Molecular analysis, biofilm formation, and susceptibility of methicillin-resistant staphylococcus aureus strains causing community- and health care-associated infections in central venous catheters

Abstract INTRODUCTION: The behavior of methicillin-resistant Staphylococcus aureus (MRSA) isolated from central venous catheter-related infection was evaluated to determine its biofilm potential, antimicrobial resistance, and adhesion genes. METHODS: A total of 1,156 central venous catheters (CVC) were evaluated to screen for pathogens. Antimicrobial sensitivity, biofilm formation potential, and molecular analysis of MRSA were examined following standard guidelines. RESULTS: Of the 1,156 samples, 882 (76%) were colonized by bacteria or candida. Among the infected patients, 69% were male and 36% were female with median age of 32 years. Staphylococcus aureus infected 39% (344/882) of CVCs in patients. Of the 59% (208/344) of patients with MRSA, 57% had community acquired MRSA and 43% had hospital acquired MRSA. Linezolid and vancomycin killed 100% of MRSA; resistance levels to fusidic acid, doxycycline, clindamycin, azithromycin, amikacin, trimethoprim-sulfamethoxazole, gentamycin, tobramycin, and ofloxacin were 21%, 42%, 66%, 68%, 72%, 85%, 95%, 97%, and 98% respectively. Strong biofilm was produced by 23% of samples, moderate by 27%, and weak by 50% of MRSA. The presence of adhesion genes, sdrC and sdrD (90%), eno (87%), fnbA (80%), clfA and sdrE (67%), fnbB, sdrD (61%), and cna (51%), in most MRSA samples suggested that the adhesion genes are associated with biofilm synthesis. CONCLUSIONS: The superbug MRSA is a major cause of CVC-related infection. Antibiotic resistance to major classes of antibiotics and biofilm formation potential enhanced superbug MRSA virulence, leading to complicated infection. MRSA causes infection in hospitals, communities, and livestock.

Risk factors for methicillin-resistant Staphylococcus aureus bacteremia: A multicenter matched case-control study / Factores de riesgo de la resistencia a meticilina de Staphylococcus aureus causante de bacteriemia: estudio multicéntrico de casos y controles emparejados

Abstract Introduction: Methicillin-resistant Staphylococcus aureus is a frequent pathogen at critical care services. Its presence leads to increased hospital stays and mortality risk in patients with bacteremia. However, the etiology of this resistance marker has not been fully studied. Objective: To identify risk factors associated with the emergence of methicillin-resistant S. aureus bacteremia in critically ill patients treated at intensive care units in Bogotá, Colombia. Materials and methods: We conducted a retrospective paired case-control study, nested in a cohort of patients diagnosed with S. aureus bacteremia and treated at intensive care units between 2006 and 2008 in Bogotá. Cases were patients with positive blood culture to methicillin resistance, matched in a 1:1 ratio with methicillin-sensitive controls isolated from the same institution and hospitalization year. We used conditional logistic regression to analyze the risk factors associated with the presence of resistance, with emphasis on prior antibiotic therapy. Results: We included 372 patients with S. aureus bacteremia. Factors such as the use of pre-hospital devices: vascular (OR=1.986, 95% CI 1.038 to 3.801) and urinary (OR=2.559, 95% CI: 1.170 to 5.596), along with the number of previously used antibiotics, were associated with the emergence of resistance. The number of antibiotics used previously was determined to have a gradient effect, particularly carbapenems. Conclusions: The rational use of antibiotics and surveillance of exposure to surgical procedures or use of invasive devices are interventions that could diminish the emergence of methicillin-resistant S. aureus bacteremia causes.

Resumen Introducción. Staphylococcus aureus resistente a la meticilina es uno de los agentes patógenos más frecuentes en las unidades de cuidados intensivos. Su presencia prolonga las hospitalizaciones y aumenta el riesgo de mortalidad en los pacientes con bacteriemia. Sin embargo, la etiología de este marcador de resistencia no ha sido completamente estudiada. Objetivo. Determinar los factores asociados con la aparición de S. aureus resistente a la meticilina causante de bacteriemia en pacientes atendidos en unidades de cuidados intensivos en Bogotá. Materiales y métodos. Se hizo un estudio retrospectivo de casos y controles emparejados, anidado en una cohorte de pacientes con diagnóstico de bacteriemia por S. aureus atendidos en unidades de cuidados intensivos de Bogotá entre 2006 y 2008. Los casos fueron pacientes con hemocultivo positivo para resistencia a la meticilina, emparejados 1 a 1 con controles con hemocultivos sensibles a la meticilina de la misma institución y año de hospitalización. Se analizaron mediante regresión logística condicional los factores de riesgo asociados con la presencia de resistencia, con énfasis en el tratamiento previo con antibióticos. Resultados. Se incluyeron 372 pacientes con bacteriemia por S. aureus. Factores como el uso de dispositivos previos a la hospitalización: vasculares (Odds ratio, OR=1,986; IC95% 1,038-3,801) y urinarios (OR=2,559; IC95% 1,170-5,596), así como el número de antibióticos administrado previamente, se asociaron con la aparición de resistencia. Se registró un efecto de gradiente con el número de antibióticos usados previamente, especialmente carbapenémicos. Conclusiones. El uso racional de antibióticos y la vigilancia de la exposición a procedimientos quirúrgicos o al uso de dispositivos invasivos, son intervenciones que podrían disminuir la aparición de S. aureus resistente a meticilina causante de bacteriemia.

