Long-Term Outcomes Associated With ß-Lactam Allergies.

Importance: ß-lactam (BL) allergies are the most common drug allergy worldwide, but most are reported in error. BL allergies are also well-established risk factors for adverse drug events and antibiotic-resistant infections during inpatient health care encounters, but the understanding of the long-term outcomes of patients with BL allergies remains limited. Objective: To evaluate the long-term clinical outcomes of patients with BL allergies. Design, Setting, and Participants: This longitudinal retrospective cohort study was conducted at a single regional health care system in western Pennsylvania. Electronic health records were analyzed for patients who had an index encounter with a diagnosis of sepsis, pneumonia, or urinary tract infection between 2007 and 2008. Patients were followed-up until death or the end of 2018. Data analysis was performed from January 2022 to January 2024. Exposure: The presence of any BL class antibiotic in the allergy section of a patient's electronic health record, evaluated at the earliest occurring observed health care encounter. Main Outcomes and Measures: The primary outcome was all-cause mortality, derived from the Social Security Death Index. Secondary outcomes were defined using laboratory and microbiology results and included infection with methicillin-resistant Staphylococcus aureus (MRSA), Clostridium difficile, or vancomycin-resistant Enterococcus (VRE) and severity and occurrence of acute kidney injury (AKI). Generalized estimating equations with a patient-level panel variable and time exposure offset were used to evaluate the odds of occurrence of each outcome between allergy groups. Results: A total of 20 092 patients (mean [SD] age, 62.9 [19.7] years; 12 231 female [60.9%]), of whom 4211 (21.0%) had BL documented allergy and 15 881 (79.0%) did not, met the inclusion criteria. A total of 3513 patients (17.5%) were Black, 15 358 (76.4%) were White, and 1221 (6.0%) were another race. Using generalized estimating equations, documented BL allergies were not significantly associated with the odds of mortality (odds ratio [OR], 1.02; 95% CI, 0.96-1.09). BL allergies were associated with increased odds of MRSA infection (OR, 1.44; 95% CI, 1.36-1.53), VRE infection (OR, 1.18; 95% CI, 1.05-1.32), and the pooled rate of the 3 evaluated antibiotic-resistant infections (OR, 1.33; 95% CI, 1.30-1.36) but were not associated with C difficile infection (OR, 1.04; 95% CI, 0.94-1.16), stage 2 and 3 AKI (OR, 1.02; 95% CI, 0.96-1.10), or stage 3 AKI (OR, 1.06; 95% CI, 0.98-1.14). Conclusions and Relevance: Documented BL allergies were not associated with the long-term odds of mortality but were associated with antibiotic-resistant infections. Health systems should emphasize accurate allergy documentation and reduce unnecessary BL avoidance.

The influence of natural disasters and multiple natural disasters on self-harm and suicidal behaviour: findings from a nationally representative cohort study of Australian adolescents.

Few studies have examined the relationship between exposure to natural hazards and suicide and self-harm in youth. We extend prior research by investigating the association between multiple disasters and the risks of self-harm and suicide longitudinally in a nationally representative longitudinal cohort of adolescents 14 to 15 years to 18-19 years of age. Natural disasters were identified through parental self-reports for the local area. Different types of multiple disaster exposures were investigated including compound disasters (two or more disasters occurring in the last 12 months), cascading disasters (a disaster that leads to another disaster in the subsequent wave) and consecutive disasters (multiple disasters within the last two years or over an eight-year period). Using 8,714 person-waves of data from 2,908 adolescents, findings from random effect models suggest that parental reports of fire or floods increase the risk of self-harm ideation, self-harm, and suicidal ideation. Compound disasters of fire/flood and drought were also associated with increased risk of suicidal thoughts. Cascading disasters of drought followed by fire/flood increased the risks of self-harm but recurrent consecutive droughts were associated with lower risks of suicidal ideation. Australian adolescents are exposed to high rates of natural disasters that increase the risk of self-harm and thoughts of self-harm and suicide. Climate change will increase risk of natural disaster exposure for all countries. Despite these increased risks, there was resilience to disaster exposure particularly in the case of recurrent drought suggesting that youth, families and communities may well develop protective strategies to support mental health.

