Increased incidence of Mycoplasma pneumoniae infections and hospital admissions in the Netherlands, November to December 2023.

Mycoplasma pneumoniae is an important cause of pneumonia and extra-pulmonary manifestations. We observed a rise in admissions due to M. pneumoniae infections starting October 2023 in a regional hospital in the Netherlands and an increased incidence in national surveillance data. The incidence in the Netherlands has not been that high since 2011. The patients had a lower median age compared with 2019 and 2020 (28 vs 40 years). M. pneumoniae should be considered in patients with respiratory symptoms, especially children.

Less Is More: When to Repeat Antimicrobial Susceptibility Testing.

This study investigated the frequency of change of the antimicrobial susceptibility pattern when the same isolate was found in the same patient in various situations. We used laboratory data collected over a period of 8 years (January 2014 to December 2021) at the clinical microbiology laboratory of a tertiary hospital for Escherichia coli, Klebsiella pneumoniae, Enterobacter spp., Pseudomonas aeruginosa, and Staphylococcus aureus. Antimicrobial susceptibility tests (AST) were performed using Vitek 2 automated system. We determined essential agreement and categorical agreement, and introduced the new terms essential MIC increase and change from nonresistant to resistant to present changes in antimicrobial susceptibility over time. During the study period, 18,501 successive AST were included. The risk for S. aureus to be resistant to any antibiotic upon repeated culture was <10% during a follow-up of 30 days. For Enterobacterales, this risk was approximately 10% during a follow-up of 7 days. For P. aeruginosa, this risk was higher. The longer the follow-up period, the higher the risk that the bacteria would show phenotypic resistance. We also found that some drug-bug combinations were more likely to develop phenotypical resistance (i.e., E. coli/amoxicillin-clavulanic acid and E. coli/cefuroxime). A potential consequence of our finding is that if we regard a risk of resistance below 10% as acceptable, it may be feasible to omit follow-up AST within 7 days for the microorganisms investigated in this study. This approach saves money, time, and will reduce laboratory waste. Further studies are needed to determine whether these savings are in balance with the small possibility of treating patients with inadequate antibiotics.

16S rRNA-Based Microbiota Profiling Assists Conventional Culture Analysis of Airway Samples from Pediatric Cystic Fibrosis Patients.

16S-based sequencing provides broader information on the respiratory microbial community than conventional culturing. However, it (often) lacks species- and strain-level information. To overcome this issue, we used 16S rRNA-based sequencing results from 246 nasopharyngeal samples obtained from 20 infants with cystic fibrosis (CF) and 43 healthy infants, which were all 0 to 6 months old, and compared them to both standard (blind) diagnostic culturing and a 16S-sequencing-informed "targeted" reculturing approach. Using routine culturing, we almost uniquely detected Moraxella catarrhalis, Staphylococcus aureus, and Haemophilus influenzae (42%, 38%, and 33% of samples, respectively). Using the targeted reculturing approach, we were able to reculture 47% of the top-5 operational taxonomical units (OTUs) in the sequencing profiles. In total, we identified 60 species from 30 genera with a median of 3 species per sample (range, 1 to 8). We also identified up to 10 species per identified genus. The success of reculturing the top-5 genera present from the sequencing profile depended on the genus. In the case of Corynebacterium being in the top 5, we recultured them in 79% of samples, whereas for Staphylococcus, this value was only 25%. The success of reculturing was also correlated with the relative abundance of those genera in the corresponding sequencing profile. In conclusion, revisiting samples using 16S-based sequencing profiles to guide a targeted culturing approach led to the detection of more potential pathogens per sample than conventional culturing and may therefore be useful in the identification and, consequently, treatment of bacteria considered relevant for the deterioration or exacerbation of disease in patients like those with CF. IMPORTANCE Early and effective treatment of pulmonary infections in cystic fibrosis is vital to prevent chronic lung damage. Although microbial diagnostics and treatment decisions are still based on conventional culture methods, research is gradually focusing more on microbiome and metagenomic-based approaches. This study compared the results of both methods and proposed a way to combine the best of both worlds. Many species can relatively easily be recultured based on the 16S-based sequencing profile, and it provides more in-depth information about the microbial composition of a sample than that obtained through routine (blind) diagnostic culturing. Still, well-known pathogens can be missed by both routine diagnostic culture methods as well as by targeted reculture methods, sometimes even when they are highly abundant, which may be a consequence of either sample storage conditions or antibiotic treatment at the time of sampling.

