Enhanced diagnosis of severe bacterial and fungal respiratory infection in children using a rapid syndromic array—case report

Introduction

Lower respiratory tract infection (LRTI) is a leading cause of admissions to paediatric intensive care units (PICUs) (1). Between 2–7% of children hospitalised for community acquired pneumonia (CAP) require mechanical ventilation in high-income countries (2,3). Children requiring prolonged mechanical ventilation for other problems can develop ventilator-associated pneumonia (VAP). Although there are consensus-based definitions of pneumonia (4,5), adherence to these definitions may vary and pathogen-specific diagnoses may be dependent on laboratory methodology and whether microorganism/s detected are considered the underlying cause of infection. National audits in the UK have found that a pathogen is identified in 9–16% of cases of paediatric pneumonia (3). Microbiological cultures are limited by previously administered antimicrobials, particularly in the PICU where broad-spectrum treatment is common (6). Molecular syndromic diagnostic tests are an alternative method for obtaining a pathogen-specific diagnosis in critically ill children. Whilst there are several studies describing use of these tests in paediatric samples, descriptions of real-world application and interpretation of these tests to influence clinical decision making are limited (7-11). Here we describe three cases which demonstrate ways in which molecular diagnostics can impact on prescribing decisions in the PICU.

PICU patients with suspected respiratory infections had samples obtained via endotracheal tube aspiration (ETA) or non-bronchoscopic bronchoalveolar lavage (BAL) (mini-BAL). In this procedure, saline is instilled and re-aspirated using an in-line suction catheter at the level of the carina. Nucleic acid was extracted from the sample and underwent a 52-respiratory pathogen TaqMan array card (TAC) (Table S1). This array was custom designed for use in this institution using a combination of commercially available targets in addition to targets that were sequenced in-house (12). Each target was reported according to cycle threshold (Ct), which represents the number of cycles of polymerase chain reaction (PCR) amplification that occurred before the exponential increase in the fluorescent reporter signal. A Ct result of ≤32 was generally considered significant, given this corresponded with clinically significant growth on microbiological culture during validation (12). Importantly, the result was taken in the context of the patient regardless of Ct value. We present this article in accordance with the CARE reporting checklist (available at https://tp.amegroups.com/article/view/10.21037/tp-23-525/rc).

Case presentation

Consent and procedures

All procedures performed in the study were in accordance with the ethical standards of the institutional and/or national research committee(s) and with the Helsinki Declaration (as revised in 2013). Written informed consent was obtained from all patients’ legal guardians for publication of this case report. A copy of the written consent is available for review by the editorial office of this journal.

Case one: Candida pneumonia in a trauma patient

A 17-year-old male went for an emergency laparotomy in his local hospital after a motorcycle accident in which he sustained a grade V liver laceration, a right-sided pneumothorax, rib, and limb fractures. He was intubated, his abdomen was packed, and post-operatively he was transferred to PICU. Haemostasis was achieved on re-exploration of the abdomen and hepatic and coeliac artery embolisation. During stabilisation, >10 L of blood products were required. He initially received amoxicillin-clavulanic acid, gentamicin, and fluconazole prophylaxis peri-operatively, which was rationalised to piperacillin-tazobactam in PICU. He returned to theatre on day 3 to manage bile leak and underwent endoscopic retrograde cholangiopancreatography (ERCP) with biliary stenting, peritoneal drain insertion and removal of intra-abdominal packs. On day 5 of his admission, his fraction of inspired oxygen (FiO₂) needed to be progressively increased from 21% to 80% despite incremental increases in peak end-expiratory pressure (PEEP). He also began to develop fevers. Chest radiograph demonstrated consolidation of the left lower lobe of the lung. Following mini-BAL culture and TAC, treatment gentamicin was added, and piperacillin-tazobactam was continued. TAC results returned on day 6, after 25 hours, suggestive of Candida albicans (C. albicans) VAP (Figure 1A). The microbiology team thought that the TAC result was ‘colonising microbiota’ and not necessarily contributing to current state. Pulmonary embolism was found on computerised tomography pulmonary angiogram (CTPA), and anti-coagulation was commenced. By day 7, the patient required 100% oxygen, and PEEP on conventional mechanical ventilation was set at 10 cmH₂O. Antimicrobials were changed to meropenem, gentamicin and amphotericin-B, as part of an escalation plan based on TAC results and clinical information. On day 8, 70 hours after collection, C. albicans was identified on the mini-BAL culture. The same organism was isolated on blood culture on day 11. He received a total of 7 days of gentamicin, 12 days of meropenem and 6 days of amphotericin-B. By day 17, the patient was recovering well and was discharged from the PICU.

