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Depression of immune reactions in insects
INSECT IMMUNITY
935
DEPRESSION OF IMMUNE REACTIONS IN INSECrS M. Breh~lin
Laboratoire de Pathologie Comparde, URA CNRS 1184, place E. Bataillon, 34095 Montpellier Cedex 05
In all organisms, depression of immune reactions is one of the main mechanisms which determine the outcome of the relations between a host and its potential parasites or pathogens. As in other animals, immune reactions in insects can be divided into two main steps: - - the recognition of non-self; - - the defence reactions s.l., that is, the mean by which insects try to destroy (or inhibit the development of) foreign bodies previously recognized as non-self. The processes by which parasites or pathogens are able to escape or to depress iasc~-: immunity are aimed at ~on-self recc~gailion as well as at the defence reactions themselves. Avoidance/depression of recognition.
Two main processes were developed to avoid non-self recognition. They are the composition of parasite surface in contact with the insect immune effectors and/or the depression of opsonic-like mechanisms. Depression of recognition due to the body wall composition.
This can be "passive" as, for instance, with the cuticle composition of Steinernematidae nematodes which were never encapsulated by locust or Galleria melionella haemocytes, or "active" as in Venturia canescens eggs which were coated with "foreign" (virus-like) particles at the time of
oviposition (Feddersen et al., 1986). These particles exhibited the same antigemc determinants as a 42-kDa protein synthesized by the host haemocytes (Berg et al., 1988). Now the question arises as to whether this non-reactivity of haemocytes was due to absence of recognition of the foreign body by the insect or to an impossibility for haemoc.vtes to build a capsule. Other inert or live foreign bodies injected together with Steinernematidae nematodes or V. canescens eggs were normally encapsulated (G6tz, personal communication; Breh61in, unpublished; Berg et al., 1988), which shows that haemocyte reactivity was not disturbed. " - , . . . . . . . . ' .... . . . . . . . immune recognition, t h a t ic t h p iveic r,f coagulocytes (or granulocytes) in contact with a foreign body (Ratcliffeand Gagea, 1976; Brchdlin et al., 1975; Breh61in and Hoffmann, 1980; see N. Ratcliffe, this Forum), was not observed with Steinernematidae. Finally, axenic nematodes did not alter the total or differential haemocyte counts as they would have done if they were recognized as foreign (Dunphy and Webster, 1985). So the absence of encapsulation of Steinernematidae nematodes in locust or in G. mellonella was the result of a lack of recognition rather than a simple failure m adhesiveness. Depression of humoral factors of immune recognition.
Dunphy and Webster (1988) showed that, in the presence of G. mellonella larval serum, lipopolysaccharides (LPS) could be released from the en-
936
34 th F O R U M I N I M M U N O L O G Y
tomopathogenic bacteria Xenorhabdus nematophilus. These LPS, in turn could trigger the pro-PO activating system (see M. Ashida and K. S6derh/fll, this Forum) and enhance recognition of the pathogen by insect haemocytes. These reactions could be depressed by the live bacteria themselves (Breh61in et ai., in preparation). We evidenced, in the culture medium of X. nematophilus, a factor which inhibited the in vitro activation of pro-PO by a serine protease such as trypsin. In the same way, the trigger effect of killed X. nematophilus on the natural pro-PO activating sys)em of locust haemocytes was inhibited if live X. nematophilus were added to the killed ones. These results have been obtained in in vitro experiments and it is now necessary to search for the secretion of such inhibiting factors in vivo. If present in parasitized insects, these factors could depress the opsonizing effect evidenced in the haemolymph of Locusta migratoria (Breh61in and Boemare, 1988; Breh61in et al., 1989) and account, in part, for the weak adhesion of X. nematophilus to their host haemocytcs. Inhibition of phenoloxidase activity produced by wasp poly-DNA virus in vivo was observed by Beckage et al. (1990) in lepidoptera larvae.
was not general but limited to the foreign bodies coated with venom prior to injection with the virus (Tanaka, 1987). Several parasites or pathogens induced extensive lysis of haemocytes. In Drosophila melanogaster, a depressive factor called lamellolysin was injected by the wasp Leptopilina heterotoma at oviposition. The lamellolysin made some haemocytes (the lamellocytes) lose their adhesiveness (Rizki and Rizki, 1984) and protected the eggs from crz~psule formation. Here again, the protection did not seem to be confined to the parasite eggs, but extendea to other foreign bodies (Nappi and Streams, 1969). Inhibitors of immune polypeptides synthesized by insects upon bacterial infections were isolated from Bacillus thuringiensis by Sid6n et al. (1979). This inhibitor affects cecropin A and B specifically. Its MW is 78 kDa. Another inhibitor is secreted by the entomopathogenic nematode Steinernema carpocapsae (G6tz et al., 1981), which destroys cecropins and attacins. Trials to purify this inhibitor have been unsuccessful to date. It seems to be proteinaceous in nature, devoid of lipid components and approximately of 50-kDa MW in gel filtration (Giilzow, 1986).
