A combined polymerase chain reaction and restriction endonuclease enzyme assay for discriminating between Campylobacter coli and Campylobacter jejuni

Abstract A combined polymerase chain reaction and restriction endonuclease (RE) enzyme assay was developed to discriminate between Campylobacter coli and Campylobacter jejuni . Amplimers of the FlaA gene obtained by PCR were digested with Alu I and Hin fI to distinguish C. coli from C. jejuni . With Alu I digestion C. jejuni -specific bands were observed at 110, 140 and 160 bp and C. coli -specific bands at 293 and 147 bp. C. jejuni -specific bands of 349 and 109 bp were found by Hin fI digestion but Hin fI did not digest the Fla A amplimer of C. coli . This combined technique is fast and easy to perform, and distinguishes the two campylobacters unequivocally.     

The evolution of the Cercopithecini: a (post)modern synthesis

The Cercopithecini, or African guenon monkeys, are one of the most diverse clades of living primates and comprise the most species‐rich clade of Catarrhini. Species identity is announced by flamboyant coloration of the facial and genital regions and, more cryptically, by vigorous chromosomal rearrangements among taxa. Beneath the skin, however, these animals are skeletally conservative and show low levels of genetic sequence divergence consonant with recent divergence between congeneric species. The guenons clearly demonstrate that morphological, cytogenetic, and reproductive differentiation proceed at different rates during speciation. We review diverse kinds of data in an effort to understand this conundrum.     

High-level HIV-1 Nef transient expression in Nicotiana benthamiana using the P19 gene silencing suppressor protein of Artichoke Mottled Crinckle Virus

Background  

In recent years, different HIV antigens have been successfully expressed in plants by either stable transformation or transient expression systems. Among HIV proteins, Nef is considered a promising target for the formulation of a multi-component vaccine due to its implication in the first steps of viral infection. Attempts to express Nef as a single protein product (not fused to a stabilizing protein) in transgenic plants resulted in disappointingly low yields (about 0.5% of total soluble protein). In this work we describe a transient expression system based on co-agroinfiltration of plant virus gene silencing suppressor proteins in Nicotiana benthamiana, followed by a two-step affinity purification protocol of plant-derived Nef.     

Mesozoic tectono-sedimentary evolution of Rocca Busambra in western Sicily

The Rocca Busambra ridge in western Sicily is a shallow to pelagic Meso-Cenozoic carbonate structural unit of the Sicilian Chain with a variety of tectono-sedimentary features. Palaeofaults, unconformities (buttress unconformity, onlap, downlap), a network of neptunian dykes with several infilling generations, several large hiatuses, different facies and lateral facies changes, and erosional submarine and subaerial surfaces are observed. Detailed fieldwork and structural analyses have indicated the occurrence of fault planes with different orientations. These data, combined with facies studies and physical-stratigraphy analyses, allow for the distinction of different depositional regions. A lateral change from an open-marine carbonate platform with a stepped fault margin (located in the westernmost sector) to a deeper basinal depositional setting in the east, in the context of an upper slope scalloped margin and base-of-slope systems with talus breccias, is envisaged here. Extensional to transtensional tectonic pulses punctuated the sedimentary evolution during Early Toarcian, Late Jurassic, Early Cretaceous, Late Cretaceous, and Early Miocene times. The collected data show that most fault planes have preserved their original orientations throughout the reactivation processes. The reconstructed Meso-Cenozoic tectono-sedimentary evolution is closely related to the late syn-rift and post-rift tectonic evolution of the Tethyan continental margin.     

Tumour-released exosomes and their implications in cancer immunity

Tumour cells release vesicular structures, defined as microvesicles or exosomes, carrying a large array of proteins from their originating cell. The expression of antigenic molecules recognized by T cells has originally suggested a role for these organelles as a cell-free antigen source for anticancer vaccines. However, recent evidence shows that tumour exosomes may also exert a broad array of detrimental effects on the immune system, ranging from apoptosis in activated antitumour T cells to impairment of monocyte differentiation into dendritic cells and induction of myeloid suppressive cells. Immunosuppressive exosomes of tumour origin can be found in neoplastic lesions and sera from cancer patients, implying a potential role of this pathway in in vivo tumour progression. Through the expression of molecules involved in angiogenesis promotion, stromal remodelling, delivery of signalling pathways through growth factor/receptor transfer, chemoresistance and genetic intercellular exchange, tumour exosomes could represent a versatile tool for moulding host environment. Hence, their secretion by neoplastic cells may in the future become a novel pathway to target for therapeutic intervention in cancer patients.     

