key: cord-309733-x4crng5j authors: Dhesi, Z.; Enne, V. I.; Brealey, D.; Livermore, D. M.; High, J.; Russell, C.; Colles, A.; Kandil, H.; Mack, D.; Martin, D.; Page, V.; Parker, R.; Roulston, K.; Singh, S.; Wey, E.; Swart, A. M.; Stirling, S.; Barber, J. A.; O'Grady, J.; Gant, V. A. title: Organisms causing secondary pneumonias in COVID-19 patients at 5 UK ICUs as detected with the FilmArray test date: 2020-06-23 journal: nan DOI: 10.1101/2020.06.22.20131573 sha: doc_id: 309733 cord_uid: x4crng5j ABSTRACT Introduction. Several viral respiratory infections - notably influenza - are associated with secondary bacterial infection and additional pathology. The extent to which this applies for COVID-19 is unknown. Accordingly, we aimed to define the bacteria causing secondary pneumonias in COVID-19 ICU patients using the FilmArray Pneumonia Panel, and to determine this tests potential in COVID-19 management. Methods. COVID-19 ICU patients with clinically-suspected secondary infection at 5 UK hospitals were tested with the FilmArray at point of care. We collected patient demographic data and compared FilmArray results with routine culture. Results. We report results of 110 FilmArray tests on 94 patients (16 had 2 tests): 69 patients (73%) were male, the median age was 59 yrs; 92 were ventilated. Median hospital stay before testing was 14 days (range 1-38). Fifty-nine (54%) tests were positive, with 141 bacteria detected. Most were Enterobacterales (n=55, including Klebsiella spp. [n= 35]) or Staphylococcus aureus (n=13), as is typical of hospital and ventilator pneumonia. Community pathogens, including Haemophilus influenzae (n=8) and Streptococcus pneumoniae (n=1), were rarer. FilmArray detected one additional virus (Rhinovirus/Enterovirus) and no atypical bacteria. Fewer samples (28 % vs. 54%) were positive by routine culture, and fewer species were reported per sample; Klebsiella species remained the most prevalent pathogens. Conclusion. FilmArray had a higher diagnostic yield than culture for ICU COVID-19 patients with suspected secondary pneumonias. The bacteria found mostly were Enterobacterales, S. aureus and P. aeruginosa, as in typical HAP/VAP, but with Klebsiella spp. more prominent. We found almost no viral co-infection. Turnaround from sample to results is around 1h 15 min compared with the usual 72h for culture, giving prescribers earlier data to inform antimicrobial decisions. The emergence of SARS-CoV2 as a pandemic virus of global importance drives a need for clinical and pathological evidence upon which to base optimal therapeutic decisions. Whilst purely viral infections should not be treated with antibiotics, several respiratory viruses, notably influenza, are associated with secondary bacterial infection and additional pathology. These secondary infections reflect a combination of damage to the protective mucosa, facilitating bacterial colonisation and invasion, as well as virally-induced immunosuppression (1, 2) . Viral and bacterial respiratory coinfections exacerbate disease severity, and can prompt ICU admission (3) . The extent to which COVID-19, as the disease caused by SARS-CoV2, is associated with secondary bacterial infection of the respiratory tract is unknown (4) . Anecdotal evidence suggests that hospitalised COVID-19 patients are frequently prescribed empirical antimicrobials. Whether this is microbiologically necessary, even in severe cases, is unknown (5) . In a brief review of existing literature Rawson et al conclude that there currently are insufficient data to inform empiric or reactive antibiotic decisions in a reasonable timeframe for critically-ill COVID-19 patients (5) . Irrespective of COVD-19, investigation of clinically-suspected bacterial pneumonia is complicated by the poor sensitivity of sputum culture and by the considerable interval (typically circa 72h) from sample to susceptibility test results. Recently-developed rapid tests have the potential to improve both the speed and sensitivity of investigation (6) . INHALE (ISRCTN16483855) is a UK NIHR-funded research programme investigating the utility of rapid molecular diagnostics for the microbiological investigation of HAP/VAP in critical care (7) . The programme incorporates an RCT, run across 12 UK hospitals, in which ICU patients with suspected hospital-acquired or ventilator-associated pneumonia (HAP/VAP) are randomised to have either (a) standard empirical therapy or (b) to have the BioFire FilmArray Pneumonia Panel test (bioMérieux) to support early treatment decisions (8) . All patients have conventional microbiological investigation performed. The FilmArray is a PCRbased test with a turnaround time of 1h15min and can be performed inside the ICU. An antibiotic-prescribing algorithm is provided to support decision-making based on the results, thus going beyond a "Point of Care Test (POCT)" to provide 'Point of Decision' clinical support. The COVID-19 pandemic resulted in recruitment