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† Corresponding author 
© 2014 Conscientia Beam. All Rights Reserved. 

STUDIES ON SEASONAL PERFORMANCE OF NEWLY DEVELOPED 

BIVOLTINE SILKWORM (BOMBYX MORI L) HYBRIDS TOLERANT TO 

BMNPV AND EFFECT OF TEMPERATURE ON DISEASE INDUCTION 

 

L.Vijaya Lakshmi1† --- V. Sivaprasad2 --- P. Sujathamma3 
1,3Sri Padmavathi Mahila Viswa Vidyalayam, Tirupathi, Andhra Pradesh, India 

2Central Sericulture Research and Training Institute, Mysore, Karnataka, India 

 

ABSTRACT 

Season and region specific studies of silkworm Bombyx mori L. are of greater importance in identifying 

and understanding the adaptability of silkworm genotypes, which are largely influenced by climatic factors. 

The performance of the newly developed hybrid combinations was evaluated for twelve economic  traits 

during pre-monsoon, monsoon and post-monsoon seasons of the year to understand genotype and 

environment interactions and their stability under fluctuating tropical environmental conditions with 

Bombyx mori Nuclear Polyhedro Virus (BmNPV) inoculation @ 2x106 PIB’s/larva. The data was 

subjected to two way analysis of variance and season-wise relative merit of individual hybrids was 

computed following Multiple Trait Evaluation Index method. The index values indicated that the hybrids 

BNR9 x BNR10 and BNR9 x BNR4 with higher average cumulative EI values were ranked I and II 

and establishing their superiority over multiple traits.  Further the larvae soon after fourth moult were kept 

at room temperature (25ºC), low temperature (5ºC) and high temperature (32ºC) without feed for 6 hours 

and then inoculated perorally with BmNPV.  The newly ecdysed fifth instar larvae when exposed to low 

temperature of 5ºC and starved enhanced larval susceptibility to BmNPV while the hybrids BNR9 x 

BNR10 and BNR9 x BNR4 exhibited higher tolerance to BmNPV at 25ºC and moderate tolerance at 

32ºC. 

Keywords: Silkworm, Bombyx mori nuclear polyhedro virus (BmNPV), Seasonal performance, Tropical environment, 

Temperature effect, Disease induction. 

 

Contribution/ Originality   

This study documents the evaluation of silkworm hybrids under BmNPV inoculated 

conditions during different seasons of the year. The methodology includes the estimation of the 

tolerance of the hybrids to BmNPV during various seasons influencing the physiological activities 

affecting their growth and development as well as the expression of economic characters. 

 

 

Animal Review 
2014 Vol. 1, No. 4, pp. 57-64 
ISSN(e): 2409-6490 
ISSN(p): 2412-3382 
© 2014 Conscientia Beam. All Rights Reserved. 
 

 
 

 



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1. INTRODUCTION 

Silkworm is one of the most important domesticated insects where the growth and 

development is greatly influenced by environmental conditions. Success of silkworm 

breeds/hybrids largely depends on their adaptability to the environment in which it is destined to 

be reared. The biological as well as cocoon-related characters are influenced by ambient 

temperature, rearing seasons, quality mulberry leaf, and genetic constitution of silkworm strains. 

It is a well-established fact that under tropical conditions, unlike polyvoltines, bivoltines are more 

vulnerable to various stress like hot climatic conditions of tropics, poor leaf quality, and improper 

management of silkworm crop during summer that is not conducive for bivoltine rearing for 

technologically and economically poor farmers of India [1-3]. Though the nature of silkworm 

healthiness is unclear, healthy silkworm varieties are important for the stabilization of silkworm 

crops. The use of commercial silkworm hybrids resistant to important silkworm diseases is 

economical and better option particularly in tropical areas. Due to fluctuating climatic conditions, 

inadequate silkworm disease management practices and poor quality mulberry leaf, frequent crop 

losses are witnessed especially due to grasserie disease with the farmers in tropical areas.  

