Fitness factor genes conserved within the multi-species core genome of Gram-negative Enterobacterales species contribute to bacteremia pathogenesis.

There is a critical gap in knowledge about how Gram-negative bacterial pathogens, using survival strategies developed for other niches, cause lethal bacteremia. Facultative anaerobic species of the Enterobacterales order are the most common cause of Gram-negative bacteremia, including Escherichia coli, Klebsiella pneumoniae, Serratia marcescens, Citrobacter freundii, and Enterobacter hormaechei. Bacteremia often leads to sepsis, a life-threatening organ dysfunction resulting from unregulated immune responses to infection. Despite a lack of specialization for this host environment, Gram-negative pathogens cause nearly half of bacteremia cases annually. Based on our existing Tn-Seq fitness factor data from a murine model of bacteremia combined with comparative genomics of the five Enterobacterales species above, we prioritized 18 conserved fitness genes or operons for further characterization. Mutants were constructed for all genes in all five species. Each mutant was used to cochallenge C57BL/6 mice via tail vein injection along with each respective wild-type strain to determine competitive indices for each fitness gene. Five fitness factor genes, when mutated, attenuated mutants in four or five species in the spleen and liver (tatC, ruvA, gmhB, wzxE, arcA). Five additional fitness factor genes or operons were validated as outcompeted by wild-type in three, four, or five bacterial species in the spleen (xerC, prc, apaGH, atpG, aroC). Overall, 17 of 18 fitness factor mutants were attenuated in at least one species in the spleen or liver. Together, these findings allow for the development of a model of bacteremia pathogenesis that may include future targets of therapy against bloodstream infections.

Klebsiella pneumoniae causes bacteremia using factors that mediate tissue-specific fitness and resistance to oxidative stress.

Gram-negative bacteremia is a major cause of global morbidity involving three phases of pathogenesis: initial site infection, dissemination, and survival in the blood and filtering organs. Klebsiella pneumoniae is a leading cause of bacteremia and pneumonia is often the initial infection. In the lung, K. pneumoniae relies on many factors like capsular polysaccharide and branched chain amino acid biosynthesis for virulence and fitness. However, mechanisms directly enabling bloodstream fitness are unclear. Here, we performed transposon insertion sequencing (TnSeq) in a tail-vein injection model of bacteremia and identified 58 K. pneumoniae bloodstream fitness genes. These factors are diverse and represent a variety of cellular processes. In vivo validation revealed tissue-specific mechanisms by which distinct factors support bacteremia. ArnD, involved in Lipid A modification, was required across blood filtering organs and supported resistance to soluble splenic factors. The purine biosynthesis enzyme PurD supported liver fitness in vivo and was required for replication in serum. PdxA, a member of the endogenous vitamin B6 biosynthesis pathway, optimized replication in serum and lung fitness. The stringent response regulator SspA was required for splenic fitness yet was dispensable in the liver. In a bacteremic pneumonia model that incorporates initial site infection and dissemination, splenic fitness defects were enhanced. ArnD, PurD, DsbA, SspA, and PdxA increased fitness across bacteremia phases and each demonstrated unique fitness dynamics within compartments in this model. SspA and PdxA enhanced K. pnuemoniae resistance to oxidative stress. SspA, but not PdxA, specifically resists oxidative stress produced by NADPH oxidase Nox2 in the lung, spleen, and liver, as it was a fitness factor in wild-type but not Nox2-deficient (Cybb-/-) mice. These results identify site-specific fitness factors that act during the progression of Gram-negative bacteremia. Defining K. pneumoniae fitness strategies across bacteremia phases could illuminate therapeutic targets that prevent infection and sepsis.

The Klebsiella pneumoniae ter Operon Enhances Stress Tolerance.

Healthcare-acquired infections are a leading cause of disease in patients that are hospitalized or in long-term-care facilities. Klebsiella pneumoniae (Kp) is a leading cause of bacteremia, pneumonia, and urinary tract infections in these settings. Previous studies have established that the ter operon, a genetic locus that confers tellurite oxide (K2TeO3) resistance, is associated with infection in colonized patients. Rather than enhancing fitness during infection, the ter operon increases Kp fitness during gut colonization; however, the biologically relevant function of this operon is unknown. First, using a murine model of urinary tract infection, we demonstrate a novel role for the ter operon protein TerC as a bladder fitness factor. To further characterize TerC, we explored a variety of functions, including resistance to metal-induced stress, resistance to radical oxygen species-induced stress, and growth on specific sugars, all of which were independent of TerC. Then, using well-defined experimental guidelines, we determined that TerC is necessary for tolerance to ofloxacin, polymyxin B, and cetylpyridinium chloride. We used an ordered transposon library constructed in a Kp strain lacking the ter operon to identify the genes that are required to resist K2TeO3-induced and polymyxin B-induced stress, which suggested that K2TeO3-induced stress is experienced at the bacterial cell envelope. Finally, we confirmed that K2TeO3 disrupts the Kp cell envelope, though these effects are independent of ter. Collectively, the results from these studies indicate a novel role for the ter operon as a stress tolerance factor, thereby explaining its role in enhancing fitness in the gut and bladder.

