Epidemiology and impact of methicillin-sensitive Staphylococcus aureus with ß-lactam antibiotic inoculum effects in adults with cystic fibrosis.

Staphylococcus aureus is the most prevalent cystic fibrosis (CF) pathogen. Several phenotypes are associated with worsened CF clinical outcomes including methicillin-resistance and small-colony-variants. The inoculum effect (IE) is characterized by reduced ß-lactam susceptibility when assessed at high inoculum. The IE associates with worse outcomes in bacteremia and other high-density infections, and may therefore be relevant to CF. The prevalence of IE amongst a CF cohort (age ≥18 years), followed from 2013 to 2016, was investigated. Yearly methicillin-sensitive S. aureus (MSSA) isolates were screened at standard (5 × 105 CFU/mL) and high (5 × 107 CFU/mL) inoculum against narrow-spectrum anti-Staphylococcal ß-lactams and those with anti-pseudomonal activity common to CF. A ≥ 4-fold increase in minimum inhibitory concentration between standard and high inoculum defined IE. Isolates underwent blaZ sequencing and genotyping and were compared against published genomes. Fifty-six percent (99/177) of individuals had MSSA infection. MSSA was observed at ≥105 CFU/mL in 44.8% of entry sputum samples. The prevalence of the IE was 25.0%-cefazolin; 13.5%-cloxacillin; 0%-meropenem; 1.0%-cefepime; 5.2%-ceftazidime; and 34.4%-piperacillin-tazobactam amongst baseline MSSA isolates assessed. blaZ A associated with cefazolin IE (P = 0.0011), whereas blaZ C associated with piperacillin-tazobactam IE (P < 0.0001). Baseline demographics did not reveal specific risk factors for IE-associated infections, nor were long-term outcomes different. Herein, we observed the IE in CF-derived MSSA disproportionally for cefazolin and piperacillin-tazobactam and this phenotype strongly associated with underlying blaZ genotype. The confirmation of CF being a high density infection, and the identification of high prevalence of MSSA with IE in CF supports the need for prospective pulmonary exacerbation treatment studies to understand the impact of this phenotype.

Clinical Outcomes Associated With Escherichia coli Infections in Adults With Cystic Fibrosis: A Cohort Study.

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.

A longitudinal characterization of the Non-Cystic Fibrosis Bronchiectasis airway microbiome.

A diverse microbiota exists within the airways of individuals with non-cystic fibrosis bronchiectasis (nCFB). How the lung microbiome evolves over time, and whether changes within the microbiome correlate with future disease progression is not yet known. We assessed the microbial community structure of 133 serial sputa and subsequent disease course of 29 nCFB patients collected over a span of 4-16 years using 16S rRNA paired-end sequencing. Interestingly, no significant shifts in the microbial community of individuals were observed during extended follow-up suggesting the microbiome remains relatively stable over prolonged periods. Samples that were Pseudomonas aeruginosa culture positive displayed markedly different microbial community structures compared to those that were positive for Haemophilus influenzae. Importantly, patients with sputum of lower microbial community diversity were more likely to experience subsequent lung function decline as defined by annual change in ≥-1 FEV1% predicted. Shannon diversity values <1 were more prevalent in patients with FEV1 decline (P = 0.002). However, the relative abundance of particular core microbiota constituents did not associate with risk of decline. Here we present data confirming that the microbiome of nCFB individuals is generally stable, and that microbiome-based measurements may have a prognostic role as biomarkers for nCFB.

Prevalence and Outcomes of Achromobacter Species Infections in Adults with Cystic Fibrosis: a North American Cohort Study.

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 (≥50% 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 = 0.03). Following persistent infection, there was no difference in annual lung function decline (-1.08% [95% CI, -2.73 to 0.57%] versus -2.74% [95% CI, -4.02 to 1.46%]; P = 0.12) or the odds of PEx (OR, 1.21 [95% CI, 0.45 to 3.28]; P = 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.

Development and Validation of a PCR Assay To Detect the Prairie Epidemic Strain of Pseudomonas aeruginosa from Patients with Cystic Fibrosis.

The monitoring of epidemic Pseudomonas aeruginosa is important for cystic fibrosis (CF) infection control. The prairie epidemic strain (PES) is common in western Canadian CF clinics. Using whole-genome sequencing, we identified a novel genomic island and developed a PCR assay for PES. Against a collection of 186 P. aeruginosa isolates, the assay had 98% sensitivity and 100% specificity.

