Production of extended-spectrum β-lactamases and the potential indirect pathogenic role of Prevotella isolates from the cystic fibrosis respiratory microbiota

Abstract

Extended-spectrum β-lactamase (ESBL) production and the prevalence of the β-lactamase-encoding gene blaTEM were determined in Prevotella isolates (n=50) cultured from the respiratory tract of adults and young people with cystic fibrosis (CF). Time-kill studies were used to investigate the concept of passive antibiotic resistance and to ascertain whether a β-lactamase-positive Prevotella isolate can protect a recognised CF pathogen from the action of ceftazidime in vitro. The results indicated that approximately three-quarters (38/50; 76%) of Prevotella isolates produced ESBLs. Isolates positive for ESBL production had higher minimum inhibitory concentrations (MICs) of β-lactam antibiotics compared with isolates negative for production of ESBLs (P<0.001). The blaTEM gene was detected more frequently in CF Prevotella isolates from paediatric patients compared with isolates from adults (P=0.002), with sequence analysis demonstrating that 21/22 (95%) partial blaTEM genes detected were identical to blaTEM-116. Furthermore, a β-lactamase-positive Prevotella isolate protected Pseudomonas aeruginosa from the antimicrobial effects of ceftazidime (P=0.03). Prevotella isolated from the CF respiratory microbiota produce ESBLs and may influence the pathogenesis of chronic lung infection via indirect methods, including shielding recognised pathogens from the action of ceftazidime.

Keywords: Anaerobes; Antibiotic resistance; Cystic fibrosis; β-Lactams.

Fig. 1

Comparison of susceptibility (MICs) between ESBL-positive (n = 38) and ESBL-negative (n = 12) cystic fibrosis Prevotella isolates. In the box and whisker plot, the top and bottom boundaries of each box indicate the 75th and 25th quartile values, respectively, with the line inside the box representing the median (50th quartile). End of whiskers indicate the range. Any isolates recorded as having an MIC greater than the maximum (>256 mg/L) concentration on the Etest strip are shown as double the maximum concentration. MIC, minimum inhibitory concentration; ESBL, extended-spectrum β-lactamase.

Fig. 2

Prevalence of ESBL-producing Prevotella isolates from CF patients prescribed long-term antibiotics (flucloxacillin, azithromycin, tobramycin and colistin) compared with CF patients not prescribed chronic antibiotic treatments. ESBL, extended-spectrum β-lactamase; CF, cystic fibrosis.

Fig. 3

Comparison of ESBL production and detection of bla_TEM between CF adult (n = 25) and CF paediatric (n = 25) isolates. *P < 0.05. ESBL, extended-spectrum β-lactamase; CF, cystic fibrosis.

Fig. 4

Time–kill studies (mean ± standard deviation, n = 3) using ceftazidime at 64× the Pseudomonas aeruginosa minimum inhibitory concentration (MIC) (32 mg/L). A protective effect (antagonism) by Prevotella was defined as a ≥2 log₁₀ increase in viable count of P. aeruginosa at 48 h compared with that of the P. aeruginosa isolate alone. *P < 0.05.