A systematic analysis of hypermucoviscosity and capsule reveals distinct and overlapping genes that impact Klebsiella pneumoniae fitness.

Hypervirulent K. pneumoniae (hvKp) is a distinct pathotype that causes invasive community-acquired infections in healthy individuals. Hypermucoviscosity (hmv) is a major phenotype associated with hvKp characterized by copious capsule production and poor sedimentation. Dissecting the individual functions of CPS production and hmv in hvKp has been hindered by the conflation of these two properties. Although hmv requires capsular polysaccharide (CPS) biosynthesis, other cellular factors may also be required and some fitness phenotypes ascribed to CPS may be distinctly attributed to hmv. To address this challenge, we systematically identified genes that impact capsule and hmv. We generated a condensed, ordered transposon library in hypervirulent strain KPPR1, then evaluated the CPS production and hmv phenotypes of the 3,733 transposon mutants, representing 72% of all open reading frames in the genome. We employed forward and reverse genetic screens to evaluate effects of novel and known genes on CPS biosynthesis and hmv. These screens expand our understanding of core genes that coordinate CPS biosynthesis and hmv, as well as identify central metabolism genes that distinctly impact CPS biosynthesis or hmv, specifically those related to purine metabolism, pyruvate metabolism and the TCA cycle. Six representative mutants, with varying effect on CPS biosynthesis and hmv, were evaluated for their impact on CPS thickness, serum resistance, host cell association, and fitness in a murine model of disseminating pneumonia. Altogether, these data demonstrate that hmv requires both CPS biosynthesis and other cellular factors, and that hmv and CPS may serve distinct functions during pathogenesis. The integration of hmv and CPS to the metabolic status of the cell suggests that hvKp may require certain nutrients to specifically cause deep tissue infections.

A plasmid locus associated with Klebsiella clinical infections encodes a microbiome-dependent gut fitness factor.

Klebsiella pneumoniae (Kp) is an important cause of healthcare-associated infections, which increases patient morbidity, mortality, and hospitalization costs. Gut colonization by Kp is consistently associated with subsequent Kp disease, and patients are predominantly infected with their colonizing strain. Our previous comparative genomics study, between disease-causing and asymptomatically colonizing Kp isolates, identified a plasmid-encoded tellurite (TeO3-2)-resistance (ter) operon as strongly associated with infection. However, TeO3-2 is extremely rare and toxic to humans. Thus, we used a multidisciplinary approach to determine the biological link between ter and Kp infection. First, we used a genomic and bioinformatic approach to extensively characterize Kp plasmids encoding the ter locus. These plasmids displayed substantial variation in plasmid incompatibility type and gene content. Moreover, the ter operon was genetically independent of other plasmid-encoded virulence and antibiotic resistance loci, both in our original patient cohort and in a large set (n = 88) of publicly available ter operon-encoding Kp plasmids, indicating that the ter operon is likely playing a direct, but yet undescribed role in Kp disease. Next, we employed multiple mouse models of infection and colonization to show that 1) the ter operon is dispensable during bacteremia, 2) the ter operon enhances fitness in the gut, 3) this phenotype is dependent on the colony of origin of mice, and 4) antibiotic disruption of the gut microbiota eliminates the requirement for ter. Furthermore, using 16S rRNA gene sequencing, we show that the ter operon enhances Kp fitness in the gut in the presence of specific indigenous microbiota, including those predicted to produce short chain fatty acids. Finally, administration of exogenous short-chain fatty acids in our mouse model of colonization was sufficient to reduce fitness of a ter mutant. These findings indicate that the ter operon, strongly associated with human infection, encodes factors that resist stress induced by the indigenous gut microbiota during colonization. This work represents a substantial advancement in our molecular understanding of Kp pathogenesis and gut colonization, directly relevant to Kp disease in healthcare settings.

Prehospital Fluid Administration for Suspected Sepsis in a Large EMS System: Opportunities to Improve Goal Fluid Delivery.

