J Antimicrob Chemother 2016; 71 Suppl 1: i103 – i109 doi:10.1093/jac/dkw076
Antibiotic susceptibility in Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes in Pakistan: a review of results from the Survey of Antibiotic Resistance (SOAR) 2002–15 A. Zafar1, R. Hasan1, S. Nizamuddin2, N. Mahmood2, S. Mukhtar2, F. Ali3, I. Morrissey4, K. Barker5 and D. Torumkuney5* 1
Aga Khan University Hospital, Department of Pathology and Laboratory Medicine, Section of Microbiology, Karachi, Pakistan; 2Shaukat Khanum Memorial Cancer Hospital and Research Center, Department of Microbiology, 7A, Block R-3, Johar Town, Lahore, Pakistan; 3 GlaxoSmithKline Pakistan, 35 Dockyard Road, West Wharf, Karachi 74000, Pakistan; 4IHMA Europe Sa`rl, 9a route de la Corniche, Epalinges 1066, Switzerland; 5GlaxoSmithKline, 980 Great West Road, Brentford, Middlesex TW8 9GS, UK *Corresponding author. Tel: +44-20-8047-1671; E-mail: [email protected]
Objectives: To investigate changes in the antibiotic susceptibility of Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes from the Survey of Antibiotic Resistance (SOAR) in community-acquired respiratory tract infections (CA-RTIs) between 2002 and 2015 in Pakistan. Methods: This is a review based on previously published studies from 2002 –03, 2004– 06 and 2007– 09 and also new data from 2014–15. Susceptibility was determined by Etestw or disc diffusion according to CLSI and pharmacokinetic/pharmacodynamic (PK/PD) breakpoints. Results: A total of 706 isolates from CA-RTIs comprising 381 S. pneumoniae, 230 H. influenzae and 95 S. pyogenes were collected between 2002 and 2015 and tested against a range of antibiotics. Antibiotic resistance in S. pneumoniae rose steeply from 2002 to 2009, with isolates non-susceptible to penicillin and macrolides increasing from 10% to 34.1% and from 13% –14% to 29.7%, respectively. Susceptibility to amoxicillin/clavulanic acid (and by inference amoxicillin) remained between 99.4% and 100% from 2002 to 2015. Over the years, the prevalence of susceptibility to cefuroxime was 98% – 100% among S. pneumoniae. Resistance in S. pneumoniae to some older antibiotics between 2007 and 2009 was high (86.8% for trimethoprim/sulfamethoxazole and 57.2% for tetracycline). Between 2002 and 2015, ampicillin resistance (b-lactamase-positive strains) among H. influenzae has remained low (between 2.6% and 3.2%) and almost unchanged over the years (H. influenzae was not tested during 2004–06). For S. pyogenes isolates, macrolide resistance reached 22%; however, susceptibility to penicillin, amoxicillin/clavulanic acid and cefuroxime remained stable at 100%. Conclusions: In S. pneumoniae from Pakistan, there has been a clear reduction in susceptibility to key antibiotics since 2002, but not to amoxicillin/clavulanic acid (amoxicillin) or cefuroxime. However, susceptibility in H. influenzae has remained stable. Local antibiotic susceptibility/resistance data are essential to support informed prescribing for CA-RTIs and other infections.
Introduction Worldwide, there are concerns about rising levels of antibiotic resistance and fears that the efficacy of antimicrobial therapy may be compromised, resulting in treatment failure and reduced utility of older antibiotics.1 Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes are all commonly carried in the nasopharynx and implicated in community-acquired respiratory tract infections (CA-RTIs). These infections account for significant morbidity and mortality, including severe life-threatening infections such as pneumonia and meningitis (S. pneumoniae and H. influenzae) and rheumatic fever with risk of cardiac sequelae (S. pyogenes).2 – 7
Like many other Asian countries, in Pakistan antimicrobial therapeutic choices for RTIs are usually empirical due to the limited availability of laboratory identification and antibiotic susceptibility testing of bacterial isolates. Even when available, the cost of laboratory testing in this setting can often prove inhibitive.8 The increasing prevalence of resistant organisms threatens the current and future treatment of CA-RTIs.4,9 – 11 Surveillance studies, therefore, provide an important tool for determining local and regional susceptibility patterns and guiding empirical antimicrobial therapy.12 The Survey of Antibiotic Resistance (SOAR) studies have been undertaken in Pakistan during 2002 – 03, 2004 – 06, 2007 – 09 and 2014 – 15, as well as elsewhere in Asia, the Middle East, Africa, Latin America, Asia Pacific and the Commonwealth of
# The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: [email protected]
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Independent States countries, to provide detailed and comparative information on resistance trends among strains from documented CA-RTIs caused by S. pneumoniae, H. influenzae and S. pyogenes.13 – 16
Materials and methods Collaborating centres Isolates were collected from two sites in Pakistan: Aga Khan University Hospital (for the 2002 – 09 study period) Karachi and Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore (for the 2014– 15 study period).
Clinical isolates (from outpatients who attended the university hospitals)
susceptibility to amoxicillin alone), cefixime, cefuroxime, cefaclor, cefprozil, clarithromycin, azithromycin, erythromycin, ciprofloxacin, trimethoprim/sulfamethoxazole, tetracycline, chloramphenicol and clindamycin. S. pyogenes strains were tested against penicillin, amoxicillin, cefaclor, cefixime, erythromycin, azithromycin, clarithromycin, clindamycin, tetracycline and chloramphenicol. Finally, susceptibility of H. influenzae was assessed against ampicillin, amoxicillin/clavulanic acid, cefaclor, cefixime, cefprozil, cefuroxime, clarithromycin, azithromycin, ciprofloxacin, levofloxacin, trimethoprim/sulfamethoxazole, tetracycline and chloramphenicol. For the 2014 –15 isolates, susceptibility testing was carried out using a more limited range of antibiotics. S. pneumoniae susceptibility was tested against amoxicillin/clavulanic acid (amoxicillin), cefaclor, cefuroxime, chloramphenicol, erythromycin and penicillin. H. influenzae was tested against amoxicillin/clavulanic acid, ampicillin, azithromycin, cefaclor, cefuroxime, chloramphenicol and levofloxacin using the same CLSI breakpoints as for 2009 isolates.
Between 2002 and 2015, a total of 381 S. pneumoniae, 230 H. influenzae and 95 S. pyogenes isolates were obtained from a range of clinical samples including sputum, blood, bronchoalveolar lavage, middle ear effusion and tracheal aspirate taken from adult and paediatric patients with a confirmed CA-RTI using routine clinical collection methods. All S. pyogenes isolates were from the throat. Duplicate isolates from the same patient were not accepted. S. pneumoniae isolates were collected during 2002 – 03, 2004 – 06, 2007 – 09 and 2014 – 15. H. influenzae isolates were collected over the same time periods, except none were collected in 2004 –06. S. pyogenes isolates were collected only during 2007 –09. The isolates were identified using conventional methods (optochin susceptibility for S. pneumoniae, X and V factor requirement for H. influenzae and bacitracin antibiotic disc susceptibility for S. pyogenes strains). Isolates were stored at 2708C until tested. b-Lactamase production of H. influenzae isolates was determined by a chromogenic cephalosporin (nitrocefin) disc method.17 b-Lactamase-negative ampicillin-resistant (BLNAR) isolates were defined as those organisms having a negative b-lactamase result and an ampicillin MIC of ≥4 mg/L.18
Quality control strains S. pneumoniae ATCC 49619, H. influenzae ATCC 49247, H. influenzae ATCC 49766, Escherichia coli ATCC 25922 and E. coli ATCC 32518 were included on each day of testing. Results of susceptibility testing were accepted if the results of the control strains were within published limits. Any Etestw MIC results that were between doubling dilutions were rounded up to the next doubling dilution MIC for data analysis. MICs and zone diameters were interpreted qualitatively using CLSI interpretive standards; MICs were also analysed using PK/PD breakpoints established to predict the clinical and bacteriological efficacy of antimicrobial dosing regimens. 11,19,23 To interpret azithromycin and clarithromycin MICs for H. influenzae, revised breakpoints were used to account for incubation in a CO2 atmosphere (using the Etestw susceptibility method according to the manufacturer’s instructions; bioMe´rieux, Marcy-l’Etoile, France). The respective PK/PD breakpoints were adjusted accordingly, i.e. raised by one doubling dilution for interpretation of H. influenzae susceptibility because of incubation under 5% CO2.
