Cystic Echinococcosis in Northern New Hampshire, USA.

In April 2022 and December 2022, the New Hampshire Department of Health and Human Services confirmed 2 cases of locally acquired human pulmonary cystic echinococcosis caused by Echinococcus granulosus tapeworms. Both patients reported dressing locally hunted moose and exposure to dogs.

Multi-host infection and phylogenetically diverse lineages shape the recombination and gene pool dynamics of Staphylococcus aureus.

BACKGROUND: Staphylococcus aureus can infect and adapt to multiple host species. However, our understanding of the genetic and evolutionary drivers of its generalist lifestyle remains inadequate. This is particularly important when considering local populations of S. aureus, where close physical proximity between bacterial lineages and between host species may facilitate frequent and repeated interactions between them. Here, we aim to elucidate the genomic differences between human- and animal-derived S. aureus from 437 isolates sampled from disease cases in the northeast region of the United States. RESULTS: Multi-locus sequence typing revealed the existence of 75 previously recognized sequence types (ST). Our population genomic analyses revealed heterogeneity in the accessory genome content of three dominant S. aureus lineages (ST5, ST8, ST30). Genes related to antimicrobial resistance, virulence, and plasmid types were differentially distributed among isolates according to host (human versus non-human) and among the three major STs. Across the entire population, we identified a total of 1,912 recombination events that occurred in 765 genes. The frequency and impact of homologous recombination were comparable between human- and animal-derived isolates. Low-frequency STs were major donors of recombined DNA, regardless of the identity of their host. The most frequently recombined genes (clfB, aroA, sraP) function in host infection and virulence, which were also frequently shared between the rare lineages. CONCLUSIONS: Taken together, these results show that frequent but variable patterns of recombination among co-circulating S. aureus lineages, including the low-frequency lineages, that traverse host barriers shape the structure of local gene pool and the reservoir of host-associated genetic variants. Our study provides important insights to the genetic and evolutionary factors that contribute to the ability of S. aureus to colonize and cause disease in multiple host species. Our study highlights the importance of continuous surveillance of S. aureus circulating in different ecological host niches and the need to systematically sample from them. These findings will inform development of effective measures to control S. aureus colonization, infection, and transmission across the One Health continuum.

An unusual presentation of a rare disease: acute upper limb ischemia as the presenting symptom of Whipple's Endocarditis, a case report.

BACKGROUND: Whipple's disease is known to cause multiple varied systemic symptoms, and is a well-documented cause of culture-negative endocarditis. Endocarditis secondary to Whipple disease, however, has rarely been known to present primarily as a cause of acute limb ischemia. We describe such a case here. CASE PRESENTATION: A previously healthy 40 year old man presented to the emergency department with acute-onset right arm paresthesias. On exam, he was found to be tachycardic with a VI/VI systolic ejection murmur. He was diagnosed with critical limb ischemia and severe aortic regurgitation, and echocardiography showed a large mass on his bicuspid aortic valve. Thrombectomy was performed urgently, with aortic valve repair the following day. As blood cultures and valvular tissue culture remained unrevealing, the patient remained on empiric vancomycin and ceftriaxone for culture-negative endocarditis. 16 s rRNA nucleic acid amplification testing (NAAT) of his formalin-fixed, paraffin-embedded valvular tissue detected T. whipplei, after which the patient was transitioned to ceftriaxone and trimethoprim-sulfamethoxazole for a year of therapy. He continues to do clinically well. CONCLUSIONS: We report an unusual presentation of Whipple endocarditis as an acute upper limb ischemia, absent other classic symptoms of Whipple's disease, and with diagnosis made by 16 s rRNA NAAT of valvular tissue in the setting of culture-negative endocarditis.

Clinical Decision Support Trees Can Help Optimize Utilization of Anaplasma phagocytophilum Nucleic Acid Amplification Testing.

