Evaluation of genetic potential of the polyvoltine silkworm (Bombyx mori L.) germplasm and identification of parents for breeding programme

Evaluation of genetic potential of the polyvoltine silkworm ( Bombyx mori L.) germplasm and identification of parents for breeding programme first-author †E-mail:gprao007@rediffmail.com †E-mail:gprao007@rediffmail.com gprao007@rediffmail.com Andhra Pradesh State Sericulture Research and Development, Institute APSSRDI, Kirikera, Hindupur-515 211, India Andhra Pradesh State Sericulture Research and Development, Institute APSSRDI, Kirikera, Hindupur-515 211, India In the present study, polyvoltine germplasm stock of Andhra Pradesh State Sericulture Research and Development, Institute (APSSRDI) was evaluated for its performance based on quantitative and qualitative traits. Twenty-one oval and 10 peanut cocoon shaped lines were reared in different seasons of the year. Since the polyvoltines are non-diapausing, six generations were reared and evaluated for various economically important traits based on evaluation index and sub-ordinate function statistical methods. Ten top ranked lines obtained by using both the methods were identified as potential parental strains. Among oval lines, APM14, APM11, APM18, APMW9, and APM19, and among peanut lines APMD5, APMD1, APMD3, APMD9 and APMD8 were selected as base material. The identified high yielding lines will be used in various breeding programmes as initial parents for the synthesis of superior polyvoltine breeds/hybrids. Polyvoltine Silkworm Germplasm Performance Evaluation INTRODUCTION 1 Silkworm is not only a commercially important insect, it is also found to be an important laboratory tool. It is estimated that more than 3 000 silkworm strains are available all over the world due to various ongoing breeding programmes (Nagaraju, 2002 2002 ; Thangavelu et al., 2003 2003 ). These silkworm varieties include univoltines, bivoltine and polyvoltines. Univoltines and bivoltines are qualitatively and quantitatively superior races whereas polyvoltines are relatively inferior in both the traits but superior in their survival and hardiness. But in tropical countries like India, polyvoltine silkworm strains play important role in the production of silk, since they are well accustomed to the tropical climatic conditions. Hence, maintenance of polyvoltine silkworm genetic resources has become very important for meeting the desired objectives of the breeder for immediate or long-term utilization in silkworm breeding programmes. But, it is necessary to maintain them in their original form for their rational use in different breeding and other research purposes (Mukarjee et al., 1999 1999 ; Basavaraja et al., 2003 2003 ; Thangavelu et al., 2003 2003 ; Yamaguchi, 2003 2003 ). In addition to maintenance, systematic study of resource material is also very important, not only for classification and characterization of varieties but also for the selection of promising parents to initiate various breeding programmes. Evaluation of genetic material also helps in identification of lines with special characters like longer filament length, fine denier, fluoride resistance, disease resistance, etc. (Li et al., 2001 2001 ). Availability of diverse genetic stock, gives ample choice for the breeder in selection of initial parents of his desire. Even half of a good silkworm egg laying from a good genetic stock can potentially transform the sericulture scenario to a greater extent (Chandrashekaraiah and Ramesh Babu, 2003 2003 ). Polyvoltine silkworm gene bank of APSSRDI, Hindupur