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Item 1137. What Do We Know? Teaching Medical Students to Deal with Uncertainty as a Pandemic Unfolds(Oxford, 2020-10) Bauer, Margaret E.; Trujillo, Daniel; Brown, Cameron; Gomez, Martiza; Davidson, Darrell; Relich, Ryan F.; Allen, Bradley L.; Microbiology and Immunology, School of MedicineBackground The global COVID-19 pandemic has had a major impact on medical student education. As the pandemic spread nationwide, numerous universities shut down with only days’ notice, and medical students were removed from all patient care settings and restricted from campuses. Yet, the need and curiosity of these future physicians to understand this new disease was great, including how to interpret and integrate rapidly evolving information on the underlying viral and immune mechanisms, pathophysiology, and epidemiology. Time students spent away from patient care settings presented an opportunity to rapidly develop and deliver new curriculum covering SARS-CoV-2 and COVID-19. Methods A team of students and faculty at Indiana University developed a Fundamentals of COVID-19 course that included up-to-date information on the virology, immunology, and pathophysiology of COVID-19. The course was delivered online, with both synchronous and asynchronous activities. Virology and immunology of the coronavirus family, including current knowledge to-date of SARS-CoV-2, were delivered using a series of readings and brief videos, followed by a small group exercise that required students to choose and present to their peers a paper from the scientific literature on COVID-19. A similar approach was used to deliver content about the pathophysiology of COVID-19. To place the COVID-19 experience in context of other pandemics, students researched and educated their small group cohort on another historical pandemic. Results To measure course effectiveness, we administered a pre-course survey gauging students’ self-confidence in their knowledge of these topics; the same survey was administered after completion of the course. Surveys from 645 (89% of enrolled) 3rd and 4th year medical students who completed both surveys were analyzed. Results showed that the course elicited a 57% increase (p< 0.001) in students’ confidence in their knowledge of COVID-19 virology and immunology and a 64% increase (p< 0.001) in knowledge of the pathophysiology. Conclusion Data showed that the asynchronous content and group activities were successful in engaging and educating the students on foundational knowledge of COVID-19 and were an effective approach to rapidly evolving information when faced with a novel disease.Item Answer to August 2014 Photo Quiz(American Society for Microbiology, 2014-08) Relich, Ryan F.; Faught, Phillip R.; McCoy, Morgan H.; Alexander, Riley E.; Pritt, Bobbi S.; Pathology and Laboratory Medicine, School of MedicineItem Answer to August 2016 Photo Quiz(American Society for Microbiology, 2016-08) Relich, Ryan F.; Boyd, Kathleen M.; McCoy, Morgan H.; Kaufman, Cynthia; Simpson, Edward R.; Christenson, John C.; Department of Pathology and Laboratory Medicine, IU School of MedicineItem Antimicrobial Activity of Gallium Protoporphyrin IX against Acinetobacter baumannii Strains Displaying Different Antibiotic Resistance Phenotypes(American Society for Microbiology, 2015-12) Arivett, Brock A.; Fiester, Steven E.; Ohneck, Emily J.; Penwell, William F.; Kaufman, Cynthia M.; Relich, Ryan F.; Actis, Luis A.; Department of Pathology & Laboratory Medicine, IU School of MedicineA paucity of effective, currently available antibiotics and a lull in antibiotic development pose significant challenges for treatment of patients with multidrug-resistant (MDR) Acinetobacter baumannii infections. Thus, novel therapeutic strategies must be evaluated to meet the demands of treatment of these often life-threatening infections. Accordingly, we examined the antibiotic activity of gallium protoporphyrin IX (Ga-PPIX) against a collection of A. baumannii strains, including nonmilitary and military strains and strains representing different clonal lineages and isolates classified as susceptible or MDR. Susceptibility testing demonstrated that Ga-PPIX inhibits the growth of all tested strains when cultured in cation-adjusted Mueller-Hinton broth, with a MIC of 20 μg/ml. This concentration significantly reduced bacterial viability, while 40 μg/ml killed all cells of the A. baumannii ATCC 19606(T) and ACICU MDR isolate after 24-h incubation. Recovery of ATCC 19606(T) and ACICU strains from infected A549 human alveolar epithelial monolayers was also decreased when the medium was supplemented with Ga-PPIX, particularly at a 40-μg/ml concentration. Similarly, the coinjection of bacteria with Ga-PPIX increased the survival of Galleria