Healthy Silkworm Research Articles

COONCARE : TRANSFORMATIVE MONITERING FOR SERICULTURE

Sericulture alludes to the raising silkworms to deliver silk. India is the second biggest producer of silk in delivering 15% of the aggregate silk creation along side china. Sericulture is the base of social, economic, cultural and political progress of India. Temperature and humidity play an main role in the development of healthy silkworms in every stage, especially during the development of larva. Disinfection is one of the reproving parameter to be considered for healthy and successful silkworm rearing. Sericulture is the significant occupation in the country side of India and techniques utilized by the agriculturists are as yet obsolete. Hence there exists the need of using innovation in Sericulture cultivate. Keywords-; IoT; Sensors; Image processing; Sericulture;

Improvement of Silkworm (Bombyx mori L.) Growth and Cocoon Quality Using Nickel Sulfate Supplementation

The silkworm belongs to the order Lepidoptera and the family Bombycidae (Bombyx mori L.). The silkworm industry plays a crucial role in providing employment opportunities in India. This study investigated the effects of nickel sulfate supplementation on the growth and cocoon quality of silkworm (Bombyx mori L.). The experiment was conducted at a silkworm-rearing center in Palayamkottai, India, from December 2022 to January 2023. Silkworms were fed Victory-1 (V1) mulberry leaves soaked in different nickel sulfate solutions (100 ppm, 300 ppm, and 500 ppm). Larval weight, duration, cocoon weight, shell weight, shell ratio, and pupal weight were measured for 10 healthy silkworms with good quality cocoons selected from each treatment group. Compared to the control group, supplementation with 500-ppm nickel sulfate solution significantly improved silkworm growth performance (weight) and economic traits (cocoon weight and quality). GRAPHICAL ABSTRACT

Microbial Community Changes in Silkworms Suspected of Septicemia and Identification of Serratia sp.

Diseases that occur in silkworms include soft rot, hardening disease, digestive diseases, and sepsis. However, research on the causes of bacterial diseases occurring in silkworms and the resulting changes in the microbial community is lacking. Therefore, we examined the morphological characteristics of sepsis and changes in the microbial community between silkworms that exhibit a unique odor and healthy silkworms; thus, we established a relationship between disease-causing microorganisms and sepsis. After producing a 16S rRNA amplicon library for samples showing sepsis, we obtained information on the microbial community present in silkworms using next-generation sequencing. Compared to that in healthy silkworms, in silkworms with sepsis, the abundance of the Firmicutes phylum was significantly reduced, while that of Proteobacteria was increased. Serratia sp. was dominant in silkworms with sepsis. After bacterial isolation, identification, and reinfection through the oral cavity, we confirmed this organism as the disease-causing agent; its mortality rate was 1.8 times higher than that caused by Serratia marcescens. In summary, we identified a new causative bacterium of silkworm sepsis through microbial community analysis and confirmed that the microbial community balance was disrupted by the aberrant proliferation of certain bacteria.

Isolation and characterization of bacteria associated with silkworm gut under antibiotic-treated larval feeding.

The impact of antibiotics on growth, cocoon production was assessed in addition to isolation and characterization of bacteria associated with silkworm gut of infected larvae. Larval rearing was maintained at recommended conditions of temperature and humidity. Silkworm larvae showing abnormal symptoms were collected from the control group and dissected for gut collection. Bacteria were isolated from the gut content by spreading on agar plates and incubated at 37 °C for 48 hrs. Bacterial identification and phylogenetic analysis were carried out by 16S rRNA gene sequencing. The isolated bacteria were subjected to antimicrobial susceptibility test (disc diffusion methods) by using Penicillin (10 µg/mL), Tetracycline (30 µg/mL), Amoxicillin (25 µg/mL), Ampicillin (10 µg/mL), and Erythromycin (15 µg/mL). All isolated strains showed positive results for the catalase test. We isolated and identified bacterial strains (n = 06) from the gut of healthy and diseased silkworm larvae. Based on the 16S rRNA gene sequence, isolated bacteria showed close relation with Serratia, Bacillus, and Pseudomonas spp. Notably, 83.3% of strains were resistant to Penicillin, Tetracycline, Amoxicillin, Ampicillin, and Erythromycin but 16.6% showed antibiotic susceptibility to the above-mentioned commonly used antibiotics. Silkworm larvae fed on penicillin-treated leaves showed significant improvement in larval weight, larval length, and cocoon production. Significantly higher larval weight (6.88g), larval length (5.84cm), and cocoon weight (1.33g) were recorded for larvae fed on leaves treated with penicillin as compared to other antibiotics. Isolated bacterial strains showed close relation with Serratia spp., Bacillus spp. and Pseudomonas spp.