Anaplasmosis, a tick-borne illness caused by Anaplasma phagocytophilum (AP), presents with nonspecific clinical symptoms, including fever and headache, and is often accompanied by laboratory abnormalities of leukopenia, thrombocytopenia, and mildly elevated liver function tests (LFTs). Laboratory confirmation of acute infection occurs with nucleic acid amplification testing (NAAT). This retrospective cohort study aimed to develop a clinical decision support algorithm to aid in decision-making about test ordering. A data set was constructed with AP NAAT results and time-adjacent complete blood count and LFT results for adult patients tested for AP in a 12.5-year period. A second, smaller data set matched each patient with a positive AP NAAT to two patients with negative tests. Chart review for clinical symptoms was performed on this smaller data set. A decision tree algorithm was deployed to identify patient clusters with negative AP NAAT results. A total of 137/1,204 (11%) patients tested positive by NAAT for AP. In the larger, laboratory-only data set (n = 1,204), patients with a platelet count of >177 × 103/µl and age of <48 years had a negative AP NAAT (204/1,204, 17%, P < 0.05). In the smaller, cohorted data set with chart review (n = 402), patients with a platelet count of >188 × 103/µl and no fever or chills also did not have positive AP NAAT (58/402, 14%, P < 0.05). We generated two decision trees that can help determine the utility of AP NAAT using readily available clinical and laboratory data. These have the potential to significantly reduce unnecessary AP testing.

Recognition of Diagnostic Gaps for Laboratory Diagnosis of Fungal Diseases: Expert Opinion from the Fungal Diagnostics Laboratories Consortium (FDLC).

Fungal infections are a rising threat to our immunocompromised patient population, as well as other nonimmunocompromised patients with various medical conditions. However, little progress has been made in the past decade to improve fungal diagnostics. To jointly address this diagnostic challenge, the Fungal Diagnostics Laboratory Consortium (FDLC) was recently created. The FDLC consists of 26 laboratories from the United States and Canada that routinely provide fungal diagnostic services for patient care. A survey of fungal diagnostic capacity among the 26 members of the FDLC was recently completed, identifying the following diagnostic gaps: lack of molecular detection of mucormycosis; lack of an optimal diagnostic algorithm incorporating fungal biomarkers and molecular tools for early and accurate diagnosis of Pneumocystis pneumonia, aspergillosis, candidemia, and endemic mycoses; lack of a standardized molecular approach to identify fungal pathogens directly in formalin-fixed paraffin-embedded tissues; lack of robust databases to enhance mold identification with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; suboptimal diagnostic approaches for mold blood cultures, tissue culture processing for Mucorales, and fungal respiratory cultures for cystic fibrosis patients; inadequate capacity for fungal point-of-care testing to detect and identify new, emerging or underrecognized, rare, or uncommon fungal pathogens; and performance of antifungal susceptibility testing. In this commentary, the FDLC delineates the most pressing unmet diagnostic needs and provides expert opinion on how to fulfill them. Most importantly, the FDLC provides a robust laboratory network to tackle these diagnostic gaps and ultimately to improve and enhance the clinical laboratory's capability to rapidly and accurately diagnose fungal infections.

Considerations from the College of American Pathologists for Implementation of an Assay for SARS-CoV-2 Testing after a Change in Regulatory Status.

The U.S. Food & Drug Administration (FDA) regulates the marketing of manufacturers' in vitro diagnostic tests (IVDs), including assays for the detection of SARS-CoV-2. The U.S. government's Clinical Laboratory Improvement Amendments (CLIA) of 1988 regulates the studies that a clinical diagnostic laboratory needs to perform for an IVD before placing it into use. Until recently, the FDA has authorized the marketing of SARS-CoV-2 IVDs exclusively through the Emergency Use Authorization (EUA) pathway. The regulatory landscape continues to evolve, and IVDs will eventually be required to pass through conventional non-EUA FDA review pathways once the emergency declaration is terminated, in order to continue to be marketed as an IVD in the United States. When FDA regulatory status of an IVD changes or is anticipated to change, the laboratory should review manufacturer information and previously performed internal verification studies to determine what, if any, additional studies are needed before implementing the non-EUA version of the IVD in accordance with CLIA regulations. Herein, the College of American Pathologists' Microbiology Committee provides guidance for how to approach regulatory considerations when an IVD is converted from EUA to non-EUA status.

Limitations of current practices in detection of bacterially contaminated blood products associated with suspected septic transfusion reactions.

