Daniela Marchini

Purification and primary structure of ceratotoxin A and B, two antibacterial peptides from the female reproductive accessory glands of the medfly Ceratitis capitata (insecta:Diptera)

In the present article we report the purification and the amino acid sequence of two antibacterial peptides present in the secretion of the female reproductive accessory glands of the dipteran insect Ceratitis capitata. Both peptides consist of 29 amino acid residues, are heat stable, strongly basic and differ from each other for the substitution of two amino acids. Their primary sequence and predicted secondary structure are related to other families of peptides known to have lytic and/or antibacterial activity. We propose the name ceratotoxins (from Ceratitis) for these antibacterial peptides.

Bacteria associated with the oesophageal bulb of the medfly Ceratitis capitata (Diptera:Tephritidae)

Extracellular Gram negative bacteria were found to be commonly associated to the oesophageal bulb of Ceratitis capitata with Klebsiella oxytoca and Enterobacter agglomerans as the most common species. All the isolates tested in vitro, except one, were sensitive to the antibacterial material present on the medfly laid egg surface.

The Novel Antibacterial Peptide Ceratotoxin A Alters Permeability of the Inner and Outer Membrane of Escherichia coli K-12

Abstract. Ceratotoxins are antibacterial 3-kDa amphiphilic peptides isolated from the female reproductive apparatus of the medfly Ceratitis capitata. The antibacterial activity of a chemically synthesized ceratotoxin A (ctx A) has been investigated. Ctx A was mainly active against Gram-negative organisms, and it had a lytic effect on nongrowing Escherichia coli K-12. Data showed that ctx A alters both the outer and the inner membrane of E. coli K-12 cells.

A male sexually dimorphic trait provides antimicrobials to eggs in blenny fish

Predation and microbial infections are the major causes of natural mortality for early life stages of oviparous species. The parental traits reducing the effects of predation are rather well described, whereas antimicrobial mechanisms enhancing offspring survival are largely unexplored. In this paper, we report that a male sexually dimorphic trait, the anal glands, of the redlip blenny (Ophioblennius atlanticus atlanticus) and the peacock blenny (Salaria pavo), two fish species with paternal egg care, produce a mucus enriched with antimicrobial substances. Histological and histochemical analyses showed that the anal glands of these species are characterized by the massive presence of mucus-secreting cells. Anal gland extracts, from both the hydrophilic and the hydrophobic protein fraction, exhibited a lysozyme-like activity. Field observations demonstrated that redlip blenny males, while performing egg care, rub the anal region over the nest internal surface, probably facilitating the transfer of mucus to eggs. These results strongly indicate that this sexually dimorphic trait is involved in egg defence against microbial infections.

Ultrastructure of the male reproductive accessory glands in the medfly Ceratitis capitata (Diptera: Tephritidae) and preliminary characterization of their secretions

The morphology and the ultrastructure of the male accessory glands and ejaculatory duct of Ceratitis capitata were investigated. There are two types of glands in the reproductive apparatus. The first is a pair of long, mesoderm-derived tubules with binucleate, microvillate secretory cells, which contain smooth endoplasmic reticulum and, in the sexually mature males, enlarged polymorphic mitochondria. The narrow lumen of the gland is filled with dense or sometimes granulated secretion, containing lipids. The second type consists of short ectoderm-derived glands, finger-like or claviform shaped. Despite the different shape of these glands, after a cycle of maturation, their epithelial cells share a large subcuticular cavity filled with electron-transparent secretion. The ejaculatory duct, lined by cuticle, has epithelial cells with a limited involvement in secretory activity. Electrophoretic analysis of accessory gland secretion reveals different protein profiles for long tubular and short glands with bands of 16 and 10kDa in both types of glands. We demonstrate that a large amount of accessory gland secretion is depleted from the glands after 30min of copulation.

The female reproductive accessory glands of the medfly Ceratitis capitata : antibacterial activity of the secretion fluid

Abstract Secretion from female reproductive accessory glands of the dipteran Ceratitis capitata was found to have antibacterial properties against E. coli . At least two basic polypeptides with mol. wt 15.5 and 4.7 kDa respectively, were identified as responsible for such activity. Furthermore, the 15.5 kDa protein is active against a number of Gram-positive and -negative bacterial strains. Lysozyme activity is also present in the secretion.

Sequences of two cDNA clones from the medfly Ceratitis capitata encoding antibacterial peptides of the cecropin family

Using a back translated oligodeoxyribonucleotide probe, encoding a conserved motif in insect antibacterial peptides, we have isolated two cDNA clones from the medfly, Ceratitis capitata. Sequence determination shows that the cDNAs encode two closely related peptides which are members of the cecropin family.

Ultrastructure of the midgut and the adhering tubular salivary glands of Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae)

Abstract The ultrastructure of the midgut and the tubular salivary glands of Frankliniella occidentalis (Thysanoptera : Thripidae) is described. The microvilli have 2 different types of glycocalyx: in the anterior part of the midgut they are surrounded by a myelin-like membrane; in the posterior region, the microvilli have numerous rod-like projections arranged to form a continuous layer. Microfilaments longitudinally cross each microvillus; the microfilaments contain F-actin. Tubular salivary glands flank the midgut but do not fuse with it. The distal part of these glands have microvillated cells containing large amounts of electron-transparent material. The cells of the proximal part are lined with a thin cuticle.

Structural organization of the copulation site in the medfly Ceratitis capitata (Diptera: Tephritidae) and observations on sperm transfer and storage.

The copulation site of the medfly Ceratitis capitata was investigated at anatomical and ultrastructural levels. It consists of the anterior vagina, with a ventral fertilization chamber and a dorsal insemination pocket into which the two spermathecal ducts open. The fertilization chamber is an organ comprised of a number of alveoli that in virgin females are filled with a filamentous secretion, whereas in mated females contain sperm bundles. Through study of the internal morphology of the aedeagus, its position in the anterior vagina, and the direct observation of sperm transfer and storage, we confirmed that sperm are ejaculated through two gonopores at the top of the distiphallus and another at the base of the genital rod. The sperm flow dorsally into the insemination pocket and ventrally into the fertilization chamber. During copulation, the two spermathecae and the fertilization chamber are progressively filled with spermatozoa.

β-N-acetylhexosaminidases in the secretion of the female reproductive accessory glands of Ceratitis capitata (Diptera)

Abstract The analysis by isoelectric focusing of the secretion fluid (AGF) of the female accessory glands of Ceratitis capitata has shown the presence of seven major protein fractions and two β-N-acetylhexosaminidase isoenzymes, HEX 1 and HEX 2, focalizing at pH 4.75 and 5.25, respectively. The two isoenzymes have the same molecular weight (69 kDa) but a different hydrolytic activity against p- nitrophenyl-2-acetamido-2-deoxy -β- d -glucopyranoside and galactopyranoside, and a pH optimum of 4 at 37°C. AGF β-N-acetylhexosaminidase activity is most stable at pH values between 5 and 8. At neutral pH a rapid loss of activity is observed at temperatures higher than 50°C. Preliminary assays, carried out using chitin as substrate, indicate the presence in the secretion of an endochitinase activity not bound to the β-N-acetylhexosaminidase isoenzymes. The enzymatic modifications of specific lectin binding sites on the surface of gametic cells is most probably one of the necessary steps preceding fertilization.