Effect of Chlorine on the development of marine
biofilms dominated by diatoms
Under the Guidance
of
Dr.Jagadish.s.Patil
Marine Corrosion & Materials Research Division
National institute of oceanography
Goa, India
Jagadeesan v
08MBT03
Synopsis
• Introduction
• Materials and methods
• Results
• Summary
• References
Biofouling- An Intro
•
Biofouling is serious problem of rapid
colonization of aquatic species on immersed
surfaces in an aquatic environment.
•
It is a serious problem in ship hulls, power plant
cooling systems, aquaculture systems, fishing
nets, pipelines, submerged structures and also
oceanographic research instrumentation.
•
Many of the technologies implemented to solve
the current problem and he results are under
the investigation., and remain unsolved.
•
In which chlorination is one of the potential
method of controlling the biofouling both by
industrial and environmental applications.
•
Chlorine is biocide and it is a disinfectant used
to reduce the prevalence of pathogens in
drinking water by municipal water supply.
Aims & objectives
• To prevent biofouling using chlorine as an antifouling agent.
• To evaluate the efficiency of chlorine against mono algal and multi
algal species.
• To find the effectiveness of chlorine against the chlorophyll auto
fluorescence, Biofilm EPS and etc.,
• The evaluation of pulse chlorination on the development of
biofouling.
Diatoms are early colonizers
• Diatoms are the earliest eukaryotic colonizers of submerged surfaces and
one of the most conspicuous organisms in biofilms. The diatoms attach to
the substratum by secretion of EPS (Eg. Wetherbee et al. 1998), and
contribute significantly to the biofilm (Patil and Anil 2005).
• In the present investigation the important biofouling diatoms Amphora and
Navicula (Wetherbee 2008) were evaluated their biofouling potential against
the chlorination along the mixed natural species.
• They were subjected to grow under laboratory with the help of cover glass
substratum, and they start to colonize the substrate at a immediate effect
and they are one of the initial colonizers in the complex mechanism of
biofouling .
Images of mono & multi species biofilm
Natural biofilm
Amphora biofilm
Navicula biofilm
Materials and Methods
• Effect of chlorination on the growth of biofilm diatoms
• Effect of chlorine on chlorophyll fluorescence
• Effect of chlorine on exo polysaccharide (EPS)
• Effect of chlorine on cell structure of fouling diatoms
• Pulse chlorination
• Effect of pulse chlorination on chlorophyll, biofilm development
• Effect of chlorination in the optical transmission
Results
Cell counting Vs chlorination
EPS ASSAY- Alcian Blue
method
Effect of chlorine on the Chlorophyll- Epifluorecscence
analysis
Chlorophyll fluorescence - Navicula
Effect of chlorine on the chlorophyll
•
Effect of chlorine on the chlorophyll is
once again reflected here in the
chlorophyll estimation.
•
The biofilm cover glass are sampled and
chlorophyll analysis was achieved by
acetone 90% extraction method
•
The experiments were done by both
control and with the treatment.
•
The control group is a biofilm without the
treatment of chlorine to compare the
reduction obtained by the chlorine treated
biofilm samples
Live Amphora control
Amphora chlorine treated 0.5%
0.5% 1 min exposure
0.5% 5 min exposure
Cl 1%
1 % 1 min exposure
1 % 5 min exposure
Navicula control
Cl 0.5%
0.5% 1 min exposure
0.5% 5 min exposure
Cl 1%
1 % 1 min exposure
1 % 5 min exposure
Cell counting Vs chlorination Vs algal seedling
Species richness in Natural biofilm Vs chlorination
Pulse chlorination
• Pulse chlorination is a technique that is used to chlorinate the
biofouling candidates at a short interval of time to remove them and
effectively prevent them colonizing.
• In this study chlorine treatment was given in pulse (i.e. at every 6
hours) in order to derive optimal anti fouling results with minimal
chlorine amounts.
• The intermittent chlorination results in the continuous removal
biofouling candidates in a short successive chlorination ensuring the
removal adhered algal species on the surfaces.
Effect of pulse chlorination Biofilm biomass & chlorophyll
Effect of pulse chlorination in optical transmission
• Optical transmission is an
important property in the marine
biosensors.
• The fixed marine biosensors is
often fouled by the activity of
algal species, which is a serious
problem in monitoring the marine
environment.
• The pulse chlorination was
achieved with the clean glass
slides for the transmission
analysis, the transmission was
estimated using the image pro
software in microscope.
Summary
• Chlorination is one of the important antifouling strategy and it is
implemented easily on industrial and environmental application.
• The application of chlorination leads to the death of the algal cells
reflected in the scanning electron microscope analysis.
• which is subsequently confirmed by the cell counting experiment.
• The chlorophyll reduction and Eps analysis in electron microscopy
analysis reveals that it is a most promising agent.
• The pulse chlorination is further earning credit by application of on
the local biosensors applications, to reduce their impact on the
chlorine.