As epidemic of the viral diseases in silkworm is observed to have close link with climatic 

factors such as temperature, humidity, air current, food and the environment, to which the insect 

is exposed also plays an important role in the manifestation of resistance. The rate of disease 

induction is perhaps controlled by the host’s developmental stage, particularly moulting period 

[4], indicating that physiological changes in silkworms may also play an important role in the 

induction of viral diseases. Tropical sericulture beset with wide and sudden fluctuations in the 

environment coupled with poor quality of mulberry and management practices by the farmers 

requires more flexible hybrids with genetic plasticity to buffer these adverse situations. The 

productive commercial hybrids presently used by farmers are susceptible to grasserie disease and 

it is felt that there is an urgent need to develop bivoltine silkworm hybrids resistant to these 

conditions with increased productivity. Most of the small and marginal farmers in India are 

unable to practice complete disease management practices due to their socio-economic conditions 

where these breeds are better option and equipped to survive. In view of the above the present 

study was carried out at Andhra Pradesh State Sericulture Research Institute, Hindupur to 

evaluate the performance of newly developed bivoltine silkworm hybrids in various seasons under 

BmNPV inoculated conditions to identify the most promising combinations for commercial use.  

 

2. MATERIALS AND METHODS 

The performance of ten new hybrid combinations viz., BNR1 x BNR2, BNR1 x BNR6, BNR3 

x BNR6, BNR3 x BNR8, BNR5 x BNR6, BNR7 x BNR2, BNR7 x BNR6, BNR9 x BNR4, BNR9 

x BNR8 and BNR9 x BNR10 were evaluated under BmNPV inoculated conditions in three 

seasons namely, pre-monsoon, monsoon and post-monsoon season were evaluated in three 

seasons namely, pre-monsoon (February - May), monsoon (June - October) and post-monsoon 

(November - January) seasons along with the existing bivoltine hybrids viz., APS45 x APS12 and 

CSR2 x CSR4 as controls.  All the hybrids, their parents along with control breeds and hybrids 



Animal Review, 2014, 1(4): 57-64 

 

 

59 
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were reared under inoculated conditions with BmNPV polyhedra @ 2 x 106 PIB’s /ml in each 

season with three replications of 300 larvae per replication after 3rd moult. The larvae were fed 

with V1 mulberry leaf. The rearings were carried out in Randomized Block Design and data for 

twelve traits namely, fecundity, yield per 10,000 larvae by number, yield per 10,000 larvae by 

weight, survival rate, cocoon weight, cocoon shell weight, cocoon shell ratio and filament length, 

non-breakable filament length, raw silk percentage, denier and neatness was analysed using two 

way analysis of variance [5] for three seasons separately to find out genotype-environment 

interaction. The merit of the individual hybrids and their relative superiority was computed 

following the Multiple Trait Evaluation Index [6]. The combination which recorded highest 

average cumulative evaluation index value was assigned 1st rank and subsequent ranks were 

assigned for the combinations in descending order. Effect of temperature on induction of disease 

in hybrids was studied by exposing inoculated larvae to low temperature (50C), room temperature 

(250C) and high temperature (320C) for 6 hours on first day of fifth instar within 2 hours of out of 

fourth moult.  Ten hybrid combinations along with controls with 25 basic number of larvae in 

three replications were inoculated with BmNPV polyhedra @ 20,000 PIBs/ml (0.25ml/25 larvae).  

After 6 hours all the larvae were kept back at room temperature (250C) and reared under normal 

conditions. The rate of viral multiplication at different temperatures and mortality due to BmNPV 

was recorded up to pupal stage and the data was subjected to statistical analysis of variance. 

 

3. RESULTS 

The data on mean of three seasons pertaining to twelve economic traits of the ten hybrids 

viz., BNR9 x BNR4, BNR1 x BNR6, BNR3 x BNR8, BNR9 x BNR10, BNR5 x BNR6, BNR7 x 

BNR2, BNR1 x BNR2, BNR9 x BNR8, BNR7 x BNR6 and BNR3 x BNR6 with control hybrids, 

APS45 x APS12 and CSR2 x CSR4 under BmNPV inoculated conditions are presented in Table 1. 

Analysis of variance results for hybrids are depicted in Table 2 and evaluation index values for 

each of twelve traits are presented in Table 3. The data related to the effect of temperature on 

disease induction is depicted in Table 4. 

Fecundity (no.): The overall performance for the trait over the three seasons revealed that BNR9 

x BNR4 and BNR5 x BNR6 recorded the maximum (505) and minimum (467) eggs/laying with 

an average of 484 eggs/laying.Cocoon Yield per 10,000 Larvae by Number: The overall 

performance of the hybrids for cocoon yield per 10,000 larvae by number indicate that maximum 

(8787) and minimum (8196) cocoon yield was recorded for BNR9 x BNR7 and BNR1 x BNR2 

respectively. Cocoon yield per 10,000 larvae by weight (kg): Comparison of the newly evolved 

hybrids for overall performance indicate that maximum (16.98 kg) and minimum (13.79 kg) 

cocoon yield was recorded for BNR9 x BNR10 and BNR1 x BNR2 respectively.  

