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Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 International Journal of Current Microbiology and Applied Sciences ISSN: 2319-7706 Volume 10 Number 03 (2021) Journal homepage: http://www.ijcmas.com Original Research Article https://doi.org/10.20546/ijcmas.2021.1003.215 Effect of Natural and Artificial Diets on Growth Parameters of Kolar Gold, Silkworm M. D. Thamidela, A. S. Bagde*, U. B. Hole and P. S. Jadhav Department of Entomology, R.C.S.M. College of Agriculture, Kolhapur, (MS) India *Corresponding author ABSTRACT Keywords Artificial diets, Natural diet, Silkworm rearing, Growth parameters Article Info Accepted: 15 February 2021 Available Online: 10 March 2021 Three types of artificial diets viz., Artificial diet1, Artificial diet2 and Artificial diet3 with different compositions were used for rearing Kolar Gold race of mulberry silkworm, Bombyx mori L. where the diets were fed up to initial three instars and the later two instars were reared on natural diet (untreated leaves). Formulated diets were compared with natural diet and the observations on growth parameters were recorded. The study revealed that amongst the three formulated artificial diets, artificial diet 1 (mulberry leaf powder - 30 g, soybean powder - 28 g, cellulose powder - 15 g, corn starch -6 g, citrate - 3.5 g, salt mixture - 4 g, sucrose - 4 g, agar agar - 7 g, along with vitamin B mixture - 0.40 g, soybean oil - 1.30 g and antibiotic chloramphenicol - 1.00 g and water - 302.0 ml) was found to be promising in almost all the growth parameters of the silkworm under study and selected for further studies. Growth parameters like larval weight and larval length were significantly maximum in silkworms which were fed with artificial diet1, followed by artificial diet3. Larval weight and larval length was maximum in artificial diet 1 with 4.338 g and 7.25 cm respectively, followed by artificial diet 3 with 4.213 g and 7.00 cm respectively in final instar. In artificial diet1, larval duration was extended by 83 hr followed by artificial diet3 in which larval duration was prolonged by 94 hr in comparison with natural diet,. In artificial diet1, moulting period was curtailed by 3 hr. However, in artificial diet3 it was extended by 20 hr in comparison with natural diet. In all the instars flacherie disease occurrence was meager in artificial diet1 with 0.75 per cent, followed by artificial diet3 with 1.00 per cent in initial instar and in later instars, disease occurrence was minimum in artificial diet1 with 2.53 per cent followed by artificial diet3 with 2.55 per cent. Introduction India is the only country that produces all four kinds of silk viz., mulberry, tasar, eri and muga (Anitha, 2011). All these four varieties of silk are produced through commercial rearing of Bombyx mori, Antheraea mylitta, Philosamia ricinii and Antheraea assama, respectively. However the best quality silk is obtained from the cocoons of the larvae of mulberry silkworm and is produced mostly in many states of India accounting for 70.21 per cent (25,348 MT) followed by Eri 19.84 per cent (7,175 MT), Tasar 9.32 per cent (3,370 1725 Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 Materials and Methods diet’s ingredients with 2.8 factor. At 800 C temperature distilled water was heated and required amount of agar agar was added to bring gel formation and all the ingredients were thoroughly mixed and steamed in autoclave for 30 minutes at 1100 C. Soon after the process of steaming before its temperature lowers down to 800 C, the material was poured in pre-sterilized plastic trays that was latter covered with aluminum foil; these test diet trays were placed in deep freezer for about 1 hr and then maintained in normal refrigerated temperature until its utilization. 