Pedro Alves d'Azevedo

Acute endophthalmitis following intravitreal bevacizumab (Avastin) injection.

Purpose:To report two cases of acute endophthalmitis following intravitreal bevacizumab injection.Methods:Two patients with exudative age-related macular degeneration were treated sequentially with an intravitreal injection of bevacizumab and developed signs of severe but painless infectious endophthalmitis 2 days later. Vitreous samples were obtained, followed by the injection of vancomycin 1u2009mg/0.1u2009ml and ceftazidime 2.25u2009mg/0.1u2009ml. Pulsed-field gel electrophoresis (PFGE) was used to determine whether the isolated microorganisms were the same.Results:Coagulase-negative staphylococci were identified and isolated from the vitreous specimen of both patients. PFGE revealed different patterns of banding, excluding that interpatient contamination occured.Conclusions:Infectious endophthalmitis is a potential complication of intravitreal bevacizumab injection.

Staphylococcus hominis subsp. novobiosepticus strains causing nosocomial bloodstream infection in Brazil

OBJECTIVESnTo report the isolation of six Staphylococcus hominis subsp. novobiosepticus (SHN) strains from hospitalized patients with bloodstream infections in two Brazilian hospitals and to characterize their susceptibility profile to several antimicrobials.nnnMETHODSnSpecies identification was performed by biochemical methods and sodA gene sequencing. The MICs of antimicrobials were determined by broth and agar dilution methods and by Etest. Isolates were typed by PFGE and PCR amplification was used to detect the ccr gene complex and the mec class. Morphometric evaluation of cell wall was performed by transmission electron microscopy (TEM).nnnRESULTSnSusceptibility profiles indicated that the majority of isolates (five) were multidrug-resistant. Overlapping and multiplex PCR showed that five out of the six strains harboured SCCmec type III with class A mec and type 3 ccr. The initial vancomycin MIC value of 4 mg/L for these strains increased to 16-32 mg/L after growth for 10 days in BHI broth supplemented with this antimicrobial. TEM indicated that vancomycin resistance was associated with cell wall thickening and to another mechanism not fully elucidated. Only one SHN strain was oxacillin- and vancomycin-susceptible. The nosocomial infections in at least five of the patients from both hospitals were caused by a single clone of SHN.nnnCONCLUSIONSnIt is very important to consider SHN strains as the cause of nosocomial infections. The clinical implications resulting from the pattern of multidrug resistance in these strains may be complicated by the emergence of vancomycin resistance.

[Culture proven bacterial endophthalmitis: a 6-year review].

PURPOSEnTo assess the distribution of microorganisms isolated from patients with bacterial endophthalmitis and their antimicrobial susceptibility.nnnMETHODSnRetrospective analysis of medical and microbiological records of patients with suspected diagnosis of endophthalmitis and bacterial culture-proven at the Department of Ophthalmology, UNIFESP, between January 1 2000 and December 31 2005.nnnRESULTSn153 (33.9%) of 451 patients showed positive bacterial culture. A total of 155 microorganisms were isolated, 79.35% were gram-positive and 20.65% gram-negative. Staphylococcus (CoNS) (41.94%) were the most frequently isolated. The antimicrobial susceptibility for gram-negative microorganisms was as follows: amikacin 87.10%, tobramycin 80.65%, ciprofloxacin 96.67%, levofloxacin, gatifloxacin and moxifloxacin 100%, ceftazidime 85.0%, and gentamicin 80.65%. Vancomycin sensitivity among gram-positive microorganisms was 100%. S. aureus and CoNS showed 83.33% of susceptibility to oxacillin, 89.61% to ciprofloxacin and 100% to gatifloxacin and moxifloxacin. The main acquisition mechanism was postoperative (60.65%).nnnCONCLUSIONnWe detected a low sensitivity of vitreous/aqueous culture for the etiologic diagnosis of endophthalmitis. The empiric antimicrobial therapy or prophylaxis should be active against gram-positive bacteria, particularly staphylococci. Surveillance studies of bacterial resistance are important for a better utilization of antimicrobials in this clinical setting.

Rapid detection of Vancomycin-Resistant Enterococci (VRE) in rectal samples from patients admitted to intensive care units

The reduction in time required to identify vancomycin-resistant enterococci (VRE) has gained increased importance during hospital outbreaks. In the present study, we implemented a laboratory protocol to speed up the VRE screening from rectal samples. The protocol combines a medium for selective VRE isolation (VREBAC(R), Probac, São Paulo) and a multiplex PCR for detection and identification of vanA and vanB resistance genes. The screening performance was analyzed in 114 specimens collected from four intensive care units. The swabs were collected at two periods: (1) during a VRE outbreak (February 2006, n=83 patients) and (2) at the post-outbreak period, after adoption of infection control measures (June 2006, n=31 patients). Forty-one/83 VRE (49.4%) and 3/31(9.7%) VRE were found at the first and second period, respectively. All isolates harbored the vanA gene. In both periods, detection of the gene vanA parallels to the minimum inhibitory concentration values of >256 microg/mL and >48 microg/mL for vancomycin and teicoplanin, respectively. Multiplex PCR and conventional methods agreed in 90.2% for enterococci identification. Besides this accuracy, we also found a remarkable reduction in time to obtain results. Detection of enterococcal species and identification of vancomycin resistance genes were ready in 29.5 hours, in comparison to 72 hours needed by the conventional methods. In conclusion, our protocol identified properly and rapidly enterococci species and vancomycin-resistance genes. The results strongly encourage its adoption by microbiology laboratories for VRE screening in rectal samples.

