Deciphering the Interplay of Acute Myeloid Leukemia with the Bone Marrow Microenvironment

Acute myeloid leukemia (AML) is a hematologic malignancy that leads to the accumulation of immature blasts in the bone marrow (BM). The BM is a complex organ, consisting of several cell types, including immune cells as well as non-hematopoietic stromal cells. In AML, chemotherapy may lead to long-term remission, although alloge- neic stem cell transplantation (alloSCT) often remains the only therapeutic strategy. However, not every patient responds to alloSCT and often suffer from relapse due to chemo-resistant leukemic stem cells (LSCs). One of the hypotheses associated with therapy failure is the incapability of donor T cells to recognize and eliminate LSCs, thus escaping graft-versus-leukemia (GVL) ef- fect. The first objective of this thesis was to investigate the role of T cell in alloSCT therapy outcome. Using single-cell RNA-sequencing (scRNA-seq) on BM T lymphocytes and CD34+ hematopoietic stem and progenitor cells (HSPCs) of six AML patients 100 days after alloSCT, I identified T cell signatures associated with either relapse (REL) or complete remission (CR). Among these signatures, a higher frequency of cytotoxic CD8+ effector and gamma delta T cells was observed in CR versus REL samples. Further analyses revealed that in CR, CD8+ T cells were more mature and characterized by higher cytotoxicity, while in REL CD8+ T cells were characterized by inflammatory TNF/NF-κB signaling as well as an immunosuppressive signature. In addition, this anal- ysis identified ADGRG1/GPR56 as a surface marker enriched in CR CD8+ T cells. Ad- ditional flow cytometry analyses in independent patient cohorts suggested GPR56 as a marker of cytotoxicity as well as a marker of antigen encounter post alloSCT. Together, these data provide a single-cell reference map of BM-derived T cells post alloSCT and propose GPR56 expression dynamics as a surrogate for monitoring alloSCT. One of the key drivers of AML progression is its interaction with the BM stromal microenvironment. In addition, AML is hypothesized to remodel the BM, creating a protective environment for LSCs. Thus, the second objective of this thesis was to study the impact of AML on the microenvironment and the specific contribution of LSCs in this process. For that, I combined scRNA-seq of AML xenograft models and in vitro co- cultures of patient-derived BM mesenchymal stromal cells (MSCs) with AML. These data indicated that AML presence impacts the BM composition, leading to the expansion of Cxcl12-abundant-reticular adipocyte progenitors (Adipo-CAR), decline of osteoblasts as well as disruption of the vasculature. When comparing high LSC-frequency (LSChigh) with low LSC-frequency (LSClow) AML, changes in the abundance of several stromal subsets were detected, suggesting the importance of these populations in LSC expansion. Specifically, LSChigh AML was associated with a decrease in the osteo-lineage and an expansion of multiple fibroblast subsets marked by altered extracellular matrix signa- tures. Furthermore, in vitro co-cultures uncovered similar patterns: upon LSChigh co- culture, human MSCs suppress the expression of osteoblast lineage genes, while over- expressing fibrosis-related genes. Collectively, the findings outlined in this thesis provide novel insights into the interaction of AML with the BM microenvironment, which has implications in disease progression and therapy outcome. These insights offer new opportunities for identifying intervention targets which may improve AML patient outcome

Com1 as a Promising Protein for the Differential Diagnosis of the Two Forms of Q Fever

