Proceedings of the Clinical Microbiology Open 2022-assessing clinical laboratory and industry responses to COVID-19 pandemic testing capacity challenges (part 1).

As the COVID-19 pandemic winds down, clinical and public health laboratories, along with industry partners, reflect on the successes and failures of the pandemic response. To capture the lessons learned and better prepare for the next pandemic, the Clinical Microbiology Open (CMO) assembled key stakeholders including directors of clinical laboratories, industry partners, and state and federal agencies such as the Centers for Disease Control and Prevention and the Food and Drug Administration. Participants were asked to provide their perspectives on the initial pandemic response, supply chain constraints especially during surges, staffing challenges, test triage and communication strategies, clinical informatics needs, laboratory financial impacts of SARS-CoV-2 testing, and the emergency use authorization process. This manuscript summarizes the diagnostic laboratory and industry perspectives on these issues that were presented and discussed at CMO and proposes some steps that could be taken to improve future pandemic responses.

Proceedings of the Clinical Microbiology Open 2023: discussions about pandemic preparedness.

The 4th Clinical Microbiology Open (CMO) took place in Carlsbad, California, on 10 and 11 February 2023. This event facilitated discussion between clinical and public health laboratory directors, government agencies, and industry representatives from the companies that make up ASM's Corporate Council. While many topics were discussed, much of the discussion focused on pandemic preparedness. There were four major questions addressed: (i) When is the perfect the enemy of good in pandemic testing? (ii) What other types of pathogens might cause another pandemic and how would this affect laboratory response? (iii) What research is needed to better understand the effectiveness of the pandemic response? (iv) What have we learned about the utility of self and at-home testing in future pandemics? This review serves as a summary of these discussions.

Reporting of Antimicrobial Resistance from Blood Cultures, an Antibacterial Resistance Leadership Group Survey Summary: Resistance Marker Reporting Practices from Positive Blood Cultures.

BACKGROUND: We assessed how laboratories use and handle reporting of results of rapid diagnostics performed on positive blood culture broths, with a focus on antimicrobial resistance (AMR) markers. METHODS: A survey assembled by the Antibacterial Resistance Leadership Group Diagnostics Committee was circulated from December 2020 to May 2021. The survey was sent to local hospitals, shared on the ClinMicroNet and Division C listservs, and included in a College of American Pathologists proficiency testing survey. RESULTS: Ninety-six laboratories of various sizes across the United States (95%) and outside of the United States (5%) participated. Of the laboratories that had at least 1 rapid diagnostic in place (94%), significant heterogeneity in methods used and reporting practices was found across community (52%) and academic (40%) laboratories serving hospitals of various sizes. Respondents had implemented 1 to 6 different panels/platforms for a total of 31 permutations. Methods of reporting rapid organism identification and AMR results varied from listing all targets as "detected"/"not detected" (16-22%) without interpretive guidance, to interpreting results (23-42%), or providing therapeutic guidance comments to patient-facing healthcare teams (3-17%). CONCLUSIONS: Current approaches to reporting molecular AMR test results from positive blood culture vary significantly across clinical laboratories. Providing interpretative comments with therapeutic guidance alongside results reported may assist clinicians who are not well-versed in genetic mechanisms of AMR. However, this is currently not being done in all clinical laboratories. Standardized strategies for AMR gene result reporting are needed.

Emergomyces pasteurianus in Man Returning to the United States from Liberia and Review of the Literature.

A 65-year-old man with HIV sought treatment for fever, weight loss, and productive cough after returning to the United States from Liberia. Fungal cultures grew Emergomyces pasteurianus, and the patient's health improved after beginning voriconazole. We describe the clinical case and review the literature, treatment, and susceptibilities for E. pasteurianus.

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.

Point-Counterpoint: Should Clinical Microbiology Laboratories Report Vancomycin MICs?

INTRODUCTIONWith numerous reported challenges to reporting MICs for vancomycin, clinical laboratories are attempting to identify accurate methods for MIC testing. However, the issues of poor reproducibility, accuracy, and clinical utility remain a challenge. In this Point-Counterpoint, Dr. Sara Revolinski discusses the pros of reporting MICs for vancomycin, while Dr. Christopher Doern argues for the use of caution.