Infection control knowledge, beliefs and behaviours amongst cystic fibrosis patients with epidemic Pseudomonas aeruginosa.

BACKGROUND: Epidemic P. aeruginosa (ePA) infections are common in cystic fibrosis (CF) and have been associated with accelerated clinical decline. Factors associated with ePA are unclear, and evidence based infection control interventions are lacking. METHODS: We prospectively collect all bacterial pathogens from adult CF patients. We performed PA strain typing on retrospectively collected enrollment samples and recent isolates to identify patients infected with ePA. All patients attending our clinic were approached to complete a survey on infection control knowledge, beliefs and exposures. We analyzed responses of those with ePA relative to the entire cohort without ePA as well as those infected with unique strains of P. aeruginosa to assess for risk factors for ePA and differences in infection control knowledge, beliefs or behaviours. RESULTS: Of 144 participants, 30 patients had ePA (two Liverpool epidemic strain, 28 Prairie epidemic strain), 83 % of which had established infection prior to transition to the adult clinic. Risk of concomitant infecting pathogens was no different between groups although, Staphylococcus aureus and non-tuberculous mycobacteria were less common in those with ePA. Patients with ePA were more likely to have attended CF-camp and have a history of CF fundraising. Patients with ePA did not differ with respect to beliefs regarding pathogens or transmission risk, except they believed indirect contact posed little risk. Furthermore, patients with ePA were more likely to continue to associate with others with CF despite extensive counselling. Use of peer-peer online networking was minimal in both groups. CONCLUSION: Infections with ePA are closely linked to past exposures, now routinely discouraged. As socialization is the greatest risk factor for ePA, infection control strategies for ePA must focus on discouraging face-to-face interactions amongst CF patients. As peer support remains a desire amongst patients, investment in technologies and strategies that enable indirect communication and support are required.

Incidence, risk factors, and outcomes for Enterococcus spp. blood stream infections: a population-based study.

BACKGROUND: Enterococci are a clinically significant cause of bloodstream infections (BSI), particularly in the nosocomial setting. The purpose of this study was to characterize the incidence, risk factors for acquisition, microbiological characteristics and mortality of enterococcal BSI within the well-defined population of a large Canadian health region. METHODS: Surveillance for all episodes of enterococcal BSI occurring among residents of the Calgary Health Zone (population 1.2 million) between 2000 and 2008 was conducted using an electronic surveillance system. Clinical features, microbiology, and outcomes were obtained. RESULTS: A total of 710 incident episodes of enterococcal BSI were identified for an annual incidence of 6.9 episodes per 100,000; the incidences of E. faecalis and E. faecium BSI were 4.5, and 1.6 per 100,000, respectively. Enterococcus faecalis infections were associated with a urinary focus, genitourinary malignancy, and abnormal genitourinary anatomy. E. faecium infections were associated with a gastrointestinal focus. Resistance to ampicillin, vancomycin and ciprofloxacin was higher in E. faecium infection. The overall case fatality rate was 22.8%, and was higher for E. faecium infection. CONCLUSIONS: This is the second population-based study to assess the risk factors for enterococcal BSI and compare the characteristics of infection with E. faecalis and E. faecium. Results suggest that BSI with E. faecalis and E. faecium should be regarded as two clinically different entities with unique sets of risk factors and microbiologic characteristics.

Granulomatous PJP presenting as a solitary lung nodule in an immune competent female.

Pneumocystis jiroveci pneumonia (PJP) opportunistically targets immunosuppressed patients, most notably those with advanced HIV/AIDS. Radiologically, PJP typically appears as bilateral diffuse pulmonary infiltrates. Herein an unusual case of an immunocompetent woman developing granulomatous PJP in the absence of evident risk factors is described. PJP may be an under-recognized cause of pulmonary nodules in immune competent individuals.

Population-based assessment of the incidence, risk factors, and outcomes of anaerobic bloodstream infections.

