Heterogeneity in Staphylococcus aureus Bacteraemia Clinical Trials Complicates Interpretation of Findings.

We systematically evaluated randomized-controlled trials (RCTs) for Staphylococcus aureus bacteremia (SAB). There was intertrial heterogeneity in cohort characteristics, including bacteremia source, complicated SAB, and comorbidities. Reporting of cohort characteristics was itself variable, including bacteremia source and illness severity. Selection bias was introduced by exclusion criteria relating to comorbidities, illness severity, infection types, and source control. Mortality was lower in RCT control arms compared with observational cohorts. Differences in outcome definitions impedes meta-analysis. These issues complicate the interpretation and application of SAB RCT results. The value of these trials should be maximized by a standardized approach to recruitment, definitions, and reporting.

A Subgroup of Patients With Hospital-acquired Pneumonia Do Not Require Broad-spectrum Gram-negative Antimicrobial Coverage.

Among 200 patients developing hospital-acquired pneumonia (HAP) outside the intensive care unit, 61% were treated empirically without broad-spectrum Gram-negative coverage, with clinical cure in 69.7%. Lower disease severity markers (systemic inflammatory response syndrome, hypoxia, tachypnoea, neutrophilia) and the absence of diabetes mellitus and prior doxycycline treatment (but not the time to HAP onset) identified patients not requiring broad-spectrum Gram-negative coverage.

Comprehensive Molecular Testing for Respiratory Pathogens in Community-Acquired Pneumonia.

BACKGROUND: The frequent lack of a microbiological diagnosis in community-acquired pneumonia (CAP) impairs pathogen-directed antimicrobial therapy. This study assessed the use of comprehensive multibacterial, multiviral molecular testing, including quantification, in adults hospitalized with CAP. METHODS: Clinical and laboratory data were collected for 323 adults with radiologically-confirmed CAP admitted to 2 UK tertiary care hospitals. Sputum (96%) or endotracheal aspirate (4%) specimens were cultured as per routine practice and also tested with fast multiplex real-time polymerase-chain reaction (PCR) assays for 26 respiratory bacteria and viruses. Bacterial loads were also calculated for 8 bacterial pathogens. Appropriate pathogen-directed therapy was retrospectively assessed using national guidelines adapted for local antimicrobial susceptibility patterns. RESULTS: Comprehensive molecular testing of single lower respiratory tract (LRT) specimens achieved pathogen detection in 87% of CAP patients compared with 39% with culture-based methods. Haemophilus influenzae and Streptococcus pneumoniae were the main agents detected, along with a wide variety of typical and atypical pathogens. Viruses were present in 30% of cases; 82% of these were codetections with bacteria. Most (85%) patients had received antimicrobials in the 72 hours before admission. Of these, 78% had a bacterial pathogen detected by PCR but only 32% were culture-positive (P < .0001). Molecular testing had the potential to enable de-escalation in number and/or spectrum of antimicrobials in 77% of patients. CONCLUSIONS: Comprehensive molecular testing significantly improves pathogen detection in CAP, particularly in antimicrobial-exposed patients, and requires only a single LRT specimen. It also has the potential to enable early de-escalation from broad-spectrum empirical antimicrobials to pathogen-directed therapy.

Pulmonary Mycobacterium marinum infection: 'fish tank granuloma' of the lung.

A 65-year-old man presented with a six-month history of lethargy, weight loss and dry cough. He had a background of mild chronic obstructive pulmonary disease. Chest radiograph showed new right upper lobe cavitary opacification. Sputum cultures were acid-fast bacilli smear positive and yielded Mycobacterium marinum - a non-tuberculous mycobacterium (NTM) often found in aquatic environments and rarely associated with respiratory disease. The suspected source was silent aspiration of contaminated water, likely due to his initiating the siphon of his fish-tank by mouth. He completed a one-year course of rifampicin, ethambutol and clarithromycin, with negative repeat sputum mycobacteria cultures and radiological improvement. This case report demonstrates a successful approach to investigation and further management of Mycobacterium marinum pulmonary disease - a rare condition, particularly in immunocompetent individuals, with limited treatment guidelines.

