A Review on Clonal Propagation of Medicinal and Aromatic Plants Through Stem Cuttings for Promoting Their Cultivation and Conservation

Introduction

Medicinal and aromatic plants (MAPs) form the basis for plant based therapies including the Indian/ Chinese/ Tibetan Systems of Medicines, which are being popularized in many parts of the world. Quality of these plants largely depends on active ingredients present in them. Though most of the raw drugs are sold on fresh/dry weight basis in the domestic market,¹ during the extraction process it is observed that superior the chemotype, better is the quality of the extract, and maximum the returns obtained per unit of charge. From time immemorial,  crude drugs have been collected from wild and thus, population of most of the important MAP species is dwindling in nature.² A number of species are on the verge of extinction due  to problems such as destructive harvesting, poor seed set, low seed viability, pest and disease incidence etc.^3-4 Though the existing populations of these species are being regenerated in the nature by means of sexual reproduction, the rate of multiplication is slow and thus insufficient to meet the requirements in most of the cases.⁵ The ever-increasing demand for these products indomestic and international markets also calls for a faster multiplication strategy.⁶ Good number of improved varieties of MAPs is now available⁷  that are being popularized amongst farmers due to their higher net returns and ease of cultivation, thereby reducing extra burden on the forests. Propagation by vegetative means is not only relevant in case of improved varieties/hybrids of commercially cultivated species but also for conservation of those, which are endemic, threatened and are on the verge of extinction.³ Multiplication of these species by clonal means such as cuttings can make cultivation much economic, providing more uniform population and active ingredient yield per unit area.⁸ In some  cases  such  as  Centellaasiatica,  plants  raised  from  cuttings  have  been proven to perform better for biomass production than those raised from seeds.⁹

Understanding the skills involved in vegetative propagation techniques ensures faster multiplication tomeet the growing demand from consumers, and thereby keeping collectors away from forests for collecting such resources. Adventitious root formation has been regarded as a complex process as it is highly influenced by different internal and external factors.¹⁰ Poor adventitious root development is considered as a major hindrance in propagation through cuttings.¹¹ Some species may produce roots even without any treatments as observed in case of Cinnamomumasomicum and C. impressinervium,¹² whereas in other species propagation by cutting might not be feasible as in Aconitum heterophyllum.¹³ For example, hard wood cuttings of a threatened species Saracaasoca exhibited rooting in mere 16.67% cuttings, when treated with 500mg/lIndole-3 butyric acid (IBA) and hence, propagation via air layering was advocated for propagation with 90% success.¹⁴ Nevertheless, considering the practicality of the technique, ease of operation, cost involved etc., propagation by cuttings has remained a technique of choice in the past and will continue to be in future as well. Even though propagation through cuttings appears to be the simplest among all the methods of vegetative propagation, some aspects need to be understood to obtain better results. Considering these, factors governing the success of vegetative propagation through cuttings in MAPs (Figure 1) have been reviewed hereunder.

Figure 1: Major factors governing success of propagation by cuttings in MAPs Click here to View Figure

 

Factors governing success of propagation through cuttings in MAPs

Physiological maturity of cuttings

Stem cuttings are classified as hardwood, semi-hardwood, softwood and herbaceous cuttings based on physiological age of the wood from which they are excised.^15-16 Hardwood cuttings are taken from dormant, mature stems of more than one year and are commonly adopted in MAPs such as Indian Myrrh, Rosa spp., Henna etc. Semi-hardwood cuttings are usually prepared from partially mature wood of the current season’s growth, which is practiced in species such as Embelia spp., Salaciaspp.etc. Softwoodandherbaceous cuttings are prepared from soft, succulent new growth of plants¹⁶ and are commonly employed for the propagation of species of mints (Mentha spp.), brahmi (Bacopa monnieri) and other herbs.

Dick and Dewar¹⁷ opined that variation in carbohydrate pools could be the main factor determining rooting ability. Classically, internal factors such as auxins, rooting co- factors, C:N ratio etc. have shown to influence root initiation process.¹⁸ Hardwood stem cuttings of Gongronemalatifolia, an important African medicinal species,produced significantly higher number of roots compared to when propagated by semi-hardwood and softwood cuttings.¹⁹ Similarly, in case of J. grandiflorum, hardwood cuttings exhibited highest regeneration capacity compared to semi-hardwood and softwood cuttings.²⁰ In case of Pogostemonheyneanus, two nodal hardwood cuttings performed better in rooting parameters when compared to semi-hardwood and softwood cuttings.²¹ Positive relationship between diameter of cutting and rooting capacity in Azadirachtaindica has been previously reported,²² which could be explained by higher carbohydrate reserves in thicker cuttings.