Depression of the defence reactions.
Discussion.
The main effectors of defence reactions are cellular (haemocytes in phagocytosis or encapsulation) or humoral (antibacterial polypeptides). These two kinds of immune reactions can be depressed by parasites or pathogens. Several mechanisms could account for the depression of cellular reactions. In ichneumonid parasitoids, poly-DNA virus injected by the female at oviposition protected the eggs from encapsulation. This depression was general, which means that other foreign bodies implanted in parasitized larvae were not encapsulated either (Davi~s and Vinson, 1988). In braconids, the virus alone was not efficient and secretion of the venom gland was needed in collaboration with virus. In this case, the protection
The depression of immunity is beneficial to a parasite or pathogen only if it protects the parasite/pathogen itself from the defence reactions of the insect. The host must remain able to fight against other parasites/pathogens which could disturb its physiology and in this way endanger the development of the first parasite/pathogen. This kind of depression is achieved by avoidance of recognition by the outer surface composition of the foreign body. At present we do not know if depression of humoral factors in immune recognition (the pro-PO system) affects all foreign bodies present in haemocoele or only the parasite/pathogen which induced the depression. A limited depression is also achieved by braconid parasitoids. On the other hand, lamellolysin in
INSECT iMMUNITY D. melanogaster or poly-DNA viruses of ichneumonids seem to induce general depression. We can speculate that the knowledge of the mechanism of this depression will tell us where is the benefit to the parasite. A peculiar instance is that of entomopathogenic nematodes Steinern e m a t i d a e and their symbiotic Xenorhabditidae bacteria. The lysis of cecropins and attacins would be induced after the end of the action of these immune po!ypeptides on the bacteria, which enter into the haemocoele through the wounds made by penetration of the parasite. When these bacteria are killed, the nematodes release their symbionts which are susceptible to attacins and cecropins and, at this time, the parasite must destroy the immune polypeptides. In the course of their
937
d,~,,,q-,~,~,*"* Xenorhabditidae seem able to produce in~bitors of the pro-PO system, which reduce their recogr,ition by haemocytes, and they also secrete antibiotics which avoid the development of other bacteria. While each of these reactions has been evidenced in vitro and (for most of them) in vivo, their pathway in the course of infection of an insect by the couple Steinernema-Xenorhabdus remains to be proven. The molecular basis of all these inhibition reactions is largely unknown. In my opinion, further research in this area is of importance because it will help us in the understanding of immunity in insects; moreover, it could lead to modification of the inhibitors so as to create specialized pathogens against specific pests.
References.
BECKAGE,N.E., MEfCALF,J.S., NESBIT,D.J., SCHLEIFER,K.W., ZETLAN,S.R. & DE BURON,1. 1990), Host hevnolymphmonophenoloxidase activity in parasitized Manduca sexta rvae and evidence for inhibition by wasp polydnavirus. Insect Biochem., 20, 285-294. BERG,R., SCHUCHMANN-FEDDERSEN,I. & SCHMIDT,O. (1988), Bacterial inf~tion induces a moth (E. kuhniella) protein which has antigenic similarity to virus-tiRe particle proteins of parasitoid wasp (Venturia canescens). J. Insect PhysioL, 34, 473-480. BRERELtn,M. & BOEMAaE,N. (1988), Immune recognition in insects: conflicting effects of autologous plasma and serum. J. Comp. Physiol., B157, 759-764. BREHELIN,M. & HOFFMANN,J. (1980), Phagocytosis of inert particles in Locusta migratoria and Galleria melioneUa: study of ultrastructure and clearance. J. Insect Physiol., 26, 103-111. BREnEUn,M., DRIF,L., BAUD,L. & BOEMARE,N. (1989), Activation of pro-phenoloxidase in insect haemo!ymph: cooperation between humoral and cellular factors in Locusta migratoria. Insect Biochem., 19, 301-307. DAvms, D.H. & VINSON,S.B. (1988), Interference with function of plasmatocytes of Heliothis virescens in vivo by calyx fluid of the parasitoid Campoletis sonorensis. Cell Tiss. Res., 251, 467-475. Dunpnv, G.B. & WEBSTER,J.M. (1985), Influence of Steinernem¢ feltiae (Filipjev) DDI36 strain on the humoral and haemocytic responses of Galleria mellonella (L.) larvae to selected bacteria. Parasitol., 91, 369-380. DUNPHY,G.B. & WEBSTER,J.M. (1988), Lipopolysaccharides of Xenorhabdus nematophilus (Enterobacteriaceae)and