In situ habitat clearance rates and particle size preference of indigenous Olympia oysters (Ostrea lurida) and non-native Pacific oysters (Magallana gigas) in North American Pacific coast estuaries.

The Olympia oyster, Ostrea lurida, is the target of many restoration projects along estuaries on the North American Pacific coast, while the non-native Pacific oyster, Magallana gigas, dominates oyster aquaculture globally. Both species provide filtration functions that were investigated in three California bays using a whole-habitat, in situ approach, a laboratory particle selection experiment, and a regional physiological comparison. Measurements of chlorophyll α, temperature, salinity, and turbidity upstream and downstream, as well as point samples of seston total particulate matter and organic content to estimate habitat clearance rates (HCR, L hr-1 m-2) were collected. From February 2018 to June 2019, twenty-two trials were conducted across four sites. HCRs were highly variable within and among sites, ranging from site averages of -464 to 166 L hr-1 m-2, and not significantly different among sites, indicating field filtration performance of O. lurida habitat and M. gigas aquaculture is similar. Using a random forest regression, site was the most important predictor of HCR, with a variable importance score of 25.7 % (SD = 4.6 %). O. lurida and M. gigas had significantly different particle size selection preferences, likely affecting the quality of their filtration. This study's findings suggest that restoring O. lurida habitat may provide similar filtration benefits as M. gigas aquaculture, but the unique hydrodynamics and food quality of individual bays, as well as regional differences in filter feeder communities, must be considered in managing oyster habitat for filtration functions.

Does correction of carpal malalignment influence the union rate of scaphoid nonunion surgery?

The aim of this retrospective study was to assess the relation between carpal malalignment correction and radiological union rates in surgery for scaphoid nonunions. A total of 59 scaphoid waist fracture nonunions treated with open reduction and palmar tricortical autograft were divided according to their pre- and postoperative scapholunate (SL) and radiolunate (RL) angles. We found that carpal malalignment failed to correct in 32 of 59 (54.2%) patients despite meticulous surgical technique and placement of an appropriately sized wedge-shaped graft. In total, 43 (72.9%) fractures united at a mean of 4.47 months (range 3-11). Of the 27 fractures with postoperative SL and RL angles within the normal range, 21 united, whereas 22 of the 32 remaining fractures that failed to achieve postoperative angles within the normal range went on to union. The postoperative SL and RL angles were not related to union. Our findings suggest that in scaphoid fracture nonunion surgery, carpal malalignment may not be corrected in a substantial proportion of patients, but such correction may not be essential for bony union. Our findings also show that there is no marked collapse of the scaphoid graft in the early postoperative period. LEVEL OF EVIDENCE: IV.

A Pilot Study Investigating Plasma Protein Electrophoresis in One Anuran and Six Urodelan Species.

As threats to amphibian health increase, there is a growing need for diagnostic tools to assess and monitor their health status. Plasma protein electrophoresis has proven to be useful in other nonmammalian species. It enables quantification of protein fractions in plasma that may be altered in various disease processes, and is therefore useful in narrowing down differential diagnoses and detecting inflammation, in combination with other modalities such as biochemical and hematologic testing. The amphibian electrophoretogram must be defined before baseline reference intervals are obtained across species. Agarose gel electrophoresis was performed on plasma samples collected from presumed clinically normal individuals of one anuran and six urodelans: Osteopilus septentrionalis (n=2), Gyrinophilus porphyriticus (n=1), Notophthalmus viridescens (n=1), Eurycea guttolineata (n=2), Amphiuma tridactylum (n=2), Cryptobranchus alleganiensis (n=5), and Siren lacertina (n=6). The electrophoretograms varied in number of fractions between each species; however, the number of fractions was consistent within a species. An albumin migrating fraction was consistently observed in all species. A prealbumin migrating fraction was identified in species that primarily use organs other than skin for respiration. This study provides preliminary examples of a normal plasma protein electrophoretogram for seven amphibian species. Further studies quantifying reference intervals and identification of protein fractions will help establish protein electrophoresis as a useful tool in amphibian health investigations.

