Culture-negative Infections Research Articles

P-2120. Clinical significance of 16S rRNA sequencing identification of Legionella species

Abstract Background Incorporation of 16S rRNA sequencing into clinical diagnostics has improved the ability of clinicians to diagnose fastidious bacteria and facilitates the identification of causative organisms in culture-negative infections. However, its clinical utility is unclear, with estimates of antimicrobial management changes from 16S identification ranging from only 4-15%. Legionella species are fastidious bacteria that can be difficult to identify without prior clinical suspicion, and thus can cause culture-negative infections that lead to 16S sequencing. However, Legionella are found ubiquitously in aquatic environments and its identification on 16S could represent a clinically significant infection, a nonpathogenic sample isolate, or a contaminant from sample processing or analysis. In this study, we reviewed 16S Legionella species results from our institution and determined whether they represented a clinical infection. Methods 16S results from our institution were queried for Legionella species and a retrospective analysis of the resulting 10 cases was conducted. Multiple factors from the cases were assessed including demographics, comorbidities, type of 16S specimen, and antibiotic regimen change in response to 16S results. Additionally, we identified whether the 16S results represented clinically significant infections or contaminants. Results 70% of the Legionella identifications were indicative of a clinically relevant infection. Specimens from a pulmonary source were much more likely to represent a true infection, while identifications from extrapulmonary sources were more likely to be a contaminant. The species of Legionella identified on 16S did not impact the likelihood of a true causative agent. The 16S results affected antimicrobial regiments in 60% of the cases and were often the only diagnostic test to point towards Legionella as a causative agent. Conclusion Legionella species identification on 16S can be an important diagnostic clue to the etiology of a patient's infection, but various clinical factors should be taken into account when assessing whether the result represents a true infection or contaminant. Disclosures All Authors: No reported disclosures

P-2160. Where Does Karius Testing Have a Clinical Impact? A Retrospective Analysis of Next Generation Sequencing

Abstract Background Karius testing, a commercially available microbial cell-free DNA assay, has emerged as a new diagnostic tool for clinicians, particularly for management of culture negative infections and in immunocompromised patients. There remains uncertainty regarding the clinical impact of next-generation sequencing in patient management. Figure 1 Methods This was a retrospective multi-site observational cohort study over a five-year period from 2018 to 2023 at a community hospital system. Patients were included if they had a Karius test ordered with results available to treating clinicians during the time period. The primary outcome was a change in clinical management defined as either the addition or removal of antibiotics, based on results from Karius testing. Results 391 patients met inclusion criteria, with 283 (72%) in immunocompetent and 108 (28%) in immunocompromised patients. An infectious diseases (ID) consult was obtained in 355 (91%) cases. Fifty-two percent of Karius tests were positive, with an average of 1.1 pathogens reported. Concordance rate between positive cultures and Karius results was 12%. Pneumonia (24%), fever of unknown origin (FUO) (18%), and endocarditis (17%) were the most common indications for Karius orders (Fig 1). Testing changed management in 19% of all included individuals, and in 18% of immunocompromised patients. Karius testing was most impactful in neutropenic fever and endocarditis with 27% and 22% respectively changing management and least impactful when ordered for joint infection, where 12% resulted in a change in management (Fig 2). Conclusion The Karius test changed antibiotic treatment in 19% of patients. It was most clinically relevant in neutropenic fever and endocarditis management. Even with our sizeable sample population including both immunocompromised and immunocompetent patients across a large, multi-site community hospital system, findings remained consistent compared to other smaller studies. Further investigation would be beneficial to determine how best to incorporate this emerging tool in clinical practice. Disclosures All Authors: No reported disclosures

Is the Presence of Sinus Tract in Periprosthetic Joint Infection Still One of the Main Deciding Factors for Septic Revision?

