[INSECTS AND THEIR IMPORTANCE IN PUBLIC HEALTH].
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Mermithid nematodes parasitize invertebrates only. A general account is given of their parasitism and the consequent mortality of blackflies. The author begins with a review of records of mermithid and blackfly association from different parts of the world and comments on the global distribution of the parasite. He discusses the life-histories of the parasites, stressing their adaptations to the moving environment of the aquatic stages of the blackfly and to the aerial environment of the adult. It is noted that the parasite usually kills its host upon emergence and has a restricted host range. Mermithids must be considered as selective mortality factors against blackfly populations. A survey of European and North American literature leads to conclusions concerning the role of the parasite in the natural regulation of blackflies. The possibility of utilizing these nematodes as biological control agents is discussed.
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The insect repellents 3535 (ethyl butylacetylaminopropionate or IR3535) and deet (N,N-diethyl-3-methylbenzamide) were prepared as 20% solutions in absolute ethanol and evaluated for repellency against many mosquito species in Thailand under laboratory and field conditions using human subjects. In the laboratory, 0.1 ml was applied per 30-cm2 of exposed area on a volunteer's forearm (0.66-0.67 mg active ingredient [AI]/ cm2), whereas in the field, volunteers' legs (from knee to ankle, with a surface area of about 712-782 cm2) were treated with 3 ml per exposed area (0.76-0.84 mg AI/cm2). In the laboratory, both IR3535 and deet showed equal repellency (P > 0.05) for 9.8 and 9.7 h against Aedes aegypti, for 13.7 and 12.7 h against Culex quinquefasciatus, and for 14.8 and 14.5 h against Cx. tritaeniorhynchus, respectively. Anopheles dirus was significantly less sensitive to IR3535 than to deet (P < 0.05), with a mean protection time of 3.8 and 5.8 h, respectively. Under field conditions, both IR3535 and deet provided a high degree of protection against various mosquito vectors ranging from 94 to 100% during the test periods. Both repellents provided a high level of protection for at least 8 h against Ae. albopictus and for at least 5 h against Cx. gelidus, Cx. tritaeniorhynchus, Cx. quinquefasciatus, Mansonia dives, Ma. uniformis, Ma. annulata, Ma. annulifera, Anopheles minimus, and An. maculatus. This study clearly documents the potential of IR3535 for use as a topical treatment against a wide range of mosquito species belonging to several genera.
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The gene encoding the 65K tegument phosphoprotein (pp65) of human cytomegalovirus (HCMV) was cloned into pAc373 to construct a recombinant baculovirus (Acpp65-3) expressing pp65 in insect Sf9 cells. A baculovirus that carried a fragment of the gene, corresponding to the first 442 amino acids of pp65, was also developed, using vector pVL941 (Acpp65-2). Recombinant proteins migrating in SDS-polyacrylamide gels with an M(r) of either 65K (Acpp65-3) or 56K (Acpp65-2) were detected in cytoplasmic and nuclear extracts of infected Sf9 cells. The 56K and 65K proteins were recognized in immunoblots by monoclonal antibodies (MAbs) 28-77 and 28-19, which are specific for pp65. The insect cell-expressed antigens were also analysed on Western blots using MAbs 4D11, 7D2, 8E3, 7B4 and 8E10, which recognize the HCMV antigen GP66 in immunoblots. The truncated pp65 antigen of Acpp65-2 was reactive with MAbs 4D11, 7D2, 8E10 and 7B4. The protein expressed by Acpp65-3 reacted only with MAb 4D11. The data proved that the epitopes recognized by MAbs 4D11, 7D2, 8E3 and 7B4 mapped in the region of pp65, comprising amino acids 1 to 442, and also that GP66 and pp65 represent the same HCMV antigen. Immunoblot analysis of human sera from individuals seropositive for HCMV showed that the recombinant pp65 products were as antigenic as the native 65K phosphoprotein produced in HCMV-infected human embryonic fibroblasts.