mellonella larvae infected with ATCC 19606(T) or ACICU. Ga-PPIX was cytotoxic only when monolayers or larvae were exposed to concentrations 16-fold and 1,250-fold higher than those showing antibacterial activity, respectively. These results indicate that Ga-PPIX could be a viable therapeutic option for treatment of recalcitrant A. baumannii infections regardless of the resistance phenotype, clone lineage, time and site of isolation of strains causing these infections and their iron uptake phenotypes or the iron content of the media.Item Cepheid Xpert Xpress Flu/RSV evaluation performed by minimally trained non-laboratory operators in a CLIA-waived environment(Elsevier, 2022) Shihabuddin, Bashar S.; Faron, Matthew L.; Relich, Ryan F.; Van Heukelom, Paul; Mayne, Donna; Staat, Mary Allen; Selvarangan, Rangaraj; Hueschen, Leslie A.; Wolk, Donna M.; House, Stacey; Harnett, Glenn; McGann, Kevin; Steele, Mark T.; Romero, Jose R.; Arms, Joe; Lander, Owen; Loeffelholz, Michael; Strouts, Fiona; Cohen, Daniel; Pathology and Laboratory Medicine, School of MedicineThe COVID-19 pandemic highlighted the significance of readily available and easily performed viral testing for surveillance during future infectious pandemics. The objectives of this study were: to assess the performance of the Xpert Xpress Flu and/or RSV test, a multiplex PCR assay for detecting influenza A and B virus and respiratory syncytial virus nucleic acids in respiratory tract specimens, relative to the Quidel Lyra Influenza A+B assay and the Prodesse ProFlu+ assay, and the system's ease of use by minimally trained operators. Overall, the Xpert Xpress Flu/RSV test demonstrated a high positive and negative percent agreement with the comparator assays, and was easy to use and interpret results, based on the operators' feedback. We concluded that the Xpert Xpress Flu/RSV test is sensitive, specific, and easy to use for the diagnosis of influenza and RSV by minimally trained operators and can be a valuable tool in future infectious clusters or pandemics.Item Characterization of Pontibacter altruii, sp. nov., isolated from a human blood culture(Elsevier, 2017-09) Roiko, Marijo; May, Meghan; Relich, Ryan F.; Department of Pathology and Laboratory Medicine, IU School of MedicineThe genus Pontibacter is a recent addition to the family Cytophagaceae, phylum Bacteroidetes. Previous reports of its cultivation and molecular detection are from a variety of environmental sources, including marine and desert habitats. We report the first description of a Pontibacter sp., which was initially identified as Elizabethkingia meningoseptica, isolated from a human clinical specimen. On the basis of 16S rRNA gene sequence, unique mass spectral profile and phenotypic characterization, this isolate represents a novel species within the genus Pontibacter that has been named Pontibacter altruii, sp. nov., strain Grand Forks.Item Clinical comparison and agreement of PCR, antigen, and viral culture for the diagnosis of COVID-19: Clinical Agreement Between Diagnostics for COVID19(Elsevier, 2022) Agard, Amanda; Elsheikh, Omar; Bell, Drew; Relich, Ryan F.; Schmitt, Bryan H.; Sadowski, Josh; Fadel, William; Webb, Douglas H.; Dbeibo, Lana; Kelley, Kristen; Carozza, Mariel; Lei, Guang-Shen; Calkins, Paul; Beeler, Cole; Medicine, School of MedicineThe aim of this study is to compare the COVID-19 nasopharyngeal PCR (NP PCR) to antigen, nasal PCR, and viral culture. One-hundred-and-fourteen risk-stratified patients were tested by culture, nasal PCR, NP PCR, and Ag testing. Twenty (48%) of the high risk and 23 (32%) of the low risk were NP PCR positive. Compared with NP PCR, the sensitivity of nasal PCR, Sofia Ag, BinaxNOW Ag, and culture were 44%, 31%, 37%, and 15%. In the high risk group, the sensitivity of these tests improved to 71%, 37%, 50%, and 22%. Agreement between tests was highest between nasal PCR and both antigen tests. Patients who were NP PCR positive but antigen negative were more likely to have remote prior COVID-19 infection (p<0.01). Nasal PCR and antigen positive patients were more likely to have symptoms (p = 0.01).Item Comparison of Four Commercially Available Group B Streptococcus Molecular Assays Using Remnant Rectal-Vaginal Enrichment Broths(Elsevier, 2018) Relich, Ryan F.; Buckner, Rebecca J.; Emery, Christopher L.; Davis, Thomas E.; Pathology and Laboratory Medicine, School of MedicineThe incidence of neonatal Group B streptococcal (GBS) disease has significantly declined since the widespread implementation of prenatal screening of expectant mothers for urogenital and gastrointestinal tract GBS colonization. Screening methods have evolved from exclusively culture-based approaches to more rapid and highly sensitive molecular methods. We chose to evaluate the performance