Susceptibility testing
Results w
MICs were determined using Etest susceptibility testing methods according to the manufacturer’s instructions [AB BIODISK (Solna, Sweden) for 2002 – 09 and bioMe´rieux (Marcy-l’Etoile, France) for 2014 – 15] and interpreted using CLSI guidelines and pharmacokinetic/pharmacodynamic (PK/PD) breakpoints (for 2002 – 03, M100-S1419; for 2004 – 06, CLSI M100-S1518; for 2007 – 09, M100-S1920; and for 2014 – 15, M100-S2421). For isolates collected during 2007 – 09, susceptibility to erythromycin, tetracycline, trimethoprim/sulfamethoxazole and chloramphenicol was also determined using the CLSI disc diffusion method.22 Briefly, for Etestw MIC testing, S. pneumoniae isolates were subcultured twice on blood-supplemented Mueller–Hinton agar and H. influenzae isolates were cultured on Haemophilus Test Medium before susceptibility testing was performed. Inocula were prepared by direct colony suspension to give a 0.5 McFarland standard dilution and inoculated onto appropriate agar plates to produce a confluent lawn of growth. Etestw and antibiotic discs were applied and plates incubated for 20 – 24 h (16 – 17 h for H. influenzae agar disc diffusion) at 358C in a 5% CO2 atmosphere. For azithromycin and clarithromycin Etests, S. pneumoniae and S. pyogenes isolates were incubated in ambient air due to the adverse effect of CO2 on the activity of macrolide antibiotics. Etestw MICs and inhibition zone diameters were read in accordance with bioMe´rieux’s instructions and CLSI guidelines for disc susceptibility testing, respectively. The number of antibiotics tested varied at the different timepoints. Between 2002 and 2009, S. pneumoniae strains were tested against penicillin, amoxicillin/clavulanic acid (data from which can be used to deduce
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Quality control and data analysis
From 2002 to 2009, CLSI breakpoint interpretations for S. pneumoniae were unchanged for all of the study drugs except penicillin. PK/PD breakpoints were also in good agreement with the ‘susceptible’ category, except for cefaclor and azithromycin. For both of these antibiotics, the susceptible breakpoint from PK/PD data was lower than the CLSI breakpoint—by one dilution for cefaclor and three dilutions for azithromycin, reflecting the relatively low blood level attainable with this antibiotic. Table 1 shows the breakpoints used to interpret Etestw MICs for strains isolated in the period 2002 –15. During the course of the studies, S. pneumoniae with raised penicillin MICs were being reported with increasing frequency worldwide. The original breakpoints for penicillin of susceptible (S) ≤0.06 mg/L, intermediate (I) 0.12– 1.0 mg/L and resistant (R) ≥2.0 mg/L had been appropriate for use of oral penicillin V and for injectable penicillin used in meningitis treatment. However, with an increasing prevalence of penicillin-intermediate S. pneumoniae (PISP) strains, the use of penicillin was questioned for treatment of S. pneumoniae infections other than meningitis and particularly for pneumonia. This concern was addressed by CLSI and new penicillin breakpoints were applied for S. pneumoniae from non-meningeal sources.22 New breakpoints were recommended in 2009 specifically for use with S. pneumoniae from RTIs. High doses of penicillin and other b-lactam antibiotics,
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Table 1. MIC breakpoints (mg/L) used for S. pneumoniae and H. influenzae isolates CLSI breakpointsa PK/PD breakpoints Antimicrobial Penicillinb Penicillinc (only for 2007– 09/2014 – 15) Ampicillin Amoxicillin AMCd Cefaclor Cefixime Cefprozil Cefuroximee Azithromycinf Clarithromycinf Ciprofloxacin Levofloxacin
S. pneumoniae
H. influenzae
S
R
S
I
R
S
I
R
— — — ≤2 ≤2 ≤0.5 ≤1 ≤1 ≤1 ≤0.12 ≤0.25 ≤1 ≤2
— — — ≥4 ≥4 ≥1 ≥2 ≥2 ≥2 ≥0.25 ≥0.5 ≥2 ≥4
≤0.06 ≤2 — ≤2 ≤2 ≤1 — ≤2 ≤1 ≤0.5 ≤0.25 — ≤2
0.12– 1 4 — 4 4 2 — 4 2 1 0.5 — 4
≥2 ≥8 — ≥8 ≥8 ≥4 — ≥8 ≥4 ≥2 ≥1 ≤1 ≥8
— — ≤1 — ≤4 ≤8 ≤1 ≤8 ≤4 ≤8 ≤16 — ≤2
— — 2 — — 16 — 16 8 — 32 — —
— — ≥4 — ≥8 ≥32 — ≥32 ≥16 — ≥64 — —
AMC, amoxicillin/clavulanic acid; —, no breakpoint available. For breakpoints see CLSI.22 b CLSI 2008 breakpoints/CLSI 2009 breakpoints for oral penicillin V. c CLSI 2009 breakpoints for parenteral penicillin/non-meningeal. d Amoxicillin/clavulanic acid was tested in a 2 : 1 ratio of amoxicillin to clavulanic acid; breakpoints are expressed as the amoxicillin component. e Breakpoints used are for cefuroxime axetil. f Azithromycin and clarithromycin breakpoints are those provided by AB Biodisk for incubation in CO2. a
such as amoxicillin, were also recommended for treatment of PISP or penicillin-resistant S. pneumoniae infections. These changes are documented in Table 1 and show that S. pneumoniae with penicillin MICs ≤2.0 mg/L are regarded as susceptible, whereas previously some of these isolates would have been regarded as having intermediate susceptibility. Clinical studies showed that, provided penicillin was used in adequate dosage, the new susceptibility category could be used as a breakpoint for non-meningeal isolates.
Antibiotic resistance in S. pneumoniae Overall, non-susceptibility to penicillin among S. pneumoniae strains rose steeply, from 10% in 2002 to 34.1% in 2009. The prevalence of penicillin-non-susceptible S. pneumoniae was 10% during 2002 – 03, 18.3% during 2004 – 06 and 34.1% during 2007– 09. During 2014 –15, only 10 S. pneumoniae isolates were collected, of which two were non-susceptible to penicillin. Of the 31 isolates that were penicillin non-susceptible in the 2007 – 09 survey, all had MICs of 4 mg/L, making them intermediate (Table 2). No isolates of S. pneumoniae obtained during the study were highly resistant to penicillin (MICs ≥8.0 mg/L), although 1.1% of strains in the 2004 – 06 survey were penicillin resistant using the lower CLSI breakpoint in use at that time. Macrolide resistance rose from 13% to 29.7% (depending on the macrolide being tested) over the study period (Figure 1 and Table 2, respectively). By contrast, susceptibility to amoxicillin/ clavulanic acid (or amoxicillin alone) between 2002 and 2015 has remained between 99.4% and 100% among S. pneumoniae isolates. Ciprofloxacin was tested only during 2007 – 09, when
91.2% of S. pneumoniae isolates were found to be susceptible. S. pneumoniae strains showing reduced susceptibility to penicillin were more likely to show increased resistance to macrolides and other antimicrobial agents, particularly older agents such as first-generation cephalosporins (cefaclor), tetracycline or trimethoprim/sulfamethoxazole (Table 3). Almost all PISP strains were resistant to trimethoprim/sulfamethoxazole, .70% were resistant to tetracycline and 50% were resistant to macrolides (erythromycin and clarithromycin). This contrasted with the macrolide resistance rate in penicillin-susceptible S. pneumoniae strains, which was only 10% – 15%. In the Lahore 2014 – 15 study, the erythromycin susceptibility of all S. pneumoniae isolates was 70%. Clindamycin resistance plus macrolide resistance is a useful proxy for detection of the erm(B) resistance marker. 24 In the 2007 – 09 study period, clindamycin resistance was 22.6% among PISP strains and this was coupled with a concomitant high incidence of macrolide resistance (50%) (Table 3). This finding suggests that a number of these isolates are more likely to have macrolide resistance attributable to the erm(B) resistance marker rather than the efflux mechanism of macrolide resistance. In the 2007 – 09 study period, of the S. pneumoniae isolates, 93.4% were susceptible to chloramphenicol (Table 2).