Introduction Two of the most common complications of joint arthroplasty surgery are aseptic and septic loosening. While aseptic loosening has a well-established treatment protocol, and diagnosis is quite straightforward, bacterial colonization of the implants is associated with a more difficult diagnosis and treatment, more surgeries, and higher morbidity for the patient. Accurate diagnosis is essential in choosing the right treatment plan. The aim of the current study was to assess the current diagnostic methods for periprosthetic joint infection and the influence of clinical signs like sinus tract on the treatment algorithm and outcomes of the patients.We wanted to highlight that sinus tract is still one of the major criteria in periprosthetic joint infection diagnosis and its presence increases the probability of choosing the right therapeuticoption. Materials and methods During the three-year period of the study, we included 48 cases of patients who presented in our hospital with pain around their hip or knee prostheses. Inclusion criteria were patients diagnosed with septic or aseptic loosening of the prosthesis that required surgical revision of the implant in one stage or two stages. We excluded patients who did not require surgery yet or had major contraindications for revision surgery, patients who refused surgery, acute periprosthetic joint infections (less than 1 month since implantation), or extrinsic mechanical complications of the prosthesis like periprosthetic fractures, implant dislocations. Results Out of 48 patients, 25 underwent one-stage revision and 23 underwent two-stage revision surgery (septic revision). In the subgroup of two-stage revision, 18 patients (78.2%) presented a sinus tract communicating with the prosthesis, this clinical sign being a major characteristic of the subgroup. We managed to successfully identify 21 out of 23 cases (91.3%) of periprosthetic joint infections prior to or during the surgery. In the two cases in which we misdiagnosed the infection, the sinus tract or a positive bacterial culture was absent prior to surgery, in addition to other clinical or paraclinical findings indicating only a small probability of periprosthetic joint infection, influenced the attending medical doctor's therapeuticdecision. In these particular cases of culture-negative periprosthetic joint infections, the outcome was poor, with patients needing additional surgeries in order to eradicate the infection. Discussion When present, a clear sign of periprosthetic joint infection, such as a sinus tract, facilitates the diagnostic protocol and allows the medical staff to initiate the appropriate treatment earlier. In the absence of such obvious signs, differential diagnosis remains difficult, and we should consider the future development of faster, cheaper, and more accurate tests for periprosthetic joint infectiondiagnosis, especially for chronic low-grade infections that could be easily misdiagnosed.

Reactive arthritis

Reactive arthritis (ReA) is adisease caused by an extra-articular infection that manifests as asterile joint inflammation. In contrast to bacterial septic arthritis no pathogens can be cultured from the joint in ReA but pathogen components, such as antigens or DNA are more frequently detectable in the joint, suggesting an intra-articular culture-negative persistent infection or at least an intra-articular interaction between the host and pathogen components. The primary extra-articular infection in classical ReA is of bacterial origin and usually affects either the urogenital, gastrointestinal or, less frequently, the respiratory tract. Chlamydia (C.trachomatis and less frequently C.pneumoniae) and enterobacteria are among the most common pathogens causing ReA. The prevalence of ReA is estimated at 40/100,000 and the incidence at 5/100,000. Typical clinical manifestations are mostly self-limiting peripheral arthritis (monoarticular or oligoarticular), dactylitis and, more rarely, axial involvement and in half of the cases, there is an association with HLA-B27. Due to these similarities, classical ReA is categorized as aform of spondyloarthritis (SpA). The diagnosis is made on the basis of atypical clinical picture, evidence of aprevious or persistent infection and the exclusion of other causes of arthritis. Treatment includes physical measures, the use of anti-inflammatory agents such as nonsteroidal anti-inflammatory drugs (NSAID) or glucocorticoids, in the case of persistent arthritis, immunomodulating substances such as sulphasalazine, methotrexate and in individual cases biologics and Janus kinase inhibitors (JAKi) are used. In general, antibiotic treatment of ReA does not shorten the duration of the disease.

