Volume 8, Issue 1, January – 2023 International Journal of Innovative Science and Research Technology

ISSN No:-2456-2165

Natural Occurrence of Pink Pigmented Facultative

Methylotrophic (PPFM) Bacteria on Phyllosphere
of Mulberry (Morus Indica)
V.Suja., K. Vijila
Tamilnadu Agricultural University, Coimbatore

Abstract:- Pink Pigmented Facultative Methylotrophic Theni. In Major Mulberry Cultivating Area Experience
(PPFM) Bacteria Of The Genus Methylobacterium Are Reduce Leaf Yield And Low Quality Leaf During The
Commonly Found In Association With Plants. These Summer Season Due To Water And Heat Stress.
Bacteria Are Characterized By Being Able To Rely On
Methanol As A Sole Carbon And Energy Source. The Current Soil Management Strategies Are Mainly
Ppfms Are Highly Hydrophobic And Strongly Adhesive Dependent On Inorganic Chemical- Based Fertilizers,
On Plant Surfaces And Has The Ability To Produce Which Caused A Serious Threat To Human Health And
Cytokinin And Indole-3 Acetic Acid On The Environment. The Exploitation Of Beneficial Microbes As
Phyllosphere Region Of Mulberry. In Mulberry (Morus A Biofertilizer Has Become Paramount Importance In
Indica) Variety MR2, The Natural Population Of Agriculture Sector For Their Potential Role In Food Safety
PPFM Occupied Approximately 10 Per Cent Of Total And Sustainable Crop Production. The Eco- Friendly
Aerobic Bacteria On Phyllosphere. The Serial Dilution Approaches Inspire A Wide Range Of Application Of
Method Recorded Higher Population Than Leaf Useful Microscopic Organisms Led To Improved Nutrient
Imprinting Method. The PPFM Isolate Uptake, Plant Growth And Plant Tolerance To Various
Methylobacterium Spp. Mi7 Shown Good Growth Rate Abiotic Stresses (Bhardwaj Et Al., 2014). In Recent Years
Under In Vitro Conditions In Ammonium Mineral Salt The Use Of Methylotrophic Bacteria That Have The Ability
Broth. The Culture Entered Into Stationary Phase To Colonize Different Habitats, Has Been Suggested For
After 10 Days With One Per Cent Initial Methanol. Sustainable Agriculture. Methylotrophic Bacteria Are
The Natural Population Of PPFM Varied During The Known To Play A Significant Role In The Biogeochemical
Periods Of September To December And January To Cycle In Soil Ecosystems, Ultimately Fortifying Plants And
April On Mulberry Leaves. Heat And Water Stress Sustainable Agriculture. Methylotrophs Also Improve Air
During Summer Decreased The Natural Ppfms. Quality By Using Volatile Organic Compounds Such As
Dichloromethane, Formaldehyde, Methanol And Formic
Keywords:- PPFM, Methylobacterium, Phyllosphere Acid (Manish Kumar Et Al., 2016). Additionally
Bacteria, Mulberry Plant. Methylotrophs Are Involved In Phosphorous, Nitrogen And
Carbon Cycling Can Help Reduce Global Warming.
I. INTRODUCTION Bacteria Belonging To The Genus Methylobacterium Are
Characterized By Being Able To Rely On Methanol As A
Mulberry (Morus Spp) Is A Fast Growing Deciduous Sole Carbon And Energy Source And By Presenting A More
Wood And Perennial Plant. Mulberry Foliage Is The Only Or Less Intense Pink Reddish Pigmentation (Omer Et Al.,
Food For The Silkworm (Bombyx Mori) And Is Grown 2004). The Ppfms Are Ubiquitous In Nature And Are
Under Varied Climatic Conditions Ranging From Frequently Reported On Various Plant Species, Are A
Temperate To Tropical. Mulberry Leaf Is A Major Substantial Part Of The Aerobic, Heterotrophic Microflora
Economic Component In Sericulture Since The Quality And Of The Surface Of Young Plants. They Are Able To
Quantity Of Leaf Produced Per Unit Area Have A Direct Produce Plant Growth Regulators Such As Cytokinins And
Bearing On Silk Production. India Is The Second Largest Auxin (Nadali Et Al., 2010), Which Affect Plant Growth
Producer Of Mulberry Raw Silk (28708 Metric Ton), Next And Different Physiological Processes. The PPFM Can
To China Accounting More Than 15 Per Cent Global Raw Also, Induce Systemic Resistance Against Disease
Silk Production. In Our Country, Favorable Climatic (Madhaiyan, 2004) And Degrade A Wide Range Of Highly
Condition For Mulberry Cultivation Prevails In Five States, Toxic Compounds And Tolerate Metals (Jahan Et Al.,
Viz., Karnataka, Andhra Pradesh, Tamil Nadu, West 2013).
Bengal, Jammu Kashmir. These States Occupy 97 Per Cent
Of Total Mulberry Cultivation And Contribute 95 Per Cent II. NATURAL POPULATION OF BACTERIA AND
Of Raw Silk Production In India. Tamil Nadu Is Ranked PPFM ON MULBERRY PHYLLOSPHERE
Fourth Place Among The Silk Producing States Of Our
Country. The Annual Silk Production In Tamil Nadu Is In The Present Study The Population Of Aerobic
Around 1200 MT. The Total Area Under Mulberry Bacteria And PPFM Was Determined By Serial Dilution
Cultivation In Tamil Nadu Is Arounds 9491 Acre Technique And Leaf Imprinting Method. The Population
(Anonymous, 2015). In Tamil Nadu Major Mulberry Was Assessed During The Period Of September To
Cultivating Regions Are Krishnagiri, Dharmapuri, Erode, December, 2015. The Bacteria And PPFM Population Were
Thirupur, Coimbatore, Namakkal, Selam, Thirunelveli And Recorded After 35 Days After Pruning (DAP). Forty Five

