- Email: [email protected]
Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital
G Model
JGAR-82; No. of Pages 4 Journal of Global Antimicrobial Resistance xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Journal of Global Antimicrobial Resistance journal homepage: www.elsevier.com/locate/jgar
Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital Charlotte A. Huber a, Anna L. Sartor a, Francis McOdimba b, Reena Shah b, Patricia Shivachi b, Hanna E. Sidjabat a, Gunturu Revathi b, David L. Paterson a,* a
The University of Queensland, UQ Centre for Clinical Research, Queensland, Building 71/918, Royal Brisbane and Women’s Hospital Campus, Herston, QLD 4029, Australia Aga Khan University Hospital, Division of Microbiology, Department of Pathology, 3rd Parklands Avenue, P.O. Box 30270-00100, Nairobi, Kenya
b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 27 September 2013 Received in revised form 25 March 2014 Accepted 28 March 2014
Acinetobacter baumannii is a serious nosocomial pathogen with a high propensity to cause outbreaks. Whilst outbreaks of A. baumannii have been reported in many regions worldwide, few data are available from East Africa. In this study, 25 A. baumannii isolates derived from a single institution located in Nairobi, Kenya, between September 2010 and September 2011 were examined. Antimicrobial susceptibility testing was performed by the disc diffusion method and the relatedness among the isolates was examined by pulsed-field gel electrophoresis, repetitive sequence-based PCR (rep-PCR) and multilocus sequence typing. The examined isolates clustered into three distinct groups. The most prevalent sequence type (ST) was ST110 (17 isolates), followed by ST92 (5 isolates) and ST109 (3 isolates). All isolates exhibited resistance to cefepime, ceftazidime, ticarcillin/clavulanic acid, cefotaxime/clavulanic acid, piperacillin/tazobactam, cefoxitin, ciprofloxacin, gentamicin, nitrofurantoin, fosfomycin trometamol, trimethoprim/sulfamethoxazole, amikacin, meropenem and imipenem, with the exception of four isolates. Two isolates belonging to ST92 and two isolates belonging to ST109 were susceptible to amikacin; one of these amikacin-susceptible ST109 isolates was also susceptible to meropenem and imipenem. All isolates were positive for OXA 51-like and all carbapenem-resistant isolates were OXA-23 positive. ß 2014 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.
Keywords: Acinetobacter baumannii Antibiotic susceptibility Outbreak Sequence type
1. Introduction Acinetobacter baumannii is a Gram-negative coccobacillus that has emerged as a serious healthcare-associated pathogen in recent years. A. baumannii infection may result in pneumonia, bloodstream infection, urinary tract infection, meningitis and wound infection as well as other manifestations of infection [1]. A. baumannii has a high propensity to cause nosocomial outbreaks, possibly due to its persistence in the hospital environment and its remarkable ability to acquire drug resistance. Several studies indicate significantly increased resistance rates in epidemic A. baumannii strains compared with sporadic strains [1]. To distinguish outbreak strains from epidemiologically unrelated A. baumannii strains, typing methods such as pulsed-field gel electrophoresis (PFGE), repetitive sequence-based PCR (rep-PCR)
* Corresponding author. Tel.: +61 7 3346 5434; fax: +61 7 3346 5598. E-mail address: [email protected] (D.L. Paterson).
