MSc in Biochemistry
Dissertation Project – 2nd Cycle
Student´s Name:
Student email address:
Supervisor(s): Luisa Jordão / Aida Duarte
Supervisor(s) email address: [email protected] / [email protected]
Lab/Institution: and Instituto Nacional de Saúde Dr Ricardo Jorge (INSA), Departamento de Saúde
Ambiental / Departamento de Microbiologia e Imunologia, Instituto de Investigação do
Medicamento, Faculdade de Farmácia, Universidade de Lisboa
TITLE: Deciphering K. pneumoniae biofilms.
BACKGROUND
The role played by biofilms in bacterial pathogenesis has been highlighted in recent years. Biofilms’
relevance started with their implication in healthcare-associated infections (HAI) which are reported as
a major public health problem. Klebsiella pneumoniae is a worldwide leading cause of hospitalacquired urinary tract infections, as well as pneumonia, being responsible for many cases of pyogenic
liver abscess or endophthalmitis contracted in community patients.
In a previous study, we showed that Kp strains independently of their antibiotic (Ab) resistance profile
were able to assemble biofilms on abiotic surfaces. The kinetic of biofilm assembly differed between
strains and was related to virulence factors. Biofilm architecture was assessed by scanning electron
microscopy using backscattered electrons mode (SEM-BSED). The data analysis showed that biofilm
structure and composition was distinct and independent of kinetic assembly and Ab resistance profile.
Curiously, the bacteria with different Ab susceptibilities assembled structurally similar biofilms
following different kinetics. Another important fact arising from this study was that Kp lacking capsule,
regarded as less virulent, have a better performance as biofilm assembler and exhibited the highest
increase in Ab resistance when organized within biofilms (1). Moreover, full-sequencing and
annotation of genomes from the 3 Kp strains was performed in order to explain the differences found
in biofilm assembly. Preliminary data already revealed that the Kp strain displaying enhanced biofilmforming ability is genetically different from the others
(1) Bandeira M, Carvalho PA, Duarte A, Jordao L. (2014) Exploring Dangerous Connections between Klebsiella
pneumoniae Biofilms and Healthcare-Associated Infections. Pathogens. 2014 Aug 19;3(3):720-31. doi:
10.3390/pathogens3030720
OBJECTIVES
The main goals are:
1) Evaluate the ability of K. pneumoniae to assemble biofilms on abiotic and biotic like surfaces.
2) Characterization of extracellular matrix present in both biofilms.
3) Analyse the transcriptome of K. pneumoniae in biofilms with different ages and planktonic
1] Matos Ana, Gonçalves L.M., Rijo P., Vaz M.A., Almeida A.J., Bettencourt A.F. (2014). Minocycline-loaded acrylic
form in different growth stages.
bone cement – in vitro release, biochemical and biological assessment. J. Mater. Science and Eng: C. 38: 218–226.
4) Identify key effectors responsible for biofilm assembly (adhesion, maturation and dispersion).
In this work will be used only one K. pneumoniae strain which have shown to be the best biofilm
assembler.
The student will have the opportunity to acquire a wide range of expertise in bacterial culture, biofilm
assays, RNA technology (isolation, purification and sequencing), and sample preparation for electron 1
microscopy.
MSc in Biochemistry
Dissertation Project – 2nd Cycle
PROJECT DESCRIPTION
Working plan:
1) Literature review.
2) Biofilm assembly on abiotic and biotic like surfaces.
3) Characterization of extracellular matrix of biofilms assembled on abiotic and biotic like
surfaces.
4) RNA extraction and preparation for sequencing from planktonic and biofilms.
5) Analysis of RNA sequencing results.
6) Biofilm structure (Scanning electron microscopy).
7) Thesis writing
TIMELINE
Month 1
Month 2
Month 3
Month 4
Month 5
Month 6
Month 7
Month 8
Month 9
Month 10
Task 1
Task 2
Task 3
Task 4
Task 5
Task6
Task 7
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