OBJECTIVES: Despite EMS-implemented screening and treatment protocols for suspected sepsis patients, prehospital fluid therapy is variable. We sought to describe prehospital fluid administration in suspected sepsis patients, including demographic and clinical factors associated with fluid outcomes. METHODS: A retrospective cohort of adult patients from a large, county-wide EMS system from January 2018-February 2020 was identified. Patient care reports for suspected sepsis were included, as identified by EMS clinician impression of sepsis, or keywords "sepsis" or "septic" in the narrative. Outcomes were the proportions of suspected sepsis patients for whom intravenous (IV) therapy was attempted and those who received ≥500 mL IV fluid if IV access was successful. Associations between patient demographics and clinical factors with fluid outcomes were estimated with multivariable logistic regression adjusting for transport interval. RESULTS: Of 4,082 suspected sepsis patients identified, the mean patient age was 72.5 (SD 16.2) years, 50.6% were female, and 23.8% were Black. Median (interquartile range [IQR]) transport interval was 16.5 (10.9-23.2) minutes. Of identified patients, 1,920 (47.0%) had IV fluid therapy attempted, and IV access was successful in 1,872 (45.9%). Of those with IV access, 1,061 (56.7%) received ≥500mL of fluid from EMS. In adjusted analyses, female (versus male) sex (odds ratio [OR] 0.79, 95% confidence interval [CI] 0.69-0.90), Black (versus White) race (OR 0.57, 95% CI 0.49-0.68), and end stage renal disease (OR 0.51, 95% CI 0.32-0.82) were negatively associated with attempted IV therapy. Systolic blood pressure (SBP) <90 mmHg (OR 3.89, 95% CI 3.25-4.65) and respiratory rate >20 (OR 1.90, 95% CI 1.61-2.23) were positively associated with attempted IV therapy. Female sex (OR 0.72, 95% CI 0.59-0.88) and congestive heart failure (CHF) (OR 0.55, 95% CI 0.40-0.75) were negatively associated with receiving goal fluid volume while SBP <90 mmHg (OR 2.30, 95% CI 1.83-2.88) and abnormal temperature (>100.4 F or <96 F) (OR 1.41, 95% CI 1.16-1.73) were positively associated. CONCLUSIONS: Fewer than half of EMS sepsis patients had IV therapy attempted, and of those, approximately half met fluid volume goal, especially when hypotensive and no CHF. Further studies are needed on improving EMS sepsis training and prehospital fluid delivery.

Effectiveness of a Novel Rapid Infusion Device and Clinician Education for Early Fluid Therapy by Emergency Medical Services in Sepsis Patients: A Pre-Post Observational Study.

OBJECTIVE: Emergency medical services (EMS) clinicians are tasked with early fluid resuscitation for patients with sepsis. Traditional methods for prehospital fluid delivery are limited in speed and ease-of-use. We conducted a comparative effectiveness study of a novel rapid infusion device for prehospital fluid delivery in suspected sepsis patients. METHODS: This pre-post observational study evaluated a hand-operated, rapid infusion device in a single large EMS system from July 2021-July 2022. Prior to device deployment, EMS clinicians completed didactic and simulation-based device training. Data were extracted from the EMS electronic health record. Eligible patients included adults with suspected sepsis treated by EMS with intravenous fluids. The primary outcome was the proportion of patients receiving goal fluid volume (at least 500 mL) prior to hospital arrival. Secondary outcomes included in-hospital mortality, disposition, and length of stay. Multivariable logistic regression was used to compare outcomes between 6-month pre- and post-implementation periods (July-December 2021 and February-July 2022, respectively), adjusting for patient demographics, abnormal prehospital vital signs, and EMS transport interval. RESULTS: Of 1,180 eligible patients (552 in the pre-implementation period; 628 in the post-implementation period), the mean age was 72 years old, 45% were female, and 25% were minority race-ethnicity. Median (interquartile range) fluid volume (in mL) increased between the pre- and post-implementation periods (600 [400,1,000] and 850 [500-1,000], respectively). Goal fluid volume was achieved in 70% of pre-implementation patients and 82% of post-implementation patients. In adjusted analysis, post-implementation patients were significantly more likely to receive goal fluid volume than pre-implementation patients (adjusted odds ratio (aOR) 2.00, 95% confidence interval (CI) 1.51-2.66). Pre-post in-hospital mortality was not significantly different (aOR 0.91, 95% CI 0.59-1.39). CONCLUSION: In a single EMS system, sepsis education and introduction of a rapid infusion device was associated with achieving goal fluid volume for suspected sepsis. Further research is needed to assess the clinical effectiveness of infusion device implementation to improve sepsis patient outcomes.

