Antisporozoite Antibodies with Protective and Nonprotective Activities: In Vitro and In Vivo Correlations Using Plasmodium gallinaceum, an Avian Model

ABSTRACT. A correlation was observed between in vivo and in vitro activity of six monoclonal antibodies (mAb) against the major circumsporozoite protein of the avian malaria Plasmodium gallinaceum as follows. (1) Two mAb were protective, totally abrogating sporozoite infectivity to chicks, its natural host, in vivo; they caused 100% inhibition of sporozoite invasion (ISI) in vitro to SL-29 chicken fibroblasts and intense ISI to cultured chicken macrophages, as well as inhibited the exoerythrocytic development of sporozoites taken up by macrophages, the initial cell host of P. gallinaceum sporozoites. (2) Two mAb were partially protective in that they reduced sporozoite infectivity to chicks, caused partial ISI to SL-29 and macrophage cells and partial inhibition to the exoerythrocytic development of sporozoites in macrophages in vitro. (3) Two mAb were totally inactive in vivo although they both bound to the sporozoite antigens as detected by indirect immunofluorescence, western blot, and ELISA; they both failed to induce ISI or inhibit the exoerythrocytic development in macrophages. The possible participation of macrophages as the initial cell type involved in sporozoite destruction in the presence of anti-circumsporozoite antibodies is discussed.     

Melanization Stimulating Activity in the Skin of the Gilthead Porgy,Sparus auratus

The presence of a melanization-stimulating factor (MSF) was discovered in dorsal and/or ventral skin of Sparus auratus. Skin from this marine species was used to condition Steinberg's balanced salt solution (BSS), which was subsequently tested with the neural tube assay. BBS conditioned by dorsal and/or ventral skin of S. auratus at 25% and 50% concentrations had a profound stimulatory effect on the percentage of melanization of neural crest cells throughout the 3day assay period. In some cases 90% melanization occurred within the first 24 hr. Such stimulated cells showed a doubling of the number of dendrites per cell. assess the effects of MSF on other indices of melanization, dorsal and/or ventral skin was used to condition MEM used in the culture of B16-F10 murine melanoma cells. During the first 24 hr, B16-F10 murine melanoma cells responded to conditioned media by demonstrating a considerable increase in activities of tyrosine hydroxylase, dopa oxidase, and dopachrome tautomerase, but no effect was observed on melanin content. In contrast, melanin content increased after 48 hr of incubation, whereas the enzymatic activities were inhibited during this period. It seems that MSF activity, expressed in several ways, may be present generally among marine species.     

Development and characterization of eight microsatellite markers in bullet tuna (Auxis rochei)

The bullet tuna (Auxis rochei) is a worldwide species of high commercial value in Spain. Nevertheless, little information is currently available about the genetic characteristics of wild A. rochei populations. In this survey, we have developed eight new microsatellites for the bullet tuna using an enriched genome library protocol. Primers were screened on a total of 78 individuals from three wild populations (Mediterranean, Atlantic, and Pacific), revealing seven to 27 alleles per locus with expected heterozygosities ranging from 0.63 to 0.97. These markers can be potentially useful tools for use in population genetic studies.     

Intrinsic Pigment-Cell Stimulating Activity in the Catfish Integument

In keeping with the concept that local factors in the vertebrate integument affect the expression of pigment cells, the present study was directed toward demonstrating the existence of such factors in the skin of the channel catfish, Ictalurus punctatus. This species has a dark dorsal surface in marked contrast to an almost white midventral surface. Pieces of skin from these two surfaces were used to condition culture media, which were in turn bioassayed using the Xenopus neural tube explant system (Fukuzawa and Ide, 1988, Dev. Biol. 129:25). A certain number of neural crest cells grow out from the explant, and many of these are melanized in a culture medium of Steinberg's basic salt solution (BSS). When the BSS was conditioned with either dorsal or ventral skin, a profound increase in both the number of crest cells emigrated from the neural tubes and the percentage of melanized cells was observed. The effects of dorsal skin were stronger than those of ventral skin and were evident on a dose/response basis. Initial fractionation of conditioned BSS with DEAE ion exchange chromatography produced fractions of particular potency in the stimulation of melanogenesis. A similarly conditioned medium based upon Leibovitz's L-15 was used in the primary culture of mature chromatophores, namely, melanophores, iridophores, and xanthophores from tadpoles of Rana pipiens. Both dorsal and ventral conditioned media stimulated iridophores and xanthophores, but seemed to have little or no effect on tadpole melanophores. A melanization inhibiting factor (MIF) from the ventral surface of adult frogs has been suggested as the basis for the light colored ventrum of amphibians, and although the present experiments were not designed to study catfish MIF, the possible existence of such a factor in this species was supported by the results. The total results of this investigation are discussed in the light of the possible presence of a melanization inhibiting factor (MIF) of greater prevalence in the ventrum and a melanization stimulatory factor (MSF) of greater prevalence in the dorsal integument. It is suggested that the light-colored ventral surface of the catfish and other poikilotherms may result from the presence of higher levels of MIF than MSF. Thus, the expression of melanophores is inhibited while that of iridophores is enhanced. In contrast, higher levels of MSF over MIF in the dark dorsal surface would result in melanophore stimulation and inhibition of iridophore expression.     

