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M R Strand

Publications and source records attributed to M R Strand.

34 records · Page 2Linked to original sources
PubMed1998

Shifts in the life history of parasitic wasps correlate with pronounced alterations in early development.

Developmental processes have been traditionally viewed to be invariant within higher taxa. However, examples are known whereby closely related species exhibit alterations in early embryogenesis yet appear very similar as adults. Such developmental changes are thought to occur in response to shifts in life history. In insects, the regulation of embryonic development has been intensively studied in model species like Drosophila melanogaster. Previous comparative studies suggest that the developmental processes documented in Drosophila well describe embryogenesis of advanced, holometabolous, insects generally. There have been few attempts, however, to take into account how life history has influenced early development of insects or to characterize early development of species with life histories fundamentally different from flies. Here we compared early development of two species from the same family of parasitic wasps that exhibit very different life histories. Bracon hebetor is an ectoparasite that lays large, yolky eggs on the integument of its host that develop much like the free-living honeybee and Drosophila. In contrast, Aphidius ervi is an endoparasite that lays small and apparently yolk-free eggs that develop in the hemocoel of the host. This wasp exhibits a radically different mode of early development at both the cellular and molecular level from B. hebetor. The developmental changes in A. ervi reflect functional adaptations for its derived life history and argue that departures from the fly paradigm may occur commonly among insects whose eggs develop under conditions different from typical terrestrial species.

Animals↗
PubMed1998

Development of polyembryonic insects: a major departure from typical insect embryogenesis.

The parasitic wasp Copidosoma floridanum represents the most extreme form of polyembryonic development known, forming up to 2000 embryos from a single egg. To understand the mechanisms of embryonic patterning in polyembryonic wasps and the evolutionary changes that led to this form of development we have analyzed embryonic development at the cellular level using confocal and scanning electron microscopy. C. floridanum embryogenesis can be divided into three phases: (1) early cleavage that leads to formation of a primary morula, (2) a proliferative phase that involves partitioning of embryonic cells into thousands of morulae, and (3) morphogenesis whereby individual embryos develop into larvae. This developmental program represents a major departure from typical insect embryogenesis, and we describe several features of morphogenesis unusual for insects. The early development of polyembryonic wasps, which likely evolved in association with a shift in life history to endoparasitism, shows several analogies with mammalian embryogenesis, including early separation of extraembryonic and embryonic cell lineages, formation of a morula and embryonic compaction. However, the late morphogenesis of polyembryonic wasps proceeds in a fashion conserved in all insects. Collectively, this suggests a lack of developmental constraints in early development, but a strong conservation of the phylotypic stage.

Animals↗
PubMed1998

Follow-up of protein release from Pseudoplusia includens hemocytes: a first step toward identification of factors mediating encapsulation in insects.

Hemocyte types involved in encapsulation were characterized using monoclonal antibodies (mAbs). This approach revealed that four hemocyte types in Pseudoplusia includens could be classified into two antigenically distinct cell lines. The first line comprised granulocytes (GR) and spherulocytes (SP) and the second line comprised plasmatocytes (PL) and oenocytoids (OE). One of the mAbs labeled a subpopulation of plasmatocytes that spread on culture plate surfaces. This subclass represented approximately 70% of all plasmatocytes. The cytoplasmic punctate staining of granulocytes clearly decreased upon short term culture, suggesting the associated antigens were released into the culture medium during cell spreading. A follow-up of protein secretion into culture medium by Western blotting confirmed this hypothesis for both granulocytes and plasmatocytes. In a few cases, discharged proteins exhibited a short half-life suggesting they may behave as regulatory molecules. Among them, plasmatocyte proteins of +/-25 kDa might be mobilized at an early stage of encapsulation. The same proteins appeared to accumulate at the periphery of the median plasmatocyte multilayer in late capsules. This location coincides with where an outer monolayer of granulocytes attaches and causes termination of capsule growth. These preliminary results raise the possibility that released proteins regulate hemocyte recruitment during encapsulation.