Pathogenesis of Gram-Negative Bacteremia.

Gram-negative bacteremia is a devastating public health threat, with high mortality in vulnerable populations and significant costs to the global economy. Concerningly, rates of both Gram-negative bacteremia and antimicrobial resistance in the causative species are increasing. Gram-negative bacteremia develops in three phases. First, bacteria invade or colonize initial sites of infection. Second, bacteria overcome host barriers, such as immune responses, and disseminate from initial body sites to the bloodstream. Third, bacteria adapt to survive in the blood and blood-filtering organs. To develop new therapies, it is critical to define species-specific and multispecies fitness factors required for bacteremia in model systems that are relevant to human infection. A small subset of species is responsible for the majority of Gram-negative bacteremia cases, including Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii The few bacteremia fitness factors identified in these prominent Gram-negative species demonstrate shared and unique pathogenic mechanisms at each phase of bacteremia progression. Capsule production, adhesins, and metabolic flexibility are common mediators, whereas only some species utilize toxins. This review provides an overview of Gram-negative bacteremia, compares animal models for bacteremia, and discusses prevalent Gram-negative bacteremia species.

The ADP-Heptose Biosynthesis Enzyme GmhB is a Conserved Gram-Negative Bacteremia Fitness Factor.

Klebsiella pneumoniae is a leading cause of Gram-negative bacteremia, which is a major source of morbidity and mortality worldwide. Gram-negative bacteremia requires three major steps: primary site infection, dissemination to the blood, and bloodstream survival. Because K. pneumoniae is a leading cause of health care-associated pneumonia, the lung is a common primary infection site leading to secondary bacteremia. K. pneumoniae factors essential for lung fitness have been characterized, but those required for subsequent bloodstream infection are unclear. To identify K. pneumoniae genes associated with dissemination and bloodstream survival, we combined previously and newly analyzed insertion site sequencing (InSeq) data from a murine model of bacteremic pneumonia. This analysis revealed the gene gmhB as important for either dissemination from the lung or bloodstream survival. In Escherichia coli, GmhB is a partially redundant enzyme in the synthesis of ADP-heptose for the lipopolysaccharide (LPS) core. To characterize its function in K. pneumoniae, an isogenic knockout strain (ΔgmhB) and complemented mutant were generated. During pneumonia, GmhB did not contribute to lung fitness and did not alter normal immune responses. However, GmhB enhanced bloodstream survival in a manner independent of serum susceptibility, specifically conveying resistance to spleen-mediated killing. In a tail-vein injection of murine bacteremia, GmhB was also required by K. pneumoniae, E. coli, and Citrobacter freundii for optimal fitness in the spleen and liver. Together, this study identifies GmhB as a conserved Gram-negative bacteremia fitness factor that acts through LPS-mediated mechanisms to enhance fitness in blood-filtering organs.

Management of Cytomegalovirus, Epstein-Barr Virus, and HIV Viral Load Quality Control Data Using Unity Real Time.

Quality control (QC) rules (Westgard rules) are applied to viral load testing to identify runs that should be reviewed or repeated, but this requires balancing the patient safety benefits of error detection with the cost and inefficiency of false rejection. In this study, we identified the total allowable errors (TEa) from the literature and utilized a commercially available software program (Unity Real Time; Bio-Rad Laboratories) to manage QC data, assess assay performance, and provide QC decision support for both FDA-approved/cleared (Abbott cytomegalovirus [CMV] and HIV viral load) as well as laboratory-developed (Epstein-Barr virus [EBV] viral load) assays. Unity Real Time was used to calculate means, standard deviations (SDs), and coefficient of variation (CV; in percent) of negative, low-positive, and high-positive control data from 73 to 83 days of testing. Sigma values were calculated to measure the test performance relative to a TEa of 0.5 log10. The sigma value of 5.06 for EBV predicts ∼230 erroneous results per million individual patient tests (0.02% frequency), whereas sigma values of >6 for CMV (11.32) and HIV (7.66) indicate <4 erroneous results per million individual patient tests. The Unity Real Time QC Design module utilized these sigma values to recommend QC rules and provided objective evidence for loosening the laboratory's existing QC rules for run acceptability, potentially reducing false rejection rates by 10-fold for the assay with the most variation (EBV viral load). This study provides a framework for laboratories, with Unity Real Time as a tool, to evaluate assay performance relative to clinical decision points and establish optimal rules for routine monitoring of molecular viral load assay performance.

Proceedings of the Clinical Microbiology Open 2018 and 2019 - a Discussion about Emerging Trends, Challenges, and the Future of Clinical Microbiology.

