From the live and decomposing S. molesta 13 strains of bacteria were isolated and the growth promoting effect of the weed

extract in various concentrations in 5 selected strains of bacteria and a mixed culture was studied. Turbidity measurements

gave maximum percentaae transmission in 5 % extract. Dry weight loss of dried weed in the laboratory was 20 and 25 % after to

and 25 days of incubation respectively. Yield efficiency of 3 bacterial strains calculated for 48 hr was 20%. The results suggest

the potential value of this primary source of organic material in the food web of Cocbin backwater

Gore, P S

Radhakrishnan, E V

Raveendran, O

Unnithan, R V

CMFRI Digital Repository

Indian Journal of Marine Sciences Vol. 8. September 1979, pp. 17(1.174 Microbial Decomposition of the Floating Weed Salvinia molesta Aublet 10 C6chin Backwaters E V RADHAKRISHNAN·, P S GORE, 0 RAVEENDRAN & R V UNNITHAN NationallnstituU: of Oceanograpby, Regional Ceotre, Cochin 682018 bcei.u 18 NoW!lllbn- 1978: re.,ised receil'eti 18 May. 1919 From the lil'C and decomposing S. mo/uto 13 strains of bacteria were isolated and I:bc: growth promoting effect o.f the weed extract in various concentrations 00 5 selected strains of bacteria and a mixed culture was studied. Turbidity measurements p...e maximum percentaae transmission in 5 % extract. Dry weight loss of dried weed in the laboratory was 20 and 2S % after to aud 25 days ofincubatiOD respecti...ely. Yield efficieocy of3 bacterial strainscakulated for 48 hr was 20 %. The results suggest ~ potential value of this primary source of organic material in the food web of Cocbin backwaten. Nutritional significance of detritus along with associated micro-Qrganisms in estuarine ecosystem has been studied l - 6. The Cochin haelcwater system receives allochthonus organic matter, primarily from the floating weed Salvinia molesla during monsoon. There is no information on the microbial association and detritus formation from the weed in Cochin haelcwaters though some infollDl!tion is available on the animal association 7 and seasonal "fluctuation and chemical characteristics8 of detritus in the haelcwaters. . Present study attempts to understand (i) utilisation of soluble weed components by aerobic, heterotrophic bacteria, isolated from fresh as well as decomposing weed in the baclcwaters, (ii) microbial decomposition of dried weed in the laboratory and (iii) yield efl;iciency (conversion oflow protein weed biomass into bacterial protoplasm) of selected strains of bacteria. Materials and Metbods Fresh and decomposing S. molesta from baclcwaters was collected (September and October 1975) in sterile glass containers and carried to the laboratory in iced condition. The weed (5-10 g) was aseptically chopped into fine pieces and 6numeration of viable counts carried out on the suitably diluted samples using sea water agar (SW A) and freshwater agar (FW A) media by the pour plate methods. Morphological and biochemical tests were performed9• Large samples of the fresh weed were also collected from the same area to prepare weed extracts of desired concentrations I ° (0.5, I, 5 and 10 %) to study the growth promoting effect on selected strains of bacteria. Millipore filtered extract was dispensed (20 mlj lOO mI conical flask) and *Prc::sent address: Madras Research Centre of CMFRJ. 29 Commander·io-Chief Road. EjpDore. Madras 600008. 170 , sterilised at 15 Ib pressure for 15 min. The generic classification of 13 strains of aerobic, heterotrophic bacteria was done as per the modified scheme of Simidu and Aisoll. Morphological and biochemical . characteristics of the isolates are given in Tables I and 2 \ Each of the 13 isolates maintained on sea water agar slant were tested for their ability to grow in 5 % weed extract broth. Growth of 5 isolates (El Serratium,.E2 Flavobacterium, E3 an unidentified gram negative yellow cocci, 8 Pseudomonas and 13 Bacillus) and a mixed culture of bacteria (MC), in different concentrations of-steri1ised weed extract (0.5, I, 5 and IO%) was studied at 28±2°C (Fig. I). The extracts inoculated with the bacterial isolates were incubated for 48 hr in a waterbath shaker. Growth response after 48 hr incubation was measured at 620 nm in a 'Specometer' with corrections to exclude absorption by the pale brown weed extract (Fig. I). To study variations in growth between the strains and between concentrations of the extract tbe results were subjected to analysis of variance. Loss in dry weight of dried weed in the laboratory due to microbial activity was studied 10. For estimating conversion of weed into microbial biomass, 3 strains of bacteria (E I Serratium, 8 Pseudomonas and 13 Bacillus) were allowed to grow in 3 different concentrations (0.5, I and 1.5 %) of weed extract prepared in sea water. Weed extract dispensed (20 mil into conical flasks was sterilised and inoculated witb 2 mI of actively growing nutrient brotb culture. Turbidity was