- Email: [email protected]
Prevalence of 16S rRNA methylase conferring high-level aminoglycoside resistance in Escherichia coli in China
Letters to the Editor / International Journal of Antimicrobial Agents 37 (2011) 385–388
gentamicin, ciprofloxacin and/or third-generation cephalosporins increased from 7.8% at baseline to 49% after international travel. In 18% of individuals, prolonged colonisation for >6 months occurred [5]. These individuals may act as reservoirs for infection within the community or as sources of outbreak when they are hospitalised [2,3,5]. In many countries there are no national guidelines for admission screening of individuals who have a history of medical treatment abroad. Second, the optimal treatment for invasive carbapenem-resistant Enterobacteriaceae infections is uncertain and is complicated by the frequent occurrence of co-resistance to many other antibiotics. The agents that are potentially active against carbapenem-resistant Enterobacteriaceae are colistin, tigecycline, fosfomycin and isepamicin. In solid organ and bone marrow transplant recipients, infections by these organisms represent a tremendous threat. In response to these challenges, hospitals in Hong Kong have recently introduced active screening for carbapenem-resistant Enterobacteriaceae carriage at hospital admission. Under the arrangement, all newly admitted patients with history of hospitalisation, surgery or dialysis in an overseas institution in the previous 12 months would be screened. Funding: This work was supported by research grants from the Research Fund for the Control of Infectious Diseases (RFCID) of the Health, Welfare and Food Bureau of the Hong Kong SAR Government. Competing interests: None declared. Ethical approval: Not required. References [1] Cohen SJ, Leverstein-Van Hall MA; Dutch Working Party on the Detection of Highly Resistant Microorganisms. Guideline for phenotypic screening and confirmation of carbapenemases in Enterobacteriaceae. Int J Antimicrob Agents 2010;36:205–10. [2] Kitchel B, Rasheed JK, Patel JB, Srinivasan A, Navon-Venezia S, Carmeli Y, et al. Molecular epidemiology of KPC-producing Klebsiella pneumoniae isolates in the United States: clonal expansion of multilocus sequence type 258. Antimicrob Agents Chemother 2009;53:3365–70. [3] Qi Y, Wei Z, Ji S, Du X, Shen P, Yu Y. ST11, the dominant clone of KPC-producing Klebsiella pneumoniae in China. J Antimicrob Chemother 2010 [Epub ahead of print]. [4] Clinical and Laboratory Standard Institute. Performance standards for antimicrobial susceptibility testing; twentieth informational supplement. Document M100-S20. Wayne, PA: CLSI; 2010. [5] Kennedy K, Collignon P. Colonisation with Escherichia coli resistant to ‘critically important’ antibiotics: a high risk for international travellers. Eur J Clin Microbiol Infect Dis 2010;29:1501–6.
P.-L. Ho ∗ Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China C.W.S. Tse Department of Clinical Pathology, Kwong Wah Hospital, Hong Kong SAR, China E.L. Lai W.U. Lo K.H. Chow Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China ∗ Corresponding
author. Tel.: +852 2255 4193; fax: +852 2855 1241. E-mail address: [email protected] (P.-L. Ho) 24 November 2010
doi:10.1016/j.ijantimicag.2011.01.002
387
Prevalence of 16S rRNA methylase conferring high-level aminoglycoside resistance in Escherichia coli in China Sir Aminoglycosides are widely used in human and animal therapy both against Gram-negative and Gram-positive bacteria. However, owing to overuse of this antimicrobial class, resistance has become increasingly prevalent, compromising their therapeutic efficacy. Recently, several 16S rRNA methylase enzymes were found in multiple countries. Modification of 16S rRNA by these enzymes reduces binding to aminoglycosides, leading to high-level resistance to amikacin, kanamycin, tobramycin and gentamicin. Currently, seven 16S rRNA methylase genes have been identified (armA, rmtA, rmtB, rmtC, rmtD, rmtE and npmA). Dissemination of these genes by conjugation or natural transformation is an increasing concern