consists of geographically isolated races and breeds, obtained from indigenous and exotic origin. In the present study, an attempt was made to evaluate and characterize polyvoltine germplasm and inbred lines based on evaluation index and sub-ordinate function statistical methods, frequently used for evaluating breeds/hybrids. Germplasm evaluation was conducted to ascertain the genetic potential of various silkworm lines/strains for commercial exploitation. Since sericulture is practiced in diverse agro-climatic zones, systematic evaluation is needed for the proper utilization of the available lines. The information generated will be useful for future breeding programmes. MATERIALS AND METHODS 1 For the present study, 21 oval cocoon types (viz. APMW12, APM25, APMW10, APMW7, APM11, APMW9, APMW8, APM21, APM24, APMG1, APM27, APM4, APM14, APM3, APM20, APM22, APM28, APM19, APM18, APM26, APM23) and 10-peanut type (APMD1, APMD2, APMD3, APMD4, APMD5, APMD6, APMD7, APMD8, APMD9, APMD10) polyvoltine germplasm lines with genetically varied characters were selected. The lines were reared under standard rearing conditions (Krishnaswami, 1975 1975 ; 1983 1983 ) for six generations in various seasons of the year. The young larvae (1st~3rd instars) were reared at 27~28 °C with 85%~90% relative humidity and the late age larvae (4th and 5th instars) were maintained at 24~26 °C with a relative humidity of 70%~80%. Each breed was maintained in three replications. At the beginning of 4th instar, 300 larvae were counted from each line and retained for further studies. Rearing was carried out under hygienic conditions. At the end of 5th instar, the spinning larvae were collected manually and mounted in plastic collapsible mountages. During the rearing period, larvae and cocoons were assessed for the yield parameters like fecundity, cocoon yield, pupation rate, cocoon weight, shell weight, shell ratio, filament length, raw silk (%), reelability (%) and neatness using the following formulae: Cocoon yield/10 000 larvae=(weight of cocoons obtained (kg))×10000/(larvae retained after 3rd moult (300)); Pupation rate (%)=(No. of good cocoons+(No. of double cocoons×2))×100/(larvae retained after 3 rd moult–uzi infestated cocoons); Cocoon weight (g)=(wt. of 25 male (g)+25 female cocoons (g))/50; Cocoon shell weight (g)=(wt. of 25 male (g)+25 female cocoon shells (g))/50; Cocoon shell ratio (SR, %)=(single cocoon shell weight (g))×100/(single cocoon weight (g)); Silk filament length (m)=revolutions of aprovette×circumference of wheel (m). These data were analyzed further by using the following evaluation index method and sub-ordinate function method. Based on these values obtained, ranks were given for each breed accordingly. Evaluation index method 2 Evaluation index value (EI) for silkworm breed performance was calculated by using the following formula (Mano et al., 1993 1993 ). $$EI = (A - B)/C \times 10 + 50$$ where, A is mean of the particular trait; B is overall mean of particular trait; C is standard deviation; 50 is constant. Sub-ordinate function method 2 Sub-ordinate function is calculated by utilizing the following formula (Gower, 1971 1971 ). $$X_u = (X_i - X_{\min } ) - (X_{\max } - X_{\min } )$$ where, X u is sub-ordinate function; X i is measurement of trait of tested breed; X min is minimum value of the trait among all the tested breeds; X max is maximum value of the trait among all the tested breeds. RESULTS AND DISCUSSION 1 The germplasm lines were broadly divided into two groups based on the shape of the cocoons. Oval lines 2 Among 21 oval shaped (cocoon) lines, larvae of almost all the strains were found plain except the line APM19 that showed larval marks on the dorsal surface of their body. However these lines have produced cocoons of different colours like white (APMW12, APMW10, APMW7, APM11, APMW9, APMW8, APM21, APM24), greenish yellow (APMG1, APM27, APM4, APM14, APM3), yellow (APM20, APM22, APM28), cream colour (APM25) and flesh colour (APM19, APM18, APM26, APM23) cocoons and the cocoon grains were found to be fine to medium and coarse (Table 1 1 ). SL. No Lines Larval markings Cocoon colour Cocoon grains 5 Oval lines 1 APMW12 Plain White Medium 2 APM25 Plain Cream Coarse 3 APM10 Plain White Coarse 4 APM14 Plain Gr. yellow Fine 5 APM11 Plain White Fine 6 APMW9 Plain White Medium 7 APMW8 Plain White Fine 8 APM21 Plain White Coarse 9 APM24 Plain White Medium 10 APMG1 Plain Gr. yellow Medium/fine 11 APM27 Plain Gr. yellow Medium/fine 12 APM4 Plain Gr. yellow Fine 13 APMW7 Plain White Coarse 14 APM3 Plain Gr. yellow Fine 15 APM20 Plain Yellow Medium 16 APM22 Plain Yellow Medium 17 APM28 Plain Yellow Medium 18 APM19 Marked Flesh Coarse 19 APM18 Plain Flesh Fine 20 APM26 Plain Flesh Fine 21 APM23 Plain Flesh Medium 5 Peanut lines 1 APMD1 Plain White Medium 2 APMD2 Plain White Coarse 3 APMD3 Plain White Medium 4 APMD4 Marked White Medium 5 APMD5 Marked Yellow Fine 6 APMD6 Marked Yellow Fine 7 APMD7 Plain Flesh Fine 8 APMD8 Marked Flesh Fine 9 APMD9 Plain Flesh Fine 10 APMD10 Marked Flesh Medium The fecundity (number of eggs laid by a single female silk moth) among oval lines ranged between 438 (APMG1 and APM26) and 567 (APM14) and average fecundity was found to be 474. Lowest cocoon yield (yield/10 000 larvae by weight) was recoded in APMG1 (8.836 kg) and highest was in APM28 (12.261 kg) with an average yield of 10.85 kg. In all the lines, pupation (survival) was recorded more than 80% except in APMG1 (72.55%) and APM11 (76.73%) and the average survival rate was 86.4%. Single cocoon weight was found to be highest in APM26 (1.412 g) and lowest in APMW12 (1.199 g) with an average weight of 1.312 g. Maximum cocoon shell ratio was recorded in APM11 (18.58%), minimum in APM25 (15.60%) while average cocoon shell ratio was 16.27%. Highest silk filament length (m) was recorded in APM11 (996 m), lowest in APM25 (604 m). Raw silk (%) was maximal in APM11 (13.03%) and minimum in APM20 (10.48%). In all the oval lines, silk reelability was observed to be above 70%. Highest neatness was recorded in APM14 (83.2) followed by APMW7 (82.0) and APM25 (81.2) whereas lowest was recorded in APM20 (73.0) followed by APM28 (76.2) and APM26 (77.2) (Table 2 2 ). SL. No. Breed Fecundity (No.) Yield/10000 larvae (kg) Pupation rate (%) Cocoon weight (g) Shell weight (g) Shell ratio (%) Filament length (m) Raw silk (%) Reelability (%) Neatness (P) 1 APMW12 484 10.379 89.63 1.199 19.03 15.69 670 10.63 74 79.2 2 APM25 477 10.347 88.07 1.226 19.28 15.60 604 10.87 76 81.2 3 APM10 479 10.669 90.33 1.258 20.22 15.94 619 10.77 74 79.0 4 APM14 567 11.377 87.90 1.332 22.85 16.91 950 12.22 78 83.2 5 APM11 465 8.840 76.73 1.346 25.28 18.58 996 13.03 76 80.5 6 APMW9 490 11.455 87.67 1.364 22.98 16.63 873 11.37 74 79.3 7 APMW8 476 11.025 89.84 1.276 20.60 15.96 766 10.87 74 79.3 8 APM21 492 10.619 86.75 1.301 21.87 16.60 801 11.52 75 80.4 9 APM24 470 11.316 84.80 1.330 22.67 17.02 968 11.76 75 80.0 10 APMG1 438 8.836 72.55 1.275 21.55 16.85 803 11.62 75 79.8 11 APM27 440 10.836 88.34 1.244 20.15 16.10 665 10.82 73 78.3 12 APM4 451 11.090 89.19 1.282 21.37 16.56 711 11.12 73 78.2 13 APMW7 475 10.642 88.35 1.230 20.23 