Molecular characterization of vancomycin-resistant Enterococci strains eight years apart from its first isolation in São Paulo, Brazil

E. faecium was the first reported VRE species, carrying the vanA gene in Brazil. In spite of this, vancomycin-resistant E. faecalis has become the predominant species in Brazilian hospitals. The aim of this study was to evaluate the genetic relatedness of VREs isolated in a Brazilian teaching hospital eight years apart from its first isolation. We analyzed 38 VRE strains obtained from 81 surveillance cultures of patients admitted to the four largest intensive care units in Hospital São Paulo in February, 2006. Presence of the vanA gene was assayed by PCR and PFGE analysis was used for molecular characterization. All VRE strains carried the vanA gene. Two distinct clonal groups were observed among vancomycin-resistant E. faecalis. Vancomycin-resistant E. faecium belonged to five distinct clones were demonstrated by molecular typing. All of these clones were different from the first vancomycin-resistant enterococci clone isolated eight years ago in our hospital.

Laboratory detection methods for methicillin resistance in coagulase negative Staphylococcus isolated from ophthalmic infections

PURPOSEnTo evaluate different methods of oxacillin susceptibility testing of ocular isolates, considering polymerase chain reaction (PCR) as the gold standard, and to compare the in vitro susceptibility to oxacillin with that of other antimicrobials used in ophthalmologic practice.nnnMETHODSnThe Vitek gram-positive identification card was used to identify ocular coagulase negative Staphylococcus species. The presence of the mecA gene was determined by the polymerase chain reaction assay with a combination of two primer sets (mecA and 16S rRNA) in a single region. Results were analyzed and compared with other oxacillin susceptibility methods: PBP2a detection by rapid slide latex agglutination test (SLA); oxacillin E-test; the Vitek automated gram-positive susceptibility card (GPS-105); the oxacillin salt agar screening test (OSAS) at a concentration of 6.0, 1.0 and 0.75 microg oxacillin per ml and the cefoxitin disk diffusion test (CDD). Automated susceptibility was also determined to other antimicrobial agents (fluoroquinolones, penicillin G, amoxicillin-ampicillin, cefazolin, ampicillin-sulbactam, erythromycin, clindamycin, gentamicin, tetracycline, trimethoprim-sulfamethoxazole, vancomycin and rifampin.nnnRESULTSnOf the 69 CoNS isolates tested, 71% were mecA-positive and 29% mecA-negative. All methods tested had a statistically significant agreement with polymerase chain reaction. There was a tendency of positive polymerase chain reaction predomination among the S. epidermidis isolates in comparison to non-epidermidis isolates, although this was not statistically significant (78.3% vs. 56.5%; chi2= 2.54; P= 0.11). The oxacillin salt agar screening test (0.75 microg oxacillin/ml) showed the best performance, with 100% sensitivity and negative predictive value; 95% specificity and 98% positive predictive value. Using the E-test, the mecA-positive isolates were statistically significantly more resistant to ciprofloxacin, ofloxacin, gatifloxacin and moxifloxacin (P= 0.002; P= 0.008; P= 0.002 and P= 0.003, respectively). There was a statistically significant higher proportion of resistance of the coagulase negative Staphylococcus mecA-positives for: penicillin G, amoxicillin-ampicillin, cefazolin, ampicillin-sulbactam, erythromycin, clindamycin, gentamicin and tetracycline (P< or =0.05). All coagulase negative Staphylococcus species were susceptible to vancomycin and there was no statistically significant correlation between the mecA-positive isolates and resistance to trimethoprim-sulfamethoxazole or to rifampin.nnnCONCLUSIONSnIn the present study, we found that the E-test and the oxacillin salt agar screening test S (0.75 microg oxacillin per ml), when compared with polymerase chain reaction, were the most accurate currently available methods to phenotypically detect oxacillin resistance of coagulase negative Staphylococcus species. This study demonstrated that a good option for screening of ocular isolates for oxacillin resistance in the microbiology laboratory is the cefoxitin disk diffusion test and the automated Vitek system. We believe it is important to have available methods that accurately detect methicillin resistance of the less commonly encountered species, chiefly because of their increasing importance as opportunistic pathogens.