BACKGROUND: In the setting of suspected septic transfusion reactions, bacterial culture of both the transfused patient and the residual blood component is recommended. Primary bacterial contamination can occur at the time of component collection. Clinically insignificant "secondary contamination" can occur during post-transfusion component discard, retrieval for culture, or manipulation of the bag at the time of culture sampling. STUDY DESIGN AND METHODS: This retrospective, multi-center study analyzes positive residual component culture results and companion patient blood cultures from 15 hospitals, 1 blood center, and all cultured transfusion reactions within the province of Quebec, Canada, over a 5-year period. Imputability was assigned as "definite" (concordant growth), "possible" (discordant growth or lack of growth in patient culture), or "unable to assess" (patient not cultured). RESULTS: There were 373 positive component cultures from 360 unique transfusion reactions, with 276 (76.7%) companion patient blood cultures performed, of which 10 (2.8%) yielded the pathogen detected in the positive component. Of these 10 definite pathogens, 7 (2 Staphylococcus aureus, 3 other staphylococci, and 1 Streptococcus pyogenes and 1 Bacillus sp.) were associated with platelet and 3 (Aeromonas veronii, Staphylococcus epidermidis, and Enterococcus faecalis) with RBC transfusions. RBC and plasma components comprised 70% of positive component cultures. DISCUSSION: The process of performing residual component culture is vulnerable to secondary contamination. The significance of microorganisms recovered from component culture cannot be interpreted in isolation. In the context of low prevalence of primary contamination of blood components, the positive predictive value of a positive component culture result is very low.

Genomic epidemiology of methicillin-resistant and -susceptible Staphylococcus aureus from bloodstream infections.

BACKGROUND: Bloodstream infections due to Staphylococcus aureus cause significant patient morbidity and mortality worldwide. Of major concern is the emergence and spread of methicillin-resistant S. aureus (MRSA) in bloodstream infections, which are associated with therapeutic failure and increased mortality. METHODS: We generated high quality draft genomes from 323 S. aureus blood culture isolates from patients diagnosed with bloodstream infection at the Dartmouth-Hitchcock Medical Center, New Hampshire, USA in 2010-2018. RESULTS: In silico detection of antimicrobial resistance genes revealed that 133/323 isolates (41.18%) carry horizontally acquired genes conferring resistance to at least three antimicrobial classes, with resistance determinants for aminoglycosides, beta-lactams and macrolides being the most prevalent. The most common resistance genes were blaZ and mecA, which were found in 262/323 (81.11%) and 104/323 (32.20%) isolates, respectively. Majority of the MRSA (102/105 isolates or 97.14%) identified using in vitro screening were related to two clonal complexes (CC) 5 and 8. The two CCs emerged in the New Hampshire population at separate times. We estimated that the time to the most recent common ancestor of CC5 was 1973 (95% highest posterior density (HPD) intervals: 1966-1979) and 1946 for CC8 (95% HPD intervals: 1924-1959). The effective population size of CC8 increased until the late 1960s when it started to level off until late 2000s. The levelling off of CC8 in 1968 coincided with the acquisition of SCCmec Type IV in majority of the strains. The plateau in CC8 also coincided with the acceleration in the population growth of CC5 carrying SCCmec Type II in the early 1970s, which eventually leveled off in the early 1990s. Lastly, we found evidence for frequent recombination in the two clones during their recent clonal expansion, which has likely contributed to their success in the population. CONCLUSIONS: We conclude that the S. aureus population was shaped mainly by the clonal expansion, recombination and co-dominance of two major MRSA clones in the last five decades in New Hampshire, USA. These results have important implications on the development of effective and robust strategies for intervention, control and treatment of life-threatening bloodstream infections.

Third-Generation Sequencing in the Clinical Laboratory: Exploring the Advantages and Challenges of Nanopore Sequencing.

Metagenomic sequencing for infectious disease diagnostics is an important tool that holds promise for use in the clinical laboratory. Challenges for implementation so far include high cost, the length of time to results, and the need for technical and bioinformatics expertise. However, the recent technological innovation of nanopore sequencing from Oxford Nanopore Technologies (ONT) has the potential to address these challenges. ONT sequencing is an attractive platform for clinical laboratories to adopt due to its low cost, rapid turnaround time, and user-friendly bioinformatics pipelines. However, this method still faces the problem of base-calling accuracy compared to other platforms. This review highlights the general challenges of pathogen detection in clinical specimens by metagenomic sequencing, the advantages and disadvantages of the ONT platform, and how research to date supports the potential future use of nanopore sequencing in infectious disease diagnostics.