Bacteria in Extracorporeal Membrane Oxygenation Circuit Clots of a Patient With Persistent Bacteremia: A Case Report.

A neonate with pulmonary hypertension was supported with extracorporeal membrane oxygenation (ECMO). During ECMO support, the patient developed Enterococcus faecalis bacteremia, treated with targeted antibiotics. Despite the maximum dose of antibiotics, routine blood cultures remained positive throughout the ECMO treatment. A circuit change was performed due to buildup of thrombotic material and disseminated intravascular coagulation (DIC) inside the circuit. Thrombus formation was more extensive in the first than the second circuit. Gram-positive diplococci were present in all initial circuit clots and gram-positive masses surrounded by fibrin were found inside thrombi of the second circuit. Scanning electron microscopy (SEM) revealed a dense fibrin network with embedded red blood cells and bacteria in the first circuit. In the second circuit, SEM analysis revealed scattered micro thrombi. Polymerase chain reaction for identification of bacteria in the thrombus of the first circuit showed the same bacteria as found in blood cultures and did not achieve a sufficient signal in the second circuit. This case report shows that bacteria can nestle in thrombi of an ECMO circuit and that there is a rationale for a circuit change in a patient with persistent positive blood cultures and DIC.

Evidence of cat-to-human transmission of Staphylococcus felis.

Introduction. Staphylococcus felis is a coagulase-negative staphylococcal species that is commonly isolated from healthy cats. Like other commensal staphylococci, S. felis can cause opportunistic infections, e.g. otitis externa, skin and urinary tract infections, in cats.Gap Statement. Several studies have reported within-household transmission between humans and pets and human infections caused by coagulase-positive staphylococci. However, human infections with coagulase-negative staphylococci of zoonotic origin are relatively rare.Methodology. Culture of a surgical site infection in a 58-year-old woman who underwent a laminectomy revealed dominant growth of S. felis. The three cats owned by the patient were sampled to investigate potential within-household transmission. S. felis isolates were sequenced to investigate the relatedness of the isolates and to look for virulence factors and host specific genes.Results. All cats were colonized with S. felis. Comparative genomics of the isolates showed that each cat was colonized with a distinct genotype. The patient's isolate clustered with isolates of one of the cats. Sequence analysis of the studied isolates together with 29 publicly available S. felis genomes detected putative virulence factors that can be crucial in potential interspecies transmission.Conclusion. The current case is the first reported human infection caused by S. felis and highlights the zoonotic potential of this bacterial species. Evidence of cat-to-human transmission was shown by comparative genomics of isolates from the patient with isolates of her cats.

Confirmatory testing of Neisseria gonorrhoeae in a sexual health clinic: implications for epidemiology and treatment policy.

