Objectives: P. aeruginosa infection is a decisive turning point during the course of the disease in CF children. We investigated whether early colonization by P. aeruginosa could impact airway microbiota. Methods: 61 CF patients (mostly children) attending three CF centers were followed, and categorized at the inclusion according to the Leeds criteria as “free” (no culture of P. aeruginosa for at least the previous year) or “never” (P. aeruginosa had never been detected from sputum or cough swab). Clinical and biological data were collected at each sampling time. The majority of patients were homozygous or heterozygous for the F508del-CFTR mutation. The airway microbiotafrom 188 sputum samples was analyzed through 16S rRNA gene sequencing performed using the MiSeq Illumina technology and the MiSeq Reagent Kit V3. An original bioinformatics pipeline was written for this purpose (https://github.com/dridk/ mucobiome), based on Snakemake, VSearch, Cutadapt, Sickle and Greengene. The LEfSe (https://huttenhower.sph.harvard.edu/galaxy/) system for biomarkers discovery was applied to highlight potential genera specific of P. aeruginosa status. Results: Microbial alpha diversity in patients with P. aeruginosa infection ( patients “Free”) was found lower (p < 0.01) than in patients without any history of P. aeruginosa infection ( patients “Never”). LEfSe determined the bacterial genera most likely to explain differences between patients Never and Free, and thus most likely to be impacted by P. aeruginosa early colonization and/or anti-Pseudomonas antibiotic treatment. Conclusion: The present results may suggest that the diversity of the CF airway microbiota is directly negatively impacted by first acquisition of P. aeruginosa. The decrease in microbial diversity observed consecutively to P. aeruginosa early colonization may due to antibiotic regimen given to patients to eradicate P. aeruginosa. 170 Members of the cystic fibrosis lung microbiome influence the epithelial immune response to Pseudomonas aeruginosa C. Rigauts 1 , E. Vandeplassche 1 , T. Coenye 1 , A. Crabbé 1 . 1 Ghent University, Laboratory of Pharmaceutical Microbiology, Ghent, Belgium Pseudomonas aeruginosa is the most common pathogen colonizing the lungs of cystic fibrosis (CF) patients. Besides P. aeruginosa, a variety of other microorganisms is present in the CF lung, often as part of a polymicrobial biofilm. The impairment in microbial clearance causes a continuous stimulation of the immune system which is driven at least in part by P. aeruginosa, and eventually leads to irreversible lung damage. The role of other microbiome members in the excessive inflammation remains mostly unexplored. The aim of the present study was to evaluate the epithelial immune response induced by planktonic and sessile P. aeruginosa cells, and to investigate how this immune response is influenced by the presence of other CF lung microbiome members. The interleukin-8 (IL-8) production by organotypic three dimensional (3- D) lung epithelial cells was evaluated after infection with P. aeruginosa biofilms or planktonic cells by an IL-8 ELISA. Subsequently, we analyzed the IL-8 induction by P. aeruginosa PAO1 co-cultured with other bacteria commonly found in the CF lungs: Staphylococcus aureus, Streptococcus anginosus, Rothia mucilaginosa, Achromobacter xylosoxidans or Gemella haemolysans. Results show that epithelial IL-8 production is significantly lower after infection with sessile P. aeruginosa than after infection with planktonic cells. While the P. aeruginosa-induced IL-8 response was high, other members of the CF lung microbiome induced low or background IL-8 levels. Interestingly, the P. aeruginosa-induced IL-8 response was strongly reduced when co-cultured with R. mucilaginosa. Co-cultures of P. aeruginosa with the other tested bacteria did not show this effect. We conclude that P. aeruginosa planktonic cells induce a higher IL-8 response than sessile cells. Furthermore, R. mucilaginosa is able to significantly lower the P. aeruginosa-induced IL-8 response in lung epithelial cells. We are currently investigating the underlying mechanism for these findings. 171 A chicken and egg situation L. Pages-Monteiro 1 , C. Commun 1 , P. Reix 2 , J. Freney 1,3 , B. Cournoyer 1 , A. Doleans-Jordheim 1,3 . 