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Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada
G Model ANTAGE-4308; No. of Pages 2
ARTICLE IN PRESS International Journal of Antimicrobial Agents xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
International Journal of Antimicrobial Agents journal homepage: http://www.elsevier.com/locate/ijantimicag
Letter to the Editor
Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada Sir, The Gram-negative bacterium Bordetella pertussis causes respiratory infections in humans. Despite decreasing numbers of reported cases in Canada since the introduction of the whole-cell pertussis vaccine in 1943 and the acellular vaccine in 1997/1998, periodic outbreaks have continued to occur in the province of Ontario over the last decade. Possible contributing factors to these outbreaks have been hypothesised to be waning immunity in vaccinated individuals, more sensitive laboratory detection methods, undervaccinated subpopulations and the emergence of new strains [1]. Most recently in 2012/2013, an outbreak in southwestern Ontario involving 443 cases was monitored by local and provincial public health authorities. Case management guidelines recommended treatment with antibiotics at the discretion of the attending healthcare provider with azithromycin, clarithromycin or erythromycin as indicated by the guidelines from the Public Health Agency of Canada [2]. Since treatment of B. pertussis infection is primarily mediated by antibiotics, it is important to monitor antimicrobial susceptibility profiles in order to provide effective treatment guidance. The possibility of new strains as well as overall uncertainty about the cause and implications of the reemergence of pertussis also make the need for good surveillance of the organism a high priority, including baseline levels of antimicrobial susceptibility, which has not previously been done in Ontario. Here we investigated the antibiotic susceptibility profiles of B. pertussis isolates collected before and during an outbreak in Ontario that occurred between November 2011 and April 2013. In addition to susceptibility testing, all isolates were investigated for macrolide resistance by molecular methods. Culture-positive isolates (only PCR-positive specimens as previously described [3] were plated on charcoal media and incubated at 37 ◦ C for 7 days) were tested for antimicrobial susceptibility and mutations known to cause macrolide resistance as follows. Isolates were cultured on plain charcoal agar plates (Public Health Ontario Laboratories, Toronto, Ontario, Canada) incubated at 37 ◦ C and Etest was performed for phenotypic susceptibility as previously described [4] with the following modifications. The cell concentration was adjusted to obtain a 0.5 McFarland standard, only azithromycin and trimethoprim/sulfamethoxazole (SXT) Etest strips (bioMérieux, St-Laurent, Quebec) were used, and minimum inhibitory concentrations (MICs) were recorded after 4 days. PCR amplification was performed on a 521-bp region in domain V of 23S rRNA centred on the peptidyl transferase region (the target
for macrolide class antimicrobial agents) where a G→A nucleotide mutation at position 2058 (Escherichia coli numbering) confers macrolide resistance in B. pertussis. Sequence analysis was conducted using Vector NTI software (Life Technologies Inc., Carlsbad, CA). From September 2011 to December 2012, 8082 specimens were received by Public Health Ontario Laboratories for detection of B. pertussis, of which 720 (8.9%) were positive by PCR and 295 (3.7%) were also positive by culture. A total of 275 viable isolates (275/295; 93.2%) were tested for antimicrobial susceptibility; 112 were from the regions affected by the outbreak. All isolates were found to be susceptible to both antibiotics (azithromycin range, <0.016 g/mL; SXT range, <0.002–0.32 g/mL) (Fig. 1). Moreover, no mutations were found in the amplified 23S rRNA, including those that have been associated with the development of resistance to macrolides. Although it was well established in the early 2000s that macrolide resistance in B. pertussis isolates is rare and that routine surveillance is not necessary, continued reporting of sporadic resistant isolates [5] and the occurrence of large-scale outbreaks warrants periodic monitoring. In addition, monitoring of resistance to recommended alternative treatments, mainly SXT, has not traditionally been performed. The increase in the number of pertussis cases during outbreaks presents an opportunity for assessment of levels of resistance in the community as well as guiding further investigations into outbreak management protocols as needed. The absence of elevated MICs and mutations in 23S rRNA signifies that this outbreak was not associated with antimicrobial resistance in B. pertussis. A potential pitfall of focusing on outbreaks is that the isolates are more likely to be similar to each other and may not be representative of the full population of circulating strains. Additional genomic analysis such as multilocus sequence typing would yield additional information for linking antimicrobial resistance to outbreak or other circulating strains. The lack of standardised methods tied to clinical effectiveness for assessing antimicrobial resistance in B. pertussis is not yet problematic due to the low MICs observed. Moreover, sequencing of 23S rRNA provides an additional means of screening isolates for potential mutations known to confer high levels of macrolide resistance that can then be confirmed by phenotypic analyses. As PCR-based methods are now commonly employed for the detection of B. pertussis, it is important that bacterial culture methods be incorporated into laboratory surveillance practices along with periodic assessments for susceptibility to facilitate investigations into the potential emergence of antimicrobial resistance and other virulence characteristics. This is also important for other bacterial pathogens, including those that have not yet emerged as resistant.
