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In India during some past years, the oak tasar cocoon production has drastically reduced due to certain disease in oak tasar silkworms, which has lead to significant mortality to the batches of oak tasar silkworms. The damage caused by this disease decreases cocoon crop production up to 80%. As the disease is transmitted through mother moth, it is very essential to avoid the transmission of disease during the grainage activities. The seed sector is backbone for the development of the silk industry. Quality of oak tasar seed and efficiency of grainage depend on the quality of the cocoons, selection of healthy seed cocoon, effective microscopic examination and production of healthy & quality Dfl's. Since long, many techniques have been identified to produce disease free layings (Dfls). Among them REC, Palampur preferred cellular moth examination technique (CMET) for the production of Dfl's. During the year 2016 to 2018, REC, Palampur used CMET by using small paper box of size lenght-3.5'', width-4.5'' and height-2.5'', in which single female moth was kept for oviposition. After the completion of oviposition, each moth was microscopically examined for identification of any infection such as bacteria, pebrine etc and in case of any infection, Dfl produced was rejected and destroyed instantly. Before the year 2016, mass oviposition was practiced, in which small nylon net was used to keep 10-15 moths together. This resulted to remarkable loss/rejection of the Dfl's, leading to the reduction in Dfl's production. Pooled data of mass examination and cellular examination indicated that, cellular examination resulted with decrease in cocoon-Dfl ratio (from 13.64:1 to 4.36:1), decrease in rejection of Dfl's after microscopic examination (from 51.74% to 4.54%) and increasing trend in grainage from 48.26% to 95.21%. This trend clearly indicated that, cellular examination of egg layings is very much encouraging for production of oak tasar seeds and successful grainage activities. [ABSTRACT FROM AUTHOR]

Tropical tasar silkworm is a polyphagous silk spinning insect, endemic to India. The silkworms are commercially reared on field-raised host plants, which are geographically contiguous with the wild silkworm fauna. Ecologically, the silkworm is r-selected and evolved with complex survival strategies. Naturally, the population regulated by the interactions between host and natural enemies and their dynamics due to tri -trophic interactions. Hence, understanding the effect of biotic and abiotic factors on silkworm is most critical for increasing the productivity of the cocoons. The cocoon yield trends at different locations are an indication for local specific problems and requirements for specific technological interventions. Cocoon yield obtained at different locations during 2015 to 2017 was analysed for variation in cocoon yield. Further, predator dynamics and incidence of silkworm diseases were also measured to know their impact on silkworms. The results revealed that the highest cocoon yield was recorded in Chhattisgarh (59.97) and West Bengal (54.57) and lowest was in Maharashtra (34.92) and Madhya Pradesh (37.80). Average cocoon yield in Jharkhand and Odisha was 40.18 and 43.30, respectively. Based on the tukeys HSD test, the highest cocoon yield per dfl was recorded at Bilaspur and Pali. The next highest was recorded in Sundergarh, Boirdadar, Patelnaga and Bastar. The significant changes in the predator's population were recorded before and after brushing of the silkworm. Highest incidence was recorded during 1st, 2nd and 3rd instar silkworm and reduced in the subsequent stages. Correlation studies indicated that there was a strong positive linear relationship between silkworm mortality and predator population. Similarly, the silkworm mortality due to disease-causing pathogens was 53.70 %, 26.75 % and 10.90 % during the first crop and 29.27%, 12.10 % and 9.70 % during the second crop in Jharkhand, Madhya Pradesh and Chhattisgarh, respectively. The study showed that the spatial variation in cocoon yield wasinfluenced by the sporadic occurrence of insect-predators and disease-causing organisms at critical stages of rearing. [ABSTRACT FROM AUTHOR]

Sericulture, often known as silk farming is the practice of rearing silkworms for production of raw silk. One of the major impediments faced by the sericulture community is the silkworm diseases that cause significant crop losses. In the present study, the efficacy of phototrophic bacteria as feed supplements in improving the economic traits as well as disease resistance in mulberry silkworm, Bombyx morí L. was assessed. A total of three phototrophic bacteria were isolated and identified as belonging to the genera Maríchromatium, Rhodobacter and Rhodopseudomonas based on 16S rRNA gene sequencing analysis. Their morphological, physiological and biochemical characters were studied, grown under ambient conditions and biomass was harvested. Mulberry fortified with the aforementioned phototrophic bacteria in solitude and combinations at varying concentrations (0.5, 1, 2 and 5%) were found to be innocuous to the mulberry silkworm. The best results were observed when mulberry leaf was supplemented with Rhodopseudomonas sp. at 2% concentration which improved survival, cocoon weight, shell weight, shell%, filament length and non-breakable filament length by 5.18, 0.27, 3.86, 3.5, 1.5 and 1.2%, respectively under normal rearing conditions. Under Staphylococcus sp. infected conditions, the aforesaid diet also enhanced survival in silkworms by 16%. On the other hand, the same diet didn't exhibit any discernible influence on survival against fungal, viral, or microsporidian infections in silkworm. [ABSTRACT FROM AUTHOR]

Trichosporon asahii is a pathogenic fungus that causes severe, deep-seated fungal infections in neutropenic patients. Elucidating the infection mechanisms of T. asahii based on genetic studies requires a specific gene-targeting system. Here, we established an efficient gene-targeting system in a highly pathogenic T. asahii strain identified using the silkworm infection model. By comparing the pathogenicity of T. asahii clinical isolates in a silkworm infection model, T. asahii MPU129 was identified as a highly pathogenic strain. Using an Agrobacterium tumefaciens-mediated gene transfer system, we obtained a T. asahii MPU129 mutant lacking the ku70 gene, which encodes the Ku70 protein involved in the non-homologous end-joining repair of DNA double-strand breaks. The ku70 gene-deficient mutant showed higher gene-targeting efficiency than the wild-type strain for constructing a mutant lacking the cnb1 gene, which encodes the beta-subunit of calcineurin. The cnb1 gene-deficient mutant showed reduced pathogenicity against silkworms compared with the parental strain. These results suggest that an efficient gene-targeting system in a highly pathogenic T. asahii strain is a useful tool for elucidating the molecular mechanisms of T. asahii infection. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Trichosporon asahii is a pathogenic fungus that causes deep mycosis in patients with neutropenia. Establishing an experimental animal model for quantitatively evaluating pathogenicity and developing a genetic recombination technology will help to elucidate the infection mechanism of T. asahii and promote the development of antifungal drugs. Here we established a silkworm infection model with a transgenic T. asahii strain expressing eGFP. Injecting T. asahii into silkworms eventually killed the silkworms. Moreover, the administration of antifungal agents, such as amphotericin B, fluconazole, and voriconazole, prolonged the survival time of silkworms infected with T. asahii. A transgenic T. asahii strain expressing eGFP was obtained using a gene recombination method with Agrobacterium tumefaciens. The T. asahii strain expressing eGFP showed hyphal formation in the silkworm hemolymph. Both hyphal growth and the inhibition of hyphal growth by the administration of antifungal agents were quantitatively estimated by monitoring fluorescence. Our findings suggest that a silkworm infection model using T. asahii expressing eGFP is useful for evaluating both the pathogenicity of T. asahii and the efficacy of antifungal drugs. [ABSTRACT FROM AUTHOR]