and multilocus sequence typing (MLST) are employed [1]. PFGE and rep-PCR have been demonstrated to be more discriminatory than MLST, which is routinely used to put epidemiological information in a global context [2]. Whilst outbreaks of A. baumannii have been reported in many regions worldwide, including Europe, North America, Latin America, Australia, Asia, the Middle East and South Africa [1], few data are available for East African A. baumannii isolates. In the current study, antibiotic susceptibility testing, MLST, rep-PCR and PFGE were performed for 25 A. baumannii strains isolated at Aga Khan University Hospital, Nairobi (AKUH, N), Kenya. 2. Materials and methods 2.1. Collection and identification of A. baumannii isolates The 25 non-repetitive A. baumannii isolates investigated in this study were cultivated at the Division of Microbiology of AKUH, N from the following specimen types: 9 A. baumannii isolates were
http://dx.doi.org/10.1016/j.jgar.2014.03.007 2213-7165/ß 2014 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
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derived from tracheal aspirates, 5 isolates were from pus swabs, 3 were from nasal swabs, 2 isolates each were from sputum, cerebrospinal fluid (CSF) and urine, and 1 isolate each was derived from blood and unspecified tissue. The two CSF isolates and the tissue sample isolate were considered clinically significant and the patients were specifically treated with colistin. Both CSF samples came from patients with ventricular shunts. All isolates were collected from September 2010 to September 2011. Although a previous outbreak of multiresistant Acinetobacter in the intensive care unit (ICU) [3] had been stopped with highly stringent infection control measures, multidrug-resistant Acinetobacter continued to be isolated from patients admitted to the hospital. Fourteen of the patients had a history of prior admission to the hospital, whilst eight had a history of admission to various other hospitals in Nairobi. Thirteen of the patients were in the ICU at the time of specimen collection. However, their hospital stay was at different time periods. Identification of the isolates as A. baumannii was reconfirmed by 16S rDNA sequencing in addition to PCR detection of the blaOXA-51like oxacillinase [4,5]. 2.2. Antimicrobial susceptibility testing Antimicrobial susceptibility of the clinical isolates was determined by the disc diffusion method on Mueller–Hinton agar (Oxoid Ltd., Basingstoke, UK) according to Clinical and Laboratory Standard Institute (CLSI) recommendations [6]. Antibiotic disks were supplied by Becton, Dickinson & Co. (Franklin Lakes, NJ) and the following antibiotics were tested: amikacin; cefepime; meropenem; imipenem; ceftazidime; ticarcillin/clavulanic acid; cefotaxime/clavulanic acid; piperacillin/tazobactam; cefoxitin;
ciprofloxacin; gentamicin; nitrofurantoin; fosfomycin trometamol; and trimethoprim/sulfamethoxazole. 2.3. Bacterial genotyping Genomic DNA was extracted using an UltraClean1 Microbial DNA Isolation Kit (MO BIO Laboratories, Carlsbad, CA) as recommended by the manufacturer and was used for MLST and rep-PCR analysis. MLST was performed as previously described [7], making use of the A. baumannii MLST website (http://pubmlst.org/ abaumannii/) developed by Keith Jolley and located at the University of Oxford (Oxford, UK). rep-PCR and PFGE typing were used for the increased resolution required for investigation of clonality in outbreak analysis.rep-PCR typing was performed using a DiversiLab Acinetobacter Kit (bioMe´rieux, Marcy l’E´toile, France). DiversiLab fingerprints were analysed with DiversiLab software using the Pearson correlation statistical method to determine clonal relationships. Isolates were considered as clonal when the rep-PCR patterns showed 95% similarity. PFGE was conducted with the restriction enzyme ApaI using a CHEF-DR1 III system (Bio-Rad, Hercules, CA) as previously described [8]. 2.4. Resistance gene analysis Presence of the metallo-b-lactamase gene blaIMP as well as the extended-spectrum b-lactamase (ESBL) gene blaPER-1 and the AmpC b-lactamase gene blaCMY-2 was determined by PCR [9–11]. The occurrence of carbapenem-hydrolysing class D b-lactamase (oxacillinase) (CHDL) genes, namely blaOXA-51-like, blaOXA-23-like and blaOXA-58-like, was determined by PCR [12]. Detection of insertion
Fig. 1. Molecular typing, antimicrobial resistance profile and source of isolation of 25 Acinetobacter baumannii clinical isolates. Of note, all isolates were resistant to all antibiotics tested, including meropenem (MEM), imipenem (IMP) and amikacin (AN), with the exception of four isolates; isolates 19, 21 and 23 were susceptible to AN and isolate 24 was susceptible to MEM, IMP and AN. R, resistant; S, susceptible. ICU, intensive care unit; HDU, haemodialysis unit.