BACKGROUND: Anaerobes are a relatively uncommon but important cause of bloodstream infection. However, their epidemiology has not been well defined in non-selected populations. We sought to describe the incidence of, risk factors for, and outcomes associated with anaerobic bacteremia. METHODS: Population-based surveillance for bacteremia with anaerobic microorganisms was conducted in the Calgary area (population 1.2 million) during the period from 2000 to 2008. RESULTS: A total of 904 incident cases were identified, for an overall population incidence of 8.7 per 100,000 per year; 231 (26 %) were nosocomial, 300 (33 %) were healthcare-associated community-onset, and 373 (41 %) were community-acquired. Elderly males were at the greatest risk. The most common pathogens identified were: Bacteroides fragilis group (3.6 per 100,000), Clostridium (non-perfringens) spp. (1.1 per 100,000), Peptostreptococcus spp. (0.9 per 100,000), and Clostridium perfringens (0.7 per 100,000). Non-susceptibility to metronidazole was 2 %, to clindamycin 17 %, and to penicillin 42 %. Relative to the general population, risk factors for anaerobic bloodstream infection included: male sex, increasing age, a prior diagnosis of cancer, chronic liver disease, heart disease, diabetes mellitus, stroke, inflammatory bowel disease, human immunodeficiency virus (HIV) infection, chronic obstructive pulmonary disease (COPD), and/or hemodialysis-dependent chronic renal failure (HDCRF). The 30-day mortality was 20 %. Increasing age, nosocomial acquisition, presence of malignancy, and several other co-morbid illnesses were independently associated with an increased risk of death. CONCLUSION: Anaerobic bloodstream infection is responsible for a significant burden of disease in general populations. The data herein establish the extent to which anaerobes contribute to morbidity and subsequent mortality. This information is key in developing preventative, empiric treatment and research priorities.

Pyrosequencing reveals the complex polymicrobial nature of invasive pyogenic infections: microbial constituents of empyema, liver abscess, and intracerebral abscess.

The polymicrobial nature of invasive pyogenic infections may be underestimated by routine culture practices, due to the fastidious nature of many organisms and the loss of viability during transport or from prior antibacterials. Pyrosequencing was performed on brain and liver abscesses and pleural fluid and compared to routine culture data. Forty-seven invasive pyogenic infection samples from 44 patients [6 intracerebral abscess (ICA), 21 pyogenic liver abscess (PLA), and 18 pleural fluid (PF) samples] were assayed. Pyrosequencing identified an etiologic microorganism in 100 % of samples versus 45 % by culture, p <0.01. Pyrosequencing was also more likely than traditional cultures to classify infections as polymicrobial, 91 % versus 17 %, p <0.001. The median number of genera identified by pyrosequencing compared to culture was 1 [interquartile range (IQR) 1-3] versus 0 (IQR 0-1) for ICA, 7 (IQR 1-15) versus 1 (IQR 0-1) for PLA, and 15 (IQR 9-19) versus 0 (IQR 0-1) for PF. Where organisms were cultured, they typically represented the numerically dominant species identified by pyrosequencing. Complex microbial communities are involved in invasive pyogenic infection of the lung, liver, and brain. Defining the polymicrobial nature of invasive pyogenic infections is the first step towards appreciating the clinical and diagnostic implications of these complex communities.

Newer antibacterial agents and their potential role in cystic fibrosis pulmonary exacerbation management.

Pulmonary exacerbations in cystic fibrosis (CF) are frequent events and account for a substantial proportion of the burden of morbidity and mortality in this disease. Antibacterial therapies to treat pulmonary exacerbations are instituted empirically and are individualized based on both patient factors (severity of exacerbation, frequency of exacerbation, recent courses of anti-infectives) and pathogen factors (previously isolated pathogens and in vitro predicted susceptibilities). However, the epidemiology of pathogens infecting CF airways is changing, with increased incidence of methicillin-resistant Staphylococcus aureus (MRSA), drug-resistant Pseudomonas aeruginosa and other Gram-negative non-fermenters such as Stenotrophomonas maltophilia and Achromobacter xylosoxidans. Accordingly, a great need for new and novel agents for the management of acute exacerbations in CF exists. While several antibiotics have recently been approved or are close to approval for clinical use, frequently their emphasis has been for Gram-positive, and specifically MRSA-related, disease. Despite this, these agents may have a role in CF-related exacerbations. This article reviews the spectrum of activity, pharmacokinetics and clinical and theoretical evidence for the use of newer agents including tigecycline, doripenem and ceftobiprole in the management of CF pulmonary exacerbations. Appropriate use of these agents in CF will require detailed CF-specific pharmacokinetic and pharmacodynamic data.

Population-based study of the epidemiology and the risk factors for Pseudomonas aeruginosa bloodstream infection.

