Culture-negative Infections Research Articles

Metagenomic Diagnosis for a Culture-Negative Sample From a Patient With Severe Pneumonia by Nanopore and Next-Generation Sequencing.

Rapid and accurate etiologic diagnosis accelerates targeted antimicrobial therapy. Metagenomic analysis has played a critical role in pathogen identification. In this study, we leveraged the advantages of both the MinION and BGISEQ-500 platforms to make a bacteriologic diagnosis from a culture-negative lung tissue sample from an immunocompromised patient with severe pneumonia. Real-time nanopore sequencing rapidly identified Klebsiella pneumoniae by an 823 bp specific sequence within 1 min. Genomic analysis further identified blaSHV-12, blaKPC-2, blaTEM-1, blaCTX-M-65, and other resistance genes. The same sample was further sequenced on the BGISEQ-500 platform, which presented consistent results regarding the most top dominant pathogens and provided additional information of resistance genes. Revised antibiotic treatment was followed by the patient's clinical recovery. Though sample preparation and the interpretation of final results still need to be improved further, metagenomic sequencing contributes to the accurate diagnosis of culture-negative infections and facilitates the rational antibiotic therapy.

PCR research as an auxiliary method for diagnostics of periphresitical infection after endostreditision of joints (clinical case).

Periprosthetic infection (PPI) after arthroplasty of large joints is the third (among the main causes of unsatisfactory results of surgical treatment) a serious threat to the health of patients. The «gold standard» for the diagnosis of PPI is the bacteriological examination of samples of periprosthetic tissues and synovial fluid. In 10-30% of cases, it is impossible to isolate microorganisms, which is explained by the difficulty of cultivation and taking antibiotics before sampling. The purpose of study is to demonstrate the diagnostic value of PCR diagnostics for identifying the genetic material of an infectious pathogen of a culture-negative periprosthetic infection. Material of the study is a description of a clinical case of a culture-negative periprosthetic infection that caused a second two-stage revision of the hip joint prosthesis In the first episode of PPI that occurred 3 years after hip replacement, a microbiological examination of the puncture of the trochanteric zone of the operated joint revealed a massive increase in methicillin-resistant Staphylococcus epidermidis (MRSE). A two-stage revision joint replacement was performed. 5 years after the revision, the patient was hospitalized with clinical and radiological signs of PPI, while examining the puncture of the joint revealed characteristic PPI cytosis. Microbiological examination of punctate and intraoperative aspirate at the first stage of the repeated two-stage revision endoprosthesis replacement did not reveal aerobic and anaerobic microorganisms. In PCR studies, the DNA of methicillin-sensitive Staphylococcus aureus (MSSA) was detected in washouts from the removed components of the endoprosthesis; no resistance marker (mecA gene) was found. Given the concomitant oncological disease, this result determined the appointment of pathogenetic antibiotic therapy, the effectiveness of which was confirmed after 8 weeks at the II stage of revision. The PCR study of joint and trochanteric punctures (before surgery), flushing from the removed spacer components (after ultrasound treatment) and intraoperative aspirate from the joint did not reveal Staphylococcus aureus DNA and resistance marker (mecA gene). In some cases of periprosthetic infection, traumatologists and orthopedists deal with culturally negative results of a microbiological study of the patient's biomaterial and swabs from the components of endoprostheses in the presence of clinical manifestations of PPI, confirmed by laboratory diagnostics and X-ray examination. According to the literature, such clinical situations are observed in 10-30% of cases and are caused by previous antibiotic therapy in the early stages of an infectious complication. After surgical treatment of PPI for the selection of adequate antibiotic therapy, such patients need to at least indirectly determine the type of infection pathogen, which is achieved by the use of additional diagnostic methods, such as a PRC study. In the case described by us, after a course of antibiotic therapy, prescribed according to the results of the first PCR study, the patient's body does not contain DNA traces of the desired infectious agent. Thus, the repeated PCR not only confirmed the accuracy of the initial diagnosis of the source of infection, but also further illustrated the success of the rehabilitation of the periprosthetic infection using a correctly selected antibacterial drug at the previous stage of the study. The use of the PCR method made it possible to diagnose the pathogen and prescribe adequate antibiotic therapy for culture-negative periprosthetic infection.

Can metagenomic next-generation sequencing identify the pathogens responsible for culture-negative prosthetic joint infection?

