Sediment and water samples were collected from mangrove and estuarine biotopes at fortnightly intervals. The physico-chemical characters of the overlying water were studied. In the mangrove biotope maximum temperature (31.5°C) and in the estuarine biotope maximum salinity (35.6‰) were recorded during the summer season, whereas in post-monsoon period the sulphate content was increased to 516 p.p.m. and the pH was reduced to 7.4. Invariably both in the enriched sediment and water samples four major peaks (at wavelengths 460, 705, 772 and 850 nm) and two minor peaks (at wavelengths 580 and 663 nm) of absorption spectra were noticed. A pure culture of Chromatium sp., isolated from mangroves sediment, showed three peaks of absorption spectra at wavelengths, 500, 580 and 850 nm. The effect of sodium chloride on the growth of Chromatium sp., was also studied and it was observed that maximum growth occurred in the range 1-3% sodium chloride concentration. This isolate was also capable of utilizing various sulphur and carbon compounds. Glycerol and glucose did not show any specific effect whereas pyruvate, malate and acetate increased the growth

Dhevendaran, K.

Aquatic Commons

hotosynthetic Bacteria in the Marine 
Environment at Porto~Novo 
K. DHEVENDARAN 
School of Marine Sciences, University of Cochin, Cochin - 682 016 
Sediment and water samples were coiiected from mangrove and estuarine biotopes 
at fortnightly intervals. The physico-chemical characters of the overlying water were 
studied. In the mangrove biotope maximum temperature (31.5°C) and in the estuarine 
biotope maximum salinity (35.6% 0 ) were recorded during the summer season, whereas 
in post-monsoon period the sulfate content was increased to 516 p.p.m. and the pH was 
reduced to 7.4. Invariably both in the enriched sediment and water samples four 
major peaks (at wavelengths 460, 705, 772 and 850 nm) and two minor peaks (at wave-
lengths 580 and 663 nm) of absorption spectra were noticed. A pure culture of Chroma-
tium sp., isolated from mangroves sediment, showed three peaks of absorption spectra 
at wavelengths, 500, 580 and 850 nm. The effect of sodium chloride on the growth of 
Chromatium sp., was also studied and it was observed that maximum growth occurred 
in the range 1%-3% sodium chloride concentration. This isolate was also capable of 
utilizing various sulfur and carbon compounds. Glycerol and glucose did not show 
any specific effect whereas pyruvate, malate and acetate increased the growth. 
Photosynthetic bacteria are widely dis-
tributed in all aquatic environments. Their 
ability to grow in anaerobic environment 
depends on the radiant energy and it was 
well documented in a review by Van Niel 
(1944). They have the leading role in the 
production of organic matter and in the 
purification of sewage in aquatic environ-
ment. These bacteria can be easily recognised 
by the formation of water-bloom and the 
bloom forming photosynthetic bacteria 
have been reported earlier and also they have 
been utilized as a feed by the copepods 
(Sorokin., 1966). Besides these, Cavari, 
et al. (1973) revealed a new phenomenon 
that the secondary peak of photosynthesis 
in the thermocline layer in the Lake Kinneret, 
(Israel) and attributed it mainly to photo-
synthetic sulfer bacteria Chlorobium phaeo-
bacteriodes. As primary producers these 
photosynthetic bacteria are likely to contri-
bute to the organic load and their contri-
bution may reach 9 to 25% of the total 
primary production (Czeczuga, 1965, 
Takahashi & Ichimura, 1968). It is under-
stood that the purple bacteria are particu-
larly depending upon the degree; of pollu-
tion of water, and in Japan and Germany 
the photosynthetic bacteria were utilized 
Vol. 21, 1984 
for the treatment of organic wastes (Koba-
yashi, 1975; Siefert et a!., 1978). 
The occurrence and the functional signi-
ficance of photosynthetic bacteria along the 
coastal waters of India have not been pre-
viously studied. Truper & Pfennig (1981) 
studied the characters and identified some 
of the anoxygenic photosynthetic bacteria. 
Following their techniques in the present 
study an attempt has been made to enrich 
and isolate the photosynthetic bacteria from 
Indian coastal waters and to characterize 
their distribution on the marine environment. 
