Indian Journal of Experimental Biology Vol. 37, May 1999, pp. 499-502

Response of micropropagated banana plantlets to Glomus mosseae at varied levels of fertilizer phosphorus

B N Shashikala, B J D Reddy & D J Bagyaraj*

Department of Agricultural Microbiology, University of Agricultural Sciences, G K V K, Banglore 560065, India

Received 27 July 1998; revised 5 February 1999

This investigation, conducted under glass house conditions, to study the effect of Glomus mosseae at various levels of phosphatic fertilizer on the growth of hardened micropropagated banana plantlets indicated that high soi l P levels inhibit the extent of root colonization of G. mosseae and thus curtail the growth and nutritional benefits of these fungi on plant growth. It is concluded that plantlets raised in soil inoculated with G. mosseae and amended with 50% of recommended P fertili zer exhibit a significant increase in plant height, number of leaves, stem girth , root volume, total biomass and total P content over those grown with 100% recommended P fertilizer.

The role of arbuscular mycorrhizal (AM) fungi in improving the plant growth and nutritional status and thei r possible use in crop production has been an area of intense research over several years. Investigations conducted hitherto reveal that these fungi improve the nutrilional status of host, particularly phosphorus status, and thereby enhance their growth, development and yield l. However, the extent of realisation of plant growth benefits depends on the host and fungal species involved in the association2.3

as these fungi are known to exhibi t preferentially colonise host roots4 . Si nce, a threshold levei of colonization must occur for the efficient functioning of this assoc ia! ion apart from the host involved, several sr, ; ' ': lI d env ironmental factors are implicated in dctermm;ng the extent of colonisation and hence the efficiency of this association. Among these factors soil phosphorus status is known to monitor the per cent root colonisation to a great extent. Several workers have shown that when P availability in soil increases the extent of infection is reduced5 and at extremely low P availability plants are not benefited by AM fungi though infection prevails but because plants incur a loss in photosynthates in maintaining the fungus and as P uptake is not facilitated by infection6. Further, the level of phosphorus needed to

*Correspondent author: Gram: UNIV AGRIS Phone : 09 1-080-3330 153 Ext. 372 Fax: 091-080-3330684, 091-080-3330684

determine the efficiency of this association varies with the fungal species involved, as AM fungi are known to differ in their tolerance to soil p7; host species involved, as plants differ markedly in their external and internal P requirementS and the soil type, as soils differ in their P fixation capacities9• Hence, it is expedient to determine a specific soil P level for a particular host-fungus combination in order to harness the maximum benefits from this association. Inoculation of mlcropropagated plantlets with AM fungi is known to improve vigour and growth of some horticultural cropslO. This investigation carried out with micro propagated banana plant lets , banana being an important fruit crop used as staple or complimentary food in countries of tropical and subtropical regions, was aimed at studying the interaction effect of Glomus mosseae and fertilizer P levels on the growth of micropropagated banana plantlets so as to know the possibility of reducing external P fertilizer application through AM fungal inoculation .

Materials and Methods A pot experiment was conducted under glass house

conditions to study the response of micropropagated banana (Musa accuminata cv. Nanjanagoudu Rasabale) plantlets to Glomus mosseae under varied levels of fertilizer phosphorus. The red sandy loam soil (fine, kaolinitic, isohyperthermic, kanhaplustalfs) used in this study was collected from an uncultivated field. Soil was crushed to pass through 4 mm sieve

500 INDIAN 1 EXP BIOL, MAY 1999

and was mixed thoroughly to obtain a homogeneous mixture. The soil was mixed with sand and compost in 2: 1 : 1 proportion and this potting mixture was filled to plastic pots at the rate of 3.5 kg/pot. The potting mixture used was deficient in phosphorus ( 3 mg/ kg available P, NH~ + HCl extraction) with pH 5.7 and an indigenous arbuscular mycorrhizal population of 30 spores/50 mI.

Glomus mosseae (Nicol. & Gerd.) Gerdemann and Trappe mailltained in pot culture with sterilized sand: soil as the substrate and Rhodes grass as the host was the AM fungi used in this study. Inoculum consisted of a mixture of sand : soil, extramatrical spores, extramatrical mycelia and Glomus mosseae colonized Rhodes grass root segments. This inoculum containing 150 spores/g was added at the rate of 25 g/pot to the planting hole according to the treatments.

