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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.

Keywords:
Staphylococcus; Healthcare professionals; Vancomycin; Multiresistance; Methicillin

INTRODUCTION

Infections due to methicillin-resistant staphylococcal strains (MRS) present increased risk of treatment failure, and for some time glycopeptides, such as vancomycin and teicoplanin, have been the only therapeutic option, which justifies the crescent use of this class of drugs11. Burnham CAD, Weber CJ, Dunne Jr WM. Novel screening agar for detection of vancomycin-nonsusceptible Staphylococcus aureus. J Clin Microbiol. 2010;48(3):949-51.),(22. Melo GB, Melo MC, Carvalho KS, Gontijo Filho PP. Vancomycin-resistant Staphylococcus aureus and coagulase-negative staphylococci in a Brazilian University Hospital. J Bas Appl Pharma Scien. 2009;30(1):55-61.. Some antimicrobials that exhibit in vitro activity against Staphylococcus aureus remain active against strains resistant to glycopeptides such as rifampicin and fusidic acid33. Howden BP, Davies JK, Johnson PDR, Stinear TP, Grayson ML Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection and clinical implications. Clin Microbiol Rev. 2010;23(1):99-139.. In recent years, a limited number of new antimicrobials have been developed. Among them, the 5th generation cephalosporins, ceftaroline and ceftobiprole have been shown to be effective against MRSA isolates44. Batista BG, Rauber JM, Bruschi JS, d’Azevedo PA. New cephalosporin as an alternative for treatment of infections by methicillin-resistant Staphylococcus aureus (MRSA). Rev Epidemiol Control Infect. 2015;5(2):94-9.. Other antimicrobials not belonging to the group of beta-lactams and with activity against these micro-organisms, including linezolid, daptomycin and tigecycline, have been available since the beginning of the 21st century and are widely employed in clinical practice55. Rodvold KA, McConeghy KW. Methicillin-resistant Staphylococcus aureus therapy: past, present, and future. Clin Infect Dis. 2014;58(Suppl 1):S20-7..

The indiscriminate use of vancomycin therapy to treat hospital infections has allowed for the emergence of S. aureus isolates and other staphylococcal species with reduced sensitivity to vancomycin and other glycopeptides66. Thati V, Shivannavar CT, Gaddad SM. Vancomycin resistance among methicillin-resistant Staphylococcus aureus isolates from intensive care units of tertiary care hospitals in Hyderabad, Indian J Med Res. 2011;134(5):704-8., such as vancomycin/glycopeptide intermediate resistant S. aureus (VISA/GISA) and vancomycin/glycopeptide resistant S. aureus (VRSA/GRSA)33. Howden BP, Davies JK, Johnson PDR, Stinear TP, Grayson ML Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection and clinical implications. Clin Microbiol Rev. 2010;23(1):99-139.. Studies have reported an increased number of glycopeptide-resistant Staphylococcus epidermidis (GRSE) worldwide77. Cremniter J, Slassi A, Quincampoix JC, Tardy VS, Bauer T, Porcher R, et al. Decreased susceptibility to teicoplanin and vancomycin in coagulase-negative staphylococci isolated from orthopedic-device-associated infections. J Clin Microbiol . 2010;48(4):1428-31..

Isolates of S. aureus, especially coagulase-negative Staphylococci (CoNS) resistant to methicillin/oxacillin isolates, with reduced susceptibility to glycopeptides have been reported in Japan, the United States (US), Europe, and Asia since the end of the 80s33. Howden BP, Davies JK, Johnson PDR, Stinear TP, Grayson ML Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection and clinical implications. Clin Microbiol Rev. 2010;23(1):99-139.. In 1997, two major categories of vancomycin-resistant S. aureus had been defined: (1) vancomycin-resistant S. aureus (VRSA), carrying the vanA gene, mediating high-level resistance (MIC ≥ 16µg/ml) and (2) vancomycin-intermediate S. aureus (VISA) isolates with low-level resistance (MIC ≥ 4 to < 16µg/ml) by cell wall thickening88. Vaudaux P, Huggler E, Bernard L, Ferry T, Renzoni A, Lew DP. Underestimation of vancomycin and teicoplanin MICs by broth microdilution leads to underdetection of glicopeptide-intermediate isolates of Staphylococcus aureus. Antimicrob Agents Chemother. 2010;54(9):3861-70..