Emergence and Rapid Diagnosis of Talaromyces marneffei Infections in Renal Transplant Recipients by Next-Generation Sequencing

In the last few years, next-generation sequencing (NGS) has emerged as a technology for laboratory diagnosis of many culture-negative infections and slow-growing microorganisms. In this study, we describe the use of metagenomic NGS (mNGS) for rapid diagnosis of T. marneffei infection in a 37-year-old renal transplant recipient who presented with chronic pneumonia syndrome. Bronchoalveolar lavage for mNGS was positive for T. marneffei sequence reads. Prolonged incubation of the bronchoalveolar lavage revealed T. marneffei colonies after 6 days of incubation. Analysis of 23 cases of T. marneffei infections in renal transplant recipients from the literature revealed that the number of cases ranged from 1 to 4 cases per five years from 1990 to 2020; but increased rapidly to 9 cases from 2021 to 2023, with 7 of them diagnosed by NGS. Twenty of the 23 cases were from T. marneffei-endemic areas [southern part of mainland China (n = 9); Hong Kong (n = 4); northeastern India (n = 2); Indonesia (n = 1) and Taiwan (n = 4)]. For the 3 patients from non-T. marneffei-endemic areas [United Kingdom (n = 2) and Australia (n = 1)], they had travel histories to China and Vietnam respectively. The time taken for diagnosis by mNGS [median 1 (range 1 to 2) day] was significantly shorter than that for fungal culture [median 6 (range 3 to 15) days] (P = 0.002). mNGS is useful for picking up more cases of T. marneffei infections in renal transplant recipients as well as providing a rapid diagnosis. Talaromycosis is an emerging fungal infection in renal transplant recipients.

Usefulness of next-generation sequencing for laboratory diagnosis of rickettsiosis.

Rickettsiosis includes a diversity of culture-negative non-specific systemic infections. Laboratory diagnosis of rickettsiosis is often not easy. In this 12-month study, six patients with a variety of rickettsia infections of the spotted fever group, typhus group and scrub typhus were diagnosed directly or indirectly by metagenomic next-generation sequencing (mNGS). The patient with Japanese spotted fever was rapidly made when mNGS analysis of the patient's blood revealed Rickettsia japonica sequences. For the two patients with Rickettsia felis chest infections, the bacterium was detected in the bronchoalveolar lavage of one case and lung biopsy of the other. Both patients had underlying malignancies, carcinoma of the breast and carcinoma of the lung respectively, and were on chemotherapy with immunosuppressive effect. For the remaining three patients who presented over a period of 13 weeks, all had fever, headache and the typical eschar. They also had increased serum transaminases and responded promptly to doxycycline. However, the Weil-Felix test results of all three patients were negative. Since we considered the three cases typical of rickettsiosis, we submitted their serum samples for mNGS analysis. Results showed that Orientia tsutsugamushi sequences were present in the serum of one case. In view of the positive mNGS results, we repeated the Weil-Felix test for the residual sera of all three patients and it revealed that those of the other two cases showed OX-19 titers of 1:640 and 1:160 respectively, inferring that these two patients probably had rickettsiosis of the typhus group. As for the patient positive for O. tsutsugamushi sequences, we also detected IgM for O. tsutsugamushi in the serum, which double confirmed that it was a case of scrub typhus. mNGS is an important molecular tool and can complement serology for laboratory diagnosis of rickettsiosis.

Gram-Negative Colonization and Bacterial Translocation Drive Neonatal Sepsis in the Indian Setting