16.42 839 11.64 77 82.0 14 APM3 477 10.730 86.08 1.328 21.32 15.99 712 10.95 75 79.7 15 APM20 470 11.414 87.20 1.351 21.58 16.09 729 10.48 71 73.0 16 APM22 493 11.180 88.35 1.337 20.97 15.69 789 10.66 74 79.3 17 APM28 441 12.261 91.22 1.387 21.70 15.63 736 10.70 75 76.2 18 APM19 456 11.056 81.74 1.386 21.75 15.67 719 10.62 74 79.2 19 APM18 481 11.231 85.76 1.390 22.63 16.18 783 11.05 75 79.5 20 APM26 438 11.358 84.53 1.412 22.70 15.95 778 11.02 75 77.2 21 APM23 487 11.054 89.21 1.290 20.25 15.65 749 10.81 75 80.3 Mean 473.6 10.843 86.391 1.312 21.48 16.27 777.9 11.17 74.8 79.3 SD 28.1 0.793 4.523 0.060 1.44 0.70 114.0 0.62 1.4 2.1 Min. 438 8.836 72.55 1.199 19.03 15.60 604 10.48 71 73.0 Max. 567 12.261 91.22 1.412 25.28 18.58 996 13.03 78 83.2 Peanut lines (dumbbell lines) 2 Among ten peanut shaped lines studied, the larvae of 5 lines (APMD1, APMD2, APMD3, APMD7 and APMD9) were found to be plain while the remaining five (APMD4, APMD5, APMD6, APMD8 and APMD10) were marked. Four lines (APMD1, APMD2, APMD3 and APMD4) yielded white colour cocoons, 2 lines yielded yellow colour cocoons with remaining 4 lines (APMD7, APMD8, APMD9 and APMD10) found to be flesh colour cocoons. The cocoon grains were found to be fine to medium and coarse (Table 1 1 ). In the peanut lines, low fecundity was recorded in APMD9 (441) followed by APMD1 (443) and high fecundity was found in APMD6 (505) followed by APMD2 (487). Yield (yield/10000 larvae), was low in APMD4 (10.30 kg) and high in APMD2 (11.66 kg) followed by APMD6 (11.64 kg). Almost all the lines showed more than 80% pupation rate. Maximum cocoon weight was recorded in APMD8 (1.368 g) followed by APMD2 (1.358 g) whereas minimum was recorded in APMD7 (1.247 g). Highest cocoon shell ratio was recorded in APMD4 (17.39%) followed by APMD8 (16.99%) whereas lowest was recorded in APMD1 (14.90%), followed by APMD10 (15.28%). Maximum filament length was recorded in APMD6 (905 m), minimum measured in APMD1 (610 m). Raw silk percentage was found to be highest in APMD4 (11.95%), lowest in APMD1 (10.02%). Almost all the stock recorded more than 70% reelability and neatness (Table 3 3 ). SL. No. Breed Fecundity (No.) Yield weight (kg) Pupation rate (%) Cocoon weight (g) Shell weight (g) Shell ratio (%) Filament length (m) Raw silk (%) Reelability (%) Neatness (P) 1 APMD1 443 10.84 89.26 1.270 19.02 14.90 610 10.02 74 79 2 APMD2 487 11.66 88.91 1.358 22.13 16.12 740 11.04 75 78 3 APMD3 470 10.74 84.64 1.309 22.20 16.83 702 11.45 74 79 4 APMD4 451 10.30 87.05 1.268 22.23 17.39 866 11.95 75 80 5 APMD5 454 10.45 83.51 1.324 22.67 16.95 660 11.48 74 79 6 APMD6 505 11.64 89.02 1.345 21.65 16.03 905 10.93 74 79 7 APMD7 454 10.65 87.82 1.247 20.65 16.53 655 11.37 75 80 8 APMD8 460 10.75 81.50 1.368 23.27 16.99 696 11.64 74 79 9 APMD9 441 10.99 88.20 1.278 20.23 15.79 622 10.76 75 80 10 APMD10 445 10.60 86.23 1.275 19.53 15.28 616 10.37 74 79 Mean 461 10.86 86.61 1.304 21.36 16.28 707 11.10 74 79 SD 21 0.46 2.63 0.043 1.42 0.80 103 0.59 0.3 0.5 Min. 441 10.30 81.50 1.247 19.02 14.90 610 10.02 74 78 Max. 505 11.66 89.26 1.368 23.27 17.39 905 11.95 75 80 Analysis of data 2 Based on rearing performance, quantitative traits of lines were assessed on different parameters like fecundity, yield/10000 larvae (kg), pupation rate (%), cocoon weight, cocoon shell weight, shell ratio, filament length, raw silk (%), reelability and neatness using the evaluation index and sub-ordinate function methods and the details are as follows. Among oval lines, as per the evaluation method, the lines APM14 (64.2), APM11 (57.4), APM18 (55.9), APMW9 (54.8), APM19 (52.7), APMG1 (52.3), APM4 (52.1), APMW7 (50.2) showed higher evaluation index values (above 50 EI value) and APM28 (43.5), APMW12 (43.6) showed lower index (Table 4 4 ). Whereas six peanut lines viz. APMD5 (57.6), APMD1 (54.5), APMD3 (53.8), APMD9 (52.7), APMD8 (50.9) and APMD2 (50.1) showed higher performance index (above 50 EI) (Table 5 5 ). 