Clinical Microbiology Open (CMO), a meeting supported by the American Society for Microbiology's Clinical and Public Health Microbiology Committee (CPHMC) and Corporate Council, provides a unique interactive platform for leaders from diagnostic microbiology laboratories, industry, and federal agencies to discuss the current and future state of the clinical microbiology laboratory. The purpose is to leverage the group's diverse views and expertise to address critical challenges, and discuss potential collaborative opportunities for diagnostic microbiology, through the utilization of varied resources. The first and second CMO meetings were held in 2018 and 2019, respectively. Discussions were focused on the diagnostic potential of innovative technologies and laboratory diagnostic stewardship, including expansion of next-generation sequencing into clinical diagnostics, improvement and advancement of molecular diagnostics, emerging diagnostics, including rapid antimicrobial susceptibility and point of care testing (POCT), harnessing big data through artificial intelligence, and staffing in the clinical microbiology laboratory. Shortly after CMO 2019, the coronavirus disease 2019 (COVID-19) pandemic further highlighted the need for the diagnostic microbiology community to work together to utilize and expand on resources to respond to the pandemic. The issues, challenges, and potential collaborative efforts discussed during the past two CMO meetings proved critical in addressing the COVID-19 response by diagnostic laboratories, industry partners, and federal organizations. Planning for a third CMO (CMO 2022) is underway and will transition from a discussion-based meeting to an action-based meeting. The primary focus will be to reflect on the lessons learned from the COVID-19 pandemic and better prepare for future pandemics.

Prolonged Severe Acute Respiratory Syndrome Coronavirus 2 Replication in an Immunocompromised Patient.

We describe a case of chronic coronavirus disease 2019 (COVID-19) in a patient with lymphoma and associated B-cell immunodeficiency. Viral cultures and sequence analysis demonstrate ongoing replication of infectious severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) for at least 119 days. The patient had 3 admissions related to COVID-19 over a 4-month period and was treated twice with remdesivir and convalescent plasma with resolution of symptoms. The patient's lack of seroconversion and prolonged course illustrate the importance of humoral immunity in resolving SARS-CoV-2 infection. This case highlights challenges in managing immunocompromised hosts, who may act as persistent shedders and sources of transmission.

The Klebsiella pneumoniae citrate synthase gene, gltA, influences site specific fitness during infection.

Klebsiella pneumoniae (Kp), one of the most common causes of healthcare-associated infections, increases patient morbidity, mortality, and hospitalization costs. Kp must acquire nutrients from the host for successful infection; however, the host is able to prevent bacterial nutrient acquisition through multiple systems. This includes the innate immune protein lipocalin 2 (Lcn2), which prevents Kp iron acquisition. To identify novel Lcn2-dependent Kp factors that mediate evasion of nutritional immunity during lung infection, we undertook an InSeq study using a pool of >20,000 transposon mutants administered to Lcn2+/+ and Lcn2-/- mice. Comparing transposon mutant frequencies between mouse genotypes, we identified the Kp citrate synthase, GltA, as potentially interacting with Lcn2, and this novel finding was independently validated. Interestingly, in vitro studies suggest that this interaction is not direct. Given that GltA is involved in oxidative metabolism, we screened the ability of this mutant to use a variety of carbon and nitrogen sources. The results indicated that the gltA mutant has a distinct amino acid auxotrophy rendering it reliant upon glutamate family amino acids for growth. Deletion of Lcn2 from the host leads to increased amino acid levels in bronchioloalveolar lavage fluid, corresponding to increased fitness of the gltA mutant in vivo and ex vivo. Accordingly, addition of glutamate family amino acids to Lcn2+/+ bronchioloalveolar lavage fluid rescued growth of the gltA mutant. Using a variety of mouse models of infection, we show that GltA is an organ-specific fitness factor required for complete fitness in the spleen, liver, and gut, but dispensable in the bloodstream. Similar to bronchioloalveolar lavage fluid, addition of glutamate family amino acids to Lcn2+/+ organ lysates was sufficient to rescue the loss of gltA. Together, this study describes a critical role for GltA in Kp infection and provides unique insight into how metabolic flexibility impacts bacterial fitness during infection.

Twin arginine translocation, ammonia incorporation, and polyamine biosynthesis are crucial for Proteus mirabilis fitness during bloodstream infection.