Antibiotic resistance in H. influenzae H. influenzae isolates studied in Pakistan during the 2002 –03 winter season (n ¼ 93) were fully susceptible (100%) to amoxicillin/ clavulanic acid, cefuroxime, cefprozil, cefaclor, cefixime, clarithromycin and azithromycin (Table 4). Furthermore, 3.2% of isolates
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Table 2. Susceptibility (%) of S. pneumoniae in Pakistan to antimicrobials using CLSI or PK/PD breakpoints 2002 –03 (n¼100)a CLSI Antimicrobial Penicillin AMC Cefixime Cefuroxime Cefaclor Cefprozil Clarithromycin Azithromycin Erythromycin Clindamycin SXT Tetracycline Chloramphenicol Ciprofloxacin
2004– 06 (n¼180)b PK/PD
S
I or (R)
90.0 100 93.0d 98.0 96.0 99.0 86.0e 87.0e — — — — — —
10.0 0 7.0 2.0 4.0 1.0 14.0 13.0 — — — — — —
S NA 99.4 80.6 98.3 67.2 — 90.8 73.4 NA NA NA NA NA —
2007– 09 (n¼91)c
CLSI
PK/PD
S
I or (R)
81.7 99.4 — 98.3 89.4 — 90.8 90.2 87.2 88.9 — — — —
18.3 (1.1) 0.6 (0) — 1.7 (1.1) 10.6 (2.8) — 9.2 9.8 12.8 11.1 — — — —
S NA 100 92.3 100 69.2 — 76.9 — NA NA NA NA NA 91.2
2014– 15 (n ¼10)c
CLSI
PK/PD
S
I or (R)
S
65.9 100 — 100 94.5 — 76.9 — 70.3 90.1 13.2 42.8 93.4 —
34.1 (0) 0 — 0 5.5 — 23.1 — 29.7 9.9 86.8 57.2 6.6 —
NA 100 100 70.0 — — — — — — — — —
CLSI S 80.0 100 — 100 90.0 — — — 70.0 — — — 80.0
I or (R) 20.0 0 — 0 10.0 — — — 30.0 — — — 20.0
Resistance (%)
AMC, amoxicillin/clavulanic acid; SXT, trimethoprim/sulfamethoxazole; S, susceptible; I, intermediate; R, resistant; NA, not applicable; —, no data available. a Data for strains from Pakistan summarized by Shibl et al.13 b Data from Pakistan summarized by Sievers et al. 15 c Pakistan current SOAR study data. d PK/PD breakpoint used. e Revised breakpoints for Etestw used.
35 30 25 20 15 10 5 0
Erythromycin resistance Clarithromycin resistance Reduced penicillin susceptibility 2002–03 2004–07 2007–09 Survey timepoint
Figure 1. Penicillin and macrolide resistance in S. pneumoniae strains from CA-RTIs from Pakistan during 2002 –09.
were designated b-lactamase positive on the basis of nitrocefin disc testing.13 For 2007–09 (n ¼99), amoxicillin/clavulanic acid susceptibility remained at 100% among H. influenzae isolates in Pakistan and ampicillin resistance was 4% (4/99 isolates). All isolates were susceptible to cefuroxime and cefixime. The prevalence of resistance to macrolides was between 6.1% and 8.1%. Ciprofloxacin was tested only during 2007–09, when 95.9% of H. influenzae isolates were found to be susceptible (Table 4). Of the 2014 –15 (n¼ 38) H. influenzae isolates, 100% were susceptible to amoxicillin/clavulanic acid and 97.4% were susceptible to ampicillin. Only 1 of the 38 isolates (2.63%) from 2014–15 was b-lactamase positive. Susceptibility to chloramphenicol was low
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at 29%. Some 86.8% of H. influenzae isolates were susceptible to levofloxacin (Table 4). Except for the 2014 –15 study period, all ampicillin-resistant H. influenzae isolates were b-lactamase positive. Although the prevalence of BLNAR isolates was not reported during different study periods, based on CLSI guidelines, rare BLNAR strains of H. influenzae should be considered resistant to amoxicillin/clavulanic acid, ampicillin/sulbactam, cefaclor, cefamandole, cefetamet, cefonicid, cefprozil, cefuroxime, loracarbef and piperacillin/tazobactam, despite apparent in vitro susceptibility of some BLNAR strains to these agents.
Antibiotic resistance in S. pyogenes S. pyogenes was also studied in the 2007 – 09 survey. All strains were susceptible to penicillin. Based on penicillin susceptibility, the strains were also susceptible to amoxicillin, amoxicillin/clavulanic acid, cefuroxime, cefotaxime and cefaclor. By contrast, macrolide (erythromycin, azithromycin and clarithromycin) resistance in these Pakistani CA-RTI isolates was 22% (data not shown).
Discussion The increase in antimicrobial resistance among CA-RTI pathogens is a serious problem throughout the world and has important implications for the successful management of these infections. Since there may be variation in levels of resistance between different geographical areas, it is essential to be aware of the local
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Table 3. Susceptibility (%) of penicillin-susceptible S. pneumoniae (PSSP) and penicillin-intermediate S. pneumoniae (PISP) strains from CA-RTIs in Pakistan in SOAR during 2007 –09 PSSP (n¼60), penicillin MIC ≤2.0 mg/La Antimicrobial Penicillin AMC Cefixime Cefuroxime Cefaclor Clarithromycin Ciprofloxacin Erythromycin Clindamycinb SXT Tetracycline Chloramphenicol
PISP (n¼31), penicillin MIC ≥4 mg/La
S (PK/PD) S (CLSI) R (CLSI) S (PK/PD) S (CLSI) R (CLSI) NA 100 98.3 100 86.7 90 91.7 NA NA NA NA NA
100 100 — 100 98.3 90 — 83.3 96.7 18.3 50 93.3
0 0 — 0 0 10 — 15 3.3 78.4 40 6.7
NA 100 80.6 100 34.5 51.6 90.3 NA NA NA NA NA
0 100 — 100 87.1 51.6 — 45.2 77.4 3.2 29.1 93.5
0 0 — 0 0 48.4 — 54.8 22.6 96.8 70.9 6.5
AMC, amoxicillin/clavulanic acid; SXT, trimethoprim/sulfamethoxazole; S, susceptible; I, intermediate; R, resistant; NA, not applicable; —, no data available. a New CLSI breakpoints for penicillin applied. b Included to differentiate erm(B)-mediated macrolide resistance.
Table 4. Susceptibility (%) of H. influenzae in Pakistan to antimicrobials using CLSI breakpoints 2002 –03 (n ¼93)a Antimicrobial Ampicillin AMC Cefaclor Cefixime Cefprozil Cefuroxime Clarithromycin Azithromycin Chloramphenicol Levofloxacin Ciprofloxacin SXT Tetracycline
2007 –09 (n¼99)b
2014 –15 (n¼38)b
S
I or (R)
S
I or (R)
S
I or (R)
96.8 100 100 100 100 100 100 100 — — — — —
3.2 0 0 0 0 0 0 0 — — — — —
96 100 98 100 — 100 91.9 93.9 88.8 — 95.9 15.2 67.7
4 0 2 0 — 0 8.1 6.1 11.2 — 4.1 84.8 32.3
97.4 100 89.5 — — 97.4 — 100 29 86.8 — — —
2.6 0 10.5 — — 2.6 — 0 71 13.2 — —
AMC, amoxicillin/clavulanic acid; SXT, trimethoprim/sulfamethoxazole; S, susceptible; I, intermediate; R, resistant; —, no data available. a Data for strains from Pakistan summarized by Shibl et al.13 b Pakistan current SOAR study data.
resistance pattern before considering empirical antimicrobial therapy. SOAR was therefore established to provide information on local resistance patterns, as well as overall levels of resistance and susceptibility trends, for key CA-RTI pathogens.
