Pattern recognition proteins in Manduca sexta plasma

Yu, Xiao‐Qiang; Zhu, Yifei; Ma, Congcong; Fabrick, Jeffrey A.; Kanost, Michael R.

doi:10.1016/s0965-1748(02)00091-7

Cited by 194 publications

(124 citation statements)

References 62 publications

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“…The susceptibility of M. sexta larvae to a variety of entomopathogenic bacterial species [1][2][3][4][5] , as well as the wealth of information available regarding the insect's immune system [6][7][8] , and the pending genome sequence 9 make it a good model organism for use in studying host-microbe interactions during pathogenesis. In addition, M. sexta larvae are relatively large and easy to manipulate and maintain in the laboratory relative to other susceptible insect species.…”

Section: Introductionmentioning

confidence: 99%

“…The insect responds to infection via both humoral and cellular responses. The humoral response includes recognition of bacterial-associated patterns and subsequent production of various antimicrobial peptides 7 ; the expression of genes encoding these peptides can be monitored subsequent to direct infection via RNA extraction and quantitative PCR 13 . The cellular response to infection involves nodulation, encapsulation, and phagocytosis of infectious agents by hemocytes 6 .…”

Section: Introductionmentioning

confidence: 99%

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Rearing and Injection of <em>Manduca sexta</em> Larvae to Assess Bacterial Virulence

Hussa

Goodrich-Blair

2012

JoVE

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Manduca sexta, commonly known as the tobacco hornworm, is considered a significant agricultural pest, feeding on solanaceous plants including tobacco and tomato. The susceptibility of M. sexta larvae to a variety of entomopathogenic bacterial species [1][2][3][4][5] , as well as the wealth of information available regarding the insect's immune system [6][7][8] , and the pending genome sequence 9 make it a good model organism for use in studying host-microbe interactions during pathogenesis. In addition, M. sexta larvae are relatively large and easy to manipulate and maintain in the laboratory relative to other susceptible insect species. Their large size also facilitates efficient tissue/hemolymph extraction for analysis of the host response to infection.The method presented here describes the direct injection of bacteria into the hemocoel (blood cavity) of M. sexta larvae. This approach can be used to analyze and compare the virulence characteristics of various bacterial species, strains, or mutants by simply monitoring the time to insect death after injection. This method was developed to study the pathogenicity of Xenorhabdus and Photorhabdus species, which typically associate with nematode vectors as a means to gain entry into the insect. Entomopathogenic nematodes typically infect larvae via natural digestive or respiratory openings, and release their symbiotic bacterial contents into the insect hemolymph (blood) shortly thereafter 10 . The injection method described here bypasses the need for a nematode vector, thus uncoupling the effects of bacteria and nematode on the insect. This method allows for accurate enumeration of infectious material (cells or protein) within the inoculum, which is not possible using other existing methods for analyzing entomopathogenesis, including nicking 11 and oral toxicity assays 12 . Also, oral toxicity assays address the virulence of secreted toxins introduced into the digestive system of larvae, whereas the direct injection method addresses the virulence of wholecell inocula.The utility of the direct injection method as described here is to analyze bacterial pathogenesis by monitoring insect mortality. However, this method can easily be expanded for use in studying the effects of infection on the M. sexta immune system. The insect responds to infection via both humoral and cellular responses. The humoral response includes recognition of bacterial-associated patterns and subsequent production of various antimicrobial peptides 7 ; the expression of genes encoding these peptides can be monitored subsequent to direct infection via RNA extraction and quantitative PCR 13 . The cellular response to infection involves nodulation, encapsulation, and phagocytosis of infectious agents by hemocytes 6 . To analyze these responses, injected insects can be dissected and visualized by microscopy 13,14 . Video LinkThe video component of this article can be found at

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Rearing and Injection of <em>Manduca sexta</em> Larvae to Assess Bacterial Virulence

Hussa

Goodrich-Blair

2012

JoVE

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“…Therefore, there must be other PRRs involved in innate immune recognition in Drosophila. In the tobacco hornworm Manduca sexta, a family of immulectins, which are members of the C-type lectin superfamily, functions as important PRRs to promote hemocyte encapsulation and stimulate prophenoloxidase activation (Yu et al, 1999;Yu and Kanost, 2000;Yu et al, 2002;Ling and Yu, 2006a). C-type lectins are calcium-dependent carbohydrate-binding proteins that can bind terminal sugars on the surface of microorganisms.…”