In Brazil, staphylococcal strains with reduced susceptibility to glycopeptides were reported in hospitals in São Paulo and Rio de Janeiro a decade ago99. Melo GB, Melo MC, Gama AP, Carvalho KS, Jesus TC, Bonetti AM, Gotijo Filho PP. Analysis of the genetic diversity of vancomycin-resistant Staphylococcus aureus. Braz J Microbiol. 2005;36(2):126-30., and recently S. aureus containing the vanA and vanB genes were described in Brazil1010. Rossi FMD, Diaz L, Wollam A, Panesso D, Zhou Y, Rincon S, et al. Transferable vancomycin resistance in a community-associated MRSA lineage. N Engl J Med. 2014;370(16):1524-31.),(1111. Breves A, Miranda CAC, Flores C, Filippis I, Clementino MM. Methicillin- and vancomycin-resistant Staphylococcus aureus in health care workers and medical device. Braz J Pathol Lab Med. 2015;51(3):143-52..

Colonization of diverse body sites by CoNS, and transient colonization by S. aureus, can be a source of infection for immunocompromised patients1212. Palazzo ICV, Araujo MLC, Darini ALC. First report of vancomycin-resistant staphylococci isolated from healthy carriers in brazil. J Clin Microbiol. 2005;43(1):179-85., highlighting microbiota colonization of health professionals as a source of dissemination of health care-associated infections (HAIs)1313. Rabelo MA, Bezerra Neto AM, Silva ECBF, Oliveira WLM, Melo FL, Lopes ACS, et al. Phenotypic methods for determination of methicillin resistance in Staphylococcus spp. from health care workers. Braz J Pathol Lab Med . 2013;49(2):91-6..

The transmission chain of antimicrobial resistant microorganisms in the hospital environment involves health professionals as potential source of transmission to patients, co-workers, family and community, emphasizing their importance in the context of the HAIs1313. Rabelo MA, Bezerra Neto AM, Silva ECBF, Oliveira WLM, Melo FL, Lopes ACS, et al. Phenotypic methods for determination of methicillin resistance in Staphylococcus spp. from health care workers. Braz J Pathol Lab Med . 2013;49(2):91-6.),(1414. Silva ECBF, Maciel MAV, Melo FL, Lopes ACS, Arca IS. Epidemiological surveillance and susceptibility of Staphylococcus aureus among healthcare workers at a reference hospital: preliminary assessment. Rev Inst Adolfo Lutz. 2010;69(1):126-30..

In Brazil, resistant phenotypic profiles of different Staphylococcus species, with reduced resistance to vancomycin, were detected in microbiota samples of asymptomatic carriers 1212. Palazzo ICV, Araujo MLC, Darini ALC. First report of vancomycin-resistant staphylococci isolated from healthy carriers in brazil. J Clin Microbiol. 2005;43(1):179-85.. Considering that Staphylococcus species are important pathogens associated with HAIs and there are few studies regarding vancomycin resistance in the colonizing microbiota of health professionals, the aims of this research were to identify the microorganisms colonizing the nasopharynx of health professionals and analyze the vancomycin resistance profile of these isolates by phenotypic and genetic methods.

METHODS

Study design and bacterial samples

An experimental-based study was carried out in April to December 2014 with staphylococcal isolates obtained in a previous study1515. Rabelo MA, Bezerra Neto AM, Loibman SO, Lima JLC, Ferreira EL, Leal NC, et al. The occurrence and dissemination of methicillin and vancomycin-resistantStaphylococcusin samples from patients and health professionals of a university hospital in Recife, State of Pernambuco, Brazil. Rev Soc Bras Med Trop. 2014;47(4):437-46., originated from nasopharyngeal secretions of healthcare professionals of three sectors of the University Hospital of Pernambuco, Brazil: Intensive Care Unit (ICU), Surgical Clinics, and Hemodialysis Service/Nephrology.

