Genetic barrier to the development of resistance to rilpivirine and etravirine between HIV-1 subtypes CRF02_AG and B.

OBJECTIVES: It has been demonstrated for some drugs that the genetic barrier, defined as the number of genetic transitions and/or transversions needed to produce a resistance mutation, can differ between HIV-1 subtypes. We aimed to assess differences in the genetic barrier for the evolution of resistance to the second-generation non-nucleoside reverse transcriptase inhibitors etravirine and rilpivirine in subtypes B and CRF02_AG in antiretroviral-naive patients. METHODS: An analysis was undertaken of 25 substitutions associated with etravirine and rilpivirine resistance at 12 amino acid positions in 267 nucleotide sequences (136 HIV-1 B and 131 HIV-1 CRF02_AG subtypes) of the reverse transcriptase gene. RESULTS: The majority (7/12) of amino acid positions studied were conserved between the two HIV-1 subtypes, leading to a similar genetic barrier. Different predominant codons between the subtypes were observed in 5/12 positions (90, 98, 179, 181 and 227), with an effect on the calculated genetic barrier only at the V179D and V179F codons (2.5 versus 3.5 for V179D, and 2.5 versus 5 for V179F, respectively, for subtype B versus subtype CRF02_AG). CONCLUSIONS: The majority of amino acids involved in etravirine and rilpivirine resistance showed a high degree of conservation of the predominant codon between the B and CRF02_AG subtypes. For rilpivirine, the genetic barrier was the same between the two subtypes. Nevertheless, subtype CRF02_AG showed a higher genetic barrier to acquiring mutations V179D and V179F (mutations associated with resistance to etravirine) compared with subtype B, suggesting that it would be more difficult to produce resistance to etravirine in the CRF02_AG subtype than the B subtype.

Similar increased serum dipeptidyl peptidase IV activity in chronic hepatitis C and other viral infections.

BACKGROUND: Dipeptidyl peptidase IV is a transmembrane enzyme widely expressed in many cell types, but also present as a soluble form in biological fluids. Its abnormal activity is sometimes associated with liver disease related pathologies. OBJECTIVES: The aim of this study was to evaluate the clinical relevance of changes in serum DPPIV activity in hepatitis C and other viral infections. STUDY DESIGN: DPPIV activity was assessed by using a microplate-based colorimetric assay on serum from 88 subjects: 12 healthy uninfected controls, 10 patients with primary biliary cirrhosis (PBC) as a reference group, 36 HCV-infected patients, and patients suffering from viral infections of different etiologies. Levels of DPPIV activity were compared with: (1) those of other serum biochemical parameters such as alanine aminotransferase (ALT), aspartate aminotransferase (AST) and gamma glutamyl transpeptidase (GGT), and bilirubin concentrations; and (2) criteria representative of liver histological status. RESULTS: Compared with healthy subjects, DPPIV activity was significantly increased during viral infections and in PBC (P<0.01). In HCV-infected patients, the median activity (interquartile range, IQR), 29.78 IU/l (24.66-35.95), differed significantly (P<0.05) from that of controls: 21.42 (19.76-24.93). No correlation was observed between DPPIV activity and either ALT, AST, bilirubin, or the stage of liver fibrosis and necroinflammatory activity, although GGT was moderately correlated (r=0.58, P<0.05). CONCLUSIONS: Although we confirmed an elevation of serum DPPIV activity in PBC, it seems to be a non-specific phenomenon common to viral infections. The absence of correlation between serum DPPIV and markers of liver disease in HCV-infected patients, suggests that this activity originates not only from the liver, but also from other sources such as peripheral blood cells involved in the control of viral infections.

Comparative analysis of translation efficiencies of hepatitis C virus 5' untranslated regions among intraindividual quasispecies present in chronic infection: opposite behaviors depending on cell type.

Hepatitis C virus (HCV) RNA translation initiation is dependent on the presence of an internal ribosome entry site (IRES) that is found mostly in its 5' untranslated region (5' UTR). While exhibiting the most highly conserved sequence within the genome, the 5' UTR accumulates small differences, which may be of biological and clinical importance. In this study, using a bicistronic dual luciferase expression system, we have examined the sequence of 5' UTRs from quasispecies characterized in the serum of a patient chronically infected with HCV genotype 1a and its corresponding translational activity. Sequence heterogeneity between IRES elements led to important changes in their translation efficiency both in vitro and in different cell cultures lines, implying that interactions of RNA with related transacting factors may vary according to cell type. These data suggest that variants occasionally carried by the serum prior to reinfection could be selected toward different compartments of the same infected organism, thus favoring the hypothesis of HCV multiple tropism.

