Quantification of Human Immunodeficiency Virus Type 1 RNA Levels in Plasma by Using Small-Volume-Format Branched-DNA Assays

We have developed small-volume (50 or 250 μl)-format branched-DNA assays for human immunodeficiency virus type 1 (HIV-1) RNA for use with specimens in which the volume is limited and/or a high viral load is anticipated. These formats exhibited good correlation with the standard 1-ml format; high specificity, reproducibility, and linearity; and no significant difference in the quantification of HIV-1 subtypes.     

Enhanced factor H binding to sialylated Gonococci is restricted to the sialylated lacto-N-neotetraose lipooligosaccharide species: implications for serum resistance and evidence for a bifunctional lipooligosaccharide sialyltransferase in Gonococci

We isolated serologically identical (by serovar determination and porin variable region [VR] typing) strains of Neisseria gonorrhoeae from an infected male and two of his monogamous female sex partners. One strain (termed 398078) expressed the L1 (Galalpha1 --> 4 [corrected] Galbeta1 --> 4Glcbeta1 --> 4HepI) lipooligosaccharide (LOS) structure exclusively; the other (termed 398079) expressed the lacto-N-neotetraose (LNT; Galbeta1 --> 4GlcNAcbeta1 --> 3Galbeta1 --> 4Glcbeta1 --> 4HepI) LOS structure. The strain from the male index case expressed both glycoforms and exhibited both immunotypes. Nuclear magnetic resonance analysis revealed that sialic acid linked to the terminal Gal of L1 LOS via an alpha2 --> 6 linkage and, as expected, to the terminal Gal of LNT LOS via an alpha2--> 3 linkage. Insertional inactivation of the sialyltransferase gene (known to sialylate LNT LOS) abrogated both L1 LOS sialylation and LNT LOS sialylation, suggesting a bifunctional nature of this enzyme in gonococci. Akin to our previous observations, sialylation of the LNT LOS of strain 398079 enhanced the binding of the complement regulatory molecule, factor H. Rather surprisingly, factor H did not bind to sialylated strain 398078. LOS sialylation conferred the LNT LOS-bearing strain complete (100%) resistance to killing by even 50% nonimmune normal human serum (NHS), whereas sialylation of L1 LOS conferred resistance only to 10% NHS. The ability of gonococcal sialylated LNT to bind factor H confers high-level serum resistance, which is not seen with sialylated L1 LOS. Thus, serum resistance mediated by sialylation of gonococcal L1 and LNT LOS occurs by different mechanisms, and specificity of factor H binding to sialylated gonococci is restricted to the LNT LOS species.     

Assessment of cancer immunotherapy outcome in terms of the immune response time features.

A cytokine-based periodic immunotherapy treatment is included in a model of tumour growth with a delay. The effects of dose schedule are studied in the case of a weak immune system and a growing tumour. We find the existence of 'metastable' states (that may last for tens of years) induced by the treatment and also potentially adverse effects of the dosage frequency on the stabilization of the tumour. These two effects depend on the delay between the tumour growth and the immune system response, the cytokine dose burden, and other parameters considered in the model.     

New evidence from magnetic resonance imaging of brain changes after climbs at extreme altitude

The aim of the present study was to look for anatomical changes in climbers' brains, using magnetic resonance imaging (MRI), after extremely high-altitude climbs and to relate them to possible associated risk factors. Clinical history, neurological examinations and MRI were carried out on a group of nine climbers before and after climbing to over 7500 m without the use of supplementary oxygen. None of the subjects showed any neurological dysfunctions. In five climbers MRI abnormalities (high signal areas, cortical atrophy) were observed before the expedition. After the descent, two of them showed new high intensity signal areas recorded by MRI. Both subjects suffered severe neurological symptoms during the climb. The present study suggested that the brain changes observed by MRI could be related to the severity of clinical events at high altitude. However, we do not know the exact meaning of such MRI findings or the reason for their location, predominantly in posterior regions of the brain. The new evidence that a high percentage of climbers show MRI brain abnormalities, and especially the appearance of changes after the ascent, reinforces the possibility of a potential neurological risk in high-altitude climbing.     