The SOAR studies have been conducted using the Etestw because reliable MIC results can be obtained relatively easily and subsequently the results can be analysed with reference to different breakpoint criteria. In these studies, analyses have been performed using breakpoints defined by CLSI and also newer breakpoints defined by PK/PD criteria, which take account of both susceptibility and attainable antibiotic concentrations in the blood. Interpretation of the data has also been made with the revised breakpoints issued in 2009. Use of these new breakpoints means that direct comparison between time periods is less straightforward, but is still useful. Data from other studies in Pakistan have highlighted that although the prevalence of S. pneumoniae strains with reduced susceptibility to penicillin and macrolides may be lower than reported in some regions of Asia,6,25 it remains an important consideration for effective empirical treatment of CA-RTIs. The reduced susceptibility of S. pneumoniae isolates to commonly used macrolides, such as erythromycin, is of concern. The 2007 – 09 data showed that 29.7% of isolates had reduced susceptibility to erythromycin (Table 2), which concurs with the 28% resistance reported in 2006 isolates by Zafar et al.8 Of particular concern, in the SOAR data analysis, erythromycin resistance rose to 54.8% in PISP isolates (Table 3). The clinical treatment of pneumonia or meningitis caused by S. pneumoniae may need to take into account the increase in PISP strains when selecting appropriate antimicrobial therapy. Specifically, clinical studies have shown that doses of penicillin and other agents may need to be adjusted if it is suspected that penicillin susceptibility is reduced.26 Guidelines in several countries propose use of higher doses of some b-lactam antibiotics because of concerns that PISP strains may not be adequately treated with those doses that were originally recommended some 30 years ago. The data from the SOAR studies for Pakistan have, however, confirmed full susceptibility to amoxicillin/clavulanic acid (amoxicillin) and cefuroxime, enabling these agents to continue to be used with confidence for appropriate S. pneumoniae-associated infections. For H. influenzae, data from the SOAR studies has shown that ampicillin resistance among H. influenzae strains isolated from documented CA-RTIs in Pakistan appears to be relatively rare (4%) and almost unchanged from 2002 to 2009. Furthermore, no BLNAR isolates were reported in this survey. This type of resistance is most frequently attributable to carriage of plasmids mediating b-lactamase with consequent opportunity for rapid spread. The low level of resistance in Pakistan contrasts markedly with the situation in some countries in Asia where b-lactamase production is approaching 50%.25 Knowledge of this very fortunate position in Pakistan may provide the opportunity for this low level of resistance to be maintained for as long as possible. Group A streptococci (GAS) isolates have remained highly susceptible to b-lactam antibiotics, but the level of macrolide resistance is high: 21% in Pakistan compared with ,5% in Algeria and Egypt.16 As well as being a cause of tonsillopharyngitis, GAS, if untreated, can lead to more severe complications including rheumatic fever with cardiac sequelae. In Pakistan, cardiac heart murmur due to rheumatic heart disease is common (prevalence 5.7/1000 patients screened in one survey).7 Effective primary treatment of proven S. pyogenes is therefore crucial and, whilst penicillin susceptibility remains at 100%, compliance may be an issue for the recommended full treatment of
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10 days of oral penicillin V. The high level of macrolide resistance means that this class of antibiotic may not be an appropriate treatment for GAS in Pakistan. Shorter treatments (4 – 5 days) with amoxicillin/clavulanic acid (or amoxicillin) or cefuroxime axetil have been shown to be as effective, or superior to, standard penicillin treatment and may aid compliance.27 In conclusion, when choosing antibiotic therapy for CA-RTIs, local antibiotic susceptibility/resistance data are essential to support informed therapy choices. The SOAR studies over the period from 2002 to 2009, along with the further study in 2014–15, have confirmed that the highly effective antibiotics, amoxicillin/clavulanic acid (amoxicillin in the case of S. pneumoniae) and cefuroxime, have retained almost 100% susceptibility against common CA-RTI isolates of S. pneumoniae, H. influenzae and S. pyogenes. In addition, organizations such as the WHO have emphasized the importance of choosing the best possible drug and treating for the optimal duration to prevent the further emergence of resistant bacterial strains.
Acknowledgements SOAR 2002– 03 results were presented at the 14th European Congress of Clinical Microbiology and Infectious Diseases, Prague, Czech Republic, 2004 (abstract number: P-507). SOAR 2004 – 06 results were presented at the 8th European Congress of Chemotherapy and Infection, Budapest, Hungary, 2006 (abstract number: 279) and at the 46th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, USA, 2006 (abstract number: C2-0425). SOAR 2007 – 09 results were presented at the 49th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, USA, 2009 (abstract number: C2-1402).
Funding This study was funded by GlaxoSmithKline.
Transparency declarations This article is part of a Supplement sponsored by GlaxoSmithKline. D. Torumkuney, F. Ali and K. Barker are employees of GlaxoSmithKline. D. Torumkuney and K. Barker also hold shares in GlaxoSmithKline. I. Morrissey is an employee of IHMA, a medical communication and consultancy company, who participated in the exploration, interpretation of the results on behalf of GSK. All other authors declare that they have no conflict of interest. Livewire Editorial Communications (Stella Deane and Tracey Morris) provided medical writing support in the form of writing assistance, collating author’s comments, grammatical editing and referencing that was paid for by GSK.
References 1 Sahm DF, Brown NP, Thornsberry C et al. Antimicrobial susceptibility profiles among common respiratory tract pathogens: a GLOBAL perspective. Postgrad Med 2008; 120 Suppl 1: 16– 24. 2 File TM. Community-acquired pneumonia. Lancet 2003; 362: 1991–2001. 3 Guthrie R. Community-acquired lower respiratory tract infections: etiology and treatment. Chest 2001; 120: 2021 –34. 4 Nicolau D. Clinical and economic implications of antimicrobial resistance for the management of community-acquired respiratory tract infections. J Antimicrob Chemother 2002; 50 Suppl 1: 61– 70. 5 Bhutta ZA. Burden of H. influenzae and S. pneumoniae infections in Pakistan. J College Phys Surg Pakistan 2001; 10: 346–54.
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6 Bravo LC, Asian Strategic Alliance for Pneumococcal Disease Prevention (ASAP) Working Group. Overview of the disease burden of invasive pneumococcal disease in Asia. Vaccine 2009; 27: 7282– 9. 7 Rizvi SF, Khan MA, Kundi A et al. Status of rheumatic heart disease in rural Pakistan. Heart 2004; 90: 394–9. 8 Zafar A, Hussain Z, Lomama E et al. Antibiotic susceptibility of pathogens isolated from patients with community-acquired respiratory tract infections in Pakistan—the active study. J Ayub Med Coll Abbottabad 2008; 20: 7 –9. 9 Pallares R, Fenoll A, Linares J et al. The epidemiology of antibiotic resistance in Streptococcus pneumoniae and the clinical relevance of resistance to cephalosporins, macrolides and quinolones. Int J Antimicrob Agents 2003; 22 Suppl 1: S15–24. 10 Beekmann SE, Heilmann KP, Richter SS et al. Antimicrobial resistance in Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and group A b-haemolytic streptococci in 2002 – 2003. Results of the multinational GRASP Surveillance Program. Int J Antimicrob Agents 2005; 25: 148–56. 11 Jacobs MR, Felmingham D, Applebaum PC et al. The Alexander Project 1998 – 2000: susceptibility of pathogens isolated from communityacquired respiratory tract infection to commonly used antimicrobial agents. J Antimicrob Chemother 2003; 52: 229–46. 12 Felmingham D, Feldman C, Hryniewicz W et al. Surveillance of resistance in bacteria causing community-acquired respiratory tract infections. Clin Microbiol Infect 2002; 8 Suppl 2: 12–42. 13 Shibl AM, Daniels J, Sievers J, SOAR in AME Study Group. Antimicrobial resistance among Streptococcus pneumoniae and Haemophilus influenzae from Africa and the Middle East: 2002/2003. In: Abstracts of the Fourteenth European Congress of Clinical Microbiology and Infectious Diseases, Prague, 2004. Clin Micro Infect 2004; 10 Suppl 3: 111. Poster data on file GSK2014N216848_00.2014. 14 Sievers J, SOAR Study Group. Antibacterial resistance among Haemophilus influenzae from 8 countries in Africa and the Middle East: results from the Survey of Antibiotic Resistance (SOAR) 2004 – 2006. In: Abstracts of the Eightieth European Congress of Chemotherapy and Infection, Budapest, 2006. Abstract 279. 15 Sievers J, SOAR Study Group. Antibacterial resistance among Streptococcus pneumoniae from ten countries in Africa and the Middle East: results from the Survey of Antibiotic Resistance (SOAR) 2004 – 2006. In: Abstracts of the Forty-sixth Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, 2006. Abstract C2-0425. 16 Torumkuney-O’Brien D, SOAR Study Group. Antibacterial resistance among Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes from 9 countries in the Middle East and Africa: results from the Survey of Antibiotic Resistance (SOAR) 2007–2009. In: Abstracts of the Forty-ninth Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, 2009. Abstract C2-1402. 17 O’Callaghan CH, Morris A, Kirby SM et al. Novel method for detection of b-lactamases by using a chromogenic cephalosporin substrate. Antimicrob Agents Chemother 1972; 1: 283–8. 18 Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Fifteenth Informational Supplement M100-S15. CLSI, Wayne, PA, USA, 2005. 19 Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Fourteenth Informational Supplement M100-S14. CLSI, Wayne, PA, USA, 2004. 20 Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Nineteenth Informational Supplement M100-S19. PA, USA, 2009.