measured using a Specometer (Elico model: C4-21). After 48' hr incubation dry weight of bacteria was estimated by the filtration method 10. The values for dry weight of 3 cultures determined at each level of weed extract are presented in Fig. 2. In this ---" RADHAKRISHNAN et ai.: MICROBIAL DECOMPOSITION OF S. MOLEST A Table I-Morphological and Growth Characteristics Strain Description of colony on SW A Growth No. Nutrient broth SWA slant FWA No visible growth Round, surface, convex, rose coloured colony Uniform. turbidity. thin pellicle at top Filiform, . dry growth 2 Round. surface, white colony with complete Medium turbid, no pellicle Filiform do margin 3 Round. surface, cream coloured colony with Medium turbid, membranc:ous pellicle do Poor growth complete margin at top 4 Horse-shoe shaped, surface white colony with Uniform turbidity. no pellicle do ·lobated margin 5 7 Round, ;iUrface, pale yellow colony Medium turbid. no pellicle do Luxuriant growth No growth Round,sub-surface, white colony with smooth Uniform turbidity. thin pellicle at top do surface 8 Oval ~ped. sub-surface white colony with Uniform turbidity thin pellicle do Poor growth frilled margin 9 10 Round, surface, white colony Uniform turbidity. no pellicle do Fair growth No growth Round, sub-surface small white $X>lony with Uniform turbidity do metallic shining 11 13 Round, surface, small. white colony UWrorm turbidity thin pellicle: at top do Poor growth Thick luxuriant growth Poor growth Round. white, sub-surface. feather margined Uniform turbidity. DO pelliCle Filiform lux~riant colony growth 14 Oval, cream coloured, surface colony with Uniform turbidity, no pellicle Filiform frilled margin 15 E-I Oval, pale yellow, surface colony Uniform turbidity do do Good growth Oval, red surface colony do do Other morphological characteristics: Motility Growth at pH 6 Pigmentation, Irridescence All strains motile except 10 and I I which were passive All showed good growth excq>t strain I which showed satisfactory growth with turbidity Str~ I, rose-pink; strain 5, yellow; and strain E-I. red Strains 2 and 3, ~ and strains 8, II and 14, bluish experiment the number of bacteria per ml determined by dilution and plating after 48 hr in the medium containing 500 mg of extract per 100 ml was isolate E l' 1.03 x 108; isolate 8, 1.64 x 108 and isolate 13, 2.06 x 108. Results and Discussion Growth response in 5 % weed extract broth indicated that 8 bacterial isolates (strain Nos. 1,3,4,5, 8, 10, 13 and 15) grew well in the extract while the rest exhibited slight growth. When all the isolates were tested for their ability to grow in standard nutrient broth and sea water broth, only 3 (strains 5, 9 and 13) showed appreciable growth in standard nutrient broth, 4 showed no growth and 5 showed poor growth (Table I). All isolates showed appreciable growth in sea water broth. Of the isolates 50 % grew better in weed extract medium than in sea water nutrient broth indicating the bacterial growth promoting factor of the weed. Many of the isolates appeared to be· marine bacteria with respect to their requirements of salinity for their growth l2. The growth promoting actlVlty of the soluble components of the weed is clearly evident from the experiment- Data were analysed using analysis of variance. For framing the analysis of variance, the peICeIltage transmission data were converted to angles using angular transformation, in order to stabilise the variance. There was significant (P <0.01) variation between percentage transmission values. The variation between strains was not siguificant at 5 % level. The least significant difference (LSD) at 5 % level was formed and the mean of percentage transmission compared. PeICeIltage transmission in 5 % was highest compared to percentage transmission in 0.5 and 1%. The percentage transmission in 10% was less than that of 0.5%. The percentage loss of organic matter during the initial 10 days of incubation was 20 % whereas only 5 % decomposition was noticed in the next 15 days. The initial loss of 20 % may be attributed to the ready decomposition of protein and other soluble extractives, which were estimated 13 to be 42 %. These components are easily susceptible to microbial attack. 171 INDIAN J. MAR. SCI., VOL 8, SEPTEMBER 1979 Table 2-Biocbemical Cbaracteristics Strain Hugh .t Leifsoo's Fermentation of sugars Gram staining Bacteria No. Test (sucrose, glucose, maltose, mannitol and lactose) NF Nil ( -)Ye PseudOfrronas sp. elongated rods 2 FNG All e;u:ept lactose (-)Ye Vibrio sp.· sbort rods 3 NF Nil ( -)ve Pseudomonas sp. elongated rods 4 do Nil do Pseudomonas sp. S FNG All except lactose (+)Ye Bacillus sp." rods with central spores 7 NF Nil ( -)ve PMMlomonas ,sp. elongated rods 8 do Nil (-)ve P.seudomonas sp. short rods 9 do Nil (-)Ye , Achromobacter sp." elongated rods 10 do Nil do P~monassp.· II do Nil (-)Ye Pseudomonas sp. short rods 13 FNG All e;u:ept lactose (+)ve Bacillus sp. spo~forming rods with central spores .'