SAINS TANAH – Journal of Soil Science and Agroclimatology, 12 (1) , 2015, 26-31 RESEARCH ARTICLE
RHIZOBACTERIA AS BIOCONTROL AGENTS OF ROOT ROT DISEASE ON SHALLOT
Nunik Iriyanti Ramadhan1, Hadiwiyono2, Sudadi2
1Undergraduate Student of Department of Agrotechnology, Faculty of Agricuture, Sebelas Maret University Surakarta
2Lecturer at Department of Agrotechnology, Faculty of Agricuture, Sebelas Maret University Surakarta
3Lecturer at Department of Soil Science, Faculty of Agricuture, Sebelas Maret University Surakarta Submitted : 2015-03-19 Accepted : 2016-02-19
ABSTRACT
Shallot is a high-economic value commodity, but so far the supply is still lower than the demand. One of the production problem is “moler” disease of shallot (MDS) caused by Fusarium oxysporum f.sp. cepae (FOCe). The aim of this research was to study the potentiality of shallot rhizobacteria (SRB) from various soil ordo to inhibit (MDS). This research was held in the Laboratory of Biology and Soil Health and Greenhouse at UNS. This research was carried out by exploring rhizobacteria of shallot planted on Entisols, Andisols, and Vertisols. Rhizobacteria exploration results were tested for their ability to control Fusarium oxysporum f.sp.cepae (FOCe). Inhibitory ability test of SRB to FOCe was carried out in vitro and on shallot in the greenhouse. The green house research used a Completely Randomized Design (CDR) with two factors. The first factor was rhizobacteria combination and the second factor was various soil ordo (Andisols, Entisols, and Vertisols). Each treatment was replicated three times. It was obtained three rhizobacteria isolates from Vertisols (B15: 70%), Andisols (B12:45,55%), and Entisols (B10:46,67%) being the highest inhibition results to FOCe. The combination of rhizobacteria B12 and B10 provided the lowest intensity.
Keywords : rhizobacteria, soil ordo, moler disease, shallot Permalink/DOI : http://dx.doi.org/10.15608/stjssa.v12i1.217 INTRODUCTION
Shallots (Allium ascalonicum L.) has a high value of economics because society have many needed of them. Shallots supply was lower than the demand (BPS 2012). One of the production problem was “moler” disease of shallot (MDS) caused by Fusarium oxysporum f.sp. cepae (FOCe). According to Ara et.al. (2006), MDS pathogen causes wilting quickly, decaying root, collapse, on the basal of bulb look whitish of fungal colonies bulb, yellowing leaves, and twisting leaf. Finally, shallots will die and tubers will be rotting (Fatawi et al. 2013).
Rhizosphere was inhabited by complex microbes, including bacteria and fungi.
Rhizobacteria diversity was influenced by the
complex properties of abiotic soil. Abiotic properties land was one of the distinguishing factors of soil types. Some of the suspected bacteria can be used as biocontrol agents of plants disease. Based on the above phenomenon was needed to obtain the rhizobacteria from various soil ordo as a biocontrol agents of MDS. Therefore, the aim of this research was to study the potentiality of SRB as biocontrol of MDS.
MATERIALS AND METHODS
Rhizobacteria Diversity from Various Soil Ordo
This research was held on September- October 2013 in the Laboratory Biology and
Soil Health belong to Agricultural Faculty the University of Sebelas Maret Surakarta.
This research used surface plating exploration method. The exploration results of SRB were differentiated that were based on colony and then were observed the diversity index. Diversity index could be interpreted as a systematic depiction illustrating the structure of the community and ease the information analysis process about kind and amount (Insafitri 2010). Diversity index was determined according to Shannon Wiever index.
H’ = diversity index
ni = amount each S spesies in sample n = total amount in sample
FOCe inhibition by Rhizobacteria in vitro Testing rhizobacteria’s potention from Andisols, Entisols, and Vertisols as biocontrol agents in vitro. Each rhizobacteria isolate was planted in PDA which has planted by FOCe before. It was incubated for three days. FOCe inhibitory test by rhizobacteria was held in the Laboratory of Biology and Soil Health belong to Agricultural Faculty at the University of Sebelas Maret in Surakarta on November-Desember 2013.
