Occurrence of the vanA gene in Staphylococcus epidermidis from nasopharyngeal secretion of Health-Care Workers, Recife, Brazil

Abstract INTRODUCTION: The increasing reports of vancomycin-resistant Staphylococcus strains (VRS) haves caused concern worldwide, from the laboratory detection to patient management. This study aimed to identify the occurrence of VRS strains among healthcare professionals from a university hospital. METHODS: A total of 102 Staphylococcus sp. isolates from healthcare professionals, obtained in a previous study were evaluated according to standard techniques for VRS detection. RESULTS: After screening inoculation of plates containing 6µg/ml of vancomycin, 19 resistant isolates were identified. The susceptibility profile to other antimicrobials revealed 18 multidrug resistant isolates. The minimum inhibitory concentration (MIC) was determined by E-test and broth microdilution. According to E-tests, of 19 isolates grown in BHI-V6, four isolates presented MIC ≥ 128 µg/ml, seven with MIC ranging from 4 to 8 µg/ml, and eight with MIC ≤ 2µg/ml. By broth microdilution, 14 isolates presented MIC ≤ 2 µg/ml and five with MIC ≥ 16µg/ml. The presence of the gene vanA was determined by PCR in the five resistant isolates, and this gene was detected in one of the strains. Furthermore, among the 19 strains, the gene mecA was found in 13 (39,4%) isolates, including the strain carrying the gene vanA. CONCLUSIONS: Based on these results, we highlight the presence of one strain carrying both vanA and the mecA genes, as well as multidrug-resistant strains colonizing healthcare professionals, and their importance as potential vectors to spread strains carrying resistance genes in the hospital environment.

Molecular epidemiology of coagulase-negative bloodstream isolates: detection of Staphylococcus epidermidis ST2, ST7 and linezolid-resistant ST23

Abstract The mechanisms contributing to persistence of coagulase-negative staphylococci are diverse; to better understanding of their dynamics, the characterization of nosocomial isolates is needed. Our aim was to characterize phenotypic and molecular characteristics of Staphylococcus epidermidis and Staphylococcus haemolyticus human blood isolates from two tertiary care hospitals in Mexico, the Hospital Universitario in Monterrey and the Hospital Civil in Guadalajara. Antimicrobial susceptibility was determined. Biofilm formation was assessed by crystal violet staining. Detection of the ica operon and Staphylococcal Cassette Chromosome mec typing were performed by PCR. Clonal relatedness was determined by Pulsed-fiel gel electrophoresis and Multi locus sequence typing. Methicillin-resistance was 85.5% and 93.2% for S. epidermidis and S. haemolyticus, respectively. Both species showed resistance >70% to norfloxacin, clindamycin, levofloxacin, trimethoprim/sulfamethoxazole, and erythromycin. Three S. epidermidis and two S. haemolyticus isolates were linezolid-resistant (one isolate of each species was cfr+). Most isolates of both species were strong biofilm producers (92.8% of S. epidermidis and 72.9% of S. haemolyticus). The ica operon was amplified in 36 (43.4%) S. epidermidis isolates. SCCmec type IV was found in 47.2% of the S. epidermidis isolates and SCCmec type V in 14.5% of S. haemolyticus isolates. No clonal relatedness was found in either species. Resistance to clindamycin, levofloxacin, erythromycin, oxacillin, and cefoxitin was associated with biofilm production for both species (p < 0.05). A G2576T mutation in 23S rRNA gene was detected in an S. haemolyticus linezolid-resistant isolate. All linezolid-resistant S. epidermidis isolates belonged to ST23; isolate with SCCmec type IV belonged to ST7, and isolate with SCCmec type III belonged to ST2. This is the first report of ST7 in Mexico. There was a high genetic diversity in both species, though both species shared characteristics that may contibute to virulence.

The prevalence of genotypes that determine resistance to macrolides, lincosamides, and streptogramins B compared with spiramycin susceptibility among erythromycin-resistant Staphylococcus epidermidis

Coagulase-negative staphylococci, particularly Staphylococcus epidermidis, can be regarded as potential reservoirs of resistance genes for pathogenic strains, e.g., Staphylococcus aureus. The aim of this study was to assess the prevalence of different resistance phenotypes to macrolide, lincosamide, and streptogramins B (MLSB) antibiotics among erythromycin-resistant S. epidermidis, together with the evaluation of genes promoting the following different types of MLSB resistance:ermA, ermB, ermC,msrA, mphC, and linA/A’. Susceptibility to spiramycin was also examined. Among 75 erythromycin-resistantS. epidermidis isolates, the most frequent phenotypes were macrolides and streptogramins B (MSB) and constitutive MLSB (cMLSB). Moreover, all strains with the cMLSB phenotype and the majority of inducible MLSB (iMLSB) isolates were resistant to spiramycin, whereas strains with the MSB phenotype were sensitive to this antibiotic. The D-shape zone of inhibition around the clindamycin disc near the spiramycin disc was found for some spiramycin-resistant strains with the iMLSB phenotype, suggesting an induction of resistance to clindamycin by this 16-membered macrolide. The most frequently isolated gene was ermC, irrespective of the MLSB resistance phenotype, whereas the most often noted gene combination wasermC, mphC, linA/A’. The results obtained showed that the genes responsible for different mechanisms of MLSB resistance in S. epidermidis generally coexist, often without the phenotypic expression of each of them.