Evaluation of the inhibitory effects of human serum components on bactericidal activity of human beta defensin 3

Naturally occurring cationic antimicrobial peptides (CAPs) are an essential component of the innate immune system of multicellular organisms. At concentrations generally higher than those found in vivo, most CAPs exhibit strong antibacterial properties in vitro, but their activity may be inhibited by body fluids, a fact that could limit their future use as antimicrobial and/or immunomodulatory agents. In the present study, we evaluated the effects of human serum components on bactericidal activity of the human beta-defensin 3 (hBD-3), a CAP considered particularly promising for future therapeutic employment. Human serum diluted to 20% strongly inhibited the bactericidal activity of the peptide against both the Gram-positive species Staphylococcus aureus and the Gram-negative species Acinetobacter baumannii. Such activity was not restored in serum devoid of salts (dialyzed), pre-treated with protease inhibitors, or subjected to both of these treatments. The addition of physiological concentrations of NaCl, CaCl2, and human albumin in the bactericidal assay abolished bactericidal activity of hBD-3 against S. aureus, while it only partially inhibited the activity of the peptide against A. baumannii. Although a proteolytic activity of serum on hBD-3 was demonstrated at the protein level by Western blot, addition of physiological concentrations of trypsin to the bactericidal assay only partially affected the antibacterial properties of the peptide. Altogether, these results demonstrate a major role of mono-divalent cations and serum proteins on inhibition of hBD-3 antibacterial properties and indicate a relative lack in sensitivity of the bactericidal activity of this peptide to trypsin and trypsin-like proteases.     

Serotonergic mechanisms of the lateral parabrachial nucleus and cholinergic-induced sodium appetite

Central cholinergic mechanisms are suggested to participate in osmoreceptor-induced water intake. Therefore, central injections of the cholinergic agonist carbachol usually produce water intake (i.e., thirst) and are ineffective in inducing the intake of hypertonic saline solutions (i.e., the operational definition of sodium appetite). Recent studies have indicated that bilateral injections of the serotonin receptor antagonist methysergide into the lateral parabrachial nucleus (LPBN) markedly increases salt intake in models involving the activation of the renin-angiotensin system or mineralocorticoid hormones. The present studies investigated whether sodium appetite could be induced by central cholinergic activation with carbachol (an experimental condition where only water is typically ingested) after the blockade of LPBN serotonergic mechanisms with methysergide treatment in rats. When administered intracerebroventricularly in combination with injections of vehicle into both LPBN, carbachol (4 nmol) caused water drinking but insignificant intake of hypertonic saline. In contrast, after bilateral LPBN injections of methysergide (4 microg), intracerebroventricular carbachol induced the intake of 0.3 M NaCl. Water intake stimulated by intracerebroventricular carbachol was not changed by LPBN methysergide injections. The results indicate that central cholinergic activation can induce marked intake of hypertonic NaCl if the inhibitory serotonergic mechanisms of the LPBN are attenuated.     

Hypergonadotropic ovarian failure associated with an inherited mutation of human bone morphogenetic protein-15 (BMP15) gene

Hypergonadotropic ovarian failure is a common cause of female infertility. It is a heterogeneous disorder that, in the most severe forms, is a result of ovarian dysgenesis (OD). Most OD cases are associated with major X-chromosome abnormalities, but the pathogenesis of this disorder is still largely undefined in patients with a normal karyotype. Animal models showed the important role in female reproduction played by the product of a gene located at Xp11.2 in humans (BMP15). BMP15 is an oocyte-specific growth/differentiation factor that stimulates folliculogenesis and granulosa cell (GC) growth. We report two sisters with a normal karyotype who are affected with hypergonadotropic ovarian failure due to OD. The familial presentation suggested a genetic origin, and candidate genes were screened for mutations. A heterozygous nonconservative substitution in the pro region of BMP15 (Y235C) was identified in both sisters but not in 210 control alleles. This mutation was inherited from the father. Mutant BMP15 appears to be processed abnormally, is associated with reduced GC growth, and antagonizes the stimulatory activity of wild-type protein on GC proliferation. In conclusion, the first natural mutation in human BMP15 is associated with familial OD, indicating that the action of BMP15 is required for the progression of human folliculogenesis. This condition represents an exceptional example of X-linked human disease exclusively affecting heterozygous females who inherited the genetic alteration from the unaffected father. BMP15 defects are involved in the pathogenesis of hypergonadotropic ovarian failure in humans.