Investigation of Antibiotic Susceptibility Profiles and Beta-Lactamase Activity of Haemophilus influenzae Isolates

Öz

Haemophilus influenzae is one of the important Gram negative bacterial pathogens which can cause respiratory infections and invasive infections. In this study, antibiotic susceptibility profiles and β-lactamase activity of 480 H.influenzae isolates isolated from different invasive/noninvasive clinical cases in the Central Microbiology Laboratory of Hacettepe University Medical Faculty Hospital between January 2018 and August 2019 were investigated retrospectively. Isolates were identified by using matrix-mediated laser desorption ionization flight time mass spectrometry (MALDI-TOF MS) and antibiotic susceptibility tests were performed with disk diffusion method using Haemophilus Test Medium, according to the Clinical and Laboratory Standards Institute (CLSI) 2018 guidelines. Antibiotic susceptibility against amoxicillin-clavulanate, ampicillin, meropenem, piperacillin-tazobactam, rifampin, cefotaxime, ceftazidime, ceftriaxone, cefuroxime, ciprofloxacin, tetracycline and trimethoprim/sulfamethoxazole were tested. The β-lactamase production of isolates was investigated by using nitrocefin disk. The highest resistance rates for isolates among tested antibiotics were detected respectively against trimethoprim-sulfamethoxazole (30,4 %) and ampicillin (16,2 %). In addition, it was found that of the isolates 7,9 % were β-lactamase negative ampicillin resistant (BLNAR) origin, 8,3 % were β-lactamase positive ampicillin resistant (BLPAR) origin, and 1,9 % were β-lactamase positive amoxicillin / clavulanate resistant (BLPACR). Antibiotic resistance rates in H.influenzae isolates vary between centers and ampicillin resistance and BLNAR rates are progressively increasing. In our study, high rates of ampicillin resistance and BLNAR origin were detected in H.influenzae isolates. Our results suggest that centers should follow epidemiological data regularly in order to guide appropriate antibiotic treatment to be applied to patients in H.influenzae infections.

Anahtar Kelimeler

• ampicillin
• beta-lactamase
• BLNAR
• BLPAR
• H.influenzae

Haemophilus influenza Suşlarının Antibiyotik Duyarlılık Profillerinin ve Beta-Laktamaz Aktivitesinin Araştırılması

Öz

Haemophilus influenzae solunum yolu enfeksiyonlarına ve invaziv enfeksiyonlara neden olabilen önemli Gram negatif bakteriyel patojenlerden biridir. Çalışmamızda Hacettepe Üniversitesi Tıp Fakültesi Hastanesi Merkez Mikrobiyoloji Laboratuvarı’nda Ocak 2018-Ağustos 2019 tarihleri arasında invaziv/invaziv olmayan çeşitli klinik olgulardan izole edilen 480 H.influenzae suşunun antibiyotik duyarlılık profilleri ve β-laktamaz üretimi retrospektif olarak araştırılmıştır. Suşlar matriks aracılı lazer dezorpsiyon iyonizasyon uçuş zamanlı kütle spektrometresi (MALDI-TOF MS) ile tanımlanmış ve in vitro antibiyotik duyarlılık testleri Haemophilus Test besiyeri ile Clinical and Laboratory Standards Institute (CLSI) 2018 dokümanı referans alınarak disk difüzyon yöntemi ile yapılmıştır. Antibiyotik duyarlılık testlerinde amoksisilinklavulanat, ampisilin, meropenem, piperasilin-tazobaktam, rifampin, sefotaksim, seftazidim, seftriakson, sefuroksim, siprofloksasin, tetrasiklin ve trimetoprim-sülfametoksazol test edilmiştir. İzolatların β-laktamaz üretimi nitrosefin diski kullanılarak araştırılmıştır. Suşlar için test edilen antibiyotikler arasında en yüksek direnç oranları sırasıyla trimetoprimsülfametoksazol (% 30,4) ve ampisilinde (% 16,2) tespit edilmiştir. Suşların % 7,9’u β-laktamaz negatif ampisilin dirençli (BLNAR), % 8,3’ü β-laktamaz pozitif ampisilin dirençli (BLPAR), % 1,9’u ise β-laktamaz pozitif amoksisilin/klavulanata dirençli (BLPACR) olarak saptanmıştır. H.influenzae suşlarında antibiyotik direnç oranları merkezler arasında değişiklik göstermekte olup, bu suşlarda ampisilin direnç ve BLNAR oranları gittikçe artmaktadır. Çalışmamızda H.influenzae suşlarında yüksek oranda ampisiline dirençli ve BLNAR köken tespit edilmiştir. Sonuçlarımız H.influenzae enfeksiyonlarında hastalara uygulanacak uygun antibiyotik tedavisini yönlendirmesi açısından merkezlerin düzenli olarak epidemiyolojik verilerini takip etmesi gerekliliğini düşündürmektedir.