http://dx.doi.org/10.1016/j.ijantimicag.2014.04.001 0924-8579/© 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.
Please cite this article in press as: Marchand-Austin A, et al. Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada. Int J Antimicrob Agents (2014), http://dx.doi.org/10.1016/j.ijantimicag.2014.04.001
G Model ANTAGE-4308; No. of Pages 2
ARTICLE IN PRESS Letter to the Editor / International Journal of Antimicrobial Agents xxx (2014) xxx–xxx
2
Nader Memari 1 Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 Samir N. Patel a,b Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 a
Patrick Tang Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 Shelley L. Deeks a,b Surveillance and Epidemiology, Public Health Ontario, 480 University Avenue, Toronto, Ontario, Canada M5G 1V2 b Dalla Lana School of Public Health, University of Toronto, 155 College Street, 6th Floor, Toronto, Ontario, Canada M5T 3M7 a
Fig. 1. Distribution of minimum inhibitory concentrations (MICs) for trimethoprim/sulfamethoxazole as determined by Etest for Bordetella pertussis.
Frances B. Jamieson a,b Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 a
Acknowledgments The authors would like to acknowledge the staff of the public health units for the identification of cases and specimen submission; the respiratory bacteriology section of Public Health Ontario Laboratories for specimen workup; and the Public Health Ontario Surveillance and Epidemiology section for funding support. Funding: Research was funded by Public Health Ontario (Canada). Competing interests: None declared. Ethical approval: Not required. References
Natasha S. Crowcroft a,b,c Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Surveillance and Epidemiology, Public Health Ontario, 480 University Avenue, Toronto, Ontario, Canada M5G 1V2 c Dalla Lana School of Public Health, University of Toronto, 155 College Street, 6th Floor, Toronto, Ontario, Canada M5T 3M7 a
David J. Farrell a,b,2 Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 a
[1] Waters V, Jamieson F, Richardson S, Finkelstein M, Wormsbecker A, Halperin SA. Outbreak of atypical pertussis detected by polymerase chain reaction in immunized preschool-aged children. Pediatr Infect Dis J 2009;28:582–7. [2] National consensus conference on pertussis, Toronto, May 25–28, 2002. Can Commun Dis Rep 2003;29, 1–33 (Eng), 1–36 (Fre). [3] Guthrie JL, Robertson AV, Tang P, Jamieson F, Drews SJ. Novel duplex real-time PCR assay detects Bordetella holmesii in specimens from patients with pertussislike symptoms in Ontario, Canada. J Clin Microbiol 2010;48:1435–7. [4] Fry NK, Duncan J, Vaghji L, George RC, Harrison TG. Antimicrobial susceptibility testing of historical and recent clinical isolates of Bordetella pertussis in the United Kingdom using the Etest method. Eur J Clin Microbiol Infect Dis 2010;29:1183–5. [5] Guillot S, Descours G, Gillet Y, Etienne J, Floret D, Guiso N. Macrolideresistant Bordetella pertussis infection in newborn girl, France. Emerg Infect Dis 2012;18:966–8.
Alex Marchand-Austin a,b,∗,1 a Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1
∗ Corresponding author. Present address: Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1. Tel.: +1 416 235 5815; fax: +1 416 235 6550. E-mail address: [email protected] (A. Marchand-Austin) 1 2
These authors contributed equally to this work.
