Senu Apewokin

Clostridioides difficile is a highly methylated organism within the gut microbiome that is responsible for Clostridioides difficile infection (CDI), a common disease that is mediated by toxins production from the bacterium. C. difficile infection is ten times more common in chemotherapy patients than the average patient, but the reasons for this disparity are unclear. Conditioning chemotherapy (CC), an integral part of cancer treatments, has the ability to induce methylation changes in many cell types. We posit that CC induces methylation changes within C. difficile that may promote toxin production and consequently CDI. To test our hypothesis, we sought to identify the epigenetic changes, particularly methylation changes, within C. difficile isolates before and after chemotherapy and within isolates that express toxin and isolates that do not. After stool sampling, we isolated C. difficile by culture then sequenced and created a hybrid assembly of each isolate using nanopore long read sequencing and Illumina short read sequencing. Bioinformatics tools such as Dorado and Samtools were used to basecall and determine methylation states, while Unicycler was used for genome assembly. Methylartist was then used for data visualization. Genome-wide methylation profiling revealed distinct epigenetic signatures in Clostridioides difficile associated with toxin expression and chemotherapy exposure. Whole-genome 6mA analysis demonstrated significant differences between toxin-positive and toxin-negative isolates, with prominent methylation changes in tcdA and tcdE, while selected sporulation genes were unmethylated in toxin-negative strains. Chemotherapy was associated with a significant shift in global 6mA methylation patterns. Targeted 5mC analysis of the pathogenicity locus revealed reduced methylation around tcdB and across multiple toxin genes following chemotherapy, whereas sporulation genes remained unaffected. These findings indicate chemotherapy-associated epigenetic remodeling of toxin-associated loci in C. difficile.

Background: Antibiotic stewardship during stem cell transplantation (SCT) is challenging.. Procalcitonin (PCT) has been employed successfully in critical care patients to safely guide stewardship. However, procalcitonin guided stewardship has not been robustly assessed in SCT recipients. We sought to evaluate the potential utility of PCT to guide antimicrobial de-escalation during engraftment. Methods: 100 SCT patients were prospectively enrolled in a “strategy trial” and had infectious complications documented. Lab parameters - CBC, BMP, CRP were obtained daily as standard of care (SOC) while PCT was obtained for research purposes. Providers were blinded to PCT results. We compared duration of antimicrobial escalation between actual events (SOC model) and a proposed PCT model. In this hypothetical PCT model, antibiotic de-escalation would occur if CRP remained <100 mg/dl and PCT <0.25 ng/ml after 3 days of escalation. Escalation events were defined as a substitution or addition of an antimicrobial agent after initiation of prophylactic antimicrobials. Results: 77 patients had escalation events and of these, 33 had bacterial infections. A total of 136 antimicrobial escalations events were identified, and of these only 39(28.7%) were associated with documented infections. The standard of care model had a mean duration (+SD) of 9.08 (+ 6.08) antibiotic days. If the PCT model were employed, the mean duration (+SD) would be 4.44 (+ 6.16) days (p<0.001). The PCT model, however, would have missed 11 infections\ Conclusion: Procalcitonin-guided antimicrobial stewardship during autologous stem cell transplantation is feasible however optimization is necessitated for utilization as a tool to guide antibiotic prophylaxis during SCT.

Backgound: Enterococcu s and Akkermansia species enhance immune function through T-cell dependent mechanisms. This suggests vancomycin exposure and consequently the relative abundance of Enterococcus may have repercussions on immune-related outcomes such as GVHD. To test this hypothesis we assessed the a) association between vancomycin exposure and aGVHD b) association between stool Enterococcus abundance and aGVHD c) in-vitro effect of vancomycin on T-cell function. Methods: Two distinct cohorts were evaluated. 1) A “derivation” cohort comprising 46 patients where vancomycin exposure data was correlated with aGVHD 2) A “validation” cohort of 26 patients where metagenomic sequencing assessed the correlation between enterococcal abundance and aGVHD. Additionally, ELISPOT assays evaluated the impact of vancomycin exposure on T-cell function. Results: In the “derivation” cohort, aGVHD was associated with lower vancomycin exposure; (IQR 4.5 days) compared to (IQR 10.5 days) in patients without aGHVD (p-value 0.0072). Among five out of fourteen patients who developed aGVHD in “validation” cohort, significant genus enrichment for Enteroccocus and Akkermansia (5.75 and 7.59 log2fold change resp. p<0.05) was noted. Elispot assays demonstrated that T-cell exposure to vancomycin did not affect function. Conclusions: Vancomycin exposure is associated with protection from aGVHD through microbiome-intrinsic processes but not direct impairment of T-cell function.

Talia H Swartz, Senu Apewokin, Stephen M Carpenter, Rana Chakraborty, Ann M Dennis, Eric Houpt, Neeraj K Surana, Danielle M Zerr, Liise-anne Pirofski; The

Infectious diseases (ID) research is vital for global public health, typically led by physician-scientists. This Perspective addresses challenges in the ID workforce and suggests solutions. Physician-scientists have made key discoveries that have significantly impacted human health. The importance of ID research in understanding diseases, leading to treatments and vaccines, is emphasized, along with the need to address persistent and new infections, antimicrobial resistance, and threats like HIV and influenza. The paper analyzes the physician-scientist workforce's struggles, including funding, training, and research-practice integration gaps. We suggest increased funding, better training, and mentorship, more collaborative and interdisciplinary research, and improved recognition systems. The article stresses the urgency of supporting physician-scientists in ID, advocating for proactive prevention and preparedness, and calls for immediate action to enhance ID research and care.