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21 Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Twenty Fourth Informational Supplement M100-S24. Wayne, PA, USA, 2014. 22 Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Disk Susceptibility Tests—Tenth Edition: Approved Standard M2-A10. CLSI, Wayne, PA, USA, 2009. 23 Jacobs MR. Optimisation of antimicrobial therapy using pharmacokinetic and pharmacodynamic parameters. Clin Microbiol Infect 2001; 7: 589–96. 24 Shibl AM. Patterns of macrolide resistance determinants among S. pyogenes and S. pneumoniae isolates in Saudi Arabia. J Int Med Res 2005; 33: 349–55.
JAC 25 Wang H, Chen M, Xu Y et al. Antimicrobial susceptibility of bacterial pathogens associated with community-acquired respiratory tract infections in Asia: report from the Community-Acquired Respiratory Tract Infection Pathogen Surveillance (CARTIPS) study, 2009 – 2010. Int J Antimicrob Agents 2011; 38: 376–83. 26 Heffelfinger JD, Dowell SF, Jorgensen JH et al. Management of community-acquired pneumonia in the era of pneumococcal resistance. Arch Int Med 2000; 160: 1399–408. 27 Adam D, Scholz H, Helmerking M. Comparison of short-course (5 day) cefuroxime axetil with a standard 10 day oral penicillin V regimen in the treatment of tonsillopharyngitis. J Antimicrob Chemother 2000; 45 Suppl: 23–30.
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Antibiotic susceptibility in Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes in Pakistan: a review of results from the Survey of Antibiotic Resistance (SOAR) 2002–15 A. Zafar1, R. Hasan1, S. Nizamuddin2, N. Mahmood2, S. Mukhtar2, F. Ali3, I. Morrissey4, K. Barker5 and D. Torumkuney5* 1
Aga Khan University Hospital, Department of Pathology and Laboratory Medicine, Section of Microbiology, Karachi, Pakistan; 2Shaukat Khanum Memorial Cancer Hospital and Research Center, Department of Microbiology, 7A, Block R-3, Johar Town, Lahore, Pakistan; 3 GlaxoSmithKline Pakistan, 35 Dockyard Road, West Wharf, Karachi 74000, Pakistan; 4IHMA Europe Sa`rl, 9a route de la Corniche, Epalinges 1066, Switzerland; 5GlaxoSmithKline, 980 Great West Road, Brentford, Middlesex TW8 9GS, UK *Corresponding author. Tel: +44-20-8047-1671; E-mail: [email protected]
Objectives: To investigate changes in the antibiotic susceptibility of Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes from the Survey of Antibiotic Resistance (SOAR) in community-acquired respiratory tract infections (CA-RTIs) between 2002 and 2015 in Pakistan. Methods: This is a review based on previously published studies from 2002 –03, 2004– 06 and 2007– 09 and also new data from 2014–15. Susceptibility was determined by Etestw or disc diffusion according to CLSI and pharmacokinetic/pharmacodynamic (PK/PD) breakpoints. Results: A total of 706 isolates from CA-RTIs comprising 381 S. pneumoniae, 230 H. influenzae and 95 S. pyogenes were collected between 2002 and 2015 and tested against a range of antibiotics. Antibiotic resistance in S. pneumoniae rose steeply from 2002 to 2009, with isolates non-susceptible to penicillin and macrolides increasing from 10% to 34.1% and from 13% –14% to 29.7%, respectively. Susceptibility to amoxicillin/clavulanic acid (and by inference amoxicillin) remained between 99.4% and 100% from 2002 to 2015. Over the years, the prevalence of susceptibility to cefuroxime was 98% – 100% among S. pneumoniae. Resistance in S. pneumoniae to some older antibiotics between 2007 and 2009 was high (86.8% for trimethoprim/sulfamethoxazole and 57.2% for tetracycline). Between 2002 and 2015, ampicillin resistance (b-lactamase-positive strains) among H. influenzae has remained low (between 2.6% and 3.2%) and almost unchanged over the years (H. influenzae was not tested during 2004–06). For S. pyogenes isolates, macrolide resistance reached 22%; however, susceptibility to penicillin, amoxicillin/clavulanic acid and cefuroxime remained stable at 100%. Conclusions: In S. pneumoniae from Pakistan, there has been a clear reduction in susceptibility to key antibiotics since 2002, but not to amoxicillin/clavulanic acid (amoxicillin) or cefuroxime. However, susceptibility in H. influenzae has remained stable. Local antibiotic susceptibility/resistance data are essential to support informed prescribing for CA-RTIs and other infections.
Introduction Worldwide, there are concerns about rising levels of antibiotic resistance and fears that the efficacy of antimicrobial therapy may be compromised, resulting in treatment failure and reduced utility of older antibiotics.1 Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes are all commonly carried in the nasopharynx and implicated in community-acquired respiratory tract infections (CA-RTIs). These infections account for significant morbidity and mortality, including severe life-threatening infections such as pneumonia and meningitis (S. pneumoniae and H. influenzae) and rheumatic fever with risk of cardiac sequelae (S. pyogenes).2 – 7
Like many other Asian countries, in Pakistan antimicrobial therapeutic choices for RTIs are usually empirical due to the limited availability of laboratory identification and antibiotic susceptibility testing of bacterial isolates. Even when available, the cost of laboratory testing in this setting can often prove inhibitive.8 The increasing prevalence of resistant organisms threatens the current and future treatment of CA-RTIs.4,9 – 11 Surveillance studies, therefore, provide an important tool for determining local and regional susceptibility patterns and guiding empirical antimicrobial therapy.12 The Survey of Antibiotic Resistance (SOAR) studies have been undertaken in Pakistan during 2002 – 03, 2004 – 06, 2007 – 09 and 2014 – 15, as well as elsewhere in Asia, the Middle East, Africa, Latin America, Asia Pacific and the Commonwealth of
# The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: [email protected]
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Independent States countries, to provide detailed and comparative information on resistance trends among strains from documented CA-RTIs caused by S. pneumoniae, H. influenzae and S. pyogenes.13 – 16
Materials and methods Collaborating centres Isolates were collected from two sites in Pakistan: Aga Khan University Hospital (for the 2002 – 09 study period) Karachi and Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore (for the 2014– 15 study period).