A total of 102 Staphylococcus strains-kept as frozen stock in Brain Heart-Infusion (BHI) broth supplemented with glycerol (20%) at -20°C and in nutrient agar slants at 4ºC-were placed in BHI broth, inoculated in 5% sheep blood agar, and incubated for 24-48h at 35ºC. Colonies with macroscopic characteristics of the genus Staphylococcus were Gram stained. Following confirmation by morphology and staining, colonies were submitted for identification using deoxyribonuclease (DNase), catalase and coagulase tests, and manitol fermentation. The identification of the staphylococcal species was made through the automated system VITEK 2. Based on these readings, profile identification was established and interpreted according to a specific algorithm. The result of the profile was compared with the database, generating the identification of the unknown organism.

Vancomycin susceptibility testing

The isolates were screened by vancomycin susceptibility test using BHI supplemented with 6µg/mL of vancomycin (BHI-V6). Inoculums were adjusted to 0.5 McFarland turbidity33. Howden BP, Davies JK, Johnson PDR, Stinear TP, Grayson ML Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection and clinical implications. Clin Microbiol Rev. 2010;23(1):99-139..

Two methods were used to determine the MIC to vancomycin: broth microdilution and E-test (BIOMÉRIEUX), according to Clinical and Laboratory Standards Institute (CLSI) guidelines1616. Clinical and Laboratory Standards Institute. CLSI M07-A9 Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard. CLSI document M07-A9. Wayne, PA: CLSI; 2012.. One clinical isolate of Enterococcus faecium harboring the vanA gene was used as a positive control1717. Vilela MA, Souza SL, Palazzo ICV, Ferreira JC, Morais Jr. MA, Darini ALC, et al. Identification and molecular characterization of Van A-type vancomycin-resistant Enterococcus faecalis in Northeast of Brazil. Mem Inst Oswaldo Cruz. 2006;101(7):716-9. and the Enterococcus faecalis (ATCC 29212) strain as the negative control. The E-test was performed using a suspension of 0.5 McFarland turbidity plated onto Müller-Hinton medium and incubated at 35ºC for 24h. The interpretation was performed according to manufacturer's specifications and compared to CLSI cut off values33. Howden BP, Davies JK, Johnson PDR, Stinear TP, Grayson ML Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection and clinical implications. Clin Microbiol Rev. 2010;23(1):99-139..

For determination of MIC by Broth Microdilution, it was determined manually in Mueller Hinton broth, according to the recommendations of the CLSI1616. Clinical and Laboratory Standards Institute. CLSI M07-A9 Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard. CLSI document M07-A9. Wayne, PA: CLSI; 2012.. Assays for vancomycin were performed in medium supplemented with Ca2 + (50mg/L). Initial inoculums of bacteria (0.5 × 105 CFU/mL) were plated onto 96-well polypropylene plates, exposed to 8 dilutions (1μg/mL to 128μg/mL) of the tested compound, and incubated for 18h at 35°C. The minimum inhibitory concentration was taken as the lowest concentration of the compound in which no visible bacterial growth was observed. According to CLSI recommendations, the bacterial isolates were categorized as resistant or susceptible using interpretive criteria1616. Clinical and Laboratory Standards Institute. CLSI M07-A9 Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard. CLSI document M07-A9. Wayne, PA: CLSI; 2012..

Agreement between E-test and microdilution was defined as minimum inhibitory concentrations (MICs) that differed by ± 1-log2 dilutions or less. Categorical agreement was defined as test results within the same susceptibility. Errors were ranked as follows: very major error, false-susceptible result by the E-test; major error, false-resistant result produced by the E-test; and minor error, intermediate result by E-test method and a resistant or susceptible category for the reference method (microdilution test), according to CLSI guidelines1919. Clinical and Laboratory Standards Institute. CLSI M23-A5 Development of In Vitro Susceptibility Testing Criteria and Quality Control Parameters, 5th Edition, document M23-A5Wayne, PA: CLSI ; 2018..

