Treatment

Forty five days after pruning of mulberry plants, all the treatment were done by foliar spray @ 10ml/plant (7.2mg/ml concentration) once daily for 15 days with Cina- test solutions  ( Cina MT,  Cina 200C  &  Cina 1000C )  and  control  solution respectively. Treatments were given in such a way that all the leaves of the plants were completely drenched with solutions. During spraying, the soil surface underneath each plant was covered with polyethylene sheet. All the CinaMT, Cina 200C and Cina 1000C treated groups received 10ml/plant test solutions (7.2 mg Cina globules/ml concentration) respectively and other healthy and infected- plant groups similarly received 10ml/plant control solutions (7.2mg- 90% ethanol soaked globules/ml concentration) respectively^7,9,10. We told about healthy ( control) and infected ( control ). These controls were only treated with the solutions made from sugar globules in the alcohol medium (i.e. without medicine Cina ). At forty four days after the last treatment all the parameters of growth and nematode infection were recorded for each group. All the data were used for statistical analysis.

Analysis of residue

Mulberry leaves, collected one day after the last treatment were homogenized in a blender and extracted with ethanol. The residue run in thin layer chromatography plate (TLC) with the standard from the Cina test substances. The test substances were Cina MT,Cina200C and Cina 1000C^7,9.

Rearing of silkworms

The eggs  of a mother moth of the multivoltine ‘Nistari‘ race(Bombyx mori L.), after hatching  ( 91 % hatching rate ) and brushing 1^st stage silk worm larvae in the rearing tray, the larvae were divided into 8 batches ( 80number / batch ) and reared ^4,6,7,19. The larvae of batch 1 were fed with the leaves of healthy (control) plants, those of batch 2 with the leaves of healthy Cina MT treated plants, those of batch 3 with the leaves of healthy Cina 200C treated plants, those of batch 4 with the leaves of healthy Cina 1000C treated plants, those of batch 5 with the leaves of infected  (control) plants, those of batch 6 with the leaves of infected CinaMT treated plants, those of batch 7 with the leaves of infected, Cina 200C treated plants and those of batch 8 with the leaves of infected Cina 1000C treated plants. Fresh leaves were given to the larvae 4 times daily. Mulberry leaves were used for feeding one day after the last treatment with Cina. The larvae were kept inside the rearing chamber at 27±2°C and 70 + 15% RH. The fresh weight of the larvae and that of the leaves served were recorded daily for each batch until the larvae started spinning. The consumption of fresh leaves [ ( Fresh leaves served - Dry leaves residues -  Fresh leaves initially consumed ) X Moisture loss ], number of feeding and number of feeding day to cocoon formation, number of escaping feeding during molting, molting span days and mortality rate were recorded. The fresh silk gland weight ( before start spinning ), starting time to spinning, span of spinning, fresh cocoon weight, fresh shell weight, silk layer ratio (Shell weight / Cocoon weight X 100 ) i.e. SR% and effective rate of rearing ( Number of cocoon harvested / Number of silk worm hatched X 100 ) i.e. ERR% were determined ^6,7,19. For statistical analysis by  t- test, ten mature 5^th instar larvae for fresh silk gland weight and ten cocoons for fresh shell weight were dissected out in each treatment batches including replica of all treatment batches^6,7,19.  All the data from rearing trials were used for statistical analysis ( t- test ).

Results.

Table1 shows the effects of Cina on healthy and Meloidogyne incognita infected mulberry plants in a field trial (P<0.01 by ‘t’- test). All healthy and infected plants treated with Cina showed increased number and surface area of leaves, and higher protein content in leaves and root than infected (control) plants. It is also noted that number, surface area, leaf and root- protein content was significantly higher in all healthy Cina treated plants than healthy (control) plants. In all infected Cina treated plants, the population of root-knot nematodes decreased significantly more in rhizospheric soil as well as in roots than in infected (control) plants. The number of root galls also decreased significantly after Cina treatment. It is observed that Cina had no direct toxic effects on nematodes mortality within the exposure period of 12 hours and it left no toxic residues of Cina in the treated plants^7,9. It is interesting that treatment effects were the most pronounced with Cina 200C.