to the INHALE trial being paused and, under the exigencies of the circumstances, we developed an observational sub-study to investigate the utility of the FilmArray Pneumonia Panel for the diagnosis and characterisation of secondary bacterial infection in COVID-19 ICU patients. Here, we report the results of this sub-study for 94 patients from 5 UK ICUs. The aims were to describe secondary bacterial, viral and "atypical" pathogens in COVID-19 ICU patients and to evaluate the potential of this panel for management of these patients. is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted June 23, 2020. . guidelines. Test results were immediately delivered to the clinical ICU team along with the INHALE RCT prescribing algorithm (which allows some local variation), providing recommended treatment guidance (11) . The foundation of this prescribing algorithm is to promote antimicrobial stewardship by indicating the narrowest spectrum antibiotic likely to cover the pathogen(s) detected and compatible with the patient's penicillin allergy status. A second FilmArray test ≥ 5 days from the first Pneumonia Panel test was permitted if a new or continuing bacterial pneumonia was suspected. In parallel, a respiratory sample was sent to the hospital laboratory for routine microbiological investigation, performed according to the standard UK Laboratories Operating Procedures (12) . Baseline data including age, sex, comorbidities, date of COVID-19 diagnosis, admission to hospital, and ICU admission were collected. FilmArray test results and clinical microbiology results were recorded, along with a brief statement of the reason for FilmArray testing. Antibiotic prescribing data were also collected, but remain under analysis and will be reported separately. A bespoke REDCap database was used for data collection and storage (13) ; this provides a number of features to maintain data quality, including an audit trail, ability to query spurious data, search facilities, and validation of predefined parameters/missing data. Chi Square tests were used to compare proportions. Both the main trial and the sub-study have ethical approval from the London, Brighton and Sussex Research Ethics Committee (19/LO/0400) and the Health Research Authority. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted June 23, 2020. Up to cut-off date of this analysis (6 May, 2020), 98 patients had been recruited at the 5 ICUs (range 7-44 patients per site), with FilmArray results available for 94, all recruited after 3 April 2020. Sixteen patients had the test performed twice, giving a total of 110 reports. Demographic and background health data are summarised in Table 1 . . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. . Clinicians were asked to record the clinical indication for performing the FilmArray test. Among 69 responses provided, across the 5 participating sites, the most widely cited reason was an increase in inflammatory markers/fever (42%), followed by 'considering a change of antibiotics' (19%), suspected bacterial pneumonia (15%), increased respiratory secretions/ hypoxia (14%), or to stop antibiotics (10%). The most prevalent pathogen found was Klebsiella pneumoniae, followed by S. aureus, Enterobacter cloacae and K. aerogenes. Resistance genes were detected in 7 samples: mecA/C, which confers methicillin resistance in staphylococci, was found in 5 samples from 4 patients at 3 ICUs (one patient had two tests, with mecA/C found both times). All these source patients were positive for S. aureus, indicating an MRSA incidence of 4%. bla CTX-M genes, encoding extended-spectrum β−lactamases, were detected in 2 samples, from different patients in the same ICU; both samples were positive for K. pneumoniae, a frequent host of these enzymes. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. (including negative results but excluding multiple instances of the same species from a single patient) . Out of the 110 specimens run on the FilmArray, 6 (5.5%) were not sent to the clinical microbiology laboratory and 5 (4.5%) were not reported; leaving 99 (90%) with a routine microbiology result Three analyses were performed to compare the agreement of the FilmArray and routine results. First, we performed a concordance analysis, as shown in table 2, for tests where both results were available. Most FilmArray results (60.6%) were fully concordant with routine culture. Among those that were not fully concordant the common pattern was for FilmArray to indicate pathogens in samples where routine microbiology reported no organisms (25.3% of tests) or to flag additional organisms over and above those reported by routine microbiology (11.1%). Only 3% of cases were classified as major discordances with routine microbiology culturing an organism that was sought by the FilmArray but not found by it. One patient had an opportunist pathogen (Citrobacter koseri) not represented on the FilmArray panel. *Includes one case where