worldwide, thus resistance gene distribution data are urgently required. The aim of this study was to investigate the occurrence of 16S rRNA methylases in Escherichia coli isolates from humans and zoo animals in Sichuan province as well as from swine in seven provinces, which, based on our knowledge, has not been reported previously. From 2007 to 2008, a total of 195 E. coli isolates were systematically collected from seven provinces (Sichuan, Fujian, Jiangxi, Henan, Zhejiang, Hunan and Gansu) in China. Zoo animals included healthy Rhinopithecus roxellana (golden snub-nosed monkey), lion, white tiger, yellow tiger, wolf, yellow deer, kangaroo, spotted deer, giraffe, slow loris and zebra from Bifengxia Valley, Sichuan Province. Escherichia coli isolates were identified by the Vitek® system (bioMérieux, Durham, NC). The minimal inhibitory concentration of kanamycin (Sigma, St Louis, MO) was determined by the broth microdilution method [1]. Antimicrobial susceptibility to gentamicin, amikacin, streptomycin, tobramycin and netilmicin (Oxoid Ltd., Basingstoke, UK) was assessed by Kirby–Bauer disk diffusion and was interpreted following the guidelines of the Clinical and Laboratory Standards Institute (CLSI) [2]. Escherichia coli ATCC 25922 was used as a reference strain. The armA, npmA, rmtB and rmtC genes were detected by polymerase chain reaction (PCR) as described in a previous study [3]. PCR products were submitted to DNA sequencing and their identity was confirmed by BLAST comparison (http://www.ncbi.nlm.nih.gov) with relevant sequences in the GenBank database. Antimicrobial susceptibility and the prevalence of resistance genes are shown in Table 1. Although the strains were isolated from different regions, most showed resistance to kanamycin and streptomycin, indicating that these antimicrobials may no longer be effective for clinical therapy not only in humans but also in animals. High-level resistance to gentamicin and amikacin varied greatly amongst the provinces. These differences might be caused by the different selective pressures exerted by aminoglycoside usage, diverse farm management or the varying quality of disease control practises. The armA and rmtB genes were detected in 63.59% and 62.05% of isolates, respectively. Across all strains tested, 96.92% of isolates contained at least one type of 16S rRNA methylase gene (armA or rmtB). Amongst isolates from different sources, the most common 16S rRNA methylase gene was armA. Furthermore, the prevalence of isolates positive for both armA and rmtB was estimated to be 55.38% (108 isolates). Overall, the resistance trends of isolates derived from different origins were nearly identical, which may be explained by two reasons. Aminoglycosides may have been used frequently to treat humans, zoo animals and
388
Letters to the Editor / International Journal of Antimicrobial Agents 37 (2011) 385–388
Table 1 Results of antimicrobial susceptibility testing and prevalence of resistance genes amongst all Escherichia coli isolates. Province
Host species
Sichuan
Humans (n = 65) Zoo animals (n = 18) Swine (n = 22) Swine (n = 15) Swine (n = 25) Swine (n = 15) Swine (n = 12) Swine (n = 8) Swine (n = 15)
Fujian Henan Jiangxi Zhejiang Hunan Gansu
Antimicrobial susceptibility (% resistant)
Prevalence of resistance genes (% positive)
KAN
GEN
AMK
STM
TOB
NET
armA
npmA
rmtB
rmtC
89.23 88.89 95.45 100.00 96.00 80.00 100.00 100.00 93.33
29.23 27.78 45.45 80.00 12.00 6.67 25.00 0.00 6.67
23.08 22.22 63.63 80.00 0.00 6.67 25.00 0.00 13.33
89.23 83.33 95.45 93.33 92.00 80.00 100.00 62.50 40.00
67.69 66.67 81.82 86.67 48.00 33.33 58.33 75.00 40.00
35.38 50.00 72.73 86.67 40.00 26.67 33.33 50.00 40.00
92.31 77.78 95.45 40.00 52.00 33.33 25.00 0.00 13.33
0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00
60.00 83.33 18.18 73.33 68.00 73.33 50.00 75.00 80.00
0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00
KAN, kanamycin (disk concentration 256 g/mL); GEN, gentamicin (120 g); AMK, amikacin (30 g); STM, streptomycin (10 g); TOB, tobramycin (10 g); NET, netilmicin (30 g).