their haemocyte toxicity in non-immune Galleria melionella (Insecta Lepidoptera) larvae. J. gen. Microbiol., 134, 1017-1028. FEDDERSEn,I., SANDERS,K. & SC.MXOT,O. (1986), Virus-like particle with host protein like antigenic determinants protect an insect parasitoid from encapsulation. Experientia (Basel), 42, 1278-1280. GOTZ, P., BOMAN,A. & BOMAN,H.G. (1981), Interactions between insect immunity and an insect pathogenic nematode with symbiotic bacteria. Proc. roy. Soc. B, 212, 333-350. GOLZOW,A. (1986), Immune inhibition by the insect pathogenic nematode Neoplectana carpocapsae, in "ISDCI Conference" (P. G6tz). Develop. Com.p. Immunol., lO, 630. NApPI,A.J. & STREAMS,F.A. (1969), Haemocytic reactions of Drosophila melanogaster to me parasites Pseudochoila mellipes and P. bochei. J. Insect Physiol., 15, 1551-1566.
938
34th FOR UM IN I~IMUNOL OG Y
RATCLIFFE,N.A. & GAGEN,S.J. (1976), Cellular defense reactions of insect hemocytes in vivo: nodule formation and development in Galleria mellonella and Pieris brassicae larvae. J. Invertebr. Path., 28, 373-382. R:z~a, R.M. & RIZKI, T.M. (1984), Selective destruction of a host blood cell type by a parasitoid wasp. Proc. nat. Acad. Sci. (~ash.)~ 81, 6154-6~58. S!DEN, I., DALHAMMAR,G., TELANDER,B., BOMAN,H.G. & SOMMERVILLE,H. (1979), Virulence factors in Bacillus thuringiensis: purification and properties of a protein inhibitor of immunity in insects. J. gen. Microbiol., 114, 45-52. TANAKA, T. (1987), Effect of the venom of the endoparasitoid Apenteles kariyai on the cellular defence reaction of the host Pseudaletia separata. J. Insect Physiol., 33, . 413-420.
DUAL FUNCTIONS OF INSECT IMMUNITY PROTEINS IN DEFENCE AND DEVELOPMENT S. Natori
Faculty of Pharmaceutical Sciences, University of Tokyo, Bunkyo-ku, Tokyo 113
Sarcophaga lectin is a galactosebinding lectin induced in the haemolymph of third instar larvae of Sarcoohapa nereprina tfla~h flv~ in r e ¢ n n n c ~
to body injury. We have ~aurified this lectin and characterized itextensively. The following findings indicate that it is essential for elimination of foreign substances from the body. When sheep red blood cells were injected into the abdominal cavity of Sarcophaga larvae, they were gradually lysed, and disappeared from the haemolymph. However, when the blood cells were injected together with antibody against Sorcophaga lectin or the hapten sugar galactose, no elimination of the blood cells was detected and the cells remained in the abdominal cavity. Preimmune serum or non-hapten sugar glucose had no effect on lysis of the blood cells. Although haemolymph of uninjured larvae does not contain Sarcophagalectin, the larval body wall was inevitably injured when the red blood cells were injected. Therefore, Sarcophaga lectin is expected to be synthesized when any
foreign cells are introduced into the body. Elimination of the injected foreign cells was blocked by inactivation of this !ectin. Northern blot analysis using cDNA for Sarcophaga lectin revealed that the gene for this lectin is also expressed twice during normal ontogeny in the absence of any external stimulus: first in the embryonic stage and again in the early pupal stage. Therefore, it is likely that Sarcophaga lectin has dual functions, in defence and in development. In fact, we were able to detect Sarcophaga lectin in extracts of embryos and pupae of Sarcophaga, indicating that this defence protein plays a crucial role during development. Both vertebrates and invertebrates develop from embryos. During ontogenesis, only necessary cells proliferate to form body structures, while unnecessary cells are eliminated. So there must be some defence system that is effective for eliminating these unnecessary cells. The specific programmed cell death ob-
935
DEPRESSION OF IMMUNE REACTIONS IN INSECrS M. Breh~lin
Laboratoire de Pathologie Comparde, URA CNRS 1184, place E. Bataillon, 34095 Montpellier Cedex 05
In all organisms, depression of immune reactions is one of the main mechanisms which determine the outcome of the relations between a host and its potential parasites or pathogens. As in other animals, immune reactions in insects can be divided into two main steps: - - the recognition of non-self; - - the defence reactions s.l., that is, the mean by which insects try to destroy (or inhibit the development of) foreign bodies previously recognized as non-self. The processes by which parasites or pathogens are able to escape or to depress iasc~-: immunity are aimed at ~on-self recc~gailion as well as at the defence reactions themselves. Avoidance/depression of recognition.