Materials and Methods 
Fortnightly collections were made from 
February to July 1978 at two stations in 
two biotopes such as estuarine and man-
grove. Description of the stations have 
been given by Dhevendaran (1978). Sedi-
ment samples were collected using Peterson 
grab and were transferred aseptically to 
sterile polythene bags and the overlying 
water was collected with a Knudsen water 
sampler for estimation of physico-chemical 
parameters following the method adopted by 
127 
Dhevendaran (1978). One g of wet sedi-
ment was aseptically removed from 1 espe- · 
ctive samples and transferred into 100 ml 
of the enrichment medium. The medium 
contained per litre of 50% aged seawater, 
1.0 g of MgCo 3 , 0.6 g of K 2HP04 , 3.0 g 
of NH4cl, 1.0 g of yeast-extract, 2.58 g of 
sodium malate, 10 ml of trace elements and 
the medium was prepared as described by 
Pfenning & Lippert (1966). 1 ml of water 
from each sample was pipetted out into the 
enrichment medium with sterile 1 ml pipette. 
The medium with samples was taken in 
glass-stoppered bottles and incubated with 
continuous illumination at 20 [J. E m-2 S-1 at 
room temperature (28± 2o C) for 10 days 
unless otherwise stated, to study the 
absorption spectra of crude extracts in 
90% acetone following the methods of 
Takahashi & Ichimura (1968) and of Tru-
9er & Pfenning (1981). For the isolation 
of purple photosynthetic . bacteria, 
sediment sample was inoculated into the 
above mentioned enrichment medium and 
incubated as mentioned above. After 10 
days of incubation at room temperature 
with continuous illumination (20 E m-2 S-1) 
0. 1 ml of aliquot was taken and poured into 
the sterile petri dishes following the method 
of Siefert et a!. (1978) and mixed the above 
enrichment medium with 2% agar and 
incubated again under continous illumina-
tion for 10 days. Single pure colonies 
appeared and the colonies were identified 
according to the method of Pfenning & 
Truper (1974), Truper & Pfenning (1981). 
After the incubation period, the crude 
extracts of the sediment and water were 
filtered separately through membrane filter 
paper under vacuum. The cell residues 
were suspended separately in about 10 ml 
K. DHEVENDARAN 
of 90% acetone and incubated in dark at 
4°C, for 30 min following the method of 
Siefert, et a!. (1978) and the mixture was 
centrifuged at 10000 x g. Then the absor-
bance of the supernatant was measured in 
UNICAM Spectrophotometer at wavelength 
from 400-900 nm. The absorption spectra 
of the pure culture was also recorded. Beside 
this, the growth of the selected culture in 
different concentration of sodium chloride 
and also its growth in various sulfur and 
carbon compounds were measured by taking 
the optical density at 650 nm (Truper & 
Genovese, 1968). 
Results and Discussion 
Results of the physico-chemical para-
meters are summarized in Table 1. It is 
understood that the shallow mangrove bio-
tope recorded maximum temperature and 
the deep estuarine biotope (Fig. 1) highest 
salinity during summer and they were 31.5°C 
and 35.6 %o respectively, but the salinity was 
lowered during monsoon season and this 
has been attributed to the high influx of 
freshwater. The pH ranged from 7.4 to 
8. 3 and the lowering of pH in mangrove 
biotope could be due to the humic acid and 
phenolic acid content of the sediment (Kara-
nath et a!., 1975). The sulfate concentration 
in the mangrove biotope was higher (516 
p. p.m.) and this could be due to the presence 
of thick vegetation as well as the direct 
influence of seawater and freshwater inflow 
from the adjacent paddy fields and this in 
turn facilitate the proliferation of sulphate 
reducing bacteria, photosynthetic bacteria 
and plankters during summer and pre-mon-
soon seasons. These seasons are noted for 
higher productivity (Krishnamoorthy & 
Table 1. Physico-chemical parameters in the mangrove and the estuarine biotopes 
Overlying Mangrove Estuary 
water 
Feb. Mar. Apr. May June July Feb. Mar. Apr. May June July 
Temp., oc 28.0 29.5 31.5 31.5 30.0 27.5 29.0 29.0 31.0 31.0 30.0 29.0 
Salinity, %o 31.8 33.0 33.6 35.4 34.6 33.9 32.9 32.8 33.4 35.6 35.0 34.2 
pH 7.4 7.9 8.2 8.2 8.1 7.8 7.9 8.3 8.2 8.3 8.3 8.0 
Sulphate, 516.0 500.0 482.0 468.0 462.0 446.0 496.8 462.4 438.6 428.8 414.6 408.2 
p.p.m. 
FISHERY TECHNOLOGY 
STUDIES ON PHOTOSYNTHETIC MARINE BACTERIA 128 
N 
A 
~ Marine biological slalion 
~ Beds of mudflals 
IIJill Mangrove foresl's ex1ens1ons 
Fig. 1. The sampling stations 
Sundararaj, 1973). The absorption spe-
ctra of selective enrichment of sediments 
as well as the overlying water were taken 
separately in order to study the nature of 
photosynthetic bacterial contribution of 
various pigments towards the sediments 
and the water (Fig. 2). Four major peaks 
(at wavelengths 460, 705, 772 and 850 nm) 
0·8 
>-
:1;\ 0·6 
c: 
<lJ 
'0 
0 0·4 
() 
·.;;:: 
c. 
