Do Bacteria Behave?
- Jaikishan Advani (Sophia College, Mumbai and HBCSE
- Aug 30, 2015
- 5 min read
Behavioral modification through associative conditioning in putative-giant bacteria
Author: Jaikishan Advani (Msc Microbiology, Sophia College,Mumbai and Homi Bhabha Centre for Science Education, TIFR,Mumbai,India)
Co-authors: Wasifa Qadiri, Sidra Ashraf (Bsc, Life sciences, Sophia College,Mumbai), Imtiaz Gulami (Hbcse,TIFR), Priyanka Padmaram (Msc Life sciences, University of Mumbai), Pooja Singh, Pradnya Bhamre, Snehal Sangolkar, Rutuja Dhuri (Msc Microbiology, Somaiya College, Mumbai), Dr.Anjumanara Surti (HOD, Microbiology Dept, Sophia College), Dr.MC Arunan (Visiting Scientist, HBCSE,TIFR)
Abstract:
Bacteria are conventionally considered as a simple, hard wired organisms which hardly modify their behavior as behavior is mainly considered to be because of neurons. But there is no reason why behavior modification cannot be without neurons, as neurons work due to network of genes and biochemicals. Recent evidences show that bacteria is not as simple and can undergo various behavioral modification and even learning, where bacterial learning is a phenomenon considered across-generations. In current study, we attempt to check the directionality of putative giant bacteria towards and away from varied range of attractants and repellants, respectively and hypothesize, to focus on associative conditioning the putative-giant bacteria by giving an attractant(as reward) along with the repellent (punishment) and expect that the conditioned once will become aversive towards the attractant to which they were originally attracted. It also is an attempt to check whether this modification in behavior of bacteria is due to adaptation or learning, which can be determined by checking whether the behavioral modification is reversible.
Rationale:
The putative giant bacteria are the organisms (current focus on two types from several) which are obtained from concrete soil collected from the depression above the manhole cover lid (used for lifting it up) which are present on most of the road pavements, a “model” which is also amusingly termed as “Pagalapos” (an inverse ideogram of Darwin`s Galapagos islands, in which the small water filled cavities/ecosystems are present on widespread mass of land). Two types of organisms are obtained from Pagalapos in Sophia College campus, namely long 50-100 microns filamentous rods and 50-100 microns(about 100 times larger than an average bacteria) compact coils/ladder shaped, which have morphology similar to bacteria and are gram negative, both also possess cell wall, thus termed as putative-giant bacteria. Motility of both these organism is gliding and as they are huge in size,it can be easily used to determine whether they have directional or random motility, and whether this behavior can be altered by conditioning.
Significance and Purpose:
Behavioral modification or Learning in bacteria is a hardly considered, as bacteria are believed to have simple lifestyle, with hardly any capabilities of modification. Thus, the current study is an attempt to bring paradigm shift in our understanding about bacterial behavior.
It also is an attempt to identify and characterize the rare putative-giant bacteria which have been obtained as till date only 10-12 giant bacteria are known, among which these organisms do not resemble any.
Methodology:
Collection of sample:
Dry concrete soil sample is collected from over the manhole cover lid in sealed plastic pouch from near Bhabha hall, in sophia college campus, in month of March(Summer), and preserved.
Culturing of Filamentous rods and Compact coils/Ladder shaped (Putative Giant Bacteria):
50-100 mg of soil taken in eppendorf and level maked upto 1ml by dechlorinated water, and add 1 drop of 1:4 diluted milk.
Determination of directionality in motility of putative giant bacteria:
Slide-agar method will be used which has two cavities opposite to each other, connected by a channel, in one cavity putative giant bacteria will be loaded and in opposite attractant or repellent will be loaded, the diffusion of these chemicals will occur so as to form gradient, this gradient will be sensed by organisms and the chemotaxis towards or away from these chemicals will be checked microscopically, the end point will be the time at which, approx 70%-80% of population crossing half of the channel towards attractant, as compared to control in which their will be equal distribution of approx 50% on both the sides.
Similarly, in repellent assay, the endpoint will be the time at which approx 70%-80% of organisms are towards the repellant well, as compared to control in which their will be equal distribution of approx 50% on both the sides.