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
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Fig. 2. Time course of Acinetobacter baumannii outbreaks according to sequence type (ST). Isolates are depicted with diamonds; isolates within circles were derived from patients staying in the intensive care unit or haemodialysis unit, and isolates outside circles were derived from patients hospitalised in other wards.
sequence ISAba1 upstream of blaOXA-51-like was performed using PCR [13]. 3. Results and discussion All isolates were resistant to cefepime, ceftazidime, ticarcillin/ clavulanic acid, cefotaxime/clavulanic acid, piperacillin/tazobactam, cefoxitin, ciprofloxacin, gentamicin, nitrofurantoin, fosfomycin trometamol and trimethoprim/sulfamethoxazole. In addition, all isolates except one (AKUH AB008) showed resistance to meropenem and imipenem. In addition to being susceptible to meropenem and imipenem, isolate AKUH AB008 was also susceptible to amikacin, as were three other isolates. Resistance profiles of meropenem, imipenem and amikacin are illustrated in Fig. 1. These investigations showed that three clones of A. baumannii were found within this Kenyan institution. The 25 isolates investigated in this study grouped into three different sequence types (STs) belonging to three different clonal complexes (CCs). The most prevalent sequence type was ST110 (17 isolates), followed by ST92 (5 isolates) and ST109 (3 isolates). Worldwide, ST92 and the associated CC92, also known as European clone 2 or worldwide clonal lineage 2, represent the most widespread sequence type(s). ST109 is part of clonal cluster CC1, which is also found worldwide (http://pubmlst.org/abaumannii/). In this study, the strains of each sequence type shared similar PFGE and rep-PCR profiles. It is worth noting that all of the ST110 isolates were resistant to all of the antibiotics investigated, whilst two isolates of ST92 and two isolates of ST109 were susceptible to amikacin; one of the amikacin-susceptible ST109 isolates was also susceptible to meropenem and imipenem. Isolates of the same sequence type clustered together when analysed by the DiversiLab rep-PCR typing method and PFGE (Fig. 1). The naturally occurring blaOXA-51-like gene was detected in all 25 A. baumannii isolates and the acquired CHDL blaOXA-23 gene was present in all meropenem-resistant isolates, whilst the meropenem-susceptible isolate was negative for the blaOXA-23-like gene. The blaOXA-58-like gene was not detected, and all isolates tested negative for blaPER-1 and blaCMY-2. Two isolates (AKUH 012 and 015), both ST109, were positive for the element ISAba1–blaOXA-51like, whilst the remaining 23 isolates were negative for the presence of this element.
ST110 A. baumannii have been previously found in India [14], South Korea [15] and Australia [16]; however, in these settings ST110 did not account for the majority of strains. Although in the current study ST110 isolates were resistant to all of the antibiotics investigated, invariably producing OXA-23, antibiotic susceptibility and resistance mechanisms of ST110 were less uniform in other settings [14,15]. Whilst in the current study blaOXA-23 was the major carbapenem resistance mechanism detected, a previous report from AKUH, N has shown the antibiotic resistance gene structure ISAba1–blaOXA23 to be the main mechanism of carbapenemase resistance among 10 Kenyan A. baumannii isolates [3]. In the current study, two strains of ST109 A. baumannii harboured the ISAba1–blaOXA-51-like element. However, the carbapenemase-producing genes blaIMP and blaOXA-58 as well as the ESBL blaPER-1 and AmpC-producing gene blaCMY-2 were not detected. Thus, the molecular epidemiology of carbapenem-resistant A. baumannii isolates from a single hospital in Kenya reveals the presence of organisms associated with infection elsewhere in the world. The time course of the outbreaks is illustrated in Fig. 2. It is noteworthy that the first isolate of each sequence type was in a patient transferred from another hospital. Unfortunately, as has occurred elsewhere in the world, introduction of strains from other hospitals and person-to-person spread of hospital-adapted strains of A. baumannii appears to be occurring in Kenyan healthcare facilities.
Funding None.
Competing interests DLP has acted as a consultant to AstraZeneca, Merck and Pfizer. All other authors declare no competing interests.
Ethical approval Ethical clearance for this work was obtained from the Research Committee of Aga Khan University Hospital, Nairobi (Kenya).