Diagnostic accuracy of pulmonary host inflammatory mediators in the exclusion of ventilator-acquired pneumonia.

BACKGROUND: Excessive use of empirical antibiotics is common in critically ill patients. Rapid biomarker-based exclusion of infection may improve antibiotic stewardship in ventilator-acquired pneumonia (VAP). However, successful validation of the usefulness of potential markers in this setting is exceptionally rare. OBJECTIVES: We sought to validate the capacity for specific host inflammatory mediators to exclude pneumonia in patients with suspected VAP. METHODS: A prospective, multicentre, validation study of patients with suspected VAP was conducted in 12 intensive care units. VAP was confirmed following bronchoscopy by culture of a potential pathogen in bronchoalveolar lavage fluid (BALF) at >10(4) colony forming units per millilitre (cfu/mL). Interleukin-1 beta (IL-1ß), IL-8, matrix metalloproteinase-8 (MMP-8), MMP-9 and human neutrophil elastase (HNE) were quantified in BALF. Diagnostic utility was determined for biomarkers individually and in combination. RESULTS: Paired BALF culture and biomarker results were available for 150 patients. 53 patients (35%) had VAP and 97 (65%) patients formed the non-VAP group. All biomarkers were significantly higher in the VAP group (p<0.001). The area under the receiver operator characteristic curve for IL-1ß was 0.81; IL-8, 0.74; MMP-8, 0.76; MMP-9, 0.79 and HNE, 0.78. A combination of IL-1ß and IL-8, at the optimal cut-point, excluded VAP with a sensitivity of 100%, a specificity of 44.3% and a post-test probability of 0% (95% CI 0% to 9.2%). CONCLUSIONS: Low BALF IL-1ß in combination with IL-8 confidently excludes VAP and could form a rapid biomarker-based rule-out test, with the potential to improve antibiotic stewardship.

Non-tuberculous mycobacteria: a retrospective review of Scottish isolates from 2000 to 2010.

There is growing recognition of the clinical importance of non-tuberculous mycobacteria (NTM), a group of versatile opportunistic bacterial pathogens. We describe the characteristics of NTM isolates in Scotland over an 11-year period using data held by the Scottish Mycobacteria Reference Laboratory. American Thoracic Society microbiological criteria were used to evaluate the clinical significance of isolates. Data presented include analysis of trends across time, species/body site associations, gender and age differences, geographical variations and the association between cystic fibrosis and Mycobacterium abscessus. We emphasise the need for standardised reporting criteria for NTM isolates to ensure optimal surveillance of NTM disease.

C5a-mediated neutrophil dysfunction is RhoA-dependent and predicts infection in critically ill patients.

Critically ill patients are at heightened risk for nosocomial infections. The anaphylatoxin C5a impairs phagocytosis by neutrophils. However, the mechanisms by which this occurs and the relevance for acquisition of nosocomial infection remain undetermined. We aimed to characterize mechanisms by which C5a inhibits phagocytosis in vitro and in critically ill patients, and to define the relationship between C5a-mediated dysfunction and acquisition of nosocomial infection. In healthy human neutrophils, C5a significantly inhibited RhoA activation, preventing actin polymerization and phagocytosis. RhoA inhibition was mediated by PI3Kδ. The effects on RhoA, actin, and phagocytosis were fully reversed by GM-CSF. Parallel observations were made in neutrophils from critically ill patients, that is, impaired phagocytosis was associated with inhibition of RhoA and actin polymerization, and reversed by GM-CSF. Among a cohort of 60 critically ill patients, C5a-mediated neutrophil dysfunction (as determined by reduced CD88 expression) was a strong predictor for subsequent acquisition of nosocomial infection (relative risk, 5.8; 95% confidence interval, 1.5-22; P = .0007), and remained independent of time effects as assessed by survival analysis (hazard ratio, 5.0; 95% confidence interval, 1.3-8.3; P = .01). In conclusion, this study provides new insight into the mechanisms underlying immunocompromise in critical illness and suggests novel avenues for therapy and prevention of nosocomial infection.

Changing Incidence and Characteristics of Nontuberculous Mycobacterial Infections in Scotland and Comparison With Mycobacterium tuberculosis Complex Incidence (2011 to 2019).