BackgroundThe aims of this study were to (1) evaluate the efficacy and safety of targeted antibiotics for the treatment of culture-negative prosthetic joint infection based on metagenomic next-generation sequencing results and (2) verify the accuracy and reliability of metagenomic next-generation sequencing for identifying pathogens related to culture-negative prosthetic joint infection.MethodsNinety-seven consecutive PJI patients, including 27 patients with culture-negative prosthetic joint infection, were treated surgically at our center. Thirteen of the 27 culture-negative prosthetic joint infection patients, who were admitted before June 2017 and treated with empirical antibiotics, comprised the empirical antibiotic group (EA group), and the other 14 patients, who were admitted after June 2017 and treated with targeted antibiotics according to their metagenomic next-generation sequencing results, were classified as the targeted antibiotic group (TA group). The short-term infection control rate, incidence of antibiotic-related complications and costs were compared between the two groups.ResultsTwo of the patients in the EA group experienced debridement and prolonged antimicrobial therapy due to wound infection after the initial revision surgery. No recurrent infections were observed in the TA group; however, no significant difference in the infection control rate was found between the two groups (83.33% vs 100%, P = 0.217). More cases of antibiotic-related complications were recorded in the EA group (6 cases) than in the TA group (1 case), but the difference was not statistically significant (P = 0.0697). The cost of antibiotics obtained for the EA group was 20,168.37 Yuan (3236.38–45,297.16), which was higher than that found for the TA group (10,164.16 Yuan, 2959.54–16,661.04, P = 0.04).ConclusionsTargeted antibiotic treatment for culture-negative prosthetic joint infection based on metagenomic next-generation sequencing results is associated with a favorable outcome, and metagenomic next-generation sequencing is a reliable tool for identifying pathogens related to culture-negative prosthetic joint infection.

Heightened Immune Response and Increased Risk of Infections in Pediatric Fontan Patients after Heart Transplantation

Infectious complications are a major cause of morbidity & mortality after heart transplantation (HT). Patients who have undergone Fontan palliation may have a higher susceptibility to post HT infections given thymectomy in infancy and known depletions of T-cell subsets. This was a single-center, retrospective cohort analysis of pediatric patients undergoing HT for dilated cardiomyopathy (DCM) or failing Fontan physiology, who underwent post HT induction with anti-thymoglobulin (ATG). Repeat HTs and multi organ transplants were excluded. The primary endpoint was infection in the first 180 days post HT, defined as 1) positive blood, urine, or respiratory culture; 2) positive viral PCR (excluding CMV and EBV); 3) skin or wound infection; or 4) culture-negative infection if a full antibiotic course (≥ 5 days) was completed. Secondary endpoints included sensitivity to ATG as defined by 1) absolute lymphocyte (ALC) and CD3 counts after 5 doses; 2) the incidence of post transplant lymphoproliferative disorder (PTLD); and 3) rejection (>/= Grade 2R ACR or pAMR2) in the first 180 days post HT. From 1/2014 to 9/2019, 63 patients (30 Fontan, 33 DCM) underwent HT at a median of 15.4 (IQR 10.8, 20.1) and 11.7 (IQR 1.4, 13.6) years, respectively. The median total ATG received was 7.5 (IQR 6.7, 8.2) mg/kg vs 7.2 (IQR 4.7, 9.2) mg/kg (p=NS), respectively. The median CD3 [9 (IQR 3,14) vs 12 (IQR 6, 26); p=0.04, Figure 1A] and lymphocyte counts [172 (IQR 98, 354) vs 427 (IQR 205, 662); p=0.02] after ATG were lower in Fontan vs DCM patients. 28 patients (41%) developed at least one infection within 180 days after HT, with a higher rate of infection in Fontan patients (60% vs 24%, p=0.005; Figure 1B). There was no difference in the incidence of PTLD (10% vs 0%; p=0.1) or rejection (17% vs 27%; p=0.31) between Fontan vs DCM patients, respectively. Compared to DCM patients, Fontan patients have a more pronounced suppression of CD3 counts after induction, as well as a higher risk of infection.

The Impact of Antibiotic-Loaded Bone Cement on Antibiotic Resistance in Periprosthetic Knee Infections.