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Volume 8, Issue 1, January – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165
Days Old Leaves Recorded Highest Bacterial Population Literature On The Origin Of Phyllosphere Ppfms
Of 39.75 X 105 CFU G-1 Dry Weight Of Leaf In Serial Abounds, But It Is Still Debatable. Corpe (1985) Argues
Dilution Method. Lindow Et Al. (2003) Reported That That The Paucity Of Ppfms In The Air Makes It Unlikely
Inhabitants Of The Phyllosphere Are Termed Epiphytes And That The Atmosphere Is A Major Contributor Of The
May Consist Of A Variety Of Bacteria, Yeast And Fungi. Methylotrophs Encountered On Plant Leaves. Holland And
Bacteria Within Phyllosphere May Include Those That Are Polacco (1992) Suggested That Leaf-Inhabiting Ppfms Are
Pathogenic To The Plant, But Also Include Non- Pathogenic Probably Descendants Of Seed-Borne Bacteria Rather Than
Organisms Which Often Play Important Role In Influencing Bacteria From Air, Soil, Or Other Plants. Madhaiyan (2005)
The Immediate Environment And Health Of Host Plant And Has Reported That Ppfms Are Transmitted Mostly Through
May Also Be Involved In More Global Processes Such As Seeds. On Other Hand, Romanovskaya Et Al. (2001) Has
Contributing To The Microbial Community On Reported That Leaves Were Not Colonized After Seed
Phyllosphere Of Vegetable Crops Also Studied (Zhang Et Bacterization Or Soil Application Of A PPFM Strain, And
Al., 2010). Fungi And Archaea Are Known To Colonize Were Colonized Only After Direct Application To The
The Leaves, Bacteria Are Numerically Dominant In The Phyllosphere, Suggesting That Natural Leaf Colonization
Phyllosphere Environment (Andrews And Harris, 2000). Occurred Via Transfer Of Soil Particles. According To
Dwelling Bacteria Can Have Neutral, Negative Or Knief Et Al. (2010), Factors Specific To The Sites From
Positive Influences On Their Host Plants ByServing As Which The Plants Species Were Collected, More Than The
Pathogens Or Preventing Leaf Colonization By Pathogen Plant Species Themselves, Have A Strong Influence On The
(Kishore Et Al., 2005). Microflora Influences The Growth Composition Of The Phyllosphere Methylobacterium
And Induces The Health Of Plants Including Mulberry Community. It Is Found In The Present Study That Ppfms
(Shree And Boraiah, 1987). The PPFM Population Of Were Highly Abundant On Mulberry Leaves And The Ratio
Mulberry Phyllosphere Ranged From 28.16 X 102 (35 Of Ppfms To The Total Bacterial Count Was Also Good,
DAP) To 18.75 X 102 (75 DAP) CFU G-1 Dry Weight Of Comparing To The Literature Available On PPFM
Leaf By Serial Dilution And Plating Technique. The Population On Phyllosphere.
Population Increase In Late Spring Indicates Increasingly
Suitable Condition For Methylobacterium Spp. As The III. RESULTS
Plants Grow And Leaves Age. The Climatic Conditions
Might Also Be A Positive Selective Factor. Natural PPFM The Leaf Samples Of Mulberry Plant (Variety MR2)