Background: An increase in infections with nontuberculous mycobacteria (NTM) has been noted globally, and their incidence has overtaken that of Mycobacterium tuberculosis complex (MTBc) in many countries. Using data from a national reference laboratory, we aimed to determine if this trend could be observed in Scotland. Methods: We undertook a retrospective review of all NTM isolates received by the Scottish Mycobacteria Reference Laboratory (SMRL) over 9 years from 2011 to 2019 inclusive. Clinical episodes were defined as per 2017 British Thoracic Society and 2020 American Thoracic Society/European Respiratory Society/European Society of Clinical Microbiology and Infectious Diseases/Infectious Diseases Society of America NTM guidelines. These rates were compared with Scottish tuberculosis rates over the same period. Results: Of 8552 NTM isolates from 4586 patients in 2011 to 2019, 7739 (90.5%) were considered clinically relevant. These represented 2409 episodes of NTM infection, with M. avium, M. intracellulare, and M. abscessus complex being most common. A total of 1953 (81.1%) were pulmonary NTM infection episodes from 1470 patients and 456 extrapulmonary episodes from 370 patients. We estimated a rise in incidence from 3.4 to 6.5 per 100 000 person-years (2011-2019 inclusive), with an increase in NTM incidence over MTBc incidence in Scotland by 2017. Conclusions: The incidence of NTM infection in Scotland has overtaken MTBc incidence. NTM infection leads to a costly health care burden, possibly as much as UK£1.47 million (US$ and €1.73 million) annually. We recommend standardization of isolate referral with clinical surveillance and implementation of agreed standards of care delivered through multidisciplinary teams. This would improve diagnosis and patient management as well as assessment of diagnostics and novel treatments through clinical trials.

Performance of lipid fingerprint by routine matrix-assisted laser desorption/ionization time of flight for the diagnosis of Mycobacterium tuberculosis complex species.

OBJECTIVES: Rapid detection of bacterial pathogens at species and sub-species levels is crucial for appropriate treatment, infection control, and public health management. Currently, one of the challenges in clinical microbiology is the discrimination of mycobacterial sub-species within the M. tuberculosis complex (MTBC). Our objective was to evaluate the ability of a biosafe mycobacterial lipid-based approach to identify MTBC cultures and sub-species. METHODS: A blinded study was conducted using 90 mycobacterial clinical isolate strains comprising MTBC strains sub-cultured in Middlebrook 7H11 medium supplemented with 10% oleic-acid, dextrose, catalase growth supplement and incubated for up to 6 weeks at 37°C and using the following seven reference strains (M. tuberculosis H37Rv, M canettii, M. africanum, M. pinnipedii, M. caprae, M. bovis, and M. bovis BCG) grown under the same conditions, to set the reference lipid database and test it against the 90 MTBC clinical isolates. Cultured mycobacteria were heat-inactivated and loaded onto the matrix-assisted laser desorption/ionization target followed by the addition of the matrix. Acquisition of the data was performed using the positive ion mode. RESULTS: Based on the identification of clear and defined lipid signatures from the seven reference strains, the method that we developed was fast (<10 minutes) and produced interpretable profiles for all but four isolates, caused by poor ionization giving an n = 86 with interpretable spectra. The sensitivity and specificity of the matrix-assisted laser desorption/ionization time of flight were 94.4 (95% CI, 86.4-98.5) and 94.4 (95% CI, 72.7-99.9), respectively. CONCLUSIONS: Mycobacterial lipid profiling provides a means of rapid, safe, and accurate discrimination of species within the MTBC.

Investigating transmission of Mycobacterium bovis in the United Kingdom in 2005 to 2008.