Clinical isolates (from outpatients who attended the university hospitals)
susceptibility to amoxicillin alone), cefixime, cefuroxime, cefaclor, cefprozil, clarithromycin, azithromycin, erythromycin, ciprofloxacin, trimethoprim/sulfamethoxazole, tetracycline, chloramphenicol and clindamycin. S. pyogenes strains were tested against penicillin, amoxicillin, cefaclor, cefixime, erythromycin, azithromycin, clarithromycin, clindamycin, tetracycline and chloramphenicol. Finally, susceptibility of H. influenzae was assessed against ampicillin, amoxicillin/clavulanic acid, cefaclor, cefixime, cefprozil, cefuroxime, clarithromycin, azithromycin, ciprofloxacin, levofloxacin, trimethoprim/sulfamethoxazole, tetracycline and chloramphenicol. For the 2014 –15 isolates, susceptibility testing was carried out using a more limited range of antibiotics. S. pneumoniae susceptibility was tested against amoxicillin/clavulanic acid (amoxicillin), cefaclor, cefuroxime, chloramphenicol, erythromycin and penicillin. H. influenzae was tested against amoxicillin/clavulanic acid, ampicillin, azithromycin, cefaclor, cefuroxime, chloramphenicol and levofloxacin using the same CLSI breakpoints as for 2009 isolates.
Between 2002 and 2015, a total of 381 S. pneumoniae, 230 H. influenzae and 95 S. pyogenes isolates were obtained from a range of clinical samples including sputum, blood, bronchoalveolar lavage, middle ear effusion and tracheal aspirate taken from adult and paediatric patients with a confirmed CA-RTI using routine clinical collection methods. All S. pyogenes isolates were from the throat. Duplicate isolates from the same patient were not accepted. S. pneumoniae isolates were collected during 2002 – 03, 2004 – 06, 2007 – 09 and 2014 – 15. H. influenzae isolates were collected over the same time periods, except none were collected in 2004 –06. S. pyogenes isolates were collected only during 2007 –09. The isolates were identified using conventional methods (optochin susceptibility for S. pneumoniae, X and V factor requirement for H. influenzae and bacitracin antibiotic disc susceptibility for S. pyogenes strains). Isolates were stored at 2708C until tested. b-Lactamase production of H. influenzae isolates was determined by a chromogenic cephalosporin (nitrocefin) disc method.17 b-Lactamase-negative ampicillin-resistant (BLNAR) isolates were defined as those organisms having a negative b-lactamase result and an ampicillin MIC of ≥4 mg/L.18
Quality control strains S. pneumoniae ATCC 49619, H. influenzae ATCC 49247, H. influenzae ATCC 49766, Escherichia coli ATCC 25922 and E. coli ATCC 32518 were included on each day of testing. Results of susceptibility testing were accepted if the results of the control strains were within published limits. Any Etestw MIC results that were between doubling dilutions were rounded up to the next doubling dilution MIC for data analysis. MICs and zone diameters were interpreted qualitatively using CLSI interpretive standards; MICs were also analysed using PK/PD breakpoints established to predict the clinical and bacteriological efficacy of antimicrobial dosing regimens. 11,19,23 To interpret azithromycin and clarithromycin MICs for H. influenzae, revised breakpoints were used to account for incubation in a CO2 atmosphere (using the Etestw susceptibility method according to the manufacturer’s instructions; bioMe´rieux, Marcy-l’Etoile, France). The respective PK/PD breakpoints were adjusted accordingly, i.e. raised by one doubling dilution for interpretation of H. influenzae susceptibility because of incubation under 5% CO2.
Susceptibility testing
Results w
MICs were determined using Etest susceptibility testing methods according to the manufacturer’s instructions [AB BIODISK (Solna, Sweden) for 2002 – 09 and bioMe´rieux (Marcy-l’Etoile, France) for 2014 – 15] and interpreted using CLSI guidelines and pharmacokinetic/pharmacodynamic (PK/PD) breakpoints (for 2002 – 03, M100-S1419; for 2004 – 06, CLSI M100-S1518; for 2007 – 09, M100-S1920; and for 2014 – 15, M100-S2421). For isolates collected during 2007 – 09, susceptibility to erythromycin, tetracycline, trimethoprim/sulfamethoxazole and chloramphenicol was also determined using the CLSI disc diffusion method.22 Briefly, for Etestw MIC testing, S. pneumoniae isolates were subcultured twice on blood-supplemented Mueller–Hinton agar and H. influenzae isolates were cultured on Haemophilus Test Medium before susceptibility testing was performed. Inocula were prepared by direct colony suspension to give a 0.5 McFarland standard dilution and inoculated onto appropriate agar plates to produce a confluent lawn of growth. Etestw and antibiotic discs were applied and plates incubated for 20 – 24 h (16 – 17 h for H. influenzae agar disc diffusion) at 358C in a 5% CO2 atmosphere. For azithromycin and clarithromycin Etests, S. pneumoniae and S. pyogenes isolates were incubated in ambient air due to the adverse effect of CO2 on the activity of macrolide antibiotics. Etestw MICs and inhibition zone diameters were read in accordance with bioMe´rieux’s instructions and CLSI guidelines for disc susceptibility testing, respectively. The number of antibiotics tested varied at the different timepoints. Between 2002 and 2009, S. pneumoniae strains were tested against penicillin, amoxicillin/clavulanic acid (data from which can be used to deduce
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Quality control and data analysis
From 2002 to 2009, CLSI breakpoint interpretations for S. pneumoniae were unchanged for all of the study drugs except penicillin. PK/PD breakpoints were also in good agreement with the ‘susceptible’ category, except for cefaclor and azithromycin. For both of these antibiotics, the susceptible breakpoint from PK/PD data was lower than the CLSI breakpoint—by one dilution for cefaclor and three dilutions for azithromycin, reflecting the relatively low blood level attainable with this antibiotic. Table 1 shows the breakpoints used to interpret Etestw MICs for strains isolated in the period 2002 –15. During the course of the studies, S. pneumoniae with raised penicillin MICs were being reported with increasing frequency worldwide. The original breakpoints for penicillin of susceptible (S) ≤0.06 mg/L, intermediate (I) 0.12– 1.0 mg/L and resistant (R) ≥2.0 mg/L had been appropriate for use of oral penicillin V and for injectable penicillin used in meningitis treatment. However, with an increasing prevalence of penicillin-intermediate S. pneumoniae (PISP) strains, the use of penicillin was questioned for treatment of S. pneumoniae infections other than meningitis and particularly for pneumonia. This concern was addressed by CLSI and new penicillin breakpoints were applied for S. pneumoniae from non-meningeal sources.22 New breakpoints were recommended in 2009 specifically for use with S. pneumoniae from RTIs. High doses of penicillin and other b-lactam antibiotics,
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Table 1. MIC breakpoints (mg/L) used for S. pneumoniae and H. influenzae isolates CLSI breakpointsa PK/PD breakpoints Antimicrobial Penicillinb Penicillinc (only for 2007– 09/2014 – 15) Ampicillin Amoxicillin AMCd Cefaclor Cefixime Cefprozil Cefuroximee Azithromycinf Clarithromycinf Ciprofloxacin Levofloxacin
S. pneumoniae
H. influenzae
S
R
S
I
R
S
I
R
— — — ≤2 ≤2 ≤0.5 ≤1 ≤1 ≤1 ≤0.12 ≤0.25 ≤1 ≤2
— — — ≥4 ≥4 ≥1 ≥2 ≥2 ≥2 ≥0.25 ≥0.5 ≥2 ≥4
≤0.06 ≤2 — ≤2 ≤2 ≤1 — ≤2 ≤1 ≤0.5 ≤0.25 — ≤2
0.12– 1 4 — 4 4 2 — 4 2 1 0.5 — 4
≥2 ≥8 — ≥8 ≥8 ≥4 — ≥8 ≥4 ≥2 ≥1 ≤1 ≥8
— — ≤1 — ≤4 ≤8 ≤1 ≤8 ≤4 ≤8 ≤16 — ≤2
— — 2 — — 16 — 16 8 — 32 — —
— — ≥4 — ≥8 ≥32 — ≥32 ≥16 — ≥64 — —
AMC, amoxicillin/clavulanic acid; —, no breakpoint available. For breakpoints see CLSI.22 b CLSI 2008 breakpoints/CLSI 2009 breakpoints for oral penicillin V. c CLSI 2009 breakpoints for parenteral penicillin/non-meningeal. d Amoxicillin/clavulanic acid was tested in a 2 : 1 ratio of amoxicillin to clavulanic acid; breakpoints are expressed as the amoxicillin component. e Breakpoints used are for cefuroxime axetil. f Azithromycin and clarithromycin breakpoints are those provided by AB Biodisk for incubation in CO2. a
such as amoxicillin, were also recommended for treatment of PISP or penicillin-resistant S. pneumoniae infections. These changes are documented in Table 1 and show that S. pneumoniae with penicillin MICs ≤2.0 mg/L are regarded as susceptible, whereas previously some of these isolates would have been regarded as having intermediate susceptibility. Clinical studies showed that, provided penicillin was used in adequate dosage, the new susceptibility category could be used as a breakpoint for non-meningeal isolates.
