My presentation in XVI Maharashtra Chapter Conference of IAMM at Krishna Institute of Medical Sciences, Karad on 26th September 2010.

1. Dr. Rajesh Karyakarte MD
Professor & Head,
Department of Microbiology,
Government Medical college, Akola

2.  Molecular Biology is the application of science
and technology to living organisms, as well as
parts, products and models thereof, to alter
living or non-living materials for the production
of knowledge, goods, and services (1).
 Pathogenicity is the capacity of a microbe to
cause damage, in a host.
 Pathogenesis is the process of producing this
damage.
(1)http://www.oecd.org/document/42/0,3343,en_2649_34537_1933994_1_1_1_1,00.html

3.  Falkow (1988) proposed “Molecular Koch’s
Postulates,” a set of conceptual tools for
dissection of bacterial pathogenesis based on
the identification of the genes responsible for
causing disease (2).
 Many genes required for virulence in bacteria are
in discrete DNA segments, e.g., pathogenicity
islands. Acquisition of these genes is sufficient
for a bacterium to become virulent.
(2) Falkow S. Molecular Koch's postulates applied to microbial pathogenicity. Rev Infect Dis
1988; 10(suppl 2): S274-6.

4.  Molecular studies show that the chemistry of
the microbial surface is the major distinction
between pathogenic and nonpathogenic
microorganisms.
 Tropism of microorganisms is subdivided
into:
 Tissue and host tropism
 Species-specific tropism

5.  Uropathogenic E. coli adhere to human
urinary tissue cells by PapG adhesin, which is
a component of Pap (pyelonephritis-
associated pili).
 There are three PapG alleles (Class I, II, and
III). Class II PapG is associated with
pyelonephritis and Class III with cystitis.

6.  PapG binds Gal-α-1-4-Gal containing glycolipid
present on the surface of renal epithelial cells.
 The glycolipid has a digalactoside core linked
via a -glucose residue to a ceramide group that
anchors the receptor to cell membrane. This
receptor is named globotriasylceramide (GbO3).
 GbO3 with one additional sugar moiety (GalNAc)
becomes GbO4 (Globoside) and
 GbO3 with two additional sugar moieties
(GalNAc 2) becomes GbO5 (Forssman
antigen).(3)
(3) Dodson KW, Pinkner JS, etal. Structural basis of the interaction of the pylonephritic E. coli adhesin
to its human kidney receptor. Cell 2001; 105: 733-43.

7.  Neisseria gonorrhoeae, N. meningitidis, E. coli
(expressing CFA-I and CFA-II) and group A
Streptococci are limited to human infection
due to species-specific tropism.
 A single strain of E. coli may be able to
express several distinct adhesins encoded by
respective distinct regions of chromosomes
or of plasmids, which helps in adapting to
changing environment.

8. Organism Adhesin on the microbe Receptor on host cell Adaptation
E. coli Type I fimbriae (FimH) D-mannose residue Allelic variation in fimH gene
helps in differential binding
Neisseria Fimbriae with methylated Surface glycoprotein --
gonorrhoeae phenylalanine amino CD46 on urogenital
terminus residue cells

9.  Molecular biology has helped in elucidation of
the molecular basis of biofilm formation.
 Staphylococcus epidermidis forms biofilms
and is a major cause of medical-device
related infections.
 The intercellular adhesion in these biofilms is
provided by polysaccharide intercellular
adhesin (PIA).

10.  PIA is a linear glucosaminoglycan composed
of N-acetylglucosamine in -1,6-glycosidic
linkages containing deacetylated amino
groups and succinate and phosphate
substitution.

11.  Synthesis of PIA requires expression of icaADBC
operon.
 These genes are controlled with the help of
alternative sigma factor B, icaR, two more
regulatory loci, plus phase variation due to IS
element insertion.
 PIA production can be stopped by inactivational
IS element insertion that allows individual S.
epidermidis cells to leave the biofilm to colonize
new surfaces.(4)
(4) Dobinsky S, Kiel K, Rohde H, Bartscht K, Knobloch JK, Horstkotte MA, Mack D. Glucose-
related dissociation between icaADBC transcription and biofilm expression by Staphylococcus
epidermidis: Evidence for an additional factor required for polysaccharide intercellular
adhesin synthesis. J Bacteriol 2003; 185: 2879-86.