Survival rate (%): Considering the overall performance over the three seasons, BNR9 x BNR10 

recorded the maximum (85.20 %) value followed by BNR9 x BNR4 (83.89 %) and BNR7 x BNR2 

(83.79 %).Cocoon weight (g): Comparison of the newly evolved hybrids over the three seasons 

indicate that maximum (1.812 g) and minimum (1.560 g) values were recorded for BNR9 x 

BNR10 and BNR1 x BNR2 respectively. Cocoon shell weight (g): The overall performance of the 



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hybrids over three seasons together revealed that BNR9 x BNR10 and BNR3 x BNR6 recorded 

the maximum (0.404 g) and minimum (0.333 g) values respectively.  

Cocoon shell ratio (%): The overall performance of the hybrids for the trait indicate that BNR9 x 

BNR10 and BNR3 x BNR6 recorded the maximum (22.30 %) and minimum (21.06 %) values 

respectively. 

Filament length (m): Comparison of the newly evolved hybrids over the three seasons of the year 

expressed maximum (977 m) and minimum (806 m) values for BNR9 x BNR10 and BNR3 x 

BNR6 respectively.  

Non-breakable Filament length (m): Comparison of the newly evolved hybrids over the three 

seasons of the year revealed maximum (866 m) and minimum (694 m) values for BNR9 x BNR10 

and BNR3 x BNR6 and BNR5 x BNR6 respectively.  

Raw Silk Percentage (%): Considering the overall performance over the three seasons, BNR9 x 

BNR10 recorded the maximum (18.09 %) value followed by BNR7 x BNR6 (17.19 %) and BNR3 x 

BNR8 (16.88 %)  

Denier (d): The overall performance of the hybrids over three seasons together revealed that 

BNR7 x BNR6 and BNR7 x BNR6 recorded maximum (2.39) and minimum (2.31) values 

respectively. 

Neatness (p): The overall performance of the hybrids for the trait neatness indicate that BNR7 x 

BNR2 and BNR1 x BNR6 recorded maximum (93 p) and minimum (90 p) values respectively.  

 

Table-1. Mean performance of hybrid combinations 

 
Values represent mean of 3 seasons 
Avg = Average, S.D = Standard Deviation, S.E = Standard Error, C.V = Coefficient of Variation 

 

Table-2. Analysis of variance of hybrid combinations 

 
Values represent mean of 3 seasons 
df = degree of freedom; * Significant (p< 0.05); ** Significant (p< 0.01) 

 

The overall mean performance of the hybrids reveals that BNR9 x BNR10 and BNR9 x 

BNR4  



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recorded higher performance. The ANOVA evaluated for the hybrids revealed highly significant 

differences (p<0.01) between hybrids, seasons and season x hybrid interactions and non-

significant interaction between season x hybrid in neatness.  

Evaluation Index: The relative merit as represented by average cumulative index value 

recorded for the overall performance of the hybrids and presented in descending order showed 

that highest EI value for BNR9 x BNR10  (61.33) followed by BNR9 x BNR4 (567.48), BNR7 x 

BNR6 (56.13), BNR7 x BNR2 (52.88), BNR3 x BNR8 (52.87), BNR3 x BNR6 (50.63), and BNR1 

x BNR6 (50.28) indicate their economic superiority over others including control hybrids APS45 

x APS12 and CSR2 x CSR4  which recorded average cumulative index values of 41.13 and 31.10 

respectively over twelve multiple traits evaluated.  

 

Table-3. Evaluation index of hybrids 

 
Avg. EI= Average Evaluation Index 

 

Effect of Temperature on Disease Induction: The pupation rate was observed 0.00 % when 

the BmNPV inoculated larvae exposed to 50C. The larvae when exposed to 250C highest pupation 

rate was observed in BNR9 x BNR10 (88.00 %) followed by BNR7 x BNR6 (86.67 %) and BNR9 

x BNR4 (85.33 %). The larvae when exposed to high temperature (320C) highest pupation rate 

was observed in BNR9 x BNR10 (54.67 %) followed by BNR7 x BNR6 (52.00 %) and BNR9 x 

BNR4 (50.67 %). The analysis of variance results indicate highly significant (p<0.01) values for 

pupation. 

