promising alternatives for the control of E. coli, especially of planktonic cells.

Escherichia coli es un patógeno asociado con infecciones en lechones en la fase posterior al destete, su patogenicidad está relacionada con la susceptibilidad del animal a las enterotoxinas bacterianas. El objetivo del presente estudio fue determinar la actividad de contra E. coli, en la forma planctónico y sésil. Aunque las pruebas de difusión de disco para determinar la concentración inhibitoria mínima, no corroboran completamente con los otros análisis de este estudio, se observó inhibición de la bacteria. Las soluciones basadas en AE se prepararon al 0.4%, 0.8% y 1.0% para pruebas. Los AEs probados fueron efectivos contra las células planctónicas (p<0.05). En cuanto a las células sésiles, el resultado más significativo fue la inhibición y el 100% de las células sésiles a la concentración de 1,0% de Cymbopogon citratus. Aunque hubo resistencia en algunos tratamientos, los AEs probados demostraron capacidad de inhibición, constituyendo alternativas prometedoras para el control de E. coli, especialmente de células planctónicas.

Capacidad de Lactiplantibacillus plantarum LP5 para inhibir biopelículas de Campylobacter coli / Ability of Lactiplantibacillus plantarum LP5 to inhibit biofilms of Campylobacter coli

Resumen El objetivo de este estudio fue evaluar la capacidad inhibitoria de Lactiplantibacillus plantarum LP5 frente a Campylobacter coli en ensayos de formación de biopelículas in vitro y exclusión competitiva. La formación de biopelículas por C. coli NCTC11366, C. coli DSPV458, C. coli DSPV541 y C. coli DSPV570 fue evaluada mediante medición de DO. La capacidad inhibitoria de L. plantarum LP5 frente a C. coli fue evaluada sobre discos de vidrio, nailon y aluminio. Sobre una biopelícula de L. plantarum se adicionó C. coli para cuantificar el efecto inhibidor de L. plantarum LP5 sobre el patógeno. Las cuatro cepas de C. coli fueron clasificadas como moderadas formadoras de biopelículas. El ensayo de exclusión competitiva mostró que la formación de biopelículas de las cepas de C. coli en todos los materiales fue significativamente mayor que la formación de biopelículas de cada patógeno en presencia de biopelículas de L. plantarum LP5. Si bien es necesario realizar más pruebas para confirmar la capacidad de supervivencia de C. coli en ambientes hostiles hasta llegar al huésped, este estudio permitiría avanzar en el esclarecimiento de su comportamiento mediante la formación de biopelículas.

Abstract The objective of this study was to evaluate the inhibitory capacity of Lactiplantibacillus plantarum LP5 against Campylobacter coli in in vitro biofilm formation and competitive exclusion assays. Biofilm formation by C. coli NCTC11366, C. coli DSPV458, C. coli DSPV541 and C. coli DSPV570 was evaluated by OD measurement. The inhibitory capacity of L. plantarum LP5 against C. coli was evaluated on glass, nylon and aluminium discs, added with L. plantarum and incubated at 37°C for 72 h. C. coli was added to each washed well. The plates were incubated at 42°C for 72 h in microaerophilic conditions and the biofilms were detached for quantification. The four strains of C. coli were classified as moderate biofilm former. The competitive exclusion test showed that the biofilm formation of the C. coli strains in all materials was significantly higher than the biofilm formation of each pathogen in the presence of L. plantarum LP5 biofilms. Although it is necessary to carry out more tests to confirm the ability of C. coli to survive in hostile environments until reaching the host, this study would allow progress in the elucidation of its behaviour through the formation of biofilms.