Antimicrobial susceptibility testing

The Staphylococcus isolates were tested by disc diffusion in Mueller-Hinton agar, according to the CLSI guidelines1818. Clinical and Laboratory Standards Institute. CLSI M100-S24 Performance Standards for Antimicrobial Susceptibility Testing - Twenty-Sixth Informational Supplement. CLSI document M100-S24. Wayne, PA: CLSI ; 2014., using the following antibiotics: penicillin (10U), gentamicin (10µg), clindamycin (2µg), sulfazotrim (1.25/23.75µg), ciprofloxacin (5µg), chloramphenicol (30µg), cefoxitin (30µg), erythromycin (15µg), and linezolid (5µg). After incubation for 24 hours at 35°C, the inhibition zones were measured using a caliper.

Concordance scale analysis of phenotypic vancomycin susceptibility tests

The agreement between the phenotypic tests to assess the vancomycin resistance profile were verified by the k (kappa) index2020. Organização Pan-Americana da Saúde (PAHO). Fundação Nacional de Saúde/Centro Nacional de Epidemiologia. Métodos de Investigação Epidemiológica em Doença Transmissíveis. Volume I. Brasília: PAHO; 1997. 135p..

Molecular techniques

Deoxyribonucleic acid (DNA) was extracted as described by Oliveira2121. Oliveira WLM, Mangueira EVC, Vilela MA, Paiva Júnior S, Leal NC, Almeida AMP. Diversity of Sccmec types in ant Staphylococcus spp. causing Hospital-Associated Infections. J J Microbiol Pathol. 2015;2(3):020. from vancomycin-resistant isolates detected by the different phenotypic techniques. Subsequently, it was used in polymerase chain reaction (PCR) for amplification of the genes vanA and mecA. PCR was performed using the primers and conditions as previously described (forward-5´-TGAATAACATCGGCATTAC-3´ and reverse-5´-TTATTTAACGGGGAAATC-3´)2222. Saha B, Singh AK, Ghosh A, Bal M. Identification and characterization of a vancomycin-resistant Staphylococcus aureus isolated from Kolkata (South Asia). J Med Microbiol. 2008;57:72-9. and (P1 5´-GGTCCCATTAACTCTGAAG-3´ and P3 5´-AGTTCTGCAGTACCGGATTTGC-3´)2323. Petinaki E, Arvaniti A, Dimitracopoulos G, Spiliopoulou I. Detection of mecA, mecR1 and mecI genes among clinical isolates of methicillin-resistant staphylococci by combined polymerase chain reactions. J Antimicrob Chemother. 2001; 47(3):297-303..

Sequencing of vanA gene

A positive PCR product for the vanA gene was purified by the Wizard® SV Gel kit and PCR Clean-Up System (Promega) according to the manufacturer's protocol. Following, it was quantified by spectrophotometry using the software Chromas Lite 2.1.1, Basic Local Alignment Research Tool (BLAST), and Expert Protein Analysis System (ExPASy) algorithm. The analyzed sequences of vanA were deposited in GenBank with the following accession number: KT581638.

RESULTS

In this study, we analyzed 102 isolates collected from health professionals, 31.4% (32/102) S. aureus and 68.6% (70/102) CoNS isolates. Approximately 43.1% (44/102) of the isolates were from the Surgical/Infectious and Parasitic Diseases sector, of which 22.7% (10/44) were S. aureus and 77.3% (34/44) were CoNS. Isolates from the ICU represented 20.6% (21/102), of which 38% (8/21) were S. aureus and 62% (13/21) were CoNS.

Concerning the vancomycin susceptibility, seven out of 19 isolates were S. aureus and 12 CoNS, which grew at the vancomycin screening test at a concentration of 6µg/mL (BHI-V6). The susceptibility profile to other antimicrobials showed that 18 isolates presented resistance to more than three classes of antimicrobials and were considered multi-drug resistant (MDR) strains. It is worth noting that 16 were resistant to erythromycin. Of these, 11 were also resistant to clindamycin, indicating resistance to macrolides, lincosamide, and streptogramin-B.