Hardened, tissue cultured, healthy and uniform seedlings were planted at the rate of one plantlet per pot. The recommended dose of NPK fertilizers for banana under the soil conditions used in this experiment is 400 : 240 : 500 kg NPKlha (1 .52 : 2.62: 1.45 g NPKlpot). The recommended dose of nitrogen and potassium were applied in the form of urea and muriate of potash respectively to all pots. Phosphorus was added in the form of single super phosphate, according to the treatment level. Fertilizers were added two weeks after planting.

This experiment with four treatments namely, UPIOO = Uninoculated control with 100 per cent of recommended P, IPIOO = Inoculated with 100 % of recoI]lIllended P, IP75 = Inoculated with 75 % of recommended P, IP50 = Inoculated with 50 % recommended P, was laid out in a randomized complete block design and randomization among treatments was done once in 20 days . . The pots were watered regularly so as to maintain the potting mixture under field capacity and the plantlets were raised under natural light for 90 days.

Plant growth parameters like plant height, number of leaves per plant, stem girth, root volume and plant biomass were recorded at 90 days after planting. Plant height (cm) was measured from the surface of soil to the tip of the main stem. Number of leaves per plant represent the fully opened leaves and stem girth indicates the diameter of the stem 1 cm above the soil surface as measured by vernier callipers. Root volume (mI) is represented as the volume of water displaced upon immersing roots. Harvested shoots and roots were dried at 60°C to attain a constant weight and the

summation of the dry weight of shoot and root is indicated as the total biomass.

Fresh root samples (300 mg) were used for estimating per cent myeorrhizal colonrzation after staining by acid fuchsin (0.02%)11 . A 3 cycle clearing with 10 % KOH each for 30 min and treating the roots with sodium hypochlorite (3%) for one hour followed by treatment with ammonical hydrogen peroxide (H20 2 : H20: NH3 in the ratio of 50:50:3) for one hour and later with HCl was adapted before staining. These additional steps were used as banana roots were hard and highly pigmented. The stained roots were estimated for per cent mycorrhizal colonization using Grid line intersect method as outlined by Giovannetti and Mosse 12• Extramatrical chlamydospores in root zone soil samples (50 mI) were enumerated using stereo binocular microscope after wet sieving and decanting the soil samples13 •

Phosphorus content in shoot and root tissues was estimated calorimetrically. adapting the van ado-molybdate yellow colour method as outlined by Jackson'4. Data thus generated were subjected to statistical analysis of variance by RCBD and the treatment means were separated by Duncan 's Multiple Range Test (DMRT)'5.

Results and Discussion Arbuscular mycorrhizal fungi are well known to

enhance the nutritional status of several micro propagated plants and thereby aid io their increased growth and better establishment. Results of this investigation also clearly indicate a profound positive influence of Glomus mosseae on the growth of rnicropropagated banana plantlets with G. mosseae colonized plantlets showing an enhanced growth compared to those gr·own in the absence of G. mosseae inoculation (Table 1). Such improvement in growth and nutritional status of micropropagated plantlets in the presence of AM fungi are well documented in the case of apple '6 and grape vine17. The beneficial effects of this fungi on the growth of banana plantlets is so marked that plantlets growing at reduced fertilizer levels but in the presence of G. mosseae exhibited significantly higher plant height, number of leaves, stem girth, root volume and total biomass compared to those grown with 100 % recommended P fertilizer but in the absence of G. mosseae inoculation. Such increased growth is primarily contributed to enhanced nutrient uptake, particularly pIS apart from increased uptake of other

SHASHIKALA el al .: MICROPROPAGATED BABABA & GLOMUS MOSSEAE 501

Table I-Effec t of Glomus mosseae at different levels of fertilizer P on the growth parameters of micropropagated banana plantlets at 90 days after planting.

;r'reatment Plant hei ght No. of leaves Stem gi rth Root vol ume Total biomass (cm) (cm) (mllplant ) (g/plant)

UP IIM1 34.20d 8.80c 2.62b 175.80d 70.57c IP ml 39.20bc 12.60a 3.27a 296.50b 78.58a I P7~ 38.30c 10.90b 3. 19a 24 1.80c 75.28ab IP,,, 4 1.1 8a 10.50b 3.28a 393 .39a 81.3 1a

Means fo ll owed by the same letter within a column do not differ significantl y at P=0.05 by Duncan ' s Multiple Range Test.