Table 2 shows the effects of Cina on the healthy and M. incognita infected mulberry plants in a field trial on the feeding, growth and mortality of silkworms (P<0.05 by ‘t’-test). The average consumption of leaves by the 5^th instars, average number of feeding to cocoon formation, average number of feeding day to cocoon formation, average number of escaping – feeding during molting and average molting span days were less for Cina treated plants than for healthy and infected (control) ones. The average mortality rate was nil with all Cina treated plants and 8% with healthy (control) and 30% with infected (control ) ones.  However, the average fresh weight of the 5^th instars larvae were higher with Cina treated and healthy plants than with infected (control) one.

Table 3 shows the commercially increased silk production by effective treatment with Cina on the feeding of healthy and M. incognita infected mulberry leaves in a field trial (P<0.05 by ‘t’-test). The average fresh silk gland weight, average fresh cocoon weight, average fresh shell weight and average shell ratio (  Shell weight / Cocoon weight X 100) were higher with healthy and Cina- treated plants than with infected (control) one. It is interesting that the shell ratios were higher in healthy plants treated with Cina than with the infected Cina treated ones. It is also notable that average starting time to spinning day and average span of spinning day ( i.e. duration of span ) were fewer with the healthy and Cina treated plants than with infected ( control ) ones. Average effective rate of rearing ( ERR%) were higher with all Cina treated groups.

Click here to see the full result tables.

Discussion

The homeopathic drug Cina once again confirms that the cost effective Cina not only reduced root-knot disease but also improved the nutritive value of the treated leaves of infected plants^7,9,10. From this field trial, we once again confirm that Cina also improved the nutritive value of the healthy treated leaves^7,9,10 which directly influences the consumption of leaves, number of feeding and number of feeding day to cocoon formation, and indirectly affects molting stage in the all Cina treated groups from this trial. And due to ill development of healthy and infected  (control) batches larvae took more time to molt which is proved from the number of escaping feeding during molting. Higher nutritive value of treated plants contributed to higher growth of silkworm larvae, silk gland weight, cocoon weight and shell weight which  increased silk production significantly⁷ for commercial purpose. The improved health of the larvae, cocoon weight, silk gland and shell weight from the Cina treated groups of healthy and infected- plants might have resulted in the shortened starting time to spinning and span of spinning day and the total elimination of the mortality rate. However, Cina is too dilute to contain drug molecules¹⁰. Naturally, the drug might not have affected the nematode directly^7,9,10 and for this reason, no mortality occurs. The effective rate of rearing ( ERR% ) is very high in all Cina treated treatment batches which enriches the sericulture industry in many ways, especially for commercial purpose. The mulberry leaves did not contain any toxic residues of the Cina test substances by the thin layer chromatography (TLC). It is reported that Cina at ultra high dilution has physical basis in the form of charge transfer interaction and altered rate of tumbling in the specific part of the molecules of the diluent’s medium^9,10,17. Rather, the drug Cina might have induced natural defense response in the test plants against nematode parasites and has conferred defense response on growing larvae^{5 to 7,9,10,20}.

In fact, it is surprising that all infected Cina treated plants not only are less affected by nematodes, but also have a better growth than healthy (control) plants. It is further confirmed by the Cina treated effects on healthy plants in order to confirm the more positive effect on growth than in healthy (control) groups^7,9,10 . The positive effects of growth may be responsible for defense resistance against pathogens. Cina might have induced synthesis of many new proteins which have stimulated increase photosynthesis rate, stomatal activity and water retention capacity of healthy and Cina treated plants. The positive effects of growth on infected Cina treated plants might not only be responsible for defense resistance to the nematodes pathogen, but also improved growth of silkworm larvae and silk gland weight, cocoon weight, shell weight and effective rate of rearing (ERR%) which increase silk production for commercial purpose. It is proved from the result that silk production is higher in all the healthy Cina treated group than infected Cina treated groups.

Conclusions.

These results suggest that plant diseases might be effectively controlled by the potentized and cost effective homeopathic medicine Cina, at an extremely low dose. It commercially increases silk production which directly enriches the sericulture industry as well as the agriculture sector. Further, it is not only easily available, but also non-phytotoxic and non-pollutant and also conserves our biodiversity.