Anahtar Kelimeler

• ampisilin
• beta-laktamaz
• BLNAR
• BLPAR
• H.influenzae

Kaynakça

1. Altun B, Gür D. Hacettepe Üniversitesi Çocuk Hastanesi’nde klinik örneklerden izole edilen Haemophilus influenzae suşlarının antibiyotiklere direnç durumu (2002-2007). Çocuk Enfeksiyon Derg. 2008;2(2):50.
2. Bae S, Lee J, Lee J, et al. Antimicrobial resistance in Haemophilus influenzae respiratory tract isolates in Korea: results of a nationwide acute respiratory infections surveillance. Antimicrob Agents Chemother. 2010;54(1):65-71.
3. Barry AL, Fuchs PC, Brown SD. Identification of β-lactamase-negative, ampicillin-resistant strains of Haemophilus influenzae with four methods and eight media. Antimicrob Agents Chemother. 2001;45(5):1585-8.
4. Berkiten R. Türkiye’de Haemophilus influenzae: beta-laktamaz pozitifliği ve antibiyotiklere direnç (1987-2002). ANKEM Derg. 2004;18(1):53-60.
5. Budak F, Gür D. In vitro sensitivity to antimicrobial agents of Haemophilus influenzae strains isolated from clinical specimens. Mikrobiyol Bul. 2003;37(1):19-25.
6. Cerquetti M, Cardines R, Giufré M, Mastrantonio P. Antimicrobial susceptibility of Haemophilus influenzae strains isolated from invasive disease in Italy. J Antimicrob Chemother. 2004;54(6):1139-43.
7. Clinical Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing; Twenty-Eightth Informational Supplement. M100-S23, CLSI, Wayne, PA (2018).
8. García-Cobos S, Campos J, Lázaro E, et al. Ampicillin-resistant non-β-lactamase-producing Haemophilus influenzae in Spain: recent emergence of clonal isolates with increased resistance to cefotaxime and cefixime. Antimicrob Agents Chemother. 2007;51(7):2564-73.