BackgroudAntibiotic-loaded bone cement (ALBC) is commonly used in total knee arthroplasty (TKA), especially among high-risk patients. While previous studies have reported on the efficacy of ALBC in reducing the rate of periprosthetic joint infection (PJI), its impact on antibiotic resistance has not been determined. The purpose of this study was to investigate antibiotic resistance among organisms causing PJIs after TKA in which ALBC was utilized.MethodsA retrospective review from December 1998 through December 2017 identified 36 PJIs that met inclusion criteria. Patients with culture-negative infection and unknown cement type were excluded. Patient characteristics, infecting organism, and antibiotic susceptibilities were recorded. ABLC included an aminoglycoside in all cases.ResultsThere was no difference in the type of PJI between the 2 groups. Staphylococcus species was the most commonly isolated, with 9 of 16 cases (56.3%) using non-ALBC and 14 of 20 (65.0%) cases using ALBC. Of those infected with Staphylococcus, there was no significant difference in antibiotic susceptibilities between groups. Overall, there were only 3 cases where the infecting organism was aminoglycoside resistant (standard cement, 1; ALBC, 2).ConclusionsThese results suggest that the use of ALBC does not increase the risk of antibiotic resistance or affect the pattern of infection, even as the use of ALBC continues to increase, particularly among high-risk patients.

Can We Optimize Antibiotic Use in Norwegian Neonates? A Prospective Comparison Between a University Hospital and a District Hospital.

Background: Worldwide, a large proportion of neonates are prescribed antibiotics without having infections leading to increased antimicrobial resistance, disturbance of the evolving microbiota, and increasing the risk of various chronical diseases. Comparing practice between different hospitals/settings is important in order to optimize antibiotic stewardship.Aim: To investigate and compare the potential for improved antibiotic stewardship in neonates in two Norwegian hospitals with different academic culture, with emphasis on antibiotic exposure in unconfirmed infections, treatment length/doses, CRP values and the use of broad-spectrum antibiotics (BSA). All types of infections were investigated, but the main focus was on early-onset sepsis (EOS).Methods: We conducted a prospective observational cohort study of antibiotic use in a Norwegian university hospital (UH) and a district hospital (DH), 2017. Unconfirmed infections were defined as culture negative infections that neither fulfilled the criteria for clinical infection (clinical symptoms, maximum CRP >30 mg/L, and treatment for at least 5 days).Results: Ninety-five neonates at the DH and 89 neonates at the UH treated with systemic antibiotics were included in the study. In total, 685 prescriptions (daily doses) of antibiotics were given at the DH and 903 at the UH. Among term and premature infants (≥ 28 weeks), 82% (75% at the UH and 86% at the DH, p = 0.172) of the treatments for suspected EOS were for unconfirmed infections, and average treatment length in unconfirmed infections was 3.1 days (both hospitals). Median dose for aminoglycoside was higher for term infants at the UH (5.96, 95% CI 5.02–6.89) compared to the DH (4.98, 95% CI 4.82–5.14; p < 0.001). At the UH, all prescriptions with aminoglycosides were gentamicin, while tobramycin accounted for 93% of all prescriptions with aminoglycosides at the DH.Conclusion: There is a potential for reduction in both antibiotic exposure and treatment length in these two neonatal units, and a systematic risk/observational algorithm of sepsis should be considered in both hospitals. We revealed no major differences between the UH and DH, but doses and choice of aminoglycosides varied significantly.

391. Septic Arthritis and Acute Hematogenous Osteomyelitis in Children: Disease-Specific Antibiogram and Implications for Treatment

BackgroundOsteomyelitis and septic arthritis are common pediatric musculoskeletal infections with potential to cause significant morbidity. Empiric antibiotic selection is made prior to the availability of microbiologic data. The aim of this study was to compare the epidemiology of osteoarticular infections (OAI) to antibiotic regimens and local antibiograms.MethodsA retrospective study was performed on patients aged 6 months to 18 years with a diagnosis of septic arthritis or acute hematogenous osteomyelitis in a large, free-standing children’s hospital between July 2012 and July 2017. Exclusion criteria were chronic osteomyelitis, prior trauma or surgery at the site, noninfectious arthritis, and Lyme arthritis. Data collected from the electronic medical record included demographics, initial and discharge antibiotic therapy, and microbiologic results. Data were compared with the local antibiogram during the same time period.ResultsA total of 207 patients were included: 66 patients <4 years (< 4Y) and 141 patients ≥4 years (≥4Y). Causative pathogens were identified in 70% of patients. Staphylococcus aureus comprised 55% of positive results in children < 4Y and 73% in children ≥4Y. Among S. aureus cultures, 70–76% were methicillin sensitive (MSSA). Overall clindamycin susceptibility was 97%, with all resistant strains detected in children ≥4Y with MSSA. This is strikingly different than the institutional antibiogram showing 79% overall clindamycin sensitivity in S. aureus [82% in MSSA, 72% in methicillin resistant (MRSA)]. Kingella kingae was exclusively identified in children <4Y (21% of positives), which was also the group with the highest rate of culture-negative infection (41%). Intravenous clindamycin alone was the most frequent initial antibiotic regimen, prescribed for 41% of all patients. Initial antibiotic regimens matched organism susceptibilities in 90% of MRSA and 100% of MSSA infections.ConclusionOur study revealed high rates of clindamycin-susceptible S. aureus in older children and K. kingae and culture-negative infection in children < 4 years with OAI. Antibiotic susceptibilities differing from our institutional antibiogram suggest that disease-specific antibiograms will aid with empiric treatment decisions.Disclosures All authors: No reported disclosures.