Methicillin-resistant Staphylococcus sp. colonizing health care workers of a cancer hospital

The aim of the study was to analyze epidemiological and microbiological aspects of oral colonization by methicillin-resistant Staphylococcus of health care workers in a cancer hospital. Interview and saliva sampling were performed with 149 health care workers. Antimicrobial resistance was determined by disk diffusion and minimum inhibitory concentration. Polymerase Chain Reaction, Internal Transcribed Spacer-Polymerase Chain Reaction and Pulsed Field Gel Electrophoresis were performed for genotypic characterization of methicillin-resistant Staphylococcus. Risk factors were determined by logistic regression. Methicillin-resistant Staphylococcus colonization prevalence was 19.5%, denture wearing (p = 0.03), habit of nail biting (p = 0.04) and preparation and administration of antimicrobial (p = 0.04) were risk factors identified. All methicillin-resistant Staphylococcus were S. epidermidis, 94.4% of them had mecA gene. Closely related and indistinguishable methicillin-resistant S. epidermidis were detected. These results highlight that HCWs which have contact with patient at high risk for developing infections were identified as colonized by MRSE in the oral cavity, reinforcing this cavity as a reservoir of these bacteria and the risk to themselves and patients safety, because these microorganisms may be spread by coughing and talking.

Genotypic study documents divergence in the pathogenesis of bloodstream infection related central venous catheters in neonates

OBJECTIVE: To investigate the pathogenesis of bloodstream infection by Staphylococcus epidermidis, using the molecular epidemiology, in high-risk neonates. METHODS: We conducted a prospective study of a cohort of neonates with bloodstream infection using central venous catheters for more than 24 h. "National Healthcare Safety Network" surveillance was conducted. Genotyping was performed by DNA fingerprinting and mecA genes and icaAD were detected by multiplex-PCR. RESULTS: From April 2006 to April 2008, the incidence of bloodstream infection and central venous catheter-associated bloodstream infection was 15.1 and 13.0/1000 catheter days, respectively, with S. epidermidis accounting for 42.9% of episodes. Molecular analysis was used to document the similarity among six isolates of bloodstream infection by S. epidermidis from cases with positive blood and central venous catheter tip cultures. Fifty percent of neonates had bloodstream infection not identified as definite or probable central venous catheter-related bloodstream infection. Only one case was considered as definite central venous catheter-related bloodstream infection and was extraluminally acquired; the remaining were considered probable central venous catheter-related bloodstream infections, with one probable extraluminally and another probable intraluminally acquired bloodstream infection. Additionally, among mecA+ and icaAD+ samples, one clone (A) was predominant (80%). A polyclonal profile was found among sensitive samples that were not carriers of the icaAD gene. CONCLUSIONS: The majority of infections caused by S. epidermidis in neonates had an unknown origin, although 33.3% appeared to have been acquired intraluminally and extraluminally. We observed a polyclonal profile between sensitive samples and a prevalent clone (A) between resistant samples. .

Dissimilarity of ccrAB gene sequences between methicillin-resistant Staphylococcus epidermidis and methicillin-resistant Staphylococcus aureus among bovine isolates in Korea

The sequences of the ccrAB genes from bovine-, canine- and chicken-originating methicillin-resistant Staphylococcus (S.) epidermidis (MRSE) and bovine methicillin-resistant Staphylococcus (S.) aureus (MRSA) were compared to investigate the frequency of intra-species horizontal transfer of the staphylococcal cassette chromosome mec (SCCmec) complex. Nineteen MRSE strains were isolated from bovine milk, chickens, and dogs, and their genetic characteristics were investigated by multilocus sequence typing and SCCmec typing. Among the animal MRSE strains, the most frequent SCCmec type was type IV, which consisted of the type B mec complex and ccrAB type 2. The ccrA2 and ccrB2 genes were sequenced from the bovine, chicken and canine MRSE strains and compared with those of the bovine MRSA strains. The sequences generally clustered as MRSA and MRSE groups, regardless of the animal source. Additionally, no bovine MRSE sequence was associated with the bovine MRSA groups. Although most of the bovine MRSE and MRSA isolates possessed SCCmec type IV sequences, our results suggest that the intra-species gene transfer of the SCCmec complex between bovine S. aureus and bovine S. epidermidis strains is not a frequent event.