Antibiotic resistance in S. pneumoniae Overall, non-susceptibility to penicillin among S. pneumoniae strains rose steeply, from 10% in 2002 to 34.1% in 2009. The prevalence of penicillin-non-susceptible S. pneumoniae was 10% during 2002 – 03, 18.3% during 2004 – 06 and 34.1% during 2007– 09. During 2014 –15, only 10 S. pneumoniae isolates were collected, of which two were non-susceptible to penicillin. Of the 31 isolates that were penicillin non-susceptible in the 2007 – 09 survey, all had MICs of 4 mg/L, making them intermediate (Table 2). No isolates of S. pneumoniae obtained during the study were highly resistant to penicillin (MICs ≥8.0 mg/L), although 1.1% of strains in the 2004 – 06 survey were penicillin resistant using the lower CLSI breakpoint in use at that time. Macrolide resistance rose from 13% to 29.7% (depending on the macrolide being tested) over the study period (Figure 1 and Table 2, respectively). By contrast, susceptibility to amoxicillin/ clavulanic acid (or amoxicillin alone) between 2002 and 2015 has remained between 99.4% and 100% among S. pneumoniae isolates. Ciprofloxacin was tested only during 2007 – 09, when
91.2% of S. pneumoniae isolates were found to be susceptible. S. pneumoniae strains showing reduced susceptibility to penicillin were more likely to show increased resistance to macrolides and other antimicrobial agents, particularly older agents such as first-generation cephalosporins (cefaclor), tetracycline or trimethoprim/sulfamethoxazole (Table 3). Almost all PISP strains were resistant to trimethoprim/sulfamethoxazole, .70% were resistant to tetracycline and 50% were resistant to macrolides (erythromycin and clarithromycin). This contrasted with the macrolide resistance rate in penicillin-susceptible S. pneumoniae strains, which was only 10% – 15%. In the Lahore 2014 – 15 study, the erythromycin susceptibility of all S. pneumoniae isolates was 70%. Clindamycin resistance plus macrolide resistance is a useful proxy for detection of the erm(B) resistance marker. 24 In the 2007 – 09 study period, clindamycin resistance was 22.6% among PISP strains and this was coupled with a concomitant high incidence of macrolide resistance (50%) (Table 3). This finding suggests that a number of these isolates are more likely to have macrolide resistance attributable to the erm(B) resistance marker rather than the efflux mechanism of macrolide resistance. In the 2007 – 09 study period, of the S. pneumoniae isolates, 93.4% were susceptible to chloramphenicol (Table 2).
Antibiotic resistance in H. influenzae H. influenzae isolates studied in Pakistan during the 2002 –03 winter season (n ¼ 93) were fully susceptible (100%) to amoxicillin/ clavulanic acid, cefuroxime, cefprozil, cefaclor, cefixime, clarithromycin and azithromycin (Table 4). Furthermore, 3.2% of isolates
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Table 2. Susceptibility (%) of S. pneumoniae in Pakistan to antimicrobials using CLSI or PK/PD breakpoints 2002 –03 (n¼100)a CLSI Antimicrobial Penicillin AMC Cefixime Cefuroxime Cefaclor Cefprozil Clarithromycin Azithromycin Erythromycin Clindamycin SXT Tetracycline Chloramphenicol Ciprofloxacin
2004– 06 (n¼180)b PK/PD
S
I or (R)
90.0 100 93.0d 98.0 96.0 99.0 86.0e 87.0e — — — — — —
10.0 0 7.0 2.0 4.0 1.0 14.0 13.0 — — — — — —
S NA 99.4 80.6 98.3 67.2 — 90.8 73.4 NA NA NA NA NA —
2007– 09 (n¼91)c
CLSI
PK/PD
S
I or (R)
81.7 99.4 — 98.3 89.4 — 90.8 90.2 87.2 88.9 — — — —
18.3 (1.1) 0.6 (0) — 1.7 (1.1) 10.6 (2.8) — 9.2 9.8 12.8 11.1 — — — —
S NA 100 92.3 100 69.2 — 76.9 — NA NA NA NA NA 91.2
2014– 15 (n ¼10)c
CLSI
PK/PD
S
I or (R)
S
65.9 100 — 100 94.5 — 76.9 — 70.3 90.1 13.2 42.8 93.4 —
34.1 (0) 0 — 0 5.5 — 23.1 — 29.7 9.9 86.8 57.2 6.6 —
NA 100 100 70.0 — — — — — — — — —
CLSI S 80.0 100 — 100 90.0 — — — 70.0 — — — 80.0
I or (R) 20.0 0 — 0 10.0 — — — 30.0 — — — 20.0
Resistance (%)
AMC, amoxicillin/clavulanic acid; SXT, trimethoprim/sulfamethoxazole; S, susceptible; I, intermediate; R, resistant; NA, not applicable; —, no data available. a Data for strains from Pakistan summarized by Shibl et al.13 b Data from Pakistan summarized by Sievers et al. 15 c Pakistan current SOAR study data. d PK/PD breakpoint used. e Revised breakpoints for Etestw used.
35 30 25 20 15 10 5 0
Erythromycin resistance Clarithromycin resistance Reduced penicillin susceptibility 2002–03 2004–07 2007–09 Survey timepoint
Figure 1. Penicillin and macrolide resistance in S. pneumoniae strains from CA-RTIs from Pakistan during 2002 –09.
were designated b-lactamase positive on the basis of nitrocefin disc testing.13 For 2007–09 (n ¼99), amoxicillin/clavulanic acid susceptibility remained at 100% among H. influenzae isolates in Pakistan and ampicillin resistance was 4% (4/99 isolates). All isolates were susceptible to cefuroxime and cefixime. The prevalence of resistance to macrolides was between 6.1% and 8.1%. Ciprofloxacin was tested only during 2007–09, when 95.9% of H. influenzae isolates were found to be susceptible (Table 4). Of the 2014 –15 (n¼ 38) H. influenzae isolates, 100% were susceptible to amoxicillin/clavulanic acid and 97.4% were susceptible to ampicillin. Only 1 of the 38 isolates (2.63%) from 2014–15 was b-lactamase positive. Susceptibility to chloramphenicol was low
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at 29%. Some 86.8% of H. influenzae isolates were susceptible to levofloxacin (Table 4). Except for the 2014 –15 study period, all ampicillin-resistant H. influenzae isolates were b-lactamase positive. Although the prevalence of BLNAR isolates was not reported during different study periods, based on CLSI guidelines, rare BLNAR strains of H. influenzae should be considered resistant to amoxicillin/clavulanic acid, ampicillin/sulbactam, cefaclor, cefamandole, cefetamet, cefonicid, cefprozil, cefuroxime, loracarbef and piperacillin/tazobactam, despite apparent in vitro susceptibility of some BLNAR strains to these agents.
Antibiotic resistance in S. pyogenes S. pyogenes was also studied in the 2007 – 09 survey. All strains were susceptible to penicillin. Based on penicillin susceptibility, the strains were also susceptible to amoxicillin, amoxicillin/clavulanic acid, cefuroxime, cefotaxime and cefaclor. By contrast, macrolide (erythromycin, azithromycin and clarithromycin) resistance in these Pakistani CA-RTI isolates was 22% (data not shown).