Trichomalopsis uziae Sureshan and Narendra Kumar (Hymenoptera: Pteromalidae) is a new addition to the already reported parasitoid complex of the tachinid uzi fly, Exorista bombycis (Diptera: Tachinidae), which causes 10-20% reduction to the silkworm (Bombyx mori L.) cocoon production in the states of Karnataka, Andhra Pradesh, and Tamil Nadu. Due to the non-availability of information on its biological characteristics, an attempt has been made in the present investigation to document the parasitism impact of T. uziae female at various densities of its host and reproductive performance of parasitoid at ratios of 1 to 5:5 (Parasitoid: Host) and 1:1 to 50 (P: H). The reproductive performance of the parasitoid was also assessed based on the durations of host exposure for 1 to10 days at a constant P:H ratio of 1:5. The results revealed that there was no perceptible change in the rate of parasitism at different parasitoid densities. However, the progeny production per female was significantly higher (P < 0.01) at a P:H ratio of 1:5 in comparison with 2:5 to 5:5. Further, at a variable host density (from 1 to 50), there was a linear decrease in per cent parasitism with an increase in host density and it was more drastic from a host density in exceed from 15. However, both progeny production and sex ratio increased with an increase in host density, more so from a host density upwards of 15. The host exposure duration and parasitism rate were significantly inferior at 1day exposure, while it was considerably high at other exposure periods. The progeny production was significantly higher at 2 to 5 days of exposure period without much difference in the sex-ratio. [ABSTRACT FROM AUTHOR]

Bombyx mori Nucleopolyhedrovirus (BmNPV), which is a member of the Baculoviridae family, is a significant pathogen of the silkworm. The infection of BmNPV is often lethal and causes about 20% loss of cocoon in the silk industry annually. To explore the effects of different gene inhibition strategies on the replication cycle of baculovirus, we constructed the mutant virus to infect BmN cells directly and further identified ie0, ie1, and gp64 as the essential viral genes of BmNPV. To elucidate the significance of the inhibition effect of different interference strategies, we characterized and constructed the recombinant BmNPV that carried a single or multigene-interfering cassette. The results showed that the inhibition effect of dsie1 on target gene expression, virus titer, and silkworm mortality was significantly better than that of dsie0 and dsgp64. It also showed that the dsie1 interference produced fewer progeny virions and was less lethal, which indicates that ie1 played a more critical role in the BmNPV replication cycle. Furthermore, the inhibitory effect of the virus titer and mortality indicated that the multigene co-interference constructed by the baculovirus expression system was significantly better than the interference of any single-gene (p < 0.05). In summary, the strategy of multigene synergy can achieve the function of continuous interference and provide a new platform for the breeding of silkworm disease resistant. In addition, this strategy improves the various traits of the silkworm. [ABSTRACT FROM AUTHOR]

A silkworm-baculovirus system is particularly effective for producing recombinant proteins, including glycoproteins. However, N -glycan structures in silkworm differ from those in mammals. Glycoproteins in silkworm are secreted as pauci-mannose type N -glycans without sialic acid or galactose residues. Sialic acid on N -glycans plays important roles in protein functions. Therefore, we developed pathways for galactosylation and sialylation in silkworm. Sialylated N -glycans on proteins were successfully produced in silkworm by co-expressing galactosyltransferase and sialyltransferase and providing an external supply of a sialylation-related substrate. α2,3/α2,6 Sialylation to N -glycans was controlled by changing the type of sialyltransferase expressed in silkworm. Furthermore, the co-expression of N -acetylglucosaminyltransferase II facilitated the formation of additional di-sialylated N -glycan structures. Our results provide new information on the control of N -glycosylation in silkworm. [ABSTRACT FROM AUTHOR]

Bombyx mori nucleopolyhedrovirus (BmNPV) is the most important pathogen of Bombyx mori , silkworm and causes severe losses in the silk industry. During the virus infectious cycle, budded virus (BVs) and occlusion-derived virus (ODVs) particles, which have identical genetic content but different phenotypes, are produced. The envelope glycoprotein GP64, specific in BVs, is involved in host cell receptor binding and is sufficient to mediate membrane fusion during the viral entry. However, the host cell factors, interacting with GP64 to mediate BVs infection, are still unknown. In this study, a cDNA library of Bombyx mori cells (BmN) was constructed and yeast two-hybrid screening was used to identify the host cell factors interacting with GP64. One of the eight candidate proteins encoded the E3 ubiquitin-protein ligase SINA-like 10 (SINAL10), was further confirmed through coimmunoprecipitation assays as novel GP64 binding protein. Moreover, overexpression of SINAL10 significantly enhances viral reproduction, and conversely, silencing its expression by small interfering RNAs showed significant inhibitory effects. Collectively, we demonstrated that SINAL10 is a novel GP64-binding protein that stimulates BmNPV proliferation. [ABSTRACT FROM AUTHOR]

Microsporidia Nosema bombycis CQ1 can be vertically transmitted in silkworm Bombyx mori but Vairimorpha necatrix BM cannot. Therefore, the pathological differences in silkworm infected with these two microsporidia required clarification. Here, we compared the virulence of N. bombycis CQ1 and V. necatrix BM against silkworm. The pathological characteristics in intestine, testis and ovary were surveyed using paraffin sections, scanning electron microscopy and transmission electron microscopy. Our data firstly showed that the virulence of V. necatrix BM was weaker than that of N. bombycis CQ1. Secondly, the typical symptom of V. necatrix BM infection is making xenomas, which are full of pathogens in different stages, at the posterior of intestine. However, no xenomas were formed surrounding intestines infected with N. bombycis CQ1. Thirdly, N. bombycis CQ1 can cluster spores near the trachea while infecting ovaries. It is worth noting that N. bombycis CQ1 infected epithelial cells and connective tissues of ovaries, while V. necatrix BM did not. Although silkworm ovaries can not be infected by V. necatrix BM in vivo , it can infect embryonic and ovarian cell lines in vitro. This study is the first report about comparing infection features of N. bombycis CQ1 and V. necatrix BM in silkworm tissues and it provided elaborate and visual information of pathological characteristics which can help to explain the different transmission strategies of these two microsporidia. [ABSTRACT FROM AUTHOR]