On the contrary, in Salacia fruticosa, Salacia reticulata and Embeliaribes, use of semi-hardwood cuttings gave superior rooting response than that by hardwood cuttings.^4,23-24 Based on diameter, thin (0.43 cm) and medium (0.52 cm) cuttings were found to be superior for increasing sprouting and survival in Embeliatsjeriam-cottam and Caesalpiniabonduc than thicker cuttings.³ Secondly, portion of the stem used for excising cutting has been found to contribute in root induction. Rooting ability of cuttings taken from different parts of the stem of Ficus species indicated that higher rooting rates (>52%) were obtained with pole and nodal cuttings as against 5% in case of terminal cuttings.²⁵ Low rooting ability of apical cuttings could be attributed  to  their  herbaceous  character,  which  made  them  sensitive  to  moisture  stress, resulting in desiccation.²⁶ Middle or apical cuttings of about 25 cm length were adjudged as the best planting material for regeneration of Commiphorawightii, the Indian myrrh.²⁷ Similarly, in Damask rose (Rosa damascena), basal woody cuttings were found to give highest rooting percentage with maximum number of roots.²⁸ Nevertheless, the response is species dependent as Khosla and Pushpangadan²⁹ observed that the current year, young lateral shoots were the best planting material for rooting of clocimum (Ocimumgratissimum). Most suitable physiological maturity of the cuttings in some important MAPs have been presented in Table 1.

Table 1: Some examples of different types of cuttings based on their physiological maturity in MAPs.

Type of cutting	Species	Reference
Herbaceous cutting	Bacopa monnieri, Centellaasiatica,Pogostemon patchouli	7, 9, 90
Softwood cutting	Nothapodytesnimmoniana, Pelargonium graveolens, Tylophoraindica	83, 91, 92
Semihard wood cutting	Adhatodavasica,Bixaorellana,Boswellia serrata, Clerodendrumindicum, Crataegusoxyacantha, Embeliaribes, Ginkgo biloba; Jasminum sambac, Leptadenia reticulata, Nothapodytesnimmoniana, Piper longum, Plumbago rosea, Plumabgozeylanica,  Rutagraveolens; Salacia fruticosa	4, 7, 23, 33, 82, 83, 84, 94, 95, 96, 97
Hard wood cutting	Celastruspaniculata, Jasminumgrandiflorum, Lawsoniainermis¸ Nothapodytesnimmoniana,Premnaintegrifolia, Rauwolfia serpentina, Streblus asper, Rosa damascena, Taxusbaccata, Vitex negundo, Wrightiatinctoria	20, 83,97, 93, 99, 100, 101, 102
Terminal cutting	Baliospermummontanum, Bursera delpichiana, Coleus forskohlii, Lippiajavanica	32, 94, 95, 103
Root cuttings	Hemidesmus indicus, Chlorophytumboriviliamum, Rauwolfia serpentina, Rubiacordifolia	94

 

Length of cuttings

Went³⁰ observed that functional buds present on the cutting produced root promotingchemicals, which signifies the importance of length of cutting in inducing rooting. The length that could produce sufficient root and shoot system for the plant to grow however varies with the species. For example, about 15 cm long cuttings were rated superior in Plumbago zeylanicaunder mist,³¹ while longer cuttings of 25 cm were required in case of Bursera delpichiana³² and Rutagraveolens to obtain maximum rooting percentage, fresh root weight and higher field survival.³³ Superior rooting percentage with better survival has been reported using 12-15 cm long cuttings in Rosa damascena³⁴ and Drymis brasiliensis³⁵ under polyhouse conditions.