2 SL. No. 2 Breed 2 Evaluation index method 2 Sub-ordinate function method Value Rank Value Rank 1 APM14 64.2 1 7.711 1 2 APM11 57.4 2 6.363 2 3 APM18 55.9 3 6.030 3 4 APMW9 54.8 4 5.757 4 5 APM19 52.7 5 5.156 5 6 APMG1 52.3 6 4.976 7 7 APM4 52.1 7 5.144 6 8 APMW7 50.2 8 4.687 8 9 APM20 49.6 9 4.500 10 10 APM21 49.4 10 4.553 9 11 APM3 49.2 11 4.323 11 12 APM22 48.6 12 4.256 12 13 APMW8 48.3 13 4.177 13 14 APM23 47.3 14 4.011 14 15 APM24 47.0 15 3.953 15 16 APM25 45.8 16 3.414 17 17 APMW10 45.6 17 3.515 16 18 APM26 44.4 18 3.358 18 19 APM27 43.7 19 3.266 19 20 APMW12 43.6 20 2.989 21 21 APM28 43.5 21 3.092 20 2 SL. No. 2 Breed 2 Evaluation index method 2 Subordinate function method Value Rank Value Rank 1 APMD5 57.6 1 7.605 1 2 APMD1 54.5 2 6.626 2 3 APMD3 53.8 3 6.193 3 4 APMD9 52.7 4 5.885 4 5 APMD8 50.9 5 5.156 5 6 APMD2 50.1 6 5.097 6 7 APMD10 47.7 7 4.223 7 8 APMD4 47.2 8 4.170 8 9 APMD6 43.6 9 2.898 9 10 APMD7 42.0 10 2.455 10 The other important method used in line evaluation is sub-ordinate function method (Gower, 1971 1971 ). As per this method, among oval lines, APM14 (7.711), APM11 (6.363) and APM18 (6.030), have shown higher index values whereas APMW12 (2.989) and APM28 (3.092) showed lower index values (Table 4 4 ) and in the case of peanut lines, APMD5 (7.605), APMD1 (6.626), APMD3 (6.193), APMD9 (5.885) and APMD8 (5.156) ranked high (Table 5 5 ). After evaluation by both the statistical methods (evaluation index method and sub-ordinate function method) five oval type lines (APM14, APM11, APM18, APMW9, APM19) and five peanut shaped lines (APMD5, APMD1, APMD3, APMD9, APMD8) that stood within five ranks were identified as potential parents for further breeding programme. Constant efforts are being made to develop productive polyvoltine silkworm hybrids suitable for sericulture in tropical agro-climatic conditions of peninsular India because, more than ninety percent of the raw silk is still coming from polyvoltine silkworm hybrids (polyvoltine×bivoltine cross breeds) only. Therefore, maintenance of polyvoltine resource material and their effective utilization has become very important. Most of the quantitative traits of commercial importance in silkworm are under complicated polygenic control under the influence of the environment. For synthesizing the potential polyvoltine cross breeds, usually, the high yielding traits of bivoltine varieties and fitness traits of polyvoltine strains are hybridized as proper selection of potential and homozygous parents is very important. Effective utilization of selected germplasm also plays an important role in saving the time of the breeder in the synthesis of new hybrids. Keeping the need in view, the germplasm strains have been reared consecutively for several generations and their quantitative traits were evaluated using two reliable statistical methods, i.e. evaluation index method and sub-ordinate function method to assess the performance of the inbred lines. Earlier many breeders (Mano et al., 1993 1993 ; Gower, 1971 1971 ; Ramesh Babu et al., 2001 2001 , Rao et al., 2004 2004 ) analyzed their breeds by adopting the above methods either individually or together. 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