Effect of plant growth-promoting rhizobacteria at varying levels of N and P fertilizers on growth and yield of cauliflower in mid hills of Himachal Pradesh INTRODUCTION

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Effect of plant growth-promoting rhizobacteria at varying levels of N and P fertilizers on growth and yield of cauliflower in mid hills of Himachal Pradesh INTRODUCTION
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   Effect of plant growth-promoting rhizobacteria at varyinglevels of N and P fertilizers on growth and yield of cauliflower in mid hills of Himachal Pradesh MANOJ KAUSHAL, RAJESH KAUSHAL, B S THAKUR*   AND R S SPEHIA** * Regional Horticultural Research Station, Bajaura, Kullu ** Department of Soil ScienceDepartment of Basic SciencesDr Y S Parmar University of Horticulture and Forestry, Nauni, Solan ABSTRACT The use of plant growth promoting rhizobacteria (PGPR) is steadily increasing in agriculture andoffers an attractive way to supplement chemical fertilizers and pesticides by a wide variety of mechanisms. The indigenous isolates of PGPR were isolated from cauliflower rhizosphere soil indifferent agro-climatic zones of Himachal Pradesh. Five efficient isolates designated as MK2, MK4,MK5, MK7 and MK9 were selected and characterized after successful experiments under in vitroand net house conditions at varying levels of N and P on the growth and yield of cauliflower. All theisolates induced the production of indole acetic acid (IAA) and were able to solubilize phosphorus.The conjoint use of PGPR and N and P fertilizers resulted in a significant increase in number of non-wrapper leaves, curd diameter, curd depth and curd weight of cauliflower. Furthermore, PGPRisolates remarkably increased the yield of cauliflower. Among the five isolates, three isolates wereselected for field trials on the basis of the results obtained from net house studies in which MK5with 100 per cent N and P proved better in all aspects of cauliflower wrt increase in growth, yield,IAA production and phosphate solubilization. The present study, therefore, suggested that the useof PGPR isolate MK5 with 100 per cent N and P as inoculant biofertilizer was beneficial forcauliflower cultivation as it enhanced growth of cauliflower, and induced IAA production andphosphorus solubilization. Key words: IAA, PGPR, phosphorus solubilization, cauliflower.  Journal of Farm Sciences   1(1) : 19-26, 2011 INTRODUCTION In the context of increasinginternational concern for food andenvironmental quality, the use of plantgrowth promoting rhizobacteria (PGPR) forreducing chemical inputs in agriculture is apotentially important issue. PGPR have beenapplied in various crops to enhance seedemergence, growth and crop yield, but onlya few isolates have been commercialized.PGPR are also known to produce  Kaushal et al antibacterial compounds that are effectiveagainst certain plant pathogens and pests(Dey et al 2004, Herman et al 2008,Minorsky 2008). PGPR are directlyinvolved in increased uptake of nitrogen,synthesis of phytohormones, solubilizationof minerals such as phosphorus andproduction of siderophores that chelate ironand make it available to the plant root(Lalande et al 1989, Glick 1995, Bowenand Rovira 1999).Cauliflower (  Brassica oleracea  var botrytis  L) is a member of family cruciferaegrown as commercial crop for vegetablepurpose and as a seed crop in HimachalPradesh. The high-yielding cauliflowervarieties have resulted in an increase incauliflower production but require largequantities of chemical fertilizers which leadto health hazards and environmentalpollution. In order to make cauliflowercultivation sustainable and less dependenton chemical fertilizers, it is important to haveeffective PGPR isolates that canbiologically fix nitrogen, solubilizephosphorus and induce some substanceslike indole acetic acid (IAA) that cancontribute to the higher production of cauliflower. Therefore, the presentinvestigations were undertaken to screenthe PGPR isolates with multifariousactivities that are compatible withcauliflower for commercial cultivation inHimachal Pradesh. MATERIAL AND METHODS Collection of samples from cauliflower rhizosphere Soil and root samples were takenfrom the rhizosphere of cauliflower plantscollected from Bilaspur, Hamirpur andKangra districts of Himachal Pradesh. Therhizosphere was dug out with intact rootsystem. The samples were placed in plasticbags and stored at 4°C in the SoilMicrobiology Laboratory, Dept of SoilScience and Water Management, Dr YSParmar University of Horticulture andForestry, Nauni, Solan for further analysis.  Isolation from the soil sample  Soil sample of 1 g was taken andadded to 9 ml sterilized water blank formaking stock solution. From this stock solution 0.1 ml of sample was taken and9.9 ml sterilized water blank was added toit. It was marked as 10 -2  dilution. It wasserially diluted up to 10 -10  dilution. 0.1 mlof each of the same 10 -4 , 10 -6 , 10 -8  dilutionswere taken and added to sterilized pre-poured petriplates containing NA andPikovskaya’s medium. The plates wereincubated at 30± 2 0 C for 48 hours.  