WD40 proteins are scaffolding molecules in protein-protein interactions and play crucial roles in fundamental biological processes. Genome-wide characterization of WD40 proteins in animals has been conducted solely in humans. We retrieved 172 WD40 protein genes in silkworm (BmWD40s) and identified these genes in 7 other insects, 9 vertebrates and 5 nematodes. Comparative analysis revealed that the WD40 protein gene family underwent lineage-specific expansions during animal evolution, but did not undergo significant expansion during insect evolution. The BmWD40s were categorized into five clusters and 12 classes according to the phylogenetic classification and their domain architectures, respectively. Sequence analyses indicated that tandem and segmental duplication played minor roles in producing the current number of BmWD40s, and domain recombination events of multi-domain BmWD40s might have occurred mainly after gene duplication events. Gene Ontology (GO) analysis revealed that a higher proportion of BmWD40s was involved in processes, such as binding, transcription-regulation and cellular component biogenesis, compared to all silkworm genes annotated in GO. Microarray-based analysis demonstrated that many BmWD40s had tissue-specific expression and exhibited high and/or sex-related expression during metamorphosis. These findings contribute to a better understanding of the evolution of the animal WD40 protein family and assist the study of the functions of BmWD40s. [ABSTRACT FROM AUTHOR]

The Murshidabad District of West Bengal, India has practised traditional mulberry sericulture since time immemorial. One of the most important aspects for sustainable development of the sericulture industry is the control and prevention of silkworm diseases. The main objective of this study was to determine silkworm disease incidence over the period 1992–2011 in Murshidabad District and how environmental factors have contributed towards their occurrence. Data were collected from a crop-wise survey of silkworm diseases from 25 farmers (five farmers from five villages), who each had a separate rearing house, were progressive and had a capacity of rearing at least 100 disease free layings (dfls) in each crop. Disease incidence was expressed in percentage and calculated taking into consideration 40,000 larvae per 100 dfls. Crop-wise silkworm disease mortality data were correlated with meteorological data. Data collected from the continuous survey conducted in the district during 1992–2011 revealed that there was no set pattern of occurrence of silkworm diseases. However, maximum mortality (up to 30%) of silkworm due to diseases in all the seasons occurred mainly due to grasserie, as relative humidity remains high for most part of the year in this area. However, other than grasserie, for most of the period, disease incidence was below 10%. This observation of the dominance of grasserie over other diseases in causing silkworm mortality calls for renewed emphasis on the preventive measures and development of disease forecasting models, for disease control. Another observation was that since 1993, pebrine, the deadliest disease of the silkworm, has not been reported from the farmers’ fields. [ABSTRACT FROM AUTHOR]

Understanding the virulence and pathogenesis of human pathogens using insect models is an increasingly popular method. Francisella novicida , which is virulent in mice but non-pathogenic to immunocompetent humans, is widely used as an ideal candidate for Francisella research. In this study, we developed a silkworm ( Bombyx mori ) infection model for F. novicida by inoculating the hemocoels of silkworms with F. novicida . We found that silkworms died within 3–7 days of F. novicida infection. However, the deletion mutant of DotU, the core part of type VI secretion systems, failed to kill silkworm. In whole silkworm bodies, the bacterial load of the DotU deletion mutant was significantly less than that of the wild-type strain. Approximately 10-fold increase in bacterial load was recorded in hemolymph and subcutaneous tissues compared with that in the silk gland, Malpighian tubule, and reproductive organs. The CFU count of the DotU deletion mutant in all organs was similar results to the whole body CFU count. Confocal microscopy further confirmed the arrested growth of the mutant strain within hemocytes. The intracellular growth of F. novicida strains was also analyzed using the silkworm ovary-derived cell line BmN4. In BmN4, both CFU count assay and confocal microscopy revealed extensive growth of the wild-type strain compared with that of the mutant strain. Francisella DotU has already been proven as a virulence factor in mammals, and it was also found to be an essential virulence factor in our silkworm infection model. Therefore, this silkworm infection model is suitable for identifying new virulence factors of Francisella . [ABSTRACT FROM AUTHOR]

Context:Antibiotic resistance is a serious problem worldwide. Searching for new potential agents is, therefore, essential. The bark ofSesbania grandiflora(L.) Pers. (Fabaceae) has been used in folk medicine against various diseases. Objective:To investigate the antibacterial activity ofS. grandiflorabark 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) ofS. grandiflorabark 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 resistantStaphylococcus aureus(MRSA) and vancomycin resistant enterococci (VRE) of 1.6 and 0.4 mg/mL, respectively, and against Gram-negativeEscherichia coliandPseudomonas aeruginosaof 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 EC50of 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. [ABSTRACT FROM PUBLISHER]

Nuclear polyhedrosis virus (NPV) is the most harmful virus responsible for the manifestation of grasserie disease in the larvae of silkworm,Bombyx mori. It causes a huge economic loss in the sericulture industry. An attempt was made in the present investigation for the screening of antiviral activity using medicinal plants such asLantana camara, Phyllanthus amarusand marine seaweeds such asSargassum wightii, Turbinaria ornataagainstBmNPV. Crude extracts were prepared using different solvents, such as hexane, ethyl acetate, methanol and water. The silkworm feeding bioassay study was carried out with the crude extracts to investigate the presence of anti-BmNPV activity after inoculating fifth instar larvae of silkworm with occlusion bodies (OBs) ofBmNPV. Each extract was tested for their anti-BmNPV activity using various concentrations of crude extracts ranging from 200 μg to 1000 μg. Among the crude extracts tested, methanol and aqueous extracts ofP. amarusshowed significant anti-BmNPV activity. [ABSTRACT FROM PUBLISHER]

The development of effective drugs against fungal diseases involves performing infection experiments in animals to evaluate candidate therapeutic compounds. Cryptococcus neoformans is a pathogenic fungus that causes deep mycosis, resulting in respiratory illness and meningitis. Here we describe a silkworm system established to evaluate the safety and efficacy of therapeutic drugs against infection by Cryptococcus neoformans and the advantages of this system over other animal models. The silkworm assay system has two major advantages: 1) silkworms are less expensive to rear and their use is less problematic than that of mammals in terms of animal welfare, and 2) in vivo screenings for identifying candidate drugs can be easily performed using a large number of silkworms. The pharmacokinetics of compounds are consistent between silkworms and mammals. Moreover, the ED50 values of antibiotics are concordant between mammalian and silkworm infection models. Furthermore, the body size of silkworms makes them easy to handle in experimental procedures compared with other invertebrate infectious experimental systems, and accurate amounts of pathogens and chemicals can be injected fairly easily. These advantages of silkworms as a host animal make them useful for screening candidate drugs for cryptococcosis. [ABSTRACT FROM AUTHOR]