Suscetibilidade antimicrobiana in vitro dos Staphylococcus coagulase negativa oculares

PURPOSE: To assess the in vitro susceptibility of conjunctival and corneal coagulase negative Staphylococcus (CoNS) to methicillin, fluoroquinolones and aminoglycosides. METHODS: A total of 707 conjunctival and corneal CoNS disk diffusion test results were retrospectively analyzed, from January 2000 through December 2003. RESULTS: From 2000 to 2003, there was an increase in number of CoNS isolated from conjunctiva (n=57 to n=153) and cornea (n=28 to n=78). The frequency of conjunctival and corneal methicillin-resistant CoNS also increased (1.8 to 19.6% and 14.3 to 29.3%, respectively). There was no statistically significant difference between fluoroquinolones-resistant CoNS percentages in conjunctiva (ofloxacin: 1.8 to 7.8% and ciprofloxacin: 3.5 to 9.2%) and cornea (ofloxacin: 14.3 to 9.0% and ciprofloxacin: 14.3 to 10.3%). Evaluating the results of the conjunctival samples, there was increased resistance to tobramycin (15.8 to 34.0%) and to gentamycin (10.5 to 25.5%). There was no change in resistance of corneal isolates to tobramycin (28.6 to 26.9%) and to gentamycin (21.4 to 23.1%). CONCLUSIONS: there was a decrease in in vitro CoNS susceptibility to methicillin, tobramycin and gentamycin. Fuoroquinolones represented by ofloxacin and ciprofloxacin demonstrated stable in vitro susceptibility.

Pneumonia necrotizante por Staphylococcus aureus resistente à meticilina

The aim of this study was to describe a case of necrotizing pneumonia caused by methicillin-resistant Staphylococcus aureus. The sample was isolated from a blood culture collected less than 48 hours after hospital admission. The patient had been healthy until the infectious process started. The isolate had the mecA gene with staphylococcal cassette chromosome mec (SCCmec) type IVa. The possibility that Staphylococcus aureus harboring this genetic determinant might be present in our setting should be considered in situations of severe pneumonia within the community.

Outbreak of Staphylococcus hominis subsp. novobiosepticus bloodstream infections in São Paulo city, Brazil.

Coagulase-negative staphylococci (CoNS) are currently recognized as one of the most important causes of nosocomial infections worldwide, principally related to bloodstream infections (Pfaller et al., 1999). Staphylococcus epidermidis is the most frequent CoNS species associated with these infections (Rupp & Archer, 1994), but results of surveillance studies indicate a varying frequency, depending on the geographical region (Yamazumi et al., 2001; Sader et al., 2001). Additionally, a substantial increase in the frequency of meticillin resistance among CoNS isolates has occurred over recent decades. According to the results of the SENTRY study, about 80 % of CoNS strains isolated from bloodstream infections in Brazilian hospitals are resistant to meticillin (Sader et al., 2001). Glycopeptides are usually the treatment of choice for infections caused by these micro-organisms. However, due to the emergence of vancomycin-resistant enterococci and staphylococci, reduction in the use of this antimicrobial agent has been recommended (Hiramatsu, 1998). The accurate detection of meticillinresistant CoNS isolates by clinical microbiology laboratories is of crucial importance in guiding therapy and promoting the correct use of glycopeptides (Yamazumi et al., 2001; Hussain et al., 1998). Recently, a novel subspecies of CoNS, Staphylococcus hominis subsp. novobiosepticus, was isolated from blood and other clinical specimens (Kloos et al., 1998). The subspecies name derives from the combination of novobio, pertaining to the property of novobiocin resistance, and septicus, pertaining to the ability to cause sepsis. This pathogen usually exhibits multidrug resistance, including resistance to oxacillin and other antimicrobial agents (Kloos et al., 1998; Chaves et al., 2005). Meticillin-resistant isolates and those resistant to other antimicrobials are particularly important because they narrow therapeutic options. Recently, Chaves et al. (2005) related a nosocomial outbreak caused by an S. hominis subsp. novobiosepticus clone in a neonatal intensive care unit in the city of Madrid, Spain.

Staphylococcus cohnii spp urealyticus: relato de caso de um patógeno incomum

Coagulase-negative Staphylococcus has emerged as an important agent in nosocomial infections. In this study, we report a case of bacteremia associated with a central venous catheter, caused by Staphylococcus cohnii spp urealyticus that was isolated in blood cultures from a 53-year-old male patient who was admitted to a general hospital in the city of São Paulo. We discuss in this report the difficulty in routinely identifying this microorganism in the clinical microbiology laboratory. Staphylococcus cohnii spp urealyticus is a microorganism found in human skin as part of the normal microbiota, and it can cause serious infections in humans, in some situations.Coagulase-negative Staphylococcus has emerged as an important agent in nosocomial infections. In this study, we report a case of bacteremia associated with a central venous catheter, caused by Staphylococcus cohnii spp urealyticus that was isolated in blood cultures from a 53-year-old male patient who was admitted to a general hospital in the city of Sao Paulo. We discuss in this report the difficulty in routinely identifying this microorganism in the clinical microbiology laboratory. Staphylococcus cohnii spp urealyticus is a microorganism found in human skin as part of the normal microbiota, and it can cause serious infections in humans, in some situations.