Present address: JMI Laboratories, 345 Beaver Kreek Centre, Suite A, North Liberty, IA 52317, USA. 9 December 2013
Please cite this article in press as: Marchand-Austin A, et al. Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada. Int J Antimicrob Agents (2014), http://dx.doi.org/10.1016/j.ijantimicag.2014.04.001
ARTICLE IN PRESS International Journal of Antimicrobial Agents xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
International Journal of Antimicrobial Agents journal homepage: http://www.elsevier.com/locate/ijantimicag
Letter to the Editor
Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada Sir, The Gram-negative bacterium Bordetella pertussis causes respiratory infections in humans. Despite decreasing numbers of reported cases in Canada since the introduction of the whole-cell pertussis vaccine in 1943 and the acellular vaccine in 1997/1998, periodic outbreaks have continued to occur in the province of Ontario over the last decade. Possible contributing factors to these outbreaks have been hypothesised to be waning immunity in vaccinated individuals, more sensitive laboratory detection methods, undervaccinated subpopulations and the emergence of new strains [1]. Most recently in 2012/2013, an outbreak in southwestern Ontario involving 443 cases was monitored by local and provincial public health authorities. Case management guidelines recommended treatment with antibiotics at the discretion of the attending healthcare provider with azithromycin, clarithromycin or erythromycin as indicated by the guidelines from the Public Health Agency of Canada [2]. Since treatment of B. pertussis infection is primarily mediated by antibiotics, it is important to monitor antimicrobial susceptibility profiles in order to provide effective treatment guidance. The possibility of new strains as well as overall uncertainty about the cause and implications of the reemergence of pertussis also make the need for good surveillance of the organism a high priority, including baseline levels of antimicrobial susceptibility, which has not previously been done in Ontario. Here we investigated the antibiotic susceptibility profiles of B. pertussis isolates collected before and during an outbreak in Ontario that occurred between November 2011 and April 2013. In addition to susceptibility testing, all isolates were investigated for macrolide resistance by molecular methods. Culture-positive isolates (only PCR-positive specimens as previously described [3] were plated on charcoal media and incubated at 37 ◦ C for 7 days) were tested for antimicrobial susceptibility and mutations known to cause macrolide resistance as follows. Isolates were cultured on plain charcoal agar plates (Public Health Ontario Laboratories, Toronto, Ontario, Canada) incubated at 37 ◦ C and Etest was performed for phenotypic susceptibility as previously described [4] with the following modifications. The cell concentration was adjusted to obtain a 0.5 McFarland standard, only azithromycin and trimethoprim/sulfamethoxazole (SXT) Etest strips (bioMérieux, St-Laurent, Quebec) were used, and minimum inhibitory concentrations (MICs) were recorded after 4 days. PCR amplification was performed on a 521-bp region in domain V of 23S rRNA centred on the peptidyl transferase region (the target
for macrolide class antimicrobial agents) where a G→A nucleotide mutation at position 2058 (Escherichia coli numbering) confers macrolide resistance in B. pertussis. Sequence analysis was conducted using Vector NTI software (Life Technologies Inc., Carlsbad, CA). From September 2011 to December 2012, 8082 specimens were received by Public Health Ontario Laboratories for detection of B. pertussis, of which 720 (8.9%) were positive by PCR and 295 (3.7%) were also positive by culture. A total of 275 viable isolates (275/295; 93.2%) were tested for antimicrobial susceptibility; 112 were from the regions affected by the outbreak. All isolates were found to be susceptible to both antibiotics (azithromycin range, <0.016 g/mL; SXT range, <0.002–0.32 g/mL) (Fig. 1). Moreover, no mutations were found in the amplified 23S rRNA, including those that have been associated with the development of resistance to macrolides. Although it was well established in the early 2000s that macrolide resistance in B. pertussis isolates is rare and that routine surveillance is not necessary, continued reporting of sporadic resistant isolates [5] and the occurrence of large-scale outbreaks warrants periodic monitoring. In addition, monitoring of resistance to recommended alternative treatments, mainly SXT, has not traditionally been performed. The increase in the number of pertussis cases during outbreaks presents an opportunity for assessment of levels of resistance in the community as well as guiding further investigations into outbreak management protocols as needed. The absence of elevated MICs and mutations in 23S rRNA signifies that this outbreak was not associated with antimicrobial resistance in B. pertussis. A potential pitfall of focusing on outbreaks is that the isolates are more likely to be similar to each other and may not be representative of the full population of circulating strains. Additional genomic analysis such as multilocus sequence typing would yield additional information for linking antimicrobial resistance to outbreak or other circulating strains. The lack of standardised methods tied to clinical effectiveness for assessing antimicrobial resistance in B. pertussis is not yet problematic due to the low MICs observed. Moreover, sequencing of 23S rRNA provides an additional means of screening isolates for potential mutations known to confer high levels of macrolide resistance that can then be confirmed by phenotypic analyses. As PCR-based methods are now commonly employed for the detection of B. pertussis, it is important that bacterial culture methods be incorporated into laboratory surveillance practices along with periodic assessments for susceptibility to facilitate investigations into the potential emergence of antimicrobial resistance and other virulence characteristics. This is also important for other bacterial pathogens, including those that have not yet emerged as resistant.
http://dx.doi.org/10.1016/j.ijantimicag.2014.04.001 0924-8579/© 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.