Animals↗
PubMed1997

Isolation and identification of a plasmatocyte-spreading peptide from the hemolymph of the lepidopteran insect Pseudoplusia includens.

Insect blood cells (hemocytes) play an essential role in defense against parasites and other pathogenic organisms that infect insects. A key class of hemocytes involved in insect cellular immunity is plasmatocytes. Here we describe the isolation and identification of a peptide from the moth Pseudoplusia includens that mediates the spreading of plasmatocytes to foreign surfaces. This peptide, designated plasmatocyte-spreading peptide (PSP1), contains 23 amino acid residues in the following sequence: H-ENFNGGCLAGYMRTADGRCKPTF-OH. In vitro assays using the synthetic peptide at concentrations >/=2 nM induced plasmatocytes from P. includens to spread on the surface of culture dishes. Injection of this peptide into P. includens larvae caused a transient depletion of plasmatocytes from circulation. Labeling studies indicated that this peptide induced 75% of plasmatocytes that were double-labeled by the monoclonal antibodies 49G3A3 and 43E9A8 to spread, whereas plasma induced significantly more plasmatocytes to spread. This suggests that only a certain subpopulation of plasmatocytes responds to the peptide and that other peptidyl factors mediate plasmatocyte adhesion responses.

Amino Acid Sequence↗
PubMed1997

Two Microplitis demolitor polydnavirus mRNAs expressed in hemocytes of Pseudoplusia includens contain a common cysteine-rich domain.

Microplitis demolitor is a polydnavirus-carrying wasp that parasitizes the larval stage of Pseudoplusia includens. A previous study indicated that M. demolitor polydnavirus (MdPDV) infects primarily hemocytes in parasitized hosts. Thereafter, several alterations that compromise the immune response of P. includens toward the developing parasitoid occur in hemocytes. In this study, we identified two MdPDV mRNAs (1.0 and 1.5 kb) expressed in P. includens hemocytes that have homology to the viral genomic clone pMd-2. Corresponding 1.0- and 1.5-kb cDNA clones (MdPi455 and MdPi59) were isolated from an MdPDV-infected hemocyte cDNA library. Nucleotide sequence analysis of the cDNA clones confirmed that the 1.5- and 1.0-kb mRNAs have significant regions of homology. Sequence alignment revealed that the gene, OMd1.0, encoding the 1.0-kb mRNA is present in pMd-2. This gene contains two introns and three exons that agree with the sequence for MdPi455. In contrast, the 1.5-kb mRNA is likely encoded by a related gene located on the same MdPDV genomic DNA as is OMd1.0. The predicted peptide sequences for the 1.0- and 1.5-kb transcripts contain a cysteine-rich region at their 5' ends that have some similarity with epidermal growth factor-like motifs. Hybridization studies revealed that both mRNAs are expressed in granular cells and plasmatocytes, the primary classes of hemocytes involved in defense against M. demolitor and other parasites.

Amino Acid Sequence↗
PubMed1996

Intensive linkage mapping in a wasp (Bracon hebetor) and a mosquito (Aedes aegypti) with single-strand conformation polymorphism analysis of random amplified polymorphic DNA markers.

The use of random amplified polymorphic DNA from the polymerase chain reaction (RAPD-PCR) allows efficient construction of saturated linkage maps. However, when analyzed by agarose gel electrophoresis, most RAPD-PCR markers segregate as dominant alleles, reducing the amount of linkage information obtained. We describe the use of single strand conformation polymorphism (SSCP) analysis of RAPD markers to generate linkage maps in a haplodiploid parasitic wasp Bracon (Habrobracon) hebetor and a diploid mosquito. Aedes aegypti. RAPD-SSCP analysis revealed segregation of codominant alleles at markers that appeared to segregate as dominant (band presence/band absence) markers or appeared invariant on agarose gels. Our SSCP protocol uses silver staining to detect DNA fractionated on large thin polyacrylamide gels and reveals more polymorphic markers than agarose gel electrophoresis. In B. hebetor, 79 markers were mapped with 12 RAPD primers in six weeks; in A aygpti, 94 markers were mapped with 10 RAPD primers in five weeks. Forty-five percent of markers segregated as codominant loci in B. hebetor, while 11% segregated as codominant loci in A. aegypti. SSCP analysis of RAPD-PCR markers offers a rapid and inexpensive means of constructing intensive linkage maps of many species.