Availability of mother stock is a limiting factor in a number of species and efforts are beingmade to increase the multiplication ratio to make it cost efficient as well. For instance, in noni (Morindacitrifolia), earlier reports³⁶ recommended use of longer (20 to 40 cm) hardwood cuttings; however, subsequent reports suggested that 4 node³⁷ or even 3 node³⁸ cuttings could suffice development of healthy plantlets. Similarly, in case of black pepper (Piper nigrum), Singh and Singh³⁹obtained maximum rooting and subsequent plant development using two node cuttings than that from three and four node cuttings. However, recently, single node cuttings have been found to increase the multiplication ratio by manifolds.⁴⁰ In a related species, long pepper (Piper longum), Kempe Gowda et al.⁴¹ reported that triple node cuttings treated with IBA 1000 mg/l produced better shoots and roots. Efforts could be made to propagate long pepper using single node cuttings as in black pepper. However, reduction in length of cuttings is not always useful⁴² as significantly higher sprouting percentage with better shoot and root growth were observed, when 15 cm cuttings were used over 7.5 cm in stevia (Stevia rebaudiana). Similarly, four nodal cuttings werefound to be superior to two nodal cuttings for increasing sprouting and survival in Embeliatsjeriam- cottam and Caesalpinia bonduc.³ In Piper sarmentosum, use of double node cuttings was rated superior to single and triple node cuttings.⁴³

Leaf retention on cuttings

Retention of leaves on the stem cutting is known to have an impact on the rooting success in some species, mainly because of the presence of auxins in leaves, which are translocated to the base of cuttings for promotion of rhizogenesis.⁴⁴ Leaf is also known to provide energy to the cuttings especially those with limited food reserves.⁴⁵ However, if excessive leaves are present on the cuttings, moisture may get lost from cuttings due to evaporation resulting into dehydration of cuttings.⁴⁶ The requirement for retention of leaves on a cutting is variable, as success varies greatly amongst the species. In some species, cuttings with lesser leaves root better than more leafy ones,⁴⁷ while rooting cannot occur in other species in the absence of leaves.⁴⁸ According to Okoro and Grace,⁴⁹ the initial carbohydrate reserve in leafless hardwood cuttings may not act as a limiting factor for rootinduction but may govern the subsequent growth and development of leaves.

In case of Pipersarmentosum,  retention  of  leaves  had  no  positive  influence  on  rooting  percentage  butimproved plant growth.⁴³ Limited efforts have been made in MAPs to study the effect ofthis factor on success of propagation (Table 2).A  comparison  was  made  between  half-leaf  and  full-leaf  stem  cuttings  in  Enantiachloranthausing different auxins. Results revealed that full-leaf cuttings were superior to half-leaf cuttings as 100% rooting was noticed in those cuttings even without applicationof auxins.⁵⁰ In another study on Lavandula dentata, cuttings were planted with 1/3, 1/2 or2/3 of their leaves. It was noticed that maintaining higher number of leaves was beneficialand 2/3 leaf retention was found to be most suitable for obtaining better rooting percentage.⁵¹ In Salacia oblonga, highest regeneration percentage (72.3%) was observed, when leafless cuttings were used as against cuttings with leaves (55.5%) at 300 ppm IBA. ⁶

Table 2: Influence of leaf retention on success of propagation through stem cuttings.

Species	Remarks	Reference
Barleriaprionitis	Better survival of 81.48% was noticed in leaved cuttings than leafless cuttings (70.37%)	104
Commiphorawightii	Leafless stem cuttings are preferred for early and better rooting	94
Dilleniasuffructicosa	Juvenile cuttings with leaves gave 100% survival as against 100% mortality in leafless cuttings.	45
Enantiachlorantha	Full-leaf cuttings were superior (100% rooting) to half-leaf cuttings even without application of auxins	50
Lavandula dentata	Maintaining higher number of leaves (2/3 leaf retention) was beneficial for obtaining better rooting percentage	51
Marsdeniatenacissima	Fresh and healthy leafy cutting having two or three nodes of axillary buds gives better rooting	94
Pogostemoncablin	Removal of all the leaves was not recommended due to higher mortality percentage	54
Piper sarmentosum	Retention of leaves on the cuttings did not enhance rooting percentage but improved plant growth.	43
Plectranthus barbatus	Removal of all leaves except apical bud gives better rooting	94
Salacia oblonga	Highest regeneration percentage (72.3%) was observed, when leafless cuttings were used as against cuttings with leaves (55.5%) using 300 ppm IBA	6

 