OBJECTIVES: European guidelines advise the use of dual nucleic acid amplification tests (NAAT) in order to minimise the inappropriate diagnosis of Neisseria gonorrhoeae (Ng) in urogenital samples from low prevalence areas and in extragenital specimens. In this cross-sectional study, we investigated the effect of confirmatory testing and confirmation policy on the Ng-positivity in a population visiting the sexual health clinic in Rotterdam, the Netherlands. METHODS: Apart from urogenital testing, extragenital (oropharyngeal/anorectal) testing was performed for men who have sex with men (MSM) and according to sexual exposure for women and heterosexual men. Ng detection using NAAT was performed using BD Viper and for confirmatory testing BD MAX. Sexual transmitted infection consultation data were merged with diagnostic data from August 2015 through May 2016. RESULTS: In women (n=4175), oral testing was performed in 84% and 22% were tested anally. In MSM (n=1828), these percentages were 97% and 96%, respectively. Heterosexual men (n=3089) were tested urogenitally. After confirmatory testing, oropharyngeal positivity rates decreased from 7.3% (95% CI 6.5 to 8.2) to 1.5% (95% CI 1.1 to 1.8) in women and from 13.9% (95% CI 12.3 to 15.5) to 5.4% (95% CI 4.3 to 6.4) in MSM. Anorectal positivity rates decreased from 2.6% (95% CI 1.6 to 3.7) to 1.8% (95% CI 0.9 to 2.6) in women and from 9.3% (95% CI 7.9 to 10.7) to 7.2% (95% CI 6.0 to 8.5) in MSM. Urogenital Ng-positivity rate ranged between 3.0% and 4.4% and after confirmation between 2.3% and 3.9%. When confirming oropharyngeal samples, Ng-positivity was 3.8% in women, 3.0% in heterosexual men and 12.5% in MSM. Additional confirmation of urogenital and anorectal samples led to 3.0% Ng positivity in women, 2.7% in heterosexual men and 11.4% in MSM. CONCLUSIONS: Confirmation of urogenital and anorectal samples reduced the Ng-positivity rates, especially for women. However, as there is no gold standard for the confirmation of Ng infection, the dilemma within public health settings is to choose between two evils: missing diagnoses or overtreatment. In view of the large decrease in oropharyngeal positivity, confirmation Ng-positivity in oropharyngeal samples remains essential to avoid unnecessary treatment.

Characterization of clinical Ralstonia strains and their taxonomic position.

To improve understanding of the role of Ralstonia in cystic fibrosis (CF), whole genomes of 18 strains from clinical samples were sequenced using Illumina technology. Sequences were analysed by core genome Multi-Locus Sequence Typing, Average Nucleotide Identity based on BLAST (ANIb), RAST annotation, and by ResFinder. Phylogenetic analysis was performed for the 16S rRNA gene, and the OXA-22 and OXA-60 ß-lactamase families. The minimal inhibitory concentrations (MICs) were determined using broth microdilution. ANIb data for the 18 isolates and 54 strains from GenBank, supported by phylogenetic analysis, showed that 8 groups of clusters (A-H), as well as subgroups that should be considered as species or subspecies. Groups A-C contain strains previously identified as Ralstonia solanacearum and Ralstonia pseudosolanacearum. We propose that group A is a novel species. Group B and C are Ralstonia syzygii, Ralstonia solanacearum, respectively. Group D is composed of Ralstonia mannitolilytica and Group E of Ralstonia pickettii. Group F and G should be considered novel species. Group H strains belong to R. insidiosa. OXA-22 and OXA-60 family ß-lactamases were encoded by all strains. Co-trimoxazole generally showed high activity with low MICs (≤1 mg/l) as did ciprofloxacin (≤0.12 mg/l). MICs against the other antibiotics were more variable, but generally high. RAST annotation revealed limited differences between the strains, and virulence factors were not identified. The taxonomy of the genus Ralstonia is in need of revision, but sequencing additional isolates is needed. Antibiotic resistance levels are high. Annotation did not identify potential virulence factors.

Pre-processing tissue specimens with a tissue homogenizer: clinical and microbiological evaluation.

BACKGROUND: Tissues are valuable specimens in diagnostic microbiology because they are often obtained by invasive methods, and effort should thus be taken to maximize microbiological yield. The objective of this study was to evaluate the added value of using tissue pre-processing (tissue homogenizer instrument gentleMACS Dissociator) in detecting microorganisms responsible for infections. METHODS: We included 104 randomly collected tissue samples, 41 (39.4 %) bones and 63 (60.6 %) soft tissues, many of those (42/104 (40.4 %)) were of periprosthetic origins. We compared the agreement between pre-processing tissues using tissue homogenizer with routine microbiology diagnostic procedure, and we calculated the performance of these methods when clinical infections were used as reference standard. RESULTS: There was no significant difference between the two methods (McNemar test, p = 0.3). Among the positive culture using both methods (n = 62), 61 (98.4 %) showed at least one similar microorganism. Exactly similar microorganisms were found in 42/62 (67.7 %) of the samples. From the included tissues, 55/ 104 (52.9 %) were deemed as infected. We found that the sensitivity of homogenized tissue procedure was lower (83.6 %) than when tissue was processed using tissue homogenizer (89.1 %). Sub-analysis on periprosthetic tissues and soft or bone tissues showed comparable results. CONCLUSIONS: The added value of GentleMACS Dissociator tissue homogenizer is limited in comparison to routine tissue processing.