Isolation from the root sample The root sample (1 g) was takenand surface sterilized with 0.2 per cent 20  HgCl 2  for 3-5 minutes and then washedthoroughly (10 times) with sterilize distilledwater to make it free from HgCl 2 . Thesamples were immersed in the sterilizednutrient broth for 1-2 minutes in order todetermine surface sterility. The 1 g of rootsamples were grinded with 9 ml sterilizedwater blank in sterilized pestle and mortarto make slurry under aseptic conditions.The grinded samples were then used forisolation and enumeration of endorhizobacteria on different media byusing modified replica plating technique. Characterization of isolates Morphological characteristics(shape, elevation, surface, margin, etc) wereexamined on nutrient agar plates for thegrowth of selected bacterial isolates. Growth under different temperature and pH conditions Growth curves were drawn bygrowing the culture at various temperatures.Growth of the isolates at different pH wasstudied. The optimum pH and temperaturesuitable for the growth of bacterium wasworked out.  Methods adopted for multifarious plant growth promoting activities of  bacterial isolates Phosphate solubilizing activity,nitrogen fixing ability, siderophoreproduction, HCN production, IAAproduction and antagonistic activity weredetermined by the methods given byPikovskaya (1948), Jensen (1987), Schwynand Neilands (1987), Bric et a1 991,Bakker and Schippers (1987) and Gordenand Paleg (1957), respectively. Thebacterial inoculants were preparedaccording to the method of Vincent (1970).  Field experimentation  Three bacterial isolates viz MK5,MK7 and MK9 were selected on the basisof their multifarious plant growth promotingactivities under controlled conditions(growth chamber and net house). Theexperiment was conducted at RegionalHorticultural Research Station, Bajaura(Kullu). The seeds were treated withbacterial isolates for 8 hours. Untreatedseeds were designated as control. Theseeds were sown in nursery and one monthold seedlings were transplanted in the field.The bacterial culture was added at onemonth interval till harvesting. The treatmentscombinations viz:T1 (Control), T2 (MK5+ 50% NP), T3 (MK5 + 75% NP), T4(MK5 + 100% NP), T5 (MK7 + 50%NP), T6 (MK7+75%NP), T7 (MK7 +100% NP), T8 (MK9 + 50% NP), T9(MK9 + 75% NP) and T10 (MK9 +100% NP) were arranged in RBD designreplicated thrice. The observations wererecorded on different quantitative charactersof cauliflower viz number of non-wrapperleaves, curd diameter, curd depth, curd 21Rhizobacteria effect on cauliflower  Kaushal et al weight and curd yield. The data wereanalysed as per the method suggested byGomez and Gomez (1984). RESULTS  Morphological characteristics of PGPRisolates The morphological characteristicsof PGPR isolates (MK2, MK4, MK5,MK7 and MK9) varied widely. All theisolates produced round shaped and raisedcolonies having rough surface with undulatedto erosive margins.  Microscopic observations of PGPRisolates Microscopic observations wereperformed to investigate the characteristicsof PGPR isolates such as shape, gramreaction and motility. All isolates were rodshaped, motile and gram positive in reaction.The growth of MK5 isolate was better inthe temperature ranges of 25 to 40°C andpH 5-8 (Fig 1).  Production of multifarious activities The multifarious activities revealedthat all isolates induced the production of IAA. Isolates MK5, MK7 and MK9 werefound to be the producers of IAA,siderophores and HCN. These isolates alsohad ability to solubilize the phosphorus andantagonistic activities against  Rhizoctoniasolani , Pythium  sp and Fusarium  sp(Table 1).  Plant Parameters The PGPR isolates with differentlevels of N and P significantly affectedTable 1. Screening of bacterial isolates for multifarious plant growth promoting activities IsolatesP-N- freeAuxinSiderophore HCN Antagonism againstsolubilizationmediumproduction production production FusariumPythium    R spsp  solani MK2+++-----MK4++++-----MK5+++++++++++++++++MK7++++++++-+MK9+++++---+-+++ Very good activity, ++ good activity, + fair activity, - no activity 22  number of non wrapper leaves, curddiameter, curd weight, curd depth and curdyield/plot as well as per hectare as comparedto control (Table 2). Results revealedsignificant increase in plant parameters intreated plants over uninoculated control.The highest yield (355.56 q/ha) wasrecorded in MK5 isolate with 100 per centN and P fertilizers which was statisticallysimilar to isolates MK7 (315.80 q/ha) andMK9 (343.21 q/ha). DISCUSSION PGPR colonize plant roots and exertbeneficial effects on plant growth anddevelopment by a wide variety of mechanisms. The exact mechanism bywhich PGPR stimulate plant growth is notclearly established, although severalhypothesis such as production of phytohormones, suppression of deleteriousorganisms, activation of phosphatesolubilization and promotion of the mineralnutrient uptake are usually believed to beinvolved (Lalande et al 1989, Liu et al 1992,Glick 1995, Bowen and Rovira 1999).Of five isolates, one isolate MK5 wasfound to be good producer of IAA (Table1). It has been reported that IAA productionby PGPR can vary among different species   Incubation period (h)Incubation period (h)    G  r  o  w   t   h   (   O   D  a   t   5   4   0  n  m   )   G  r  o  w   t   h   (   O   D  a   t   5   4   0  n  m   ) Fig 1. Effect of pH and temperature on growth of  Bacillus  sp (MK5) 23Rhizobacteria effect on cauliflower
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