12.  In Enterobacteriaceae, the waa locus enzymes
required for the sequential assembly of the
core oligosaccharide on to the lipid A
acceptor. This locus consists of three operons:
 The gmhD operon that directs inner core
biosynthesis
 The central waaQ operon that is responsible
for outer core; and

13.  The waaA operon that contains the structural
genes for a bifunctional KDO
transferase, which is required for the addition
of KDO to the inner core
 With a few exceptions, the enzymes involved
in O-polysaccharide assembly are encodes by
genes at the rfb locus and are expressed
constitutively.

14.  Translocation of protein synthesized in the cell occurs
across the inner and outer membranes in Gram-
negative bacteria by one of the five main pathways:
 Type I Mechanism: Utilized for secretion of the hemolytic
toxin HlyA from E. coli.
 Type II Mechanism: Is the general secretory pathway.
 Type III Mechanism: Responsible for injection of proteins
into eukaryotic cells
 Type IV Mechanism: Also responsible for injection of
proteins into eukaryotic cells, known as TFSS (Type four
secretion system)
 Type V Mechanism: Also called autotransportation

15.  Pathogenic Yersinia spp. utilize the YopH
proteins to cause dephosphorylation of
several macrophage proteins to prevent
phagocytosis.
 Transient tyrosine phosphorylation of
macrophage proteins is required for normal
phagocytosis.

16.  Salmonellae on the other hand possess two
type III secretion systems. These systems are
encoded by two distinct gene clusters termed
Salmonella pathogenicity island (SPI-1 and
SPI-2).
 These secretion systems have different roles
with SPI-1 involved in mucosal penetration
and SPI-2 with systemic spread.

17.  Pathogenic bacteria encounter countless
different environments and therefore need to
constantly monitor diverse physical and
chemical signals to tailor their responses with
the help of regulatory proteins.

18.  The 6.3-MB genome of P. aeruginosa, for
example, contains around 5500 genes, some
10% of which encode such regulatory
proteins. These regulatory proteins help
expression or repression of specific genes
depending on environmental cues.

19.  For example, if iron is plentiful in the
environment, synthesis of siderophores and
their associated export and import pathways
can be switched off.
 V. cholerae have an inverse correlation
between motility and virulence gene
expression as it switches between aquatic
and human host environment.

20.  In E. coli, the 17-kDa Fur repressor protein
controls transcription from iron-responsive
promoters in an iron-dependant manner, i.e.
to be active, Fur requires ferrous ion as a
corepressor.

21.  When iron is plentiful, the Fur-Fe2+ complex
interacts with an operator sequence called
Fur box and prevents gene expression.
 During iron scarcity, ferrous ion is removed
from the Fur-Fe2+ complex by the cell. Fur
alone cannot act as repressor for expression
of iron-repressed structural genes. The entire
iron absorption system becomes active.

22.  An enormous number of genes, including
many virulence determinants, regulate this
system.
 This system consists of two regulator
proteins, a cytoplasmic-membrane
associated sensor and a response regulator in
the bacterial cell.

23.  TCSTC sense a variety of different factors
including oxygen, hydrogen, quorum-sensing
signal molecules, Mg2+ and phosphate
limitation.

24.  It is crucial in many bacterial infections that
the bacterial population attains a particular
cell density to overcome host defense and
establish an infection.
 Pathogens have an ability to communicate
between themselves to orchestrate collective
attack against the host immune system.

25.  The term ‘quorum sensing’ is commonly used
in describing the phenomenon whereby
accumulation of a diffusible, low-molecular-
weight signal molecule (sometimes called
‘autoinducer’) enables individual bacterial
cell to sense when the minimal number, or
‘quorum’ of bacterial cell has been achieved
for a concerted response to be initiated.

26.  Bacteria employ a number of different quorum
sensing ‘languages’, and several families of
signal molecules have been characterized.
 At molecular level, quorum sensing requires a
synthase plus a signal transduction system for
producing and responding to the signal
molecule.
 Many bacterial species have quorum sensing
system consisting of the luxS and a furanone-
related molecule AI-2 (for autoinducer-2).

27.  P. aeruginosa has a multisystem quorum
sensing. It possesses two AHL (N-
acylhomoserine lactone) dependant quorum-
sensing systems.
 Apart from AHLs, P. aeruginosa has a third
quorum-sensing molecule, the pseudomonas
quinolone signal (PQS).
 Staphylococci produce quorum-sensing
molecules termed autoinducing peptides
(AIP).