routine microbiology did not report S. aureus and Klebsiella spp., which were found by FilmArray, but did report Citrobacter koseri, which is not sought by FilmArray Secondly, we reviewed agreement by species group in relation to the organism load reported by FilmArray (Table 3 ). Only 6 of 34 organisms reported by the FilmArray at a load of 10 4 or 10 5 CFU/ml were reported by routine microbiology, but this proportion rose to 19/44 for organisms found at a load of 10 6 or 10 7 CFU/ml (p=0.014, chi square test). Thirdly, we considered agreements between phenotypic resistance and detection of corresponding resistance genes by FilmArray. MRSA was reported by routine microbiology from 3 of the 5 samples where FilmArray flagged mecA/C and S. aureus; the remaining 2 samples were reported by microbiology as yielding 'no growth'. Culture did not detect ESBL-producing organisms in either of the two samples where FilmArray found bla CTX-M. genes and K. pneumoniae but, curiously, did find . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. . https://doi.org/10.1101/2020.06.22.20131573 doi: medRxiv preprint an ESBL-producing K. pneumoniae in another sample (where FilmArray was negative) from the same ICU on the same day, raising a possible confusion of samples, though this could not be confirmed. Routine microbiology identified one K. oxytoca isolate with a phenotype suggesting hyper-production of K1 chromosomal β-lactamase; FilmArray detected the K. oxytoca, but does not seek the mutations that cause hyper-production of this enzyme. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. The age, sex and co-morbidity profiles of these patients are in keeping with those reported by others in severe COVID-19 disease (14) . Although the inclusion criteria permitted testing of patients at any point during their hospital and ICU admission, the majority (95%) of tests were performed at least 5 days after hospital admission. In context it should be noted that hospitalised COVID-19 patients in the UK are typically admitted to a general ward then, after several days, if necessary, are transferred to ICU, where our testing was conducted (14) . Although sites had the option of using the test earlier during the hospitalisation, they reported anecdotally that most recently-hospitalised COVID-19 patients were unproductive for sputum and thus not eligible (data not shown). This observation may, of itself, suggest that early-onset bacterial secondary infections are uncommon in COVID-19 illness, as they would be expected to provoke sputum production. Nonetheless, it would be pertinent to examine the microbiology earlier in the patient's hospital journey, e.g. on the day of admission, to see whether there were more negative results or if different organisms are isolated. However, the lack of sputum production may constrain such studies, especially where deliberate sampling, such as collecting BAL or induced sputum, is viewed as unnecessarily hazardous to staff and invasive for patients. Among 110 FilmArray tests, representing 94 patients, 56% recorded bacteria or, in one case, a second virus. The microbiology found resembled that typical of HAP/VAP, in being dominated by Enterobacterales, P. aeruginosa and S, aureus (15) . Organisms typically associated with communityacquired pneumonia were much less prominent: nevertheless H. influenzae was detected in 8 patients . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. . and S. pneumoniae in one, with all these detections relating to patients who had been hospitalised for at least 5 days, and for 22 days in the case of the S. pneumoniae patient. Similar observations in other studies of (COVID-19-unrelated) HAP suggest that these 'community' organisms can, on occasion, be hospital-acquired (15) . Despite the dominance of pathogens typically associated with HAP/VAP the species distribution differed from that seen in INHALE's earlier evaluation study of the FilmArray test in HAP/VAP patients, conducted prior to the COVID-19 pandemic (16, 17) . In particular, Klebsiella spp. (both K. pneumoniae and K. aerogenes) were significantly more prevalent (35/89 isolates versus 85/775, p <0.001; chi square test) in the COVID-19 patients, whereas P. aeruginosa and E. coli were underrepresented, with the overall species distributions also significantly different (p <0.001; chi square test). An altered species distribution in HAP/VAP may reflect the particular thrombotic lung pathology associated with COVID-19 (18) . The distribution of bacteria differed even more markedly from that typically seen following influenza, which is dominated by community-acquired pathogens such as S. pneumoniae and H. influenzae, with S. aureus also prominent (1). In China, Zhu et al. found S. pneumoniae to be the most prevalent bacterial pathogen in COVD-19 patients, followed by K. pneumoniae and H. influenzae (19) . In contrast to our study they mostly sampled early in the course of COVID-19 