All 19 resistant strains, previously detected by the screening test, were evaluated by E-test for quantitative determination of the MIC to vancomycin. The seven isolates of S. aureus presented the following MIC ranges: two isolates with MIC ≤ 2μg/mL (sensitive), three with MIC between 4 and 8μg/mL, and two with MIC > 256μg/mL. In addition, of the 12 CoNS, 10 isolates presented MIC ≤ 4μg/mL and two isolates MIC ≥ 128μg/mL.

According to the MIC values obtained by broth microdilution, of the seven S. aureus isolates, five were sensitive (MIC ≤ 2µg/mL) and two were resistant (MIC ≥ 16µg/mL). Regarding the 12 CoNS isolates, nine showed MIC ≤ 2µg/mL and three MIC ≥ 32µg/mL, as shown in Table 1.

TABLE 1:
Susceptibility profile of the Staphylococcus isolates.

The kappa index coefficient of 0.96 was obtained by comparing the E-test to Broth Microdilution, indicating very good agreement between these two methods.

The agreement within 1 two-fold dilution between E-test and the broth microdilution reference method was 84%. Seven (36%) minor errors were found comparing E-test with microdilution in broth for vancomycin. There was no occurrence of major or very major errors. The categorical concordance was 93%.

Of the five isolates with MICs considered resistant, only one isolate (S. epidermidis) carried the vanA gene, which was confirmed by sequencing and showed 100% of similarity with ten sequences from Enterococcus deposited in GenBank (KT581638).

In parallel to obtaining vancomycin susceptibility, a cefoxitin susceptibility test was performed using the disc-diffusion technique, from which it was possible to observe that all the isolates were resistant to cefoxitin. After the disc-diffusion cefoxitin test, the presence of the mecA gene was investigated.

In order to genetically characterize these isolates, mecA gene detection was performed by PCR, and the gene was found in 13/19 (39.4%) isolates, including one isolate of S. aureus (MRSA) from the Nephrology/Hemodialysis Service sector. Of the 12 MRCoNS, five were obtained from the Nephrology/Hemodialysis Service sector, four from Surgical/Infectious and Parasitic Diseases sector, and three from ICU. The isolate that presented the gene vanA also exhibited the gene mecA. As for the remaining noncarrier isolates of the vanA gene, five contained the mecA gene (all MRCoNS) and 13 did not present this gene (6 MSCoNS and 7 MSSA).

DISCUSSION

In this study, we found a higher incidence of mecA gene in CoNS strains. Regarding the susceptibility profile analyses, the MRS strains were more resistant to multiple classes of antimicrobial agents than MRSA. Similar results were obtained by Costa44. Batista BG, Rauber JM, Bruschi JS, d’Azevedo PA. New cephalosporin as an alternative for treatment of infections by methicillin-resistant Staphylococcus aureus (MRSA). Rev Epidemiol Control Infect. 2015;5(2):94-9.; however, Fadeyi2525. Fadeyi A, Bolaji BO, Oyedepo OO, Adesiyun OO, Adeboye MAN, Olanrewaju TO, et al. Methicilin resistant Staphylococcus aureus carriage amongst Healthcare Workers of the Critical Care Units in a Nigerian Hospital. Am J Infect Dis. 2010;6(1):18-23. described MDR in MRSA isolates colonizing the nasopharynx of health professionals in Nigeria.