BACKGROUND: Detailed population-based data on the epidemiology of Pseudomonas aeruginosa bloodstream infections are sparse. We sought to describe the incidence rate, risk factors, and outcomes associated with P. aeruginosa bacteremia in a large Canadian health region. PATIENTS AND METHODS: A retrospective population-based surveillance for P. aeruginosa bacteremia was conducted in the Calgary Health Region (CHR, population:approx. 1.2 million) during the period from 2000 to 2006. RESULTS: A total of 284 incident cases of P. aeruginosa bacteremia were identified in CHR residents, corresponding to an annual incidence rate of 3.6/100,000.Nosocomial acquisition accounted for 45% of cases,healthcare-associated community onset for 34% of cases,and community-acquired (CA) cases for 21%. Relative to the general population, risk factors for blood stream infection included male sex, increasing age, hemodialysis,solid organ transplant, diagnosis of cancer, heart disease, HIV infection, diabetes mellitus, and/or chronic obstructive airway disease (COPD). Overall mortality was 29%. Factors associated with mortality in univariate analysis included pulmonary focus of infection and co-morbidities, including chronic liver disease, substance abuse, heart disease, COPD, and cancer, and increased with the burden of co-morbidities. Despite those patients with CA disease having fewer co-morbidities,they had a significantly higher mortality rate than either healthcare-associated cases or nosocomial cases(RR 1.88, p = 0.05). CONCLUSIONS: This study documents that P. aeruginosa bacteremic disease is responsible for a significant burden of illness in general populations and identifies those groups at increased risk of infection and subsequent mortality. This information can be used to identify those individuals likely to benefit from empiric anti-pseudomonal therapies.

Population-based laboratory surveillance for Serratia species isolates in a large Canadian health region.

A population-based laboratory surveillance was conducted during a six-year period to define the incidence, demographic risk factors for acquisition, and anti-microbial susceptibilities of Serratia species isolates. A total of 715 incident Serratia species isolates were identified for an annual incidence of 10.8 per 100,000 residents; bacteremic disease occurred in 0.9 per 100,000 residents annually. The incidence increased with advancing age and males were at the highest risk. Ninety-two percent of the isolates were Serratia marcescens, and the majority (65%) of incident Serratia species isolates were of community onset. Ninety-five percent of isolates were susceptible to ciprofloxacin, 98% to gentamicin, 98% to trimethoprim/sulfamethoxazole, and >99% to imipenem. No yearly increase in resistance was observed. Serratia species isolation is most commonly of community onset and older patients and males are at increased risk. Despite reports of increasing resistance among Serratia species, the incidence in our region remains at a low stable rate.

Population-based laboratory surveillance for tribe Proteeae isolates in a large Canadian health region.

The tribe Proteeae comprises the genera Proteus, Morganella and Providencia. Few studies have specifically investigated the epidemiology of infections caused by the Proteeae, and none has been conducted in a large non-selected population. The present study was a population-based laboratory surveillance in the Calgary Health Region (population 1.2 million), Canada during 2000-2005 that aimed to define the incidence, demographical risk-factors for acquisition and antimicrobial susceptibilities of Proteeae isolates. In total, 5047 patients were identified from whom Proteeae isolates were obtained (an annual incidence of 75.9/100 000), with females and the elderly being at highest risk. Incidence rates were 64.8, 7.7 and 3.4/100,000/year for the genera Proteus, Morganella and Providencia, respectively. Overall, 85% of infections were community-onset, and the overall rate of bacteraemic disease was 2.0/100,000. Compared with other species, Proteus mirabilis occurred at a much higher frequency, especially among females, and was less likely to be isolated from hospital-onset infections or to be part of a polymicrobial infection. Among isolates from community-onset infections, Providencia spp. were less likely to be from outpatients and more likely to be from nursing home residents. There were low overall rates of resistance to ciprofloxacin (4%) and gentamicin (5%), with Prot. mirabilis generally being the most susceptible. Members of the Proteeae were isolated frequently in both the community and hospital settings, but were infrequent causes of invasive disease. The occurrence, demographical risk-factors and microbiology of Proteeae isolates varied according to the individual species.

Treatment of infections caused by metallo-beta-lactamase-producing Pseudomonas aeruginosa in the Calgary Health Region.