1 hr. prior to the feeding, test diets were taken out of the refrigerator. Temperature and relative humidity for first three and later two instars were maintained at 30 (+1)0 C and 85 (+5) per cent and 25 (+1)0 C and 75 (+5) per cent, respectively. Last two larval instars were then fed with natural diet (Bhojne, 2013). Artificial diets Method of recording observation Three diets were formulated and were fed to initial three instars and later two instars were fed with natural diet. Ingredients and the quantity of ingredients used to formulate the diets are given below Larval weight (g) MT) and Muga 0.66 per cent (240 MT) of the provisional total raw silk production of 36,152 MT (CSB, 2020). Mulberry leaves are complete diet for silkworm sometimes it is possible that some deficiencies occur due to different reasons. The supplementation of extra nutrients along with mulberry leaves results higher yield because the production of superior quality and quantity of silk depends mainly on nutritional status and healthiness of the larvae (Rahmathulla et al., 2007). Studies on the nutrition experienced real progress due to the introduction and development of artificial diets that enhances nutritional status to obtain good cocoon yield and shell ratio (Zah Cristina and Marghitas, 2011). Weight of single larvae = Weight of 10 larvae (g) --------------------------------Total number of larvae (10) Larval length (cm) Procedure for formulating test diet The procedure used for formulating the test diets of the silkworm consists of a series of steps including measuring the requisite amounts of the feed ingredients, mixing the feed ingredients (namely, dried diet), mixing water with the dried diets and steaming them, pouring them in trays and cooling them in deep freezer and transferring the prepared diet to the rearing rooms whenever required. Young age mulberry leaves of var. Victory 1 were used in powder form as a constituent of the diet. Required amount of ingredients were measured by making dry to green factor. Volume of distilled water added to the dried diet was adjusted by multiplying the weighed Length of the larvae was measured in mm with the help of the vernier caliper at initiation and at the end of each instar when the larvae were in relaxed state taking utmost care not to disturb the larvae. Disease incidence (%) Disease incidence = Total number of diseased larvae ------------------------------------------- X 100 Total number of larvae Moulting duration (hours) Moulting duration was recorded from the time worms started settling for moult up to 50 per cent worms came out of moult. 1726 Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 larval weights than those raised on natural diet. Instar larval duration (hours) Larval duration was recorded from the time worms came out of the moult up to settling for next moult, for each instar up to spinning. Results and Discussion In T4 (natural diet), mean larval weight recorded for first, second, third, fourth and fifth larval instars were as 0.006, 0.058, 0.382, 1.140 and 4.033 g, respectively. In first instar all the remaining three treatments T1, T2 and T3 (0.005 g) recorded same larval weight. In second instar, weight was significantly highest in T4 (0.058 g) which was at par with T1 (0.057 g), followed by T3 (0.055 g). In third instar, highest weight was recorded in T1 (0.383 g) that was found at par with T4 (0.382 g), followed by T3 (0.379 g). In fourth instar, significantly lowest weight was recorded in T2 (1.045 g), while the highest weight was recorded in T1 (1.288 g), followed by T3 (1.205 g). In fifth instar T1 (4.338 g) was found to be superior among all the treatments, next highest weight in order of statistical significance was exhibited in T3 (4.213 g) and T4 (4.033 g) and the lowest weight was recorded in T2 (3.996 g). In initial two instars, larval weight of diets was observed to be lesser in comparison with T4 (natural diet), in third instar it was noticed almost similar in range with T4 (natural diet). However in later two instars fed with leaves, the weight was remarkably higher in T1 in comparison with T4 and other formulated diets. These findings are in harmony with the results of Roychoudhury et al., (1994) found that the larvae reared on artificial diet has