The studies regarding the analysis of susceptibility to vancomycin started approximately three decades ago, when environmental strains of S. aureus were detected with reduced susceptibility (intermediate) to this drug2626. Hiramatsu K. Vancomycin-resistant Staphylococcus aureus: a new model of antibiotic resistance. Lancet Infect Dis. 2001;1(3):147-55.. Additionally, the emergence of hetero-VRSA strains occurred in the 80s after the introduction of vancomycin use for treatment of staphylococcal infections in Japan2727. Watanakukorn C. Mode of action and in-vitro activity of vancomycin. J Antimicrob Chemother . 1984;14(Suppl D):7-18.. Several research studies related to the molecular analysis of heteroresistance vancomycin-intermediate S. aureus worldwide have been performed, following associations of these strains with persistent infections and treatment failure2828. Silveira ACO, Cunha GR, Caierão J, Cordova CMM, D’azevedo PA. Molecular epidemiology of heteroresistant vancomycin-intermediate Staphylococcus aureus in Brazil. Braz J Infect Dis. 2015;19(5):466-72..

Results of MIC to vancomycin evaluated by E-test and broth microdilution techniques demonstrated divergence. Three isolates of S. aureus showed MICs between 4 and 8μg/mL (VISA) with the microdilution method but no isolates presented similar MIC with E-test. In addition, this technique is a screening tool for heterogeneous VISA (hVISA) and VISA, but does not apply to vancomycin or teicoplanin, and the results obtained with this technique should not be reported as true MIC2929. Prakash V, Lewis JS, Jorgensen JH. Vancomycin MICs for methicillin-resistant Staphylococcus aureus isolates differ based upon the susceptibility test method used. Antimicrob. Agents Chemother. 2008;52(12):4528..

In the present study, four resistant vancomycin isolates (two VRSA and two VRS) were detected by both the E-test and broth microdilution. There was reasonable correlation between these two methods. Using comparisons between Broth Microdilution and E-test MICs results for vancomycin, it was possible to observe that essential and categorical agreements presented at 84% and 93%, respectively. Yet, a minor error of 36% was detected: however, major error and very major error were 0%.

Genetic characterization these vancomycin resistant isolates was performed by PCR for the vanA gene. During the period that the present study was conducted, vancomycin resistance was not described in other genes in Brazil1010. Rossi FMD, Diaz L, Wollam A, Panesso D, Zhou Y, Rincon S, et al. Transferable vancomycin resistance in a community-associated MRSA lineage. N Engl J Med. 2014;370(16):1524-31.. Among the isolates only one harbored the vanA gene, which was a specific isolate Staphylococcus epidermidis from HCW microbiota.

The presence of the vanA gene was not found in the four resistant vancomycin strains (VRSA and VRS), as a result, the precise genetic mechanism for vancomycin resistance in these staphylococcal strains awaits elucidation. The cell wall thickening has been reported for glycopeptide-resistant VRS and VRSA1212. Palazzo ICV, Araujo MLC, Darini ALC. First report of vancomycin-resistant staphylococci isolated from healthy carriers in brazil. J Clin Microbiol. 2005;43(1):179-85.),(1717. Vilela MA, Souza SL, Palazzo ICV, Ferreira JC, Morais Jr. MA, Darini ALC, et al. Identification and molecular characterization of Van A-type vancomycin-resistant Enterococcus faecalis in Northeast of Brazil. Mem Inst Oswaldo Cruz. 2006;101(7):716-9.),(2828. Silveira ACO, Cunha GR, Caierão J, Cordova CMM, D’azevedo PA. Molecular epidemiology of heteroresistant vancomycin-intermediate Staphylococcus aureus in Brazil. Braz J Infect Dis. 2015;19(5):466-72..