This study reviewed 56 patients with significant metallo-beta-lactamase (MBL)-producing Pseudomonas aeruginosa infections between May 2002 and March 2004 to identify features associated with mortality. Immunosuppression (p 0.002), bacteraemia (p 0.08) and inadequate antimicrobial therapy (p <0.001) were associated with death. Among those patients treated with adequate therapy, the use of multiple drug treatment regimens (two or three active agents) was associated with a non-significant two-fold increase in survival (p 0.45). Further prospective studies are warranted to determine the optimal treatment of MBL-producing P. aeruginosa infections.

Overnight storage of autologous stem cell apheresis products before cryopreservation does not adversely impact early or long-term engraftment following transplantation.

To reduce costs and avoid inconvenient overtime work, our institution changed policy in September 2000 so that autologous stem cell apheresis products were stored overnight before cryopreservation rather than immediately processed. This retrospective review was conducted to evaluate the possible impact of this policy change on hematopoietic engraftment following autologous stem cell transplantation (ASCT). In total, 229 consecutive lymphoma patients who underwent a single, unpurged ASCT in Calgary between January 1995 and November 2003 were evaluated. Of these patients, 131 patients' autografts underwent immediate processing and cryopreservation before September 2000, and 98 patients' autografts underwent next-day cryopreservation after overnight storage following this date. Results of univariate and multivariate analyses demonstrated no adverse effect of overnight storage before cryopreservation on the number of days to initial engraftment of platelets or neutrophils, on the proportion of patients with low blood counts 6 months post-ASCT, or on lymphoma relapse rates or overall survival post-ASCT. These data suggest that overnight storage of the autograft before cryopreservation does not adversely affect graft viability or influence long-term disease status, and support the continued use of overnight storage of stem cells before cryopreservation as a convenient, cost reduction measure.

Pseudomonas aeruginosa GacA, a factor in multihost virulence, is also essential for biofilm formation.

We have investigated a potential role for GacA, the response regulator of the GacA/GacS two-component regulatory system, in Pseudomonas aeruginosa biofilm formation. When gacA was disrupted in strain PA14, a 10-fold reduction in biofilm formation capacity resulted relative to wild-type PA14. However, no significant difference was observed in the planktonic growth rate of PA14 gacA(-). Providing gacA in trans on the multicopy vector pUCP-gacA abrogated the biofilm formation defect. Scanning electron microscopy of biofilms formed by PA14 gacA(-) revealed diffuse clusters of cells that failed to aggregate into microcolonies, implying a deficit in biofilm development or surface translocation. Motility assays revealed no decrease in PA14 gacA(-) twitching or swimming abilities, indicating that the defect in biofilm formation is independent of flagellar-mediated attachment and solid surface translocation by pili. Autoinducer and alginate bioassays were performed similarly, and no difference in production levels was observed, indicating that this is not merely an upstream effect on either quorum sensing or alginate production. Antibiotic susceptibility profiling demonstrated that PA14 gacA(-) biofilms have moderately decreased resistance to a range of antibiotics relative to PA14 wild type. This study establishes GacA as a new and independent regulatory element in P. aeruginosa biofilm formation.

Multidrug efflux pumps: expression patterns and contribution to antibiotic resistance in Pseudomonas aeruginosa biofilms.

Pseudomonas aeruginosa biofilms are intrinsically resistant to antimicrobial chemotherapies. At present, very little is known about the physiological changes that occur during the transition from the planktonic to biofilm mode of growth. The resistance of P. aeruginosa biofilms to numerous antimicrobial agents that are substrates subject to active efflux from planktonic cells suggests that efflux pumps may substantially contribute to the innate resistance of biofilms. In this study, we investigated the expression of genes associated with two multidrug resistance (MDR) efflux pumps, MexAB-OprM and MexCD-OprJ, throughout the course of biofilm development. Using fusions to gfp, we were able to analyze spatial and temporal expression of mexA and mexC in the developing biofilm. Remarkably, expression of mexAB-oprM and mexCD-oprJ was not upregulated but rather decreased over time in the developing biofilm. Northern blot analysis confirmed that these pumps were not hyperexpressed in the biofilm. Furthermore, spatial differences in mexAB-oprM and mexCD-oprJ expression were observed, with maximal activity occurring at the biofilm substratum. Using a series of MDR mutants, we assessed the contribution of the MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY efflux pumps to P. aeruginosa biofilm resistance. These analyses led to the surprising discovery that the four characterized efflux pumps do not play a role in the antibiotic-resistant phenotype of P. aeruginosa biofilms.