The Gram-negative bacterium Proteus mirabilis is a common cause of catheter-associated urinary tract infections (CAUTI), which can progress to secondary bacteremia. While numerous studies have investigated experimental infection with P. mirabilis in the urinary tract, little is known about pathogenesis in the bloodstream. This study identifies the genes that are important for survival in the bloodstream using a whole-genome transposon insertion-site sequencing (Tn-Seq) approach. A library of 50,000 transposon mutants was utilized to assess the relative contribution of each non-essential gene in the P. mirabilis HI4320 genome to fitness in the livers and spleens of mice at 24 hours following tail vein inoculation compared to growth in RPMI, heat-inactivated (HI) naïve serum, and HI acute phase serum. 138 genes were identified as ex vivo fitness factors in serum, which were primarily involved in amino acid transport and metabolism, and 143 genes were identified as infection-specific in vivo fitness factors for both spleen and liver colonization. Infection-specific fitness factors included genes involved in twin arginine translocation, ammonia incorporation, and polyamine biosynthesis. Mutants in sixteen genes were constructed to validate both the ex vivo and in vivo results of the transposon screen, and 12/16 (75%) exhibited the predicted phenotype. Our studies indicate a role for the twin arginine translocation (tatAC) system in motility, translocation of potential virulence factors, and fitness within the bloodstream. We also demonstrate the interplay between two nitrogen assimilation pathways in the bloodstream, providing evidence that the GS-GOGAT system may be preferentially utilized. Furthermore, we show that a dual-function arginine decarboxylase (speA) is important for fitness within the bloodstream due to its role in putrescine biosynthesis rather than its contribution to maintenance of membrane potential. This study therefore provides insight into pathways needed for fitness within the bloodstream, which may guide strategies to reduce bacteremia-associated mortality.

Opportunities for Emergency Medical Services Intervention to Prevent Opioid Overdose Mortality.

OBJECTIVES: The opioid crisis is a growing cause of mortality in the United States and may be mitigated by innovative approaches to identifying individuals at-risk of fatal opioid overdose. We examined Emergency Medical Services (EMS) utilization among a cohort of individuals who died from opioid overdose in order to identify potential opportunities for intervention. Methods: Individuals who died of unintentional opioid overdose in a large North Carolina county between 01/01/2014 and 12/31/2016 were studied in a retrospective cohort. Death records obtained from North Carolina Vital Records were linked to EMS patient care records obtained from the county EMS System in order to describe the EMS encounters of each decedent in the year preceding their death. Patient demographics and EMS encounters were assessed to identify encounter characteristics that may be targeted for intervention. Chi-square tests and odds ratios were used to evaluate and characterize the statistical significance of differences in EMS utilization. Results: Of the 218 individuals who died from unintentional opioid overdose in the study interval, 30% (n = 66) utilized EMS in the year before their death and 17% (n = 38) had at least one EMS encounter with documented drug or alcohol use (i.e. "drug-related encounter"). The mean age at death was 38 (range 19-74) years, 30% were female, 89% were White, and 8% were Black/African American. Factors associated with higher incidence of EMS utilization included age (P<.001), gender (P=.006), and race (P<.001). Decedents aged 56-65 had the highest EMS utilization (47%) and patients aged <25 and 25-35 had more drug-related EMS encounters (29% and 20%, respectively). The most common reasons for EMS utilization were "other medical" (27%), "non-traumatic pain" (20%), "traumatic injury" (16%), and "poisoning/drug ingestion" (14%). Drug or alcohol use was documented by EMS in 33% of all encounters and an opioid prescription was reported in 22% of encounters. Conclusions: Nearly one-third of individuals who died from accidental opioid overdose utilized EMS in the year before their death and nearly one-fifth had a drug-related encounter. EMS encounters may present an opportunity to identify individuals at-risk of opioid overdose and, ultimately, reduce overdose mortality.

Predictive Utility of End-Tidal Carbon Dioxide on Defibrillation Success in Out-of-Hospital Cardiac Arrest.