The Slow March toward Rapid Phenotypic Antimicrobial Susceptibility Testing: Are We There Yet?

Antimicrobial susceptibility testing (AST) provides critical information for the management of patients with infections. The gold standard methods for assessing organism susceptibility are still based on growth and require incubation over relatively long periods of time. Until now, little progress has been made in developing rapid, growth-based, phenotypic AST systems. This commentary puts the recently FDA-cleared Accelerate PhenoTest (P. Pancholi et al., J Clin Microbiol 56:e01329-17, 2018, https://doi.org/10.1128/JCM.01329-17) in context by providing a historical perspective on attempts to accelerate phenotypic susceptibility results. In addition, some promising new innovations that promise to shorten the turnaround time for phenotypic AST will be briefly reviewed.

The Confounding Role of Antimicrobial Stewardship Programs in Understanding the Impact of Technology on Patient Care.

Numerous studies have demonstrated the benefit of the combination of antimicrobial stewardship program (ASP) intervention and rapid diagnostic testing (RDT). However, few studies have attempted to study the incremental benefit of ASP and RDT, making it difficult to understand the true benefits of each intervention. This issue is discussed in the context of an article by S. H. McVane and F. S. Nolte (J Clin Microbiol 54:2476-2484, 2016, http://dx.doi.org/doi:10.1128/JCM.00996-16), with suggestions about how the findings of this study can be applied to other areas of clinical microbiology.

Diagnosis of Urinary Tract Infections in Children.

Urinary tract infections (UTIs) are a common occurrence in children. The management and laboratory diagnosis of these infections pose unique challenges that are not encountered in adults. Important factors, such as specimen collection, urinalysis interpretation, culture thresholds, and antimicrobial susceptibility testing, require special consideration in children and will be discussed in detail in the following review.

Defining the Clinical Significance of Alloscardovia omnincolens in the Urinary Tract.

The clinical significance of Alloscardovia omnincolens in the urinary tract has not been thoroughly evaluated. In this study, 15 patients with A. omnincolens present in their urine cultures were identified. A. omnincolens is only rarely associated with urinary tract symptoms and in some patients may play a commensal role.

Investigation of Linezolid Resistance in Staphylococci and Enterococci.

The objective of this study was to investigate an apparent increase in linezolid-nonsusceptible staphylococci and enterococci following a laboratory change in antimicrobial susceptibility testing from disk diffusion to an automated susceptibility testing system. Isolates with nonsusceptible results (n = 27) from Vitek2 were subjected to a battery of confirmatory testing which included disk diffusion, Microscan broth microdilution, Clinical and Laboratory Standards Institute (CLSI) reference broth microdilution, gradient diffusion (Etest), 23S rRNA gene sequencing, and cfr PCR. Our results show that there is poor correlation between methods and that only 70 to 75% of isolates were confirmed as linezolid resistant with alternative phenotypic testing methods (disk diffusion, Microscan broth microdilution, CLSI broth microdilution, and Etest). 23S rRNA gene sequencing identified mutations previously associated with linezolid resistance in 16 (59.3%) isolates, and the cfr gene was detected in 3 (11.1%) isolates. Mutations located at positions 2576 and 2534 of the 23S rRNA gene were most common. In addition, two previously undescribed variants (at positions 2083 and 2345 of the 23S rRNA gene) were also identified and may contribute to linezolid resistance.

Low Utility of Pediatric Isolator Blood Culture System for Detection of Fungemia in Children: a 10-Year Review.