One Health Approach to Globalizing, Accelerating, and Focusing Amphibian and Reptile Disease Research-Reflections and Opinions from the First Global Amphibian and Reptile Disease Conference.

The world's reptiles and amphibians are experiencing dramatic and ongoing losses in biodiversity, changes that can have substantial effects on ecosystems and human health. In 2022, the first Global Amphibian and Reptile Disease Conference was held, using One Health as a guiding principle. The conference showcased knowledge on numerous reptile and amphibian pathogens from several standpoints, including epidemiology, host immune defenses, wild population effects, and mitigation. The conference also provided field experts the opportunity to discuss and identify the most urgent herpetofaunal disease research directions necessary to address current and future threats to reptile and amphibian biodiversity.

Attitudes and Behavioral Intentions of Pet Amphibian Owners About Biosecurity Practices.

Global trade has been linked with the emergence of novel pathogens and declines in amphibian populations worldwide. The potential for pathogen transmission within and between collections of captive amphibians and spillover to wild populations makes it important to understand the motivations, knowledge, attitudes and behaviors of pet amphibian owners. We surveyed US pet amphibian owners to understand their characteristics and evaluated whether and how they were associated with behavioral intentions to adopt biosecurity practices. We found that the majority of pet amphibian owners are aware of the threat of emerging pathogens, concerned about potential spillover of pathogens from captive to wild populations and willing to adopt biosecurity practices to mitigate pathogen threats. Intentions to adopt such practices were driven more by psychosocial constructs such as attitudes, perceptions and beliefs than demographic characteristics. Pet amphibian owners also expressed a strong interest in acquiring, and willingness to pay a price premium for, certified disease-free animals. These findings advance our understanding of the characteristics, motivations and behaviors of pet owners, a key stakeholder in global amphibian trade, which could help to inform new policies and outreach strategies to engage them in mitigating pathogen threats. Moreover, our results imply the economic viability of a market-based program to promote pathogen-free, sustainable trade of amphibians.

Broad host susceptibility of North American amphibian species to Batrachochytrium salamandrivorans suggests high invasion potential and biodiversity risk.

Batrachochytrium salamandrivorans (Bsal) is a fungal pathogen of amphibians that is emerging in Europe and could be introduced to North America through international trade or other pathways. To evaluate the risk of Bsal invasion to amphibian biodiversity, we performed dose-response experiments on 35 North American species from 10 families, including larvae from five species. We discovered that Bsal caused infection in 74% and mortality in 35% of species tested. Both salamanders and frogs became infected and developed Bsal chytridiomycosis. Based on our host susceptibility results, environmental suitability conditions for Bsal, and geographic ranges of salamanders in the United States, predicted biodiversity loss is expected to be greatest in the Appalachian Region and along the West Coast. Indices of infection and disease susceptibility suggest that North American amphibian species span a spectrum of vulnerability to Bsal chytridiomycosis and most amphibian communities will include an assemblage of resistant, carrier, and amplification species. Predicted salamander losses could exceed 80 species in the United States and 140 species in North America.

Assessing adverse drug reaction reports for antidiabetic medications approved by the food and drug administration between 2012 and 2017: a pharmacovigilance study.