In one case, rooting and successful establishment in patchouli (Pogostemoncablin) was found to be superior with 6 leaves + IBA (2000 mg/l),⁵² while other report suggested useof  2-4 leaves + IBA (150 mg/l) combination.⁵³ These variations could be attributed to thedifferences in the growing environments and the varieties used in these studies. Removal ofall the leaves in patchouli cuttings was not recommended due to higher mortality percentage.⁵⁴

Season and environmental conditions

Season plays an important role onsuccess of different methods of vegetativepropagation including cuttings (Table 3). Day temperature, cloud cover and relative humidity have great influence on the success of rooting, sprouting and growthof propagules.^25,55 These variations largely govern the physiological activities in a plantsystem, including the sugar levels (due to  retention of leaves on the cuttings, which iscommon in evergreen species) and temperature of the substrate.⁵⁵ It has been reported thatwet season with high relative humidity is congenial for rapid callus production and earlyrooting.⁵⁶ In Salacia fruticosa and Embeliaribes, plant propagation by semi-hardwoodcuttings was most successful when cuttings were collected during January-April.^4,23

Table 3: Influence of season and growing environment on success of propagation in MAPs.

Species	Remarks	Reference
Caralluma edulis, Leptadenia reticulata and Tylophoraindica	Aeroponics system was found to be superior in terms of rooting percentage, plant growth and survival than the conventional soil planting	68
Commiphorawightii	Four to five days seasoning of cuttings in the month of August supported better sprouting and field survival (50%)	105
Clerodendrumserratum	Stem cuttings treated with IBA 400 ppm for early sprouting	106
Embeliaribes	Semi-hardwood cuttings collected during January-April were most suitable	23
Ginkgo biloba	Cuttings maintained in  polyhouse (25 °C and 70% RH) gave 56.7% rooting after 6 months as against  two years under open condition	84
Ginkgo biloba	Cuttings taken during July rooted better (90%) than those taken during April (40%) with the same dose of IAA	107
Jasminum sambac, J. auriculatumand J. grandiflorum	Better rooting under mist than in open condition	59, 60
Magnolia fuscata	Propagation under mist reported highest rooting (%) and survival of rooted cuttings	58
Operculinaturpethum	Stem cuttings with two nodes may either be planted directly in the field during monsoon (July) or may be rooted in mist chamber during March-June	93
Pelargonium graveolens	Shade of Putranjeeva roxburghii was equally suitable as shadenet house for rooting	108
Piper longum	Growth and field establishment was improved in greenhouse than those in natural shade	66
Plumbago zeylanica	15 cm long cuttings were rated superior under mist conditionsBasal stem cuttings up to seventh to ninth nodes under mist chamber gives 80-100% rooting success	3193
Salacia fruticosa	Semi-hardwood cuttings collected during January-April were most suitable	4
Stevia rebaudiana	Growth and field establishment was improved in greenhouse than those in natural shade	65
Stevia rebaudiana	Better rooting in autumn season with 50 ppm NAA + 500 ppm IBA, while 2000 ppm NAA +  2000 ppm IBA is required during kharif season	109, 110
Tinosporacordifolia	Obtaining stem cuttings from mother plant during June-July gives better rooting and field survival	94
Vanilla planifolia	Growing cuttings under 50% shade was found to be cost effective	61
Vanilla planifolia	Greenhouse was most suitable for producing vigorous plants, early rooting with highest rooting percentage	64
Zanthoxylumaramatum	Stem cuttings planted in nursery during monsoon (July – August) gives good rooting	94

 

Under open conditions, ideal conditions for propagation cannot be maintained throughout the year; however, manipulation of environment is possible under protected conditions. Kindof structuresusedforraising nursery also governs the success as light penetration, temperature, relative humidity and gaseous composition inside the structure varies with thematerial used. Several workers have reported good results with cuttings under mist. Raines⁵⁷was first to report the use of mist chamber for induction of rooting in stem cuttings.Balakrishna and Bhattacharjee⁵⁸ reported in Magnolia fuscata that shoot tip cuttings withtwo leaves treated with IBA (6000 ppm) gave the highest rooting percentage and survival ofrooted cuttings under mist. Stem cuttings of Jasminumsambac, J.auriculatum and J.grandiflorum were rooted better under mist than in open condition.^59-60 Under costeffective growing conditions, Konedenedo⁶¹ obtained best results with one meter long vanilla (Vanilla planifolia) cuttings with 50% shade.Under  Indian  conditions,  use  of  low  cost  polyhouse  with  ambient  condition  wasrecommended for propagation of MAPs through cuttings as against shade net conditions.⁶² Medium cost greenhouse was also found to be ideal due to maintenance of relatively high temperature and humidity,⁶³ wherein the extent of rooting of MAPs was better (76.3%)than that in shade net (25.0%).

