The “Crisis and Concerns: A cross-sectional study on Junior Doctors during COVID-19 pandemic”.

Introduction: Doctors committed to the care of patients are trying to fulfill their duty in a difficult situation, but the consequential impacts of COVID-19 outbreak on Junior Doctors mental health are far too complex. Objective: The objective of this study was to gain insight into Junior Doctors’levels of concerns during a global pandemic of COVID-19. Methods: A quantitative cross-sectional study, utilizing a 23-item validated questionnaire was conducted on two hundred and fifty doctors of various disciplines and nationalities working on training and non-training posts. Results: The response rate was (77 %) and the levels of concern score among Junior doctors during COVID-19 pandemic in the NHS was found to be (41.35±4.9). With regard to gender differences, 32.5% of males and 27.5% females had a moderate concern score of 40 to 47. In regard to safety at work (85%) agreed that they are not safe, (100%) agreed that they are at risk to contract a COVID-19 infection at work and (100%) felt that they will transmit COVID-19 to their families but (95%) agreed that were obliged to take care of patients. In addition, (97.5%) doctors were not confident with the current infection control measures and (77.5%) felt frustrated with poor infection control training offered to them. Conclusion: Therefore, measures to strengthen personal protection and adequate support to Junior Doctors should be addressed urgently by the Healthcare system. KEYWORDS: COVID-19, Doctors, PPE, Concerns, Psychological impact.  

Role of Type Vi Secretion System in Stress Adaptations and Developing Control Strategies for Campylobacter Jejuni

The human Gram-negative enteropathogen Campylobacter jejuni is the leading cause of gastroenteritis, with 25% of the cases worldwide being attributed to this organism. C. jejuni, due to emergence of multidrug resistances, has also been classified as “high priority pathogens” thus making surveillance system as an essential key for the control of disease. However, in developing countries like Pakistan, the prevalence and source prediction of MDR C. jejuni isolates has been under reported due to lack of proper surveillance program. Therefore, the first section of the present study was conducted to determine the prevalence of C. jejuni isolates among pediatric diarrheal cases in Pakistan, their origin through source tracking as well as associated antibiotic resistance patterns. C. jejuni was detected in 54.6% of the total samples and more than 75% of the isolated strains were resistant to panel of 8 out of 13 antibiotics tested however; high level of susceptibility was observed against imipenem (12.2%) and tigecycline (9.7%). Moreover, six isolates (7.3%) were metallo-beta lactamase (MBL) producer and were positive for at least one of the five screened metallo-beta lactamase genes. Source tracking using source predictive PCR, showed that 57.3% of the isolates belonged to livestock associated cluster (C1 to C6) and 42.8% were assigned to nonlivestock/environmental clusters (C7-C9). Isolates belonging to livestock cluster had high MAR index as compared to non-livestock suggesting possible transmission of antimicrobial resistant C. jejuni strains to human population via food chain. Moreover, it also suggests that extensive use of antibiotics for disease control and growth promotion in livestock especially in poultry industry results in the emergence and spread of AMR C. jejuni and the genes associated with resistance. C. jejuni, although being a fastidious microorganism, is able to survive and maintain its pathogenesis in the harsh environments. However, the mechanisms of stress adaptations and pathogenesis is C. jejuni are not fully elucidated. Type VI secretion system among several gram-negative bacteria have been associated with virulence and stress xii adaptations but its role in stress tolerance and pathogenesis of C. jejuni is still unknown. Moreover, in contrast to the other T6SS positive bacteria, C. jejuni has only one hemolysin coregulated protein (Hcp) protein, the hallmark of a functional T6SS system, whose structure and secretory function was not known. Therefore, the second aim of this study was to determine the role of type VI secretion system in biofilm formation (a stress adaptive mechanism) and pathogenesis. Moreover, the structure and function of Hcp protein was also elucidated. Comparative analysis of effect of different sub-lethal stresses on biofilm forming potential of isolates having fully functional T6SS (Hcp+) and those lacking functional T6SS (Hcp-) showed that the biofilm formation was significantly more enhanced in T6SS positive (Hcp+) groups as compared to T6SS negative (Hcp-) group under heat (55ºC) and oxidative stresses (8mM of H2O2). Structural analysis of Hcp showed similarities between the hexameric ring structure of Hcp-Cj and that of Hcp3 from Pseudomonas aeruginosa. Through functional studies, two roles for Hcp-Cj were identified i.e., in cytotoxicity toward HepG2 cells and in biofilm formation in C. jejuni. Structure-based mutational analyses showed that an Arginine to Alanine mutation at position 30 within the extended loop of Hcp-Cj resulted in a significant decrease in cytotoxicity. However, biofilm formation function remained unaffected by this mutation. Collectively, this study supports the dual role of Hcp-Cj as a structural and effector protein in C. jejuni. High prevalence of C. jejuni associated diarrhea in both developed and developing countries can be attributed towards three major factors i) emergence of multidrug resistant C. jejuni isolates making WHO recommended antibiotics ineffective for treatment; ii) lack of control strategies at poultry level (as poultry meat consumption is the major source of infection in human); iii) persistence of C. jejuni as biofilm in poultry processing units and water reservoirs.. Therefore, third part of the present study was to develop control strategies against C. jejuni at three different levels i.e., 1) In-silico drug target identification was done using comparative proteomics and metabolomics approach by which ten essential non-homologous drug targets were identified which can be used in future to develop safe and effective drug against MDR C. jejuni ; 2) pH sensitive alginate coated chitosan nanoparticles were designed for targeted delivery of anti-campylobacter hexane fraction (PE-CANP) of Trachyspermum ammi (ajwain) into the chicken caecum without loss of its activity in gizzard to control C. jejuni at poultry farm level. The PE-CANP nanoparticles were able to reduce C. jejuni load to 6 logs (in CFU/g of caecal content) as compared to control group in chicken after 21 days of post infection and can therefore be used as a good alternative to conventional antibiotics for on-farm control of C. jejuni; 3) Three different metallic nanoparticles i.e., Silver/Graphene/TiO2, Erbium doped Li-Ni Ferrites and ZnO nanoparticle were tested for their antibacterial activity against C. jejuni which showed excellent antibacterial and anti-biofilm activity. All these nanoparticles can serve as an excellent coating material for control of C. jejuni biofilms in food processing setups and hence may control the transmission of C. jejuni to humans. Overall the present study helps in better understanding the disease transmission patterns and associated drug resistance traits as well as provides a better insight into the role of T6SS in C. jejuni in stress adaptation and thereafter developing control strategies for tackling C. jejuni both at farm and food processing level to reduce the burden of campylobacteriosis in humans.