Modulations of Haemolymph Parameters in Fifth Instar Silkworm Bombyx Mori L. Parasitized with Beauveria Bassiana

The study focused on understanding the impact of Beauveria bassiana infection on the haemotological parameters such as pH, specific gravity and rate of pulsation of the economically significant silkworm, Bombyx mori. Silkworms, due to centuries of domestication, have developed sensitivity to environmental conditions and various micro-pathogens affecting their nutrition, metabolism, and physiological aspects of haemolymph. The researchers used the bivoltine double hybrid silkworm race (CSR2 X CSR27) X (CSR6 X CSR 26) for the investigation. The results revealed that the pH levels of the haemolymph on the first day of the fifth instar in healthy silkworms were 6.615, and on the sixth day, they were 6.596. In the experimental batch, there was a drastic reduction from the first day (6.590) to the second day (6.253), which gradually increased until the fourth day (6.426) of the instar and gradually decreased on the fifth (6.398) and sixth days (6.374). A lower level of specific gravity of the haemolymph was noticed on the first day (1.024) of the 5th instar in silkworms inoculated with Beauveria bassiana. Then a gradual increase was observed till the sixth day (1.038) of the 5th instar. Conversely, in the healthy silkworms, there was a gradual decline in the specific gravity of the haemolymph from the first day (1.025) to the sixth day (1.012) of the fifth instar. The pulse rate in silkworms parasitized by Beauveria bassiana showed a significant increase from the first day (35) to the second day (48), followed by a gradual decrease to the sixth day (32) of the instar. But in control silkworms, the heart rate increased gradually from the first day (35) to the fifth day (45) of the instar and then suddenly decreased on the sixth day (38).

Regulation and Characterization of Amylase Enzyme Secretion In the Digestive Tract of <i>Antheraea Assamensis</i> Helfer

Antheraea assamensis Helfer is an endemic non-mulberry lepidopteran species of North East India with high commercial demand for its golden hued silk. Antheraea assamensis suffers from a protozoan disease pebrine which lead to death of the silkworm. In this study, the effect of various factors (starvation, feeding, temperature, pH, Ca ion and larval stage) in the regulation of amylase secretion is compared in healthy and diseased muga silkworm. Moreover, enzyme zymography and purification of amylase from the midgut is also performed. Result shows that, the secretion of amylase in Antheraea assamensis in starved condition, significantly decreases. Refeeding experiments (after 2 days of starvation) suggest that amylase secretion sharply increases due to feeding stimulus. The purified enzyme molecular weight is confirmed as 56 kDa by SDS PAGE. The purification fold of purified enzyme is 34 times higher than the crude enzyme extract.

CA-YOLOv5: Detection model for healthy and diseased silkworms in mixed conditions based on improved YOLOv5

CA-YOLOv5: Detection model for healthy and diseased silkworms in mixed conditions based on improved YOLOv5

Effects of Microbial Transfer during Food-Gut-Feces Circulation on the Health of Bombyx mori.