Mucosal inflammation in Candida esophagitis has distinctive features that may be helpful diagnostically.

The diagnosis of Candida esophagitis can be challenging when the epithelium containing Candida filamentous forms is not readily seen or is entirely sloughed away. Mucosal inflammation could be helpful diagnostically, if distinctive. However it is thought to be nonspecific in Candida esophagitis. The goal of this retrospective study was to identify features of mucosal inflammation helpful in alerting a pathologist to the possibility of Candida esophagitis when Candida mycelia are not readily observed. The study group consisted of 99 consecutive cases of Candida esophagitis and a control group of 64 consecutive cases of reflux esophagitis diagnosed at our institution from 2008-2016. Band-like superficial intraepithelial neutrophils and increased intraepithelial lymphocytes were observed in 75 and 67% of Candida esophagitis cases, respectively and only in 14 and 19% of reflux esophagitis cases, respectively (p < .0001). Intraepithelial lymphocytes were peripapillary or CD4-predominant in 75% of Candida esophagitis cases with increased lymphocytes, in contrast to 17% of reflux esophagitis cases (p = .0011). Concurrent presence of intraepithelial neutrophils and increased lymphocytes showed increased specificity for Candida esophagitis and was observed in 61% of patients with Candida esophagitis and only in 2% of patients with reflux esophagitis (p < .0001). In addition, superficial band-like neutrophils were observed concurrently with increased peripapillary lymphocytes or CD4-predominant lymphocytes in 35 and 50% of Candida esophagitis cases, respectively, in contrast to no reflux esophagitis cases. Basal cell hyperplasia and elongation of stromal papillae were frequent in both groups. The data suggest that when Candida microorganisms are not readily observed, concurrent presence of superficial band-like neutrophils and increased lymphocytes may be indicative of Candida etiology of active esophagitis.

The first reported case of concurrent trimethoprim-sulfamethoxazole-induced immune hemolytic anemia and thrombocytopenia.

BACKGROUND: Drug-induced immune hemolytic anemia (DIIHA) and drug-induced immune thrombocytopenia (DIIT) are rare but dangerous complications of pharmacotherapy that may be underrecognized in hematopoietic stem cell transplant (HSCT) patients due to overlap of signs and symptoms with those of more common disease processes. CASE REPORT: A 61-year-old woman with NK-cell deficiency and GATA-2-associated myelodysplastic syndrome, status post-recent allogeneic HSCT (Day +58), presented with 3 days of acute-onset severe back pain, muscle cramps, and increasingly dark urine. She was found to be anemic, thrombocytopenic, and in acute renal failure. On admission, the direct antiglobulin test was positive for complement (C3) only. After careful review of her medication list, the possibility of DIIHA was raised. She had started taking trimethoprim-sulfamethoxazole (TMP-SMX) for Pneumocystis jiroveci pneumonia prophylaxis 24 days prior on a weekend dose schedule. Serologic tests on peripheral blood samples were performed using standard methods. Drug studies were performed at an immunohematology reference laboratory. RESULTS: The patient's serum showed hemolysis of donor red blood cells in the presence of TMP-SMX and also TMP-SMX-induced platelet antibodies. The patient was treated with transfusions, hemodialysis, and immunosuppressive agents. Her clinical condition improved and she was discharged after 8 days in stable condition. CONCLUSION: This case describes the first reported concurrent DIIHA and DIIT due to TMP-SMX-induced antibodies in an HSCT patient. DIIHA and DIIT can present a diagnostic challenge in the setting of intermittent medication dosing.

Therapeutic plasma exchange as a steroid-sparing therapy in a patient with limbic encephalitis due to antibodies to voltage-gated potassium channels.

