Os reservatórios animais de Enterococos Resistentes à Vancomicina (VRE) têm um importante papel na epidemiologia destas bactérias e dos respectivos genes de resistência. O presente estudo examinou a presença de VRE em amostras fecais obtidas de primatas não-humanos. Foram analisados os perfis de resistência, as características de virulência e a variabilidade genética dos isolados. A amostragem incluiu macacos Prego (Cebus apella, n=28) e Sagüis do cerrado (Callithrix penicillata, n=37) alojados no Centro de Primatologia da Universidade de Brasília, Brasil. A maioria dos indivíduos amostrados foram macacos apreendidos na região Centro-Oeste e Sudeste do Brasil (n=48). Assim, foram coletados swabs retais e realizado o isolamento seletivo, seguido da Reação de Polimerização em Cadeia (PCR) multiplex para identificar espécies e genes de resistência. Não foram isolados enterococos contendo os genes vanA ou vanB. A porcentagem de enterococos variou de 1,5% para E. casseliflavus e E. gallinarum VanC até 12,3% (n=8) para Enterococcus faecalis. A totalidade dos isolados da espécie E. faecalis demonstrou sensibilidade aos antimicrobianos vancomicina, teicoplanina, ampicilina, gentamicina e estreptomicina. Os genes de virulência ace e esp foram prevalentes (100%, 87.5%). A análise em multilocus de repetições em tandem de número variável (MLVA) revelou diversidade no número de repetições entre os isolados de E. faecalis, que foi mais alta para espC, efa5 e efa6. Foram identificados seis diferentes genotipos de MVLA, divergindo daqueles já descritos em humanos. Os genotipos foram ainda agrupados em dois genogrupos, demonstrando especificidade de hospedeiro para as espécies Cebus apella ou Callithrix penicillata. Concluindo, não foram isoladas linhagens de enterococos contendo os genes vanA ou vanB colonizando as espécies de primatas analisadas. O presente estudo demonstrou que os isolados de E. faecalis obtidos de primatas não-humanos apresentam determinantes de virulência e possuem a habilidade de disseminar linhagens entre diferentes indivíduos.

The animal reservoirs of vancomycin-resistant enterococci (VRE) have important role in the epidemiology of the bacteria and resistant genes. The present work searched fecal samples taken off nonhuman primates for the presence of VRE. Resistance profiles, virulence traits, and genetic variability among enterococci isolates were also analyzed. The samples included Capuchin monkeys (Cebus apella, n=28) and Common marmoset (Callithrix penicillata, n=37) housed in the Primate Center of the University of Brasília, Brazil. Most individuals were captive monkeys from the Central-West and South-East regions of Brazil (n=48). We collected rectal swabs and carried out selective isolation followed by multiplex Polymerase Chain Reaction (PCR) to identify species and resistance genes. No vanA or vanB-containing enterococci were found. The carriage rates ranged from 1.5% for the VanC-type E. casseliflavus and E. gallinarum until 12.3% (n=8) for Enterococcus faecalis. All E. faecalis isolates showed susceptibility to vancomycin, teicoplanin, ampicillin, gentamicin, and streptomycin. The virulence genes ace and esp were prevalent (100.0%, 87.5%). Multilocus variable number of tandem repeats (MLVA) revealed diversity in the number of repeats among E. faecalis isolates and targets, which was higher for espC, efa5, and efa6. We identified six different MLVA genotypes that were divergent from those described in human beings. Also, they were clustered into two genogroups that showed host-specificity for the species Cebus apella or Callithrix penicillata. In conclusion, no vanA- or vanB-containing enterococci were found colonizing those primate individuals. This finding suggested that the primate individuals investigated in our study are not directly involved in the epidemiological chain of high-level vancomycin-resistant genes vanA or vanB in Brazil. Our study also showed that E. faecalis isolated from nonhuman primates carry virulence traits and have ability to spread their lineages among different individuals.

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®, 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 µg/mL and >48 µg/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 screenning in rectal samples.

We studied the antimicrobial resistance and the molecular epidemiology of 99 enterococcal surveillance isolates from two hospitals of Brasília, Brazil. Conventional biochemical tests were used to identify the enterococcal species and the disk diffusion method was used to determine their resistance profiles. Enterococcus faecalis (76%) and E. faecium (9%) were the most prevalent species. No enterococci showed the vanA or vanB vancomycin resistance phenotypes or genotypes. Only the intrinsically resistant species E. gallinarum (n=2) and E. casseliflavus (n=3) harbored the vancomycin-resistance genes vanC1 and vanC2/3, respectively. We found E. faecalis isolates with high-level resistance to gentamicin (22%) and streptomycin (8%) and both E. faecalis and E. faecium isolates with resistance to more than two antimicrobials (84% and 67%, respectively). Nine E. faecalis isolates (12%) were resistant to ampicillin; the minimal inhibitory concentration (MIC) values were 16µg/mL (n=6) and 32µg/mL (n=3). Among these ampicillin-resistant E. faecalis, seven were also resistant to gentamicin, ciprofloxacin, rifampin, penicillin, chloramphenicol, tetracycline and erythromycin. Pulsed-field gel electrophoresis classified those isolates in three different genotypes, suggesting dissemination of genetically related ampicillin-resistant E. faecalis strains among different patients.