An Update on Eight "New" Antibiotics against Multidrug-Resistant Gram-Negative Bacteria.

Infections in the ICU are often caused by Gram-negative bacteria. When these microorganisms are resistant to third-generation cephalosporines (due to extended-spectrum (ESBL) or AmpC beta-lactamases) or to carbapenems (for example carbapenem producing Enterobacteriales (CPE)), the treatment options become limited. In the last six years, fortunately, there have been new antibiotics approved by the U.S. Food and Drug Administration (FDA) with predominant activities against Gram-negative bacteria. We aimed to review these antibiotics: plazomicin, eravacycline, temocillin, cefiderocol, ceftazidime/avibactam, ceftolozane/tazobactam, meropenem/vaborbactam, and imipenem/relebactam. Temocillin is an antibiotic that was only approved in Belgium and the UK several decades ago. We reviewed the in vitro activities of these new antibiotics, especially against ESBL and CPE microorganisms, potential side effects, and clinical studies in complicated urinary tract infections (cUTI), intra-abdominal infections (cIAI), and hospital-acquired pneumonia/ventilator-associatedpneumonia (HAP/VAP). All of these new antibiotics are active against ESBL, and almost all of them are active against CPE caused by KPC beta-lactamase, but only some of them are active against CPE due to MBL or OXA beta-lactamases. At present, all of these new antibiotics are approved by the U.S. Food and Drug Administration for cUTI (except eravacycline) and most of them for cIAI (eravacycline, ceftazidime/avibactam, ceftolozane/tazobactam, and imipenem/relebactam) and for HAP or VAP (cefiderocol, ceftazidime/avibactam, ceftolozane/tazobactam, and imipenem/relebactam).

Frequency of Positive Aspergillus Tests in COVID-19 Patients in Comparison to Other Patients with Pulmonary Infections Admitted to the Intensive Care Unit.

The aim of this study was to describe the frequency of positive Aspergillus tests in COVID-19 patients and investigate the association between COVID-19 and a positive Aspergillus test result. We compared the proportion of positive Aspergillus tests in COVID-19 patients admitted to the intensive care unit (ICU) for >24 h with two control groups: patients with community-acquired pneumonia with (i) a PCR-confirmed influenza infection (considered a positive control since the link between influenza and invasive aspergillosis has been established) and (ii) Streptococcus pneumoniae pneumonia (in whom positive Aspergillus tests are mostly considered as colonization). During the study period, 92 COVID-19 patients (mean [standard deviation] age, 62 [14] years; 76.1% males), 48 influenza patients (55 [14]; 56.2% males), and 65 pneumococcal pneumonia patients (58 [15], 63,1% males) were identified. Any positive Aspergillus test from any respiratory sample was found in 10.9% of the COVID-19 patients, 6.2% of the patients with pneumococcal pneumonia, and 22.9% of those infected with influenza. A positive culture or PCR or galactomannan test on bronchoalveolar lavage (BAL) fluid only was found in 5.4% of COVID-19 patients, which was lower than in patients with influenza (18.8%) and comparable to that in the pneumococcal pneumonia group (4.6%). Using logistic regression analysis, the odds ratio (OR) (95% confidence interval) for a positive Aspergillus test on BAL fluid for COVID-19 patients was 1.2 (0.3 to 5.1; P = 0.8) compared to the pneumococcal pneumonia group, while it was 0.2 (0.1 to 0.8; P = 0.02) compared to the influenza group. This difference remained significant when corrected for age and sex. In conclusion, in COVID-19 patients, the prevalence of a positive Aspergillus test was comparable to that in patients admitted for pneumococcal pneumonia but substantially lower than what we observed in patients with influenza.