Caracterización fenotípica de Staphylococcus epidermidis aislado de pacientes con endoftalmitis / Phenotypic characterization of Staphylococcus epidermidis isolated from patients with endophthalmitis

Objetivo: Conocer las características fenotípicas de Staphylococcus epidermidis aislado de endoftalmitis relacionadas con el implante de lente intraocular de metilmetacrilato, su capacidad para formar biofilm y adherencia a proteínas de matriz extracelular y poliestireno. Métodos: Se estudiaron cinco cepas de Staphylococcus epidermidis aisladas de enfermos con endoftalmitis posterior la extracción del cristalino con implante de lente intraocular. Se investigó si estas cepas se adhieren a poliestireno, a colágena tipo I y a fibronectina, así como si las bacterias eran formadoras de biofilm. Al final se extrajeron las proteínas de superficie de las bacterias y se analizaron por electroforesis en gel de poliacrilamida. Resultados: Se encontró que las cinco cepas se unieron al poliestireno y que lo hicieron con mayor eficacia en la fase de crecimiento exponencial, con máxima adherencia a los 105 minutos; las cinco cepas se adhirieron a fibronectina y solo dos (CV y EN) a colágena. Dos cepas (CV y EN) fueron débiles formadoras de biofilm. Se identificaron proteínas que por peso molecular corresponden con las informadas en la literatura como proteínas de unión a biomateriales. Conclusiones: Las cepas estudiadas al no ser formadoras de biofilm tendrían que ser consideradas no patógenas, pero cumplen con el paso inicial de la patogenicidad, la adherencia, además de que fueron aisladas de un proceso infeccioso intraocular y produjeron endoftalmitis cuando fueron inoculadas en ojos de conejo.

OBJECTIVE: To carry out the phenotypic characterization of Staphylococcus epidermidis isolated from endophthalmitis developed after cataract extraction and implantation of an intraocular lens. This bacteria produces a biofilm, adheres to polystyrene and host proteins such as collagen and fibronectine, significant virulence factors. METHODS: Five S. epidermidis strains were isolated from cases of endophthalmitis, they developed after crystalline extraction and implantation of an intraocular lens. We assessed if these strains adhere to polystyrene, to Type I collagen and to fibronectine and if bacteria produced biofilm. Finally, the bacterial surface proteins were obtained and analyzed using polyacrylamide gel electrophoresis. RESULTS: All five bacterial strains adhered to polystyrene, with a maximum adherence time of 105 min; they also displayed adherence to fibronectine but only two to collagen. Only two strains were weak biofilm producers. We identified proteins that by molecular weight are similar to those identified in the literature as proteins binding to biomaterials. CONCLUSIONS: As the strains that we studied were not biofilm-forming they should be considered as non-pathogenic. Nevertheless, they meet the initial criteria of pathogenicity and adherence, aside from being isolated from an intraocular infectious process and being able to provoke endophtalmitis when inoculated in rabbit eyes.

Hospital strain colonization by Staphylococcus epidermidis

The skin and mucous membranes of healthy subjects are colonized by strains of Staphylococcus epidermidis showing a high diversity of genomic DNA polymorphisms. Prolonged hospitalization and the use of invasive procedures promote changes in the microbiota with subsequent colonization by hospital strains. We report here a patient with prolonged hospitalization due to chronic pancreatitis who was treated with multiple antibiotics, invasive procedures and abdominal surgery. We studied the dynamics of skin colonization by S. epidermidis leading to the development of catheter-related infections and compared the genotypic profile of clinical and microbiota strains by pulsed field gel electrophoresis. During hospitalization, the normal S. epidermidis skin microbiota exhibiting a polymorphic genomic DNA profile was replaced with a hospital-acquired biofilm-producer S. epidermidis strain that subsequently caused repetitive catheter-related infections.

Phenotypic and plasmid pattern analysis of Staphylococcus epidermidis in bacterial keratitis.

BACKGROUND: Staphylococcus epidermidis, a commensal of the conjunctival sac has been incriminated as the commonest etiological agent of bacterial keratitis. However, the pathogenic potential of this commensal organism is not clearly known. AIM: To determine any phenotypic, molecular markers of S. epidermidis pathogenicity in bacterial keratitis. MATERIALS AND METHODS: A total of 382 corneal ulcer isolates of S. epidermidis and 87 S. epidermidis isolates from healthy eyes (controls) were studied. Speciation, biotyping and antibiotic sensitivity testing were performed by conventional methods. Tube slime and adherence tests were carried out by recommended techniques. Plasmid analysis was conducted by a standard protocol. STATISTICAL ANALYSIS: Chi-square test was employed for calculations. RESULTS: Out of 382 corneal ulcer isolates (Pathogens) 284 (74.3%) belonged to biotypes I and II. Slime was detected in 164 (42.9%) of 382 pathogens vs. 21 (24.1%) of 87 controls (P<0.001). Sixty-five (39.6%) of 164 slime positive isolates were multidrug-resistant as compared to only 49 (22.4%) of 218 slime negative isolates (P<0.001). A significantly higher number i.e, 73.1% (120/164) of slime-producers possessed a 21 Kb plasmid in contrast to only 53.2% (116/218) of nonslime-producers (P<0.001). Presence of this plasmid had a statistical correlation of low significance with multidrug resistance (P=0.04). One hundred and seventy-two (45.0%) of 382 pathogens and 24 (27.6%) of the 87 controls were adherent to artificial surfaces (P=0.003) and the majority of the adherent organisms (99/172, 57.6%) were slime producers (P<0.001). CONCLUSIONS: Slime was associated with multidrug resistance in corneal ulcer isolates of S. epidermidis. The 21 Kb plasmid could determine virulence as it was responsible for slime production and adherence.