BackgroundThe gut microbiota, comprising billions of microorganisms, plays a pivotal role in health and disease. This study aims to investigate the effect of sepsis on gut microbiome of neonates admitted to the Neonatal Intensive Care Unit.MethodsA prospective cohort study was carried out in the NICU of tertiary care hospital in Karnataka, India, from January 2021 to September 2023. Preterm neonates with birth weight < 1500 g and gestational age < 37 weeks were recruited, excluding those with congenital gastrointestinal anomalies, necrotizing enterocolitis, or blood culture-negative infections. The study population was divided into three groups: healthy neonates (Group A), neonates with drug-sensitive GNB sepsis (Group B), and neonates with pan drug-resistant GNB sepsis (Group C). Stool samples were collected aseptically, snapped in liquid nitrogen, and stored at -80⁰C for extraction of DNA and microbiome analysis.ResultsThe gut microbiota of healthy neonates (Group A) was dominated by Proteobacteria (24.04%), Actinobacteria (27.13%), Firmicutes (12.74%), and Bacteroidetes (3%). Predominant genera included Bifidobacterium (55.17%), Enterobacter (12.55%), Enterococcus (50.69%), Streptococcus (7.92%), and Bacteroides (3.58%).Groups B and C, the microbiota exhibited higher Proteobacteria abundance (57.16% and 66.58%, respectively) and reduced diversity of beneficial bacteria. Notably, the presence of sepsis was associated with an increase in pathogenic bacteria and a decrease in beneficial commensal bacteria.ConclusionNeonates with sepsis exhibited significant gut microbiome dysbiosis, characterized by increased Proteobacteria and reduced beneficial bacteria diversity. These findings highlight the potential of microbiome profiling as a diagnostic tool and underscore the importance of gut microbiota modulation in managing neonatal sepsis.

Can pre-analytical procedures improve microbiological culture yield in patients with periprosthetic infections?

BackgroundThe detection of causative pathogens plays a crucial role in the diagnosis and targeted treatment of periprosthetic joint infections (PJI). While there have been improvements in analytic methods in the past, pre-analytical procedures have not yet been sufficiently investigated. The objective of this study was to compare the culture yield of four different pre-analytical procedures.MethodsPatients with perioperative diagnosis of PJI were included in a single center cross-sectional study (2021–2022). Tissue samples (n = 20) of each patient were randomly and equally distributed to each of the four study arms. Tissue samples were either send to the laboratory without culture medium (group A) or were transported in thioglycolate medium immediately after sampling at three different temperatures (room temperature, 4 °C, 37° for 24 h; group B-D). Culture media were investigated for growth on days 1, 3, 7, 12, 14. All organisms, the number of positive samples and the time to positivity were recorded and compared between the study arms. Single positive cultures were considered as contamination.ResultsIn total, 71 patients were included. The proportions of culture negative samples (10–15%) and polymicrobial infections (51–54%) were comparable between the four arms. Seven patients (10%) were culture-negative in group A, but showed growth in thioglycolate media (group B-D). Furthermore, 13% of patients showed growth in all groups, but additional organisms were cultured in thioglycolate. There was growth beyond day 7 of culturing only in thioglycolate, but not in group A. A storage temperature of 4 °C showed a longer time to positivity compared to the other groups (p < 0.001).ConclusionsPre-analytical storage of tissue samples in thioglycolate broth did not improve the culture yield and did not detect additional cases of infection compared to the standard (pre-analytical storage in sterile containers). However, including a thioglycolate medium to the sampling algorithm reduced the rate of culture-negative infections and helped to identify additional organisms.

Evaluation of 16S-Based Metagenomic NGS as Diagnostic Tool in Different Types of Culture-Negative Infections.

Bacterial infections pose significant global health challenges, often underestimated due to difficulties in accurate diagnosis, especially when culture-based diagnostics fail. This study assesses the effectiveness of 16S-based metagenomic next generation sequencing (NGS) for identifying pathogens in culture-negative clinical samples across various medical settings. Overall, 48% of samples were collected from orthopedics, 15% from neurosurgery, and 12% in cardiac surgery, among others. The detection rate of monomicrobial infections was 68.6%, and 5.7% for polymicrobial infections. In addition, NGS detected bacteria in all samples from the lungs, head and neck, and eye specimens. Cutibacterium acnes (11%, 12/105) was the most frequent microorganism, followed by Staphylococcus epidermidis (10.4%, 11/105), and Staphylococcus aureus (9.5%, 10/105). In conclusion, 16S-targeted metagenomic sequencing enhances pathogen detection capabilities, particularly in instances where traditional cultures fail. By the combination of NGS and bacterial cultures, microbiologists might provide a more accurate diagnosis, guiding more effective treatments and potentially reducing healthcare costs associated with empirical treatments.