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
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Acknowledgments The authors are thankful to the team in the Division of Microbiology, Department of Pathology, at Aga Khan University Hospital, Nairobi (Kenya). References [1] Peleg AY, Seifert H, Paterson DL. Acinetobacter baumannii: emergence of a successful pathogen. Clin Microbiol Rev 2008;21:538–82. [2] Runnegar N, Sidjabat H, Goh HM, Nimmo GR, Schembri MA, Paterson DL. Molecular epidemiology of multidrug-resistant Acinetobacter baumannii in a single institution over a 10-year period. J Clin Microbiol 2010;48:4051–6. [3] Revathi G, Siu LK, Lu PL, Huang LY. First report of NDM-1-producing Acinetobacter baumannii in East Africa. Int J Infect Dis 2013;17:e1255–58. [4] Lane DJ. 16S/23S rRNA sequencing. In: Stackebrandt E, Goodfellow M, editors. Nucleic acid techniques in bacterial systematics. Chichester, UK: Wiley; 1991. p. 115–75. [5] Coelho J, Woodford N, Afzal-Shah M, Livermore D. Occurrence of OXA-58-like carbapenemases in Acinetobacter spp. collected over 10 years in three continents. Antimicrob Agents Chemother 2006;50:756–8. [6] Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing; twenty-first informational supplement.Document M100-S21 Wayne, PA: CLSI; 2011. [7] Bartual SG, Seifert H, Hippler C, Luzon MA, Wisplinghoff H, Rodrı´guez-Valera F. Development of a multilocus sequence typing scheme for characterization of clinical isolates of Acinetobacter baumannii. J Clin Microbiol 2005;43:4382–90.
[8] Seifert H, Dolzani L, Bressan R, van der Reijden T, van Strijen B, Stefanik D, et al. Standardization and interlaboratory reproducibility assessment of pulsedfield gel electrophoresis-generated fingerprints of Acinetobacter baumannii. J Clin Microbiol 2005;43:4328–35. [9] Poirel L, Naas T, Nicolas D, Collet L, Bellais S, Cavallo JD, et al. Characterization of VIM-2, a carbapenem-hydrolyzing metallo-b-lactamase and its plasmidand integron-borne gene from a Pseudomonas aeruginosa clinical isolate in France. Antimicrob Agents Chemother 2000;44:891–7. [10] Poirel L, Cabanne L, Vahaboglu H, Nordmann P. Genetic environment and expression of the extended-spectrum b-lactamase blaPER-1 gene in Gramnegative bacteria. Antimicrob Agents Chemother 2005;49:1708–13. [11] Koeck JL, Arlet G, Philippon A, Basmaciogullari S, Thien HV, Buisson Y, et al. A plasmid-mediated CMY-2 b-lactamase from an Algerian clinical isolate of Salmonella senftenberg. FEMS Microbiol Lett 1997;152:255–60. [12] Adams-Haduch JM, Paterson DL, Sidjabat HE, Pasculle AW, Potoski BA, Muto CA, et al. Genetic basis of multidrug resistance in Acinetobacter baumannii clinical isolates at a tertiary medical center in Pennsylvania. Antimicrob Agents Chemother 2008;52:3837–43. [13] Turton JF, Ward ME, Woodford N, Kaufmann ME, Pike R, Livermore DM, et al. The role of ISAba1 in expression of OXA carbapenemase genes in Acinetobacter baumannii. FEMS Microbiol Lett 2006;258:72–7. [14] Rynga D, Shariff M, Deb M. Multi-locus sequence types of Acinetobacter baumanii clinical isolates from India. J Infect Dev Ctries 2013;7:358–60. [15] Park YK, Jung SI, Park KH, Kim DH, Choi JY, Kim SH, et al. Changes in antimicrobial susceptibility and major clones of Acinetobacter calcoaceticus– baumannii complex isolates from a single hospital in Korea over 7 years. J Med Microbiol 2012;61:71–9. [16] Hamidian M, Hall RM. ISAba1 targets a specific position upstream of the intrinsic ampC gene of Acinetobacter baumannii leading to cephalosporin resistance. J Antimicrob Chemother 2013;68:2682–3.
