Achromobacter species are increasingly being detected in cystic fibrosis (CF) patients, with an unclear epidemiology and impact. We studied a cohort of patients attending a Canadian adult CF clinic who had positive sputum cultures for Achromobacter species in the period from 1984 to 2013. Infection was categorized as transient or persistent (Ն50% positive cultures for 1 year). Those with persistent infection were matched 2:1 with age-, sex-, and time-matched controls without a history of Achromobacter infection, and mixed-effects models were used to assess pulmonary exacerbation (PEx) frequency and lung function decline. Isolates from a biobank were retrospectively assessed, identified to the species level by nrdA sequencing, and genotyped using pulsed-field gel electrophoresis (PFGE). Thirty-four patients (11% of those in our clinic), with a median age of 24 years (interquartile range [IQR], 20.3 to 29.8 years), developed Achromobacter infection. Ten patients (29%) developed persistent infection. Persistence did not denote permanence, as most patients ultimately cleared infection, often after years. Patients were more likely to experience PEx at incident isolation than at prior or subsequent visits (odds ratio [OR], 2.7 [95% confidence interval {CI}, 1.2 to 6.7]; P ϭ 0.03). Following persistent infection, there was no difference in annual lung function decline (Ϫ1.08% [95% CI, Ϫ2.73 to 0.57%] versus Ϫ2.74% [95% CI, Ϫ4.02 to 1.46%]; P ϭ 0.12) or the odds of PEx (OR, 1.21 [95% CI, 0.45 to 3.28]; P ϭ 0.70). Differential virulence among Achromobacter species was not observed, and no cases of transmission occurred. We demonstrated that incident Achromobacter infection was associated with a greater risk of PEx; however, neither transient nor chronic infection was associated with a worsened long-term prognosis. Large, multicenter studies are needed to clarify the clinical impact, natural history, and transmissibility of Achromobacter.

Background Analysis of “emerging” pathogens in cystic fibrosis (CF) lung disease has focused on unique pathogens that are rare in other human diseases or are drug resistant. Escherichia coli is recovered in the sputum of up to 25% of patients with CF, yet little is known about the epidemiology or clinical impact of infection. Methods We studied patients attending a Canadian adult CF clinic who had positive sputum cultures for E coli from 1978 to 2016. Infection was categorized as transient or persistent (≥3 positive sputum cultures, spanning >6 months). Those with persistent infection were matched 2:1 with age, sex, and time-period controls without history of E coli infection, and mixed-effects models were used to assess pulmonary exacerbation (PEx) frequency, lung function decline, hospitalization, and intravenous antibiotic days. Results Forty-five patients (12.3%) had E coli recovered from sputum samples between 1978 and 2016, and 18 patients (40%) developed persistent infection. Nine patients (24%) had PEx at incident infection, and increased bioburden was predictive of exacerbation (P = .03). Risk factors for persistent infection included lower nutritional status (P < .001) and lower lung function (P = .009), but chronic infection with Pseudomonas aeruginosa was protective. There was no difference in annual lung function decline, need for hospitalization or intravenous antibiotics, or risk of PEx in patients with persistent infection. Conclusions Persistent E coli infection was frequent and was more common in CF patients with low nutritional status and lung function. However, this does not predict clinical decline. Multicenter studies would allow better characterization of the epidemiology and clinical impact of E coli infection.

Escherichia coli is frequently isolated from the respiratory secretions of cystic fibrosis (CF) patients yet is not considered a classical CF pathogen. Accordingly, little is known about the natural history of this organism in the CF airways, as well as the potential for patientto-patient transmission. Patients attending the Calgary Adult CF Clinic (CACFC) between January 1983 and December 2016 with at least one E. coli-positive sputum culture were identified by retrospective review. Annual E. coli isolates from the CACFC biobank from each patient were typed by pulsed-field gel electrophoresis (PFGE) and isolates belonging to shared pulsotypes were sequenced. Single nucleotide polymorphism (SNP) and phylogenetic analysis were used to investigate the natural history of E. coli infection and identify potential transmission events. Forty-five patients with E. coli-positive sputum cultures were identified. Most patients had a single infection episode with a single pulsotype, while replacement of an initial pulsotype with a second was observed in three patients. Twenty-four had E. coli recovered from their sputum more than once and 18 patients had persistent infections (E. coli carriage >6 months with ≥3 positive cultures). Shared pulsotypes corresponded to known extraintestinal pathogenic E. coli strains: ST-131, ST-73, and ST-1193. Phylogenetic relationships and SNP distances among isolates within shared pulsotypes were consistent with independent acquisition of E. coli by individual patients. Most recent common ancestor date estimates of isolates between patients were inconsistent with patient-to-patient transmission. E. coli infection in CF is a dynamic process that appears to be characterized by independent acquisition within our patient population and carriage of unique sets of strains over time by individual patients.