Aedes↗
PubMed1996

Granular cells are required for encapsulation of foreign targets by insect haemocytes.

Haemocytes play an essential role in defending invertebrates against pathogens and parasites that enter their haemocoel. A primary defense response is encapsulation; a process in which haemocytes attach to the foreign organism and kill it. Whether encapsulation requires cooperation between specific subpopulations of haemocytes is unknown. Using purified subpopulations of haemocytes and an in vitro encapsulation assay, we investigated the process of capsule formation in the insect Pseudoplusia includens. Immunocytochemical staining revealed that capsule formation involves a three step process. Encapsulation began when granular cells attached to the foreign target. This was followed by attachment of multiple layers of plasmatocytes. Termination of capsule formation occurred when a subpopulation of granular cells formed a monolayer around the periphery of the capsule. Neither granular cells nor plasmatocytes were capable of forming a capsule independently. However, plasmatocytes encapsulated targets if granular cells were present or if targets were preincubated in medium conditioned by granular cells. The effect of granular cell-conditioned medium could be blocked by the addition of the cell adhesion recognition sequence, RGDS, but not by RGES. These results demonstrate experimentally that granular cells are required for encapsulation of foreign targets by plasmatocytes in vitro, and that the role of granular cells in this process involves an RGD-dependent cell adhesion mechanism.

Animals↗
PubMed1995

Microplitis demolitor polydnavirus induces apoptosis of a specific haemocyte morphotype in Pseudoplusia includens.

Microplitis demolitor polydnavirus (MdPDV) is associated with Microplitis demolitor, a parasitic wasp that attacks the larval stage of the lepidopteran Pseudoplusia includens. Previously, we observed that MdPDV induced several alterations in the granular cells and plasmatocytes of P. includens, the primary haemocytes involved in regulating the cellular immune response toward M. demolitor and other parasites. In examining the mechanisms underlying immunosuppression of this host, we found that MdPDV induced apoptosis of granular cells. Granular cells underwent apoptosis both when virus was injected into the haemocoel of P. includens larvae and after infection with MdPDV in vitro. Characteristics of MdPDV-induced apoptosis included condensation of chromatin, cell surface blebbing and fragmentation of DNA into a 200 bp ladder. Although MdPDV induced changes in the ability of plasmatocytes to adhere to foreign surfaces, apoptosis of this morphotype was not observed. Examples from the literature suggest that some viruses promote their own survival by suppressing apoptosis of host cells. However, since polydnaviruses are likely to be transmitted vertically, we suggest that MdPDV promotes its own survival by inducing apoptosis of host immune cells which would otherwise kill the developing M. demolitor egg.

Animals↗
PubMed1995

Immunological basis for compatibility in parasitoid-host relationships.

The insect immune system serves as a key defense against attack by parasitoids. Incompatible hosts often eliminate parasitoids by encapsulation, a process in which hemocytes form a multilayered envelope around the invading organism. Capsule formation involves cooperation between one or more classes of hemocytes and is likely mediated by cytokines and adhesion molecules. Reciprocally, parasitoids have evolved a variety of strategies for overcoming host immune responses. Some parasitoids passively avoid elimination by developing in locations inaccessible to host hemocytes or by possessing surface features that fail to elicit an immune response. Other species actively disrupt the host immune system by injecting specific factors into the host at oviposition. In particular, polydnaviruses associated with several taxa of parasitoids disrupt capsule formation by killing hemocytes or altering their ability to adhere to foreign surfaces. These symbionts have likely played a critical role in evolution of host range and in defining parasitoid-host compatibility.

Animals↗
PubMed1994

Development and partial characterization of monoclonal antibodies to venom of the parasitoid Microplitis demolitor.