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
JGAR-82; No. of Pages 4 Journal of Global Antimicrobial Resistance xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Journal of Global Antimicrobial Resistance journal homepage: www.elsevier.com/locate/jgar
Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital Charlotte A. Huber a, Anna L. Sartor a, Francis McOdimba b, Reena Shah b, Patricia Shivachi b, Hanna E. Sidjabat a, Gunturu Revathi b, David L. Paterson a,* a
The University of Queensland, UQ Centre for Clinical Research, Queensland, Building 71/918, Royal Brisbane and Women’s Hospital Campus, Herston, QLD 4029, Australia Aga Khan University Hospital, Division of Microbiology, Department of Pathology, 3rd Parklands Avenue, P.O. Box 30270-00100, Nairobi, Kenya
b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 27 September 2013 Received in revised form 25 March 2014 Accepted 28 March 2014
Acinetobacter baumannii is a serious nosocomial pathogen with a high propensity to cause outbreaks. Whilst outbreaks of A. baumannii have been reported in many regions worldwide, few data are available from East Africa. In this study, 25 A. baumannii isolates derived from a single institution located in Nairobi, Kenya, between September 2010 and September 2011 were examined. Antimicrobial susceptibility testing was performed by the disc diffusion method and the relatedness among the isolates was examined by pulsed-field gel electrophoresis, repetitive sequence-based PCR (rep-PCR) and multilocus sequence typing. The examined isolates clustered into three distinct groups. The most prevalent sequence type (ST) was ST110 (17 isolates), followed by ST92 (5 isolates) and ST109 (3 isolates). All isolates exhibited resistance to cefepime, ceftazidime, ticarcillin/clavulanic acid, cefotaxime/clavulanic acid, piperacillin/tazobactam, cefoxitin, ciprofloxacin, gentamicin, nitrofurantoin, fosfomycin trometamol, trimethoprim/sulfamethoxazole, amikacin, meropenem and imipenem, with the exception of four isolates. Two isolates belonging to ST92 and two isolates belonging to ST109 were susceptible to amikacin; one of these amikacin-susceptible ST109 isolates was also susceptible to meropenem and imipenem. All isolates were positive for OXA 51-like and all carbapenem-resistant isolates were OXA-23 positive. ß 2014 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.
Keywords: Acinetobacter baumannii Antibiotic susceptibility Outbreak Sequence type
1. Introduction Acinetobacter baumannii is a Gram-negative coccobacillus that has emerged as a serious healthcare-associated pathogen in recent years. A. baumannii infection may result in pneumonia, bloodstream infection, urinary tract infection, meningitis and wound infection as well as other manifestations of infection [1]. A. baumannii has a high propensity to cause nosocomial outbreaks, possibly due to its persistence in the hospital environment and its remarkable ability to acquire drug resistance. Several studies indicate significantly increased resistance rates in epidemic A. baumannii strains compared with sporadic strains [1]. To distinguish outbreak strains from epidemiologically unrelated A. baumannii strains, typing methods such as pulsed-field gel electrophoresis (PFGE), repetitive sequence-based PCR (rep-PCR)
* Corresponding author. Tel.: +61 7 3346 5434; fax: +61 7 3346 5598. E-mail address: [email protected] (D.L. Paterson).