Recent advances in bacterial characterization methodologies have made taxonomic categorization significantly more accurate. Here, we re-evaluated the position of bacterial strains (532 and 652) belonging to the genus Hafnia, isolated from flacherie-diseased silkworms in 1965. Phylogenetic analysis based on the 16S rRNA gene sequences of these strains suggests that they belong to genus Enterobacter. Using multilocus sequence analysis (MLSA), these strains were further classified to MLSA group A, which is a "core" group of Enterobacter containing E. cloacae (the type species of the genus). Although these strains were closely related to E. mori, E. tabaci, and E. asburiae, they also had other MLSA characteristics that distinguished them from these neighboring bacterial species. These data were supported by further biochemical analysis. Thus, it appears that the 532 and 652 strains isolated almost half a century ago belong to genus Enterobacter, and their unique characteristics strongly suggest that they are a novel bacterial species. [ABSTRACT FROM AUTHOR]

Synthetic compounds are a vital source of antimicrobial agents. To uncover therapeutically effective antimicrobial agents from a chemical library, we screened over 100,000 synthetic compounds for in vitro antimicrobial activity against methicillin-resistant Staphylococcus aureus and evaluated the in vivo therapeutic effectiveness of the hits in S. aureus-infected silkworms. Three antimicrobial agents exhibited therapeutic effects in the silkworm infection model. One of these, GPI0363, a novel spiro-heterocyclic compound, was bacteriostatic and inhibited RNA synthesis in S. aureus cells. GPI0363-resistant S. aureus strains harbored a point mutation in the gene encoding the primary sigma factor, SigA, of RNA polymerase, and thismutation was responsible for the resistance to GPI0363. We further revealed that GPI0363 could bind to SigA, inhibit promoter-specific transcription in vitro, and prolong the survival of mice infected with methicillin-resistant S. aureus. Thus, GPI0363 is an attractive candidate therapeutic agent against drug-resistant S. aureus infections. [ABSTRACT FROM AUTHOR]

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 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. [ABSTRACT FROM AUTHOR]

Viral microRNAs (miRNAs) have been demonstrated to play important roles in virus-host interactions. Some RNA virus-encoded miRNAs have been reported to promote viral replication and may be used as potential drug targets. Bombyx mori cypovirus (BmCPV), an important pathogen of silkworm, is a double-stranded RNA virus frequently causing serious damages in sericulture. Research on miRNA encoded by BmCPV may be useful to elucidate the BmCPV-host interaction and to develop new anti-viral methods. In our previous study, small RNA libraries of the midgut of BmCPV-infected silkworm have been generated by deep sequencing and several BmCPV-encoded putative miRNAs were predicted. In this study, two putative miRNAs encoded by BmCPV were identified and then validated by stem-loop qRT-PCR and northern blot. They are BmCPV-miR-3 encoded by the third genomic RNA segment of BmCPV (478–497 bp) and BmCPV-miR-5 encoded by the fifth genomic RNA segment (2481–2500 bp), both are 20 bp and encoded by ORF regions. miRNA expression could be detected as early as 5 h after BmCPV infection, and the expression level of BmCPV-miR-3 is much higher than that of BmCPV-miR-5 in the course of infection. Three potential target genes were predicted in the host genome, two for BmCPV-miR-3 and one for BmCPV-miR-5, but just one in the virus genome for BmCPV-miR-3 only, with the binding sites all in coding regions. Dual luciferase assay and qRT-PCR indicated that BmCPV-miR-3 could down-regulate the expression of both its two target genes, but no regulatory effect by BmCPV-miR-5 on its target gene was detected. In contrast, BmCPV-miR-3 could up-regulate the viral target. This is the first report that an insect double stranded RNA virus may generate miRNAs and the results obtained will benefit the future study of the functions of BmCPV-encoded miRNAs on viral replication and virus-host interaction. [ABSTRACT FROM AUTHOR]

The microsporidia Nosema bombycis is the insect pathogen of pebrine disease severely destructive to sericulture production. Here, we describe the use of Escherichia coli HT115 strain ( DE3) to express double-strand RNAs targeting the gene encoding ADP/ ATP protein in N. bombycis. The results showed that ds RNAs deferentially suppressed the gene expression during N. bombycis infection in the silkworm, and the effect waned gradually. Our results, for the first time, provide a tool to utilize the ds RNA expressed by recombinant E. coli to control the pebrine disease of the domestic silkworm. [ABSTRACT FROM AUTHOR]

BACKGROUND Silkworm ( Bombyx mori) is an economically important insect. It is relatively less resistant to certain chemicals and environment exposures such as pesticides and pathogens. After pesticide exposures, the silkworms are more susceptible to microbial infections. The mechanism underlying the susceptibility might be related to immune response and oxidative stress. RESULTS A sublethal dose of phoxim combined with Bombyx mori nucleopolyhedrovirus (BmNPV) elevated the silkworm mortality at 96 h. We found a higher content of H2O2 and increased levels of genes related to oxidative stress and immune response after treatment with a sublethal dose of phoxim for 24 h or 48 h. However, such response decreased with longer pesticide treatment. Mortality increased by 44% when B. mori was exposed to combined treatment with BmNPV and phoxim rather than BmNPV alone. The level of examined immune-related and oxidative-stress-related genes significantly decreased in the combined treatment group compared with the BmNPV group. Our results indicated that, with long-term exposure to pesticides such as OPs, even at sublethal dose, the oxidative stress response and immune responses in silkworm were inhibited, which may lead to further immune impairment and accumulation of oxidative stress, resulting in susceptibility to the virus and harm to the silkworm. CONCLUSION Our study provided insights for understanding the susceptibility to pathogen after pesticide exposures, which may promote the development of better pesticide controls to avoid significant economic losses. © 2016 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

The silkworm infection model has the potential to replace conventional animal models for evaluation of the efficacy and toxicity of investigational antifungal agents. Silkworms are relatively inexpensive, can be simply grown in large numbers and can be easily infected with pathogenic fungi, including mutant strains. Antifungal agents can then be injected into the silkworm either via the hemolymph to mimic intravenous administration or directly into the gut for oral administration, and their antifungal effect can be evaluated. Common features regarding the mechanisms of pharmacokinetics between the silkworm and mammals result in consistent therapeutic effectiveness of antifungal agents. ASP2397, a promising new antifungal agent, was discovered using the silkworm model. The conclusion is that silkworms can be a more ethical and less expensive alternative to standard animal models, particularly for the identification and testing of new antifungal agents. [ABSTRACT FROM AUTHOR]