Coxiella burnetii is the causative agent of acute and chronic Q fever in humans. Although the isolates studied so far showed a difference in virulence potential between those causing the two forms of the disease, implying a difference in their proteomic profile, the methods used so far to diagnose the two forms of the disease do not provide sufficient discriminatory capability, and human infections may be often misdiagnosed. The aim of the current study was to identify the outer membrane Com1 (CBU_1910) as a candidate protein for serodiagnostics of Q fever. The protein was cloned, expressed, purified, and used as an antigen in ELISA. The protein was then used for the screening of sera from patients suffering from chronic Q fever endocarditis, patients whose samples were negative for phase I immunoglobulin G (IgG), patients for whom at least one sample was positive for phase I IgG, and patients suffering from any kind of rheumatoid disease. Blood donors were used as the control group. Following statistical analysis, 92.4% (122/132) of the samples tested agreed with the negative clinical diagnosis, and 72.2% (26/36) agreed with the positive clinical diagnosis. Moreover, a significant correlation to the presence of the disease (p = 0.00) was calculated. The results support the idea that a Com1 antigen-based serodiagnostic test may be useful for differential diagnosis of chronic Q fever. Further studies are required to compare more immunogenic proteins of the bacterium against samples originating from patients suffering from different forms of the disease

Eljotine

Sepia tone portrait of brown mare Eljotine being held by an unidentified man near the sheds at Skowhegan Fairgrounds, Thursday, August 19, 1937, Skowhegan, Maine.https://digitalcommons.library.umaine.edu/kendall_images/3089/thumbnail.jp

Doctors’ Perceptions, Attitudes and Practices towards the Management of Multidrug-Resistant Organism Infections after the Implementation of an Antimicrobial Stewardship Programme during the COVID-19 Pandemic

Background: Greece is among the European countries with the highest consumption of antibiotics, both in community and hospital settings, including last-line antibiotics, such as carbapenems. We sought to explore doctors’ perceptions, attitudes and practices towards the management of patients with multidrug-resistant organism (MDRO) infections after the implementation of an antimicrobial stewardship programme (ASP) in a tertiary academic hospital during the COVID-19 pandemic. Methods: A self-administered, internet-based questionnaire survey was completed by doctors of the University Hospital of Heraklion in Crete, Greece. Results: In total, 202 (59.1%) hospital doctors fully completed the questionnaire. Most of them agreed that the prospective audit and feedback ASP strategy is more effective and educational than the preauthorization ASP strategy. ASP implementation prompted most respondents to monitor the continuously evolving microbiological data of their patients more closely and affected them towards a multidisciplinary and personalised care of patients with infections caused by MDROs and towards a more rigorous implementation of infection prevention and control measures. The vast majority of participants (98.5%) stated that ASP must be continued and further developed during the COVID-19 pandemic. Conclusion: The ASP implementation in our hospital had a beneficial impact on doctors’ perceptions, attitudes and practices with regard to the management of infections due to MDROs

Antimicrobial Prescribing before and after the Implementation of a Carbapenem-Focused Antimicrobial Stewardship Program in a Greek Tertiary Hospital during the COVID-19 Pandemic

Background: Irrational use of antimicrobials poses a significant risk for public health by aggravating antimicrobial resistance. The aim of this repeated point prevalence survey (PPS) was to evaluate the impact of a carbapenem-focused antimicrobial stewardship program (ASP) on overall antimicrobial use and quality of antimicrobial prescribing during the COVID-19 pandemic. Methods: All adult inpatients in the University Hospital of Heraklion in Greece were audited twice, before and after the implementation of the ASP, in October 2019 and October 2020, respectively. Patient characteristics, indications and diagnoses for antimicrobial administration, antimicrobials prescribed, and compliance with treatment guidelines were recorded. Results: Of 743 adult inpatients on the days of the two surveys, 398 (53.6%) were on antimicrobials for 437 diagnoses. Following implementation of the ASP, there was substantial decrease in the utilization of carbapenems (4.9% of all antibacterials prescribed in the second PPS compared to 10.3% in the first PPS). A significant improvement was observed for all indicators of the quality of antimicrobial prescribing. Conclusions: Our study demonstrated a positive impact of an ASP implementation during the first stages of the COVID-19 pandemic on reducing the use of last-line antimicrobials and improving overall quality of antimicrobial prescribing