The chemicals which will be used as attractants or repellants are:
Sugars: Milk(crude) or Lactose, Maltose,Mannose, Ribose, Glucose, Galactose (range: ng/ml to mg/ml)
Organic compounds: Acetate,Succinate, Propionate,Benzoate, Ethanol, Isopropanol, Glycerol, Phenol, Trimethylamine (range: micro moles to millimoles)
Inorganic compounds: Nickel, Cobalt (pg/ml to mg/ml)
Ph: Acid, Alkali (range: 0.01 N to 0.1 N)
Others: Indole, Mercaptans (range: micromoles to millimoles)
Associative conditioning to modify the motility/chemotactic behavior:
The suitable concentrations of attractant and repellents selected will be used in 1:1 proportion to condition the organisms to averse from the previous attractant, and will be tested after appropriate rounds of conditioning(at least 10), with normal attractant alone, to check whether the normal chemotactic behavior towards attractant is altered.
Tentative Results:
Following are the images attached of;
a) Wet mount of filamentous rods and ladder/compact coils in 40X objective lens.
b) Gram negative nature of filamentous rods and ladder/compact coils in 40X objective, compared with positive control Yeast and negative control E.coli(100X objective).
c) Haemocytometeric measurements showing both types are covering 2 RBC squares(each square of 50 microns), hence they are of nearly 100 microns(100 times larger than an average bacteria).
d) Cell wall of filamentous rods and compact coils.
Interpretation of Results:
The initial studies on filamentous rods and compact coils suggest that they are giant around 50-100 microns and most likely bacteria as they have cell wall, though the pure culture of them is being attempted and not being achieved so far. This also suggests that these organisms could be novel giant bacteria as in current literature no mention of these morphologies have been found (Caveat References: 2007, chapter of H.N Schulz on Giant Bacteria and 2001 review on Big Bacteria by H.N Schulz, Bergey's` Manual of Systematic Bacteriology, Introductory Version and Gram Negative: Proteobacteria,Volume-2 and Protist Classification).
References:
1) Predictive Behavior Within Microbial Genetic Networks
Ilias Tagkopoulos1,2, *Yir-Chung Liu2,3, *,Saeed Tavazoie2,3
1 Department of Electrical Engineering, Princeton University, Princeton, NJ 08544, USA.
2 Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, NJ 08544, USA.
3 Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.
http://www.sciencemag.org/content/320/5881/1313
2) Molecular circuits for associative learning in single-celled organisms
Chrisantha T. Fernando,1,2,* Anthony M.L. Liekens,3 Lewis E.H. Bingle,1 Christian Beck,4 Thorsten Lenser,4 Dov J. Stekel,1 and Jonathan E. Rowe5
1Systems Biology Centre, University of Birmingham, Birmingham B15 2TT, UK
2MRC National Institute for Medical Research, Mill Hill, London NW7 1AA, UK
3TU/e Techniche Universiteit Eindhoven, 5600 MB Eindhoven, The Netherlands
4Bio Systems Analysis Group, Friedrich Schiller University Jena, Jena 07743, Germany
5School of Computer Science, University of Birmingham, Birmingham B15 2TT, UK
*Author and address for correspondence: Mathematical Biology, National Institute for Medical Research, Mill Hill, London NW7 1AA, UK.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2582189/
3) Bacteria 'can learn', Michael Hopkin, Published online 8 May 2008 | Nature | doi:10.1038/news.2007.360
http://www.nature.com/news/2008/080508/full/news.2007.360.html
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Esther R Angert, Cornell University, Ithaca, New York, USA, Ferran Garcia‐Pichel, Arizona State University, Tempe, Arizona, USA , Published online: September 2007
http://www.els.net/WileyCDA/ElsArticle/refId-a0020371.html
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Max-Planck-Institute for Marine Microbiology, Celsiusstrasse 1, D-28359 Bremen, Germany.Annual Review of Microbiology (Impact Factor: 13.02). 02/2001; 55(1):105-37. DOI: 10.1146/annurev.micro.55.1.105. Source: PubMed
http://www.researchgate.net/publication/11802808_Big_Bacteria
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http://www.researchgate.net/publication/230236347_Giant_Bacteria
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8) Kingdom Protista http://www.nicholls.edu/biol-ds/Biol156/Lectures/Protista.pdf
9) Protist, Wikipedia, https://en.wikipedia.org/wiki/Protist
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