Change in habitual diet may negatively affect health. The domestic silkworm (Bombyx mori) is an economically important oligophagous insect that feeds on mulberry leaves. The growth, development, and immune-disease resistance of silkworms have declined under artificial dietary conditions. In this study, we used B. mori as a model insect to explore the relationship between changes in diet and balance of intestinal microbes due to its simpler guts compared with those of mammals. We found that artificial diets reduced the intestinal bacterial diversity in silkworms and resulted in a simple intestinal microbial structure. By analyzing the correlations among food, gut, and fecal microbial diversity, we found that an artificial diet was more easily fermented and enriched the lactic acid bacteria in the gut of the silkworms. This diet caused intestinal acidification and microbial imbalance (dysbiosis). When combined with the artificial diet, Enterococcus mundtii, a colonizing opportunistic pathogen, caused dysbiosis and allowed the frequent outbreak of bacterial diseases in the silkworms. This study provides further systematic indicators and technical references for future investigations of the relationship between diet-based environmental changes and intestinal microbial balance. IMPORTANCE The body often appears unwell after habitual dietary changes. The domestic silkworm (Bombyx mori) raised on artificial diets is a good model to explore the relationship between dietary changes and the balance of intestinal microbes. In this study, the food-gut-feces microbial model was established, and some potential key genera that could regulate the balance of intestinal microbiota were screened out. Our findings will provide a reference for future research to further our understanding of healthy silkworm development and may even be useful for similar research on other animals.

INVESTIGATION ON CONVERSION EFFICIENCIES OF MULBERRY LEAVES IN SILKWORM Bombyx mori L. DURING THE PROGRESS OF ENTOMO-PATHOGENIC FUNGUS Beauveria bassiana (Bal) Vuill.

Silkworm Bombyx mori is an economically important monophagous insect which is thoroughly domesticated by providing all the requisites for its growth and survival. Due to domestication over centuries, silkworms have become very gentle and delicate to the environmental conditions and different types of micro-pathological agents. The microorganisms such as virus, bacteria, fungi and protozoa cause diseases such as grasserie, flacherie, muscardine and pebrine which are the major threat to the sericulture industry. Chandrasekaran and Nataraju [1] stated that, out of the total crop loss due to different diseases, 10 to 40 per cent of crop loss occurs due to fungi, Beauveria bassiana in India. Nutrition is the basic unit of silkworm growth and development. Feeding and conversion efficiency of mulberry leaves such as leaf to cocoon ratio, leaf to shell ratio etc., in silkworm is one of the commercially important factors to obtain good dividends. Silkworm acquires its essential nutrients and water from the mulberry leaves it consumes. Various nutritional parameters such as ingestion of mulberry leaf, digestion, Efficiency of Conversion of Ingesta to Larva, Efficiency of Conversion of Digesta to Larva, Efficiency of Conversion of Ingesta to Cocoon, Efficiency of Conversion of Digesta to Cocoon, Efficiency of Conversion of Ingesta to Shell, Efficiency of Conversion of Digesta to Shell, I/g- Cocoon, D/g- Cocoon, I/g-Shell, D/g-Shell were estimated in the present investigation in Beauveria bassiana infected silkworms compared to untreated silkworms to understand the conversion efficiencies in infected silkworms. The results indicated a drop in ingesta, digesta, conversion efficiencies of ingesta and digesta to larva, cocoon and shell in the inoculated silkworms. On the contrary, ingesta and digesta required to produce one gram cocoon and one gram shell was higher in inoculated silkworms than in healthy silkworms. So, there is need to understand the various mechanisms involved and physiological changes occurring in the silkworm due to diseases which may be useful in developing prophylactic measures to overcome the attack of diseases.

INFLUENCE OF Beauveria bassiana INFECTION ON RESPIRATORY AND REPRODUCTIVE PARAMETERS OF SILKWORM Bombyx mori L

The mulberry silkworm, Bombyx mori is highly susceptible to diseases caused by viruses, bacteria, fungi and protozoa. The most virulent and contagious disease is muscardine, which is caused by Beauveria bassiana. The present study is focused to understand the modulations in the oxygen consumption, fecundity, hatching percentage, unfertile eggs and dead egg percentage of silkworms i.e., in bivoltine double hybrid (CSR 2 X CSR 27 and CSR6 X CSR26) infected with Beauveria bassiana. The daily oxygen consumption of healthy batch showed a steady increase in the oxygen consumption from first day (0.615 ml of O2/hr/gm wet weight) to fifth day (0.827 ml of O2/hr/gm wet weight) in the control batch. It is higher on the second day (0.671 ml of O2/hr/gm wet weight) in the infected batch with gradual decline till the sixth day of the fifth instar (0.598 ml of O2/hr/gm wet weight) compared to healthy silkworms. Significant reduction in the number of eggs (595) and hatching percentage (75.8%) was recorded in inoculated silk moths compared to number of eggs laid (695) and hatching percentage (99.1%) in control. Significantly high percentage of unfertilized eggs (16.8%) and dead egg percentage (4.37%) was noticed in the experimental animals compared to unfertilized egg percentage (0.14%) and dead egg percentage (0.72%) in control. So, it is necessary to prevent the disease occurrence, especially during silkworm seed rearing to obtain quality commercial silkworm seed necessary for the progress of the industry. Therefore, there is a dire need to carry out research in basic and applied aspects to develop strategies to control the disease.