Bacterial infection causes enhanced reactive oxygen species (ROS) levels in insects. Oxidation resistance 1 (OXR1) plays an antioxidant role in eukaryotic organisms, including insects. In this report, we demonstrated that Pseudomonas aeruginosa and Staphylococcus aureus infection and hydrogen peroxide (H2O2) injection induced the expression of specific transcriptional isoforms of OXR1 in larval silkworms. We further showed that a Jun kinase (JNK) pathway inhibitor, SP600125, down-regulated expression of OXR1 during infection, leading to elevated H2O2 levels in the hemolymph, resulting in lower viability of the injected bacteria inside the silkworm larvae. Our study suggests that OXR1 participates in protecting larval silkworms from oxidative stress and bacterial infection through the JNK pathway. [ABSTRACT FROM AUTHOR]

Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) specifically infects the epithelial cells in the midgut of silkworm and causes them to death, which negatively affects the sericulture industry. In order to determine the midgut response at the protein levels to the virus infection, differential proteomes of the silkworm midgut responsive to BmCPV infection were identified with isobaric tags for relative and absolute quantitation (iTRAQ) labeling followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). 193, 408, 189 differentially expressed proteins (DEPs) were reliably quantified by iTRAQ analysis in the midgut of BmCPV-infected and control larvae at 24, 48, 72 h post infection (hpi) respectively. KEGG enrichment analysis showed that Oxidative phosphorylation, amyotrophic lateral sclerosis, Toll-like receptor signaling pathway, steroid hormone biosynthesis were the significant pathways (Q value ≤ 0.05) both at 24 and 48 hpi. qRT-PCR was used to further verify gene transcription of 30 DEPs from iTRAQ, showing that the regulations of 24 genes at the transcript level were consistent with those at the proteomic level. Moreover, the cluster analysis of the three time groups showed that there were seven co-regulated DEPs including BGIBMGA002620-PA, which was a putative p62/sequestosome-1 protein in silkworm. It was upregulated at both the mRNA level and the proteomic level and may play an important role in regulating the autophagy and apoptosis (especially apoptosis) induced by BmCPV infection. This was the first report using an iTRAQ approach to analyze proteomes of the silkworm midgut against BmCPV infection, which contributes to understanding the defense mechanisms of silkworm midgut to virus infection. Significance The domesticated silkworm, Bombyx mori , is renowned for silk production as well as being a traditional lepidopteron model insect served as a subject for morphological, genetic, physiological, and developmental studies. Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) specifically infects the epithelial cells in the midgut of silkworm and causes the silkworm to death, which negatively affects the sericulture industry. Studies on insect antiviral immunity and on interactive mechanisms between host cells and BmCPV are in their infancy and remain insufficient. In order to obtain an overall view of silkworm response to BmCPV infection, we performed a proteomic analysis of the midgut of silkworm responses to BmCPV infection by iTRAQ. This was the first report using an iTRAQ approach to analyze proteomes of the silkworm midgut against BmCPV infection, which contributes to understanding the defense mechanisms of silkworm midgut to virus infection. [ABSTRACT FROM AUTHOR]

Bombyx mori nucleopolyhedrosis virus (BmNPV) is a major pathogen that threatens the growth and sustainability of the sericulture industry. Since microRNAs (miRNAs) have been shown to play important roles in host-pathogen interactions, in this study we investigated the effects of BmNPV infection on silkworm microRNAs expression profile. To achieve this, we constructed and deep-sequenced two small RNA libraries generated from BmNPV infected and un-infected larvae. The results revealed that 38 silkworm miRNAs were differentially expressed after BmNPV infection. Based on the GO analysis, their predicted target genes were found to be involved in diverse functions such as binding, catalytic, virion and immune response to stimulus suggesting their potential roles in host-virus interactions. Using the dual-luciferase reporter assay, we confirmed that Bmo-miR-277-5p, up-regulated in BmNPV-infected larvae, targeted the B. mori DNA cytosine-5 methyltransferase (Dnmt2) gene which may play potential role in silkworm-BmNPV interaction. These results provide new insights into exploring the interaction mechanism between silkworm and BmNPV. [ABSTRACT FROM AUTHOR]

The halophilic marine bacterium, Vibrio vulnificus, occasionally causes fatal septicemia in immunocompromised patients. Mice are commonly used as experimental animals to investigate the virulence of V. vulnificus, however, a large number of mice are generally required for bioassays. The present study examined whether the invertebrate species, silkworms, can be used instead of mice to investigate V. vulnificus virulence. When the silkworms were inoculated with 1.2x107 colony forming units of V. vulnificus OPU1-Rf, a virulent strain of V. vulnificus, all injected silkworms died within 48 h, however, those injected with culture filtrate or diluent did not. This silkworm infection model was then used to isolate attenuated V. vulnificus mutants from 1,016 transposon-inserted mutants. Consequently, a harmless mutant, SW998, was isolated. In this strain, the transposon was inserted into the rtxA gene, which is a known V. vulnificus virulence gene. In conclusion, the present study demonstrated that silkworms are useful animals for investigating the virulence of V. vulnificus. [ABSTRACT FROM AUTHOR]

Bombyx mori nucleopolyhedrovirus (BmNPV) orf5 ( Bm5 ) is a core gene of lepidopteran baculoviruses and encodes the protein with the conserved amino acid residues (DUF3627) in its C-terminus. Here, we found that Bm5 disruption resulted in lower titers of budded viruses and fewer numbers of occlusion bodies (OBs) in B. mori cultured cells and larvae, although viral genome replication was not affected. Bm5 disruption also caused aberrant expression of various viral genes at the very late stage of infection. Immunocytochemical analysis revealed that BM5 localized to the nuclear membrane. We also found that DUF3627 is important for OB production, transcriptional regulation of viral genes, and subcellular localization of BM5. Compared with wild-type BmNPV infection, larval death was delayed when B. mori larvae were infected with Bm5 mutants. These results suggest that BM5 is involved in progeny virus production and regulation of viral gene expression at the very late stage of infection. [ABSTRACT FROM AUTHOR]