The nasopharynx microbiota of health professionals harboring resistant strains to vancomycin have already been described in the literature1313. Rabelo MA, Bezerra Neto AM, Silva ECBF, Oliveira WLM, Melo FL, Lopes ACS, et al. Phenotypic methods for determination of methicillin resistance in Staphylococcus spp. from health care workers. Braz J Pathol Lab Med . 2013;49(2):91-6.),(3030. Cui L, Ma X, Sato K, Okuma K, Tenover FC, Mamizuka EM, Gemmell CG, Kim MN, Ploy MC, El-Solh N, Ferraz V, Hiramatsu K. Cell wall thickening is a common feature of vancomycin resistance in Staphylococcus aureus. J Clin Microbiol . 2003;41:5-14.),(3131. Daef EA, Elsherbiny NM, Ibrahim MA, Ahmed EH. Decolonization of Methicillin resistant Staphylococcus areus Nasal Carriage Among Health Care Workers. Life Scien J. 2012;9(4):4496-501.; however, the first Brazilian report of S. epidermidis, harboring the vanA gene, and colonizing a health professional from ICU occurred at an University Hospital in Recife, Brazil1212. Palazzo ICV, Araujo MLC, Darini ALC. First report of vancomycin-resistant staphylococci isolated from healthy carriers in brazil. J Clin Microbiol. 2005;43(1):179-85.. Breves1111. Breves A, Miranda CAC, Flores C, Filippis I, Clementino MM. Methicillin- and vancomycin-resistant Staphylococcus aureus in health care workers and medical device. Braz J Pathol Lab Med. 2015;51(3):143-52. reported the occurrence of one S. aureus isolate obtained from the hands of a health professional, which harbored resistance genes to vancomycin (vanB) and methicillin (mecA). On the other hand, there are many studies reporting methicillin-resistant staphylococcal isolates from nasopharyngeal of health professionals and microbiota of patients3232. Saito G, Thom J, Wei Y, Gnanasuntharam P, Kreiswirth N, Willey B, et al. Methicillin-resistant Staphylococcus aureus colonization among health care workers in a downtown emergency department in Toronto, Ontario. Can J Infect Dis Med Microbiol. 2013;24(3):57-60.

33. Ruiz A, Mora M, Zurita C, Larco D, Toapanta Y, Zurita J. Prevalence of methicillin-resistant Staphylococcus aureus among health care workers of intensive care units in Ecuador. J Infect Dev Ctries. 2014;8(1):116-19.
-3434. Goud R, Gupta S, Neogi U, Agarwal D, Naidu K, Chalannavar R, Subhaschandra G. Community prevalence of methicillin and vancomycin resistant Staphylococcus aureus in and around Bangalore, southern India. Rev Soc Bras Med Trop . 2011;44(3):309-12..

Cases reports regarding microbiota colonization of vancomycin-resistant staphylococcal strains obtained from patients at clinics or hospitals are scarce. In India, two studies have reported patients harboring S. aureus with the vanA gene3535. Banerjee T, Anupurba S. Colonization with Vancomycin-Intermediate Staphylococcus aureus strains containing the vanA resistance gene in a Tertiary-Care Center in North India. J Clin Microbiol . 2012;50(5):1730-32.),(3636. Center for Disease Control and Prevention (CDC). Staphylococcus aureus resistant to vancomycin - United States, 2002. Morb Mortal Wkly Rep MMWR. 2002;51:565-7..

The occurrence of HAIs caused by Staphylococcus isolates carrying the vanA gene has been reported worldwide, associated with different patterns of infections, mainly in the US and in Brazil1111. Breves A, Miranda CAC, Flores C, Filippis I, Clementino MM. Methicillin- and vancomycin-resistant Staphylococcus aureus in health care workers and medical device. Braz J Pathol Lab Med. 2015;51(3):143-52.),(2222. Saha B, Singh AK, Ghosh A, Bal M. Identification and characterization of a vancomycin-resistant Staphylococcus aureus isolated from Kolkata (South Asia). J Med Microbiol. 2008;57:72-9.),(3737. Chang S, Sievert DM, Hageman JC, Boulton ML, Tenover FC, Downes FP, et al. Infection with vancomycin-resistant Staphylococcus aureus containing the vanA resistance gene. N Engl J Med . 2003;348(14):1342-47.

38. Aligholi M, Emaneini M, Jabalameli F, Shahsavan S, Dabiri H, Sedaght H. Emergence of high-level vancomycin-resistant Staphylococcus aureus in the Imam Khomeini Hospital in Tehran. Med Princ Pract. 2008;17(5):432-4.