Moler Disease Intensity and Shallots Growth The test in vivo was held in greenhouse of Agricultural Faculty the University of Sebelas Maret Surakarta on March-July 2014.
The experiment used Completely Randomized Design (CDR) with two factors. The first factor was rhizobacteria combination and the second factor was various soil ordo (Andisols, Entisols, and Vertisols). Each treatment was replicated three times. Plant scoring was made with 5 orders. Score 0 : healthy plants. Score 1 : plants with wilted leaves. Score 2 : plants started to loose root due to decayed. Score 3 : shallot with no root due to decayed, wilted leaves but still had green leaves. Score 4 representated plants which died due to moler disease. The data were analyzed using F (Fisher) test at a level confidence 95% and Duncan’s Multiple Range Test (DMRT) at a level confidence 95 %. The diversity index was analyzed by Independent-Sample T test at a level confidence 95%. The assessment of antagonism potentiality of SRB to FOCe in vitro was based on the capability of inhibiting on the culture medium. The selected isolates of SRB potential were used for inoculating shallots with 10 ml (109 CFU/ml) planted on Vertisols, Andisols, and Entisols inoculated by FOCe 10 ml (109 spore/ml).
RESULTS AND DISCUSSIONS
Rhizobacteria Diversity from Various Soil Ordo
The results of rhizosphere exploration from various soil ordo were 75 isolates. The Isolates were grouped based on the colony morphology and obtained 17 isolates then be analysed the diversity index (Table 1).
Tabel 1. Diversity index of rhizobacteria of healthy and diseased shallots from various
Andisols, Entisols, and Vertisols used surface plating exploration method.
Plant condition Diversity index
Entisols Andisols Vertisols
Soil environment factors (physics, chemistry, biology) influence soil microbes for growing (Chau et. al 2011). According to the exploration results, Vertisols isolates had the highest diversity index for both from healthy and diseased shallots. Notohadiprawiro (1988) explained that high diversity microorganism participate to suppression the soil pathogens.
FOCe on the diseased shallots rhizosphere, grew well because of the low diversity SRB. The diversity index of SRB from Vertisols was higher than the others, possibly it was related to the pH being closest to neutral (6,4) where as the soil texture was clay. This was due to the texture of the soil known not to affect the diversity index, but only affect the richness (Chau et.al 2011). Soesanto (2006) explained that rhizobacteria could live optimally at neutral pH (6,6-7,0), where as some rhizobacteria could live at acid to alkali environment. The diversity of SRB Andisols and Entisols index were lower than Vertisols because of the pH which was more acid. Soil microbes on low pH was dominated by fungal than bacteria. Despite of that, the presence of rhizobacteria was known to be increasing with high CEC. CEC influences nutrient availability in soil that was needed by rhizobacteria (Eichorst et.al. 2007). This suggestion as supported by
the analysis of N, P, and K that were high in increase CEC.
FOCe Inhibition by Rhizobacteria in Vitro Results of in vitro test showed that some rhizobacteria could inhibit FOCe. (Figure 1). The highest inhibition percentage was showed by isolate B15 (70%). This rhizobacteria could be found in all soil ordo. SRB isolate B02, B03, B04, B09, and B17 could not inhibit the pathogen.
According Soesanto (2006) disease inhibiton mechanism was generally differentiated to competition, parasitism, ISR (Induced Systemic Resistance), and antibiosis.
Possibly, Fe competition and antibiosis contributed on antagonism of SRB to FOCe. Fe was needed to physiological process for any microorganisms (Weber et.al 2006). In poor nutrient Fe (in media) rhizobacteria would synthesize siderophore which had better holding ability for Fe than fungi. Antibiosis was a secondary metabolit compounds that harmful to others organism. FOCe hyphae changed from purple into red. This mechanism is called antibiosi. Some antibiotics which produced by some bacteria were volatil (Yuan 2012), 2-4- diacetylphoroglucinol (Sessittsch et al 2004),HCN, salicylic acid, and siderophore (pyoverdine) (Djatnika 2012).