Insect infection models have been used increasingly to study various pathogenic agents in evaluations of pathogenicity and drug efficacy. In this study, we demonstrated that larvae of the silkworm Bombyx mori are useful for studying Listeria monocytogenes infections in insects. Infection with the L. monocytogenes wild-type strain induced silkworm death. Infection by a listeriolysin O (LLO) deletion mutant also induced silkworm death, but the bacterial numbers in silkworms were lower than those of the wild-type strain. Intracellular growth was observed when the silkworm ovary-derived cell line BmN4 was infected with the wild-type strain. Explosive replication was not observed in BmN4 cells infected with the LLO mutant and the bacterial numbers of the LLO mutant were lower than those of the wild-type strain. Pretreatment with vitamin A did not affect silkworm mortality after bacterial infection, but the efficiency of infecting the hemocytes and BmN4 cells was decreased with vitamin A treatment. Our results indicate that silkworm larvae are a useful insect infection model for L. monocytogenes and that vitamin A has protective effects against bacterial infection in silkworms. [ABSTRACT FROM AUTHOR]

Nosema bombycis, a microsporidium, is a pathogen of pebrine disease of silkworms, and its genomic DNA sequences had been determined. Thus far, the research of gene functions of microsporidium including N. bombycis cannot be performed with gain/loss of function. In the present study, we targeted to construct transgenic N. bombycis. Therefore, hemocytes of the infected silkworm were transfected with a non-transposon vector pIZT/V5-His vector in vivo, and the blood, in which the hemocyte with green fluorescence could be observed, was added to the cultured BmN cells. Furthermore, normal BmN cells were infected with germinated N. bombycis, and the infected cells were transfected with pIZT/V5-His. Continuous fluorescence observations exposed that there were N. bombycis with green fluorescence in some N. bombycis-infected cells, and the extracted genome from the purified N. bombycis spore was used as templates. PCR amplification was carried out with a pair of primers for specifically amplifying the green fluorescence protein (GFP) gene; a specific product representing the gfp gene could be amplified. Expression of the GFP protein through Western blotting also demonstrated that the gfp gene was perfectly inserted into the genome of N. bombysis. These results illustrated that exogenous gene can be integrated into N. bombycis genome by mediating with a non-transposon vector. Our research not only offers a strategy for research on gene function of N. bombycis but also provides an important reference for constructing genetically modified microsporidium utilized for biocontrol of pests. [ABSTRACT FROM AUTHOR]

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 (10 4 CFU/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 (LC 50 ) values of the pathogenic strains were calculated at different time intervals (24, 48, 72 and 96 h) within the range from 1.38 × 10 2 to 3.63 × 10 7 CFU/ml. The pathogenic groups demonstrated inhibition of phenoloxidase activity and decreased in total hemocyte count after 48 h 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. [ABSTRACT FROM AUTHOR]

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Transforming growth factor (TGF)-β superfamily members inhibit Bombyx mori nucleohedrovirus (BmNPV) multiplication in silkworm are not determined. In this study, we first found that BmNPV RNA transcription and protein expression level were regulated by TGF-β members, Decapentaplegic (Bmdpp) and Dawdle (Bmdaw) in the domesticated silkworm, B. mori and silkworm ovary-derived cells. Furthermore, subcellular localization showed that Bmdpp and Bmdaw were mainly presented in cytomembrane of the cultured BmN cells. Tissues expression pattern analysis found that the highest expression levels of Bmdpp and Bmdaw genes were in the hemocyte of fifth instar larvae. During the immune response, the expression level of Bmdpp gene was elevated and Bmdaw gene was declined in BmNPV infected BmN cells and silkworm. The multiplication of BmNPV was inhibited by overexpression of Bmdpp and Bmdaw genes in BmN cells. RNA interference experiments found that the multiplication of BmNPV was raised with specific siRNAs of Bmdpp and Bmdaw genes in BmN cells. The antiviral immune pathways were not significantly regulated by the TGF-β superfamily members. Taken together, these findings provided a clue to understand the function of Bmdpp and Bmdaw gene in response to the BmNPV infection in silkworm. [ABSTRACT FROM AUTHOR]

Viruses and high temperature (HT) are the primary threats to silkworms. Changes in the expression of stress-response genes can be measured using quantitative polymerase chain reaction (qPCR) after exposure to viruses or HT. However, appropriate reference genes (RGs) for qPCR data normalization have not been established in this organism. In this study, we summarized the RGs used in the previous silkworm studies after infection with Bombyx mori nucleopolyhedrovirus (BmNPV), B. mori cytoplasmic polyhedrosis virus (BmCPV), or B. mori densovirus (BmDNV) or after HT treatment. The expression levels of these RGs were extracted from silkworm transcriptome data to screen for candidate RGs that were unaffected by the experimental conditions. Actin-1 ( A1), actin-3 ( A3), glyceraldehyde-3-phosphate dehydrogenase ( GAPDH), and translation initiation factor 4a ( TIF- 4A) were selected for further qPCR verification. The results of RNA-seq and qPCR showed that GAPDH and TIF- 4A were suitable RGs after BmNPV challenge or HT stress, whereas TIF- 4A was an appropriate RG for BmCPV or BmDNV-Z challenge in silkworms. These results suggested that TIF-4A may be the most appropriate RG for gene expression analysis after challenge with viruses or HT in silkworms. [ABSTRACT FROM AUTHOR]

Genes in the signal transducer and activator of transcription (STAT) family are vital for activities including gene expression and immune response. To investigate the functions of the silkworm Bombyx mori STAT ( Bm-STAT ) gene in antiviral immunity, two Bm-STAT gene isoforms, Bm-STAT -L for long form and Bm-STAT- S for short form, were cloned. Sequencing showed that the open reading frames were 2313 bp encoding 770 amino acid residues for Bm-STAT -L and 2202 bp encoding 734 amino acid residues for Bm-STAT- S. The C-terminal 42 amino acid residues of Bm-STAT-L were different from the last 7 amino acid residues of Bm-STAT-S. Immunofluorescence showed that Bm-STAT was primarily distributed in the nucleus. Transcription levels of Bm-STAT in different tissues were determined by quantitative PCR, and the results revealed Bm-STAT was mainly expressed in testes. Western blots showed two bands with molecular weights of 70 kDa and 130 kDa in testes, but no bands were detected in ovaries by using anti-Bm-STAT antibody as the primary antibody. Expression of Bm-STAT in hemolymph at 48 h post infection with B. mori macula-like virus (BmMLV) was slightly enhanced compared with controls, suggesting a weak response induced by infection with BmMLV. Hemocyte immunofluorescence showed that Bm-STAT expression was elevated in B. mori nucleopolyhedrovirus (BmNPV)-infected cells. Moreover, resistance of BmN cells to BmNPV was reduced by downregulation of Bm-STAT expression and increased by upregulation. Resistance of BmN cells to BmCPV was not significantly improved by upregulating Bm-STAT expression. Therefore, we concluded that Bm-STAT is a newly identified insect gene of the STAT family. The JAK-STAT pathway has a more specialized role in antiviral defense in silkworms, but JAK-STAT pathway is not triggered in response to all viruses. [ABSTRACT FROM AUTHOR]