Objective: Between 2012 and 2017, the U.S. Food and Drug Administration (FDA) approved 10 antidiabetic indicated therapies. Due to the limited literature on voluntarily reported safety outcomes for recently approved antidiabetic drugs, this study investigated adverse drug reactions (ADRs) reported in the FDA Adverse Event Reporting System (FAERS). Research Design and Methods: A disproportionality analysis of spontaneously reported ADRs was conducted. FAERS reports from January 1, 2012 to March 31, 2022 were compiled, allowing a 5-year buffer following drug approval in 2017. Reporting odds ratios were calculated for the top 10 ADRs, comparing new diabetic agents to the other approved drugs in their therapeutic class. Results: 127,525 reports were identified for newly approved antidiabetic medications listed as the primary suspect (PS). For sodium-glucose co-transporter-2 (SGLT-2) inhibitors, the odds of blood glucose increased, nausea, and dizziness being reported was greater for empagliflozin. Dapagliflozin was associated with greater reports of weight decreased. Canagliflozin was found to have a disproportionally higher number of reports for diabetic ketoacidosis, toe amputation, acute kidney injury, fungal infections, and osteomyelitis. Assessing glucagon-like peptide-1 (GLP-1) receptor agonists, dulaglutide and semaglutide were associated with greater reports of gastrointestinal adverse drug reactions. Exenatide was disproportionally associated with injection site reactions and pancreatic carcinoma reports. Conclusion: Pharmacovigilance studies utilizing a large publicly available dataset allow an essential opportunity to evaluate the safety profile of antidiabetic drugs utilized in clinical practice. Additional research is needed to evaluate these reported safety concerns for recently approved antidiabetic medications to determine causality.

Adverse drug reactions reported for antidiabetic medications Introduction: This study investigated the trends in voluntary reporting of adverse drug reactions for recently approved antidiabetic medications. Methods: Data from the FDA Adverse Events Reporting System were evaluated. The top 10 adverse drug reactions were compared between antidiabetic medications in the same therapeutic class. Results: We identified 127,525 adverse drug reaction reports for the newer approved antidiabetic medications. For SGLT-2 inhibitors, empagliflozin was associated with greater reports of blood glucose increase, nausea, and dizziness; weight decreased was reported more often for dapagliflozin; and diabetic ketoacidosis, toe amputation, acute kidney injury, fungal infections, and osteomyelitis were reported more commonly for canagliflozin. Assessing GLP-1 receptor agonists, the odds of gastrointestinal adverse drug reactions being reported was greater for dulaglutide and semaglutide. Exenatide was disproportionally associated with injection site reactions and pancreatic carcinoma reports. Conclusion: Medication safety studies using a large publicly available dataset allows an essential opportunity to evaluate the safety profile of antidiabetic drugs in the real-world setting. Additional research is needed to determine if the reported safety concerns for recently approved antidiabetic medications to determine causality.

Multi-dimensional and region-specific planning for coal retirements.

Early retirement of coal-fired power is essential for remaining in line with the 2°C target set in the Paris Agreement. Plant age plays the major role in designing retirement pathways, however, this overlooks the economic and health costs associated with coal-fired power. We introduce multi-dimensional retirement schedules that account for age, operating cost, and air pollution hazards. Results show that regional retirement pathways vary substantially with different weighting schemes. Schedules based on age would retire capacity mostly in the US and EU, whereas those based on cost or air pollution would shift the majority of near-term retirements to China and India, respectively. Our approach emphasizes that a "one-size-fits-all" strategy is ineffective in addressing global phase-out pathways. It provides the opportunity for devising region-specific pathways that are sound to the local context. Our results involve emerging economies and highlight incentives for early retirement that surpass climate change mitigation and address regional priorities.

Coccidiosis in Wild Eastern Newts (Notophthalmus viridescens): An Unexpected Finding in Consecutive Mortality Events in Tennessee, USA.