Smart Automated Sericulture Based On Image Processing Technique And Embedded System

Sericulture is the process of growing silkworms for the purpose of producing silk. India is the world’s second-largest producer of silk. Sericulture is central to India’s socioeconomic, economic, cultural, and political development. Temperature and humidity are important factors in the growth of healthy silkworms. at all stages, particularly during larval development. We’re utilising the webcam to detect silkworms that aren’t doing well. Image processing aids us in recognising infection, ill health, and non-identical phases of silkworms in our study. The colour change in the silkworm’s body is mostly determined via image processing techniques. It denotes non-identical phases, such as black worms and swallow worms, as well as diseases caused by worms.

Comparison of Gut Microbiota between Midgut of Healthy and Tiger Band Disease Infected Oak Tasar Silkworm, Antheraea proylei J

Antheraea proylei J., an economically significant silkworm in the Northeastern region of India, is exclusively domesticated for Tasar silk production. This silkworm is susceptible to various diseases caused by bacteria, viruses, and fungi, including the recently dreadful viral disease dubbed tiger band disease. This viral infection causes damage to silkworm larvae affecting cocoon production, which causes significant losses to the economy of the silk industry. The gut microbiota plays a crucial

Italy, East Asia and Silk. One hundred years of a relationship (1830–1940)

ABSTRACT During the early nineteenth century Italy became the second biggest world exporter of silk thread. To improve productivity in silk cultivation a number of Chinese and Japanese silk production techniques were tried and when Europe was struck by a ruinous and incurable silkworm disease (1845–1880) to keep domestic silk production running Italian experts went to Inner and East Asia to purchase large quantities of healthy silkworm eggs. Eventually Japanese silkworm eggs were seen as the best choice and exported consecutively to Italy for some twenty years in a row. As well as trade connections, cultural ties between Italy and Japan were developed as well while from the late nineteenth century Italy showed much interest in Chinese silk production too. Two expeditions were sent there, the first 1914 lasted for about one year while the second was from 1932 to 1937 and was headed by an Italian expert who was recruited by the League of Nations to revive and modernize Chinese silk production.

Quality Formation Mechanism of Stiff Silkworm, Bombyx batryticatus Using UPLC-Q-TOF-MS-Based Metabolomics.

Bombyx batryticatus is a well-known animal in traditional Chinese medicine. The aim of the research was to reveal the quality formation mechanism of B. batryticatus and to screen out the characteristic component used for the quality control. The anticonvulsant effects of B. batryticatus with a stiff time of one, five, and nine days (D1, D5 and D9, respectively) and healthy silkworm of the same developmental stage (SW) were determined by animal experiment. The dynamic changes in chemical composition were analyzed using UPLC-Q-TOF-MS-based metabolomics. D5 and D9 B. batryticatus exhibited significant anticonvulsant effects (p < 0.05 and p < 0.01, respectively). Accordingly, principal component analysis (PCA) and partial least squares discrimination analysis (PLS-DA) indicated that the chemical composition of D5 and D9 B. batryticatus changed significantly. The different metabolites mainly consisted of primary metabolites such as lipids and amino acids and secondary metabolites such as flavonoids, beauvericin, and glycolipids. Interestingly, the relative abundance of quercetin-7-O-β-d-4-O-methylglucoside, the characteristic component of B. batryticatus, increased with stiff time and was promised to be used as an index component of quality control. The results expand our understanding of the quality formation mechanism of B. batryticatus. In addition, it highlights the potential of UPLC-Q-TOF-MS-based metabolomics for the quality control purpose of TCMs.

Development and comparison of real-time and conventional PCR tools targeting β-tubulin gene for detection of Nosema infection in silkworms.