of four commercially available GBS molecular tests for detection of GBS colonization using 299 antepartum rectal-vaginal specimens submitted to our laboratory for routine GBS screening. In 97% of instances, there was agreement between all three systems. When testing 1, 6, and 12 samples simultaneously, all methods performed comparably, but the ARIES® GBS assay required the least total hands-on time and the illumigene® Group B Streptococcus assay required the most hands-on time.Item Comparison of nucleocapsid antigen with strand-specific reverse-transcription PCR for monitoring SARS-CoV-2 infection(Elsevier, 2023) Chang-Graham, Alexandra L.; Sahoo, Malaya K.; Huang, ChunHong; Solis, Daniel; Sibai, Mamdouh; August, Gianna; Calayag, Lira; Kenji, Obadia M.; Shi, Run-Zhang; Mostafa, Heba H.; Lei, Guang-Sheng; Relich, Ryan F.; Pinsky, Benjamin A.; Pathology and Laboratory Medicine, School of MedicineBackground: Tests that sensitively detect the presence of actively replicating SARS-CoV-2 may improve patient care by allowing the safe and timely discontinuation of isolation. Correlates of active replication include nucleocapsid antigen and virus minus-strand RNA. Methods: Qualitative agreement of the DiaSorin LIAISON SARS-CoV-2 nucleocapsid antigen chemiluminescent immunoassay (CLIA) with minus-strand RNA was determined using 402 upper respiratory specimens from 323 patients previously tested using a laboratory-developed SARS-CoV-2 strand-specific RT-qPCR. Nucleocapsid antigen levels, minus-strand and plus-strand cycle threshold values, as well as virus culture, were used to evaluate discordant specimens. Receiver operating characteristic curves were also used to identify virus RNA thresholds for active replication, including values harmonized to the World Health Organization International Standard. Results: Overall agreement was 92.0% [95% confidence interval (CI): 89.0 - 94.5], positive percent agreement was 90.6% (95% CI: 84.4 - 95.0), and negative percent agreement was 92.8% (95% CI: 89.0 - 95.6). The kappa coefficient was 0.83 (95% CI: 0.77 - 0.88). Discordant specimens contained low levels of nucleocapsid antigen and minus-strand RNA. 84.8% (28/33) were negative by culture. Sensitivity-optimized plus-strand RNA thresholds for active replication were 31.6 cycles or 3.64 log10 IU/mL; resulting in 100.0% sensitivity (95% CI: 97.6 to 100.0) and 55.9 specificity (95% CI: 49.7 to 62.0). Conclusions: Detection of nucleocapsid antigen by CLIA performs equivalently to minus-strand detection via strand-specific RT-qPCR, though these methods may overestimate replication-competent virus compared to culture. Careful implementation of biomarkers for actively replicating SARS-CoV-2 has the potential to inform infection control decision-making and patient management.Item A Comprehensive Systems Biology Approach to Studying Zika Virus(Plos, 2016-09) Relich, Ryan F.; May, Meghan; Department of Pathology and Laboratory Medicine, IU School of MedicineZika virus (ZIKV) is responsible for an ongoing and intensifying epidemic in the Western Hemisphere. We examined the complete predicted proteomes, glycomes, and selectomes of 33 ZIKV strains representing temporally diverse members of the African lineage, the Asian lineage, and the current outbreak in the Americas. Derivation of the complete selectome is an 'omics' approach to identify distinct evolutionary pressures acting on different features of an organism. Employment of the M8 model did not show evidence of global diversifying selection acting on the ZIKV polyprotein; however, a mixed effect model of evolution showed strong evidence (P<0.05) for episodic diversifying selection acting on specific sites. Single nucleotide polymorphisms (SNPs) were predictably frequent across strains relative to the derived consensus sequence. None of the 9 published detection procedures utilize targets that share 100% identity across the 33 strains examined, indicating that ZIKV escape from molecular detection is predictable. The predicted O-linked glycome showed marked diversity across strains; however, the N-linked glycome was highly stable. All Asian and American strains examined were predicted to include glycosylation of E protein ASN154, a modification proposed to mediate neurotropism, whereas the modification was not predicted for African strains. SNP diversity, episodic diversifying selection, and differential glycosylation, particularly of ASN154, may have major biological implications for ZIKV disease. Taken together, the systems biology perspective of ZIKV indicates: a.) The recently emergent Asian/American N-glycotype is mediating the new and emerging neuropathogenic potential of ZIKV; and b.) further divergence at specific sites is predictable as endemnicity is established in the Americas.