0 
0·2 
,•' 
.. ·· 
...... ·· 
.. 
.... 
/o.· .. 
I , 
t 
/ 
,.l 
o~-A~~~5±oo~~6~o~o~?~o~o~~s~o~o~~9o~o 
Wavelength (n m) 
Fig. 2. Absorption spectra of enrichment cultures 
inoculated with sediment and water sam-
ples collected from mangrove and estuarine 
biotopes 
Curve a 
Curve a1 
Curve b 
Curve b1 
Mangrove water 
Mangrove sediment 
Estuarine water 
Estuarine sediment 
Vol. 21, 1984 
and two minor peaks (at wavelengths 580 
and 663 nm) were noticed both in sediment 
and water. There is no marked variation 
in the absorption spectra but for the qu1;1n-
tity. The absorption maximum at 460 nm is 
due to the contribution by typical carotenoids 
and the peaks CJ.t 705 and 850 nm are mainly 
by bacteriochlorophylls d and a respectively, 
thus indicating the predominance of Thior-
. hodaceae in sediments and water and the 
peculiar peak at 772 nm is also by Thior-
hodaceae as suggested by Jimbo, (1938) 
and Truper & Genovese (1968). The pre-
sence of H 2 S in the reductive zone of sedi-
ment attracted the photosynthetic sulfur 
bacteria. Besides this, they are present at 
the boundary between the oxidative and 
reductive zone thus confirming the report 
of Kuznetsov (1958). It is understood that 
the features of absorption spectra reveal 
that the photosynthetic bacteria are verti-
cally distributed and they play an important 
role in the sulfur cycle. The suspension 
was pink, microscopically it showed Chro-
matium like bacteria of high motility in addi-
tion to a few non-motile Chlorobium-like 
forms (Truper & Genovese, 1968). 
One pure culture was isolated which is 
short, rod like, motile with sulfur globules 
inside the cell. The colour of the culture 
is pink and is Chromatium-like (Truper & 
Genovese, 1968). The absorption spectrum 
of the pure culture (Fig. 3) showed three 
peaks at 510, 580 and 850 nm. The Chroma-
than like bacterium contains typical caro-
tenoids (maximum absorption at 510 and 
l·ol --.--·---,.--~--.--·---,----·-,-----] 
0·8 
:>, 
·1;;o-6 
c 
(j) 
u 
o0-4 (.) 
·.;:: 
a. 
0 
0·2. 
0 
-4bo-·50o Goo --=?:::o~o::----=a-=o-=o--9::-0-=--:o::-:-::ro-=o-='o 
Wavelength (n m) 
Fig. 3. Absorption spectra of pure culture of 
Chromatium sp. 
129 
580 nm) and bacteriochlorophyll a (850 mn) 
as shown in the absorption spectra of Chro-
matium warmingii and Cromatium sp. strain 
3311 (Schlegel & Pfenning, 1961; Truper & 
Genovese, 1968). 
The result of the sodium chloride concent-
ration on the growth of the pure culture 
( Chromatium sp.) is given in Table 2. 
Table 2. Growth of the isolated strain at 
Chroma-
tium sp. 
+ -
different sodium chloride concen-
trations 
Nacl Nacl Nacl Nacl Nacl 
free 1% 2% 3% 5% 
++ +++ +++ +++ + 
Slight growth; ++= moderate 
growth; + + + = profuse growth 
Sodium chloride was added to the medium 
at the concentrations of 1, 2, 3 and 5 g/100 
ml of the medium. An inoculated control 
without sodium chloride for its growth was 
maintained to ascertain the importance of 
sodium chloride for its growth. The most 
distinct characteristic of the marine isolates, 
compared to the terrestrial counter parts, is a 
requirement of seawater. It is obvious that 
marine Chromatium gradually reduce their 
100% demand of salinities but they need at 
least 10% seawater, whereas terrestrial and 
freshwaterChromatium would not tolerate sea-
water (Genovese et a!., 1963). In the pre-
sent study, it was shown, that the maximum 
growth occurred from 1 to 3% sodium chlo-
ride concentration. However, in the absence 
of sodium chloride, growth still occurred to 
a acertain extent in this culture. Similar 
reports have already been made by Geno-
vese et a!. (1963) and Truper & Genovese 
(1968). Addition of sodium chloride to 
optimum levels substantially increased the 
growth rate and it may, therefore, be pre-
sumed that there is a demand for sodium 
chloride for its normal metabolic process. 