Discussion The increase in antimicrobial resistance among CA-RTI pathogens is a serious problem throughout the world and has important implications for the successful management of these infections. Since there may be variation in levels of resistance between different geographical areas, it is essential to be aware of the local
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Table 3. Susceptibility (%) of penicillin-susceptible S. pneumoniae (PSSP) and penicillin-intermediate S. pneumoniae (PISP) strains from CA-RTIs in Pakistan in SOAR during 2007 –09 PSSP (n¼60), penicillin MIC ≤2.0 mg/La Antimicrobial Penicillin AMC Cefixime Cefuroxime Cefaclor Clarithromycin Ciprofloxacin Erythromycin Clindamycinb SXT Tetracycline Chloramphenicol
PISP (n¼31), penicillin MIC ≥4 mg/La
S (PK/PD) S (CLSI) R (CLSI) S (PK/PD) S (CLSI) R (CLSI) NA 100 98.3 100 86.7 90 91.7 NA NA NA NA NA
100 100 — 100 98.3 90 — 83.3 96.7 18.3 50 93.3
0 0 — 0 0 10 — 15 3.3 78.4 40 6.7
NA 100 80.6 100 34.5 51.6 90.3 NA NA NA NA NA
0 100 — 100 87.1 51.6 — 45.2 77.4 3.2 29.1 93.5
0 0 — 0 0 48.4 — 54.8 22.6 96.8 70.9 6.5
AMC, amoxicillin/clavulanic acid; SXT, trimethoprim/sulfamethoxazole; S, susceptible; I, intermediate; R, resistant; NA, not applicable; —, no data available. a New CLSI breakpoints for penicillin applied. b Included to differentiate erm(B)-mediated macrolide resistance.
Table 4. Susceptibility (%) of H. influenzae in Pakistan to antimicrobials using CLSI breakpoints 2002 –03 (n ¼93)a Antimicrobial Ampicillin AMC Cefaclor Cefixime Cefprozil Cefuroxime Clarithromycin Azithromycin Chloramphenicol Levofloxacin Ciprofloxacin SXT Tetracycline
2007 –09 (n¼99)b
2014 –15 (n¼38)b
S
I or (R)
S
I or (R)
S
I or (R)
96.8 100 100 100 100 100 100 100 — — — — —
3.2 0 0 0 0 0 0 0 — — — — —
96 100 98 100 — 100 91.9 93.9 88.8 — 95.9 15.2 67.7
4 0 2 0 — 0 8.1 6.1 11.2 — 4.1 84.8 32.3
97.4 100 89.5 — — 97.4 — 100 29 86.8 — — —
2.6 0 10.5 — — 2.6 — 0 71 13.2 — —
AMC, amoxicillin/clavulanic acid; SXT, trimethoprim/sulfamethoxazole; S, susceptible; I, intermediate; R, resistant; —, no data available. a Data for strains from Pakistan summarized by Shibl et al.13 b Pakistan current SOAR study data.
resistance pattern before considering empirical antimicrobial therapy. SOAR was therefore established to provide information on local resistance patterns, as well as overall levels of resistance and susceptibility trends, for key CA-RTI pathogens.
The SOAR studies have been conducted using the Etestw because reliable MIC results can be obtained relatively easily and subsequently the results can be analysed with reference to different breakpoint criteria. In these studies, analyses have been performed using breakpoints defined by CLSI and also newer breakpoints defined by PK/PD criteria, which take account of both susceptibility and attainable antibiotic concentrations in the blood. Interpretation of the data has also been made with the revised breakpoints issued in 2009. Use of these new breakpoints means that direct comparison between time periods is less straightforward, but is still useful. Data from other studies in Pakistan have highlighted that although the prevalence of S. pneumoniae strains with reduced susceptibility to penicillin and macrolides may be lower than reported in some regions of Asia,6,25 it remains an important consideration for effective empirical treatment of CA-RTIs. The reduced susceptibility of S. pneumoniae isolates to commonly used macrolides, such as erythromycin, is of concern. The 2007 – 09 data showed that 29.7% of isolates had reduced susceptibility to erythromycin (Table 2), which concurs with the 28% resistance reported in 2006 isolates by Zafar et al.8 Of particular concern, in the SOAR data analysis, erythromycin resistance rose to 54.8% in PISP isolates (Table 3). The clinical treatment of pneumonia or meningitis caused by S. pneumoniae may need to take into account the increase in PISP strains when selecting appropriate antimicrobial therapy. Specifically, clinical studies have shown that doses of penicillin and other agents may need to be adjusted if it is suspected that penicillin susceptibility is reduced.26 Guidelines in several countries propose use of higher doses of some b-lactam antibiotics because of concerns that PISP strains may not be adequately treated with those doses that were originally recommended some 30 years ago. The data from the SOAR studies for Pakistan have, however, confirmed full susceptibility to amoxicillin/clavulanic acid (amoxicillin) and cefuroxime, enabling these agents to continue to be used with confidence for appropriate S. pneumoniae-associated infections. For H. influenzae, data from the SOAR studies has shown that ampicillin resistance among H. influenzae strains isolated from documented CA-RTIs in Pakistan appears to be relatively rare (4%) and almost unchanged from 2002 to 2009. Furthermore, no BLNAR isolates were reported in this survey. This type of resistance is most frequently attributable to carriage of plasmids mediating b-lactamase with consequent opportunity for rapid spread. The low level of resistance in Pakistan contrasts markedly with the situation in some countries in Asia where b-lactamase production is approaching 50%.25 Knowledge of this very fortunate position in Pakistan may provide the opportunity for this low level of resistance to be maintained for as long as possible. Group A streptococci (GAS) isolates have remained highly susceptible to b-lactam antibiotics, but the level of macrolide resistance is high: 21% in Pakistan compared with ,5% in Algeria and Egypt.16 As well as being a cause of tonsillopharyngitis, GAS, if untreated, can lead to more severe complications including rheumatic fever with cardiac sequelae. In Pakistan, cardiac heart murmur due to rheumatic heart disease is common (prevalence 5.7/1000 patients screened in one survey).7 Effective primary treatment of proven S. pyogenes is therefore crucial and, whilst penicillin susceptibility remains at 100%, compliance may be an issue for the recommended full treatment of
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10 days of oral penicillin V. The high level of macrolide resistance means that this class of antibiotic may not be an appropriate treatment for GAS in Pakistan. Shorter treatments (4 – 5 days) with amoxicillin/clavulanic acid (or amoxicillin) or cefuroxime axetil have been shown to be as effective, or superior to, standard penicillin treatment and may aid compliance.27 In conclusion, when choosing antibiotic therapy for CA-RTIs, local antibiotic susceptibility/resistance data are essential to support informed therapy choices. The SOAR studies over the period from 2002 to 2009, along with the further study in 2014–15, have confirmed that the highly effective antibiotics, amoxicillin/clavulanic acid (amoxicillin in the case of S. pneumoniae) and cefuroxime, have retained almost 100% susceptibility against common CA-RTI isolates of S. pneumoniae, H. influenzae and S. pyogenes. In addition, organizations such as the WHO have emphasized the importance of choosing the best possible drug and treating for the optimal duration to prevent the further emergence of resistant bacterial strains.
Acknowledgements SOAR 2002– 03 results were presented at the 14th European Congress of Clinical Microbiology and Infectious Diseases, Prague, Czech Republic, 2004 (abstract number: P-507). SOAR 2004 – 06 results were presented at the 8th European Congress of Chemotherapy and Infection, Budapest, Hungary, 2006 (abstract number: 279) and at the 46th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, USA, 2006 (abstract number: C2-0425). SOAR 2007 – 09 results were presented at the 49th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, USA, 2009 (abstract number: C2-1402).
Funding This study was funded by GlaxoSmithKline.
Transparency declarations This article is part of a Supplement sponsored by GlaxoSmithKline. D. Torumkuney, F. Ali and K. Barker are employees of GlaxoSmithKline. D. Torumkuney and K. Barker also hold shares in GlaxoSmithKline. I. Morrissey is an employee of IHMA, a medical communication and consultancy company, who participated in the exploration, interpretation of the results on behalf of GSK. All other authors declare that they have no conflict of interest. Livewire Editorial Communications (Stella Deane and Tracey Morris) provided medical writing support in the form of writing assistance, collating author’s comments, grammatical editing and referencing that was paid for by GSK.
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