Student Projects

Discover what Master projects you can carry out in the Laboratory of Microbiology and Microtechnology. Available to students of all universities, each of them is a perfect opportunity to acquire a deep understanding of the biological context and engineering aspects of development of new technologies in Life Sciences.
| © EPFL Thomas Simonet

To apply for a student project, please contact us as specified below, indicating which one you are interested in, and attaching your resume and your cover letter. The project can then be tailored to your interests.

Bachelor Project / Master Lab Immersion Project / Master Semester Project / Master Thesis Project

Description:

Many strains of E. coli are beneficial commensals while others (pathogenic strains) can cause several gastro-intestinal and urinary tract infections (UTIs). The majority of UTIs are caused by uropathogenic E. coli (UPEC), which affects 150 million people worldwide. UTIs are an especially important cause of morbidity and mortality among infants, older people, and women (Flores-Mireles et al., 2015). UPEC infection of the urinary bladder is countered by innate defences of the host, including influx of anti-bacterial neutrophils and epithelial exfoliation at the infected site. UPEC has been known to replicate inside bladder cells in biofilm-like communities within bulging globular structures called “pods”. The formation of intracellular bacterial pods has been shown to protect UPEC from killing mediated by host neutrophils (Justice et al., 2004). In vitro tissue culture models of infection are often too simplistic; typically, they do not capture the diversity of host-pathogen interactions and are unable to recreate the heterogeneous physical niches that pathogens typically encounter. In this project, we have established differentiated mouse organoid cultures for studying the host-pathogen dynamics of UPEC pathogenesis. In the bladder organoid model system, we use microinjection to infect the orgaoids with UPEC in order to study UPEC growth and responses to antibiotic treatment using time-lapse fluorescence microscopy.

Bladder organoids | © EPFL UPKIN Kunal Sharma Bladder organoids | © EPFL UPKIN Kunal Sharma
Figure 1: Confirmation that mouse bladder organoids express biomarkers of mouse uroepithelium: CK13 (left, red) and CK7 (right, red). Cellular DNA was stained with DAPI (cyan). Scale bar is 20 μm.

 

Bacterial growth in the lumen of a bladder organoid | © EPFL UPKIN Kunal Sharma Bacterial growth in the lumen of a bladder organoid | © EPFL UPKIN Kunal Sharma Bacterial growth in the lumen of a bladder organoid | © EPFL UPKIN Kunal Sharma
Figure 2: Bacterial growth in the lumen of a bladder organoid. Mouse bladder epithelial cells express tdTomato (red). UPEC cells express YFP (yellow). (A-B) Bacterial growth between the initial stage of infection (A) and after 3h45 (B). (C) At 4h00 ampicillin was added to the culture medium and the infected organoid was imaged after 3 hours of ampicillin exposure.

 

Planned research:

The next stage of the project will involve doing time-lapse microscopy experiments with bladder organoids infected with wild-type and mutant strains of UPEC followed by immunofluorescence staining and scanning electron microscopy of the infected organoids.

Learning skills acquired during this project:

  • Working with organoid cultures, immune cells, and pathogenic bacteria (Biosafety Level 2)
  • Performing time-lapse experiments on infected organoids with a confocal microscope
  • Immunofluorescence staining of infected organoids
  • Scanning electron microscopy of infected organoids

Prerequisites:

  • Mandatory: Strong motivation for learning new things and troubleshooting

The project could be tailored to the interests of the student. Anyone interested in pursuing this project should contact Kunal Sharma ([email protected]) and Prof. John McKinney ([email protected]).

References

A. Mireles et. al , Nature Reviews Microbiology, 13, 269-284, (2015).

S. Justice et. al , PNAS, 101 (5), 1333-1338, (2004).

Bachelor Project / Master Lab Immersion Project / Master Semester Project / Master Thesis Project

Description:

Academic and pharmaceutical companies have invested substantial resources in developing predictive models for human physiology and disease states. However, conventional 2D cell culture models in tissue culture flasks are simplistic and do not recapitulate tissue-level architecture and organ microenvironments. More complex in-vitro systems such as trans-well inserts and organoids are more complex. but they also lack tissue-tissue interfaces, immune cells, and microscale environmental cues that are essential for the normal functioning of organ systems. These limitations can be overcome with organ-on-a-chip devices, which are microfluidic cell culture systems that mimic some aspects of tissue-level and organ-level physiology. Organ-on-a-chip technology has been developed for several organs, including: lung, gut, liver, kidney, and blood-brain barrier. We have developed a bioinspired bladder-on-a-chip (BoC) system using a commercially available organ-on-a-chip microfluidic platform. We have established a 3D co-culture of human bladder epithelial cells and human bladder endothelial cells with urine and nutrition perfusion that can be maintained for long-term differentiation and infection experiments.

Organ-on-a-chip device | © EPFL UPKIN Kunal Sharma

Figure 1. Organ-on-a-chip device.

Co-culture of bladder epithelial cells and bladder endothelial cells | © EPFL Kunal SharmaBladder organoids | © EPFL UPKIN Kunal Sharma

Figure 2. Co-culture of bladder epithelial cells and bladder endothelial cells on a trans-well insert. The cells become stratified and achieve uniform stratification on exposure to calcium on the basolateral side and urine on the apical side.

We are using BoC system to study different stages of UPEC pathogenesis (bacterial attachment, bacterial multiplication, bacterial shedding, IBC formation, epithelial cell exfoliation, etc.) and to study UPEC persistence after exposure to clinically relevant antibiotics. The BoC system can also mimic mechanical cycles of bladder filling and voiding with corresponding changes in shear stress and stretching experienced by the cells.

Different forms of uropathogenic Escherichia coli (UPEC) | © EPFL UPKIN Kunal Sharma

Figure 3. Different forms of uropathogenic Escherichia coli (UPEC) observed within urinary tract infection of mouse uroepithelium. The schematic also shows the different layers of uroepithelium: umbrella cells (orange), intermediate cells (light green), and basal cells (brown).

Planned research:

The next stage of the project will involve fabrication of the BoC devices with PDMS and PETE membranes. The devices will be used for differentiation of human bladder epithelial cells with the conditions established in Figure 2 and characterized by immunofluorescence staining and scanning electron microscopy. The validated BoC system will be used for studying UPEC pathogenesis (Figure 3) and UPEC dormant reservoirs within deeper layer of epithelial cells with time-lapse confocal microscopy. We will also use different UPEC mutants and reporters to better understand the persistence and underlaying heterogeneity between host-pathogen interactions.

Learning skills acquired during this project:

  • Working with mammalian cell culture and pathogenic bacteria (Biosafety Level 2)
  • Microfabrication of organ-on-a-chip devices
  • Time-lapse confocal microscopy
  • Immunofluorescence staining
  • Scanning electron microscopy

Prerequisites:

  • Mandatory: Strong motivation for learning new things and troubleshooting

The project could be tailored to the interests of the student. Anyone interested in pursuing this project should contact Kunal Sharma ([email protected]) and Prof. John McKinney ([email protected]).