and multilocus sequence typing (MLST) are employed [1]. PFGE and rep-PCR have been demonstrated to be more discriminatory than MLST, which is routinely used to put epidemiological information in a global context [2]. Whilst outbreaks of A. baumannii have been reported in many regions worldwide, including Europe, North America, Latin America, Australia, Asia, the Middle East and South Africa [1], few data are available for East African A. baumannii isolates. In the current study, antibiotic susceptibility testing, MLST, rep-PCR and PFGE were performed for 25 A. baumannii strains isolated at Aga Khan University Hospital, Nairobi (AKUH, N), Kenya. 2. Materials and methods 2.1. Collection and identification of A. baumannii isolates The 25 non-repetitive A. baumannii isolates investigated in this study were cultivated at the Division of Microbiology of AKUH, N from the following specimen types: 9 A. baumannii isolates were
http://dx.doi.org/10.1016/j.jgar.2014.03.007 2213-7165/ß 2014 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
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derived from tracheal aspirates, 5 isolates were from pus swabs, 3 were from nasal swabs, 2 isolates each were from sputum, cerebrospinal fluid (CSF) and urine, and 1 isolate each was derived from blood and unspecified tissue. The two CSF isolates and the tissue sample isolate were considered clinically significant and the patients were specifically treated with colistin. Both CSF samples came from patients with ventricular shunts. All isolates were collected from September 2010 to September 2011. Although a previous outbreak of multiresistant Acinetobacter in the intensive care unit (ICU) [3] had been stopped with highly stringent infection control measures, multidrug-resistant Acinetobacter continued to be isolated from patients admitted to the hospital. Fourteen of the patients had a history of prior admission to the hospital, whilst eight had a history of admission to various other hospitals in Nairobi. Thirteen of the patients were in the ICU at the time of specimen collection. However, their hospital stay was at different time periods. Identification of the isolates as A. baumannii was reconfirmed by 16S rDNA sequencing in addition to PCR detection of the blaOXA-51like oxacillinase [4,5]. 2.2. Antimicrobial susceptibility testing Antimicrobial susceptibility of the clinical isolates was determined by the disc diffusion method on Mueller–Hinton agar (Oxoid Ltd., Basingstoke, UK) according to Clinical and Laboratory Standard Institute (CLSI) recommendations [6]. Antibiotic disks were supplied by Becton, Dickinson & Co. (Franklin Lakes, NJ) and the following antibiotics were tested: amikacin; cefepime; meropenem; imipenem; ceftazidime; ticarcillin/clavulanic acid; cefotaxime/clavulanic acid; piperacillin/tazobactam; cefoxitin;
ciprofloxacin; gentamicin; nitrofurantoin; fosfomycin trometamol; and trimethoprim/sulfamethoxazole. 2.3. Bacterial genotyping Genomic DNA was extracted using an UltraClean1 Microbial DNA Isolation Kit (MO BIO Laboratories, Carlsbad, CA) as recommended by the manufacturer and was used for MLST and rep-PCR analysis. MLST was performed as previously described [7], making use of the A. baumannii MLST website (http://pubmlst.org/ abaumannii/) developed by Keith Jolley and located at the University of Oxford (Oxford, UK). rep-PCR and PFGE typing were used for the increased resolution required for investigation of clonality in outbreak analysis.rep-PCR typing was performed using a DiversiLab Acinetobacter Kit (bioMe´rieux, Marcy l’E´toile, France). DiversiLab fingerprints were analysed with DiversiLab software using the Pearson correlation statistical method to determine clonal relationships. Isolates were considered as clonal when the rep-PCR patterns showed 95% similarity. PFGE was conducted with the restriction enzyme ApaI using a CHEF-DR1 III system (Bio-Rad, Hercules, CA) as previously described [8]. 2.4. Resistance gene analysis Presence of the metallo-b-lactamase gene blaIMP as well as the extended-spectrum b-lactamase (ESBL) gene blaPER-1 and the AmpC b-lactamase gene blaCMY-2 was determined by PCR [9–11]. The occurrence of carbapenem-hydrolysing class D b-lactamase (oxacillinase) (CHDL) genes, namely blaOXA-51-like, blaOXA-23-like and blaOXA-58-like, was determined by PCR [12]. Detection of insertion
Fig. 1. Molecular typing, antimicrobial resistance profile and source of isolation of 25 Acinetobacter baumannii clinical isolates. Of note, all isolates were resistant to all antibiotics tested, including meropenem (MEM), imipenem (IMP) and amikacin (AN), with the exception of four isolates; isolates 19, 21 and 23 were susceptible to AN and isolate 24 was susceptible to MEM, IMP and AN. R, resistant; S, susceptible. ICU, intensive care unit; HDU, haemodialysis unit.
