Application of Trichoderma harzianum and Bacillus thuringiensis in increasing plant growth of Aceh patchouli (Pogostemon cablin Benth)

Aceh Patchouli (Pogostemon cablin Benth), an essential oil crop, has been famous for a long time. The best quality patchouli producing area in Indonesia is Aceh Province. Trichoderma harzianum is one of the PGPFs that has been successfully used on a commercial scale while Bacillus thuringiensis as a plant growth-promoting rhizobacteria (PGPR). Improving plant growth, one of its roles, especially seed germination, and shoot elongation. The use of these two microbes indicates that plants grow well and healthy, this is part of the efficiency of plant disease management. In connection with the large number of patchouli plants in Blang Tingkeum Village, Mukim Lamteuba, Seulimum District, Aceh Besar, it is necessary to know whether Application of T. harzianum and B. thuringiensis in the field can increase the growth of aceh patchouli plants (Pogostemon cablin Benth), Plant height, number of shoots, and crown width were observed. The study used a non-factorial Randomized Group Design with 3 treatments and 7 blocks. PGPR and PGPF from both microbes are concluded to play a role in increasing plant growth and production based on the parameters mentioned above.


Introduction
Aceh Patchouli (Pogostemon cablin Benth) has been recognized as an essential oil-producing plantation crop.Indonesian patchouli oil exports have dominated the world market with a volume of 1,200-1,500 tons/year, compared to other essential oil producing plants [1].Patchouli plants have been popular in the village of Blang Tingkeum, a village located in Mukim Lamteuba, Seulimum District, Aceh Besar Regency, then farmers have stopped growing patchouli due to problems in cultivation and it is difficult to get a guarantee of a stable price for patchouli crops.
The rhizosphere has an important role in interactions between plants and microorganisms.Some species of fungi and rhizosphere bacteria can be utilized and have potential as plant growth promoting fungi (PGPF) and plant growth promoting rhizobacteria (PGPR).Trichoderma harzianum as a PGPF has been successfully used on a commercial scale for biological control of phytopathogens and plant growth promoters, Trichoderma helps plants be more resistant to environmental stresses such as salinity and drought by strengthening plant growth [2].B. thuringiensis is a plant growth-promoting rhizobacterium (PGPR) that promotes growth, seed germination, and shoot elongation.Compared to chemical control methods, biological control using PGPR has shown to be a more efficient technique [3].
From the description above, the two biological control agents compared to chemical control, biological control using PGPR shows efficient disease management [3].From the description above, the two biological control agents are able to increase plant growth.Tests on patchouli plants have been carried out in the field using local varieties and the effect of T. harzianum and B. thuringiensis increases the growth of patchouli plants in the field.

Materials and methods
The research was conducted at the Plant Disease Laboratory of the Faculty of Agriculture, Syiah Kuala University and in Blang Tingkeum Mukim Lamteuba Village, Seulimeum District, Aceh Besar Regency.Patchouli seedlings, pure culture isolates of antagonistic fungus T. harzianum and bacterium B. thuringiensis were used (both isolates are collections of Plant Disease Laboratory, Faculty of Agriculture, Syiah Kuala University).Golden snail and corn extracts have also been used as microbial propagation media.
Polybags filled with topsoil and compost (2:1) were placed in the nursery house.Cuttings of patchouli plants with a length of 20 cm were cut (4 stem segments), the leaves were removed leaving only the top leaves.The cuttings were planted in polybags with a depth of 5 -7 cm, placed in the nursery house.Plants are watered, weeds are weeded by pulling weeds that grow around the plants.
Rejuvenation of endophytic bacteria B. thuringiensis was carried out on NA media, then endophytic bacterial isolates were propagated on 100 ml of gold snail broth media, while the fungus was rejuvenated using PDA media and propagated on 100 g of cracked corn media.In order to be free of weeds, the land is stirred with a hoe, for fertilization using basic fertilizer (manure).In order for planting to be carried out properly, the size of the bed is 80 cm, 40 cm wide and 30 cm high.The 1-month-old patchouli seedlings are transplanted on the land, watered first and then the polybags are removed from the seedlings, ensuring that the seedlings roots are not disturbed.Seedlings were placed in the planting hole and then covered with surrounding soil.
Application of biological microbes to patchouli plants by dissolving T. harzianum isolates (100 g of corn media in 100 ml), B. thuringiensis isolates with gold snail media (15 L of water).Then applied as much as 240 ml per plant according to the treatment.Observations of plant height, number of shoots and crown width at 20, 40 and 60 days after application of biological control agents were carried out.