Due to an increase in bovine tuberculosis in cattle in the United Kingdom, we investigated the characteristics of Mycobacterium bovis infection in humans and assessed whether extensive transmission of M. bovis between humans has occurred. A cross-sectional study linking demographic, clinical, and DNA fingerprinting (using 15-locus mycobacterial interspersed repetitive-unit-variable-number tandem-repeat [MIRU-VNTR] typing) data on cases reported between 2005 and 2008 was undertaken. A total of 129 cases of M. bovis infection in humans were reported over the period, with a decrease in annual incidence from 0.065 to 0.047 cases per 100,000 persons. Most patients were born pre-1960, before widespread pasteurization was introduced (73%), were of white ethnicity (83%), and were born in the United Kingdom (76%). A total of 102 patients (79%) had MIRU-VNTR typing data. A total of 31 of 69 complete MIRU-VNTR profiles formed eight distinct clusters. The overall clustering proportion determined using the n - 1 method was 33%. The largest cluster, comprising 12 cases, was indistinguishable from a previously reported West Midlands outbreak strain cluster and included those cases. This cluster was heterogeneous, having characteristics supporting recent zoonotic and human-to-human transmission as well as reactivation of latent disease. Seven other, smaller clusters identified had demographics supporting recrudescence rather than recent infection. A total of 33 patients had incomplete MIRU-VNTR profiles, of which 11 may have yielded 2 to 6 further small clusters if typed to completion. The incidence of M. bovis in humans in the United Kingdom remains low, and the epidemiology is predominantly that of reactivated disease.

Reducing ventilator-associated pneumonia in intensive care: impact of implementing a care bundle.

OBJECTIVES: Ventilator-associated pneumonia is the most common intensive care unit-acquired infection. Although there is widespread consensus that evidenced-based interventions reduce the risk of ventilator-associated pneumonia, controversy has surrounded the importance of implementing them as a "bundle" of care. This study aimed to determine the effects of implementing such a bundle while controlling for potential confounding variables seen in similar studies. DESIGN: A before-and-after study conducted within the context of an existing, independent, infection surveillance program. SETTING: An 18-bed, mixed medical-surgical teaching hospital intensive care unit. PATIENTS: All patients admitted to intensive care for 48 hrs or more during the periods before and after intervention. INTERVENTIONS: A four-element ventilator-associated pneumonia prevention bundle, consisting of head-of-bed elevation, oral chlorhexidine gel, sedation holds, and a weaning protocol implemented as part of the Scottish Patient Safety Program using Institute of Health Care Improvement methods. MEASUREMENTS AND MAIN RESULTS: Compliance with head-of-bed elevation and chlorhexidine gel were 95%-100%; documented compliance with "wake and wean" elements was 70%, giving overall bundle compliance rates of 70%. Compared to the preintervention period, there was a significant reduction in ventilator-associated pneumonia in the postintervention period (32 cases per 1,000 ventilator days to 12 cases per 1,000 ventilator days; p<.001). Statistical process control charts showed the decrease was most marked after bundle implementation. Patient cohorts staying ≥6 and ≥14 days had greater reduction in ventilator-associated pneumonia acquisition and also had reduced antibiotic use (reduced by 1 and 3 days; p=.008/.007, respectively). Rates of methicillin-resistant Staphylococcus aureus acquisition also decreased (10% to 3.6%; p<.001). CONCLUSIONS: Implementation of a ventilator-associated pneumonia prevention bundle was associated with a statistically significant reduction in ventilator-associated pneumonia, which had not been achieved with earlier ad hoc ventilator-associated pneumonia prevention guidelines in our unit. This occurred despite an inability to meet bundle compliance targets of 95% for all elements. Our data support the systematic approach to achieving high rates of process compliance and suggest systematic introduction can decrease both infection incidence and antibiotic use, especially for patients requiring longer duration of ventilation.

False detection of rifampicin resistance using Xpert® MTB/RIF Ultra assay due to an A451V mutation in Mycobacterium tuberculosis.