Wildlife diseases are a major threat for species conservation, and there is a growing need to implement more comprehensive disease response programs to better identify these diseases of concern. During March 2017, we observed moribund and dead eastern newts, Notophthalmus viridescens, in a single pond in middle Tennessee. All moribund individuals were emaciated. We euthanized and processed all individuals immediately on-site and later performed histopathology and quantitative PCR for ranavirus, the protist Perkinsea, and chytrid fungi Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans. One newt was positive for ranavirus. Histopathology showed no evidence of ranavirosis but did reveal overwhelming coccidiosis. Overlapping partial sequences of coccidian 18S subunit DNA showed a 96.4% match with Eimeria steinhausi, suggesting that lesions were due to a previously undescribed Eimeria sp. In 2019, two more moribund newts were encountered at the same pond. Histopathology revealed the same suspicious parasitic organisms, and one individual was positive for B. dendrobatidis. Further research on how seasonal and other environmental parameters may influence coccidia-associated morbidity and mortality is warranted. These events highlight the importance of histopathologic evaluation of mortality events and provide guidance for investigation of future outbreaks.

Corrigendum: Electrolyte imbalances and dehydration play a key role in Batrachochytrium salamandrivorans chytridiomycosis.

[This corrects the article DOI: 10.3389/fvets.2022.1055153.].

Validation of Ligand Tetramers for the Detection of Antigen-Specific Lymphocytes.

The study of Ag-specific lymphocytes has been a key advancement in immunology over the past few decades. The development of multimerized probes containing Ags, peptide:MHC complexes, or other ligands was one innovation allowing the direct study of Ag-specific lymphocytes by flow cytometry. Although these types of study are now common and performed by thousands of laboratories, quality control and assessment of probe quality are often minimal. In fact, many of these types of probe are made in-house, and protocols vary between laboratories. Although peptide:MHC multimers can often be obtained from commercial sources or core facilities, few such services exist for Ag multimers. To ensure high quality and consistency with ligand probes, we have developed an easy and robust multiplexed approach using commercially available beads able to bind Abs specific for the ligand of interest. Using this assay, we have sensitively assessed the performance of peptide:MHC and Ag tetramers and have found considerable batch-to-batch variability in performance and stability over time more easily than using murine or human cell-based assays. This bead-based assay can also reveal common production errors such as miscalculation of Ag concentration. This work could set the stage for the development of standardized assays for all commonly used ligand probes to limit laboratory-to-laboratory technical variation and experimental failure caused by probe underperformance.

Cross-protective antibodies against common endemic respiratory viruses.

Respiratory syncytial virus (RSV), human metapneumovirus (HMPV), and human parainfluenza virus types one (HPIV1) and three (HPIV3) can cause severe disease and death in immunocompromised patients, the elderly, and those with underlying lung disease. A protective monoclonal antibody exists for RSV, but clinical use is limited to high-risk infant populations. Hence, therapeutic options for these viruses in vulnerable patient populations are currently limited. Here, we present the discovery, in vitro characterization, and in vivo efficacy testing of two cross-neutralizing monoclonal antibodies, one targeting both HPIV3 and HPIV1 and the other targeting both RSV and HMPV. The 3 × 1 antibody is capable of targeting multiple parainfluenza viruses; the MxR antibody shares features with other previously reported monoclonal antibodies that are capable of neutralizing both RSV and HMPV. We obtained structures using cryo-electron microscopy of these antibodies in complex with their antigens at 3.62 Å resolution for 3 × 1 bound to HPIV3 and at 2.24 Å for MxR bound to RSV, providing a structural basis for in vitro binding and neutralization. Together, a cocktail of 3 × 1 and MxR could have clinical utility in providing broad protection against four of the respiratory viruses that cause significant morbidity and mortality in at-risk individuals.

Transferability of robotic console skills by early robotic surgeons: a multi-platform crossover trial of simulation training.