This indicates that this culture can be grou-
p;':d under m1rine Chromr:ttium. Further 
increase in the concentration of sodium 
chloride (5%) reduced the growth but it 
could tolerate that concentration. Similar 
halotolerant and halophilic strains of. Chlo-
ribium and Chromatium have been reported 
K. DHEVENDARAN 
(Genovese et al., 1963; Truper & Pfenning, 
1981). 
Thiele (1966), Truper & Genovese (1968) 
and Truper & Pfenning (1981) studied the 
ability of Thiorhodacease to use a variety of 
different carbon and sulfur compounds. 
Following their technique we have done 
parallel experiments with our culture. Table 3 
Table 3. Utilization of sulfur and carbon 
compounds added to the enrich-
ment medium 
Chromatium sp. 
Additions Percen- 3311 Chro-
tage (Refer- matium 
ence) sp. 
Sulfide 0.05 Control Control 
Thiosulphate 0.1 + + 
Elemental 
sulfur 0.1 + + 
Glycerol 0.1 + + 
Glucose 0.1 + + 
Fructose 0.1 + + 
Casamino acids 0.1 + + 
Py1uvate 0.1 + + 
Acetate 0.1 + + 
Lactate 0.05 + + 
Malate 0.1 + + 
Succinate 0.1 + + 
Citrate 0.1 + + 
Nitrate 0.1 + + 
- =no growth; = slight growth; 
++ = profuse growth 
gives the results compared with those of 
Truper & Genovese (1968) for Chromatium 
sp. Their strain Chromatium 3311 has the 
strongest heterotrophic tendency. The simi-
larity in the use of carbon and sulfur com-
pounds between our strain and their strain 
3311 is noticed. When Chromatium strain 
was supplied with fructose, pyruvate, ace-
tate, malate and succinate, growth was mar-
kedly stimulated, while glucose, glycerol, 
citrate and nitrate showed no variation in 
growth, thereby indicating that there is not 
much difference in the metabolic activity. 
Thus the metabolism of Chromatium seems 
to be similar to that of Chromatium 3311 
(Truper & Genovese, 1968) and that of 
Chromatium sp. (Truper & Pfenning, 1981). 
FISHERY TECHNOLOGY 
STUDIES ON PHOTOSYNTHETIC MARINE BACTERIA 130 
The occurrence of Chromatium sp. in the 
marine environment shows that these bacteria 
are widely distributed. It was isolated from 
the anoxic, H 2S containing layer. But its 
role in the biogeochemistry and the primary 
production of the seawater is yet to be fully 
understood. Sulfate is the second most 
abundant anion in the sea forming a large 
reserve of bound oxygen and sulfur (Red-
field, 1958). Beside, these photosynthetic 
sulfur bacteria, various other groups of sul-
fur oxidizing and sulfate reducing bacteria 
are known to take part in the sulfur cycle 
of marine environment. However, photo-
synthetic bacteria can also serve as food 
for zooplankton such as copepods as sugges-
ted by Sorok:in (1966) and Takahashi & 
Ichimura (1968) and the work on this line 
is m progress. 
Thanks are due to Prof. R. Natarajan, Director 
of Marine Biological Station, Porto Novo for con-
stant encouragement and critically going through 
the manuscript. The facilities provided by Annamalai 
University, the financial assistance by UGC are 
gratefully aknowledged. I am also thankful to 
Prof. N. R. Menon, Professor and Head of the 
Division of Marine Biology, Microbiology and 
Biochemistry, University of Cochin for his en-
couragement and facilities provided. 
References 
Cavari, B. Z.,. Uriel, 0., Gophen, M. & 
Berman, T. (1973) 1st Int. Cong. 
Bacterial. Israel. 11, 147 
Czeczuga, B. (1965) Hydrobiologia. 25, 
412 
Dhevendaran, K. (1978) Ph.D. Thesis, 
Annamalai University, p. 115 
Geonvese, S., Pichinoty, P. & Macri, G. 
(1963) Atti. Soc. Peloritana Sci. Fis. 
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Jimbo, T. (1938) Sci. Rept. Tohoku Univ. 
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Karanath, N. G., Santha Nair, K. & Loka-
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Kobayashi, M. (1975) Prog. Water Tech-
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Redfield, A. A. (1958) Am. SCientist. 
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Schlagel, H. G. & Pfenning, N. (1961) 
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Photosynthetic bacteria in the marine environment at Porto-Novo

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Photosynthetic bacteria in the marine environment at Porto-Novo

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