Note: UPm l = Uninocul ated + 100% P fertilizer ; IP m ) = Inoculated + 100% P fertilizer ; IP75 = Inoculated + 75% P fertilizer ; IP50 = Inocu lated + 50% P ferti li zer

micronutrients 19 . Plantlets colonized by C. mosseae also had a higher content of shoot, root and hence total phosphorus compaf(~d to those raised either in the presence or absence of C . mosseae inoculation but at 100 % recommended P fertilizer (Table 2) demonstrating the ability of thi s fungu s to efficiently trans locate phosphorus from soil to plant through their ramified hyphae which contributes to the increased surface area for absorption6. Among the plantlets colonized by C. mosseae, those raised in the presence of 100 % P fertilizer have harnessed significant ly lesser benefits both in terms of growth and nutritional status compared to plantlets raised at 75 and 50 % recommended P fertilizers (Tables I and 2). Plantlets raised in the presence of C. mosseae and at 50 % recommended level showed the highest improvement in growth and phosphorus content compared to all other treatments. This indicates that the Hficiency of the fungu s-host association was maximum when phosphatic fertilizer was applied at the half the recommended level. This is correlated to the observation that banana plantlets growing in the presence uf 50 % recommended P fertilizer exhibiting the highest % root colonization and their rhizosphere containing the highest number of spores per 50 mJ of soil compared to all other treatments used in this study (Ta~le 3) .

It can be clearly observed that with an increase in the amount of phosphatic fertilizer application there results a significant reduction in the % root colonization and spore numbers (Table 3) . Such detrimenta l effect of high soil P on root colonization and extramatrical spores has been well documented20. It is also known that the efficiency of AM fungi measured e ithe r in terms of dry matter increase or increase in P uptake, is in verse ly re lated to the soil P status 21 . Further, under high so il P conditions the host is able to meet its phosphoru s requirements on its

Table 2-EITect of Glollllls mosseae at different levels of fertilizer P on the tissue P content of micropropagated banana plantlets at

90 days i)fter planting.

Treatment

UP 11K) lPliMI

lP75 IP,,,

ShootP content

(mg/plant)

80.60d IOY .70c 136. IOb 170.75a

RootP Total P content content

(mg/plant) (mg/plant )

20.95d 101.55d 45.26b 154.33c 35.43c 171.53b 58.26a 229.2 1 a

Means fo ll owed by the same letter within a column do not di ffer significantl y at P=0.05 by Duncan' s Multipie Range Test.

Note : UP 1(KJ=U ninoculated+ I 00% P fe rti lizer; IP ICK) = Inoculated + I 00% P ferti li zer; IP75=lnoculated+ 75% P fertilizer ; IPslF Inocu lated +50% P fertilizer

Table 3- Effect of fertilizer P on per cent root colonization and spore numbers of G. mosseae in root zone soi l of

mi cropropagated banana pl antlets 90 days after planting.

Treatment % root colonization Spore numbers/SO ml soil

UPICK) 35.00d 101.1 Od IP IIXl 63. ll c 38 1.20c IP7S 75.49b 425.39b IPso 83.37a 516.00a

Means foll owed by the same letter within a column do not differ significantly at P=0.05 by Duncan' s Multiple Range Test.

Note: UPIIK1= Uninocul ated+ I 00% P fertili zer ; IPltKI=lnocul ated+ 100% P fertilizer ; IP7, = Inocu lated+ 75% P fertilizer ; IPso= Inoculated+50% P fertili zer

own, which agai n depends on its root characteristics, and 'thus curtail s the extent of root colonizatic1n. However, banana plant lets raised in the presence of 100 and 75 % recommended P fertilizer though significantly cu rta iled root colonization by C. l110sseae compared to those raised in the presence of half the recommended level, the extent of colonization at these two soil P levels were very high (63. 1 I and 75.49 % respecti ve ly). Under such

502 INDIAN J EXP BIOL, MAY 1999

conditions where plant species fails to significantly curtail the extent of root colonization by mycorrhizal fungus the growth depression has been attributed to the utilization of photosynthates by the fungi22 and this observation is supported in the present study (Tables 1 and 2). Hence, the present study indicates that maximum growth benefits are realised by micropropagated banana plantlets when the soils are amended with half the recommended level of fertilizer phosphorus and inoculated with the AM fungus Glomus mosseae. This suggests that there is a possibility of reduction in the applicati.on of phosphatic fertilizers by 50 % when Glomus mosSeae is inoculated to the soils in which micropropagated banana plantlets are raised.

The results of this study clearly indicates that micropropagated banana plantlets are benefited when they are raised in the presence of AM fungus Glomus mosseae. Further, maximum benefits of this fungus-host can be harnessed when the soils are amended with half the recoIl1l11ended phosphatic fertilizers, thus resulting a saving in P fetilizer application and subsequently reduction in cost of cultivation.

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