Figure 1 This figure demonstrates the investigation results of three children with suspected respiratory infection admitted to the pediatric intensive care unit. In case one, Candida albicans was identified on TAC prior to blood cultures, prompting earlier treatment when the patient had early signs of deterioration. In case two, an unusual diagnosis of psittacosis was made based on TAC results. In case three, Haemophilus influenzae respiratory was diagnosed, whilst the pathogen was not identified on routine investigations. BAL, bronchoalveolar lavage; CFU, colony forming unit; Ct, cycle threshold; MIS-C, multisystem inflammatory syndrome in children; ETA, endotracheal aspirate; CSF, cerebrospinal fluid; TAC, TaqMan array card.

Case two: Chlamydia psittaci (C. psittaci) pneumonia in an undifferentiated adolescent

A 16-year-old female presented to her local hospital with a 3-day history of fever, cough, and increased work of breathing. She required FiO₂ 70% to maintain SaO₂ >90% on high-flow nasal cannulae running at 60 L/min. Her initial inflammatory markers were significantly elevated (Figure 1B). Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) combined nasopharyngeal-oropharyngeal swab quantitative PCR (qPCR) was negative. On chest radiograph, she had bilateral basal consolidation and parapneumonic effusion. Given worsening respiratory distress, she was intubated and transferred to PICU. The leading differential diagnosis was multisystem inflammatory syndrome in children (MIS-C), a latent response to coronavirus disease 2019 (COVID-19). MIS-C was suspected given the history of fever, inflammatory presentation of the patient, and a high incidence of this problem in UK PICUs at the time (13).

TAC, undertaken on a mini-BAL sample obtained at the time of admission to PICU, identified C. psittaci, which was reported back the following day. On further questioning, the family said they had purchased two pet cockatiels, one of which had recently died. The patient initially received meropenem and clindamycin, later rationalised to a 10-day course of doxycycline. She required 3 days of mechanical ventilation and was discharged to the ward. The C. psittaci result was confirmed on C. psittaci/abortus serology undertaken in the national reference laboratory, with results available after 10 days.

Case three: Haemophilus influenzae (H. influenzae) pneumonia in a neonate with suspected sepsis and meningitis

A 2-week-old male, born at 35⁺⁵ weeks gestation, was taken to his local hospital with increased work of breathing and apnoeas. He had received 5 days of antimicrobial therapy after birth for suspected sepsis with no organism identified. In the present admission, he had a short history of cough and nasal congestion. Despite treatment with high-flow nasal cannulae, intubation was required due to the persistence of apnoeas. He then needed 40 mL/kg in fluid boluses for hypotension and tachycardia. Ceftriaxone, amoxicillin and acyclovir were given to treat possible sepsis and meningitis. He was transferred to PICU overnight. The patient was producing moderate, thick yellow secretions and there were scattered crackles on chest auscultation. Chest radiograph demonstrated diffuse interstitial change. Mini-BAL was undertaken the following morning given suspicion of a primary respiratory source of infection. H. influenzae was reported on TAC 5 hours after sample collection (Figure 1C). Amoxicillin and acyclovir were accordingly ceased. Ceftriaxone was rationalised to a 10-day course of amoxicillin-clavulanate, and he was extubated and discharged to the ward on day 3 of PICU admission. There were no organisms reported on microbiological cultures when results became available later.