1 UMR CNRS UCBLVetAgroSup 5557 (équipe 6), Lyon, France, 2 Service de pneumologie-allergologie pédiatrique, Centre de ressources et de compétence de la mucoviscidose pédiatrique, Hospices Civils de Lyon, Lyon, France, 3 Laboratoire de Bactériologie - Institut des Agents Infectieux, Hospices Civils de Lyon, Lyon, France Objectives: In the presence of Pseudomonas aeruginosa (Pa) in the lungs of patients with Cystic Fibrosis (CF), a modification in the ratio of bacteria is observed. This phenomenon, called dysmicrobism, is either the cause or the consequence of the implantation of Pa. Indeed, a restructuration of CF communities could lead to the pulmonary invasion by Pa, or this dysmicrobism could be explained by the invasive behavior of this bacterium. Faced with these two hypotheses, we realized a longitudinal study of the microbiota of CF patients before, during and after the Pa colonization of lungs. Methods: Over 3 years, we collected28 expectorations from 4 CF patients (7 expectorations/patient) and divided them in 3 groups: expectoration before (group FC1), during (group FC2) and after (group FC3) the first colonization by Pa. We analyzed the microbiota structure (richness and diversity) of each expectoration by fingerprint method (Ribosomal Intergenic Spacer Analysis-RISA). As controls, we included expectorations of 4 CF patients with no Pa identified during the 3 years (N group) as well as 4 patients chronically infected by Pa (CC group). The impact of the antibiotics as confounding factor was evaluated. Results: Differences between expectorations were observed over time for each patient. However, microbiota evolved less for the CC group which present the lowest diversity and richness. The diversity of the three FC groups was equivalent to the N group, but different from the CC group. The richness was equivalent between the N group and the FC1 and FC2 groups. However, it was different between the N group and the FC3 group. Conclusion: These results suggest that modifications of microbiota could be due to the implantation of Pa. Parameters such as richness and diversity cannot predict its colonization but might reflect a previous as well as a chronic infection with this pathogen. 172 Does the presence of “oral flora” in sputum cultures affect the clinical outcomes of people with cystic fibrosis lung disease? S.A.H. Higgi 1 , T.W.V. Daniels 2 . 1 University of Southampton, Southampton, United Kingdom; 2 Southampton Adult CF Centre, Southampton, United Kingdom Background: Lung disease is the leading cause of mortality in Cystic Fibrosis (CF). The presence of P. aeruginosa correlates with a more rapid decline in lung function. Laboratory studies show that P. aeruginosa’s behaviour is modulated by “avirulent” Oral Flora (OF) species via quorum sensing, allowing bacterial gene expression alteration to favourably suit the environment. This potentially leads to increased virulence as shown in Drosophila models where polymicrobial infection resulted in increased fly killing, with increased production of virulence factors enhancing the proteolytic ability of P. aeruginosa, further increasing lung function decline. The clinical relevance of these findings is unknown. Aims: To determine whether, if in those with CF and chronic P. aeruginosa, there is a difference in the rate of change of FEV1 over a three-year period between those with frequent OF and those with less frequent OF growth. Methods: Patient specific data was collected electronically. Subjects were divided into case – ≥75% OF alongside P. aeruginosa, and controls -<75% OF growth in P. aeruginosa positive sputum samples. Sixty-nine patients were recruited and divided into case (n = 40) and control (n = 29) groups for analysis via an independent samples t test, where the percentage change in FEV1 values was assessed. Results: No statistically significant difference was detected between case and control groups, p = 0.198. However, there was a greater mean decline in FEV1 values, mean = 5.45% ± 7.27 in the case group compared to the control group, mean = 3.14% ± 7.34 over three years, suggestive of clinical significance. Poster Sessions / Journalof Cystic Fibrosis 16S1 (2017) S63–S174 S110