Asymptomatic Bacteriuria in Patients with Type 2 Diabetes Mellitus

Objectives: Asymptomatic bacteriuria (ASB) is a common finding in patients with diabetes. Moreover, patients with diabetes and ASB have a greater risk for symptomatic urinary tract infections and associated severe complications. The aim of this study was to estimate the prevalence of ASB, as well as to identify independent risk factors and related pathogens associated with ASB in female and male patients with type 2 diabetes mellitus (T2D). Methods: This prospective case-control study was performed at the University hospital, and the Venezeleion General Hospital, Heraklion, Greece between 2012 and 2019. All patients with T2D attending the diabetes and hypertension outpatient clinics at both hospitals were enrolled, and data regarding their medical history and clinical and laboratory profiles were recorded. Asymptomatic patients with positive urine cultures were assigned as cases while those with negative urine cultures were designated as controls. Results: A total of 437 adult patients of which 61% were female and 39% were male patients with a mean age of 70.5 ± 9.6 years, were enrolled. The prevalence of ASB was 20.1%, in total. ASB was noted in 27% of female participants and 9.4% of male participants. Higher glycated hemoglobin (OR = 3.921, 95%CI: 1.521–10.109, p < 0.001) and urinary tract infection within the previous year (OR = 13.254, 95%CI: 2.245–78.241, p < 0.001) were independently positively associated with ASB, while higher levels of vitamin B12 were independently negatively associated with ASB (OR = 0.994 per ng/mL, 95%CI: 0.989–0.999, p < 0.001). Conclusions: Development of ASB was associated with specific factors, some of which may be modifiable. Interestingly, high B12 was found to be negatively associated with ASB

Asymptomatic Bacteriuria in Patients with Type 2 Diabetes Mellitus

Objectives: Asymptomatic bacteriuria (ASB) is a common finding in patients with diabetes. Moreover, patients with diabetes and ASB have a greater risk for symptomatic urinary tract infections and associated severe complications. The aim of this study was to estimate the prevalence of ASB, as well as to identify independent risk factors and related pathogens associated with ASB in female and male patients with type 2 diabetes mellitus (T2D). Methods: This prospective case-control study was performed at the University hospital, and the Venezeleion General Hospital, Heraklion, Greece between 2012 and 2019. All patients with T2D attending the diabetes and hypertension outpatient clinics at both hospitals were enrolled, and data regarding their medical history and clinical and laboratory profiles were recorded. Asymptomatic patients with positive urine cultures were assigned as cases while those with negative urine cultures were designated as controls. Results: A total of 437 adult patients of which 61% were female and 39% were male patients with a mean age of 70.5 ± 9.6 years, were enrolled. The prevalence of ASB was 20.1%, in total. ASB was noted in 27% of female participants and 9.4% of male participants. Higher glycated hemoglobin (OR = 3.921, 95%CI: 1.521–10.109, p p p < 0.001). Conclusions: Development of ASB was associated with specific factors, some of which may be modifiable. Interestingly, high B12 was found to be negatively associated with ASB

Knowledge and Perceptions about COVID-19 among Health Care Workers: Evidence from COVID-19 Hospitals during the Second Pandemic Wave

Health care workers (HCWs) face a higher risk of infection, since they work at the front line of COVID-19 patients’ management. Misinterpretations of current scientific evidence among HCWs may impact the delivery of appropriate care to COVID-19 patients and increase the risk of SARS-CoV-2 transmission in the hospital setting. Moreover, knowledge may affect HCWs perceptions depending on their broad beliefs and past experiences. The aim of this study was to explore the knowledge and perceptions of HCWs regarding COVID-19 issues during the second wave of the pandemic. A cross-sectional survey, involving a printed questionnaire, was conducted from 21 October 2020 to 31 January 2021 in four tertiary care hospitals located at four distant geographical regions in Greece. In total, 294 HCWs participated in this study. The majority of HCWs provided precise responses regarding general knowledge, perceptions, and practices concerning the COVID-19 pandemic. However, responses on hand hygiene and antimicrobial use in HCWs with COVID-19 were mistaken. This study reveals a certain degree of misconceptions and knowledge gaps in HCWs everyday practice, especially regarding hand hygiene and antimicrobial use in COVID-19 patients