Population In Red Clover Was Recorded As 106 No. Of Cultivated In The Mulberry Field Of Tamil Nadu
Cells G-1 FW Of Red Clover Leaves By Omer Et Al. (2004). Agricultural University, Coimbatore During The Period Of
The Mean PPFM Population In Clover Was Reported As September To December, 2015 Was Used For This Study.
3.18 Log CFU G-1 FW And In Wheat The Population Was The Results Of The Bacterial Population Analyzed By The
2.67 Log CFU G-1 FW Of Leaf. It Is Possible That Different Serial Dilution And Plating Method And Leaf Imprintment
Plants Possess Varying Carrying Capacity For PPFM And Method Are Presented In Table 1. The Results Indicated
The Reason For This Is So Far Unknown. Hirano And That There Is A Significant Difference In Bacterial
Upper (1991) Found That Ppfms, Unlike Other Population During The Period Of 35th Day To 75th Days
Phyllosphere-Inhabiting Bacteria, Exhibit Population Levels After Pruning (DAP).
Derived From Single Leaf Sample Similar To The Ones A Low Increase In Population Was Observed Upto 45
Derived From Bulk Samples. As An Ecological Niche, The DAP And A Decline With Age Of The Plant After 45th
Plant Phyllosphere Supports Highly Abundant DAP. The Population Ranged From 31.25 X 105 To 21.25
Methylobacterium Species Of 104-107 CFU Per Leaflet X 105 CFU G-1 Dry Weight Of Leaf. The Maximum
(Austin Et Al., 1978). The Bacterial Genus Population Was Recorded On 45th DAP (39.75 X 105 CFU
Methylobacterium Is A Well-Studied Example Of Pink- G-1 Dry Weight Of Leaf) Followed By 55th DAP (33.16 X
Pigmented Facultative Methylotrophs (Ppfms) That Belong 105 CFU G-1 Dry Weight Of Leaf).
To The Class Α- Proteobacteria, And Use Methanol As
Sole Carbon And Energy Source. These Bacteria Are Not The Population Of Bacteria As Determined By Leaf
Considered To Be Passive Passengers On Plant Leaves, But Imprintment Method Is Shown In Table 1. The Population
Are Known To Stimulate Seed Germination And Plant Of Bacteria Showed Variation From 41.17 To 26.58 CFU
Development, And Contribute Towards The Aroma Of Cm-2 Of Leaf. The Maximum Population Was Recorded In
Strawberry (Bligh Et Al., 1959). Mizuno Et Al. (2012) 45th DAP (51.52 CFU Cm-2 Of Leaf) Followed By 55th
Reported Commercial Green Perilla, Perilla Frutescens DAP (45.75 CFU Cm-2 Of Leaf). The Slight Decline In The
Viridis (Makino), Gave The Highest Counts Of Ppfms (2.0- Population Was Observed From 55th Day To 75th DAP.
4.1 X 107 CFU G-1 Of Leaf) Of All The Commercial The Bacterial Communities In Mulberry Phyllosphere
Vegetable Leaves Tested, Amounting To 15 Per Cent Of Isolated Using Nutrient Agar Media By Both The Method
Total Microbes On The Leaves. The Authors Analyzed The Are Exhibited In Plate 1 And Plate 2. Most Of The Bacterial
Green Vegetables For The Ratio Of PPFM To Total Colonies Appeared On The Nutrient Agar Plates Were
Bacteria. Creamy Coloured And Irregular With Undulate Margin.