In Vitro Development of Exoerythrocytic Forms of Plasmodium Gallinaceum Sporozoites In Avian Macrophages

ABSTRACT Exoerythrocytic forms of Plasmodium gallinaceum were cultured in vitro using salivary gland sporozoites extracted from experimentally infected Aedes fluviatilis mosquitoes. the host cells were macrophage precursors from chicken bone marrow. At various times after introduction of Sporozoites, the cultures were stained by Giemsa or by immunofluorescence assay (IFA) using anti-sporozoite-specific monoclonal antibodies (MAb). the time to complete parasite development in vitro was 50-70 h. By 70 h, ruptured segmenters and free merozoites were visible within the cells. Inoculation of normal chickens with infected cultures induced parasitemia after a pre-patent period of 10-11 days. In vitro young exoerythrocytic forms, late schizonts that include the matured segmenters, and free merozoites shared common antigens with the sporozoites as revealed by IFA using anti-sporozoite-specific MAbs. Our data indicate that macrophages support development of P. gallinaceum sporozoites and that the circumsporozoite proteins are present until Ac end of the primary exoerythrocytic schizogony.     

Isolation and characterization of ten microsatellite loci for Senegal sole (Solea senegalensis Kaup)

Senegal sole (Solea senegalensis Kaup) is a high‐value marine flatfish exploited in both fisheries and aquaculture. Here we describe the isolation of seven tetranucleotide and three dinucleotide polymorphic microsatellite loci. Characteristics of a captive broodstock and a wild population are described. Three loci displayed a significant departure from Hardy–Weinberg expectations in both populations, suggesting a substantial frequency of null alleles. The remaining seven loci were found to be in equilibrium in the wild population, whereas only four of them were in the cultured population. Cross‐species amplification was successful for three loci in other five flatfishes.     

Melanocytes in the Stria Vascularis and Vestibular Labyrinth of the Mongolian Gerbil (Meriones unguiculatus)

Inner ear melanocytes are mainly present in the cochlea, vestibular organ, and endolymphatic sac, but their exact biological function has not been determined. In this investigation, we study the pigment cells in the membranous labyrinth of the gerbil. The inner ear melanocytes of M. unguiculatus show an irregular dendritic shape with cytoplasmic processes. These cells are disposed following the distribution of striai marginal and vestibular dark cells that have an important metabolic activity. Gerbil inner ear melanocytes are characterized by the presence of melanosomes, which are homogeneously dense organelles, of variable size and shape, that are surrounded by a membrane. In these cells, the Golgi apparatus plays a important role in melanin synthesis. When melanocytes were incubated in L-DOPA solution, the vesicles and cisterns of the Golgi apparatus exhibited a positive tyrosinase reaction. An interesting observation is the relation between melanocytes and inner ear capillaries. Sometimes, near to sensory vestibular areas, the melanocytes were in contact with Schwann cells and with myelinated fibres of vestibular nerve. The ultrastructural findings of this investigation are consistent with the hypothesis that melanocytes may have functional significance in the inner ear.     

Characterization of microsatellite loci for the redbanded seabream,Pagrus auriga (Teleostei,Sparidae)

The redbanded seabream Pagrus auriga (Teleostei, Sparidae) is a species of a high commercial value in Spain. There is currently little information available about the genetic characteristics of both wild and cultured populations. In this survey, we have developed eight polymorphic microsatellites for the redbanded seabream using an enriched genome library protocol. All of them were polymorphic in the 64 individuals tested, 22 of which were wild specimens, and 42 were individuals from a captive reproductive broodstock. These markers can potentially be useful tools for use in population genetic studies.