The accuracy of four commercial broth microdilution tests in the determination of the minimum inhibitory concentration of colistin.

Colistin is considered as one of the last-resort antibiotics and reliable antimicrobial susceptibility testing is therefore crucial. The reference standard for AST according to EUCAST and CLSI is broth microdilution (BMD). However, BMD is labor intensive to perform. Commercial antimicrobial susceptibility tests derived from BMD method are available. We investigated the performance of four different commercial tests: Sensititre™, SensiTest™ Colistin, Micronaut MIC Strip Colistin and UMIC Colistin using 70 clinical isolates (half of them was deemed by VITEK2 as resistant), including isolates from cystic fibrosis patients and mcr-1 bearing isolates. We used two reference standards: BMD and composite MIC as determined by all four tests. Sensititre™ had essential agreement (EA, defined as minimum inhibitory concentration within ± 1 dilution) of 87% and 89% compared to BMD and composite reference standard, respectively. For SensiTest™, the EA's were 93% and 90%. For UMIC, 87% and 90%, and for Micronaut, 83% and 84%. All four tests demonstrated categorical agreement (CA) above 90%. CA for SensiTest™ and Micronaut was both 96%, UMIC 94%, and Sensititre™ 93%. All tests were reproducible as tested in two quality control isolates. In conclusion, in clinical isolates from a large referral center, the four commercial tests for determination of colistin minimum inhibitory concentrations showed acceptable performance.

Development of the gut microbiota in early life: The impact of cystic fibrosis and antibiotic treatment.

OBJECTIVES: Patients with Cystic Fibrosis (CF) suffer from pancreatic insufficiency, lipid malabsorption and gastrointestinal complaints, next to progressive pulmonary disease. Altered mucosal homoeostasis due to malfunctioning chloride channels results in an adapted microbial composition of the gastrointestinal and the respiratory tract. Additionally, antibiotic treatment has the potential to distort resident microbial communities dramatically. This study aims to investigate early life development of the gut microbial community composition of children with CF compared to healthy infants and to study the independent effects of antibiotics taking into account other clinical and lifestyle factors. STUDY DESIGN: Faecal samples from 20 infants with CF and 45 healthy infants were collected regularly during the first 18 months of life and microbial composition was determined using 16S rRNA based sequencing. RESULTS: We observed significant differences in the overall microbiota composition between infants with CF and healthy infants (p<0.001). Akkermansia and Anaerostipes were significantly more abundant in control infants, whereas Streptococci and E. coli were significantly more abundant in infants with CF, also after correction for several clinical factors (p<0.05). Antibiotic use in infants with CF was associated with a lower alpha diversity, a reduced abundance of Bifidobacterium and Bacteroides, and a higher abundance of Enterococcus. CONCLUSION: Microbial development of the gut is different in infants with CF compared to healthy infants from the first months of life on, and further deviates over time, in part as a result of antibiotic treatment. The resulting dysbiosis may have significant functional consequences for the microbial ecosystem in CF patients.

Successful High-Dosage Monotherapy of Tigecycline in a Multidrug-Resistant Klebsiella pneumoniae Pneumonia-Septicemia Model in Rats.