Periprosthetic joint infection in patients with rheumatoid arthritis: case series

Introduction The differential diagnosis of periprosthetic joint infection (PJI) is challenging in patients with systemic diseases due to identical clinical and laboratory patterns and activity of the inflammatory process.The objective was to evaluate the diagnostic data and results of debridement of PJI in patients with rheumatoid arthritis using a case series.Material and methods A retrospective analysis of surgical treatment of PJI was produced in patients with rheumatoid arthritis between 2014 and 2022. PJI was verified based on ICM criteria. A poor outcome included the presence of clinical and laboratory signs of infection on admission to the second stage of treatment and recurrence after successful debridement.Results Among the 524 cases of PJI, 35 (6.7 %) were patients with rheumatoid arthritis with 48.6 % receiving antibiotics prior to admission. Culture-negative infection was recorded in 38.4 %. PJI was not confirmed in five cases (14.3 %). High average values of inflammatory markers were registered in the blood (ESR, CRP and D-dimer) before and after debridement; decreased ESR and leukocyte count in the synovial fluid was statistically significant. Favorable outcomes were obtained in 82.9 % of cases at mid term with every fifth patient treated with a spacer or arthrodesis.Discussion The incidence of culture-negative infection in patients with systemic diseases was reported as much as 27–37 %. A systematic review of the literature showed that the percentage of band neutrophils in synovial fluid has a sensitivity of 95.2 % and a specificity of 85.0 %, with an optimal threshold of 78 % sufficient to verify infection. The poor outcomes we identified resulted from two- or three-stage surgical treatment. Other authors reported better outcomes with two-stage debridement.Conclusion Culture-negative infection was common in cases of PJI observed in patients with rheumatoid arthritis. Favorable outcomes were seen mostly with two-stage surgical treatment. Inflammatory markers ESR, CRP and D-dimer did not reach normal values during diagnosis and treatment of infection indicating the inapplicability of standard diagnostic criteria for PJI in patients with rheumatoid arthritis.

Development and external validation of a model to predict multidrug-resistant bacterial infections in patients with cirrhosis.

We included patients with cirrhosis and bacterial infections from two prospective studies: a transcontinental study was used for model development and internal validation (n = 1302), and a study from Argentina and Uruguay was used for external validation (n = 472). All predictors were measured at the time of infection. Both culture-positive and culture-negative infections were included. The model was developed using logistic regression with backward stepwise predictor selection. We externally validated the optimism-adjusted model using calibration and discrimination statistics and evaluated its clinical utility. The prevalence of MDRO infections was 19% and 22% in the development and external validation datasets, respectively. The model's predictors were sex, prior antibiotic use, type and site of infection, MELD-Na, use of vasopressors, acute-on-chronic liver failure, and interaction terms. Upon external validation, the calibration slope was 77 (95% CI .48-1.05), and the area under the ROC curve was .68 (95% CI .61-.73). The application of the model significantly changed the post-test probability of having an MDRO infection, identifying patients with nosocomial infection at very low risk (8%) and patients with community-acquired infections at significant risk (36%). This model achieved adequate performance and could be used to improve the selection of empiric antibiotics, aligning with other antibiotic stewardship program strategies.

The Impact of Culture Negativity on the Outcomes of Revision Total Knee Arthroplasty for Chronic PJI.

Culture-positive (CP) and culture-negative (CN) periprosthetic joint infections (PJI) remain a crucial area of research; however, current studies comparing these infections rely on unstandardized outcome reporting tools. Our study aimed to compare the outcomes of two-stage revision of CP and CN PJI using the standardized Musculoskeletal Infection Society (MSIS) outcome reporting tool. We retrospectively reviewed 138 patients who were diagnosed with PJI and indicated for two-stage revision total knee arthroplasty (rTKA). The majority of patients in both CP and CN cohorts achieved infection control without the need for reoperation (54.1% and 62.5%, respectively). There was a significant difference in the overall distribution of MSIS outcomes (p = 0.043), with a significantly greater rate of CN patients falling into Tier 1 (infection control without the use of suppressive antibiotics) (52.5% versus 29.6%, p = 0.011). There was also a significant difference in the distribution of septic versus aseptic reoperations after 2nd stage (p = 0.013), with more CP reoperations being septic and more CN reoperations being aseptic. The duration from first to second stage was significantly shorter in the CN cohort (p = 0.002). While overall infection control was similar between cohorts, these data suggest that the outcomes of two-stage rTKA are favorable in cases of CN PJI.