--14 NF Nil (-)ve Achromobacter sp. short rods IS do Nil ( -)ve Achromobacter sp. elongated rods B-1 do All except maltose & lactose ( -)Ye Serralium sp. NF = noD" fermentative and FNG = fermentative, DO gas *Deoitrifier Otber biocbemica1 characteristics: Strain No.2 positive; rest all negative short rods Indole production VP reaction Gelatin-liquefaction Nitrate reduction Starch digestion Sensitivity towards penicillin Strain Nos. 5, 13, and E-I positive, rest all negative Eight strains (1 , 2, 5,9, 13, 14, 15 and E-I) positive; rest negative All strains positive S~ 'Strains (1,2, 9, 13, 14 and E·1) positive; rest negative . Five strains (5, 9, 13, 14 and ~1) only sensitive The remammg 5 % loss may be due to the slow decomposition of more complex fractions such as ceUulose and hemicellulose (27 % of the dry weight)14, which are less susceptible to microbial attack. The yield efficiency for the isolates El' 8 and 13 for 48 hr is 20 %. This is comparable to the yield obtained for salt marsh grass I 0 for 48 hr incubation. According to Gosselink and Kirby I S yield efficiency calculated for 48 hr may not reflect the maximum bacterial biomass that has been achieved during the period. They have further shown that yield efficiency can be reached even up to 46 % for a longer time interval as decomposition of more refractory fractions were expected within that period. 172 Burkholder and Bomsbyl0 assumed that 11.2 % of the annual crop of marsh grass may be available for rapid production of bacterial biomass if 20 % of the soluble extractives and protein is readily converted. Using the same relationship it may also be assumed that 8.4 % of the weed will be available for bacterial biomass production in Cochin backwater system also (0.42 x 0.2=0.084, i.e. 8.4 %). Role of allochthonous organic matter as a major energy source in salt marsh ecosystems and streams have been well emphasisedJ.6. AccordIng to TaitI 6 in detritus based ecosystem, some of the energy is directly available to the benthic fauna by digestion and assimilation of detritus, but many detritic materials RADHAKRISHNAN e/ al.: MICROBIAL DECOMPOSITION OF S. MOLESTA • 0-f-, 0 0 EI E3 StraIn numbers Fig. I-Growth of 6 bacterial isolates in different concentrations of weed extract in sea water reaching the bottom cannot be digested by animals. From the experimental evidence available he further assumed that 25 % of the energy intake of the benthic herbivores come directly from detritus and 75 % from the consumption of associated bacteria. From the present study it is seen that Salvinia detritus of Cochin estuary is acted upon by bacteria whIch may contribute to a significant proportion of the food of benthic animals in the estuary. Detailed siudies of energy flow in natural ecosystems are available3. l7" 8. But studies of the energetics of a tropical estuarine ecosystem like Cochin backwater system are limited. From the studies of organic producti9n and the food chain of Cochin backwaters8. l9 it is known that a major portion of energy in the form of detritus is available. The detritus 250 ~ • E 20 / 0 2 • • -a> E 150 • 8 ~ 3' >-./: E, L 100 '0 .<:! L • .. ~ u 0 50 '" 500 1000 1500 Weed extroct mg/,ooml Fig. 2-Dry weight of bacterial culture grown in different weed extract concentrations constitutes an additional pathway between organic production and animal nutrition increasing the efficiency of energy transfer from one trophic level to other. The production of Salvinia detritus is seasonal in that the fresh weeds come to the estuarine system in monsoon months, and undergo decomposition leaving the detritus, which accumulates in the bottom. If a large proportion of detritus exists in the sediments then even a slow rate of decomposition would be sufficient, to support the coilsumer populations. Fenchal6 has stilted that the existence of a large quantity of slowly decomposing plant body may enable the ecosystem to continue functioning even when the primary producers are temporarily removed. Similarly the existence of decomposing Salvinia to some extent may prove beneficial, especially in the absence of other primary producers. Acknowledgement The authors are grateful to Dr S. Z. Qasim, Director, for encouragement. Thanks are due to Prof. N.K. Velankar, Consultant & Head of the Department of Industrial Fisheries, University of Cochin, for going through the manuscript and also to Shri H. Krishna Iyer for help in the statistical analysis. One of the authors (EVR) is grateful to CSIR for the award of SRF. References 1 Darnell R M. in Estuaries, edited by G H Lauff (Pub!, No. 83, AAAS, Washington) 1967,376. 173 

Microbial decomposition of the floating weed Salvinia molesta Aublet in Cochin backwaters

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Microbial decomposition of the floating weed Salvinia molesta Aublet in Cochin backwaters

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