The use of the Wampole Isolator 1.5-ml pediatric blood culture tube for the detection of fungemia in children was assessed by a 10-year retrospective review at two pediatric hospitals, The Hospital for Sick Children in Toronto, Canada, and the Children's Medical Center of Dallas, Texas. Over this period, a total of 9,442 pediatric Isolator specimens were processed, with yeast or yeast-like organisms recovered in 297 (3.1%) of the specimens (151 [1.6%] unique clinical episodes) and filamentous or dimorphic fungi recovered in 31 (0.3%) of the specimens (25 unique clinical episodes). Only 18 of the 151 clinical episodes of fungemia attributable to yeast were not detected by automated blood culture systems. The majority of isolated yeast were Candida spp., which were usually detected by automated systems, whereas the most common non-Candida yeast was Malassezia furfur, which the automated system failed to detect. Filamentous or dimorphic fungi were detected in 25 episodes, of which only 9 (36%) episodes were deemed clinically significant after chart review, indicating that in the majority of cases (16/25, 64%) fungal isolation represented contamination. In five of the nine clinically significant episodes, the isolated fungus (Histoplasma capsulatum, Coccidioides immitis/posadasii, Fusarium oxysporum, Aspergillus spp., and Bipolaris spp.) was also identified in other clinical specimens. Over the 10-year study period, the use of the pediatric Isolator system, at the discretion of the treating physician, only rarely provided useful clinical information for the diagnosis of fungemia in children, with the exception of M. furfur and possibly endemic mycoses.

Helicobacter pylori Clarithromycin Resistance and Treatment Failure Are Common in the USA.

BACKGROUND: Helicobacter pylori antibiotic resistance leads to frequent treatment failure. However, the current US prevalence of H. pylori clarithromycin resistance and treatment failure is unknown. AIMS: To determine the prevalence of clarithromycin-resistant H. pylori and its impact on treatment failure in the USA. METHODS: A multicenter, retrospective, cohort study for clarithromycin-resistant H. pylori was conducted over four academic medical centers in different geographic regions of the USA. Gastric biopsy material, residual from standard clinical pathologic examination, was examined for clarithromycin resistance by DNA sequencing of H. pylori 23S rRNA. RESULTS: One hundred and twenty-four cases of H. pylori gastritis were examined from medical centers in four different geographic regions of the USA. The overall prevalence of clarithromycin resistance was 32.3 % (range 23.1-45.8 %). There was no significant difference in the prevalence of clarithromycin resistance by study site, gender, age, or race/ethnicity. In a subset of 67 patients that had clinical follow-up data, the overall prevalence of clarithromycin resistance was 31.3 %. There was a 2.9-fold increase (p = 0.002) in treatment failure for cases with clarithromycin resistance (57.1 %) compared to wildtype H. pylori (19.6 %). CONCLUSIONS: H. pylori clarithromycin resistance in the USA exceeds the estimated 20 % prevalence compatible with successful empiric antibiotic therapy. This resistance resulted in a significant rate of treatment failure in all sites surveyed. Empiric therapy in the USA should be used with caution until there is better regional or local determination of H. pylori antibiotic resistance.

Diagnostic Algorithm for the Diagnosis of Pediatric Parasitic Gastroenteritis.

BACKGROUND: Current practices for ordering stool studies in patients with abdominal and gastrointestinal symptoms are not standardized. We hypothesized that an algorithm involving first-line use of a Cryptosporidium/Giardia combination antigen test and stricter use of ova and parasite (O&P) examinations would be clinically and cost effective. METHODS: In this study, stool O&P test results for pediatric patients in Dallas, Texas, were reviewed. All results obtained between 2009 and 2012 were included. Patient charts were reviewed to determine test results, symptoms, treatment, travel, and past medical history. Using these data, a retrospective modeling study was done to evaluate the utility of a diagnostic algorithm that limits O&P testing to those patients who are immunocompromised or have travelled outside the United States. RESULTS: Over the 3-year period of this study, we found that the prevalence of gastrointestinal parasitic disease in children was 1.9%. Analysis of the diagnostic algorithm for the judicious use of stool O&P showed that as much as 65% of testing may be unnecessary and could be eliminated. CONCLUSIONS: Our findings show that the prevalence of pediatric gastrointestinal parasitic disease in Texas may be lower than expected. In addition, these data show that a diagnostic algorithm limiting O&P testing may be both clinically and cost effective in low-prevalence settings. However, such an algorithm would miss a significant number of infections due to Dientamoeba fragilis and Blastocystis hominis.

The Clinical Usefulness of Polymerase Chain Reaction as a Supplemental Diagnostic Tool in the Evaluation and the Treatment of Children With Septic Arthritis.