Seasonal variation and preoperative risk factors for polymicrobial infection following open fracture

IntroductionDespite modern approaches to open fracture management, fracture-related infection (FRI) rates remain high. Recent studies demonstrated the seasonal and regional variation of causative organisms in FRI. This study aims to better understand the causative organisms and identify preoperative risk factors for the primary outcome of FRIs at a Level I trauma center. Materials and methodsThis retrospective cohort study examined all patients that underwent irrigation and debridement of an open fracture at a single Level I trauma center between 2007 and 2019. Exclusion criteria included gunshot wounds, hand injuries, and follow-up less than 3 months. Patients that developed FRI were compared by season, injury characteristics, patient demographics, initial management, and causative organisms. ResultsAmong 695 patients with open fractures, 78 patients (11.2 %) developed infection, of which eight were Gustilo-Anderson (GA) Type I, 16 were GA Type II, 25 were GA Type IIIA, 26 were GA Type IIIB, and three were GA Type IIIC. Gram-positive FRIs were most common (81.1 %), followed by 56.8 % polymicrobial, 54.1 % gram-negative, and 10.1 % culture-negative infections. More than half (55.1 %) of the infections were from open tibial fractures and occurred after a motorcycle (32.1 %) or motor vehicle collision (23.1 %). Patients were more likely to have high FRI rates in the summer (12.8 %, n = 29) and fall (15.8 %, n = 32) in comparison of spring (4.7 %, n = 7) and winter (8.5 %, n = 10) (p < 0.01). Staphylococcus infections were more common in fall and winter (73.8 %, n = 31) versus spring and summer (44.4 %, n = 16) (p = 0.01). Patients that were transferred from outside hospitals had significantly higher rates of polymicrobial infection when compared to those who arrived from the field (63.6 % vs 41.2 %, p = 0.03). No differences were observed in infection causative organisms based on GA type. ConclusionsTwo preoperative risk factors for polymicrobial infection following open fracture include inter-hospital transfers and warm/humid weather. Broadening antibiotic prophylaxis during spring/summer months or for transferred patients may enhance antibiotic coverage and reduce infections.

Culture-negative periprosthetic joint infections: Do we have an issue?

Culture-negative periprosthetic joint infection (PJI) poses a significant challenge in clinical settings. The lack of information on causative organism(s) leads to uncertainties regarding the choice of antimicrobial treatment, which can potentially adversely influence the outcome. Recent advances in molecular-based diagnostic methods have the potential to address the difficulties associated with culture-negative PJIs. These technologies offer a solution to the existing clinical dilemma by providing identification of pathogens and guiding appropriate antimicrobial treatment.In this narrative review, we provide information regarding: 1) incidence and risk factors for culture-negative PJI; 2) the optimal antimicrobial therapy and duration of treatment for culture-negative PJI; 3) outcome comparison between culture-positive and culture-negative PJI; and 4) utilization of novel molecular diagnostic methods in culture-negative PJI, including pathogen identification, and the implementation of an antibiotic stewardship program.

Efficacy and safety of two-stage revision for patients with culture-negative versus culture-positive periprosthetic joint infection: a single-center retrospective study