swine infections, contributing to widespread resistance. Alternatively, since humans, zoo animals and swine have close contact with one another, resistance genes may have been transferred by conjugation or transformation. Thus, horizontal dissemination of plasmids conferring aminoglycoside resistance may have led to equivalent resistance rates amongst isolates from different sources. Of the swine and human isolates chosen for sequencing, very little sequence diversity was observed for the armA and rmtB genes. The sequences from swine and human strains were identical to one another and were nearly identical to the published sequence, suggesting the genes may have come from the same or a very close source. The armA prevalence rate in humans in this study was much higher (92.31%) than that in Europe and South Korea (10.53% and 0.8%, respectively) [4,5]. Amongst human isolates, the detection rate of armA was higher than that of rmtB (60.00%). This finding differs from studies conducted in South Korea where the rmtB gene was the most frequently present single resistance gene. In the current study, no isolates carried npmA or rmtC. To the best of our knowledge, there have been no reports of npmA- and rmtC-positive E. coli isolates in China at this time. This suggests that armA and rmtB are the main 16S rRNA methylase genes causing high-level resistance in China. The high-level aminoglycoside resistance produced by methyltransferases is a troubling development that will require advanced infection control practises to limit continued spreading as well as more effective laboratory methods for detection and surveillance. In addition, efforts to identify new aminoglycoside antibiotics that can bind to methylated ribosomes are urgently required to help address this emerging antimicrobial resistance. Funding: This research was supported by research grants from the National Science and Technology Pillar Program for Food Safety Key Technology Project during the eleventh Five-Year Plan (No. 2006BAK02A19). Competing interests: None declared. Ethical approval: Not required. References [1] Sahm DF, Washington II JA. Manual of clinical microbiology. 5th ed. Washington, DC: American Society for Microbiology; 1991.
[2] Clinical and Laboratory Standards Institute. Methods for antimicrobial susceptibility testing of anaerobic bacteria; approved standard. 7th ed. Document M11-A7. Wayne, PA: CLSI; 2007. [3] Fritsche TR, Castanheira M, Miller GH, Jones RN, Armstrong ES. Detection of methyltransferases conferring high-level resistance to aminoglycosides in Enterobacteriaceae from Europe, North America, and Latin America. Antimicrob Agents Chemother 2008;52:1843–5. [4] Kang HY, Kim KY, Kim J, Lee JC, Lee YC, Cho DT, et al. Distribution of conjugative-plasmid-mediated 16S rRNA methylase genes among amikacinresistant Enterobacteriaceae isolates collected in 1995 to 1998 and 2001 to 2006 at a university hospital in South Korea and identification of conjugative plasmids mediating dissemination of 16S rRNA methylase. J Clin Microbiol 2008;46:700–6. [5] Galimand M, Sabtcheva S, Courvalin P, Lambert T. Worldwide disseminated armA aminoglycoside resistance methylase gene is borne by composite transposon Tn1548. Antimicrob Agents Chemother 2005;49:2949–53.