INTRODUCTION: Culture-negative septic arthritis occurs frequently in children. The supplemental use of polymerase chain reaction (PCR) techniques improves the detection of bacteria in the joint fluid. This study evaluates the clinical utility of PCR at a tertiary pediatric medical center. METHODS: Children with septic arthritis were studied prospectively from 2012 to 2014. Culture results and clinical infection parameters were recorded. PCR was performed whenever sufficient fluid was available from the joint aspiration. A statistical comparison was made for the rates of identification of the causative organism by these methods. A subgroup analysis was performed to assess the correspondence of clinical and laboratory parameters with the results of joint fluid culture and PCR. RESULTS: Ninety-nine children with septic arthritis were enrolled consecutively. A broad range of parameter results was identified among these children with an average of 3.6 of 6 parameters per child that met thresholds of infection. Joint fluid cultures were positive in 34 of 97 (35.1%) children from whom they were sent. Among the 68 children from whom the material was sent for PCR, the result was positive in 32 (47.1%). The combination of blood culture, joint fluid culture, and PCR resulted in bacterial detection in 49 of 97 (50.5%) children. PCR improved the rate of detection of Kingella kingae markedly when compared with joint fluid culture. PCR results were available at an average of 14.6 days after the acquisition of joint fluid. 16S PCR results were reported at an average of 17.5 days, whereas Kingella PCR took 5.1 days. DISCUSSION: PCR provides supplemental information for diagnostic confirmation through an increased rate of detection of bacteria. The timing of results and the inability to provide antibiotic sensitivity are factors that limit its clinical usefulness currently.

GRP78/Dna K Is a Target for Nexavar/Stivarga/Votrient in the Treatment of Human Malignancies, Viral Infections and Bacterial Diseases.

Prior tumor cell studies have shown that the drugs sorafenib (Nexavar) and regorafenib (Stivarga) reduce expression of the chaperone GRP78. Sorafenib/regorafenib and the multi-kinase inhibitor pazopanib (Votrient) interacted with sildenafil (Viagra) to further rapidly reduce GRP78 levels in eukaryotes and as single agents to reduce Dna K levels in prokaryotes. Similar data were obtained in tumor cells in vitro and in drug-treated mice for: HSP70, mitochondrial HSP70, HSP60, HSP56, HSP40, HSP10, and cyclophilin A. Prolonged 'rafenib/sildenafil treatment killed tumor cells and also rapidly decreased the expression of: the drug efflux pumps ABCB1 and ABCG2; and NPC1 and NTCP, receptors for Ebola/Hepatitis A and B viruses, respectively. Pre-treatment with the 'Rafenib/sildenafil combination reduced expression of the Coxsackie and Adenovirus receptor in parallel with it also reducing the ability of a serotype 5 Adenovirus or Coxsackie virus B4 to infect and to reproduce. Sorafenib/pazopanib and sildenafil was much more potent than sorafenib/pazopanib as single agents at preventing Adenovirus, Mumps, Chikungunya, Dengue, Rabies, West Nile, Yellow Fever, and Enterovirus 71 infection and reproduction. 'Rafenib drugs/pazopanib as single agents killed laboratory generated antibiotic resistant E. coli which was associated with reduced Dna K and Rec A expression. Marginally toxic doses of 'Rafenib drugs/pazopanib restored antibiotic sensitivity in pan-antibiotic resistant bacteria including multiple strains of blakpc Klebsiella pneumoniae. Thus, Dna K is an antibiotic target for sorafenib, and inhibition of GRP78/Dna K has therapeutic utility for cancer and for bacterial and viral infections.

Lack of clinical utility of urine gram stain for suspected urinary tract infection in pediatric patients.