Discussion

These clinical cases demonstrate that TAC can diagnose bacterial and fungal infections in critically ill children faster than routine investigations, and in some instances, identify pathogens that would not be identified using first-line tests. Diagnostic syndromic arrays for pneumonia have been widely commercially available since 2018, but their potential role in children has not been described broadly (14,15).

In our first case study, C. albicans pneumonia was identified earlier by TAC than routine microbiological cultures. Candida causes between 2.4–11.9% of paediatric VAP (16). There are no targets for Candida on commercial pneumonia syndromic arrays. Whilst C. albicans can be a lung commensal, pulmonary candidiasis can occur in severely unwell and immunocompromised patients—a group that frequently requires PICU admission. This pneumonia likely occurred secondary to disseminated candidiasis. The presence of candidaemia can increase mortality by 10.0% (95% confidence interval: 6.2–13.8%) in children; hence its prompt identification is essential (17). If found on a respiratory sample alone, clinicians must decide whether it requires treatment (18). In this case, whilst anti-fungal treatment was not given immediately, identification of Candida using TAC allowed treatment to be escalated promptly when the patient demonstrated signs of deterioration.

In the second case study, an unexpected diagnosis of C. psittaci was made due to the detection on the TAC, which would have been unlikely to have been identified with routine tests. Psittacosis occurs due to contact with aerosols generated from the urine or faeces of birds, particularly those belonging to the Psittaciformes—parrots, parakeets, budgerigars and cockatiels (19,20). It is an intracellular, Gram-negative infection that is rare in children; however, is expected to be identified more frequently due to the increasing availability of molecular diagnostic tests (19,20). In this case, TAC significantly impacted treatment as the patient initially had poor Gram-negative antimicrobial cover. A benefit of TAC is that it contains a wide range of pathogens that may help detect unusual respiratory pathogens, potentially offering a diagnosis in situations of uncertainty.

In the last case, H. influenzae pneumonia was identified as the primary cause of infection, resulting in neonatal apnoeas. In this case, early identification of the organism with TAC allowed antimicrobial cover for presumed central nervous system infection to be rationalised sooner than if decision making was based on microbiological culture. The prevalence of invasive paediatric Haemophilus has reduced significantly since the introduction of the H. influenzae type B (Hib) vaccine, which is administered in repeated doses from eight weeks of life.

The three cases highlight the role and potential limitations of syndromic diagnostic arrays in children. The TAC identified the cause of pneumonia faster than routine cultures and provided a cause where it may not have been otherwise identified. Respiratory cultures in the PICU are typically obtained after children have received broad-spectrum antimicrobial therapy, when initial signs of bacterial LRTI, or deterioration is identified. This reduces the viable number of organisms in specimens, reducing the yield of microbiological culture. Molecular diagnostic tests are advantageous in that they do not require a live organism, but instead detect nucleic acid to assist clinicians in making a pathogen-specific diagnosis. Molecular diagnostics tests are capable of identifying respiratory pathogens within 1–5 hours (21-24), offering the potential for clinicians to make earlier antimicrobial decisions.

As is the case for microbiological cultures, it is possible to identify organisms that may be commensals/contaminants with rapid molecular diagnostic arrays. This was initially suspected in case one, which is why the clinical oversight by the multidisciplinary team is essential. Without this input there is a risk of unnecessary escalation of antimicrobial therapy should low consequence microorganisms be detected. Diagnostic arrays, such as TAC, are limited to detection of pathogens pre-specified by design. Therefore, it is important for clinicians to be aware that a negative test result does not indicate absence of an off-panel pathogen. This is critical to consider where tests are used in international settings, where there may be different patterns of epidemiology. Finally, there is still progress to be made in implementing molecular diagnostic tests into clinical practice. Institutions must have the laboratory infrastructure and personnel, and familiarity of clinicians in interpreting the tests for their routine use to be effective (25).