Qingqing Xia 1 Hongning Wang ∗ Anyun Zhang Ting Wang Yunfei Zhang School of Life Science, Sichuan University, Animal Disease Prevention and Food Safety Key Laboratory of Sichuan Province, ‘985 Project’ Science Innovative Platform for Resource and Environment Protection of Southwestern China, Chengdu, Sichuan 610064, China ∗ Corresponding
author. Present address: 29 Wangjiang Road, School of Life Science, Sichuan University Chengdu, Sichuan 610064, China. Tel.: +86 288 547 1599; fax: +86 288 547 1599. E-mail address: [email protected] (H. Wang) 1
Present address: 1113 Vet-Med VMPM, Iowa State University, Ames, IA 50010, USA. 14 October 2010
doi:10.1016/j.ijantimicag.2011.01.004
gentamicin, ciprofloxacin and/or third-generation cephalosporins increased from 7.8% at baseline to 49% after international travel. In 18% of individuals, prolonged colonisation for >6 months occurred [5]. These individuals may act as reservoirs for infection within the community or as sources of outbreak when they are hospitalised [2,3,5]. In many countries there are no national guidelines for admission screening of individuals who have a history of medical treatment abroad. Second, the optimal treatment for invasive carbapenem-resistant Enterobacteriaceae infections is uncertain and is complicated by the frequent occurrence of co-resistance to many other antibiotics. The agents that are potentially active against carbapenem-resistant Enterobacteriaceae are colistin, tigecycline, fosfomycin and isepamicin. In solid organ and bone marrow transplant recipients, infections by these organisms represent a tremendous threat. In response to these challenges, hospitals in Hong Kong have recently introduced active screening for carbapenem-resistant Enterobacteriaceae carriage at hospital admission. Under the arrangement, all newly admitted patients with history of hospitalisation, surgery or dialysis in an overseas institution in the previous 12 months would be screened. Funding: This work was supported by research grants from the Research Fund for the Control of Infectious Diseases (RFCID) of the Health, Welfare and Food Bureau of the Hong Kong SAR Government. Competing interests: None declared. Ethical approval: Not required. References [1] Cohen SJ, Leverstein-Van Hall MA; Dutch Working Party on the Detection of Highly Resistant Microorganisms. Guideline for phenotypic screening and confirmation of carbapenemases in Enterobacteriaceae. Int J Antimicrob Agents 2010;36:205–10. [2] Kitchel B, Rasheed JK, Patel JB, Srinivasan A, Navon-Venezia S, Carmeli Y, et al. Molecular epidemiology of KPC-producing Klebsiella pneumoniae isolates in the United States: clonal expansion of multilocus sequence type 258. Antimicrob Agents Chemother 2009;53:3365–70. [3] Qi Y, Wei Z, Ji S, Du X, Shen P, Yu Y. ST11, the dominant clone of KPC-producing Klebsiella pneumoniae in China. J Antimicrob Chemother 2010 [Epub ahead of print]. [4] Clinical and Laboratory Standard Institute. Performance standards for antimicrobial susceptibility testing; twentieth informational supplement. Document M100-S20. Wayne, PA: CLSI; 2010. [5] Kennedy K, Collignon P. Colonisation with Escherichia coli resistant to ‘critically important’ antibiotics: a high risk for international travellers. Eur J Clin Microbiol Infect Dis 2010;29:1501–6.
P.-L. Ho ∗ Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China C.W.S. Tse Department of Clinical Pathology, Kwong Wah Hospital, Hong Kong SAR, China E.L. Lai W.U. Lo K.H. Chow Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China ∗ Corresponding
author. Tel.: +852 2255 4193; fax: +852 2855 1241. E-mail address: [email protected] (P.-L. Ho) 24 November 2010
doi:10.1016/j.ijantimicag.2011.01.002
387