Robotic surgical training is undergoing a period of transition now that new robotic operating platforms are entering clinical practice. As this occurs, training will need to be adapted to include strategies to train across various consoles. These new consoles differ in multiple ways, with some new vendors using flat screen open source 3D enhanced vision with glasses and differences in design will require surgeons to learn new skills. This process has parallels with aviation credentialling across different aircraft described as type rating. This study was designed to test the hypothesis that technical robotic console operating skills are transferrable across different robotic operating platforms. Ten participants sequentially completed four Mimic®(Surgical Science) simulation exercises on two different robotic operating platforms (DaVinci®, Intuitive Surgical and HUGO™ RAS, Medtronic). Ethical approval and informed consent were obtained for this study. Groups were balanced for key demographics including previous robotic simulator experience. Data for simulation metrics and time to proficiency were collected for each attempt at the simulated exercise and analysed. Qualitative feedback on multi-platform learning was sought via unstructured interviews and a questionnaire. Participants were divided into two groups of 5. Group 1 completed the simulation exercises on console A first then repeated these exercises on console B. Group 2 completed the simulated exercises on console B first then repeated these exercises on console A. Group 1 candidates adapted quicker to the second console and Group 2 candidates reached proficiency faster on the first console. Participants were slower on the second attempt of the final exercise regardless of their allocated group. Quality and efficiency metrics and risk and safety metrics were equivalent across consoles. The data from this investigation suggests that console operating skills are transferrable across different platforms. Overall risk and safety metrics are within acceptable limits regardless of the order of progression of console indicating that training can safely occur across multiple consoles contemporaneously. This data has implications for the design of training and certification as new platforms progress to market and supports a proficiency-based approach.

Recommendations for the design of video-based educational interventions as instructional tools in robotic surgical training.

Video-based instructional tools are an important emerging component of robotic surgical education. However, there is a paucity of literature on the structure of video as an instructional tool in robotic surgery. This paper reviews the literature on video-based instructional tools and provides a series of recommendations for the structure of a video- based training tool in robotic surgery. Key educational aspects to consider in designing a video training tool are student engagement, content validity, cognitive load, and active learning. Key recommendations for structure of instructional videos are that videos should be short, presented in modular blocks with appropriate narration and engagement strategies.

Adjuvants influence the maturation of VRC01-like antibodies during immunization.

Once naive B cells expressing germline VRC01-class B cell receptors become activated by germline-targeting immunogens, they enter germinal centers and undergo affinity maturation. Booster immunizations with heterologous Envs are required for the full maturation of VRC01-class antibodies. Here, we examined whether and how three adjuvants, Poly(I:C), GLA-LSQ, or Rehydragel, that activate different pathways of the innate immune system, influence the rate and type of somatic mutations accumulated by VRC01-class BCRs that become activated by the germline-targeting 426c.Mod.Core immunogen and the heterologous HxB2.WT.Core booster immunogen. We report that although the adjuvant used had no influence on the durability of plasma antibody responses after the prime, it influenced the plasma VRC01 antibody titers after the boost and the accumulation of somatic mutations on the elicited VRC01 antibodies.

Identification and Optimization of a Ligand-Efficient Benzoazepinone Bromodomain and Extra Terminal (BET) Family Acetyl-Lysine Mimetic into the Oral Candidate Quality Molecule I-BET432.

The bromodomain and extra terminal (BET) family of proteins are an integral part of human epigenome regulation, the dysregulation of which is implicated in multiple oncology and inflammatory diseases. Disrupting the BET family bromodomain acetyl-lysine (KAc) histone protein-protein interaction with small-molecule KAc mimetics has proven to be a disease-relevant mechanism of action, and multiple molecules are currently undergoing oncology clinical trials. This work describes an efficiency analysis of published GSK pan-BET bromodomain inhibitors, which drove a strategic choice to focus on the identification of a ligand-efficient KAc mimetic with the hypothesis that lipophilic efficiency could be drastically improved during optimization. This focus drove the discovery of the highly ligand-efficient and structurally distinct benzoazepinone KAc mimetic. Following crystallography to identify suitable growth vectors, the benzoazepinone core was optimized through an explore-exploit structure-activity relationship (SAR) approach while carefully monitoring lipophilic efficiency to deliver I-BET432 (41) as an oral candidate quality molecule.

