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C-type lectins (CTLs) function as pattern recognition receptors (PRRs) and play an important role in the innate immunity of insects. To investigate the role of CTLs in the antifungal responses, we analyzed expression profiles of 36 CTLs of Helicoverpa armigera in the tissues (hemocytes, fat bodies, and midgut) after the infection by entomopathogenic fungus Metarhizium rileyi. The expression levels of many HaCTLs were found to be up-regulated after the infection. Four recombinant HaCTLs (rHaCTL11, rHaCTL12, rHaCTL27, and rHaCTL45) were expressed and purified. Analysis of the purified rHaCTLs revealed that rHaCTLs were able to bind to conidia and hyphal bodies of M. rileyi, and the affinity of rHaCTL11 and rHaCTL27 for hyphal bodies was weaker than for conidia. All these rHaCTLs agglutinate conidia and hyphal bodies in a calcium (Ca

Rab3, a member of the Rab GTPase family, has been found to be involved in innate immunity. However, the precise function of this GTPase in innate immunity remains unknown. In this study, we identified a Rab3 gene (Ha-Rab3) from the cotton bollworm, Helicoverpa armigera and studied its roles in innate immune responses. Expression of Ha-Rab3 was upregulated in the hemocytes of H. armigera larvae after the injection of Escherichia coli or chromatography beads. The dsRNA-mediated knockdown of Ha-Rab3 gene in H. armigera larval hemocytes led to significant reduction in the phagocytosis and nodulation activities of hemocytes against E. coli, significant increase in the bacterial load in larval hemolymph, and significant reduction in the encapsulation activities of hemocytes toward invading chromatography beads. Furthermore, Ha-Rab3 knockdown significantly suppressed spreading of plasmatocytes. These results suggest that Ha-Rab3 plays important roles in H. armigera cellular immune responses, possibly by mediating spreading of hemocytes.

C-type lectins play important roles in innate immunity as pattern recognition receptors (PRRs). We have previously reported a novel C-type lectin HaCTL7 from the cotton bollworm (Helicoverpa armigera) which contains two carbohydrate-recognition domains (CRDs), namely N-terminal CRD1 and C-terminal CRD2. Interestingly, there are four but not six of conserved cysteine residues in CRD2 of HaCTL7, which is different from that of other dual CRD C-type lectins. In the current study, we expressed and purified recombinant HaCTL7 (rHaCTL7) as well as rCRD1 and rCRD2, and demonstrated that both rHaCTL7 and rCRD2, but not rCRD1, owned the agglutinate ability against both Gram-negative and Gram-positive bacteria in a calcium dependent manner. In addition, both rHaCTL7 and rCRD2, but not rCRD1, could bind to various bacteria, and enhanced haemocytes mediated encapsulation and melanization processes. HaCTL7 secreted from fat bodies is able to bind to granulocytes, plasmatocytes and oenocytoids, but not to spherulocytes. Recombinant HaCTL7 and rCRD2 are capable of binding to both granulocytes and oenocytoids, while rCRD1 can only bind to granulocytes. Our data suggest that as a PRR HaCTL7 enhances encapsulation and melanization likely through its C-terminal CRD2, but not N-terminal CRD1, which imply that the characteristic four cysteine structure of CRD2 plays key roles in innate immunity.

SRP gene was first identified from the fall webworm, Hyphantria cunea as one of genes up-regulated after bacteria injection. A rent study in Spodoptera litura showed that stress-induced elevation of SRP expression highly correlates with reduced feeding activities and growth retardation of larvae. In this study, we identified a SRP gene from the cotton bollworm, Helicoverpa armigera, namely Ha-SRP, and studied its precise roles in insect immunity. Expressions of Ha-SRP were upregulated in H. armigera larval hemocytes after injection of Escherichia coli. When the expression of Ha-SRP in H. armigera larval hemocytes was inhibited by dsHa-SRP injection, the transcription of prophenoloxidase genes in hemocytes was repressed, phenoloxidase activity in bacteria-challenged larval hemolymph was significantly decreased, and nodule formation in bacteria-injected larvae was reduced. More importantly, RNAi-treated insects infected with E. coli showed higher bacterial growth in hemolymph compared with infected controls. These results suggest that Ha-SRP gene plays importance roles in H. armigera innate immunity, possibly by mediating prophenoloxidase activation and nodulation response.

C-type lectins can act as pattern recognition receptors (PRRs) and play an important role in innate immunity. Two C-type lectins (HaCTL1 and HaCTL2) have been previously identified in the cotton bollworm (Helicoverpa armigera). Here we isolate six C-type lectins from H. armigera (HaCTL3, 4, 5, 6, 7 and 8). All six new HaCTLs encode a signal peptide (or partial signal peptide) and complete tandem carbohydrate-recognition domains (CRDs). HaCTL4, 5, 6, 7 and 8 mRNA increased in the fat body after injection with both killed Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus, whereas HaCTL3 mRNA was upregulated following E. coli injection only. Recombinant HaCTL3 exhibited agglutinating activity against both Gram-negative and Gram-positive bacteria in a calcium-dependent manner. Agglutination inhibitory analysis indicated that rHaCTL3 recognizes maltose, trehalose, peptidoglycan and lipopolysaccharides. HaCTL3 and HaCTL8 mRNA showed upregulation while HaCTL4, 5, and 6 mRNA downregulation post 20-Hydroxyecdysone (20E) injection. Our results indicate that the six novel C-type lectins of H. armigera may play important roles in defending against bacteria as PRRs and the hormone 20E can function in regulating immunity through lectins.