Prevalence of 16S rRNA methylase conferring high-level aminoglycoside resistance in Escherichia coli in China Sir Aminoglycosides are widely used in human and animal therapy both against Gram-negative and Gram-positive bacteria. However, owing to overuse of this antimicrobial class, resistance has become increasingly prevalent, compromising their therapeutic efficacy. Recently, several 16S rRNA methylase enzymes were found in multiple countries. Modification of 16S rRNA by these enzymes reduces binding to aminoglycosides, leading to high-level resistance to amikacin, kanamycin, tobramycin and gentamicin. Currently, seven 16S rRNA methylase genes have been identified (armA, rmtA, rmtB, rmtC, rmtD, rmtE and npmA). Dissemination of these genes by conjugation or natural transformation is an increasing concern worldwide, thus resistance gene distribution data are urgently required. The aim of this study was to investigate the occurrence of 16S rRNA methylases in Escherichia coli isolates from humans and zoo animals in Sichuan province as well as from swine in seven provinces, which, based on our knowledge, has not been reported previously. From 2007 to 2008, a total of 195 E. coli isolates were systematically collected from seven provinces (Sichuan, Fujian, Jiangxi, Henan, Zhejiang, Hunan and Gansu) in China. Zoo animals included healthy Rhinopithecus roxellana (golden snub-nosed monkey), lion, white tiger, yellow tiger, wolf, yellow deer, kangaroo, spotted deer, giraffe, slow loris and zebra from Bifengxia Valley, Sichuan Province. Escherichia coli isolates were identified by the Vitek® system (bioMérieux, Durham, NC). The minimal inhibitory concentration of kanamycin (Sigma, St Louis, MO) was determined by the broth microdilution method [1]. Antimicrobial susceptibility to gentamicin, amikacin, streptomycin, tobramycin and netilmicin (Oxoid Ltd., Basingstoke, UK) was assessed by Kirby–Bauer disk diffusion and was interpreted following the guidelines of the Clinical and Laboratory Standards Institute (CLSI) [2]. Escherichia coli ATCC 25922 was used as a reference strain. The armA, npmA, rmtB and rmtC genes were detected by polymerase chain reaction (PCR) as described in a previous study [3]. PCR products were submitted to DNA sequencing and their identity was confirmed by BLAST comparison (http://www.ncbi.nlm.nih.gov) with relevant sequences in the GenBank database. Antimicrobial susceptibility and the prevalence of resistance genes are shown in Table 1. Although the strains were isolated from different regions, most showed resistance to kanamycin and streptomycin, indicating that these antimicrobials may no longer be effective for clinical therapy not only in humans but also in animals. High-level resistance to gentamicin and amikacin varied greatly amongst the provinces. These differences might be caused by the different selective pressures exerted by aminoglycoside usage, diverse farm management or the varying quality of disease control practises. The armA and rmtB genes were detected in 63.59% and 62.05% of isolates, respectively. Across all strains tested, 96.92% of isolates contained at least one type of 16S rRNA methylase gene (armA or rmtB). Amongst isolates from different sources, the most common 16S rRNA methylase gene was armA. Furthermore, the prevalence of isolates positive for both armA and rmtB was estimated to be 55.38% (108 isolates). Overall, the resistance trends of isolates derived from different origins were nearly identical, which may be explained by two reasons. Aminoglycosides may have been used frequently to treat humans, zoo animals and
388
Letters to the Editor / International Journal of Antimicrobial Agents 37 (2011) 385–388
Table 1 Results of antimicrobial susceptibility testing and prevalence of resistance genes amongst all Escherichia coli isolates. Province
Host species
Sichuan
Humans (n = 65) Zoo animals (n = 18) Swine (n = 22) Swine (n = 15) Swine (n = 25) Swine (n = 15) Swine (n = 12) Swine (n = 8) Swine (n = 15)
Fujian Henan Jiangxi Zhejiang Hunan Gansu
Antimicrobial susceptibility (% resistant)
Prevalence of resistance genes (% positive)
KAN
GEN
AMK
STM
TOB
NET
armA
npmA
rmtB
rmtC
89.23 88.89 95.45 100.00 96.00 80.00 100.00 100.00 93.33
29.23 27.78 45.45 80.00 12.00 6.67 25.00 0.00 6.67
23.08 22.22 63.63 80.00 0.00 6.67 25.00 0.00 13.33
89.23 83.33 95.45 93.33 92.00 80.00 100.00 62.50 40.00
67.69 66.67 81.82 86.67 48.00 33.33 58.33 75.00 40.00
35.38 50.00 72.73 86.67 40.00 26.67 33.33 50.00 40.00
92.31 77.78 95.45 40.00 52.00 33.33 25.00 0.00 13.33
0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00
60.00 83.33 18.18 73.33 68.00 73.33 50.00 75.00 80.00
0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00
KAN, kanamycin (disk concentration 256 g/mL); GEN, gentamicin (120 g); AMK, amikacin (30 g); STM, streptomycin (10 g); TOB, tobramycin (10 g); NET, netilmicin (30 g).