Silkworm shows hyperglycemia after intake of diet containing large amount of glucose. The hyperglycemic silkworm model is useful for evaluation of anti-diabetic drugs. A hot water extract of Rehmanniae Radix, an herbal medicine, showed hypoglycemic effect against the hyperglycemic silkworms. This method is applicable for quick and simple evaluation of the hypoglycemic activities of different batches of Rehmanniae Radix. Our findings suggest that silkworms have a lot of merit as experimental animals for evaluation of various herbal medicines. [ABSTRACT FROM AUTHOR]

Bombyx mori bidensovirus (BmBDV) and Bombyx mori nucleopolyhedrovirus (BmNPV) are serious pathogens of Bombyx mori. In this study, we reported the changes of transcription level of several immune genes, including bmi, argo, dicer, cap1, cap3 and car, in Bombyx mori midgut after exposure to BmBDV or BmNPV. Silkworm strains 798 (anti-BmBDV) and 306 (susceptible to BmBDV) were subjected to BmBDV infection, and NB (anti-BmNPV) and HUABA (35) (susceptible to BmNPV) were subjected to BmNPV infection. The results showed that the transcription levels differ largely among different silkworm strains, and that the extent to which the gene transcriptions were affected by the viruses was different. However, both BmNPV and BmBDV viruses can reverse the transcription patterns of these genes when the silkworms were administered with the viruses compared with those control groups. The transcript levels of bmi and dicer were decreased in 798 and 306 strains that were inoculated with BmBDV compared with their respective controls, but were increased in NB and HUABA (35) inoculated with BmNPV. The transcript levels of argo and cap3 were risen in 798, 306 and NB strains when inoculated with their respective viruses, but were decreased in HUABA (35) strain. The transcript levels of cap1 were risen in all silkworm strains, while the levels of car were decreased in 798, 306 and HUABA (35) strains, and increased in NB strain when inoculated with their respective viruses. These findings may contribute to more in-depth understanding on functions of these genes in virus infection and proliferation. [ABSTRACT FROM AUTHOR]

We aimed to identify virulence-associated genes of Serratia liquefaciens FK01 against Bombyx mori. Among 1,200 transconjugants from a transposon library, 4 (ET0234, ET0373, ET0418, and ET0964) showed decreased virulence towards B. mori. Southern hybridization revealed that the transposon was inserted at a single site of the S. liquefaciens genome. The flanking sequences of the transposon indicated that it disrupted the lipopolysaccharide (LPS) synthesis gene in all mutants. The complemented strain restored virulence completely or partially. Thus, LPS contributes to the virulence of S. liquefaciens against B. mori. Serum killing assays indicated that the bacterium was probably killed by the complement system. Since the innate immunity of a host is triggered by bacterial recognition, LPS might inhibit recognition by modifying the bacterial surface in B. mori. [ABSTRACT FROM AUTHOR]

We previously established two silkworm cell lines, BmN-SWU1 and BmN-SWU2, from Bombyx mori ovaries. BmN-SWU1 cells are susceptible while BmN-SWU2 cells are highly resistant to BmNPV infection. Interestingly, we found that the entry of BmNPV into BmN-SWU2 cells was largely inhibited. To explore the mechanism of this inhibition, in this study we used isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative protein expression profiling and identified 629 differentially expressed proteins between the two cell lines. Among them, we identified a new membrane protein termed BmREEPa. The gene encoding BmREEPa transcribes two splice variants; a 573 bp long BmREEPa-L encoding a protein with 190 amino acids and a 501 bp long BmREEPa-S encoding a protein with 166 amino acids. BmREEPa contains a conserved TB2/DP, HVA22 domain and three transmembrane domains. It is localized in the plasma membrane with a cytoplasmic C-terminus and an extracellular N-terminus. We found that limiting the expression of BmREEPa in BmN-SWU1 cells inhibited BmNPV entry, whereas over-expression of BmREEPa in BmN-SWU2 cells promoted BmNPV entry. Our results also indicated that BmREEPa can interact with GP64, which is the key envelope fusion protein for BmNPV entry. Taken together, the findings of our study revealed that BmREEPa is required for BmNPV to gain entry into silkworm cells, and may provide insights for the identification of BmNPV receptors. [ABSTRACT FROM AUTHOR]

This study assessed the effect of the pesticides on activity and expression of glutathione S-transferases (GSTs) in the midgut of the silkworm after intense selections by phoxim and fenpropathrin for five and four generations, respectively. GSTs activity towards cumene hydroperoxide (CHP), a substrate of GSH-dependent peroxidase, was significantly increased in the crude homogenate of midguts after long term exposure to the insecticides. An epsilon-class GST gene ( BmGSTe2 ) was identified and showed elevated expression in the midguts of phoxim- and fenpropathrin-selected strains. Expression of BmGSTe2 was only detected in the midgut at transcriptional and translational levels. The recombinant BmGSTE2 possessed peroxidase activity and significantly inhibited by fenpropathrin, phoxim and chlorpyrifos in vitro. These results indicated that BmGSTE2 might be one of the enzymes involved in enhancing larval tolerance to the insecticides used. Furthermore, GST activity and expression level of BmGSTe2 might be used as biomarkers of organophosphorus and pyrethroid insecticide exposures. [ABSTRACT FROM AUTHOR]

Baculoviruses have been known to induce hyperactive behavior in their lepidopteran hosts for over a century. As a typical lepidopteran insect, the silkworm Bombyx mori displays enhanced locomotor activity (ELA) following infection with B . mori nucleopolyhedrovirus (BmNPV). Some investigations have focused on the molecular mechanisms underlying this abnormal hyperactive wandering behavior due to the virus; however, there are currently no reports about B . mori . Based on previous studies that have revealed that behavior is controlled by the central nervous system, the transcriptome profiles of the brains of BmNPV-infected and non-infected silkworm larvae were analyzed with the RNA-Seq technique to reveal the changes in the BmNPV-infected brain on the transcriptional level and to provide new clues regarding the molecular mechanisms that underlies BmNPV-induced ELA. Compared with the controls, a total of 742 differentially expressed genes (DEGs), including 218 up-regulated and 524 down-regulated candidates, were identified, of which 499, 117 and 144 DEGs could be classified into GO categories, KEGG pathways and COG annotations by GO, KEGG and COG analyses, respectively. We focused our attention on the DEGs that are involved in circadian rhythms, synaptic transmission and the serotonin receptor signaling pathway of B . mori . Our analyses suggested that these genes were related to the locomotor activity of B . mori via their essential roles in the regulations of a variety of behaviors and the down-regulation of their expressions following BmNPV infection. These results provide new insight into the molecular mechanisms of BmNPV-induced ELA. [ABSTRACT FROM AUTHOR]