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Today, many thousands of recombinant proteins, ranging from cytosolic enzymes to membrane-bound proteins, have been successfully produced in baculovirus-infected insect cells. Yet, in addition to its value in producing recombinant proteins in insect cells and larvae, this viral vector system continues to evolve in new and unexpected ways. This is exemplified by the development of engineered insect cell lines to mimic mammalian cell glycosylation of expressed proteins, baculovirus display strategies and the application of the virus as a mammalian-cell gene delivery vector. Novel vector design and cell engineering approaches will serve to further enhance the value of baculovirus technology.
Biological parameters of Triatoma brasiliensis and T. pseudomaculata that could influence the epidemiological importance of these insects as vectors of Trypanosoma cruzi were compared. The parameters studied were incubation period, interval between hatching or moulting and first feeding, number of blood meals, development time, mortality, net reproductive rate, instantaneous daily reproductive rate, time-lapse before starting feeding, duration of feeding, blood ingestion capacity, occurrence of defecation and blood ingestion velocity. Most aspects of feeding were similar for the two species, although T. pseudomaculata had a longer life cycle than T. brasiliensis producing one and two generations per year, respectively. The two species had similar instantaneous daily rates of population growth.
A baculovirus transfer vector was constructed containing an entire cDNA copy of the chick nicotinic acetylcholine receptor (nAChR) alpha-subunit under control of the Autographa californica nuclear polyhedrosis virus (AcNPV) polyhedrin gene promoter. Recombinant baculovirus was obtained by co-transfection of Spodoptera frugiperda cells with infectious, wild-type AcNPV DNA and the transfer vector. Polyhedrin-negative, recombinant viruses were identified which expressed the nAChR alpha-subunit. The insect cell-expressed alpha-subunit protein had a molecular mass of 42 kDa and was shown to be targeted to the plasma membrane by fluorescence microscopy and toxin-binding assays. The levels of expression were low, approximately 1-2% of cell proteins, when compared with the levels of natural polyhedrin protein. The expressed receptor alpha-subunit was recognised by polyclonal antisera raised against purified Torpedo nAChR alpha-subunit and carried the binding site for the snake venom toxin, alpha-bungarotoxin. Bound alpha-bungarotoxin was displaced in competition binding assays by alpha-cobra toxin, carbamylcholine and d-tubocurarine, and thus had a similar pharmacological profile to that obtained with authentic receptors in muscle cells and receptors expressed in other systems i.e. Xenopus oocytes and mammalian cells. We have also shown that when the chick nAChR alpha-subunit is expressed in the absence of other receptor subunits, unexpectedly high concentrations of nicotine (10 mM) were required to displace bound alpha-bungarotoxin.
Baculovirus expression vectors are used routinely for foreign gene expression and are under intense development as improved biological pesticides. Conventional baculovirus expression vectors are recombinant viruses that can express a foreign gene in insect cells under the control of the polyhedrin promoter, which provides high-level transcription during the very late phase of infection. For some applications, including foreign glycoprotein production and insect pest control, it might be advantageous to have baculovirus vectors that could express foreign gene products in uninfected cells or earlier after infection. To fulfill this need, we have constructed a new set of plasmids that can be used to clone and express foreign genes under the control of a baculovirus ie1 promoter, which is active in uninfected insect cells and throughout infection. We used a subset of these new plasmids to isolate recombinant baculoviruses containing various foreign genes and compared expression of these genes by the resulting immediate-early baculovirus vectors and by conventional baculovirus vectors. As expected, the immediate-early vectors began to express each foreign gene earlier in infection but, by 36-48 h postinfection, the conventional vectors had produced more of each foreign protein. Conventional baculovirus vectors also produced more enzymatic activity from two different procaryotic genes than the immediate-early baculovirus vectors. However, immediate-early vectors produced as much or more enzymatic activity from two different eucaryotic genes encoding secretory pathway proteins than the conventional vectors, even at 48 h postinfection. Hence, this report describes a new set of plasmids that can be used to clone and express foreign genes under the control of the baculovirus ie1 promoter and suggests that immediate-early baculovirus vectors might be as useful as conventional baculovirus expression vectors for producing biologically active eucaryotic secretory pathway proteins.