Autoantibodies to the voltage-gated potassium channel (VGKC) complex cause a spectrum of non-paraneoplastic neurologic syndromes including limbic encephalitis (LE). We report a case of a man with LE who underwent a course of therapeutic plasma exchange (TPE) in addition to other immunomodulatory therapies and experienced sustained clinical resolution of his symptoms. This report adds to the existing literature supporting TPE in cases of LE due to VGKC complex autoantibodies.

Rapid ADAMTS13 availability impacts treatment for microangiopathic hemolytic anemia and thrombocytopenia.

Thrombotic thrombocytopenic purpura (TTP) can present with a spectrum of clinical manifestations. When TTP is in a patient's clinical differential diagnosis, therapeutic plasma exchange (TPE) should be initiated emergently. Enzyme activity level of A Disintegrin And Metalloproteinase with a Thrombospondin type 1 motif, member 13 (ADAMTS13) in conjunction with the evolving clinical picture can guide further therapy, including duration and frequency of TPE and choice of fluid replacement. Our experience switching reference laboratories to obtain a more rapid turnaround time of ADAMTS13 activity level resulted in significant changes in clinical management, including fewer overall TPE procedures and the occasional use of albumin for a portion of the replacement fluid in patients without severe deficiency of ADAMTS13 and a low index of clinical suspicion for TTP. J. Clin. Apheresis 31:419-422, 2016. © 2015 Wiley Periodicals, Inc.

Demographic fluctuations in bloodstream Staphylococcus aureus lineages configure the mobile gene pool and antimicrobial resistance.

Staphylococcus aureus in the bloodstream causes high morbidity and mortality, exacerbated by the spread of multidrug-resistant and methicillin-resistant S. aureus (MRSA). We aimed to characterize the circulating lineages of S. aureus from bloodstream infections and the contribution of individual lineages to resistance over time. Here, we generated 852 high-quality short-read draft genome sequences of S. aureus isolates from patient blood cultures in a single hospital from 2010 to 2022. A total of 80 previously recognized sequence types (ST) and five major clonal complexes are present in the population. Two frequently detected lineages, ST5 and ST8 exhibited fluctuating demographic structures throughout their histories. The rise and fall in their population growth coincided with the acquisition of antimicrobial resistance, mobile genetic elements, and superantigen genes, thus shaping the accessory genome structure across the entire population. These results reflect undetected selective events and changing ecology of multidrug-resistant S. aureus in the bloodstream.

Analytical validation of a semi-automated methodology for quantitative measurement of SARS-CoV-2 RNA in wastewater collected in northern New England.

Wastewater-based epidemiology (WBE) can be used to monitor the community presence of infectious disease pathogens of public health concern such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Viral nucleic acid has been detected in the stool of SARS-CoV-2-infected individuals. Asymptomatic SARS-CoV-2 infections make community monitoring difficult without extensive and continuous population screening. In this study, we validated a procedure that includes manual pre-processing, automated SARS-CoV-2 RNA extraction and detection workflows using both reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR) and reverse transcriptase droplet digital PCR (RT-ddPCR). Genomic RNA and calibration materials were used to create known concentrations of viral material to determine the linearity, accuracy, and precision of the wastewater extraction and SARS-CoV-2 RNA detection. Both RT-qPCR and RT-ddPCR perform similarly in all the validation experiments, with a limit of detection of 50 copies/mL. A wastewater sample from a care facility with a known outbreak was assessed for viral content in replicate, and we showed consistent results across both assays. Finally, in a 2-week survey of two New Hampshire cities, we assessed the suitability of our methods for daily surveillance. This paper describes the technical validation of a molecular assay that can be used for long-term monitoring of SARS-CoV-2 in wastewater as a potential tool for community surveillance to assist with public health efforts.IMPORTANCEThis paper describes the technical validation of a molecular assay that can be used for the long-term monitoring of SARS-CoV-2 in wastewater as a potential tool for community surveillance to assist with public health efforts.

Phylogenetic lineages and antimicrobial resistance determinants of clinical Klebsiella oxytoca spanning local to global scales.