swine infections, contributing to widespread resistance. Alternatively, since humans, zoo animals and swine have close contact with one another, resistance genes may have been transferred by conjugation or transformation. Thus, horizontal dissemination of plasmids conferring aminoglycoside resistance may have led to equivalent resistance rates amongst isolates from different sources. Of the swine and human isolates chosen for sequencing, very little sequence diversity was observed for the armA and rmtB genes. The sequences from swine and human strains were identical to one another and were nearly identical to the published sequence, suggesting the genes may have come from the same or a very close source. The armA prevalence rate in humans in this study was much higher (92.31%) than that in Europe and South Korea (10.53% and 0.8%, respectively) [4,5]. Amongst human isolates, the detection rate of armA was higher than that of rmtB (60.00%). This finding differs from studies conducted in South Korea where the rmtB gene was the most frequently present single resistance gene. In the current study, no isolates carried npmA or rmtC. To the best of our knowledge, there have been no reports of npmA- and rmtC-positive E. coli isolates in China at this time. This suggests that armA and rmtB are the main 16S rRNA methylase genes causing high-level resistance in China. The high-level aminoglycoside resistance produced by methyltransferases is a troubling development that will require advanced infection control practises to limit continued spreading as well as more effective laboratory methods for detection and surveillance. In addition, efforts to identify new aminoglycoside antibiotics that can bind to methylated ribosomes are urgently required to help address this emerging antimicrobial resistance. Funding: This research was supported by research grants from the National Science and Technology Pillar Program for Food Safety Key Technology Project during the eleventh Five-Year Plan (No. 2006BAK02A19). Competing interests: None declared. Ethical approval: Not required. References [1] Sahm DF, Washington II JA. Manual of clinical microbiology. 5th ed. Washington, DC: American Society for Microbiology; 1991.
[2] Clinical and Laboratory Standards Institute. Methods for antimicrobial susceptibility testing of anaerobic bacteria; approved standard. 7th ed. Document M11-A7. Wayne, PA: CLSI; 2007. [3] Fritsche TR, Castanheira M, Miller GH, Jones RN, Armstrong ES. Detection of methyltransferases conferring high-level resistance to aminoglycosides in Enterobacteriaceae from Europe, North America, and Latin America. Antimicrob Agents Chemother 2008;52:1843–5. [4] Kang HY, Kim KY, Kim J, Lee JC, Lee YC, Cho DT, et al. Distribution of conjugative-plasmid-mediated 16S rRNA methylase genes among amikacinresistant Enterobacteriaceae isolates collected in 1995 to 1998 and 2001 to 2006 at a university hospital in South Korea and identification of conjugative plasmids mediating dissemination of 16S rRNA methylase. J Clin Microbiol 2008;46:700–6. [5] Galimand M, Sabtcheva S, Courvalin P, Lambert T. Worldwide disseminated armA aminoglycoside resistance methylase gene is borne by composite transposon Tn1548. Antimicrob Agents Chemother 2005;49:2949–53.
Qingqing Xia 1 Hongning Wang ∗ Anyun Zhang Ting Wang Yunfei Zhang School of Life Science, Sichuan University, Animal Disease Prevention and Food Safety Key Laboratory of Sichuan Province, ‘985 Project’ Science Innovative Platform for Resource and Environment Protection of Southwestern China, Chengdu, Sichuan 610064, China ∗ Corresponding
author. Present address: 29 Wangjiang Road, School of Life Science, Sichuan University Chengdu, Sichuan 610064, China. Tel.: +86 288 547 1599; fax: +86 288 547 1599. E-mail address: [email protected] (H. Wang) 1
Present address: 1113 Vet-Med VMPM, Iowa State University, Ames, IA 50010, USA. 14 October 2010
doi:10.1016/j.ijantimicag.2011.01.004