The venom of Microplitis demolitor consists of a mixture of proteins. On native PAGE gels three major proteins designated a, b, and g were detected, while on SDS-PAGE gels two major proteins of M(r) 64.5 and 30.8 kD and several minor proteins were detected. No proteins smaller than M(r) 30.8 kD were present. Murine monoclonal antibodies were generated against different venom components. Analysis by Western blot of venom proteins separated on native and SDS-PAGE gels confirmed that antibodies from seven hybridoma lines recognized venom components. Two of the seven hybridoma lines reacted specifically with protein g on native PAGE gels and the M(r) 30.8 k protein on SDS-PAGE gels, while four other lines cross-reacted with these and other venom proteins. The final hybridoma line reacted with protein a when venom was separated on native PAGE gels and an array of proteins when venom was separated on SDS-PAGE gels. Using an enzyme-immunoassay and specific monoclonal antibodies, M. demolitor females were estimated to inject 0.02-0.05 venom gland reservoir equivalents into its host, Pseudoplusia includens, at oviposition. Venom proteins persisted in host hemolymph for 6-12 h before dropping to undetectable levels.

Animals↗
PubMed1994

Separation and behavior in vitro of hemocytes from the moth, Pseudoplusia includens.

Hemocytes collected from larvae of Pseudoplusia includens (Lepidoptera:Noctuidae) were separated by centrifugation on Percoll cushions. The procedure resulted in 95% purity of plasmatocytes and greater than 99% purity of granular and spherule cells. Medium supplemented with chicken serum enhanced cell viability and promoted spreading of plasmatocytes. Cell-free plasma and medium preconditioned by plasmatocytes or granular cells stabilized cells in vitro and also accelerated spreading of plasmatocytes relative to medium supplemented with chicken serum. Oenocytoids were the only morphotype that exhibited endogenous phenoloxidase activity, while granular cells and plasmatocytes were the only cells that endocytosed fluorescent beads in vitro. Granular cells and plasmatocytes ingested fluorescently labelled beads, both in mixed populations of hemocytes and after separation. Plasmatocytes were the only morphotype that encapsulated large foreign targets in vitro following separation. Separated granular cells attached and spread on the surface of foreign targets but never formed a multilayered capsule.

Animals↗
PubMed1994

Microplitis demolitor polydnavirus infects and expresses in specific morphotypes of Pseudoplusia includens haemocytes.

Microplitis demolitor is a polydnavirus-carrying wasp that parasitizes the larval stage of Pseudoplusia includens. M. demolitor eggs are never encapsulated by host haemocytes when coinfected with its associated polyndnavirus (MdPDV) whereas eggs are encapsulated within 36 h when injected into hosts without virus. In this study, infection of specific classes of P. includens haemocytes by MdPDV was examined. Electron microscopic studies indicated that MdPDV entered all haemocyte morphotypes. Northern blot analysis revealed that similar size classes of viral mRNAs were produced in granular cells, plasmatocytes and spherule cells. Expression of a 1.6 kb MdPDV mRNA in haemocytes from parasitized hosts was detectable by in situ hybridization at 2 h post-parasitism (p.p.) and continued through until day 6 p.p. By 12 h p.p., viral expression was detected in greater than 80% of the haemocytes in circulation but thereafter the percentage of haemocytes exhibiting a hybridization signal declined. Similar patterns were observed in haemocytes from larvae injected with calyx fluid or MdPDV plus venom. Granular cells and plasmatocytes from unparasitized larvae were purified on Percoll cushions and maintained in vitro. Both morphotypes were successfully infected with MdPDV and exhibited changes in morphology and adhesiveness very similar to cells from parasitized hosts. Cell-free plasma from parasitized larvae had a variable effect on haemocyte adhesion. Haemocytes cultured in plasma from 1 or 4 day p.p. larvae rapidly spread whereas cells cultured in 7 day p.p. plasma did not. Reciprocally, adhesion of haemocytes from parasitized larvae could not be rescued by cell-free plasma from unparasitized larvae. Together, these data suggest that disruption of the host encapsulation response is medicated primarily by direct infection of granular cells and plasmatocytes by MdPDV.