Antibiotic Resistance Patterns and Plasmid Profiles of Methicillin Resistant Staphylococcus aureus Isolates from Human Samples.

Antibiograms and plasmid profiles are commonly used to characterizemethicillin-resistant Staphylococcus aureus (MRSA) inepidemiologic studies. However, antibiograms are frequently inadequate to accomplish the differentiation. Plasmid profile being more informative has been reported to be useful in tracing the epidemiology of antibiotic resistance. Antibiotic resistance patterns and plasmid profiles of methicillin-resistant Staphylococcus aureus (MRSA) isolates from human specimens were investigated to determine the discriminatory power of plasmid profile analysis in conjunction with antibiotic susceptibility pattern. Specimens were analyzed using disc diffusion assay and restriction enzymes analysis of plasmid DNA procedure. The 51 methicillin-resistant Staphylococcus aureus (MRSA) isolates were grouped into 18 groups using their resistogram. Twenty four (47.1%) strains out the 51 methicillin-resistant Staphylococcus aureus (MRSA) isolates harbored plasmids. Single plasmid isolates were 14(27.5%), double plasmid isolates were 6(11.8%) while tripple plasmid isolates were 4(7.8%). The 24 isolates containing plasmids were categorized into 14 groups based on their resistogram. Plasmid profile showed greater similarity between isolates (10 profiles) than antibiotic resistance pattern which showed a higher disparity (14 patterns). However, resistance to various antimicrobial agents was not consistent with the presence of plasmids. No particular molecular size plasmid could be associated with any particular antimicrobial resistance patterns. Resistance was observed in isolates with various molecular size plasmids as well as in those that had no plasmids. Nonetheless, 2 pairs of isolates with the same plasmid profile also had similar (almost the same) resistance pattern. Plasmid profile analysis in conjunction with the antibiotic resistance typing is valuable in the epidemiological investigation of methicillin-resistant Staphylococcus aureus (MRSA).

Newly-synthesized chalcones-inhibition of adherence and biofilm formation of methicillin-resistant Staphylococcus aureus

Biofilm formation and adherence of bacteria to host tissue are one of the most important virulence factors of methicillin-resistant strains of Staphylococcus aureus (MRSA). The number of resistant strains is seriously increasing during the past years and bacteria have become resistant, not only to methicillin, but also to other commonly used antistaphylococcal antibiotics. There is a great need for discovering a novel antimicrobial agent for the treatment of staphylococcal infections. One of the most promising groups of compounds appears to be chalcones. In present study we evaluated the in vitro effect of three newly synthesized chalcones: 1,3-Bis-(2-hydroxy-phenyl)-propenone, 3-(3Hydroxy-phenyl)-1-(2-hydroxy-phenyl)-propenone and 3-(4-Hydroxy-phenyl)-1-(2-hydroxyphenyl)-propenone on glycocalyx production, biofilm formation and adherence to human fibronectin of clinical isolates and laboratory control strain of MRSA (ATCC 43300). Subinhibitory concentrations of the tested compounds reduced the production of glycocalyx, biofilm formation and adherence to human fibronectin of all MRSA strains. Inhibition of biofilm formation was dose dependent and the most effective was 1,3-Bis-(2-hydroxy-phenyl)-propenone. In our study we demonstrated that three newly-synthesized chalcones exhibited significant effect on adherence and biofilm formation of MRSA strains. Chalcones may be considered as promising new antimicrobial agents that can be used for prevention of staphylococcal infections or as adjunct to antibiotics in conventional therapy.

Actividad bactericida de superficies de cobre frente a bacterias asociadas a infecciones nosocomiales, en un modelo in vitro de adherencia y sobrevivencia / Adherence to copper and stainless steel metal coupons of common nosocomial bacterial strains

Background: Copper has a bactericidal activity against a series of bacterial strains. Aim: To measure resistance to bacterial adherence of copper (Cu) and stainless steel (SS) metal coupons. Material and Methods: Bacterial strains causing nosocomial infections in Chile were analyzed. Bacterial adherence was studied using apreviously described method based on a system of metal coupons that are immersed in culture media containing the bacteria ofinterest at room temperature. Results: Adherence to Cu and SS coupons was differentfor Methicillin-resistant Staphylococcus aureus (MRSA), Klebsiella pneumoniae and Acinetobacter baumannii strains. For these strains, no adherence to Cu coupons occurred during the 48 h observation period compared to a rapidly increasing adherence to SS coupons, with a final colony count of 1.00E + 07 cfu/mL. For two different Pseudomonas aeruginosa clinical strains, inhibition of adherence was not observed on Cu coupons, and colony counts were similar for Cu and SS using the standard inoculum (2-3 xlO7 cfu).Apartial decrease in adherence was observed for Cu but not for SS coupons, when a lower inoculum was used. Conclusions: Copper surfaces represent an interesting option to reduce bacterial contamination in the hospital environment due to its resistance to bacterial adhesión ofmost ofthe common nosocomial bacterial strains.