In comparison to commercially available molecular diagnostic arrays, TAC has a longer turnaround time given the need for laboratory staff to perform extraction of nucleic acid manually. However, the test is capable of a broader range of pathogens that other tests and is customisable. Currently, use of the respiratory pathogen TAC is limited to this centre, but in future may become available if validated through a larger scale study. We demonstrate through these cases how this syndromic diagnostic test could influence the management of PICU patients.

Conclusions

Respiratory infection is frequently encountered in the PICU, but a bacterial or fungal cause is often not identified due to the limitations of microbiological cultures. Use of a custom TAC can deliver rapid and sensitive results in mechanically ventilated children with suspected LRTI. If demonstrated to be reliable and cost-effective in a dedicated paediatric randomised control trial, TAC could become a routine adjunct to microbiological cultures in the PICU.

Acknowledgments

We wish to thank the families, children and young people who kindly agreed to be involved in this case report.

Funding: This project was funded by the Addenbrooke’s Charitable Trust, Cambridge University Hospitals (No. 900240 to J.A.C., N.P., and S.B.), in addition to the NIHR Cambridge Biomedical Research Centre; the Gates Cambridge Trust (No. OPP1144 to J.A.C.); Wellcome Trust (No. 215515 to S.B.); Wellcome Trust Clinical Research Career Development Fellowship (No. WT 2055214/Z/16/Z to A.C.M.) and Medical Research Council (MRC) Clinician Scientist Fellowship (No. MR/V006118/1 to A.C.M.); and Action Medical Research (to N.P., S.B.) (Nos. GN2751 and GN2903). This work was also supported, in whole or in part, by the Bill & Melinda Gates Foundation (No. OPP1144). Under the grant conditions of the Foundation, a Creative Commons Attribution 4.0 Generic License has already been assigned to the Author Accepted Manuscript version that might arise from this submission.

Footnote

Reporting Checklist: The authors have completed the CARE reporting checklist. Available at https://tp.amegroups.com/article/view/10.21037/tp-23-525/rc

Peer Review File: Available at https://tp.amegroups.com/article/view/10.21037/tp-23-525/prf

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tp.amegroups.com/article/view/10.21037/tp-23-525/coif). J.A.C. received PhD personal funding through the Gates Cambridge Trust and funding for the consumables for the research were provided by the Addenbrooke’s Charitable Trust, Cambridge University Hospitals. T.G. receives a salary from the NIHR Cambridge Biomedical Research Campus and participates in monitoring the PROMISE study. A.C.M. receives support from the Medical Research Council Clinician Scientist Fellowship and Wellcome Trust Clinical Research Career Development Fellowship, and has received speaker fees from BioMérieux, Boston Scientific and Fisher and Paykel. M.D.C. is the inventor of a patent held by the Secretary of State for Health (UK government) EP2788503, which covers some of the genetic sequences used in this study. V.N. is a founder, director, and shareholder in Cambridge Infection Diagnostics (CID), which is a commercial company aimed at developing molecular diagnostics in infection and antimicrobial and AMR stewardship. A.C.M. and S.B. are advisory board members of the CID. S.B. has received funding from Addenbrooke’s Charitable Trust, Cambridge University Hospitals, Wellcome Trust and Action Medical Research. N.P. has received funding from Action Medical Research, the Addenbrooke’s Charitable Trust, Cambridge University Hospitals and the Cambridge NIHR Biomedical Research Centre, and has received speaker fees from BioMérieux. The other authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All procedures performed in the study were in accordance with the ethical standards of the institutional and/or national research committee(s) and with the Helsinki Declaration (as revised in 2013). Written informed consent was obtained from the patients’ legal guardians for publication of this case report. A copy of the written consent is available for review by the editorial office of this journal.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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