Investigation altered Penicillin Binding Protein 3 (PBP3) in Non-type Haemophilus influenza (NTHi) Isolated from Chronic Obstructive Pulmonary Disease (COPD) patients

Abstract

Non-typeable Haemophilus influenzae (NTHi) is commonly isolated from the lower airways of chronic obstructive pulmonary disease (COPD). Empirical antibiotic treatment is usually prescribed for eradicating the colonising bacteria and for subsequent prevention exacerbations. This empirical prescription of antibiotics has led to the emergence of resistant strains, particularly β-lactamase negative ampicillin resistance (BLNAR) strains, which can reduce susceptibility to penicillins and cephalosporins. Resistant phenotypes are sometimes incorrectly expressed presenting confused laboratory results, which increases difficulty in distinguishing them from other susceptible strains. This makes the genetic identification (molecular sequencing) is the most effective method for identifying mutations within these BLNAR strains. The present study examined the molecular mechanisms and susceptibility to β-lactams in BLNAR isolates from selected COPD patients. Isolates preliminarily identified as BLNARs by disc diffusion susceptibility and non β-lactamase production were examined for phenotypic minimum inhibitory concentrations (MIC) confirmation and genotypic confirmation by sequencing ftsI gene encoding Penicillin Binding Protein3 (PBP3). This PBP3 may contribute to reducing affinity to specific β-lactam antibiotics. Our results reveal that all samples (100%) were resistant to ampicillin (2 µg), amoxicillin (2 µg) and co-amoxiclav (2:1 µg) under EUCAST interpretive criteria. Of the four isolates sequenced, three isolates were identified as genetic BLNAR strains belonging to group II because of the substitution at Ans526Lys and the ampicillin MICs ranged between (0.5 and 1 µg/ml). One isolate, categorised as subgroup IIb low-BLNAR strain, was carrying Ans526Lys and Ala502Val substitutions near to the KTG Motif and had ampicillin MICs (2 µg/ml), which was higher than the remaining isolates. Although no high-BLNAR strains were found in this study, for the first time, we showed the prevalence of low-BLNARs in NTHi isolated from COPD patients raising concerns over prolonged empirical antibiotic prescription