Animals↗
PubMed1993

Ecdysteroid induction of embryonic morphogenesis in a parasitic wasp.

Development of the parasitic wasp Copidosoma floridanum (Hymenoptera: Encyrtidae) is unusual in two ways. As many as 3000 embryos are formed from a single egg and embryonic morphogenesis is closely synchronized with the onset of the metamorphosis of its host. Given this extreme synchrony between parasite and host development, we undertook a series of experiments to determine whether host endocrine factors regulate C. floridanum embryonic morphogenesis. Here we report that C. floridanum embryos must develop for 9 days before acquiring the competence to undergo morphogenesis. Furthermore, several pieces of evidence suggest that ecdysteroids of host origin regulate induction of C. floridanum morphogenesis. First, competent embryos initiated morphogenesis when transplanted into host larvae and pupae, host stages possessing elevated ecdysteroid titers, but not when transplanted into adult moths. Second, morphogenesis was arrested by ablation of the host's source of ecdysone, but could be rescued by injection of 20-hydroxyecdysone in a dose-dependent manner. Finally, a segment of DNA encoding a zinc finger nearly identical in sequence to a portion of the ecdysone receptor of Drosophila melanogaster was isolated and characterized from C. floridanum. This putative ecdysone receptor probe indicated that expression of this gene was correlated with the initiation of C. floridanum embryonic morphogenesis. The temporal pattern of putative receptor RNA accumulation increased in association with the onset of morphogenesis, while the spatial pattern of expression was associated with the invagination of cells forming the gastrula. Together, these data suggest that ecdysone of host origin is directly involved in the induction of C. floridanum embryonic morphogenesis.

Amino Acid Sequence↗
PubMed1992

Persistence and expression of Microplitis demolitor polydnavirus in Pseudoplusia includens.

Persistence and expression of Microplitis demolitor polydnavirus (MdPDV) was examined in parasitized and virus-injected Pseudoplusia includens larvae. Viral DNA persisted in P. includens larvae for 6 days, but no increase in the amount of viral DNA present was detected. Viral transcripts were observed in parasitized and virus-injected larvae 4 h post-parasitism and expression continued for 6 days. When specific host tissues were examined, more viral DNA and RNA was detected in haemocytes than in the gut, nervous system and fat body. 32P-labelled MdPDV DNA hybridized to approximately six different size classes of mRNAs on Northern blots of RNA from haemocytes of parasitized larvae. MdPDV transcription was first detected in haemocytes at 4 h post-parasitism and continued for 6 days. Similar transcripts were observed in haemocytes from larvae that had been injected with calyx fluid or MdPDV plus venom. First-strand cDNA probes of haemocyte-specific MdPDV transcripts hybridized to only certain MdPDV viral DNAs, suggesting that only part of the MdPDV genome is expressed in this host cell type.

Animals↗
PubMed1991

Developmental disruption of Pseudoplusia includens and Heliothis virescens larvae by the calyx fluid and venom of Microplitis demolitor.

Calyx fluid and venom from the braconid parasitoid Microplitis demolitor differentially affected the development of Pseudoplusia includens and Heliothis virescens. P. includens exhibited delays in larval development, supernumerary instars, and formed larval-pupal intermediates when injected with 0.01-0.10 wasp equivalents of calyx fluid. In contrast, H. virescens was relatively unaffected by calyx fluid regardless of dose. Venom did not affect the development of either host species, but appeared to synergize the activity of calyx fluid. This was particularly evident in H. virescens, where injection of 0.10-0.20 wasp equivalents of calyx fluid and venom induced the formation of a large number of intermediates while the same amount of calyx fluid did not. The particulate portion of M. demolitor calyx fluid was the only component that caused developmental delays and the formation of intermediates in both host species. Purified virus caused developmental alterations in P. includens, while trioxsalen treated calyx fluid did not affect development of P. includens or H. virescens. These data suggest the requirement for venom in parasitism may differ between host species, and that dosage plays an important role in interpreting the interaction between calyx and venom components.

Animals↗