Urinary tract infection (UTI) is one of the most common infections in children. Urine culture remains the gold standard for diagnosis, but the utility of urine Gram stain relative to urinalysis (UA) is unclear. We reviewed 312 pediatric patients with suspected UTI who had urine culture, UA, and urine Gram stain performed from a single urine specimen. UA was considered positive if ≥10 leukocytes per oil immersion field were seen or if either nitrates or leukocyte esterase testing was positive. Urine Gram stain was considered positive if any organisms were seen. Sensitivity, specificity, and positive and negative predictive values were calculated using urine culture as the gold standard. Thirty-seven (12%) patients had a culture-proven UTI. Compared to urine Gram stain, UA had equal sensitivity (97.3% versus 97.5%) and higher specificity (85% versus 74%). Empirical therapy was prescribed before the Gram stain result was known in 40 (49%) patients and after in 42 (51%) patients. The antibiotics chosen did not differ between the two groups (P=0.81), nor did they differ for patients with Gram-negative rods on urine Gram stain compared to those with Gram-positive cocci (P=0.67). From these data, we conclude that UA has excellent negative predictive value that is not enhanced by urine Gram stain and that antibiotic selection did not vary based on the urine Gram stain result. In conclusion, the clinical utility of urine Gram stain does not warrant the time or cost it requires.

Detection of intracellular parasites by use of the CellaVision DM96 analyzer during routine screening of peripheral blood smears.

Conventional microscopy is the gold standard for malaria diagnosis. The CellaVision DM96 is a digital hematology analyzer that utilizes neural networks to locate, digitize, and preclassify leukocytes and characterize red blood cell morphology. This study compared the detection rates of Plasmodium and Babesia species on peripheral blood smears utilizing the CellaVision DM96 with the rates for a routine red blood cell morphology scan. A total of 281 slides were analyzed, consisting of 130 slides positive for Plasmodium or Babesia species and 151 negative controls. Slides were blinded, randomized, and analyzed by CellaVision and microscopy for red cell morphology scans. The technologists were blinded to prior identification results. The parasite detection rate was 73% (95/130) for CellaVision and 81% (105/130) for microscopy for positive samples. The interobserver agreement between CellaVision and microscopy was fair, as Cohen's kappa coefficient equaled 0.36. Pathologist review of CellaVision images identified an additional 15 slides with parasites, bringing the total number of detectable positive slides to 110 of 130 (85%). Plasmodium ovale had the lowest rate of detection at 56% (5 of 9); Plasmodium malariae and Babesia spp. had the highest rate of detection at 100% (3/3 and 6/6, respectively). The detection rate by CellaVision was 100% (23/23) when the parasitemia was ≥2.5%. The detection rate for <0.1% parasitemia was 63% (15/24). Technologists appropriately classified all negative specimens. The percentage of positive specimens detectable by CellaVision (73%) approaches results for microscopy on routine scan of peripheral blood smears for red blood cell morphology.

Empiric Antibiotics for Serious Bacterial Infection in Young Infants: Opportunities for Stewardship.

OBJECTIVES: To evaluate the causative agents of serious bacterial infection (SBI) in young infants and the optimal approach to empiric antibiotic therapy for infants with SBI. METHODS: From May 1, 2011, to December 1, 2013, pertinent clinical data were collected on previously well infants 60 days or younger with SBI as defined by a positive bacterial culture from a sterile site. Infants were identified by prospective surveillance of admissions and daily review of microbiology records. RESULTS: Two hundred sixty-five infants with SBI were identified. Mean age was 32 days (SD ±16.6 days). Twenty-nine infants had meningitis, 66 had bacteremia (37 with concomitant urinary tract infection), and 170 had urinary tract infection alone. No methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus sp., or penicillin-resistant Streptococcus pneumoniae were identified. Four extended-spectrum ß-lactamase-producing gram-negative bacilli were seen. Empiric therapy was ampicillin and gentamicin (n = 116, 44%) or third-generation cephalosporin based (n = 149, 56%). Ampicillin and gentamicin, with third-generation cephalosporins reserved for cases where meningitis is suspected, would have provided effective coverage for 98.5% of infants and unnecessarily broad therapy for 4.3%. Third-generation cephalosporins with ampicillin would have been effective for 98.5% of infants and unnecessarily broad for 83.8%. Third-generation cephalosporin monotherapy was less effective than either combination (P < 0.001). Fifty-seven percent of broad spectrum empiric therapy was continued despite culture results allowing de-escalation. CONCLUSIONS: Ampicillin/gentamicin remains an effective empiric regimen for infants 60 days or younger with suspected SBI. Use of a third-generation cephalosporin for suspected meningitis is appropriate, but cerebrospinal fluid must be obtained promptly to guide appropriate therapy.