BACKGROUND: In a 12 month period, three Irish-born adult cases with pulmonary TB were initially diagnosed by Xpert® MTB/RIF Ultra assay, which detected a rifampicin resistance-conferring mutation prompting treatment as potential MDR cases. METHODS: Further laboratory investigations on the cultured isolates included GenoType MTBDRplus assay, phenotypic drug susceptibility tests using the BD BACTEC MGIT culture system and MIC broth microdilution tests. Sequencing of the rpoB gene was performed using Sanger sequencing and WGS. RESULTS: Phenotypic drug susceptibility tests determined the isolates to be rifampicin susceptible. Molecular investigations identified an A451V (codon 532) mutation in the Mycobacterium tuberculosis rpoB gene that has not previously been found to cause rifampicin resistance. Genome sequencing revealed that the three isolates' genomes differed by ≤5 SNPs, indicating a high likelihood of recent transmission events. Furthermore, a cluster of six related M. tuberculosis isolates from our in-house typing database showed four were highly related; all were rifampicin susceptible and lacked this mutation. CONCLUSIONS: False detection of rifampicin resistance, albeit rare, should be considered possible with Xpert® MTB/RIF Ultra assay, particularly in low TB incidence settings. Confirmatory sequencing methods should be performed to prevent the unnecessary use of second-line anti-tuberculous drugs.

Diagnostic importance of pulmonary interleukin-1beta and interleukin-8 in ventilator-associated pneumonia.

BACKGROUND: Ventilator-associated pneumonia (VAP) is the most commonly fatal nosocomial infection. Clinical diagnosis of VAP remains notoriously inaccurate. The hypothesis was tested that significantly augmented inflammatory markers distinguish VAP from conditions closely mimicking VAP. METHODS: A prospective, observational cohort study was carried out in two university hospital intensive care units recruiting 73 patients with clinically suspected VAP, and a semi-urban primary care practice recruiting a reference group of 21 age- and sex-matched volunteers. Growth of pathogens at >10(4) colony-forming units (cfu)/ml of bronchoalveolar lavage fluid (BALF) distinguished VAP from "non-VAP". Inflammatory mediators were quantified in BALF and serum. Mediators showing significant differences between patients with and without VAP were analysed for diagnostic utility by receiver operator characteristic (ROC) curves. RESULTS: Seventy-two patients had recoverable lavage-24% had VAP. BALF interleukin-1beta (IL-1beta), IL-8, granulocyte colony-stimulating factor and macrophage inflammatory protein-1alpha were significantly higher in the VAP group (all p<0.005). Using a cut-off of 10 pg/ml, BALF IL-1beta generated negative likelihood ratios for VAP of 0.09. In patients with BALF IL-1beta <10 pg/ml the post-test probability of VAP was 2.8%. Using a cut-off value for IL-8 of 2 ng/ml, the positive likelihood ratio was 5.03. There was no difference in cytokine levels between patients with sterile BALF and those with growth of <10(4) cfu/ml. CONCLUSIONS: BALF IL-1beta and IL-8 are amongst the strongest markers yet identified for accurately demarcating VAP within the larger population of patients with suspected VAP. These findings have potential implications for reduction in unnecessary antibiotic use but require further validation in larger populations.

Sensitivity of RT-PCR testing of upper respiratory tract samples for SARS-CoV-2 in hospitalised patients: a retrospective cohort study.

Background: This study aimed to determine the sensitivity and specificity of reverse transcription PCR (RT-PCR) testing of upper respiratory tract (URT) samples from hospitalised patients with coronavirus disease 2019 (COVID-19), compared to the gold standard of a clinical diagnosis. Methods: All URT RT-PCR testing for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in NHS Lothian, Scotland, United Kingdom between the 7 th of February and 19 th April 2020 (inclusive) was reviewed, and hospitalised patients were identified. All URT RT-PCR tests were analysed for each patient to determine the sequence of negative and positive results. For those who were tested twice or more but never received a positive result, case records were reviewed, and a clinical diagnosis of COVID-19 allocated based on clinical features, discharge diagnosis, and radiology and haematology results. For those who had a negative RT-PCR test but a clinical diagnosis of COVID-19, respiratory samples were retested using a multiplex respiratory panel, a second SARS-CoV-2 RT-PCR assay, and a human RNase P control. Results: Compared to the gold standard of a clinical diagnosis of COVID-19, the sensitivity of a single upper respiratory tract RT-PCR for COVID-19 was 82.2% (95% confidence interval 79.0-85.1%).   The sensitivity of two upper respiratory tract RT-PCR tests increased sensitivity to 90.6% (CI 88.0-92.7%). A further 2.2% and 0.9% of patients who received a clinical diagnosis of COVID-19 were positive on a third and fourth test; this may be an underestimate of the value of further testing as the majority of patients 93.0% (2999/3226) only had one or two URT RT-PCR tests. Conclusions: The sensitivity of a single RT-PCR test of URT samples in hospitalised patients is 82.2%. Sensitivity increases to 90.6% when patients are tested twice.  A proportion of cases with clinically defined COVID-19 never test positive on URT RT-PCR despite repeat testing.