Microsporidiosis (Pebrine) caused by the microsporidian parasite is one of the important devastating disease which affect the silk production leading to an unprofitable harvest. Till date ribosomal RNA (rRNA) gene was used as a target for detection of microsporidian species. In this study, we describe conventional and SYBR green based real-time PCR techniques alternatively targeting β-tubulin gene for quantitative detection ofmicrosporidia infecting both the mulberry and non-mulberry silkworms. The modified DNA extraction method followed in our study was found to be easy, economical and could be used for both conventional and real time PCR as template. The real time qPCR revealed the expression of β-tubulin gene in different infected tissues of the silkworm Bombyx mori. The sensitivity of the SYBR green based real time PCR was found to be 100 times more than the conventional PCR and PCR was found more sensitive than the microscopic examination. The developed method did not produce any false positive results with the other silkworm pathogens and healthy silkworm. The data suggest that both the developed PCR methods targeting β-tubulin gene could be used effectively in quarantine process at seed centres for early detection of microsporidian infection in silkworms.

Serine protease Bm-SP142 was differentially expressed in resistant and susceptible Bombyx mori strains, involving in the defence response to viral infection.

Bm-SP142 is a 35 kDa protease in the silkworm, but its exact functions remain unknown. In this study, sequence alignment revealed that the His-Asp-Ser catalytic triad is embedded in the TAAHC-DIAL-GDSGGP sequence motif, establishing Bm-SP142 as a serine protease. Soluble recombinant GST-BmSP142 was expressed and purified, and serine protease activity was confirmed in vitro. RT-qPCR results indicated that Bm-SP142 was mainly expressed in the middle part of the silkworm midgut, and Bm-SP142 transcripts were significantly up-regulated at 24 hours post infection (hpi) in BmBDV-resistant strains (798) inoculated with BmBDV and BmNPV-resistant strains (NB) inoculated with BmNPV, but not in BmBDV-susceptible strains (306). Surprisingly, transcripts were significantly down-regulated at 12 hpi in BmNPV-susceptible strains (HuaBa 35) inoculated with BmNPV, compared with healthy silkworms. Recombinant BmNPV treated with purified Bm-SP142 effectively impaired its ability to infect BmN cells, and Bm-SP142 decreases the efficiency of BmNPV and BmBDV propagation in silkworms. Furthermore, overexpression of Bm-SP142 in BmN cells inhibited viral propagation.

In vitro antibacterial activity and in vivo therapeutic effect of Sesbania grandiflora in bacterial infected silkworms

Context: Antibiotic resistance is a serious problem worldwide. Searching for new potential agents is, therefore, essential. The bark of Sesbania grandiflora (L.) Pers. (Fabaceae) has been used in folk medicine against various diseases. Objective: To investigate the antibacterial activity of S. grandiflora bark and explore the therapeutic effect of the highest potent fraction. Materials and methods: Bacteria and healthy silkworms were exposed to three fractionated extracts (3.1–400 mg/mL) of S. grandiflora bark from hexane (HXF), chloroform (CFF), and ethyl acetate (EAF). The sets of bacteria were incubated at 37 °C while silkworms were kept at 27 °C for 24 h. To evaluate the therapeutic effect, silkworms infected with bacteria were exposed to the extracts (0.5–60 mg/mL) and incubated at 27 °C for 52 h. Qualitative analysis of the most potent extract was done using HPLC. Results: EAF showed the highest activity with MIC against methicillin resistant Staphylococcus aureus (MRSA) and vancomycin resistant enterococci (VRE) of 1.6 and 0.4 mg/mL, respectively, and against Gram-negative Escherichia coli and Pseudomonas aeruginosa of 6.2 and 3.1 mg/mL, respectively. It is nontoxic to silkworms with LC50 >400 mg/mL and has high therapeutic effect on infected silkworms with EC50 of 1.9 mg/mL. EAF consists of at least five major compounds, one of them is gallic acid. The activity of EAF is higher than the sum of individual activities of separated compounds. Discussion and conclusion: These results suggest that EAF is a promising antibacterial extract, suitable for further investigation in rodents infected with drug resistant bacteria.