9. Gazi H, Kurutepe S, Surucuoglu S, Teker A, Ozbakkaloglu B. Antimicrobial susceptibility of bacterial pathogens in the oropharynx of healthy school children in Turkey. Indian J Med Res. 2004;120(5):489-94.
10. Gonullu N, Catal F, Kucukbasmaci O, Ozdemir S, Torun MM, Berkiten R. Comparison of in vitro activities of tigecycline with other antimicrobial agents against Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis in two university hospitals in Istanbul, Turkey. Chemotherapy. 2009; 55(3):161-7.
11. Hoban D, Felmingham D. The PROTEKT surveillance study: antimicrobial susceptibility of Haemophilus influenzae and Moraxella catarrhalis from community-acquired respiratory tract infections. J Antimicrob Chemother 2002;50(Suppl 2):49-59.
12. Ilki A, Sağiroğlu P, Elgörmüş N, Söyletir G. Trends in antibiotic susceptibility patterns of Streptococcus pneumoniae and Haemophilus influenzae isolates: four years follow up. Mikrobiyol Bul. 2010;44(2):169-75.
13. Johnson D, Sader H, Fritsche T, Biedenbach D, Jones RN. Susceptibility trends of Haemophilus influenzae and Moraxella catarrhalis against orally administered antimicrobial agents: fiveyear report from the SENTRY Antimicrobial Surveillance Program. Diagn Microbiol Infect Dis. 2003;47(1):373-6.
14. Kaczmarek FS, Gootz TD, Dib-Hajj F, Shang W, Hallowell S, Cronan M. Genetic and molecular characterization of β-lactamase-negative ampicillin-resistant Haemophilus influenzae with unusually high resistance to ampicillin. Antimicrob Agents Chemother. 2004;48(5):1630-9.
15. Kocabeyoğlu Ö Bİ, Koşan E. Haemophilus influenzae suşlarında beta-laktamaz aktivitesi ve çeşitli antibiyotiklere duyarlılık. ANKEM Derg. 1996;10(2):119.
16. Morrissey I, Maher K, Williams L, Shackcloth J, Felmingham D, Reynolds R. Non-susceptibility trends among Haemophilus influenzae and Moraxella catarrhalis from community-acquired respiratory tract infections in the UK and Ireland, 1999–2007. J Antimicrob Chemother. 2008;62(Suppl 2):ii97-ii103.
17. Moxon ER. The carrier state: Haemophilus influenzae. J Antimicrob Chemother. 1986;18(Supplement A):17-24.
18. Nevzat Ü, Güney AK, Yanik K, Eroğlu C, Günaydin M. Comparison of in vitro susceptibility of Haemophilus influenzae strains to various antimicrobial drugs. J Exp Clin Med. 2013;30(2):165-9.
19. Özkul H, Özbek Ö, Çoban H, Gülay Z. Dokuz Eylül Üniversitesi Hastanesinde 2003-2006 yıllarında üretilen Haemophilus influenzae suşlarının antibiyotik duyarlılıkları. ANKEM Derg. 2007;21(2):86-90.
20. Pérez-Trallero E, García-De-La-Fuente C, GarcíaRey C, et al. Geographical and ecological analysis of resistance, coresistance, and coupled resistance to antimicrobials in respiratory pathogenic bacteria in Spain. Antimicrob Agents Chemother. 2005;49(5):1965-72.
21. Rashid H, Shoma S, Rahman M. Prevalence of β-lactamase positive ampicillin resistant H. influenzae from children of Bangladesh. J Infect Dis Epidemiol. 2016;2(2):1-5.
22. Sener B, Tunckanat F, Ulusoy S, et al. A survey of antibiotic resistance in Streptococcus pneumoniae and Haemophilus influenzae in Turkey, 2004 2005. J Antimicrob Chemother. 2007;60(3):587-93.
23. Torun M, Aksun E, Özcan N, Başaran G. Alt solunum yolu infeksiyonlarından izole edilen S. aureus kökenlerinin çeşitli antimikrobik maddelere duyarlılıkları. ANKEM Derg. 1998;12(2):102.
24. Torun M, Namal N, Demirci M, Bahar H. Nasopharyngeal carriage and antibiotic resistance of Haemophilus influenzae, Streptococcus pneumoniae and Moraxella catarrhalis in healthy school children in Turkey. Indian J Med Microbiol. 2009;27(1):86-8.
25. Torun MM, Namal N, Demirci M, Bahar H, Kocazeybek B. Pharyngeal carriage and antimicrobial resistance of Haemophilus influenzae in non-type-b-vaccinated healthy children attending day care centers in Turkey. Chemotherapy. 2007;53(2):114-7.
26. Tristram S, Jacobs MR, Appelbaum PC. Antimicrobial resistance in Haemophilus influenzae. Clin Microbiol Rev. 2007;20(2):368-89.
27. Tsang RS, Shuel M, Whyte K, et al. Antibiotic susceptibility and molecular analysis of invasive Haemophilus influenzae in Canada, 2007 to 2014. J Antimicrob Chemother. 2017;72(5):1314-9.
28. Wang H-J, Wang C-Q, Hua C-Z, et al. Antibiotic resistance profiles of Haemophilus influenzae isolates from children in 2016: a multicenter study in China. Can J Infect Dis Med Microbiol. 2019;2019.
29. Yamada S, Seyama S, Wajima T, et al. β-Lactamase-non-producing ampicillin-resistant Haemophilus influenzae is acquiring multidrug resistance. J Infect Public Health. 2019.