INTRODUCTION: The likelihood of survival from ventricular fibrillation (VF) declines 7%-10% per minute until successful defibrillation. When VF duration is prolonged, immediate defibrillation of the ischemic myocardium is less likely to result in ROSC, and repeated unsuccessful defibrillations are associated with post-resuscitation myocardial dysfunction. Thus, the timing of defibrillation should be based upon the probability of shock success-a function of VF duration. Unfortunately, VF duration is often unknown in out-of-hospital cardiac arrest (OHCA) and a better predictor of shock success is needed. OBJECTIVE: To assess the ability of end-tidal carbon dioxide (EtCO2) to predict successful defibrillation in OHCA. METHODS: This retrospective study included adult patients among four EMS systems who experienced non-traumatic OHCA from August, 2015-July, 2017 and received one or more defibrillations. First and succedent shocks were analyzed separately. First shocks represented EMS-attempted defibrillation of patients who had not received a prior AED shock, whereas succedent shocks included all shocks subsequent to the first. Logistic regression provided odds ratios (OR) for first shocks resulting in ROSC, while a generalized estimating equation was used to analyze succedent shocks. RESULTS: Among 324 patients, 869 shocks were delivered by EMS (153 first and 716 succedent shocks). Layperson CPR was performed in 48.1% of cases and 21.6% received an AED shock before EMS arrival. First defibrillation ROSC was more likely with layperson CPR (OR = 4.41;p = 0.01) and increasing EtCO2 (OR = 1.03/mmHg;p = 0.01). No other variables were statistically significant. Notably, only one patient with EtCO2 < 20 mmHg was successfully defibrillated on the first shock. The probability of ROSC was higher with increasing values of EtCO2 when layperson CPR was provided, yet remained relatively unchanged across all values of EtCO2 ≥ 20 mmHg without layperson CPR. The optimal threshold first shock EtCO2 was 27 and 32 mmHg for those with/without layperson CPR, respectively. EtCO2 was not a predictor of ROSC for succedent shocks. CONCLUSIONS: An optimal defibrillation threshold EtCO2 of 27 and 32 mmHg was observed for patients with and without layperson CPR, respectively. Further studies are warranted to verify these results and to evaluate the clinical effect of delaying defibrillation in favor of chest compressions until these values are attained.

Evaluation of an Integrated Rescue Task Force Model for Active Threat Response.

OBJECTIVE: An integrated response to active threat events is essential to saving lives. Coordination of law enforcement officer (LEO) and emergency medical services (EMS) roles requires joint training, as maximizing survival is a shared responsibility. We sought to evaluate the performance of an integrated LEO-EMS Rescue Task Force (RTF) response to a simulated active shooter incident utilizing objective performance measures. METHODS: Following prior didactic training, we conducted a series of evaluation scenarios for EMS providers and patrol officers in our urban/suburban advanced life support EMS system (pop. 1,000,000). The scenario-tested command staff, LEOs tasked with neutralizing an active shooter threat, and two RTFs of LEOs and EMS providers each tasked with triage and treatment of 11 simulated casualties scattered over 2 office building floors totaling 13,000 square feet. Trained evaluators recorded performance on 30 objective data elements related to LEO-EMS operations/communication, time intervals, and trauma care. Data were analyzed using descriptive statistics and t-tests for between group comparisons. RESULTS: Over 18 days, 69 scenario events evaluated 388 EMS providers and 468 LEOs. Overall median (90th percentile) times in minutes from dispatch were: unified command established 4.1 (5.5), RTF assembled 9.4 (13.5), first victim contact 11.9 (16.5), first victim to internal casualty collection point (CCP) 16.6 (20.8), all victims ready for evacuation 21.6 (26.0). Life-saving interventions included tourniquet placed: 96% (95% CI 92-99) and LEO placed tourniquet: 88% (79-94). Clinical delays included inappropriate chest decompression: 4% (2-9) and unnecessary initial treatment: 17% (12-25). Correct operational actions included communication with LEO to ensure EMS was safe to treat: 70% (61-77) and appropriate CCP selection: 84% (74-91). Incorrect operational actions included failure to maintain protective LEO-EMS formation: 49% (45-62) and inappropriate single patient evacuation: 20% (14-28). Limitations included the lack of a pre-training control group for this novel program. CONCLUSIONS: We described the performance of an integrated LEO-EMS Rescue Task Force response to a simulated active shooter event in a large city. In general, clinical care was appropriate while operational targets can be improved. Objective measurement of response goals may be used for benchmarking and performance improvement for active threat events.

Improving Decisions About Transport to the Emergency Department for Assisted Living Residents Who Fall.