References
•
Abarzua S, Jakubowski S, (1995) Biotechnological investigation for the prevention of biofouling. I. Biological
and biochemical principles for the prevention of biofouling. Marine Ecology Progress Series, 123: 301-312
•
Alliance for Coastal Technologies (ACT) (2003) Biofouling Prevention Technologies for Coastal
Sensors/Sensor Platforms Solomons, Maryland, Nov. 2003, ACT No. 03-05/UMCES Technical Report Series
TS-426-04-CML, http://www.actonline.ws/Technologies.html
•
Alzieu C, (1998) Tributyltin: case study of a chronic contaminant in the coastal environment. Ocean and
Coastal Management, 40: 23–36
•
Baier RE, Meyer AE, DePalma VA, King RW, Fornalik MS (1983) Surface microfouling during the induction
period. Journal of Heat Transfer-Transactions of the Asme, 105, 618-624
•
Bakus GJ, Targett NM, Schulte B (1986) Chemical ecology of marine organisms: an overview. Journal of
Chemical Ecology, 12: 951-987
•
Characklis WG (1981) Microbial fouling: A process analysis. In E.F.C. Somerscales and JG Knudsen (eds.),
Fouling of heat transfer equipment, Hemisphere Publ. Co., Washington, D.C. p251-291
•
Chamberlian AHL (1976). Algal settlement and secretion of adhesive material. In: Sharpley JM, Kaplan AM
(eds.) Proc 3rd Int Biodegradation Symp. Applied Science, London, pp 417-432
•
Champ MA (2003) The new IMO treaty to ban TBT and other harmful antifouling systems for ships. Marine
Pollution Bulletin. Vol. 45.
•
Cooksey KE, Wigglesworth-Cooksey B. 1995. Adhesion of bacteria and diatoms to surfaces in the sea: a
review. Aquatic Microbial Ecology 9: 87–96
•
Evans SM (1999) Tributyltin pollution: the catastrophe that never happened. Marine Pollution Bulletin, 38:
629–636.
•
Granhag LM, Finlay JA, Jonsson PR, Callow JA, Callow ME (2004) Roughness-dependent removal of settled
spores of the green alga Ulva (syn Enteromorpha) exposed to hydrodynamic forces from a water jet.
Biofouling 20, 117–122
•
Guillard RRL, Ryther JH (1962) Studies of marine planktonic diatoms. I. Cyclotella nana Hustedt, and
Detonula confervaceae (Cleve Gran). Cannadian Journal of Microbiology 8: 229-239.
•
Hoagland KD, Rosowski JR, Gretz MR, Roemer SC. 1993. Diatom extracellular polymeric substances:
function, fine structure,chemistry and physiology. Journal of Phycology 29: 537-566.
•
Little BJ (1984) Succession in microfouling. In: Costlow JD, Tipper RC (Eds). Marine bideterioration: An
interdisciplinary study. Naval Institute Press, Annapolis, Maryland p63-67
•
Passow U (1995) A dye-binding assay for the spectrophotometric measurement of transparent exopolymer
particles (TEP). Limnology and Oceanography 40(7), 1995, 1326-1335.
•
Patil JS, Anil AC (2005a) Biofilm diatom community structure: Influence of temporal and substratum variability.
Biofouling, 21(3/4), 189–206
•
Patil JS, Anil AC (2005b) Quantification of diatoms in biofilm: standardization of methods. Biofouling, 21(3/4),
181–188
•
Patil JS, Kimoto H, Kimoto T, Saino T (2007) Ultraviolet radiation (UV-C): a potential tool for the control of
biofouling on marine optical instruments. Biofouling , 23(4): 215-230
•
Phe MH, Dossot M, Block JC (2004) Chlorination effect on the fluorescence of nucleic acid staining dyes.
Water Research 38: 3729-3737.
•
Poornima, E. H.; Rajadurai, M.; Rao, T. S.; Anupkumar, B.; Rajamohan, R.; Narasimhan, S. V.; Rao, V. N. R.;.
Venugopalan, V. P. Impact of thermal discharge from a tropical coastal power plant on phytoplankton. Journal
of Thermal Biology, 30, 307-316.
•
Phe MH, Dossot M, Guilloteau H, Block JC (2005) Nucleic acid fluorochromes and flow cytometry prove
useful in assessing the effect of chlorination on drinking water bacteria. Water Research 39: 3618-3628.
Railkin AI (2004). Marine Biofouling: Colonization Processes and Defenses, CRC Press, Boca Raton, Fl, USA
(2004) 303 pp.
•
 Ramus J. (1977). Alcian blue: a quantitative aqueous assay for algal acid and sulfated
polysaccharides. Journal of Phycology 13:345-8.
•
Saby S, Sibille I, Mathieu L, Paquin JL, Block JC (1997) Influence of water chlorination on the counting of
bacteria with DAPI (4¢,6-diamidino-2-phenylindole). Applied and Environmental Microbiology, 63: 1564-1569.
•
Vishwa kiran Y, Anil AC (1999) Record of imposex in Cronia konkanensis (Gastropoda, Muricidae) from Indian
waters. Marine Environmental Research 48:123-130
•
Vishwakiran Y, Anil AC, Venkat K, Sawant SS (2005) Gyrineum natator: A potential indicator of imposex along
the Indian coast. Chemosphere, 62: 1718-25.
•
Von Oertzen JA, Scharf EM, Arndt EA, Sandrock S, Dettmann L, Holzapfel H, Ringstorf H, Kohn H, Günther
R (1989) Spezialstudie ‘Alternative Antifouling Systeme’. Fachbereich Biologie, Universität Rostock
Wetherbee R, Lind JL, Burke J (1998) The first kiss: Establishment and control of initial adhesion by raphid
diatoms. Journal of Phycology 34: 9-15
•