660. Extraction-Free 16S Ribosomal RNA (rRNA) Gene Amplification and Sequencing from Resected Cardiac Implantable Electronic Device (CIED) Sonicate Fluid

BackgroundWe recently demonstrated that 16S rRNA PCR/sequencing performed on biofilms dislodged from extracted CIEDs into a salt solution—referred to as “sonicate fluid’ (SF)—may be used to detect pathogens in culture-negative CIED infection (Clinical Infectious Diseases, ciz266, doi:10.1093/cid/ciz266). The method we described included a DNA extraction/purification step, which can be time consuming and costly. Here, we evaluated an extraction-free approach to 16S rRNA gene PCR/sequencing.Methods28 SF samples derived from explanted clinically-infected CIEDs were tested. Cases were categorized as “culture-positive” (C-P) if ≥20 cfu/10 mL were recovered, and as “culture-negative” (C-N), if <20 cfu/10 mL were detected in SF culture. The extraction-free method consisted of a single step of lysis at 1,000 rpm, 95°C for 5 minutes using an Eppendorf ThermoMixer®. DNA extraction (the comparator method) was performed using the ZymoBIOMICSTM Kit with modifications. Samples were processed using both methods, followed by amplification of the 16S rRNA gene and bidirectional Sanger sequencing. Crossing points (CPs) generated by the two approaches were compared. Organisms detected by the two PCR methods were compared with those detected with culture.ResultsOf the 28 samples tested, 13 were C-N and 15 C-P. The extraction-free method generated an amplicon in 13/15 C-P cases, with CPs ranging from 26 to 36 cycles vs. 100% (15/15) detected with the DNA extraction method and Cps of 19 to 32. Usable sequence length for the extraction-free method was of 359 (interquartile range, 307–390) vs. 390 (interquartile range, 308–396) base pairs with DNA extraction. Genus-level concordance between bacteria detected by culture in C-P samples and those found using the extraction-free and extraction methods was 92% (12/13) and 93% (14/15), respectively. Bacteria were detected by the extraction method in 2/13 C-N specimens, with none detected with an extraction-free method.ConclusionThe described extraction-free method may be suitable for testing SF derived from CIEDs using 16S rRNA gene PCR/sequencing, saving time and cost. More studies are needed to establish clear cutoffs for interpretation of results and to assess for PCR inhibitors in the studied specimen-type.DisclosuresRobin Patel, MD, ASM and IDSA: Other Financial or Material Support, Travel reimbursement, editor’s stipends; CD Diagnostics, Merck, Hutchison Biofilm Medical Solutions, Accelerate Diagnostics, ContraFect, TenNor Therapeutics Limited, Shionogi: Grant/Research Support; Curetis, Specific Technologies, NextGen Diagnostics, PathoQuest, Qvella: Consultant; NBME, Up-to-Date, the Infectious Diseases Board Review Course: Honorarium recipient, Other Financial or Material Support; Patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication with royalties paid by Samsung to Mayo Clinic, and a patent on an anti-biofilm substance issued: Other Financial or Material Support, Patents.

Culture-negative periprosthetic joint infections.

Culture-negative periprosthetic joint infections (CN-PJI) pose a significant challenge in terms of diagnosis and management. The reported incidence of CN-PJI is reported to be between 7% and 15%.Fungi and mycobacterium are thought to be responsible for over 85% of such cases with more fastidious bacteria accounting for the rest.With the advent of polymerase chain reaction, mass spectrometry and next generation sequencing, identifying the causative organism(s) may become easier but such techniques are not readily available and are very costly.There are a number of more straightforward and relatively low-cost methods to help surgeons maximize the chances of diagnosing a PJI and identify the organisms responsible.This review article summarizes the main diagnostic tests currently available as well as providing a simple diagnostic clinical algorithm for CN-PJI.Cite this article: EFORT Open Rev 2019;4:585-594. DOI: 10.1302/2058-5241.4.180067

Severe Sepsis in Pediatric Liver Transplant Patients: The Emergence of Multidrug-Resistant Organisms.