Acknowledgements.

The work described here has been supported by Rtd. Prof. N.C.Sukul, Dept. of Zoology, Visva-Bharati and Joint Director, Sriniketan Sericultural Composite Unit, Sriniketan, Govt. of West Bengal and Mr. Achintya Mondal, Secretary, Unit of Oriental Association for Education and Research (BIMS, BMS,BIMLS & OIST),West Bengal, India. Lastly, for help in statically analysis we are immensely indebted to Mr.Ashis Mondal, Asst. teacher of Ajodhya High School.

References.

1.Sasser JN, Freckman DW. Pro of 25^th Ann Meet Soc Nemat (abst), Orlando, Florida 1986; 32.

2.Datta SC. Possible use of amaranth as catch crop for root-knot nematodes intercropped   with mulberry. J Environ & Sociobiol 2005b;2(2&3):61-65.

3.Datta SC. Possible use of amaranth as catch crop for root-knot nematodes intercropped  with okra. Phytomorph. 2006b;56(3&4):1-3.

4.Paul A, Sinhababu SP, Sukul NC. Effect of Meloidogyne incognita infected mulberry plants and their treatment with nematicides of feeding silkworms, Bombyx mori L. Sericol 1995; 35:577-581.

5.Datta SC. Plant parasitic nematodes- an agricultural problem and its solution. The Visva-Bharati Quartery 2005a; 11 (3&4): 89-100.

6.Datta SC, Sinhababu SP, Sukul NC. Improved growth of silk worms from effective treatment of mulberry diseases by Acacia auriculiformis extract. Sericol 1997; 37(4): 707-712.

7.Datta SC, Datta (Nag) R. Liquid homeopathic medicine Cina enriching sericulture industry. J Environ & Sociobiol 2006;3(1):55-60.

8.Halfter G. Biodiversity conservation and protected areas in tropical countries. In Biodiver Sci and Dev Towards a New partnership (eds) Dicastri F and Younis T USA: AB Inter in associated with IUBS 1996;212-223.

9.Datta SC. Effects of Cina on root-knot disease of mulberry. Homeopath 2006;95(2):98:102.

10.Sukul NC, Sinhababu SP, Datta SC, et al. Nematotoxic effect of Acacia auriculiformis and Artemisia nilagirica against root-knot nematodes. Allelopath J 2001; 8:65-72.

11.Christie JR, Perry VG. Removing nematodes from soil. Proc Helm Soc Wash 1951; 18:106-108.

12.Sukul NC.  Soil and plant nematodes. West Bengal State Book Board Publisher, 1987: 1-271.

13.Das S, Sukul NC. Biochemical changes of some crop plants due to root-knot nematode infection. Proc Nat Symp of New Dimension in Parasitology, Allahabad, India 1986.

14.Lowry OH, Rossebrough NJ, Farr AR, Randall RJ, Protein measurement with the Folin-phenol reagent. J  Biol Chem 1951; 193: 265-275.

15.Chatterjee A, Sukul NC. Total protein of galled roots as an index of root-knot nematode infestation of lady’s finger plants. Phytopathol 1981; 31: 372-374.

16.Anonymous. The American Homeopathic Pharmacopoeia. 9^th edn, Philadelphia, USA: Boericke and Tafel,1920.

17.Sukul NC. High Dilution pharmacology and Homeopathy. Sukul, A.,Calcutta,1997: 109.

18.Fenner LM. Determination of nematode mortality. Plant  Dis Rep 1962; 46:383.

19.Krishnaswamy S, Narasimhanna MN, Suryanarayana SK, kumararaj S. Manual of sericulture, vol 2 silkworm rearing. Agri Services Bull 15, FAO, Rome,1972.

20.Klessig, Daniel F, Malamy J. The salicylic acid signal in plants. Plant Mol Biol 1994; 26:1439-1458.

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Dr. Subhas Chandra Datta, Researcher & Assistant teacher, C/O- Rajendranath Nag, Bajeprotappur (Katwa Road), Burdwan- 713101,West Bengal, India.