BackgroundThe safety and efficacy of two-stage revision for culture-negative PJI remain controversial. This study analyzed outcomes after two-stage revision in patients with culture-negative and culture-positive periprosthetic joint infection (PJI) during follow-up lasting at least two years.MethodsData were retrospectively analysed patients who underwent hip or knee revision arthroplasty from January 2008 to October 2020 at our medical center. The primary outcome was the re-revision rate, while secondary outcomes were the rates of reinfection, readmission, and mortality. Patients with culture-negative or culture-positive PJI were compared in terms of these outcomes, as well as survival time without reinfection or revision surgery, based on Kaplan‒Meier analysis.ResultsThe final analysis included 87 patients who were followed up for a mean of 72.3 months (range, 24–123 months). The mean age was 58.1 years in the culture-negative group (n = 24) and 59.1 years in the culture-positive group (n = 63). The two groups (culture-negative versus culture-positive) did not differ significantly in rates of re-revision (0.0% vs. 3.2%, p > 0.05), reinfection (4.2% vs. 3.2%, p > 0.05), readmission (8.4% vs. 8.0%, p > 0.05), or mortality (8.3% vs. 7.9%, p > 0.05). They were also similar in survival rates without infection-related complications or revision surgery at 100 months (91.5% in the culture-negative group vs. 87.9% in the culture-positive group; Mantel‒Cox log-rank χ2 = 0.251, p = 0.616).ConclusionThe two-stage revision proves to be a well-tolerated and effective procedure in both culture-negative and culture-positive PJI during mid to long-term follow-up.

Plasma Microbial Cell-Free DNA Sequencing for Pathogen Detection and Quantification in Children With Musculoskeletal Infections.

Nearly half of all pediatric musculoskeletal infections (MSKIs) are culture negative. Plasma microbial cell-free DNA (mcfDNA) sequencing is noninvasive and not prone to the barriers of culture. We evaluated the performance of plasma mcfDNA sequencing in identifying a pathogen, and examined the duration of pathogen detection in children with MSKIs. We conducted a prospective study of children, aged 6 months to 18 years, hospitalized from July 2019 to May 2022 with MSKIs, in whom we obtained serial plasma mcfDNA sequencing samples and compared the results with cultures. A pathogen was recovered by culture in 23 of 34 (68%) participants, and by initial mcfDNA sequencing in 25 of 31 (81%) participants. Multiple pathogens were detected in the majority (56%) of positive initial samples. Complete concordance with culture (all organisms accounted for by both methods) was 32%, partial concordance (at least one of the same organism(s) identified by both methods) was 36%, and discordance was 32%. mcfDNA sequencing was more likely to show concordance (complete or partial) if obtained prior to a surgical procedure (82%), compared with after (20%), (RR 4.12 [95% CI 1.25, 22.93], p = .02). There was no difference in concordance based on timing of antibiotics (presample antibiotics 60% vs no antibiotics 75%, RR 0.8 [95% CI 0.40, 1.46], p = .65]). mcfDNA sequencing was positive in 67% of culture-negative infections and detected a pathogen for a longer interval than blood culture (median 2 days [IQR 1, 6 days] vs 1 day [1, 1 day], p < .01). Plasma mcfDNA sequencing may be useful in culture-negative pediatric MSKIs if the sample is obtained prior to surgery. However, results must be interpreted in the appropriate clinical context as multiple pathogens are frequently detected supporting the need for diagnostic stewardship.

Relevance of cardiac imaging in the evolving landscape of infective endocarditis management.

Infective endocarditis (IE) is an increasingly recognized condition with high morbidity. Patients with atypical symptoms, culture-negative infections, and prosthetic cardiac devices and implants represent challenging populations to evaluate and manage. Recent major society guidelines have recommended the appropriate incorporation of multimodality imaging in the evaluation of these more complex IE cases. This article draws on the available literature regarding the different cardiac imaging modalities and discusses the role of multimodality imaging in IE.

Implementation of full-length 16S nanopore sequencing for bacterial identification in a clinical diagnostic setting

This study describes the implementation of 16S nanopore sequencing in a diagnostic lab for pathogen identification without prior enrichment. First, the universality of the test and taxonomic resolution was evaluated for 78 clinically relevant bacteria (69 known and 9 unknown bacterial cultures). Next, the diagnostic value of the test was evaluated based on clinical samples. It was shown that 16S sequencing can be used both for identification of unknown cultures and to find bacteria directly in the clinical sample without cultivation. All culture-positive samples (n=11) tested positive with 16S sequencing directly performed on the sample, but bacteria were found as well in 15/30 culture-negative samples. Pathogenic bacteria were found in a background of commensal flora, and even complex polymicrobial infections could be unraveled. This study demonstrates the feasibility of implementing 16S nanopore sequencing in a clinical diagnostic setting and demonstrates its value for the diagnosis of culture-negative and polymicrobial infections.