To describe characteristics of liver transplant patients with severe sepsis in the PICU. Retrospective descriptive analysis. Tertiary children's hospital PICU. Liver transplant recipients admitted January 2010 to July 2016 for pediatric severe sepsis. None. Between January 2010 and July 2016, 173 liver transplants were performed, and 36 of these patients (21%) were admitted with severe sepsis (54 episodes total). Median age at admission was 2 years (1-6.5 yr), 47.2% were male. Bacterial infections were the most common (77.8%), followed by culture negative (12.9%) and viral infections (7.4%). Fungal infections accounted for only 1.9%. Median time from transplant for viral and culture negative infections was 18 days (8.25-39.75 d) and 25 days (9-41 d), whereas 54.5 days (17-131.25 d) for bacterial infections. Bloodstream and intra-abdominal were the most common bacterial sites (45% and 22.5%, respectively). Multidrug-resistant organisms accounted for 47.6% of bacterial sepsis. Vancomycin-resistant Enterococcus and extended-spectrum beta-lactamase producers were the most frequently identified multidrug-resistant organisms. Patients with multidrug-resistant organism sepsis demonstrated higher admission Pediatric Logistic Organ Dysfunction scores (p = 0.043) and were noted to have an odds ratio of 3.8 and 3.6 for mechanical ventilation and multiple organ dysfunction syndrome, respectively (p = 0.047 and p = 0.044). Overall mortality was 5.5% (n = 2 patients), with both deaths occurring in multidrug-resistant organism episodes. We report that multidrug-resistant organisms are increasingly being identified as causative pathogens for sepsis in pediatric liver transplant recipients and are associated with significantly higher odds for mechanical ventilation and higher organ failure. The emergence of multidrug-resistant organism infections in pediatric liver transplant patients has implications for patient outcomes, antibiotic stewardship, and infection prevention strategies.

Validation and Retrospective Clinical Evaluation of a Quantitative 16S rRNA Gene Metagenomic Sequencing Assay for Bacterial Pathogen Detection in Body Fluids

Next-generation sequencing-based 16S rRNA gene metagenomic sequencing (16S MG) technology has tremendous potential for improving diagnosis of bacterial infections given its quantitative capability and culture-independent approach. We validated and used a quantitative 16S MG assay to identify and quantify bacterial species in clinical samples from a wide spectrum of infections, including meningitis, septic arthritis, brain abscess, intra-abdominal abscess, soft tissue abscess, and pneumonia. Twenty clinical samples were tested, and 16S MG identified a total of 34 species, compared with 22 species and three descriptive findings identified by culture. 16S MG results matched culture results in 75% (15/20) of the samples but detected at least one more species in five samples, including one culture-negative cerebrospinal fluid sample that was found to contain Streptococcus intermedius. Shotgun metagenomic sequencing verified the presence of all additional species. The 16S MG assay is highly sensitive, with alimit of detection of 10 to 100 colony-forming units/mL. Other performance characteristics, including linearity, precision, and specificity, all met the requirements for a clinical test. This assay showed theadvantages of accurate identification and quantification of bacteria in culture-negative and polymicrobial infections for which conventional microbiology methods are limited. It also showed promises to serve unmet clinical needs for solving difficult infectious diseases cases.

Clinical Experience with Tigecycline in the Treatment of Prosthetic Joint Infections.

Purpose: The purpose of this study was to examine the use of tigecycline in the treatment of prosthetic joint infection (PJI).Methods: This is a retrospective review performed from 2008 to 2017, examining adult patients with PJI at a tertiary medical referral center who received tigecycline for 75% or greater of the treatment course. Failure was defined as need to return to the operating room for an infectious complication or persistent drainage from the joint.Results: A total of 37 patients met inclusion criteria. The median age was 65 years, and 65% of patients were female. The most common reasons for tigecycline use were culture negative infection, polymicrobial infection, and renal failure, but other reasons included antimicrobial allergies and resistant organisms. The mean duration of tigecycline therapy was 40 days (range 28-52 days). Treatment success was documented in 16 cases (43%).Conclusions: Tigecycline is a glycylcycline approved for treatment of a variety of infections including skin and soft tissue infections, but little is known about its use in the treatment of PJI. We found that tigecycline is well-tolerated for prolonged durations. Our success rate was 43%, but the majority of patients in this study had complicated infectious surgical histories and had received prior prolonged courses of antimicrobial therapy which likely affected treatment outcome. We concluded that tigecycline should be reserved as an alternative when other antimicrobials for PJI have been exhausted. More studies are needed to assess tigecycline's use in the treatment of PJI.