disease and, using throat swabs, examined patients who varied greatly in disease severity, meaning that comparability is tenuous. Also of note, only one of our 94 patients had an additional respiratory virus whereas 15.2% (94/620) of adult patients were positive for viruses in earlier INHALE work (16) . This contrasts with data from China and California, where 22.6 -31.5% of COVID-19 patients had co-infection with other viruses (19, 20) . The key difference may be that we specifically examined ICU patients, many of whom had been hospitalised for prolonged periods, whereas these authors examined broader groups of COVID-19 patients with more recent community residency. Alternatively, the difference may be that these studies were done up to March 2020, and so overlapped the winter respiratory season, whereas we recruited later, in April and May. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. . In general, FilmArray identified a larger proportion of samples as positive for bacteria than routine culture (54 % vs. 28%); moreover, FilmArray more often indicated multiple bacteria in a sample. These findings accord with our previous observations, where we demonstrated that various PCR systems including FilmArray, Curetis Unyvero and also 16S rDNA analysis, all tended to find more organisms than are reported by routine culture from respiratory samples and that they tended also to find the same additional organisms as one another, implying that the vast majority of these additional detections represent organisms genuinely present in the sample (16) . A curious discrepancy was that routine serology, only obtained from one hospital, reported 6 cases positive for mycoplasma among the cohort (Table 3) . M. pneumoniae was not detected by FilmArray PCR in the corresponding specimens, suggesting either that the serology represented a false positive result, perhaps owing to an anamnestic response, as seen with Dengue serology (21) , or that mycoplasma species other than M. pneumoniae were present. ICU clinicians have welcomed this new diagnostic platform to aid the rapid detection (or not) of bacteria in their patients' lower respiratory tracts, and as a guide to treatment. The hazard is, however, that the greater diagnostic yield compared with culture may lead to treatment of patients who merely had a few colonising bacteria. The significance of organisms detected at low population densities (10 4 to 10 5 CFU/ml) remains open to debate; those found at higher densities were more often reported also by routine microbiology, with this differentiation stronger than in the main INHALE trial. More generally, we would underscore that the clinical context must be taken into account and that, as with many microbiological results, detection of an organism does not prove that it is causing infection. Balancing these factors will need careful liaison between ICUs, microbiology and other antimicrobial stewards; furthermore, clinical antibiotic prescribing decisions are subject to factors beyond a valid test result, as demonstrated in the VAPrapid study (22) . That said, preliminary observational data from INHALE's earlier work suggested that treatment of additional organisms detected by PCR may have the potential to improve patient outcomes (17) . To examine the impact of FilmArray results in the context of COVID-19, we are collecting and analysing additional data at the . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 23, 2020. . https://doi.org/10.1101/2020.06.22.20131573 doi: medRxiv preprint 2 largest of the present 5 sites, assessing consequences for antimicrobial prescribing and patient outcomes. A behavioural sub-study is also underway to investigate how antibiotic decision making has been affected during the COVID-19 pandemic, and how this is influenced by the FilmArray. In summary we have shown, first, that the bacteria causing secondary pneumonias in severely-ill COVID-19 patients mostly are Enterobacterales, S. aureus and P. aeruginosa, as is typical of HAP/VAP. The organism distribution is different from 'typical' HAP/VAP, with K. pneumoniae and K. aerogenes more prominent and E. coli and P. aeruginosa less prominent. Secondly, severe COVID-19 patients do not appear to progress to secondary bacterial infection in the same way as do severe influenza patients and do not have the same pathogens; rather, invasive ventilation seems likely to be the main driver for secondary infections in COVID-19. Thirdly, we have shown that FilmArray had a higher diagnostic yield than culture-as reported also in INHALE's pre-COVID-19 work (16, 17) . Turnaround from sample to results was around 1h 15 min compared with the usual 72h for culture, giving prescribers earlier data to inform antimicrobial decisions. Further work is required to establish the contribution of secondary infections to the overall clinical outcome in severely ill COVID-19 patients. . 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