Table-4. Effect of temperature on disease induction in Hybrids 

 
Values represent mean of three replications 

 

 



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62 
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4. DISCUSSION 

Success of silkworm breeds/hybrids depends largely on their adaptability to the environment 

in which it is destined to be reared as opined by Sato [7]. It is clear from the results, that under 

BmNPV inoculated conditions the survival rate and cocoon characters of newly evolved hybrids 

are significantly superior to that of controls during different seasons of the year. The peer 

analysis of the data on the performance of hybrids in pre-monsoon, monsoon and post-monsoon 

seasons of the year revealed maximum expression of economic traits during favourable post-

monsoon season followed by monsoon and pre-monsoon season. These findings corroborate with 

[8-19]. However, the performance of the hybrids during three different seasons revealed 

marginal differences in the expression of economic traits.  

The higher survival rate and quantitative traits under BmNPV inoculated conditions were 

noticed in the newly evolved hybrids even during unfavourable months. The higher cocoon 

weight observed in the new hybrids, which is positively co-related with the cocoon yield indicates 

higher productivity under diversified environmental and BmNPV inoculated conditions reflecting 

their overall superiority and cocoon crop stability. The overall mean performance of the hybrids 

reveals that BNR9 x BNR10 and BNR9 x BNR4 recorded higher performance. The ANOVA 

evaluated for the hybrids revealed highly significant differences (p<0.01) between hybrids, 

seasons and season x hybrid interactions and non-significant interaction between season x hybrid 

in neatness. Further the evaluation index also confirmed the superiority of BNR9 x BNR10 

(61.33) and BNR9 x BNR4 (57.48).  

Extensive work has been done on the induction of viral diseases. Among them, the important 

physical agents, which induce diseases in silkworms, are temperature and pathogenic virulence. 

Temperature is the most important external physical factor for both silkworm susceptibility and 

multiplication of viruses in the host. Most silkworm varieties have been adapted to rearing at 

250C, which is most suitable for their development.  Accordingly, temperatures much higher or 

lower then 250C tend to act as a stress and increase the larval susceptibility to viral infections 

[4].  Exposure of silkworm larvae to low temperature (50C) before peroral infection with NPV 

enhanced susceptibility to each virus [20]. Many workers were also successful in inducing NPV 

with cold treatment soon after the ecdysis of IV instar [21]. However, in an experiment the virus 

concentration below 103 particles/ml injected into V instar larvae at 25ºC went hidden (occult) 

but were reactivated by cold treatment [22]. It has also been observed that the incidence of 

nuclear polyhedro virus (NPV) was higher in sudden changes from room temperature to low 

temperature, i.e., 5ºC than in gradual changes [23, 24]. The results of the present study on 

disease induction corroborates with the above findings showing that the newly ecdysed fifth 

instar larvae when exposed to low temperature of 5ºC and starved enhanced larval susceptibility 

to BmNPV. Environmental factor such as low temperature would activate the latent virus and 

once the occult virus reaches the infective state, the virus multiplication proceeds, in the same 

manner as in the case of normal/natural infections [25]. Changes in the resistance to the 

infection with NPV was investigated by subjecting 4th an 5th instar larvae to low temperature (50 

and 10°C) and high temperature (330, 350 and 37°C) treatments for various periods of time 



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immediately after ecdysis [26]. In this study, the larvae when exposed to 250C highest pupation 

rate was observed in BNR9 x BNR10 (88.00 %) followed by BNR7 x BNR6 (86.67 %) and BNR9 

x BNR4 (85.33 %) exhibiting higher level of tolerance at optimum temperature. The larvae when 

exposed to high temperature (320C) highest pupation rate was observed in BNR9 x BNR10 (54.67 

%) followed by BNR7 x BNR6 (52.00 %) and BNR9 x BNR4 (50.67 %) expressing moderate 

tolerance showing that increase in temperature will decrease tolerance to BmNPV.   

Based on the mean, index values, stability parameters and temperature effect on disease 

induction the newly evolved hybrids BNR9 × BNR10 and BNR9 × BNR4 tolerant to Bombyx mori 

Nuclear Polyhedro Virus (BmNPV) performed well for most of the characters and found suitable 

to rear throughout the year were recommended for commercial exploitation under tropical 

environmental conditions of Andhra Pradesh. 

 

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