Background: Recent scientific reports on the use of high dose tigecycline monotherapy as a "drug of last resort" warrant further research into the use of this regimen for the treatment of severe multidrug-resistant, Gram-negative bacterial infections. In the current study, the therapeutic efficacy of tigecycline monotherapy was investigated and compared to meropenem monotherapy in a newly developed rat model of fatal lobar pneumonia-septicemia. Methods: A Klebsiella pneumoniae producing extended-spectrum ß-lactamase (ESBL) and an isogenic variant producing K. pneumoniae carbapenemase (KPC) were used in the study. Both strains were tested for their in vitro antibiotic susceptibility and used to induce pneumonia-septicemia in rats, which was characterized using disease progression parameters. Therapy with tigecycline or meropenem was initiated at the moment that rats suffered from progressive infection and was administered 12-hourly over 10 days. The pharmacokinetics of meropenem were determined in infected rats. Results: In rats with ESBL pneumonia-septicemia, the minimum dosage of meropenem achieving survival of all rats was 25 mg/kg/day. However, in rats with KPC pneumonia-septicemia, this meropenem dosage was unsuccessful. In contrast, all rats with KPC pneumonia-septicemia were successfully cured by administration of high-dose tigecycline monotherapy of 25 mg/kg/day (i.e., the minimum tigecycline dosage achieving 100% survival of rats with ESBL pneumonia-septicemia in a previous study). Conclusions: The current study supports recent literature recommending high-dose tigecycline as a last resort regimen for the treatment of severe multidrug-resistant bacterial infections. The use of ESBL- and KPC-producing K. pneumoniae strains in the current rat model of pneumonia-septicemia enables further investigation, helping provide supporting data for follow-up clinical trials in patients suffering from severe multidrug-resistant bacterial respiratory infections.

Low gonorrhoea antimicrobial resistance and culture positivity rates in general practice: a pilot study.

OBJECTIVE: In the Netherlands, the Gonococcal Resistance to Antimicrobials Surveillance (GRAS) programme is carried out at Centres for Sexual Health (CSH), which provide care for sexual high-risk populations. However, half of gonorrhoea infections are diagnosed in general practice (GP). We performed a pilot study to explore expanding GRAS to GPs using laboratory-based surveillance. Additionally, antimicrobial resistance patterns of GP and CSH patients were compared. METHODS: Three laboratories from different regions were included, which all perform gonorrhoea diagnostics for GPs and used ESwab for patient sampling. Additional culturing for all GP patients with gonorrhoea took place from February to July 2018. After positive PCR-nucleic acid amplification test, residual ESwab material was used for culture. In positive cultures, susceptibility testing was performed for azithromycin, ciprofloxacin, cefotaxime and ceftriaxone using Etest. RESULTS: During the study period, 484 samples were put in culture. 16.5% of cultures were positive (n=80). Antimicrobial resistance levels were low, with 2.6% resistance to azithromycin, 21.5% to ciprofloxacin and 0.0% to cefotaxime and ceftriaxone. Resistance levels in CSH GRAS data (first half of 2018) were 19.2% for azithromycin, 31.5% for ciprofloxacin, 1.9% for cefotaxime and 0.0% for ceftriaxone. CONCLUSIONS: Culture positivity rates for GP patients were low, probably due to long transportation times and awaiting PCR test results before attempting culture. Positivity rates might be improved by making changes in sampling and/or transportation methods, but that would require involvement of GPs and patients instead of keeping the surveillance lab based. Resistance levels appeared to be lower at GPs than at the CSH, indicating that resistance might emerge first in more high-risk populations. It is important to consider all potentially relevant patient populations when establishing a gonococcal antimicrobial resistance surveillance programme. However, based on the findings from this study the current GRAS programme will not be extended to GPs.

Antimicrobial susceptibility of non-fermenting Gram-negative pathogens isolated from cystic fibrosis patients.