Images contain a wealth of information that is often under analyzed in biological studies. Developmental models of vascular disease are a powerful way to quantify developmentally regulated vessel phenotypes to identify the roots of the disease process. We present vessel Metrics, a software tool specifically designed to analyze developmental vascular microscopy images that will expedite the analysis of vascular images and provide consistency between research groups. We developed a segmentation algorithm that robustly quantifies different image types, developmental stages, organisms, and disease models at a similar accuracy level to a human observer. We validate the algorithm on confocal, lightsheet, and two photon microscopy data in zebrafish. The tool accurately segments data taken by multiple scientists on varying microscopes. We validate vascular parameters such as vessel density, network length, and diameter, across developmental stages, genetic mutations, and drug treatments, and show a favorable comparison to other freely available software tools. Vessel Metrics reduces the time to analyze experimental results, improves repeatability within and between institutions, and expands the percentage of a given vascular network analyzable in experiments.

Vascular malformations develop when growth pathway signaling goes awry in the endothelial cells lining blood vessels. Arteriovenous malformations (AVMs) arise where arteries and veins abnormally connect in patients with loss of RASA1, a Ras GTPase activating protein, and, as we show here, in zebrafish rasa1 mutants. Mutant fish develop massively enlarged vessels at the connection between artery and vein in the tail vascular plexus. These AVMs progressively enlarge and become filled with slow-flowing blood and have a greater drop in pulsatility from the artery to the vein. Expression of the flow responsive transcription factor klf2a is diminished in rasa1 mutants, suggesting changes in flow velocity and pattern contribute to the progression of vessel malformations. Migration of endothelial cells is not affected in rasa1 mutants, nor is cell death or proliferation. Early developmental artery-vein patterning is also normal in rasa1 mutants, but we find that MEK/ERK signaling is ectopically activated in the vein as compared to high arterial activation seen in wildtype animals. MEK/ERK signaling inhibition prevents AVM development of rasa1 mutants, demonstrating venous MEK/ERK drives the initiation of rasa1 AVMs. Thus, rasa1 mutants show overactivation of MEK/ERK signaling causes AVM formation, altered blood flow and downstream flow responsive signaling.

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BackgroundDespite a growing interest in emerging pathogens in CF, research has largely overlooked commonplace organisms. Escherichia coli has been reported in up to 50% of CF respiratory samples, yet little is known about its clinical impact. We sought to investigate outcomes of E. coli infection in CF.MethodsWe undertook a retrospective cohort study of patients (≥18 years) attending a Canadian CF clinic between 1978 and 2016 with at least one E. coli positive sputum culture. Infection was classified as transient (≥1 isolate) or persistent (≥3 isolates over a period ≥6 months). Clinical and demographic data were collected from patient charts 2 years pre- and post-incident infection. For each patient with persistent infection, we collected data on two age (±3 years), sex, and time-matched control patients for comparison. Outcomes sought included risk of pulmonary exacerbation (PEx), lung function decline (FEV1), antibiotic days, and progression to transplant or death. Susceptibility testing was performed as per CLSI standards.ResultsA total of 45 (12.3%) patients (median age 23.5 (IQR 20.0–34.8), 52% male) cultured E. coli in their sputum at least once. At incident infection, 24% had PEx but this was not increased relative to prior visits (RR 0.9, P = 1.00). Of the cohort, 18 (40%) developed persistent infection. Persistent infection developed in patients with lower nutritional scores (BMI) (−2.6 kg/m2, P < 0.001) and lung function (FEV1%; 57.2 vs. 74.2, P = 0.02). Compared with matched controls, those with persistent infection had no increase in mean annual lung function decline (difference −1.06%/year, P = 0.24), odds of PEx (OR 1.4, P = 0.26), or mean annual hospital IV days (difference: 0.31 days, 95% CI −4.97 to 5.59 days, P = 0.91). Five patients underwent lung transplantation and three died at 5-year follow-up, but this did not differ between transient and persistent infection (P = 0.63 and P = 0.25, respectively). TMP-SMX resistance (P = 0.05), but not ESBL production in incident isolates, was predictive of persistence (P = 0.56).ConclusionIn this Canadian CF cohort, E. coli infection was common and occurred more frequently in patients with compromised nutrition and lung function. Persistent infection with E. coli did not portend worse clinical outcomes. Multi-centre studies are merited to further understand the epidemiology and clinical impact of E. coli infection.Disclosures All authors: No reported disclosures.

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