Klebsiella oxytoca is an opportunistic pathogen causing serious nosocomial infections. Knowledge about the population structure and diversity of healthcare-associated K. oxytoca from a genomic standpoint remains limited. Here, we characterized the phylogenetic relationships and genomic characteristics of 20 K. oxytoca sensu stricto isolates recovered from bloodstream infections at the Dartmouth-Hitchcock Medical Center, New Hampshire, USA from 2017 to 2021. Results revealed a diverse population consisting of 15 sequence types (STs) that together harbored 10 variants of the intrinsic beta-lactamase gene bla OXY-2, conferring resistance to penicillins. Similar sets of antimicrobial resistance (AMR) determinants reside in multiple distinct lineages, with no one lineage dominating the local population. To place the New Hampshire K. oxytoca in a broader context, we compared them to 304 publicly available genomes of clinical isolates from 18 countries. This global clinical K. oxytoca sensu stricto population is represented by over 65 STs that together harbored resistance genes against 14 antimicrobial classes, including eight bla OXY-2 variants. Three dominant STs in the global population (ST2, ST176, ST199) circulate across multiple countries and were also present in the New Hampshire population. The global K. oxytoca population is genetically diverse, but there is evidence for broad dissemination of a few lineages carrying distinct set of AMR determinants. Our findings reveal the clinical diversity of K. oxytoca sensu stricto and its importance in surveillance efforts aimed at monitoring the evolution of this drug-resistant nosocomial pathogen. IMPORTANCE The opportunistic pathogen Klebsiella oxytoca has been increasingly implicated in patient morbidity and mortality worldwide, including several outbreaks in healthcare settings. The emergence and spread of antimicrobial resistant strains exacerbate the disease burden caused by this species. Our study showed that clinical K. oxytoca sensu stricto is phylogenetically diverse, harboring various antimicrobial resistance determinants and bla OXY-2 variants. Understanding the genomic and population structure of K. oxytoca is important for international initiatives and local epidemiological efforts for surveillance and control of drug-resistant K. oxytoca.

Antimicrobial Activity of Tebipenem and Comparators against Enterobacterales from diverse Outpatient Centers and Nursing Homes in the United States.

INTRODUCTION: Tebipenem is a potential option for the treatment of a range of infections because of its oral dosing coupled with the safety profile of the ß-lactam antimicrobial class. OBJECTIVES: To evaluate tebipenem in vitro activity against a challenge set of clinical Enterobacterales collected from outpatient and community settings. METHODS: 618 Enterobacterales isolates were submitted by 11 geographically dispersed U.S medical centers that processed cultures from affiliated outpatient centers in 2022. Susceptibility tests for tebipenem and comparator agents were performed by broth microdilution. Extended-spectrum-ß-lactamase (ESBL)-like isolates were identified phenotypically. Multidrug-resistant isolates were non-susceptible to ≥1 agent in ≥3 antimicrobial classes. Genotypic testing (CarbaR) was conducted on select isolates. RESULTS: Isolates (59% Escherichia coli) were recovered from patients seen predominantly in urology/nephrology (24%), nursing home/long-term care (21%), and ambulatory/primary care (21%) clinics. Comparator agent susceptibility rates against all isolates were as follows: levofloxacin (67.5%), amoxicillin/clavulanate (73.6%), cefixime (70.4%), cefpodoxime (70%), cephalexin (61.7%), ceftriaxone (74.4%), cefazolin (63.8%), ertapenem (97.6%), meropenem (99.7%), nitrofurantoin (64.9%), and sulfamethoxazole/trimethoprim (70.9%). Overall, 90.3% (558/619) of isolates were inhibited at a tebipenem MIC of ≤0.125 mg/L (MIC50/90, 0.016/0.125 mg/L), including 85.7% inhibition of ESBL-phenotype isolates (n=161; MIC50/90, 0.03/0.25 mg/L), 86.3% of levofloxacin and sulfamethoxazole/trimethoprim co-resistant isolates (n=95; MIC50/90, 0.016/0.25 mg/L) and 84.3% of multidrug-resistant isolates (n = 172; MIC50/90, 0.03/0.25 mg/L). Carbapenemase genes were observed in 2 ESBL-phenotype isolates with a tebipenem MIC of ≥0.5 mg/L. CONCLUSION: Relative to common oral comparators, these data demonstrate excellent tebipenem in vitro activity against Enterobacterales isolated from patients receiving care in outpatient settings, including urology clinics and nursing homes.