Figure 1. Grow inhibition percentage of FOCe by rhizobacteria. Value which is followed by same letter is not significant.
Disease Intensity and Shallots Growth.
Plant Height
S
oil ordo significantly influenced to plant height, but SRB combinations did not (Figure 2). Shallots planted in Entisols had highest plant height. This was due to Entisols had loam texture. Shallot was most suitable to plant in soil with good aeration-draenation (porous) (Samadi dan Cahyono 2005).Vertisols was dominated by clay fraction. Type of this clay was montmorilonit (type 2:1) which could expand and contract (Foth 1943).
The condition can disturb the roots to grow.
Andisols had sandy loam texture.
According to Foth (1943) sand which composed this ordo was pseudosand.
Pseudosand formed when the soils was in dry conditions (water-film evaporate) so the bonds between particles would be closer then the particles size change to be big with allophone aggregation with others particles.
When it was run into drought, it was difficult to bind the water again though it was dampened because irreversible. This case would always make plant growth obstructed.
Rhizobacteria combinations were showed their influence to the difference plant height.
This case was caused by rhizobacteria isolates inoculated could not provide nutrient which supported the growth. Generally, shallots had soft root so sensitive to the soil conditions,
especially texture. Chau et.al. (2011) explained that richness increase had positive correlation with soil coarseness increase.
Richness increase in coarse soil was suspected causing amount of water-films where was isolated in soil. According to this explanation, rhizobacteria inoculated in Entisols could grow well.
Dry Weight Plant
Dry weight of plant was a plant biomass.
Dry weight were obtained after plant was roasted in oven with temperature at 700C for 3 days. Results of F test showed that the effect of rhizobacteria combinations and various soil ordo were not significant dry weight of plant (Table 5).
Various soil ordo was not showing significant impact to dry weight plant though it had positive correlation with plant height.
This case was caused by contents of kalium in the soil was low and very low so insufficient for plant needs. Foth (1943) explained that kalium took a role in growing cell wall thickness so tuber crops need a lot of kalium.
But, shallots in Entisols tended to have heavier dry weight.
Rhizobacteria combination did not give a significant impact to dry weight plant. This case was caused by rhizobacteria isolate was not known could provide nutrient increasing
Figure 2. Plant height of shallot planted on three soil ordo. Value which is followed by the same letter is not significant.
dry weight, such as kalium. But, there was difference in some isolates. Shallots inoculated by B15 gave dry weight more heavier in all soil ordo.
Disease Intensity
According to Fatawi et al (2003), MDS would cause the shallots to be dead or wilting leaves quickly and decay on the root sand plate. This study used scoring method with key field for observing the disease intensity.
Based on key field, the results of observation of the disease intensity were represented at Figure 3.
F analysis results was showed that rhizobacteria combination gave significant
effect to disease intensity. The highest disease intentsity was showed by B15 and B10 combination, and the lowest was showed by B12 and B10 combination. Isolates B15 had the highest inhibition ability than the others and could be founded in all soil ordo. This test results was inconsistant with the results of greenhouse experiment. Notohadiprawiro (1988) explained that the factor suppressive disease were diversity of biological communities and antagonism. So, it is needed an intact soil ecosystem to suppress the disease. Despite of that, antagonism between SRB which had been inoculated on the shallot was possible.
Tabel 6. Effects of rhizobacteria and soil ordo to dry weight shallots inoculated by FOCe.
Rhizobacteria isolate Dry weight (g)*
Vertisols(V) Andisols(A) Entisols(E)
Without FOCe 2,96±0,28 a 2,27±0,69 a 2,66±0,16 a
Without Rhizobacteria 2,76±0,73 a 2,80±0,75 a 2,33±0,13 a
B15 3,59±0,43 a 3,45±0,71 a 3,49±0,33 a
B12 2,01±0,29 a 2,86±0,25 a 2,87±0,28 a
B10 2,90±0,12 a 2,78±0,14 a 3,17±0,60 a
B15 B12 2,31±0,29 a 2,84±0,17 a 3,04±0,85 a
B15 B10 2,34±0,19 a 2,11±0,43 a 2,57±0,05 a
B10 B12 2,62±0,31 a 2,70±0,32 a 3,52±0,58 a
B15B10B12 2,87±0,29 a 2,40±0,29 a 3,16±0,24 a
*average ± standard error
Figure 3. Effect of rhizobacteria SRB to disease intensity of MDS. Value which is followed by the same letter is not significant.