Efficacy of Plant-Derived Fungicides at Inhibiting Batrachochytrium salamandrivorans Growth.

The emerging fungal amphibian pathogen, Batrachochytrium salamandrivorans (Bsal), is currently spreading across Europe and given its estimated invasion potential, has the capacity to decimate salamander populations worldwide. Fungicides are a promising in situ management strategy for Bsal due to their ability to treat the environment and infected individuals. However, antifungal drugs or pesticides could adversely affect the environment and non-target hosts, thus identifying safe, effective candidate fungicides for in situ treatment is needed. Here, we estimated the inhibitory fungicidal efficacy of five plant-derived fungicides (thymol, curcumin, allicin, 6-gingerol, and Pond Pimafix®) and one chemical fungicide (Virkon® Aquatic) against Bsal zoospores in vitro. We used a broth microdilution method in 48-well plates to test the efficacy of six concentrations per fungicide on Bsal zoospore viability. Following plate incubation, we performed cell viability assays and agar plate growth trials to estimate the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of each fungicide. All six fungicides exhibited inhibitory and fungicidal effects against Bsal growth, with estimated MIC concentrations ranging from 60 to 0.156 µg/mL for the different compounds. Allicin showed the greatest efficacy (i.e., lowest MIC and MFC) against Bsal zoospores followed by curcumin, Pond Pimafix®, thymol, 6-gingerol, and Virkon® Aquatic, respectively. Our results provide evidence that plant-derived fungicides are effective at inhibiting and killing Bsal zoospores in vitro and may be useful for in situ treatment. Additional studies are needed to estimate the efficacy of these fungicides at inactivating Bsal in the environment and treating Bsal-infected amphibians.

Culture-Free Phylogenetic Analysis of Legionella pneumophila Using Targeted CRISPR/Cas9 Next-Generation Sequencing.

Currently available methods for the laboratory investigation of Legionella pneumophila outbreaks require organism culture. The ability to sequence L. pneumophila directly from clinical samples would significantly reduce delays. Here, we develop a method for targeted next-generation sequencing (NGS) of selected L. pneumophila genes utilizing a CRISPR/Cas9-based target enrichment system. We determine the method's utility by typing cultured L. pneumophila isolates and subsequently apply the method directly to patient samples. We sequenced 10 L. pneumophila isolates by 2 methods, (i) whole-genome sequencing (WGS) and (ii) targeted (CRISPR/Cas9-based) finding low-abundance sequences by hybridization (FLASH)-NGS, sequencing 57 selected genes. The targeted NGS of 57 genes was more efficient than WGS, and phylogenetic analysis of the 57 genes yielded the same classification of the L. pneumophila isolates as that based on analysis of whole-genome data. Furthermore, targeted NGS of L. pneumophila performed directly on patient respiratory samples correctly classified the patients according to their corresponding cultured isolates. This provides proof of concept that targeted NGS can be used to sequence L. pneumophila directly from patient samples. Studies on a larger number of patient samples will further validate this method. Nonetheless, CRISPR/Cas9 targeted NGS methods have the potential to be widely applicable to microbial-outbreak investigations in the future, particularly in the context of difficult and slow-growing organisms. IMPORTANCE The bacterium Legionella pneumophila is responsible for outbreaks of serious and life-threatening pneumonia called Legionnaires' disease. There is a need for new molecular methods that allow investigation of Legionella outbreaks directly from patient samples, without the need for prior microbiological culture, which causes delays. Our study aims to address this problem. We have utilized a CRISPR/Cas9-based targeted next-generation sequencing (NGS) method that can be applied directly on human specimens. Furthermore, we show that analysis of the sequences of a small number of targeted genes offers the same classification of L. pneumophila as that based on data derived from the whole genome. Given the rising interest globally in sequencing pathogens directly from human samples, CRISPR/Cas9 targeted NGS methods have the potential to be widely applicable to microbial-outbreak investigations in the future, particularly in the context of difficult and slow-growing organisms.