Presented by

NISHANTH. S

2016601106II M.Sc.(Ag.) in Microbiology*

Department of Agricultural MicrobiologyTNAU CBE - 3

TAMIL NADU AGRICULTURAL UNIVERSITY

The Bacterium or a Bacteria

Bacterial biofilm

Extracellular matrix (ECM)

Bacterial cells

A cause for infection !!!

What are biofilms?

Biofilms are defined as highly structured communities of

microorganisms that are either surface associated or attached to one

another and are enclosed within a self-produced protective extracellular

matrix (ECM)

What does ECM comprises !!!

Polysaccharides

Proteins

Lipids

e- DNA

High cell densiy

10 8 to 10 11 cells g-1 wet weight

(Ramage et al. 2009)

Cont…

Planktonic bacteria vs biofilms???

1. The intercellular signaling system (QS) - regulationBhardwaj et al (2013)

2. Cyclic nucleotide second messengers – cyclic GMPHengge et al (2015)

3. bap protein – assembles matrix scaffold proteins & builds the

biofilm matrix Taglialegna et al (2016)

Cont…

(Fleming et al. 2016)

Why microbes form biofilms???

Prevention of biofilms

How to overcome biofilm effect???

(Zhao et al. 2017)

Phage therapy

Why Phages not antibiotics !!!

Lytic cycle – A key to phage therapy

Multiple defence strategies – an overview

Surface modification

(Kimberley D. Seed 2015)

Pre-infection strategy

Prevention of phage adsorption

1. Blocking of phage receptors

2. Production of extracellular matrix

3. Production of competitive inhibitors

Three categories(Labrie et al. 2010)

Pre-infection strategy

Prevention of phage adsorption

1. Blocking of phage receptors

2. Production of extracellular matrix

3. Production of competitive inhibitors

Three categories(Labrie et al. 2010)

Pre-infection strategy

Prevention of phage adsorption

1. Blocking of phage receptors

2. Production of extracellular matrix

3. Production of competitive inhibitors

Three categories

E. coli

FhuAAn iron transporter

Entry of coliphages (T5)

No Compete with Microcin J25 (an antibiotic)

(Labrie et al. 2010)

NUTRIENT RICH ENVIRONMENT

Pre-infection strategy

Prevention of phage adsorption

1. Blocking of phage receptors

2. Production of extracellular matrix

3. Production of competitive inhibitors

Three categories

E. coli

FhuAAn iron transporter

NUTRIENT LESS ENVIRONMENT

Entry of coliphages (T5)

Compete with Microcin J25

(Labrie et al. 2010)

Infection strategy

Prevention phage DNA entry

Sie (superinfection exclusion) – proteins that blocks the entry of phage DNA

Rapid inhibition of DNA injection into the cells

Imm – changes the conformation of the injection sites

Sp – inhibits the activity of T4 lysozyme (gp5)

(Labrie et al. 2010)

Cutting phage nucleic acid

Defence island system associated with restriction modification

DISARM allows phage adsorption but prevents phage

replication.

Composed of five genes1. DNA methylase

2. Helicase domain

3. Phospholipase D (PLD) domain

4. DUF1998 domain and

5. A gene of unknown function

R/M system (3 activities)

1. Restriction endonuclease activity

2. DNA modification activity

3. Target recognition activity

DISARM

(Ofir et al. 2017)

Cont…

(Labrie et al. 2010)

Cont…

(Labrie et al. 2010)

Cutting phage nucleic acid

Clustered Regularly Interspaced Short Palindromic Repeats

CRISPR Cas system

Lysogeny dependent inhibition of biofilm formation

(Labrie et al. 2010)

Abortive infection system

Rex system found in phage λ – lysogenic E. coli strains

(Labrie et al. 2010)

State of phage and host !!!

Phage lives Phage dies

Bact

eri

um

liv

esB

act

eri

um

die

s

Adsorption resistance

Blocking of phage receptors

Production of ECM

Production of competitive inhibitors

Superinfection exclusion

DISARM

CRISPR CAS

Absence of resistance

Abortive infection

Transcription blocks

Translation blocks

Replication blocks

Newly discovered defence systems

Prokaryotic argonaute

BREX system

(Swarts et al. 2017)

Key protein in RNA interference system

pAgos – NA guided host defence against

invading NAs

Complete phage resistance

Involves DNA methylation of the host cell

Methylation on non-palindromic TAGGAG

motifs (bacterial genome)

Case study

Engineering the Bacillus paralicheniformis 9945a DISARM system into Bacillus subtilis has

rendered the engineered bacteria protected against phages from all three major families of

tailed double-stranded DNA phages

phi3T

Three core genes - drmA, drmB, drmC

Methyltransferase – drmMI or drmMII

Additional gene - drmD or drmE

Conclusion

Department of Agricultural MicrobiologyTNAU CBE - 3

To understand the interactions between phages and their

bacterial hosts:

i. in order to fully exploit their antimicrobial potential and

ii. to effectively control their populations in bio-industries.

Future thrust

Department of Agricultural MicrobiologyTNAU CBE - 3

1. The efficacy of phage resistance mechanisms against

genetically diverse phages – need to be measured.

2. Multiple anti-phage barriers.

3. Systems targeting phages with ss DNA, ss RNA and ds RNA.