Results and discussion
The application of T. harzianum and B. thuringiensis had a significant effect on patchouli plant height at 20 DAP and very significant at 40 and 60 DAP (Table 1).The numbers in the same column followed by the same letter are not significantly different at the Least Significant Different test (α = 0.05).
Patchouli plant growth was highest in the T. harzianum treatment while the lowest patchouli plant height was in the control treatment.T. harzianum has a role as a plant growth-promoting fungus that acts as a trigger for excellent plant growth.It is suspected that T. harzianum can also act as a growth stimulant that can stimulate the growth of patchouli plants by producing phytohormones in the form of growth regulators or IAA hormones (stimulating the growth of plant roots and stems).Optimal root growth correlates with the optimal process of absorption of nutrients and water, thus the effect of plant height grows differently than the control treatment [4,5].
Changes in the height of patchouli plants aged 40 DAP as a result of the application of B. thuringiensis showed a significant difference compared to the treatment of T. harzianum and control.This is thought to be because B. thuringiensis is a group of PGPR (plant growth promoting rhizobacteria) that can increase plant growth in addition to biocontrol agents.Previous studies, this bacterium was found to reduce pathogen nematode attacked tomato roots [6] and Fusarium wilt [7].B. thuringiensis also can produce secondary metabolites that have a beneficial role for plants.Secondary metabolites produced by B. thuringiensis are hydrolytic enzymes (cellulase, protease, and chitinase), hydrogen cyanide (HCN), indole 3-acetic acid (IAA), 1-aminocyclopropane 1-carboxylic acid (ACC) deaminase, and siderophores (8).The ability of B. thuringiensis as PGPR enhances plant growth through several actions, both direct and indirect.This bacterium increases nutrient availability, phytohormone production in the form of indole acetic acid hormone, shoot and root development, protection against several phytopathogens to reduce disease.The increase in nutrient availability is because B. thuringiensis as PGPR is able to fix nitrogen and dissolve bound phosphorus [9,10].
The results of the application of T. harzianum and B. thuringiensis had a very significant effect on the growth and development of patchouli plants at 20, 40 and 60 DAP (Table 2).The number of shoots of patchouli plants in various treatments was significantly different compared to the control.The application of B. thuringiensis was able to increase the growth of shoots of this local variety of patchouli plants.These two microbes stimulate plant growth through the production of phytohormones such as auxins, or by affecting the plant's ethylene balance (ET), encourage nodulation in plants and can increase mineral nutrients (i.e.iron, phosphorus) obtained by plants [11].Some strains of B. thuringiensis possess PGPR activity in plant roots and have been shown to be beneficial for crop sustainability [12,13].The numbers in the same column followed by the same letter are not significantly different at the Least Significant Different test (α = 0.05).
Although the application of B. thuringiensis can increase the growth of patchouli plant shoots is not different from the treatment of T. harzianum, but the best treatment is the application of T. harzianum because it produces the most shoots compared to the others.T. harzianum acts as a high growth regulator that stimulates the growth of patchouli plants and finally patchouli plants can grow optimally [4].T. harzianum can accelerate plant growth by producing IAA compounds and increasing plant defense mechanisms [14].
In this study, the application of T. harzianum and B. thuringiensis had an effect on the total crown width of patchouli plants.The average crown width of patchouli plants at the age of 20, 40 and 60 DAP due to the application of T. harzianum and B. thuringiensis in Table 3.The numbers in the same column followed by the same letter are not significantly different at the Least Significant Different test (α = 0.05).
The application of T. harzianum and B. thuringiensis, both respectively on patchouli plants at 20 and 60 DAP were both significantly different of crown width then the control, however, at 40 DAP the B. thuringiensis treatment and the control were not different.The lowest crown width was found in the control treatment while the widest crown was in the T. harzianum treatment.This is strongly suspected because T. harzianum produces the auxin phytohormone indole-3-acetic acid (IAA) and its production can spur root and stem growth [5].As a result of optimal root and stem growth, the crown width expands.Plant canopy (crown) is also influenced by height, and plant branching, the higher the plant and plant branching will be directly proportional to the width of the plant crown in chili [15].The application of T. harzianum and B. thuringiensis can increase the number of leaves and crowns of Patchouli leaves compared to the control treatment [16], increasing plant size because the application of Trichoderma sp. can increase the C-organic content in the soil and ultimately improve crop quality [17].

Conclusions
The application of T. harzianum and B. thuringiensis in patchouli plants can increase the growth and development of patchouli plants.T. harzianum and B. thuringiensis are able to act as plant growth stimulators.Patchouli plants in the treatment of T. harzianum and B. thuringiensis at 40 DAP have an average plant height, and the number of shoots, and crown width were higher.In this study, the T. harzianum treatment showed the best results compared to the B. thuringiensis treatment.

Table 1 .
Average height of patchouli plants due to application of T. harzianum and B. thuringiensis at 20, 40 and 60 Day After Planting (DAP).

Table 2 .
Average number of shoots of patchouli plants due to the application of T. harzianum and B. thuringiensis at 20, 40 and 60 DAP.

Table 3 .
Average crown width of patchouli plants due to the application of T. harzianum and B. thuringiensis at 20, 40 and 60 DAP.