Propagating structuresviz.  greenhouse,  shade  net  and  natural  shade  of  Muntingia calabura were evaluated for propagation of vanilla (Vanilla planifolia), which revealed that greenhouse was most suitable for producing vigorous plants, early rooting and maximumrooting percentage.⁶⁴ Similarly, rooting, shoot growth and field establishment of stevia (Stevia  rebaudiana)  and  long  pepper  cuttings  were  improved  when  experiments  wereconducted in greenhouse than those in natural shade.^65,66 Higher temperature and relativehumidity prevailing in such structures compared to outside condition could result in enhancedphotosynthetic efficiency due to early sprouting and leaf production, better root growth andestablishment of cuttings.⁶⁷Recently, aeroponics system has become popular in the multiplication of plantingmaterial. In three medicinal species viz. Carallumaedulis,  Leptadenia  reticulata  and Tylophoraindicathe  aeroponics  system  was  found  to  be  superior in terms of rootingpercentage, plant growth and survival than the conventional soil  planting.⁶⁸ Majoradvantage of this system is production of completely disease free planting material, andhence could be exploited on commercial scale.

Influence of rooting media/ substrate

Various characteristics of rooting mediumviz. structure, texture, porosity, chemicalcomposition, water holding ability of the media and pH have pronounced effect on rootingability as well as the quality of root system formed.^18,69-70 Hence, adequate attentionshould be paid, while selecting a medium for the concerned species. In a number of medicinalplant species, such studies are lacking, however spice species having numerous medicinalproperties such as black pepper are adequately studied (Table 4). In  black  pepper  variety  Panniyur-1,  highest  rooting  (30.9%)  and  plant  growth  wereobtained in the medium containing sand + farmyard manure (FYM), followed by sand alone.⁷¹ Bogantes-Arias⁷² tested six substrates as rooting media for black pepper cuttings, wherein lowestmortality with better root growth was observed in soil + Bio-rigi + sawdust (1:1:1) and soil +sand (1:1) combinations, while soil was not recommended. Thankamani et al.,⁷³ reported that use of vermicompost as a substrate supported better plant growth over standard pottingmixture comprising of soil, sand and farmyard manure (3:1:1). Mixture containing topsoil,sand and vermicompost (1:1:1) was found to be the best for root induction and recoverypercentage in black pepper var. Panniyur-1 under rapid multiplication technique that couldproduce 23.5 rooted cuttings per vine within three and a half months.⁷⁴

Table 4: Effect of substrate on rooting, survival and plant growth.

Species	Substrate	Remarks	Reference
Barleriaprionitis	Sand+ soil+ vermicompost (1:1:1)	Better (97%) survival and plant growth	104
Cinnamomumverum var. Navashree and Nityashree	Sand and coir dust (1:1)	Superior rooting parameters	78
Leptadenia reticulata	Sand : FYM : Red soil	Good rooting and field establishment	94
Morindacitrifolia	Forest soil + sand, volcanic cinder + compost	Superior rooting parameters	36
Piper longum	Soil + sand + vermicompost (1:1:1)	Better shoot and root growth parameters	66
Piper nigrum	Soil + Bio-rigi + sawdust (1:1:1) and soil + sand (1:1); while soil was not recommended	Lowest mortality with better root growth	72
Piper nigrum	Vermicompost	Better plant growth	73
Piper nigrum var. Panniyur-1	Topsoil, sand and vermicompost (1:1:1)	Better multiplication ratio (23.5 cuttings per vine) under rapid multiplication technique	74
Pogostemonheyneanus	Top soil: sand: compost (1:1:1)	Better root growth	21
Premnaintegrifolia	Sand : soil : FYM (1:1:1)	Better rooting	94
Salacia reticulata	Top soil and compost (1:1)	Highly congenial for establishment of cuttings	24
Stevia rebaudiana	Sand: perlite (1:3)	Better rooting and growth	75
Stevia rebaudiana	Soil + sand + vermicompost (1:1:1)	Better shoot and root growth parameters	65
Vanilla planifolia	Vermicompost and coir pith compost	Most efficient and cost effective	76