39. Sievert DM, Rudrick JT, Patel JB, McDonald LC, Wilkins MJ, Hageman JC. Vancomycin-resistant Staphylococcus aureus in United States, 2002-2006. Clin Infect Dis .2008;46(5):668-74.

40. Foucault ML, Courvalin P, Grillot-Courvalin C. Fitness cost of vanA-type vancomycin resistance in methicillin-resistant Staphylococcus aureus. Antimicrob Agents Chemother . 2009;53(6):2354-9.
-4141. Perichon B, Courvalin P. VanA-type vancomycin-resistant Staphylococcus aureus. Antimicrob Agents Chemother . 2009;53(11):4580-7.),(2424. Costa DM, Kipnis A, Vasconcelos LSL, Vilefort LOR, Telles S A, André MC, et al. Staphylococcus sp. colonizing health care workers of a cancer hospital. Braz J Microbiol . 2014;45(3):799-805..

Some studies correlate methicillin resistance to vancomycin tolerance due to vancomycin treatment failures in cases of infections caused by methicillin-resistant microorganisms77. Cremniter J, Slassi A, Quincampoix JC, Tardy VS, Bauer T, Porcher R, et al. Decreased susceptibility to teicoplanin and vancomycin in coagulase-negative staphylococci isolated from orthopedic-device-associated infections. J Clin Microbiol . 2010;48(4):1428-31.),(4242. Soriano A, Marco F, Martínez JA, Pisos E, Almela M, Dimova VP, et al. Influence of vancomycin minimum inhibitory concentration on the treatment of methicillin-resistant Staphylococcus aureus bacteremia. Clin Infect Dis . 2008;46(2):193-200.

43. Kos VN, Desjardins CA, Griggs A, Cerqueira G, Van Tonder A, Holden MTG, et al. Comparative genomics of vancomycin-resistant Staphylococcus aureus strains and their positions within the clade most commonly associated with methicillin-resistant S. aureus hospital-acquired infection in the United States. mBio. 2012;3(3):e-00112-12
-4444. Domínguez VC, Córdova AC, Ochoa AS, Escalona G, Galindo JA, Leviz AR, Castro RH, et al. Vancomycin tolerant, methicillin-resistant Staphylococcus aureus reveals the effects of vancomycin on cell wall thickening. PLoS One. 2015;10(3):1-16.. In 2006, three strains were reported, two S. aureus and one CoNS, with resistance to both antibiotics vancomycin and methicillin4545. Tiwari HK, Sen MR. Emergence of vancomycin resistant Staphylococcus aureus (VRSA) from a tertiary care hospital from northern part of India. BMC Infect Dis. 2006;6(156):1-6..

A potential emergence of vancomycin resistance may occur in hospitals in Brazil, as reported in recent studies99. Melo GB, Melo MC, Gama AP, Carvalho KS, Jesus TC, Bonetti AM, Gotijo Filho PP. Analysis of the genetic diversity of vancomycin-resistant Staphylococcus aureus. Braz J Microbiol. 2005;36(2):126-30.),(2828. Silveira ACO, Cunha GR, Caierão J, Cordova CMM, D’azevedo PA. Molecular epidemiology of heteroresistant vancomycin-intermediate Staphylococcus aureus in Brazil. Braz J Infect Dis. 2015;19(5):466-72.. This suggests the need for constant monitoring of susceptibility patterns to vancomycin, application of molecular methods and heteroresistance detection, as well as adoption of control measures to avoid the spread of these strains in the hospital environment.

Acknowledgments

We thank our collaborator Dr. Marcia Maria Camargo de Morais who provided the positive control enterococcal strain for the vanA gene.

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  • Financial support: This work was financially supported by Fundação de Amparo Ciência do Estado de Pernambuco (FACEPE) to MAVM (Process APQ 0579-2.12/06) and the Conselho Nacional Científico e Tecnológico (CNPq) Grant to MAVM (Process #474131/2011-4).

Publication Dates

  • Publication in this collection
    May-Jun 2018

History

  • Received
    04 May 2017
  • Accepted
    06 June 2018
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