Section: Introductionmentioning

confidence: 99%

Drosophila C-type lectins enhance cellular encapsulation

Ling

2007

Molecular Immunology

132

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C-type lectins are calcium-dependent carbohydrate binding proteins, and animal C-type lectins participate in innate immunity and cell-cell interactions. In the fruit fly Drosophila melanogaster, more than 30 genes encode C-type lectin domains. However, functions of Drosophila C-type lectins in innate immunity are not well understood. This study is to investigate whether two Drosophila Ctype lectins, CG33532 and CG335333 (designated as DL2 and DL3, respectively), are involved in innate immune responses. Recombinant DL2 and DL3 were expressed and purified. Both DL2 and DL3 agglutinated Gram-negative Escherichia coli in a calcium-dependent manner. Though DL2 and DL3 are predicted to be secreted proteins, they were detected on the surface of Drosophila hemocytes, and recombinant DL2 and DL3 also directly bound to hemocytes. Coating of agarose beads with recombinant DL2 and DL3 enhanced their encapsulation and melanization by Drosophila hemocytes in vitro. However, hemocyte encapsulation was blocked when the lectin-coated beads were preincubated with rat polyclonal antibody specific for DL2 or DL3. Our results suggest that DL2 and DL3 may act as pattern recognition receptors to mediate hemocyte encapsulation and melanization by directly recruiting hemocytes to the lectin-coated surface.

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“…Pattern recognition molecules also abound in insects (14), and recently, it has been shown that apolipophorin III (apoLp-III), a major exchangeable lipid transport molecule found in the blood (hemolymph), may also play a crucial role in the innate immune response and act in pattern recognition (15). ApoLp-III binds to Gram-positive bacteria and to lipoteichoic acid (LTA) (16,17), and as in mammals, apoLp-III can also bind and detoxify LPS (18,19) and promote phagocytosis (20).…”

mentioning

confidence: 99%

A Novel Role for an Insect Apolipoprotein (Apolipophorin III) in β-1,3-Glucan Pattern Recognition and Cellular Encapsulation Reactions

Whitten

Tew

Lee

et al. 2004

The Journal of Immunology

203

163

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Lipoproteins and molecules for pattern recognition are centrally important in the innate immune response of both vertebrates and invertebrates. Mammalian apolipoproteins such as apolipoprotein E (apoE) are involved in LPS detoxification, phagocytosis, and possibly pattern recognition. The multifunctional insect protein, apolipophorin III (apoLp-III), is homologous to apoE. In this study we describe novel roles for apoLp-III in pattern recognition and multicellular encapsulation reactions in the innate immune response, which may be of direct relevance to mammalian systems. It is known that apoLp-III stimulates antimicrobial peptide production in insect blood, enhances phagocytosis by insect blood cells (hemocytes), and binds and detoxifies LPS and lipoteichoic acid. In the present study we show that apoLp-III from the greater wax moth, Galleria mellonella, also binds to fungal conidia and β-1,3-glucan and therefore may act as a pattern recognition molecule for multiple microbial and parasitic invaders. This protein also stimulates increases in cellular encapsulation of nonself particles by the blood cells and exerts shorter term, time-dependent, modulatory effects on cell attachment and spreading. All these responses are dose dependent, occur within physiological levels, and, with the notable exception of β-glucan binding, are only observed with the lipid-associated form of apoLp-III. Preliminary studies also established a beneficial role for apoLp-III in the in vivo response to an entomopathogenic fungus. These data suggest a wide range of immune functions for a multiple specificity pattern recognition molecule and may provide a useful model for identifying further potential roles for homologous proteins in mammalian immunology, particularly in terms of fungal infections, pneumoconiosis, and granulomatous reactions.

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Pattern recognition proteins in Manduca sexta plasma

Cited by 194 publications

References 62 publications

Rearing and Injection of <em>Manduca sexta</em> Larvae to Assess Bacterial Virulence

Rearing and Injection of <em>Manduca sexta</em> Larvae to Assess Bacterial Virulence

Drosophila C-type lectins enhance cellular encapsulation

A Novel Role for an Insect Apolipoprotein (Apolipophorin III) in β-1,3-Glucan Pattern Recognition and Cellular Encapsulation Reactions

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