Background: Residents of assisted living facilities who fall may not be seriously ill or injured, but policies often require immediate transport to an emergency department regardless of the patient's condition. Objective: To determine whether unnecessary transport can be avoided. Design: Prospective cohort study. Setting: One large county with a single system of emergency medical services. Participants: Convenience sample of residents in 22 assisted living facilities served by 1 group of primary care physicians. Intervention: Paramedics providing emergency medical services followed a protocol that included consulting with a physician by telephone. Measurements: The number of transports after a fall and the number of time-sensitive conditions in nontransported patients. Results: Of the 1473 eligible residents, 953 consented to participate in the study (mean age, 86 years; 76% female) and 359 had 840 falls in 43 months. The protocol recommended nontransport after 553 falls. Eleven of these patients had a time-sensitive condition. At least 7 of them received appropriate care: 4 requested and received transport despite the protocol recommendation, and 3 had minor injuries that were successfully managed on site. Three additional patients had fractures that were diagnosed by outpatient radiography. The final patient developed vomiting and diarrhea, started palliative care, and died 60 hours after the fall. At least 549 of the 553 patients (99.3% [95% CI, 98.2% to 99.8%]) with a protocol recommendation for nontransport received appropriate care. Limitation: The resources required for this program will preclude use in some locations. Conclusion: Shared decision making between paramedics and primary care physicians can prevent transport to the emergency department for many residents of assisted living facilities who fall. Primary Funding Source: None.

Measuring a Crisis: Questioning the Use of Naloxone Administrations as a Marker for Opioid Overdoses in a Large U.S. EMS System.

OBJECTIVE: The United States is currently experiencing a public health crisis of opioid overdoses. To determine where resources may be most needed, many public health officials utilize naloxone administration by EMS as an easily-measured surrogate marker for opioid overdoses in a community. Our objective was to evaluate whether naloxone administration by EMS accurately represents EMS calls for opioid overdose. We hypothesize that naloxone administration underestimates opioid overdose. METHODS: We conducted a chart review of suspected overdose patients and any patients administered naloxone in Wake County, North Carolina, from January 2013 to December 2015. Patient care report narratives and other relevant data were extracted from electronic patient care records and the resultant database was analyzed by two EMS physicians. Cases were divided into categories including "known opioid use," "presumed opioid use," "no known opioid," "altered mental status," "cardiac arrest with known opioid use," "cardiac arrest with no known opioid use," or "suspected alcohol intoxication," and then further separated based on whether naloxone was administered. Patient categories were compared by patient demographics and incident year. Using the chart review classification as the gold standard, we calculated the sensitivity and positive predictive value (PPV) of naloxone administration for opioid overdose. RESULTS: A total of 4,758 overdose cases from years 2013-15 were identified. During the same period, 1,351 patients were administered naloxone. Of the 1,431 patients with known or presumed opioid use, 57% (810 patients) received naloxone and 43% (621 patients) did not. The sensitivity of naloxone administration for the identification of patients with known or presumed opioid use was 57% (95% CI: 54%-59%) and the PPV was 60% (95% CI: 57%-63%). CONCLUSION: Among patients receiving care in this large urban EMS system in the United States, the overall sensitivity and positive predictive value for naloxone administration for identifying opioid overdoses was low. Better methods of identifying opioid overdose trends are needed to accurately characterize the burden of opioid overdose within and among communities.

Acute Crisis Care for Patients with Mental Health Crises: Initial Assessment of an Innovative Prehospital Alternative Destination Program in North Carolina.

OBJECTIVE: Emergency Departments (ED) are overburdened with patients experiencing acute mental health crises. Pre-hospital transport by Emergency Medical Services (EMS) to community mental health and substance abuse treatment facilities could reduce ED utilization and costs. Our objective was to describe characteristics, treatment, and outcomes of acute mental health crises patients who were transported by EMS to an acute crisis unit at WakeBrook, a North Carolina community mental health center. METHODS: We performed a retrospective cohort study of patients diverted to WakeBrook by EMS from August 2013-July 2014. We abstracted data from WakeBrook medical records and used descriptive statistics to quantify patient characteristics, diagnoses, length of stay (LOS), and 30-day recidivism. RESULTS: A total of 226 EMS patients were triaged at WakeBrook. The median age was 38 years, 55% were male, 58% were white, and 38% were uninsured. The most common chief complaints were suicidal ideation or self-harm (46%) and substance abuse (19%). The most common diagnoses were substance-related and addictive disorders (42%), depressive disorders (32%), and schizophrenia spectrum and other psychotic disorders (22%). Following initial evaluation, 28% of patients were admitted to facilities within WakeBrook, 40% were admitted to external psychiatric facilities, 18% were stabilized and discharged home, 5% were transferred to an ED within 4 hours for further medical evaluation, and 5% refused services. The median LOS at WakeBrook prior to disposition was 12.0 hours (IQR 5.4-21.6). Over a 30-day follow-up period, 60 patients (27%) had a return visit to the ED or WakeBrook for a mental health issue. CONCLUSIONS: A dedicated community mental health center is able to treat patients experiencing acute mental health crises. LOS times were significantly shorter compared to regional EDs. Successful broader programmatic implementation could improve care quality and significantly reduce the volume of patients treated in the ED for acute mental health disorders.