Serum Hepcidin-25 Levels Reflect the Presence of Bacteremia in Patients with Systemic Inflammatory Response Syndrome.

Hepcidin-25 is a key regulatory hormone of iron homeostasis in humans, and its production is greatly upregulated by inflammation as well as iron overload. The aim of this study was to investigate the pathophysiological role of hepcidin-25 in patients with systemic inflammatory response syndrome (SIRS). We enrolled 113 consecutive patients (aged 63.4±21, 50 men, 63 women), with 2 or more SIRS criteria, who were admitted to our department of general medicine between August 1, 2015 and August 31, 2017. We measured complete blood cell count and serum levels of hepcidin-25, iron, iron-binding capacity, ferritin, blood urea nitrogen, creatinine, albumin, and C-reactive protein (CRP) on admission. The patients were divided into 3 group: a bacteremia group (27 patients), a culture-negative bacterial infection group (60 patients), and a non-bacterial infection group (26 patients). Hepcidin-25 levels were found to be comparable in terms of SIRS criteria: 162 [2.8-579], 193 [2.24-409], and 180 [89.2-421] ng/mL in patients with 2, 3, and 4 criteria, respectively (P=0.533). However, hepcidin-25 levels were significantly higher in the bacteremia group (209 [56.7-579] ng/mL) than in either the culture-negative bacterial infection group (168 [2.24-418] ng/mL) or the non-bacterial infection group (142 [2.8-409] ng/mL). A significant positive correlation between hepcidin-25 and CRP levels was noted in the bacteremia group (r=0.528, P=0.005) and non-bacterial infection group (r=0.648, P<0.001). Moreover, iron and ferritin levels were significantly lower in the bacteremia group than in the non-bacterial infection group. Our findings suggest that hepcidin-25 level may reflect the presence of bacteremia as well as the severity of inflammation in patients with SIRS.

UltraStrain: An NGS-Based Ultra Sensitive Strain Typing Method for Salmonella enterica.

In the last few years, advances in next-generation sequencing (NGS) technology for whole genome sequencing (WGS) of foodborne pathogens have provided drastic improvements in food pathogen outbreak surveillance. WGS of foodborne pathogen enables identification of pathogens from food or environmental samples, including difficult-to-detect pathogens in culture-negative infections. Compared to traditional low-resolution methods such as the pulsed-field gel electrophoresis (PFGE), WGS provides advantages to differentiate even closely related strains of the same species, thus enables rapid identification of food-source associated with pathogen outbreak events for a fast mitigation plan. In this paper, we present UltraStrain, which is a fast and ultra sensitive pathogen detection and strain typing method for Salmonella enterica (S. enterica) based on WGS data analysis. In the proposed method, a noise filtering step is first performed where the raw sequencing data are mapped to a synthetic species-specific reference genome generated from S. enterica specific marker sequences to avoid potential interference from closely related species for low spike samples. After that, a statistical learning based method is used to identify candidate strains, from a database of known S. enterica strains, that best explain the retained S. enterica specific reads.Finally, a refinement step is further performed by mapping all the reads before filtering onto the identified top candidate strains, and recalculating the probability of presence for each candidate strain. Experiment results using both synthetic and real sequencing data show that the proposed method is able to identify the correct S. enterica strains from low-spike samples, and outperforms several existing strain-typing methods in terms of sensitivity and accuracy.

Procalcitonin and Other Common Biomarkers Do Not Reliably Identify Patients at Risk for Bacterial Infection After Congenital Heart Surgery.