Rifampicin fails to eradicate mature biofilm formed by methicillin-resistant Staphylococcus aureus / Rifampicina falha na erradicação de biofilmes maduros formados por Staphylococcus aureus resistentes à meticilina

INTRODUCTION: Antimicrobial activity on biofilms depends on their molecular size, positive charges, permeability coefficient, and bactericidal activity. Vancomycin is the primary choice for methicillin-resistant Staphylococcus aureus (MRSA) infection treatment; rifampicin has interesting antibiofilm properties, but its effectivity remains poorly defined. METHODS: Rifampicin activity alone and in combination with vancomycin against biofilm-forming MRSA was investigated, using a twofold serial broth microtiter method, biofilm challenge, and bacterial count recovery. RESULTS: Minimal inhibitory concentration (MIC) and minimal bactericidal concentration for vancomycin and rifampicin ranged from 0.5 to 1mg/l and 0.008 to 4mg/l, and from 1 to 4mg/l and 0.06 to 32mg/l, respectively. Mature biofilms were submitted to rifampicin and vancomycin exposure, and minimum biofilm eradication concentration ranged from 64 to 32,000 folds and from 32 to 512 folds higher than those for planktonic cells, respectively. Vancomycin (15mg/l) in combination with rifampicin at 6 dilutions higher each isolate MIC did not reach in vitro biofilm eradication but showed biofilm inhibitory capacity (1.43 and 0.56log10 CFU/ml reduction for weak and strong biofilm producers, respectively; p<0.05). CONCLUSIONS: In our setting, rifampicin alone failed to effectively kill biofilm-forming MRSA, demonstrating stronger inability to eradicate mature biofilm compared with vancomycin.

INTRODUÇÃO: A atividade dos antimicrobianos em biofilmes depende do seu peso molecular, de cargas positivas, coeficiente de permeabilidade e atividade bactericida. Vancomicina é a escolha primária para o tratamento de infecções causadas por Staphylococcus aureus resistentes à meticilina (MRSA) e rifampicina possui interessante propriedade antibiofilme, apesar da sua efetividade ainda ser fracamente definida. MÉTODOS: Foi investigada a atividade da rifampicina sozinha e em combinação com vancomicina frente à MRSA formadores de biofilme, utilizando o método das microplacas com diluição seriada e recuperação bacteriana em biofilme após exposição antimicrobiana. RESULTADOS: Concentração inibitória minima (MIC) e concentração bactericida mínima (MBC) para vancomicina e rifampicina foi de 0,5-1mg/l e 0,008-4mg/l; 1-4mg/l e 0,06-32mg/l, respectivamente. Biofilmes maduros foram expostos à vancomicina e rifampicina, e a concentração mínima para erradicar o biofilme (MBEC) foi 64-32.000 e 32-512 vezes maior do que para células planctônicas, respectivamente. A combinação de vancomicina (15mg/l) com rifampicina (6-diluições maior do que o MIC de cada isolado) não atingiu erradicação do biofilme in vitro, porém apresentou capacidade inibitória do biofilme formado (redução de 1,43 e 0,56log10 UFC/ml para produtores fracos e fortes, respectivamente; p<0,05). CONCLUSÕES: Rifampicina sozinha falhou em efetivamente matar MRSA formadores de biofilme, demonstrando fraca habilidade para erradicação de biofilmes maduros comparado com vancomicina.

Typing of Methicillin resistant Staphylococcus aureus: A technical review.

Increasing prevalence of Methicillin-resistant Staphylococcus aureus (MRSA) worldwide is a growing public health concern. MRSA typing is an essential component of an effective surveillance system to describe epidemiological trends and infection control strategies. Current challenges for MRSA typing are focused on selecting the most appropriate technique in terms of efficiency, reliability, ease of performance and cost involved. This review summarises the available information on application, potential and problems of various typing techniques in discriminating the strains and understanding the epidemiology of MRSA strains. The phenotypic methods in general are easier to perform, easier to interpret, cost effective and are widely available, however less discriminatory. The genotypic methods are expensive and technically demanding, however more discriminatory. Newer technologies involving sequencing of various genes are coming up as broadly applicable and high throughput typing systems. Still there is no consensus regarding the single best method for typing of MRSA strains. Phage typing is recommended as first line approach in epidemiological investigation of MRSA strains. PFGE remains the gold standard for characterisation of outbreak strains. DNA sequencing methods including MLST, spa typing, SCCmec typing and toxin gene profile typing are more practical methods for detecting evolutionary changes and transmission events. The choice of typing technique further depends on the purpose of the study, the facilities available and the utility of data generated to answer a desirable research question. A need for harmonisation of typing techniques by following standard protocols is emphasised to establish surveillance networks and facilitate global MRSA control.