Two main processes were developed to avoid non-self recognition. They are the composition of parasite surface in contact with the insect immune effectors and/or the depression of opsonic-like mechanisms. Depression of recognition due to the body wall composition.
This can be "passive" as, for instance, with the cuticle composition of Steinernematidae nematodes which were never encapsulated by locust or Galleria melionella haemocytes, or "active" as in Venturia canescens eggs which were coated with "foreign" (virus-like) particles at the time of
oviposition (Feddersen et al., 1986). These particles exhibited the same antigemc determinants as a 42-kDa protein synthesized by the host haemocytes (Berg et al., 1988). Now the question arises as to whether this non-reactivity of haemocytes was due to absence of recognition of the foreign body by the insect or to an impossibility for haemoc.vtes to build a capsule. Other inert or live foreign bodies injected together with Steinernematidae nematodes or V. canescens eggs were normally encapsulated (G6tz, personal communication; Breh61in, unpublished; Berg et al., 1988), which shows that haemocyte reactivity was not disturbed. " - , . . . . . . . . ' .... . . . . . . . immune recognition, t h a t ic t h p iveic r,f coagulocytes (or granulocytes) in contact with a foreign body (Ratcliffeand Gagea, 1976; Brchdlin et al., 1975; Breh61in and Hoffmann, 1980; see N. Ratcliffe, this Forum), was not observed with Steinernematidae. Finally, axenic nematodes did not alter the total or differential haemocyte counts as they would have done if they were recognized as foreign (Dunphy and Webster, 1985). So the absence of encapsulation of Steinernematidae nematodes in locust or in G. mellonella was the result of a lack of recognition rather than a simple failure m adhesiveness. Depression of humoral factors of immune recognition.
Dunphy and Webster (1988) showed that, in the presence of G. mellonella larval serum, lipopolysaccharides (LPS) could be released from the en-
936
34 th F O R U M I N I M M U N O L O G Y
tomopathogenic bacteria Xenorhabdus nematophilus. These LPS, in turn could trigger the pro-PO activating system (see M. Ashida and K. S6derh/fll, this Forum) and enhance recognition of the pathogen by insect haemocytes. These reactions could be depressed by the live bacteria themselves (Breh61in et ai., in preparation). We evidenced, in the culture medium of X. nematophilus, a factor which inhibited the in vitro activation of pro-PO by a serine protease such as trypsin. In the same way, the trigger effect of killed X. nematophilus on the natural pro-PO activating sys)em of locust haemocytes was inhibited if live X. nematophilus were added to the killed ones. These results have been obtained in in vitro experiments and it is now necessary to search for the secretion of such inhibiting factors in vivo. If present in parasitized insects, these factors could depress the opsonizing effect evidenced in the haemolymph of Locusta migratoria (Breh61in and Boemare, 1988; Breh61in et al., 1989) and account, in part, for the weak adhesion of X. nematophilus to their host haemocytcs. Inhibition of phenoloxidase activity produced by wasp poly-DNA virus in vivo was observed by Beckage et al. (1990) in lepidoptera larvae.
was not general but limited to the foreign bodies coated with venom prior to injection with the virus (Tanaka, 1987). Several parasites or pathogens induced extensive lysis of haemocytes. In Drosophila melanogaster, a depressive factor called lamellolysin was injected by the wasp Leptopilina heterotoma at oviposition. The lamellolysin made some haemocytes (the lamellocytes) lose their adhesiveness (Rizki and Rizki, 1984) and protected the eggs from crz~psule formation. Here again, the protection did not seem to be confined to the parasite eggs, but extendea to other foreign bodies (Nappi and Streams, 1969). Inhibitors of immune polypeptides synthesized by insects upon bacterial infections were isolated from Bacillus thuringiensis by Sid6n et al. (1979). This inhibitor affects cecropin A and B specifically. Its MW is 78 kDa. Another inhibitor is secreted by the entomopathogenic nematode Steinernema carpocapsae (G6tz et al., 1981), which destroys cecropins and attacins. Trials to purify this inhibitor have been unsuccessful to date. It seems to be proteinaceous in nature, devoid of lipid components and approximately of 50-kDa MW in gel filtration (Giilzow, 1986).