Following surgery, it is difficult to distinguish a postoperative inflammatory reaction from infection. This study examined the predictive value of the biomarkers; procalcitonin, C-reactive protein, lactate, neutrophils, lymphocytes, platelets, and the biphasic activated partial thromboplastin time waveform in diagnosing bacterial infection following cardiac surgery. Prospective, observational study. A regional, PICU in the United Kingdom. Three-hundred sixty-eight children under the age of 16 admitted to the PICU for elective cardiac surgery were enrolled in the study. All biomarker measurements were determined daily until postoperative day 7. Children were assessed for postoperative infection until day 28 and divided into four groups: bacterial infection, culture-negative sepsis, viral infection, and no infection. We used the Kruskal-Wallis test, chi-square test, analysis of variance, and area under the curve in our analysis. In total, 71 of 368 children (19%) developed bacterial infection postoperatively, the majority being surgical site infections. In those with bacterial infection, procalcitonin was elevated on postoperative days 1-3 and the last measurement prior to event compared with those without bacterial infection. The most significant difference was the last measurement prior to event; 0.72 ng/mL in the bacterial infection group versus 0.13 ng/mL in the no infection group (for all groups; p < 0.001). Longitudinal profiles of all biomarkers were indistinct in the bacterial infection and nonbacterial infection groups except in those with culture-negative infections who had distinct procalcitonin kinetics on postoperative days 1-4. Children with culture-negative sepsis required longer ventilatory support and PICU stay and were more likely to develop complications than the other groups. None of the biomarkers studied within 3 days of infection distinguished between infection and postoperative inflammatory reaction. However, procalcitonin kinetics peaked on postoperative day 2 and fell more sharply than C-reactive protein kinetics, which peaked at postoperative day 3. The monitoring of procalcitonin kinetics following cardiac surgery may help guide rational antimicrobial use.

Shoulder Joint Infections with Negative Culture Results: Clinical Characteristics and Treatment Outcomes.

Background The incidence of septic arthritis of the shoulder joint is increasing as the population ages. The prevalence of shoulder infection is also increasing because of the growing use of arthroscopy and expansion of procedures in the shoulder. However, cultures do not always identify all microorganisms, even in symptomatic patients. The incidence of negative cultures ranges from 0% to 25%. Few studies have reported clinical features and treatment outcomes of culture-negative shoulder infections. This cohort study addresses culture-negative shoulder joint infections in nonarthroplasty patients. This study aimed to compare clinical characteristics and treatment outcomes of patients with culture-negative results to those with culture-positive results. Our hypothesis was that culture-negative infections would have more favorable outcomes than culture-positive infections. Methods We retrospectively reviewed data of 36 patients (17 culture-negative and 19 culture-positive) with shoulder infections between June 2004 and March 2015. The minimum follow-up duration was 1.2 years (mean, 5 ± 3.8 years; range, 1.2-11 years). We assessed preoperative demographic data and characteristics, laboratory markers, imaging and functional scores, intraoperative findings, and postoperative findings of both groups. Results Culture-negative patients (17/36, 47.2%) had a significantly lower occurrence of repeated surgical debridement (culture-negative vs. culture-positive: 1.2 ± 0.4 vs. 2.4 ± 1.7, p = 0.002) without osteomyelitis. In the multiple logistic regression analysis, the presence of osteomyelitis [odds ratio (OR) = 9.7, 95% confidence interval (CI): 1.0-91.8, p=0.04)] and the number of surgical debridements (OR = 5.3, 95% CI: 1.3-21.6, p=0.02) were significantly associated with culture-positive infections. Conclusions Culture-negative infections without osteomyelitis are less severe than culture-positive infections. Culture-negative infections can be controlled more easily and are not necessarily a negative prognostic factor for shoulder joint infections.

Sonication to improve the yield in culture-negative peri-prosthetic joint infection

ABSTRACTPeri-prosthetic joint infection (PJI) is a catastrophic complication, after joint arthroplasty. Culture-negative PJI presented 7–39% of PJI cases at most health-care centers. We aimed to investigate the diagnostic accuracy of sonication fluid culture (SFC) with the respective peri-prosthetic tissue cultures (PTC). Also, to evaluate the efficacy of sonication in PTC-negative PJI. Peri-prosthetic tissue specimens were subjected to microbiological culture and histo-pathological examination. Retrieved prosthesis components were sonicated. SFC was examined. One hundred and seventy patients had PJI and 95 were aseptic failures. SFC was positive in 120 patients (70.5%). For PTC, 80 of 170 samples (47.1%) were positive; all reveal the same bacterial species with the corresponding SFC. In 50 patients, both PTC and SFC cultures of the prosthesis were negative. There were 40 negative PTC samples proved to be positive by SFC, 39 displayed Staphylococcus epidermidis, and only one sample yielded Propionibacterium acne. Biofilm formation was detected in 22 (56.4%) out of the 39 S. epidermidis isolates. The overall sensitivity of SFC compared to PTC was (70.6% versus 47.1%). SFC represents a sensitive, accurate, easy diagnostic strategy representing increased sensitivity contrasted with PTC. S. epidermidis, although an important commensal, is considered the most significant pathogen in the context of PJI.