recorded accelerated larval growth and weight of the larvae was higher, Rajaram et al., (2012) reported that the larval growth was better in silkworms reared on artificial diet in early instars than those reared on mulberry leaves, Jatuporn and Nijura (2017) reported that silkworms raised on artificial diets had higher Mean larval length for first, second, third, fourth and fifth instars were recorded in T4 (natural diet) as 0.500, 1.352, 1.550, 4.85 and 7.00 cm, respectively. In first instar, larval length was observed to be same in T1, T3 and T4 (0.50 cm). In second instar significantly longer length was recorded in T4 (1.352 cm) which was at par with T1 (1.334 cm), followed by T3 (1.313 cm) and the shorter length was recorded in T2 (0.950 cm). In third instar, significantly more length was observed in T4 (1.550 cm), followed by T1 (1.400 cm) that was found at par with T3 (1.390 cm) and less length was recorded in T2 (1.200 cm). In fourth instar significantly longer length was recorded in T1 (5.00 cm) which was at par with T3 (4.90 cm), followed by T4 (4.85 cm). In fifth instar longer length was recorded in T1 (7.25 cm), followed by T3 and T4 both with same length (7.00 cm) while the shorter length was recorded in T2 (6.50 cm). In all the races of silkworm, larval length in initial two instars was observed to be lesser, in intermediate instar in similar range with natural diet and in later two instars remarkably higher than T4 (natural diet). These findings are in harmony with the results of Roychoudhury et al., (1994) found that the larvae reared on artificial diet has recorded accelerated larval growth, Rajaram et al., (2012) reported that the larval growth was better in case of silkworms reared on artificial diet in early instars than those reared on mulberry leaves, Moise et al., (2020) reported that the length of the larvae grown on artificial diet was remarkably higher than those larvae which were reared on natural diet. Mean larval duration for first, second, third, fourth and fifth instars were found significantly lowest in T4 (natural diet) amongst the treatments recording 72, 84, 96, 1727 Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 114 and 140 hr, respectively. In first instar, duration was significantly prolonged in T2 (131 hr) extended by 59 hr, followed by T3 (124 hr) extended by 52 hr. In second instar, duration was significantly extended by 10 hr in T2 (94 hr) that was at par with T1 (92 hr) extended by 8 hr, and at par with T3 (91 hr) extended by 7 hr. In third instar significantly enhanced larval duration was observed in T2 (120 hr) extended by 24 hr, followed by T3 (114 hr) extended by 18 hr, followed by T1 (112 hr) that was extended by 16 hr. In fourth instar, duration was significantly extended by 16 hr in T2 (130 hr), followed by T3 (124 hr) extended by 10 hr, followed by T1 (120 hr) extended by 6 hr. In fifth instar, duration was significantly extended by 8 hr in T2 (148 hr) that was at par with T3 (147 hr) extended by 7 hr, followed by T1 (144 hr) extended by 4 hr. In initial two instars, larval duration of the treatments was found to be extended as compared to T4 (natural diet) similar trend was observed in third instar. However in later two instars, duration was not much extended or prominently extended in T1 when compared to T4 (natural diet). These findings are in accordance with Mottaghitlab and Pourali (2004) found that larval duration was recorded to be highest in all the artificial diets formulated, Bhojne (2013) found that the larval duration was extended for the larvae reared on semi- synthetic diets in comparison to natural diet, Jatuporn and Nijura (2017) reported that larval periods were extended on artificial diets as compared to those reared on natural diet (Table 1–8). Table.1 Treatment details Treatment T1 T2 T3 T4 Details Artificial diet1 Artificial diet2 Artificial diet3 Natural diet Table.2 Composition of the artificial diet Sr.No Ingredients 1 2 3 4 5 6 7 8 9 10 11 12 13 Mulberry leaf powder (g) Soybean powder (g) Cellulose powder (g) Corn starch (g) Citrate (g) Salt mixture (g) Sucrose (g) Agar agar (g) Ascorbic acid (g) Vitamin B mixture (g) Soybean oil Chloramphenicol (ug) Water (ml) 1728 Artificial diets Diet1 Diet2 Diet3 30.00 30.00 45.00 28.00 20.00 20.00 15.00 25.00 10.00 06.00 06.00 06.00 03.50 03.50 03.50 04.00 04.00 04.00 04.00 02.00 02.00 07.00 07.00 07.00 00.50 00.50 00.50 00.40 00.40 00.40 01.30 01.30 01.30 01.00 01.00 01.00 302.0 302.0 302.0 Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 Table.3 Effect of natural and artificial diets on mean larval weight of Kolar Gold Treatment First Mean + SD 0.005+0.00032 T1 0.005+0.00022 T2 0.005+0.00021 T3 0.006+0.00010 T4 0.00011 SE + 0.00035 C.D at 5% 4.13 C.V. % Mean larval weight(g) Initial instars Later instars Second Third Fourth Fifth Mean + SD Mean + SD Mean + SD Mean + SD 0.057+0.0017 0.383+0.0024 1.288+0.010 4.338+0.025 0.050+0.0027 0.359+0.0085 1.045+0.064 3.996+0.101 0.055+0.0005 0.379+0.0021 1.205+0.021 4.213+0.115 0.058+0.0017 0.382+0.0008 1.140+0.018 4.033+0.052 0.0009 0.0022 0.017 0.040 0.0029 0.0070 0.054 0.125 3.38 1.21 3.01 1.97 Table.4 Effect of natural and artificial diets on mean larval length of Kolar Gold Treatment T1 T2 T3 T4 SE + C.D at 5% C.V. % First Mean + SD 0.500 + 0.008 0.452 + 0.005 0.500 + 0.014 0.500 + 0.008 0.004 0.014 1.93 Mean larval length (cm) Initial instars Later instars Second Third Fourth Fifth Mean + SD Mean + SD Mean + SD Mean + SD 1.334 + 0.011 1.400 + 0.041 5.00 + 0.183 7.25 + 0.041 0.950 + 0.035 1.200 + 0.058 4.50 + 0.041 6.50 + 0.041 1.313 + 0.003 1.390 + 0.013 4.90 + 0.082 7.00 + 0.183 1.352 + 0.003 1.550 + 0.030 4.85 + 0.041 7.00 + 0.183 0.009 0.019 0.052 0.066 0.02 0.059 0.160 0.203 1.50 2.80 2.16 1.90 Table.5 Effect of natural and artificial diets on mean larval duration of Kolar Gold Treatment T1 T2 T3 T4 SE + C.D at 5% C.V. % First Mean + SD 121 + 0.82 131 + 1.15 124 + 2.31 72 + 1.63 0.790 2.44 1.41 Mean larval duration Initial instars Second Third Mean + SD Mean + SD 92 + 1.63 112 + 2.45 94 + 0.82 120 + 1.63 91 + 2.58 114 + 0.82 84 + 1.63 96 + 1.63 0.889 0.866 2.74 2.67 1.97 1.56 1729 (hr) Later instars Fourth Fifth Mean + SD Mean + SD 120 + 1.63 144 + 1.83 130 + 1.41 148 + 1.63 124 + 2.16 147 + 0.82 114 + 2.45 140 + 1.63 0.978 0.763 3.01 2.35 1.60 1.05 Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 Table.6 Effect of natural and artificial diets on mean moulting period of Kolar Gold Treatment T1 T2 T3 T4 SE + C.D at 5% C.V. % First Mean + SD 25 + 1.41 28 + 0.81 27 + 1.08 24 + 1.63 0.637 1.96 4.90 Mean moulting period (hr) Second Third Mean + SD Mean + SD 26 + 0.81 27 + 1.08 30 + 1.25 34 + 1.63 28 + 1.15 28 + 0.50 26 + 0.81 28 + 1.15 0.515 0.581 1.59 1.79 3.73 3.96 Fourth Mean + SD 27 + 0.81 36 + 1.63 28 + 1.15 30 + 1.41 0.645 1.99 4.26 Table.7 Effect of natural and artificial diets on total larval phase of Kolar Gold Treatment T1 T2 T3 T4 Duration of test treatments in comparison with natural diet (hr) Feeding phase Non- feeding phase Larval Extended Curtailed Moulting Extended Curtailed 589.00 83 105 3 623.00 117 128 20 600.00 94 111 3 506.00 108 Table.8 Effect of natural and artificial diets on disease occurrence of Kolar Gold Treatment T1 T2 T3 T4 SE + C.D at 5% C.V. % Disease occurrence (%) Initial three instars Later two instars 0.75 (4.96) 2.53 (09.15) 1.75 (7.60) 4.60 (12.39) 1.00 (5.73) 2.55 (09.19) 1.50 (7.00) 3.01 (10.00) 0.007 0.016 0.02 0.05 1.18 1.01 (Figures in parenthesis are mean arcsine transformed values) In T4 (natural diet), moulting period required for first, second, third and fourth moults was recorded as 24, 26, 28 and 30 hr, respectively. In first moult, the period required for moulting was significantly extended by 4 hr in T2 (28 hr) that was found to be at par with T3 (27 hr) extended by 3 hr wherein T1 (25 hr) was observed to be extended by only 1 hr when compared to natural diet. In second moult, requisite time was extended by 4 hr in T2 (30 hr) followed by T3 (28 hr) extended by 2 hr and T1 reported similar time as that of natural diet (26 hr). In third moult, the