Depression of the defence reactions.
Discussion.
The main effectors of defence reactions are cellular (haemocytes in phagocytosis or encapsulation) or humoral (antibacterial polypeptides). These two kinds of immune reactions can be depressed by parasites or pathogens. Several mechanisms could account for the depression of cellular reactions. In ichneumonid parasitoids, poly-DNA virus injected by the female at oviposition protected the eggs from encapsulation. This depression was general, which means that other foreign bodies implanted in parasitized larvae were not encapsulated either (Davi~s and Vinson, 1988). In braconids, the virus alone was not efficient and secretion of the venom gland was needed in collaboration with virus. In this case, the protection
The depression of immunity is beneficial to a parasite or pathogen only if it protects the parasite/pathogen itself from the defence reactions of the insect. The host must remain able to fight against other parasites/pathogens which could disturb its physiology and in this way endanger the development of the first parasite/pathogen. This kind of depression is achieved by avoidance of recognition by the outer surface composition of the foreign body. At present we do not know if depression of humoral factors in immune recognition (the pro-PO system) affects all foreign bodies present in haemocoele or only the parasite/pathogen which induced the depression. A limited depression is also achieved by braconid parasitoids. On the other hand, lamellolysin in
INSECT iMMUNITY D. melanogaster or poly-DNA viruses of ichneumonids seem to induce general depression. We can speculate that the knowledge of the mechanism of this depression will tell us where is the benefit to the parasite. A peculiar instance is that of entomopathogenic nematodes Steinern e m a t i d a e and their symbiotic Xenorhabditidae bacteria. The lysis of cecropins and attacins would be induced after the end of the action of these immune po!ypeptides on the bacteria, which enter into the haemocoele through the wounds made by penetration of the parasite. When these bacteria are killed, the nematodes release their symbionts which are susceptible to attacins and cecropins and, at this time, the parasite must destroy the immune polypeptides. In the course of their
937
d,~,,,q-,~,~,*"* Xenorhabditidae seem able to produce in~bitors of the pro-PO system, which reduce their recogr,ition by haemocytes, and they also secrete antibiotics which avoid the development of other bacteria. While each of these reactions has been evidenced in vitro and (for most of them) in vivo, their pathway in the course of infection of an insect by the couple Steinernema-Xenorhabdus remains to be proven. The molecular basis of all these inhibition reactions is largely unknown. In my opinion, further research in this area is of importance because it will help us in the understanding of immunity in insects; moreover, it could lead to modification of the inhibitors so as to create specialized pathogens against specific pests.
References.
BECKAGE,N.E., MEfCALF,J.S., NESBIT,D.J., SCHLEIFER,K.W., ZETLAN,S.R. & DE BURON,1. 1990), Host hevnolymphmonophenoloxidase activity in parasitized Manduca sexta rvae and evidence for inhibition by wasp polydnavirus. Insect Biochem., 20, 285-294. BERG,R., SCHUCHMANN-FEDDERSEN,I. & SCHMIDT,O. (1988), Bacterial inf~tion induces a moth (E. kuhniella) protein which has antigenic similarity to virus-tiRe particle proteins of parasitoid wasp (Venturia canescens). J. Insect PhysioL, 34, 473-480. BRERELtn,M. & BOEMAaE,N. (1988), Immune recognition in insects: conflicting effects of autologous plasma and serum. J. Comp. Physiol., B157, 759-764. BREHELIN,M. & HOFFMANN,J. (1980), Phagocytosis of inert particles in Locusta migratoria and Galleria melioneUa: study of ultrastructure and clearance. J. Insect Physiol., 26, 103-111. BREnEUn,M., DRIF,L., BAUD,L. & BOEMARE,N. (1989), Activation of pro-phenoloxidase in insect haemo!ymph: cooperation between humoral and cellular factors in Locusta migratoria. Insect Biochem., 19, 301-307. DAvms, D.H. & VINSON,S.B. (1988), Interference with function of plasmatocytes of Heliothis