Characterization and pathogenicity assessment of gut-associated microbes of muga silkworm Antheraea assamensis Helfer (Lepidoptera: Saturniidae)

Antheraea assamensis Helfer (muga silkworm) is an economically important endemic insect species of North Eastern Region of India. The silkworm is often susceptible to infection by pathogenic bacteria, leads to a disease commonly known as flacherie which causes 40% loss per annum to the silk industry. In this study, we have reported isolation, characterization and pathogenicity assessment of gut-associated bacteria of healthy and diseased muga silkworms. Thirty five bacterial isolates were screened from the gut of healthy and diseased silkworms by morphological observation and biochemical tests. 11 and 5 strains from healthy and diseased silkworm respectively were identified by 16S rRNA gene sequencing and analyzed. Pseudomonas aeruginosa (DRK1), Ornithinibacillus bavariensis (DRK2), Achromobacter xylosoxidans (KH3) and Staphylococcus aureus (FLG1) strains were commonly found in healthy as well as diseased larvae whereas, Bacillus thuringiensis (MK1) was found only in diseased larvae. Survivability analysis was performed with the identified strains by injection and oral administration (104CFU/ml). The immune response of the silkworm against the pathogen was also studied by phenoloxidase and lysozyme enzyme activity assay, total and differential hemocyte count and phagocytic activity of hemocytes. It was observed that S. aureus, P. aeruginosa and B. thuringiensis significantly reduced the survivability of silkworm (p<0.001) hence found highly pathogenic. The lethal concentrations (LC50) values of the pathogenic strains were calculated at different time intervals (24, 48, 72 and 96h) within the range from 1.38×102 to 3.63×107CFU/ml. The pathogenic groups demonstrated inhibition of phenoloxidase activity and decreased in total hemocyte count after 48h of infection. However, the lysozyme activity increased significantly in the pathogenic groups compared to the control (p<0.05). Granulocytes and plasmatocytes showed phagocytosis whereas; other types of cells did not show any phagocytic activity. Increasing granulocytes and plasmatocytes counts corroborates the results of phagocytic activity. The present study might be helpful in understanding the disease prognosis and colonization of bacteria causing the disease in muga silkworm.

Effects of BmCPV Infection on Silkworm Bombyx mori Intestinal Bacteria.

The gut microbiota has a crucial role in the growth, development and environmental adaptation in the host insect. The objective of our work was to investigate the microbiota of the healthy silkworm Bombyx mori gut and changes after the infection of B. mori cypovirus (BmCPV). Intestinal contents of the infected and healthy larvae of B. mori of fifth instar were collected at 24, 72 and 144 h post infection with BmCPV. The gut bacteria were analyzed by pyrosequencing of the 16S rRNA gene. 147(135) and 113(103) genera were found in the gut content of the healthy control female (male) larvae and BmCPV-infected female (male) larvae, respectively. In general, the microbial communities in the gut content of healthy larvae were dominated by Enterococcus, Delftia, Pelomonas, Ralstonia and Staphylococcus, however the abundance change of each genus was depended on the developmental stage and gender. Microbial diversity reached minimum at 144 h of fifth instar larvae. The abundance of Enterococcus in the females was substantially lower and the abundance of Delftia, Aurantimonas and Staphylococcus was substantially higher compared to the males. Bacterial diversity in the intestinal contents decreased after post infection with BmCPV, whereas the abundance of both Enterococcus and Staphylococcus which belongs to Gram-positive were increased. Therefore, our findings suggested that observed changes in relative abundance was related to the immune response of silkworm to BmCPV infection. Relevance analysis of plenty of the predominant genera showed the abundance of the Enterococcus genus was in negative correlation with the abundance of the most predominant genera. These results provided insight into the relationship between the gut microbiota and development of the BmCPV-infected silkworm.

Studies on the Pathogenicity of Pebrine Spores Isolated from Ichneumon Fly ( Xanthopimpla Pedator) Infesting Tropical Tasar Silkworm on Healthy Silkworm Larvae

Studies on the Pathogenicity of Pebrine Spores Isolated from Ichneumon Fly ( Xanthopimpla Pedator) Infesting Tropical Tasar Silkworm on Healthy Silkworm Larvae