Non-fermenting Gram-negative bacteria (NFGNB) are increasingly cultured in respiratory samples from cystic fibrosis (CF) patients. This study determined the antimicrobial susceptibility of clinical CF respiratory isolates from distinct geographical regions. A total of 286 isolates (106 Stenotrophomonas maltophilia, 100 Burkholderia spp., 59 Achromobacter spp., 12 Pandoraea spp., 9 Ralstonia spp.) from the Netherlands, Northern Ireland, Spain, USA and Australia were tested. MIC50/90 values and susceptibility categorisation were determined. Trimethoprim/sulfamethoxazole (SXT) was the most active compound for all micro-organisms (MIC50, 0.12-4 mg/L; MIC90, 1-16 mg/L). For S. maltophilia, 47% and 62% of isolates were susceptible to SXT according to CLSI and EUCAST breakpoints, respectively. Ceftazidime presented lower susceptibility (35%; MIC50, 32 mg/L; MIC90, 256 mg/L). MIC90 values for tobramycin and colistin were >128 mg/L and >16 mg/L, respectively. Regarding Burkholderia, 72%, 56% and 44% were susceptible to SXT, ceftazidime and meropenem, respectively. For both ceftazidime and meropenem, MIC50 and MIC90 values were within the intermediate or resistant category. The most active antibiotics for Achromobacter spp. were SXT (MIC50, 0.5 mg/L; MIC90, 8 mg/L) and imipenem (MIC50, 2 mg/L; MIC90, 8 mg/L). SXT, imipenem and ciprofloxacin were active against 12 Pandoraea spp. (MIC50, 0.12-4 mg/L; MIC90, 1-8 mg/L). Ciprofloxacin (MIC50, 4 mg/L) and SXT (MIC50, 1 mg/L) were the only active antibiotics for Ralstonia spp. There were no statistically significant differences in susceptibility rates between countries. NFGNB other than Pseudomonas aeruginosa are potential pathogens in CF. SXT was demonstrated to be the most active compound against these isolates.

Patient-specific modelling of regional tobramycin concentration levels in airways of patients with cystic fibrosis: can we dose once daily?

BACKGROUND: Inhaled tobramycin is important in the treatment of Pseudomonas aeruginosa (Pa) infections in cystic fibrosis (CF). However, despite its use it fails to attenuate the clinical progression of CF lung disease. The bactericidal efficacy of tobramycin is known to be concentration-dependent and hence changing the dosing regimen from a twice-daily (q12h) inhalation to a once-daily (q24h) inhaled double dose could improve treatment outcomes. OBJECTIVES: To predict local concentrations of nebulized tobramycin in the airways of patients with CF, delivered with the small airway-targeting Akita® system or standard PARI-LC® Plus system, with different inspiratory flow profiles. METHODS: Computational fluid dynamic (CFD) methods were applied to patient-specific airway models reconstructed from chest CT scans. The following q12h and q24h dosing regimens were evaluated: Akita® (150 and 300 mg) and PARI-LC® Plus (300 and 600 mg). Site-specific concentrations were calculated. RESULTS: Twelve CT scans from patients aged 12-17 years (median = 15.7) were selected. Small airway concentrations were 762-2999 mg/L for the q12h dosing regimen and 1523-5997 mg/L for the q24h dosing regimen, well above the MIC for WT Pa strains. Importantly, the q24h regimen appeared to be more suitable than the q12h regimen against more resistant Pa strains and the inhibitory effects of sputum on tobramycin activity. CONCLUSIONS: CFD modelling showed that high concentrations of inhaled tobramycin are indeed delivered to the airways, with the Akita® system being twice as efficient as the PARI-LC® system. Ultimately, the q24h dosing regimen appears more effective against subpopulations with high MICs (i.e. more resistant strains).

Concordance between upper and lower airway microbiota in infants with cystic fibrosis.

Nasopharyngeal and oropharyngeal samples are commonly used to direct therapy for lower respiratory tract infections in non-expectorating infants with cystic fibrosis (CF).We aimed to investigate the concordance between the bacterial community compositions of 25 sets of nasopharyngeal, oropharyngeal and bronchoalveolar lavage (BAL) samples from 17 infants with CF aged ∼5 months (n=13) and ∼12 months (n=12) using conventional culturing and 16S-rRNA sequencing.Clustering analyses demonstrated that BAL microbiota profiles were in general characterised by a mixture of oral and nasopharyngeal bacteria, including commensals like Streptococcus, Neisseria, Veillonella and Rothia spp. and potential pathogens like Staphylococcus aureus, Haemophilus influenzae and Moraxella spp. Within each individual, however, the degree of concordance differed between microbiota of both upper respiratory tract niches and the corresponding BAL.The inconsistent intra-individual concordance between microbiota of the upper and lower respiratory niches suggests that the lungs of infants with CF may have their own microbiome that seems seeded by, but is not identical to, the upper respiratory tract microbiome.