virescens in vivo by calyx fluid of the parasitoid Campoletis sonorensis. Cell Tiss. Res., 251, 467-475. Dunpnv, G.B. & WEBSTER,J.M. (1985), Influence of Steinernem¢ feltiae (Filipjev) DDI36 strain on the humoral and haemocytic responses of Galleria mellonella (L.) larvae to selected bacteria. Parasitol., 91, 369-380. DUNPHY,G.B. & WEBSTER,J.M. (1988), Lipopolysaccharides of Xenorhabdus nematophilus (Enterobacteriaceae)and their haemocyte toxicity in non-immune Galleria melionella (Insecta Lepidoptera) larvae. J. gen. Microbiol., 134, 1017-1028. FEDDERSEn,I., SANDERS,K. & SC.MXOT,O. (1986), Virus-like particle with host protein like antigenic determinants protect an insect parasitoid from encapsulation. Experientia (Basel), 42, 1278-1280. GOTZ, P., BOMAN,A. & BOMAN,H.G. (1981), Interactions between insect immunity and an insect pathogenic nematode with symbiotic bacteria. Proc. roy. Soc. B, 212, 333-350. GOLZOW,A. (1986), Immune inhibition by the insect pathogenic nematode Neoplectana carpocapsae, in "ISDCI Conference" (P. G6tz). Develop. Com.p. Immunol., lO, 630. NApPI,A.J. & STREAMS,F.A. (1969), Haemocytic reactions of Drosophila melanogaster to me parasites Pseudochoila mellipes and P. bochei. J. Insect Physiol., 15, 1551-1566.
938
34th FOR UM IN I~IMUNOL OG Y
RATCLIFFE,N.A. & GAGEN,S.J. (1976), Cellular defense reactions of insect hemocytes in vivo: nodule formation and development in Galleria mellonella and Pieris brassicae larvae. J. Invertebr. Path., 28, 373-382. R:z~a, R.M. & RIZKI, T.M. (1984), Selective destruction of a host blood cell type by a parasitoid wasp. Proc. nat. Acad. Sci. (~ash.)~ 81, 6154-6~58. S!DEN, I., DALHAMMAR,G., TELANDER,B., BOMAN,H.G. & SOMMERVILLE,H. (1979), Virulence factors in Bacillus thuringiensis: purification and properties of a protein inhibitor of immunity in insects. J. gen. Microbiol., 114, 45-52. TANAKA, T. (1987), Effect of the venom of the endoparasitoid Apenteles kariyai on the cellular defence reaction of the host Pseudaletia separata. J. Insect Physiol., 33, . 413-420.
DUAL FUNCTIONS OF INSECT IMMUNITY PROTEINS IN DEFENCE AND DEVELOPMENT S. Natori
Faculty of Pharmaceutical Sciences, University of Tokyo, Bunkyo-ku, Tokyo 113
Sarcophaga lectin is a galactosebinding lectin induced in the haemolymph of third instar larvae of Sarcoohapa nereprina tfla~h flv~ in r e ¢ n n n c ~
to body injury. We have ~aurified this lectin and characterized itextensively. The following findings indicate that it is essential for elimination of foreign substances from the body. When sheep red blood cells were injected into the abdominal cavity of Sarcophaga larvae, they were gradually lysed, and disappeared from the haemolymph. However, when the blood cells were injected together with antibody against Sorcophaga lectin or the hapten sugar galactose, no elimination of the blood cells was detected and the cells remained in the abdominal cavity. Preimmune serum or non-hapten sugar glucose had no effect on lysis of the blood cells. Although haemolymph of uninjured larvae does not contain Sarcophagalectin, the larval body wall was inevitably injured when the red blood cells were injected. Therefore, Sarcophaga lectin is expected to be synthesized when any
foreign cells are introduced into the body. Elimination of the injected foreign cells was blocked by inactivation of this !ectin. Northern blot analysis using cDNA for Sarcophaga lectin revealed that the gene for this lectin is also expressed twice during normal ontogeny in the absence of any external stimulus: first in the embryonic stage and again in the early pupal stage. Therefore, it is likely that Sarcophaga lectin has dual functions, in defence and in development. In fact, we were able to detect Sarcophaga lectin in extracts of embryos and pupae of Sarcophaga, indicating that this defence protein plays a crucial role during development. Both vertebrates and invertebrates develop from embryos. During ontogenesis, only necessary cells proliferate to form body structures, while unnecessary cells are eliminated. So there must be some defence system that is effective for eliminating these unnecessary cells. The specific programmed cell death ob-