The lowest disease intensity that could be suppressed by SRB was different at different soil ordo because various soil ordo did not give significant impact for the disease intensity. Various soil ordo were not give significant impact to plant height, like as discussing before. Appropriate to this research, B10 and B12 combination that were applied in Entisols tended to suppress disease intensity and gave positive impact for plant height.
CONCLUSION
The most potential rhizobacteria from shallots rhizosphere to suppressed “moler”
disease intensity in vitro was isolate B15 that could be found in all soil ordo with inhibition ability to the pathogen up to 70 %.
Combination isolate B10 and B12 that was applied at Entisols was the most capable to suppress the disease intensity up to 11,11%
and gave positive impact for plant height.
ACKNOWLEDGEMENTS
This research was funded by an Excellent Research Grants of Sebelas Maret University in 2013 entitled “Integration of Functional Microbial Consortia of Providers Nutrient and Soil Borne Prevention of Biological Agents as Biofilmed Biofertilizer”.
REFERENCES
Ara K et al. 2006. Foot odor due to microbial metabolism and its control. J Microbiol 52(4)357-64.
BPS (Badan Pusat Statistik) 2012. Produksi sayuran di Indonesia, 1997-2012.
http://www.bps.go.id. Diakses pada tanggal 26 September 2013.
Chau JF, Bagtzoglou AC, Willig MR 2011. The effect of soil texture on richness and diversity of bacterial communities. J Env Forens 12:333-341.
Djatnika I 2012. Seleksi bakteri antagonis untuk mengendalikan penyakit layu fusarium pada tanaman phalaenopsis. J Hort 22(3):276-284.
Eichorst SA, Breznak JA, Schmidt TM 2007.
Isolation and characterization of soil bacteria that define “terriglobis” gen in the phylum “acidobacteria”. J App and Env Microbiol 73(8): 2708-2717.
Fatawi ZD, HS Gutomo, Hadiwiyono 2003.
Studi lini dasar terjadinya epidemi penyakit busuk pangkal bawang putih di tawangmangu. Laporan Hasil Penelitian Penelitian Sumber Dana DUE-Like
TA.2003. PS. Agronomi. F.
Pertanian.UNS. 45 Hal.
Foth HD 1943. Fundamental of soil sciences.
Yogyakarta (ID): Gadjah Mada University Press.
Insafitri 2010. Keanekaragaman, keseragaman, dan dominasi, bivalvia di area buangan lumpur lapindo muara suangai porong. J Kelautan 3(1): 54-59.
Notohadiprawiro Tejoyuwono 1988. Tanah dan lingkungan. Direktorat Jenderal Pendidikan Tinggi Departemen Pendidikan dan Kebudayaan.
Samadi Budi, Cahyono Bambang 2005.
Bawang merah. Yogyakarta (ID):
Kanisius.
Sessitsch A, Reiter B, Berg G 2004. Endophytic bacterial communities of field-grown potatoplants and their plant- growth- promoting and antagonistic abilities. J Microbiol 50:239-249.
Soesanto L 2006. Pengantar pengendalian hayati penyakit tanaman. Jakarta (ID):
Rajawali Press.
Weber KA, Achenbach LA, Coates JD 2006.
Microorganism pmping iron: anaerobic microbial iron oxidation and reduction. J Nat Rev Microbiol 4: 752-764.
Yuan J, Raza W, Shen Q, Huang Q 2012.
Antifungal activity of bacillus amyloliquetaciens NJN-6 volatile compounds against Fusarium
oxysporum f.sp. cubense. J App Env Microbiol 78(16): 5942-5944.
