Electron microscopy at the National Institute of Cholera and Enteric
Diseases started in the early 80’s with the installation of a
Philips state-of-the-art transmission electron microscope, model
420T. One high vacuum evaporator (shadow caster), Polaron E6100, was
also installed along with the EM. Later on one LKB Nova
ultramicrotome, a LKB 7800 knifemaker , one De Vere enlarger, one
JEOL JEE-400 high vacuum evaporator along with a JEOL HDT400
hydrophilic treatment apparatus were also installed. At the
beginning of the new millennium the laboratory was upgraded to a
cryoEM laboratory. To this end, one FEI Tecnai 12 BioTwin
transmission electron microscope with a Gatan cryostage, one Leica
EM CPC universal cryo-workstation, one Leica Ultracut UCT
ultramicrotome with FC6 cryo attachment were installed in the
The electron microscope is used primarily for research and
occasionally for diagnosis. The techniques in routine use are
negative-staining analysis, Kleinschmidt’s protein monolayer
technique of DNA, partial denaturation mapping of DNA, heteroduplex
analysis of DNA, protein-free spreading methods of DNA and RNA,
immunoelectron microscopy, ferritin labellig, ultramicrotomy,
darkfield electron microscopy and electron diffraction, cryoelectron
microscopy, three-dimensional image reconstruction and tomography,
environmental scanning electron microscopy and atomic force
There are several projects going on in the laboratory.
Characterization of several choleraphages is being done by electron
microscopy. This laboratory, for the first time, showed the
filamentous nature of RS1-KmΦ phage of V. cholerae. The morphology
of different choleraphages and their DNA have been characterized.
This laboratory, for the first time, constructed partial
denaturation maps of DNA of vibriophages which have been used for
the determination of cohesive ends, circular permutation, terminal
redundancy, packaging pattern of DNA in the phage head. Also for the
first time, three-dimensional structure of vibriophages has been
determined using cryoelectron microscopy and single-particle
analysis methods. Also packaging of DNA inside these phages has been
determined using cryoelectron microscopy.
Also the structure of several hemagglutinins of Vibrio cholerae and
Shigella dysenteriae has been determined using negative staining
methods. Now the 3-D structures of these molecules are being worked
out using cryoelectron microscopy. Hydrodynamic properties of the
flagella of Vibrios have been studied as well.
Fimbriae or pili play a vital role in the attachment of bacteria to
the human intestinal cell wall. These have been studied extensively
in different serovars of Vibrio cholerae and Escherichia coli.
Mechanism of pathogenesis of V. cholerae and E. coli has also been
studied. Presence of a capsular layer in a new strain of Vibrio
cholerae has been confirmed with the help of ferritin labelling
Histopathological changes caused by different enteric pathogens have
been studied by light microscopy. Surface structural changes and
in-depth ulltrastructural changes are being studied using scanning
and transmission electron microscopes. Few of the important
enteropathogens studied so far are: Vibrio cholerae, Helicobacter
pylori, Shigella and Aeromonas hydrophila.
Some collaborative research was also carried out in the laboratory.
Amongst these, studies on chlorpromazine induced aggregation of
normal human hemoglobin and protein folding activity of ribosomal
RNA merit special mention.
Several national workshops on electron microscopy were also
organized by this division with great success.