 

Better rooting and growth response have been reported in stevia (Stevia rebaudiana),when stem cuttings were planted in sand: perlite (1:3) media under polythene film.⁷⁵ Inanother study on stevia, use of soil + sand + vermicompost media (1:1:1) was found toproduce longer and thicker sprouts, better root girth, dry weight of shoot and roots.⁶⁵ Samemedium combination was also found to be promising in long pepper (Piper longum) also.⁶⁶ In Pogostemonheyneanus, a combination of top soil, sand and compost (1:1:1) wasfound to be the most suitable potting medium.²¹ In Salacia reticulata, use of top soil andcompost  (1:1)  was  found  to  be  highly  congenial  for  establishment  of  cuttings.²⁴Siddagangaiah et al., ⁷⁶ evaluated various rooting substrates and found that vermicompostand  decomposed  coir  pith  were  most  efficient,  cost  effective  and  hence  ideal  for  theproduction of vanilla stem cuttings. Vermicompost is commonly incorporated in the rootingmedium due to various properties such as its richness in nutrients and growth stimulants,higher water holding capacity and improved soil texture that facilitate the root growth.^27,77 Krishnamoorthy et al.,⁷⁸ reported that sand and coir dust (1:1) was the most appropriaterooting medium for two cinnamon (Cinnamomumverum) varieties viz. Navashreeand Nityashree as highest rooting percentage, length of primary roots and secondary roots percutting were obtained in this combination. Nelson³⁶ recommended weed and nematodefree  forest  soil  mixed  with  sand,  volcanic  cinder  and  composted  organic  matter  forpropagation in noni (Morindacitrifolia).

Growth Regulators

Inherent capability of a cutting to induce rooting could largely be altered by chemicaltreatments. Many synthetic growth substances have been used to aid the rooting of cuttingssince long time.⁷⁹ Of those, auxins are the most commonly used as they are known to help in accumulation of metabolites, synthesis of new proteins, cell enlargement and increasenitrogen in roots.⁸⁰ They also regulate different aspects of plant growth and developmentby affectingphysiological processes including cell division, cellelongationanddifferentiation.⁸¹Indole-3-butyric acid (IBA), naphthaleneacetic acid (NAA) and indole-3-acetic acid (IAA)are the most commonly employed auxins due to their ability to initiate roots, stability and lowmobility in plants. Although the induction of rooting by auxin application has been reportedin different plant species, reports on the mechanisms of this response are contradictory.⁸¹

In Bixaorellana, use of 4000 mg/l concentration of IAA supported root induction in 22%cuttings, whereas none of the IBA concentrations could induce rooting in these cuttings.⁸²It is noticed that the concentration of growth regulators required for rooting varies accordingto nature of the plant and as a thumb rule, woody plants require higher concentration than theherbaceous ones. The optimum kind and concentration of auxin are species-specific e.g. adose of 2000 mg/l of IBA is recommended for Jasminum grandiflorum, whereas double doseof 4000 mg/l was required in case of Jasminum auriculatum.⁶⁰ Hormonal requirement also differs with the type of cutting used for propagation e.g. in Nothapodytesnimmoniana, IBA concentrations of 2000 mg/l, 3000 mg/l and 4000 mg/l were optimum for soft wood, semihardwood and hardwood cuttings, respectively.⁸³ Hence, a number of other factors need tobe studied together with growth regulators to get comprehensive results.

Unlike other aspects discussed in this article, role of plant growth regulators have beenstudied in multiplication of a number of MAPs (Table 5). In a number of instances, rootingcould not be obtained in the absence of external application of growth hormones, therebyindicating their pivotal role in the propagation of these species. Apart from auxins,compounds such as catechin 5 mg/L and gallic acid 10 mg/L have been reported to inducerooting in semi hardwood cuttings of Ginkgo biloba with 53 and 57% success.⁸⁴

Table 5: Interactions between type of cuttings and different growth regulators in propagation of MAPs.