In the present study, we examined whether microorganisms collaterally ingested by insects with their food activate the innate immune system to confer systemic resistance against subsequent bacterial invasion. Silkworms orally administered heat-killed Pseudomonas aeruginosa cells showed resistance against intra-hemolymph infection by P. aeruginosa. Oral administration of peptidoglycans, cell wall components of P. aeruginosa, conferred protective effects against P. aeruginosa infection, whereas oral administration of lipopolysaccharides, bacterial surface components, did not. In silkworms orally administered heat-killed P. aeruginosa cells, P. aeruginosa growth was inhibited in the hemolymph, and mRNA amounts of the antimicrobial peptides cecropin A and moricin were increased in the hemocytes and fat body. Furthermore, the amount of paralytic peptide, an insect cytokine that activates innate immune reactions, was increased in the hemolymph of silkworms orally administered heat-killed P. aeruginosa cells. These findings suggest that insects sense bacteria present in their food by peptidoglycan recognition, which activates systemic immune reactions to defend the insects against a second round of infection. [ABSTRACT FROM AUTHOR]

Mulberry silkworm, Bombyx mori L. is susceptible to different diseases viz., Grasserie, Flacherie, Muscardine and Pebrine. To formulate silkworm disease management strategies, understanding the factors responsible for causing the diseases and the impact of silkworm diseases on cocoon productivity at farmers' conditions are essential. The silkworm hybrids reared at present in South Indian states are having the potential to yield 60-75 kg of cocoons/100 dfls of rearing. In Tamil Nadu, majority of the farmers are harvesting >60 kg cocoons/100 dfls of rearing but in certain areas, farmers are still harvesting <60 kg cocoons/100 dfls (less than potential of the hybrid). Hence, to find out the prevalence of silkworm diseases in the selected areas of Tamil Nadu viz., Salem, Erode and Tiruppur and their impact on cocoon productivity and the factors responsible for low cocoon yield (<60 kg/100 dfls of rearing), the present study was conducted for two years (October, 2012 to September, 2014). The average data on the point prevalence of diseases in two years indicated that the prevalence of 1.65 % of diseases in which grasserie was 0.46%, flacherie was 1.19% and no incidence of muscardine and pebrine throughout the period. Among 720 farmers crops surveyed during the period, 568 farmers (78.89 %) were harvested >60 kg cocoons/100 dfls of rearing and 152 farmers (21.11 %) were harvested <60 kg cocoons/100 dfls of rearing. The major reasons for harvesting <60 kg cocoons/100 dfls (less than potential of the hybrid) are poor quality leaf (16.11%) and high temperature with low humidity (1.25%). [ABSTRACT FROM AUTHOR]

This study established the genetic characterisation of 10 microsporidian isolates infecting non-mulberry silkworms (Antheraea assamensis and Samia cynthia ricini) collected from biogeographical forest locations in the State of Assam, India, using PCR-based markers assays: inter simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD). A Nosema type species (NIK-1s_mys) was used as control for comparison. The shape of mature microsporidian spores were observed oval to elongated, measuring 3.80 to 4.90 μm in length and 2.60 to 3.05 μm in width. Fourteen ISSR primers generated reproducible profiles and yielded 178 fragments, of which 175 were polymorphic (98%), while 16 RAPD primers generated reproducible profiles with 198 amplified fragments displaying 95% of polymorphism. Estimation of genetic distance coefficients based on dice coefficients method and clustering with un-weighted pair group method using arithmetic average (UPGMA) analysis was done to unravel the genetic diversity of microsporidians infecting Indian muga and eri silkworm. The similarity coefficients varied from 0.385 to 0.941 in ISSR and 0.083 to 0.938 in RAPD data. UPGMA analysis generated dendrograms with two microsporidian groups, which appear to be different from each other. Based on Euclidean distance matrix method, 2-dimensional distribution also revealed considerable variability among different identified microsporidians. Clustering of these microsporidian isolates was in accordance with their host and biogeographic origin. Both techniques represent a useful and efficient tool for taxonomical grouping as well as for phylogenetic classification of different microsporidians in general and genotyping of these pathogens in particular. [ABSTRACT FROM AUTHOR]

The fungal pathogen Beauveria bassiana causes serious economic losses in sericulture. Its origin is usually attributed to the release of B. bassiana insecticides against pine caterpillars ( Dendrolimus punctuatus ). In the present study, 488 B. bassiana isolates obtained from silkworm ( Bombyx mori ) collected from 13 Chinese provinces, and 327 B. bassiana isolates obtained from D. punctatus collected from 9 provinces, were analyzed for population genetic structure using the ISSR technique based on genetic distance. A UPGMA dendrogram clustered them into three independent clades: two B. mori clades and one D. punctatus clade. A 3-D principal component analysis further divided them into two completely independent host groups, revealing high host-specificity. This suggested that white muscardine occurring in B. mori populations throughout southern China was not caused by any B. bassiana strain either naturally prevailing in D. punctatus populations or by any strain artificially released as a fungal insecticide against D. punctatus . We further investigated the genetic differentiation coefficient G st and gene flow between B. mori -pathogenic and D. punctatus -pathogenic B. bassiana isolates from across China and from five provinces inhabited by both B. mori and D. punctatus . The G st value across China was computed as 0.410, while the values of the five provinces ranged from 0.508 to 0.689; all above 0.25, which is the threshold for significant genetic differentiation. This suggests that B. bassiana strains isolated from the two different hosts maintained their respective heredity without a convergent homogenization trend, and reduces the possibility that the host range of the caterpillar isolates could expand and enhance their virulence in B. mori . These findings indicate that the use of B. bassiana does not threaten the safety of sericulture. [ABSTRACT FROM AUTHOR]

To obtain therapeutically effective new antibiotics, we first searched for bacterial culture supernatants with antimicrobial activity in vitro and then performed a secondary screening using the silkworm infection model. Through further purification of the in vivo activity, we obtained a compound with a previously uncharacterized structure and named it 'lysocin E'. Lysocin E interacted with menaquinone in the bacterial membrane to achieve its potent bactericidal activity, a mode of action distinct from that of any other known antibiotic, indicating that lysocin E comprises a new class of antibiotic. This is to our knowledge the first report of a direct interaction between a small chemical compound and menaquinone that leads to bacterial killing. Furthermore, lysocin E decreased the mortality of infected mice. To our knowledge, lysocin E is the first compound identified and purified by quantitative measurement of therapeutic effects in an invertebrate infection model that exhibits robust in vivo effects in mammals. [ABSTRACT FROM AUTHOR]