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
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Fig. 2. Time course of Acinetobacter baumannii outbreaks according to sequence type (ST). Isolates are depicted with diamonds; isolates within circles were derived from patients staying in the intensive care unit or haemodialysis unit, and isolates outside circles were derived from patients hospitalised in other wards.
sequence ISAba1 upstream of blaOXA-51-like was performed using PCR [13]. 3. Results and discussion All isolates were resistant to cefepime, ceftazidime, ticarcillin/ clavulanic acid, cefotaxime/clavulanic acid, piperacillin/tazobactam, cefoxitin, ciprofloxacin, gentamicin, nitrofurantoin, fosfomycin trometamol and trimethoprim/sulfamethoxazole. In addition, all isolates except one (AKUH AB008) showed resistance to meropenem and imipenem. In addition to being susceptible to meropenem and imipenem, isolate AKUH AB008 was also susceptible to amikacin, as were three other isolates. Resistance profiles of meropenem, imipenem and amikacin are illustrated in Fig. 1. These investigations showed that three clones of A. baumannii were found within this Kenyan institution. The 25 isolates investigated in this study grouped into three different sequence types (STs) belonging to three different clonal complexes (CCs). The most prevalent sequence type was ST110 (17 isolates), followed by ST92 (5 isolates) and ST109 (3 isolates). Worldwide, ST92 and the associated CC92, also known as European clone 2 or worldwide clonal lineage 2, represent the most widespread sequence type(s). ST109 is part of clonal cluster CC1, which is also found worldwide (http://pubmlst.org/abaumannii/). In this study, the strains of each sequence type shared similar PFGE and rep-PCR profiles. It is worth noting that all of the ST110 isolates were resistant to all of the antibiotics investigated, whilst two isolates of ST92 and two isolates of ST109 were susceptible to amikacin; one of the amikacin-susceptible ST109 isolates was also susceptible to meropenem and imipenem. Isolates of the same sequence type clustered together when analysed by the DiversiLab rep-PCR typing method and PFGE (Fig. 1). The naturally occurring blaOXA-51-like gene was detected in all 25 A. baumannii isolates and the acquired CHDL blaOXA-23 gene was present in all meropenem-resistant isolates, whilst the meropenem-susceptible isolate was negative for the blaOXA-23-like gene. The blaOXA-58-like gene was not detected, and all isolates tested negative for blaPER-1 and blaCMY-2. Two isolates (AKUH 012 and 015), both ST109, were positive for the element ISAba1–blaOXA-51like, whilst the remaining 23 isolates were negative for the presence of this element.
ST110 A. baumannii have been previously found in India [14], South Korea [15] and Australia [16]; however, in these settings ST110 did not account for the majority of strains. Although in the current study ST110 isolates were resistant to all of the antibiotics investigated, invariably producing OXA-23, antibiotic susceptibility and resistance mechanisms of ST110 were less uniform in other settings [14,15]. Whilst in the current study blaOXA-23 was the major carbapenem resistance mechanism detected, a previous report from AKUH, N has shown the antibiotic resistance gene structure ISAba1–blaOXA23 to be the main mechanism of carbapenemase resistance among 10 Kenyan A. baumannii isolates [3]. In the current study, two strains of ST109 A. baumannii harboured the ISAba1–blaOXA-51-like element. However, the carbapenemase-producing genes blaIMP and blaOXA-58 as well as the ESBL blaPER-1 and AmpC-producing gene blaCMY-2 were not detected. Thus, the molecular epidemiology of carbapenem-resistant A. baumannii isolates from a single hospital in Kenya reveals the presence of organisms associated with infection elsewhere in the world. The time course of the outbreaks is illustrated in Fig. 2. It is noteworthy that the first isolate of each sequence type was in a patient transferred from another hospital. Unfortunately, as has occurred elsewhere in the world, introduction of strains from other hospitals and person-to-person spread of hospital-adapted strains of A. baumannii appears to be occurring in Kenyan healthcare facilities.
Funding None.
Competing interests DLP has acted as a consultant to AstraZeneca, Merck and Pfizer. All other authors declare no competing interests.
Ethical approval Ethical clearance for this work was obtained from the Research Committee of Aga Khan University Hospital, Nairobi (Kenya).