Development of the Nasopharyngeal Microbiota in Infants with Cystic Fibrosis.

RATIONALE: Cystic fibrosis (CF) is characterized by early structural lung disease caused by pulmonary infections. The nasopharynx of infants is a major ecological reservoir of potential respiratory pathogens. OBJECTIVES: To investigate the development of nasopharyngeal microbiota profiles in infants with CF compared with those of healthy control subjects during the first 6 months of life. METHODS: We conducted a prospective cohort study, from the time of diagnosis onward, in which we collected questionnaires and 324 nasopharynx samples from 20 infants with CF and 45 age-matched healthy control subjects. Microbiota profiles were characterized by 16S ribosomal RNA-based sequencing. MEASUREMENTS AND MAIN RESULTS: We observed significant differences in microbial community composition (P < 0.0002 by permutational multivariate analysis of variance) and development between groups. In infants with CF, early Staphylococcus aureus and, to a lesser extent, Corynebacterium spp. and Moraxella spp. dominance were followed by a switch to Streptococcus mitis predominance after 3 months of age. In control subjects, Moraxella spp. enrichment occurred throughout the first 6 months of life. In a multivariate analysis, S. aureus, S. mitis, Corynebacterium accolens, and bacilli were significantly more abundant in infants with CF, whereas Moraxella spp., Corynebacterium pseudodiphtericum and Corynebacterium propinquum and Haemophilus influenzae were significantly more abundant in control subjects, after correction for age, antibiotic use, and respiratory symptoms. Antibiotic use was independently associated with increased colonization of gram-negative bacteria such as Burkholderia spp. and members of the Enterobacteriaceae bacteria family and reduced colonization of potential beneficial commensals. CONCLUSIONS: From diagnosis onward, we observed distinct patterns of nasopharyngeal microbiota development in infants with CF under 6 months of age compared with control subjects and a marked effect of antibiotic therapy leading toward a gram-negative microbial composition.

Patient-specific modeling of regional antibiotic concentration levels in airways of patients with cystic fibrosis: are we dosing high enough?

BACKGROUND: Pseudomonas aeruginosa (Pa) infection is an important contributor to the progression of cystic fibrosis (CF) lung disease. The cornerstone treatment for Pa infection is the use of inhaled antibiotics. However, there is substantial lung disease heterogeneity within and between patients that likely impacts deposition patterns of inhaled antibiotics. Therefore, this may result in airways below the minimal inhibitory concentration of the inhaled agent. Very little is known about antibiotic concentrations in small airways, in particular the effect of structural lung abnormalities. We therefore aimed to develop a patient-specific airway model to predict concentrations of inhaled antibiotics and to study the impact of structural lung changes and breathing profile on local concentrations in airways of patients with CF. METHODS: In- and expiratory CT-scans of children with CF (5-17 years) were scored (CF-CT score), segmented and reconstructed into 3D airway models. Computational fluid dynamic (CFD) simulations were performed on 40 airway models to predict local Aztreonam lysine for inhalation (AZLI) concentrations. Patient-specific lobar flow distribution and nebulization of 75 mg AZLI through a digital Pari eFlow model with mass median aerodynamic diameter range were used at the inlet of the airway model. AZLI concentrations for central and small airways were computed for different breathing patterns and airway surface liquid thicknesses. RESULTS: In most simulated conditions, concentrations in both central and small airways were well above the minimal inhibitory concentration. However, small airways in more diseased lobes were likely to receive suboptimal AZLI. Structural lung disease and increased tidal volumes, respiratory rates and larger particle sizes greatly reduced small airway concentrations. CONCLUSIONS: CFD modeling showed that concentrations of inhaled antibiotic delivered to the small airways are highly patient specific and vary throughout the bronchial tree. These results suggest that anti-Pa treatment of especially the small airways can be improved.