Phenotypic and molecular detection methods for carbapenemase-producing organisms and their clinical significance at two Scottish tertiary care hospitals.

PURPOSE: This study evaluated in-house PCR testing for local identification of bacteria carrying the major carbapenemase genes (blaOXA-48-like, blaVIM, blaNDM, blaKPC and blaIMP). METHODOLOGY: Carbapenemase-producing organisms (CPOs) isolated from patients managed in two tertiary care hospitals in Scotland from September 2014-January 2017 were investigated. A combination of chromogenic screening agar (ChromID CARBA SMART), a carbapenem hydrolysis test (Rapidec Carba NP) and in-house real-time PCR for the blaOXA-48-like, blaVIM, blaNDM, blaKPC and blaIMP genes were utilized. All isolates were sent to the AMRHAI reference unit for confirmatory testing. RESULTS: During the 29-month study period 39 CPO were isolated from 34 patients. The average turnaround time for a workflow involving phenotypic and molecular testing was 4.2 days. PCR had a sensitivity and specificity of 100 %. The most common carbapenemase genes were blaOXA-48-like (31%), blaVIM (23%) and blaNDM (20%). Resistance to antimicrobials other than beta-lactams was common; the most active agents were colistin, amikacin and fosfomycin. Twenty-seven patients were considered to be colonized (although CPO detection influenced empiric antimicrobials in five) and a CPO was implicated in infection in seven patients (bacteraemia in immunocompromised patients, n=2; surgical site infections, n=2; osteomyelitis in a patient with diabetes mellitus, n=1; and urinary tract infections, n=2). All patients survived infection. CONCLUSION: In a lowincidence setting we demonstrate the efficacy of a combined local laboratory workflow for rapid detection of CPOs, incorporating phenotypic and molecular testing. In 7/34 patients the CPO was implicated as a pathogen and detection influenced antimicrobial decision-making in five colonized patients.

Tractable targets for meropenem-sparing antimicrobial stewardship interventions.

BACKGROUND: As meropenem is a restricted antimicrobial, lessons learned from its real-life usage will be applicable to antimicrobial stewardship (AMS) more generally. OBJECTIVES: To retrospectively evaluate meropenem usage at our institution to identify targets for AMS interventions. METHODS: Patients receiving meropenem documented with an 'alert antimicrobial' form at two tertiary care UK hospitals were identified retrospectively. Clinical records and microbiology results were reviewed. RESULTS: A total of 107 adult inpatients receiving meropenem were identified. This was first-line in 47% and escalation therapy in 53%. Source control was required in 28% of cases after escalation, for predictable reasons. Those ultimately requiring source control had received more prior antimicrobial agents than those who did not (P = 0.03). Meropenem was rationalized in 24% of cases (after median 4 days). Positive microbiology enabled rationalization (OR 12.3, 95% CI 2.7-55.5, P = 0.001) but rates of appropriate sampling varied. In cases with positive microbiology where meropenem was not rationalized, continuation was retrospectively considered clinically and microbiologically necessary in 8/40 cases (0/17 empirical first-line usage). Rationalization was more likely when meropenem susceptibility was not released on the microbiology report (OR 5.2, 95% CI 1.3-20.2, P = 0.02). Input from an infection specialist was associated with a reduced duration of meropenem therapy (P < 0.0001). Early review by an infection specialist has the potential to further facilitate rationalization. CONCLUSIONS: In real-life clinical practice, core aspects of infection management remain tractable targets for AMS interventions: microbiological sampling, source control and infection specialist input. Further targets include supporting rationalization to less familiar carbapenem-sparing antimicrobials, restricting first-line meropenem usage and selectively reporting meropenem susceptibility.