Species	Type of cutting	Growth regulators	Remarks	Reference
Adhatodavasica	SHWC	1500 ppm IBA	Higher (67.5%) rooting than control (30.8%)		96
Azadirachtaindica	SC	1000 ppm IBA	Superior (51%) rooting than control (35%)		112
Baliospermummontanum	SHWC	IBA, NAA, Boric acid	Improved sprouting percentage to nearly 90 to 100 %		94
Berberis aristata	TC	5000 ppm IBA	Significantly higher (50%) rooting as against no rooting in control		98
Bursera delpichiana	HWC	2000 ppm IBA	Better rooting (67%) than control (15.3%)		32
Caesalpiniabonduc	SC	1000 ppm IBA	Superior sprouting and field survival		3
Celastruspaniculata	HWC	2000 ppm IBA	Improved (72.6%) rooting percentage as against 54% in control		102
Caralluma edulis	SC	2000 ppm IBA	100 % root induction in hydroponics system		68
Coleus forskohlii	TC	500 ppm IAA	Superior rooting response		103
Commiphorawightii	SC	100 ppm IBA	Hasten rooting and better survival		94
Embeliaribes	SHWC	3000 ppm IBA	100 percent rooting		23
Embeliatsjerium-kottam	SC	1000 ppm NAA	Superior sprouting and Survival		3
Glycyrrhiza glabra	SC	500 ppm IBA	90% rooting as compared to 67% in control		113
Gymnemasylvestre	BC	500 ppm IAA 100 PPM IBA	Superior rooting, root length and survival		94, 95
Jasminum auriculatum	SC	4000 ppm IBA	Better rooting percentage (13.3%) than control (6.7%)		60
Jasminum grandiflorum	SC	2000 ppm IBA	Better rooting percentage (50%)		60
Jasminum sambac	SC	2000 ppm IBA	Better rooting and survival		114
Lavendula angustifolia	BC	2000 ppm IBA	Superior rooting percentage		111
Leptadenia reticulata	SC	3000 ppm IBA	Better root induction (97.7 %) in hydroponics system
Lippiajavanica	BC and TC	3000 ppm IBA	Better rooting percentage		103
Magnolia fuscata	SC	6000 ppm IBA	Superior rooting and survival under mist		58
Marsdeniatenacissima	SC	100 ppm IBA	Better rooting		94
Nothapodytesnimmoniana	1. SWC 2. SHWC 3. HWC	1. 2000 ppm IBA 2. 3000 ppm IBA 3. 4000 ppm IBA	Superior rooting, survival and growth parameters		83
Pelargonium graveolens	SC	2000 ppm IBA	Better rooting percentage and field survival		92
Picrorhizakurroa	Runners	50 µM IBA for 12 h	Superior rooting (87%) than control (53%)		115
Piper longum	BC	500 ppm IAA	Superior rooting percentage, root length and survival		95
Piper longum	SC	500 ppm IBA	Better rooting and field survival		116
Piper nigrum	NC	2000 ppm IBA	Superior root growth parameters		117
Piper sarmentosum	SWC	1000 mg/l	Superior rooting (92.5%)		43
Plumbago zeylanica	SC	500 ppm NAA	Promote quick rooting		94
Pogostemon patchouli	LHC	1500 ppm IBA	Superior rooting percentage and survival		90
Pogostemon patchouli	SC	IBA and NAA	4-6 folds increased rooting over control		118
Premnaintegrifolia	HWC	1000 ppm IBA	Superior rooting percentage, growth and field survival		101
Rauwolfiaserpentina	HWC	30 ppm IAA	75-95% success with better rooting		119
Rosa damascena	SC	50 ppm NAA	Maximum rooting and shoot growth		99
Rutagraveolens	SHWC	300 ppm IBA	Maximum rooting and field survival		33
Salacia fruticosa	SHWC	3000 ppm IBA	Superior rooting percentage and root length		4
Taxusbaccata,	HWC	500 ppm IBA	Higher callusing percentage, rooting percentage, number of roots		93
Terminalia arjuna	SC	2000 ppm IBA	Superior rooting parameters, shoot proliferation, maximum shoot and root biomass		120
Terminalia chebula	Juvenile SC	4000 ppm IBA	Better (55%) rooting than control (25%)		121
Tinosporacordifolia	SC	100 ppm IBA	Better rooting and plant growth		122
Thymus vulgaris	SC	150 ppm IBA	Early and better rooting		123
Tylophoraindica	SWC	1000 ppm IBA	Maximum rooting and survival		124
Tylophoraindica	SC	3000 ppm IBA	Better RP (93.3 %) in hydroponics		68
Vitex negundo	SC	3000 ppm IBA	100% sprouting		125
Wrightiatinctoria	HWC	4000 ppm IBA	Higher root induction		100