Background: Viral infections such as adenovirus (ADV), BK virus (BKV), and cytomegalovirus (CMV) after kidney transplantation negatively impact outcomes in transplant recipients despite advancements in screening and antiviral therapy. We describe our experience of using the virus‐specific T cell therapy (VSTs) in kidney transplant recipients (KTR) at our transplant center.Methods: This is a retrospective, single center review of KTR with ADV, BKV and CMV infections between June 2021 and December 2022. These patients received third party VSTs as part of the management of infections. The immunosuppression, details of infection and outcome data were obtained from electronic medical records.Results: Two cases of ADV infection resolved after one infusion of VSTs. The response rate of BKV and CMV infection was not as robust with close to 50% reduction in median viral load after VSTs. Out of 23 patients, two patients developed chronic allograft nephropathy from membranoproliferative glomerulonephritis and acute rejection.Conclusion: Patients that are resistant to antivirals or who have worsening viremia despite conventional management may benefit from VSTs therapy to treat underlying viral infection. Additional studies are needed to ascertain efficacy and short‐ and long‐term risks secondary to VSTs. image

Background and Aims: Colorectal cancer (CRC) incidence is increasing in young patients without a clear etiology. Emerging data have implicated the fecal microbiome in CRC carcinogenesis. However, its impact on young onset CRC is poorly defined.Methods: We performed a meta‐analysis of fecal metagenomics sequencing data from n = 692 patients with CRC and n = 602 healthy controls from eleven studies to evaluate features of the fecal metagenome associated with CRC. We hypothesized that known carcinogenic virulence factors (colibactin, fadA) and species abundance may be differentially enriched in young CRC patients relative to older CRC patients and controls.Results: Summary odds ratios (OR) for CRC were increased with the presence of colibactin (OR 1.92 95% CI 1.08–3.38), fadA (OR 4.57 95% CI 1.63–12.85), and F. nucleatum (OR 6.93 95% CI 3.01–15.96) in meta‐analysis models adjusted for age, gender, and body mass index. The OR for CRC for the presence of E.coli was 2.02 (0.92–4.45). An increase in the prevalence of Fusobacterium nucleatum (OR = 1.40 [1.18; 1.65]) and Escherichia coli (OR = 1.14 [1.02; 1.28]) per 10‐year increase in age was observed in models including samples from both CRC and healthy controls. Species relative abundance was differentially enriched in young CRC patients for five species‐Intestinimonas butyriciproducens, Holdemania filiformis, Firimicutues bacterium CAG 83, Bilophilia wadsworthia, and Alistipes putredinis.Conclusion: In this study, we observed strong associations with CRC status for colibactin, fadA, and Fusobacterium nucleatum with CRC relative to controls. In addition, we identified several microbial species differentially enriched in young colorectal cancer patients. Studies targeting the young CRC patients are warranted to elucidate underlying preclinical mechanisms.

Ocular candidiasis is a known complication of candidemia. Given the poor ocular penetration of echinocandins, there is some concern that the increasing use of echinocandins may portend an increased incidence of ophthalmic complications. We examined the changing trends in antifungal prescribing patterns and the incidence of ophthalmic complications after candidemia. Patients with blood cultures positive for Candida species between January 2014 and June 2020 who underwent screening fundoscopic examination by an ophthalmologist were analyzed. The χ² analysis was used to compare antifungal prescriptions and ocular exam findings before and after 2016. Trend analysis was also performed to assess temporal changes in prescribing practices and eye exam findings. There were 226 candidemia cases during the study period, 129 (57.1%) of which underwent screening eye exams. From 2014 to 2015, 24 of 37 (64.5%) patients received eye-penetrating antifungals compared to 36 of 92 (39.1%) from 2016 to 2020 (P = .008). Overall, 30 of 129 (23.3%) patients had abnormal eye exams with the prevalence of abnormal findings being 7 of 37 (18.9%) before 2016 compared to 23 of 92 (25%, P = .46) thereafter. A trend analysis revealed an increase in abnormal eye findings over the study period (P = .008). Of the 30 patients who had abnormal eye exams, 9 (30%) had a change in systemic antifungal therapy from echinocandins to eye-penetrating antifungals. Echinocandin use was associated with abnormal eye findings. Prescription of eye-penetrating antifungals for candidemia has trended down since 2016. This was associated with a concomitant increase in abnormal findings on screening fundoscopy. Abnormal eye exams were not uncommon throughout our study period.

Pulmonary arterial hypertension (PAH) is a progressive, ultimately fatal cardiopulmonary disease associated with a number of physiologic changes, which is believed to result in imbalances in the intestinal microbiota. To date, comprehensive investigational analysis of the intestinal microbiota in human subjects is still limited. To address this, we performed a pilot study of the intestinal microbiome in 20 PAH and 20 non‐PAH healthy control subjects, recruited from a single center, with each PAH subject recruited simultaneously with a cohabitating non‐PAH control subject. Shotgun metagenomic sequencing was used to analyze the microbiome profiles. There were no differences between PAH and non‐PAH subjects across several measures of microbial abundance and diversity (Alpha Diversity, Beta Diversity, F/B ratio). The relative abundance of Lachnospiraceae bacterium GAM79 was lower in PAH stool samples as compared to non‐PAH control subject' stool. There was no strong or reproducible association between PAH disease severity and global microbial abundance, but several bacterial species (a relative abundance of Anaerostipes rhamnosivorans and a relative deficiency of Amedibacterium intestinale, Ruminococcus bicirculans, and Ruminococcus albus species were associated with disease severity (most proximal right heart catheterization hemodynamics and six‐minute walk test distance) in PAH subjects. Our results support further investigation into the presence, significance, and potential physiologic effects of a PAH‐specific intestinal microbiome.