The study aimed to boost the efficiency of clearance of the respiratory pathobionts in children with CF based on the study of the spectrum of colonizers, their sensitivity to antimicrobial agents, probiotic microorganisms, and environmental conditions.
For the first time using the expanded cultural method the respiratory microbiome of children with CF living in Dnipro region and the local chemotherapeutic susceptibility of pathobionts was described in the study. The repiratory microbiome in children with CF was characterized by a suppression of normal microbiota and an excessive diversity of pathogens that are usually not characteristic of the general population. The diversity of respiratory tract commensals was poor in children with CF, the species composition included representatives of the generas Corynebacterium, Streptococcus, and Aerococcus. The qualitative and quantitative characteristics of commensal microbiota changed negatively with age, the aggregated proportion of coagulase-negative Staphylococcus spp., Streptococcus, Aerococcus spp., Corynebacterium spp. Lactobacillus spp. and Neisseria spp. occupied 61.1% of isolates in children under 2 years of age, 48.0% - from 6 to 12 years, and 35.3% - over 15 years (p < 0.05). The prevalence of the mentioned commensals also negatively correlated with age. So, under 2 years old, the prevalence of commensals was 111.1%,
2 to 6 years – 64.2%, 6 to 12 years – 51.4%, above 15 years – 43.8% (p<0.001). The main pathogens in childhood included Staphylococcus aureus, Haemophilus influenzae, Pseudomonas aeruginosa and Aspergillus fumigatus.
The pathognomic changes of the main pathobionts associated with CF were described in the study. The share of small colony varianst (SCVs) was 11.1%. Auxotrophic cultures were isolated after treatment with aminoglycosides and/or sulfonamides and in case of P. aeruginosa infection (p˂0.001). Only 20.4% (95%CI 53.7-79.6) of S. aureus isolates were sensitive to all β-lactams. The share of methicillin-resistant S. aureus (MRSA) was 5.3%. Auxotrophic S. aureus were more resistant to β-lactams and gentamicin (p<0.05).
The share of P. aeruginosa mucoids was 27.2% (n = 26), SCVs – 5.83% (n = 6). The chemotherapeutic sensitivity profile of the pathogen was variable: only 62.1% (95% CI 52.1-71.5) of the isolates were susceptible to all penicillins, while the sensitivity with increased exposure to ceftazidime and cefepime was 82.5% and 72.8%, respectively. Mucoid isolates were more resistant than non-mucoid to penicillins (p˂0.001) and cephalosporins (p˂0.05).
In our study we described adhesive and biofilm-forming potential of the main pathobionts associated with CF. S. aureus P. aeruginosa had significant adhesive potential and biofilm-forming ability. There was a direct correlation between the adhesive potential of S. aureus and P. aeruginosa and their biofilm-forming activity, rs = 0.84 (p˂0.001) and rs = 0.95 (p˂0.001), respectively.
In our study, the activity of autoprobiotics and nonsymbiotic bacteria against pathobionts associated with CF was described for the first time. The symbionts Aerococcus viridans, the museum strain A. viridans 167 and Bacillus clausii had a pronounced antagonistic potential against opportunists and pathogens isolated from the respiratory tract of children with CF. When comparing the antagonistic potential of the symbionts of A. viridans and the museum strain A. viridans 167, it was established that the endogenous microbiota is more active against S. aureus and E. coli.
At the next stage of our work, for the first time we evaluated the levels of antimicrobial peptides (AMPs) hBD-2 and hCAP-18/LL-37 in the sputum of children with CF and their correlations with microbial profile of lungs. The concentration of cathelicidin hCAP-18/LL-37 in the CF-sputum correlated with the colonization of
P. aeruginosa between the levels of determined AMPs weren`t found (rs = 0.63; p < 0.001).
We have confirmed the importance of microbiological monitoring in cystic fibrosis as one of the key aspects of medical follow up. The results obtained in this work highlight the main microbiological features of pathobionts commonly associated with cystic fibrosis. Considering the results of the study and basing on the world best practices, for the first time a guideline for working with biological material from patients with cystic fibrosis at all stages of microbiological research were developed in Ukraine to improve the effectiveness of surveillance and quality of care. In addition, data on the local profile of antimicrobial susceptibility was obtained and is constantly considered by clinicians when prescribing antimicrobial therapy. The dynamics of non-specific immunity depending on the microbial profile of the respiratory tract and the density and type of cellular infiltration can be used by clinicians in planning anti-inflammatory therapy for children with cystic fibrosis