Please cite this article in press as: Marchand-Austin A, et al. Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada. Int J Antimicrob Agents (2014), http://dx.doi.org/10.1016/j.ijantimicag.2014.04.001
G Model ANTAGE-4308; No. of Pages 2
ARTICLE IN PRESS Letter to the Editor / International Journal of Antimicrobial Agents xxx (2014) xxx–xxx
2
Nader Memari 1 Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 Samir N. Patel a,b Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 a
Patrick Tang Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 Shelley L. Deeks a,b Surveillance and Epidemiology, Public Health Ontario, 480 University Avenue, Toronto, Ontario, Canada M5G 1V2 b Dalla Lana School of Public Health, University of Toronto, 155 College Street, 6th Floor, Toronto, Ontario, Canada M5T 3M7 a
Fig. 1. Distribution of minimum inhibitory concentrations (MICs) for trimethoprim/sulfamethoxazole as determined by Etest for Bordetella pertussis.
Frances B. Jamieson a,b Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 a
Acknowledgments The authors would like to acknowledge the staff of the public health units for the identification of cases and specimen submission; the respiratory bacteriology section of Public Health Ontario Laboratories for specimen workup; and the Public Health Ontario Surveillance and Epidemiology section for funding support. Funding: Research was funded by Public Health Ontario (Canada). Competing interests: None declared. Ethical approval: Not required. References
Natasha S. Crowcroft a,b,c Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Surveillance and Epidemiology, Public Health Ontario, 480 University Avenue, Toronto, Ontario, Canada M5G 1V2 c Dalla Lana School of Public Health, University of Toronto, 155 College Street, 6th Floor, Toronto, Ontario, Canada M5T 3M7 a
David J. Farrell a,b,2 Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1 a
[1] Waters V, Jamieson F, Richardson S, Finkelstein M, Wormsbecker A, Halperin SA. Outbreak of atypical pertussis detected by polymerase chain reaction in immunized preschool-aged children. Pediatr Infect Dis J 2009;28:582–7. [2] National consensus conference on pertussis, Toronto, May 25–28, 2002. Can Commun Dis Rep 2003;29, 1–33 (Eng), 1–36 (Fre). [3] Guthrie JL, Robertson AV, Tang P, Jamieson F, Drews SJ. Novel duplex real-time PCR assay detects Bordetella holmesii in specimens from patients with pertussislike symptoms in Ontario, Canada. J Clin Microbiol 2010;48:1435–7. [4] Fry NK, Duncan J, Vaghji L, George RC, Harrison TG. Antimicrobial susceptibility testing of historical and recent clinical isolates of Bordetella pertussis in the United Kingdom using the Etest method. Eur J Clin Microbiol Infect Dis 2010;29:1183–5. [5] Guillot S, Descours G, Gillet Y, Etienne J, Floret D, Guiso N. Macrolideresistant Bordetella pertussis infection in newborn girl, France. Emerg Infect Dis 2012;18:966–8.
Alex Marchand-Austin a,b,∗,1 a Department of Laboratory Medicine and Pathobiology, Faculty of Medicine, University of Toronto, Medical Sciences Building, 1 King’s College Circle, 6th Floor, Toronto, Ontario, Canada M5S 1A8 b Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1
∗ Corresponding author. Present address: Public Health Ontario Laboratories, Public Health Ontario, 81 Resources Road, Toronto, Ontario, Canada M9P 3T1. Tel.: +1 416 235 5815; fax: +1 416 235 6550. E-mail address: [email protected] (A. Marchand-Austin) 1 2
These authors contributed equally to this work.
Present address: JMI Laboratories, 345 Beaver Kreek Centre, Suite A, North Liberty, IA 52317, USA. 9 December 2013
Please cite this article in press as: Marchand-Austin A, et al. Surveillance of antimicrobial resistance in contemporary clinical isolates of Bordetella pertussis in Ontario, Canada. Int J Antimicrob Agents (2014), http://dx.doi.org/10.1016/j.ijantimicag.2014.04.001