(BC: basal cuttings; HWC: hard wood cuttings; LHC: leafy herbaceous cuttings; NC: nodal cuttings; SC: stem cuttings; SHWC: semi hard wood cuttings; SWC: soft wood cuttings; TC: terminal cuttings)

Role of Microbial inoculants

Microbial inoculation could help in getting healthy and vigorous transplants with well-developed root system. This in turn, helps in reducing the transplant injury and improving field establishment. Further, a number of microbial species are known to offer protection against soil borne nursery diseases. Microorganisms such as Trichoderma, Azotobacter, Azospirillum, Bacillus, Rhizobacteria, Pseudomonas, Phosphate Solubilising Bacteria (PSB),Vesicular Arbuscular Mycorrhiza (VAM) etc. have been used to induce rooting in severalspecies.

The mechanism of action of these species is diverse. Trichoderma strains are known to assist the process of decomposition of plant residues in the soil⁸⁵ and also act as bio-control agent.⁸⁶ Root colonization by Trichoderma spp. enhances root growth and development, crop productivity and the uptake and use of nutrients.⁸⁷ Incorporation of VAM fungus, Glomus fasciculatus into the rooting medium enhanced rooting and increased the plant biomass mainly by increasing concentration of endogenous hormone level in blackpepper cuttings.⁸⁸ Similarly, increased feeder root production and absorptive surface areadue to colonization of Pseudomonas fluorescens has been reported.⁸⁹ Table 2 represents the applications of different microbial strains in nursery propagation of MAPs. It is evidentthat a good number of reports are available on black pepper; however, studies on otherspecies are limited (Table 6).

Table 6: Examples of use of microbial inoculants in propagation of MAPs.

Species	Microbial inoculant	Remarks	Reference
Coleus aromaticus	G. fasciculatum, T. harzianum	Maximum growth, total biomass and nutrients in plants	126
Piper longum	T. viride	Improved plant growth and yield	66
Piper nigrum	Trichoderma viride (1 g/kg)	Superior growth, survival percentage  and lower disease incidence	127
Piper nigrum	Azospirillumsp. and Phosphobacteria	Superior plant growth and biomass	128
Piper nigrum	Trichoderma spp. and VAM	Robust, disease free cuttings	129
Piper nigrum	T. harzianum and Pseudomonas fluorescens	Improved growth, reduced incidence of soil borne diseases	130
Piper nigrum	P. fluorescence or T. harzianum (1g/kg)	Superior plant growth and biomass	131
Stevia rebaudiana	T. viride	Superior growth and field establishment compared to growth regulator treatments	65
Vanilla planifolia	T. harzianum (2 g/polybag)	Better rooting and plant growth than in growth regulators treated cuttings	67

 

Conclusion

In recent years, there has been an ever increasing demand for MAPs from national and international markets leading to over-exploitation of their wild sources, resulting in theirdwindling population in the wild. Hence, there is an urgent need to bring these plants undercultivation  for  which  standardization  of  propagation  techniques  is  of  prime  importance.Though  sexual  propagation  through  seeds  helps  in  maintaining  the  diversity  in  nature,commercial scale exploitation of these species demands uniformity, thereby necessitating thestandardization of an alternative vegetative propagation method. Considering the work donein various species as described in the article, there is tremendous scope for studying variousfactors influencing rooting of stem cuttings in MAPs. Once standardized, it could increase theefficiency of this method for mass multiplication in the MAPs for their cultivation andconservation.

Acknowledgements

Authors are thankful to the staff and officials of Department of Horticulture, University of Agricultural Sciences, Bengaluru for the support provided during the conduct of various studies compiled herein.

Conflict of Interest

Authors declare that there is no conflict of interest.

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Abbreviations

Indole-3-acetic acid (IAA); Indole-3 butyric acid (IBA); Farmyard manure (FYM); Medicinal and aromatic plants (MAPs); Naphthaleneacetic acid (NAA); Vesicular Arbuscular Mycorrhiza (VAM)