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Volume 8, Issue 1, January – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165
Population Of Bacteria
S.No. Days After Serial Dilution And Plating
Pruning Method Leaf Imprinting Method
(CFU G-1 Dry Weight OfLeaf) (CFU Cm-2 Of Leaf)
1 31.25 X 105 41.17
35
2 39.75 X 105 51.52
45
3 33.16 X 105 45.75
55
4 26.12 X 105 33.46
65
5 21.25 X 105 26.58
75
2.68 1.37
SE D
5.84 2.93
CD (P= 0.05)
Table 1: Occurrence Of Bacteria On Mulberry Phyllosphere

35 Days After Pruning 45 Days After Pruning

Plate 1: Appearance Of Bacterial Communities Of Mulberry Phyllosphere On Nutrient Agar Plates- Serial Dilution Method

35 Days After Pruning 45 Days After Pruning

Plate 2: Bacterial Flora Of Mulberry Phyllosphere On Nutrient Agar Plates- Leaf ImprintmentMethod

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Volume 8, Issue 1, January – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165
The PPFM Population In The Phyllosphere Region Of The Population Of PPFM As Determined By Leaf
Mulberry Was Analyzed By Serial Dilution Technique And Imprintment Method Showed Variation Ranging From
Leaf Imprintment Method On Ammonium Mineral Salt 24.26 To 12.38 CFU Cm-2 Of Leaf With Age Of Plants. The
(AMS) Agar Plates And Results Presented In Table 2. In Maximum Population Was Recorded In 55th DAP (36.25
Serial Dilution Technique, Population Sizes Of The PPFM CFU Cm-2 Of Leaf) Followed By 45th DAP (31.61 CFU Cm-
2
On Mulberry Leaf Showed Variation Ranging From 28.16 X Of Leaf). Slight Decline In The Population Was Observed
102 To 18.75 X 102 CFU G-1 Dry Weight Of Leaf. The From 65th Day To 75th DAP. The Serial Dilution Technique
Maximum Population Was Recorded On 45th DAP (34.20 X Yielded High Population Of PPFM Compared To Leaf
102 CFU G-1 Dry Weight Of Leaf) Followed By 55th DAP Imprintment Method (Plate 3 And Plate 4). However The
(29.51 X 102 CFU G-1 Dry Weight Of Leaf). Statistically Stability Of Bacteria And Natural Population Of PPFM Was
There Is A Significant Difference In Population Of PPFM Noted On Mulberry Phyllosphere During 35th To 75th DAP.
On Mulberry Phyllosphere.

Population Of PPFM
S.No. Days After Serial Dilution And Plating
Pruning Method Leaf Imprinting Method
(CFU G-1 Dry Weight OfLeaf) (CFU Cm-2 Of Leaf)

1 28.16 X 102 24.26

35
2 34.20 X 102 31.61
45
3 29.51 X 102 36.25
55
4 23.25 X 102 17.27
65
5 18.75 X 102 12.38
75
2.14 1.24
SE D
4.67 2.66
CD (P=0.05)
Table 2: Population Of Naturally Occurring Pink Pigmented Facultative Methylotrophic (PPFM) Bacteria On Mulberry
Phyllosphere

Plate 3: Appearance Of PPFM Communities Of Mulberry Phyllosphere On AMS Agar Plates-Serial Dilution Method

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Volume 8, Issue 1, January – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165

Plate 4: PPFM Communities Of Mulberry Phyllosphere On AMS Agar Plates- LeafImprintment Method

IV. CONCLUSION [5.] Bligh, E.G. And W.J. Dyer. 1959. A Rapid Method
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