requisite time was extended by 6 hr in T2 (34 hr) while T3 (28 hr) recorded similar period as that of natural diet. Wherein, the time period required for moulting in T1 (27 hr) was curtailed by 1 hr. In fourth moult, the 1730 Int.J.Curr.Microbiol.App.Sci (2021) 10(03): 1725-1732 requisite time was extended by 6 hr in T2 (36 hr). It was curtailed by 2 and 3 hr in T3 (28 hr) and T1 (27 hr), respectively. Moulting period was more or less extended in almost all the treatments except in T1 wherein, it was prominently curtailed in last two moults than T4 ( natural diet). The requisite period for moulting was observed to be more or less extended in all the treatments when compared to the natural diet except for T1. These findings are in accordance with Bhojne et al., (2013) reported that larval and moulting period was prolonged in the artificial diet in comparison with natural diet, however the results do not correspond the findings of Roychoudhury et al., (1994) who observed it to be reduced. Larval feeding duration was extended by 117 hr in T2 (623 hr), followed by T3 (600 hr) extended by 94 hr, followed by T1 (589 hr) extended by 83 hr. Moulting period was extended by 20 and 3 hr in T2 and T3, respectively, whereas curtailed by 3 hr in T1 during non-feeding phase. In initial instars, flacherie disease occurrence was significantly lowest in T1 (0.75 %) that was followed by T3 (1.00 %), followed by T4 (1.50 %), however the highest disease occurrence in initial instars was reported in T2 (1.75 %). In later instars, significantly highest disease occurrence was reported in T2 (4.60 %), followed by T4 (3.01 %) that was followed by T3 (2.55 %) and minimum disease occurrence was reported in T1 (2.53 %). The results are in accordance with Matsubara et al., (1989) reported that aseptically prepared diet in bioclean room led to meager disease occurrence, Gomma et al., (1976) reported feeding of the larvae on artificial diet from the beginning of second of second instar had least percentage of mortality, Sannamvong and Quiniones (1990) reported that formulated diet recorded lesser larval mortality in comparison with natural diet. In conclusion, amongst all the three formulated diets of all the races viz., Kolar Gold, FC1 x FC2 and FC2 x FC1; T1 (artificial diet1) exhibited better performance. In the entire instars disease incidence was meager. When compared to the natural diet both larval weight and larval length was found to be lower in initial instars and remarkably higher in later instars. Larval duration was prolonged in silkworms fed on artificial diet. References Anitha, R. (2011). Indian silk industry in the global scenario. International Journal of Multidisciplinary Management Studies, 1(3): 100-110. Bhojne, I. R. (2013). Investigation on economic parameters of Bombyx mori L. as influenced by leaf supplementations and semi-synthetic diets. Ph.D. (Entomology) Thesis, MPKV, Rahuri, pp. 31-37. CSB, (2020). http://www.csb.gov.in/assets/uploads/do cuments/note-on-sericulture.pdf Gomma, A. A., EL- Shaarawy, M. F., Salam, Y. S. and Rizk, M. A. (1976). 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Roychoudhury, N., Basu, R., Shamsuddin, M. and Sen, S. K. (1994). Rearing of Bombyx mori on artificial diet after chawki rearing on leaf. CSR and TI, Berhumpore-742101, West Bengal, India. Sericologia. 34(1): 67-76. Sannamvong. K. And Quiniones, A. C. (1990). Comparative study of natural and artificial diet of Bombyx mori. Inst. Of Graduate studies, Central Luzon State Univ., Munoz, Nueva, (Philippines) CLSU Scientific journal. 11(1), 27-38. Zah Cristina and Marghitas. (2011). Nutrition research on silkworms Bombyx mori L., Analele Universitatii din Oradea, Fascicula: Ecotoxicologies technologii de Industriesi Alimentara, 399-402. How to cite this article: Thamidela, M. D., A. S. Bagde, U. B. Hole and Jadhav, P. S. 2021. Effect of Natural and Artificial Diets on Growth Parameters of Kolar Gold, Silkworm. Int.J.Curr.Microbiol.App.Sci. 10(03): 1725-1732. doi: https://doi.org/10.20546/ijcmas.2021.1003.215 1732