Inhibition of poly(ADP-ribose) polymerase attenuates the severity of acute pancreatitis and associated lung injury

The severity of acute pancreatitis results from the transmigration and activation of leukocytes within the pancreas and the local synthesis and release of proinflammatory-soluble mediators that transform a local injury into a systemic inflammatory response. Poly(ADP-ribose)polymerase-1 (PARP-1) is a nuclear DNA-binding protein that has been shown to play a relevant role in cell necrosis and organ failure in various diseases associated with inflammation. Therefore, we set out to investigate whether the genetic deletion of PARP-1 or PARP-2 (a new member of the PARP family) genes, or pharmacological inhibition of PARP activity might affect the development and severity of acute pancreatitis and pancreatitis-associated lung injury. Secretagogue-induced acute pancreatitis was achieved by 12 hourly intraperitoneal injections of cerulein in mice deficient in PARP-1 or PARP-2 genes, and wild-type (WT) littermate mice untreated or treated with PARP activity inhibitors. The severity of pancreatitis was assessed by measurements of serum amylase, lipase, interleukin-1beta and IL-6, pancreatic water content, histologic grading and pancreas myeloperoxidase (MPO) activity. Lung injury was evaluated by quantifying MPO activity and morphological changes. We found that the severity of acute pancreatitis and pancreatitis-associated lung injury was significantly attenuated in mice lacking PARP-1, but not PARP-2, compared with WT mice. Interestingly, administration of PARP inhibitors, 3-aminobenzamide or PJ34 (N-(6-oxo-5,6-dihydro-phenanthridin-2-yl)-N,N-dimethyacetamide HCl), in WT mice markedly decreased acute pancreatitis severity and pulmonary-associated injury in a larger extension than genetic deletion of PARP-1. Our results support the potential therapeutic application of PARP inhibitors in the development and severity of acute pancreatitis and associated lung injury.     

Induction of cellular immune response and anti-Salmonella enterica serovar typhi bactericidal antibodies in healthy volunteers by immunization with a vaccine candidate against typhoid fever

Typhoid fever remains a serious public health problem. We have developed a vaccine from Salmonella enterica serovar typhi (S. typhi) outer-membrane proteins (OMPs) known as porins. A single subcutaneous dose of 10 microg of porins induced a five-fold (P = 0.05) seroconversion index consisting of IgM and IgG at 7 and 15 days after vaccination as well as the production of IgG1 and IgG2 isotypes. The porins-based vaccine induced a two-fold increase (P = 0.05) in bactericidal titres in volunteers, whom also developed a T-cell response characterized by the production of interferon-gamma (INF-gamma). Side effects after vaccination were mild and transient. The data showed that our S. typhi porins-based candidate vaccine is safe and immunogenic in healthy humans.     

Molecular characterization of a carnation etched ring virus isolate from India

Incidence of the Carnation etched ring virus (CERV), the only DNA virus reported to date on carnation, was investigated by a bioassay using a partially purified virus as inoculum and then by a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). Out of 61 carnation cultivars analyzed 41 (67%) were found positive. The virus positivity was verified by polymerase chain reaction (PCR) and nucleotide sequencing. The amplified 1349 bp fragment was by about 98% and 96% identical with respect to coat protein (CP) and enzymatic polyprotein genes, respectively, as compared to the sequences available in the database. In terms of amino acid sequence similarity, the homology values were 99% and 97%, respectively. Comparison with other caulimoviruses revealed that CERV is most closely related to the Cauliflower mosaic virus (CaMV). High genetic stability of CERV may be attributed to the fact that it has evolved from the same initial sequence in an original host. Because of global market of cut flowers and vegetative propagation it has been dispersed around the world.     

Cytopathology of uncommon malignant renal neoplasms in the pediatric age group

Malignant renal neoplasms are common solid tumors in pediatric oncology practice. These include the common Wilms' tumor/nephroblastoma and the uncommon neoplasms such as clear-cell sarcoma of the kidney (CCSK), rhabdoid tumor, renal-cell carcinoma, and others. The aim of this study was to describe in detail the cytopathological features of the histopathologically proven uncommon pediatric renal tumors. Aspirates from Wilms' tumor, which are mesenchyme predominant, show clusters of spindle cells associated with the matrix material. Evidence of rhabdomyoblastic differentiation may be present. CCSK, classic subtype, is characterized by round to oval cells arranged perivascularly and also in sheets and clusters intimately associated with a metachromatic matrix mucopolysaccharide material better appreciated in May-Grunwald-Giemsa (MGG)-stained smears. The cells also have more abundant cytoplasm and may show nuclear grooves. Spindle-cell pattern of CCSK is difficult to diagnose on aspiration cytology. Renal-cell carcinoma of childhood shows similar cytological features as its adult counterpart. Rhabdoid tumor of the kidney is characterized by a monomorphic population of cells with abundant cytoplasm, eccentric nuclei with prominent nucleoli. Intrarenal yolk sac tumor is a rare neoplasm and shows severely pleomorphic cells on aspiration.Awareness of these entities is important for the practicing cytopathologist. Further, non-Wilms' renal malignant neoplasms must be distinguished from the common Wilms' tumor so that appropriate chemotherapy protocols may be instituted in cases where the tumor is in an advanced stage of malignancy.