Bacterial bone and joint infections in children with leukaemia or following haematopoietic cell transplant: A systematic review of published cases

BackgroundBone and joint infections (BJI) are common in immunocompetent children, however in children with acute leukaemia the epidemiology, optimal diagnostic pathway and medical and surgical treatment is poorly described. We review the presentation, management and outcomes of BJI in this special population. MethodsSystematic literature review of BJI in children with acute leukaemia was performed using Medline, EMBASE and PubMed databases from 1946 to June, 2023. Titles and abstracts were independently screened by 3 reviewers. Studies were included if they reported data on children (≤18 years) with leukaemia who had a diagnosis of osteomyelitis (radiological diagnosis) or septic arthritis (culture proven or managed as infection by treating clinician). ResultsTwenty-two articles describing 33 episodes of confirmed or suspected bacterial BJI were included. Of these, microbiological diagnosis was confirmed in 27 episodes (82%) and included Gram-positive (9/33), Gram-negative (8/33), mycobacterial (5/33), Mycoplasmatota (1/33) and polymicrobial (4/33). The femur was the most common site of infection (33%) and there were 18 episodes (55%) of multifocal disease. Duration of treatment varied from 3 weeks to 2.5 years in patients with culture confirmed infection (82%) and from 3 to 8 weeks in patients with culture-negative infection (18%). Clinical cure was documented in 31 episodes (94%). ConclusionBone and joint infections in children with acute leukaemia are often caused by atypical pathogens, with a higher predilection for multifocal disease and/or involvement of unusual sites in comparison with immunocompetent children. It is unclear whether this population requires prolonged antibiotic treatment due to periods of intensive chemotherapy or neutropenia. Monitoring response to therapy may be challenging as inflammatory markers such as CRP and ESR may take longer to normalise, making it difficult to assess for clinical cure.

Comparison of Outcomes in Neonates Receiving Cefepime or Ceftazidime.

Cefepime and ceftazidime are alternatives to cefotaxime for management of Gram-negative infections in neonates. The objective was to evaluate neonatal outcomes when receiving cefepime or ceftazidime. This was a single center, retrospective analysis of neonates exposed to at least 24 hours of cefepime or ceftazidime between June 1, 2018, and June 1, 2021. The primary outcome was incidence of culture-positive, late-onset sepsis after initial exposure. Secondary outcomes included culture-negative, respiratory, urinary tract, and resistant infections; necrotizing enterocolitis; length of stay; age at discharge; mortality; and adverse effects. A total of 105 neonates were included (cefepime, n = 50; ceftazidime, n = 55). Baseline characteristics were similar except more cumulative days of antibiotics (25.0 [IQR, 9.3-47.0] versus 9.0 [IQR, 4.0-23.5], p = 0.01), central line days (11.0 [IQR, 6.0-40.0] versus 6.5 [IQR, 0.0-11.5], p = 0.001), and ventilator days (13.0 [IQR, 2.3-48.0] versus 4.0 [IQR, 0.0-25.0], p = 0.02) were found in the cefepime group than in the ceftazidime group. There was no difference in culture-positive sepsis after the initial antibiotic course (8.0% versus 3.6%, p = 0.42). Statistical differences were seen in select secondary outcomes including treated respiratory infections (16.0% versus 1.8%, p = 0.01), length of stay greater than 30 days (72.0% versus 50.9%, p = 0.03), and mortality (26.0% versus 9.1%, p = 0.02). These differences were not observed in analyses adjusted for ventilator days. This analysis found no difference in culture-positive sepsis in neonates exposed to cefepime versus ceftazidime. Moreover, there were no differences in secondary outcomes in adjusted analyses. Further research is needed to assess neonatal outcomes in a larger analysis.