Early Outcome of Culture-Negative Infection in Open Fractures of the Lower Limb: A Prospective Study

AbstractBackground: Culture-negative infections in open long bone fractures are frequently encountered in clinical practice. We aimed to identify the rate and outcome of culture-negative infections in open long bone fractures of lower limb. Methodology: A prospective cohort study was conducted from November 2015 to May 2017 on Gustilo and Anderson Grade III open long bone fractures of the lower limb. Demographic data, injury details, time from injury to receiving antibiotics and index surgical procedure were noted. Length of hospital stay, number of additional surgeries and occurrence of complications were also noted. Patients with infected open fractures were grouped as culture positive or culture negative depending on the isolation of infecting microorganisms in deep intraoperative specimen. The clinical outcome of these two groups was statistically analysed. Results: A total of 231 patients with 275 open fractures involving the femur, tibia or fibula were studied. There was clinical signs of infection in 84 patients (36.4%) with 99 fractures (36%). Forty-three patients (51.2%) had positive cultures and remaining 41 patients had negative cultures (48.8%). The rate of culture-negative infection in open type III long bone fractures in our study was 17.7%. There was no statistical difference in the clinical outcome between culture-negative and culture-positive infections. Conclusion: Failure to identify an infective microorganism in the presence of clinical signs of infection is routinely seen in open fractures and needs to be treated aggressively.

Heart Rate Characteristic Index Monitoring for Early Detection of Infections in Very Low Birth Weight Infants

The aim of this study is 1) to determine the sensitivity and specificity of continuous heart rate characteristics (HRC) monitoring in detection of infections and 2) to evaluate whether HRC monitoring detects infections prior to onset of clinical symptoms in very low birth weight (VLBW) infants. A retrospective cohort study was conducted analyzing HRC scores and episodes of infection for VLBW infants in the Neonatal Intensive Care Unit (NICU) at Cincinnati Children’s Hospital Medical Center from January 2015 through May 2016. HRC scores were acquired using the HRC monitor system and entered into the electronic medical record by bedside staff. Culture-positive and culture-negative clinical infections were recorded. Positive HRC scores were defined as an increase 1 point above the baseline or the first rise above 2. HRC scores within 24 hours and also within the 5-day period before the start of antibiotics for infections were analyzed for sensitivity, specificity, positive predictive value and negative predictive value for detection of neonatal infection. The HRC score increase 1 point above the baseline or the first rise above 2 in the 24 hours before the start of antibiotics for infectious events had a sensitivity of 68.0%, a specificity of 10.8%. Positive predictive value (PPV) and negative predictive value (NPV) were 34.0% and 33.3%, respectively. The PPV and NPV were modestly higher for elevated HRC scores during the 5-day period before infections, 41.1% and 66.7%, respectively. In our single-center retrospective study, elevated HRC scores had limited ability to detect infection. More than half of the positive monitor events were not related to infection. The potential clinical impact of the monitor to detect infection before the onset of clinical symptoms was limited and the risk for unnecessary evaluation and treatment was high.

Rapid Rule Out of Culture-Negative Bloodstream Infections by Use of a Novel Approach to Universal Detection of Bacteria and Fungi.

Currently it can take up to 5 days to rule out bloodstream infection. With the low yield of blood cultures (approximately 10%), a significant number of patients are potentially exposed to inappropriate therapy that can lead to adverse events. More rapid rule out can accelerate deescalation or cessation of antimicrobial therapy, improving patient outcomes. A method is described, termed enzymatic template generation and amplification (ETGA), that universally and sensitively detects DNA polymerase activity liberated from viable bacteria and fungi isolated from blood culture samples as a measure of bloodstream infection. ETGA was applied in a diagnostic test format to identify negative blood cultures after an overnight incubation. Performance data for a prototype (Cognitor) and automated (Magnitor) version of the test are presented. The Cognitor manual assay displayed analytical reactivity for a panel of the 20 most prevalent causes of bloodstream infection, with a detection range of 28-9050 CFU/mL. Validation with 1457 clinical blood cultures showed a negative predictive value of 99.0% compared to blood culture incubation for 5 days. Magnitor showed an improved detection range of 1-67 CFU/mL, allowing for detection of bacteria-supplemented blood cultures after 2-8 h incubation, and Candida albicans-supplemented blood cultures at 16-22 h, 5-15 h faster than blood culture. Removing an aliquot from a blood culture bottle and replacing the bottle into the incubator was shown not to result in contaminating organisms being introduced. The described method displays excellent breadth and detection for microbial cells and demonstrates the capability of confirming negative blood cultures after an overnight incubation in a blood culture instrument.