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007
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Acknowledgments The authors are thankful to the team in the Division of Microbiology, Department of Pathology, at Aga Khan University Hospital, Nairobi (Kenya). References [1] Peleg AY, Seifert H, Paterson DL. Acinetobacter baumannii: emergence of a successful pathogen. Clin Microbiol Rev 2008;21:538–82. [2] Runnegar N, Sidjabat H, Goh HM, Nimmo GR, Schembri MA, Paterson DL. Molecular epidemiology of multidrug-resistant Acinetobacter baumannii in a single institution over a 10-year period. J Clin Microbiol 2010;48:4051–6. [3] Revathi G, Siu LK, Lu PL, Huang LY. First report of NDM-1-producing Acinetobacter baumannii in East Africa. Int J Infect Dis 2013;17:e1255–58. [4] Lane DJ. 16S/23S rRNA sequencing. In: Stackebrandt E, Goodfellow M, editors. Nucleic acid techniques in bacterial systematics. Chichester, UK: Wiley; 1991. p. 115–75. [5] Coelho J, Woodford N, Afzal-Shah M, Livermore D. Occurrence of OXA-58-like carbapenemases in Acinetobacter spp. collected over 10 years in three continents. Antimicrob Agents Chemother 2006;50:756–8. [6] Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing; twenty-first informational supplement.Document M100-S21 Wayne, PA: CLSI; 2011. [7] Bartual SG, Seifert H, Hippler C, Luzon MA, Wisplinghoff H, Rodrı´guez-Valera F. Development of a multilocus sequence typing scheme for characterization of clinical isolates of Acinetobacter baumannii. J Clin Microbiol 2005;43:4382–90.
[8] Seifert H, Dolzani L, Bressan R, van der Reijden T, van Strijen B, Stefanik D, et al. Standardization and interlaboratory reproducibility assessment of pulsedfield gel electrophoresis-generated fingerprints of Acinetobacter baumannii. J Clin Microbiol 2005;43:4328–35. [9] Poirel L, Naas T, Nicolas D, Collet L, Bellais S, Cavallo JD, et al. Characterization of VIM-2, a carbapenem-hydrolyzing metallo-b-lactamase and its plasmidand integron-borne gene from a Pseudomonas aeruginosa clinical isolate in France. Antimicrob Agents Chemother 2000;44:891–7. [10] Poirel L, Cabanne L, Vahaboglu H, Nordmann P. Genetic environment and expression of the extended-spectrum b-lactamase blaPER-1 gene in Gramnegative bacteria. Antimicrob Agents Chemother 2005;49:1708–13. [11] Koeck JL, Arlet G, Philippon A, Basmaciogullari S, Thien HV, Buisson Y, et al. A plasmid-mediated CMY-2 b-lactamase from an Algerian clinical isolate of Salmonella senftenberg. FEMS Microbiol Lett 1997;152:255–60. [12] Adams-Haduch JM, Paterson DL, Sidjabat HE, Pasculle AW, Potoski BA, Muto CA, et al. Genetic basis of multidrug resistance in Acinetobacter baumannii clinical isolates at a tertiary medical center in Pennsylvania. Antimicrob Agents Chemother 2008;52:3837–43. [13] Turton JF, Ward ME, Woodford N, Kaufmann ME, Pike R, Livermore DM, et al. The role of ISAba1 in expression of OXA carbapenemase genes in Acinetobacter baumannii. FEMS Microbiol Lett 2006;258:72–7. [14] Rynga D, Shariff M, Deb M. Multi-locus sequence types of Acinetobacter baumanii clinical isolates from India. J Infect Dev Ctries 2013;7:358–60. [15] Park YK, Jung SI, Park KH, Kim DH, Choi JY, Kim SH, et al. Changes in antimicrobial susceptibility and major clones of Acinetobacter calcoaceticus– baumannii complex isolates from a single hospital in Korea over 7 years. J Med Microbiol 2012;61:71–9. [16] Hamidian M, Hall RM. ISAba1 targets a specific position upstream of the intrinsic ampC gene of Acinetobacter baumannii leading to cephalosporin resistance. J Antimicrob Chemother 2013;68:2682–3.
Please cite this article in press as: Huber CA, et al. Outbreaks of multidrug-resistant Acinetobacter baumannii strains in a Kenyan teaching hospital. J Global Antimicrob Resist (2014), http://dx.doi.org/10.1016/j.jgar.2014.03.007