Preferential associations of alleles of three distinct genes argue for the existence of two prototype variants of human herpesvirus 7.

We had previously described six distinct alleles of the glycoprotein B (gB) gene of human herpesvirus 7 (HHV-7). The genetic changes corresponding to these alleles did not affect gB gene transcription or translation in in vitro assays. The study of distinct HHV-7-positive human samples showed preferential associations of some gB alleles with some alleles of two other genes, distantly located on the HHV-7 genome, coding for the phosphoprotein p100 (p100) and the major capsid protein (MCP). Two allele combinations, corresponding to 44 and 31% of the samples studied, respectively, were interpreted as the genetic signatures of two major prototype HHV-7 variants.

Yellow fever 5' noncoding region as a potential element to improve hepatitis C virus production through modification of translational control.

The lengthy 5' noncoding region (5' NCR) of hepatitis C virus (HCV) RNA forms a highly ordered secondary structure, very conserved among different strains. It includes an internal ribosome entry site (IRES) element, responsible for the cap-independent translation initiation of HCV RNA. Similarly to the IRES of hepatitis A virus (HAV), another human hepatitis virus, HCV IRES, activity in internal initiation of translation is weak. Furthermore, both viruses exhibit a poor growth phenotype that may result at least partially from an inhibitory control of translation. To enhance HCV translation, as a preliminary step in designing constructs for improvement in viral production, we sought to evaluate a chimeric construct containing the yellow fever virus (YFV) 5' NCR fused to the initiation codon of the HCV coding sequence. YF viral RNA, as the majority of eukaryotic messenger RNAs, is translated by a ribosome scanning mechanism in a cap-dependent manner. The efficiency of translation initiation of the parental HCV construct was compared in vitro in rabbit reticulocyte lysates with that of the chimeric construct containing YFV 5' NCR. Surprisingly, the related distanced YFV 5' NCR was fivefold more active than was the wild-type HCV IRES in directing that function. Furthermore, chimeric transcripts were shown to be effective in vivo after transfection of eukaryotic cells. Taken together, these results raise the following question: why has the HCV genus evolved to the acquisition of an IRES element within its 5' NCR among the Flaviviridae family?

Use of inverse polymerase chain reaction to characterize a novel human herpesvirus 7 isolate.

Human herpesvirus-7 (HHV-7) is a T-lymphotropic virus detected in peripheral blood mononuclear cells and in saliva, but no reliable link between this agent and a disease has been demonstrated so far. Starting from a 186 bp-fragment described previously, we used an inverse polymerase chain reaction to clone and sequence the adjacent sequences of this known region. A 1062 bp-fragment containing two ORFs was characterized and its sequence was compared with those of two other beta-herpesviruses, human cytomegalovirus (HCMV) and human herpesvirus-6 (HHV-6). With respect to the first ORF, amino-acid identity was estimated to be 22% between HHV-7 and HCMV, and 47% between HHV-7 and HHV-6. In contrast, only a weak homology between HHV-7 and the two other beta-herpesviruses was demonstrated for the second ORF. The newly characterized 1062 bp-fragment was used to define a novel HHV-7-specific PCR assay. Preliminary data indicate that this region is highly conserved among HHV-7 isolates.

"In vivo" and "in vitro" evaluation of four antihistamines (astemizole, azatadine, mequitazine, terfenadine).

A group of 32 patients with perennial allergic rhinitis sensitized exclusively to house dust and mites were evaluated for "in vivo" modifications (skin tests for histamine, house dust and mites) and "in vitro" alterations (seric IgE, specific IgE, histamine release curve to different concentrations of Dermatophagoides pteronyssinus and histamine release curve to this pneumoallergen after incubation with H1 receptor antagonists) to four antihistamines (astemizole, azatadine, mequitazine and terfenadine). The patients were divided into 4 groups of 8 and the "in vivo" and "in vitro" tests were performed on days -1, +15 and +30 of administration of these drugs. The reduction in the size of the wheal (prick test) for histamine when comparing days -1 and +15 and -1 and +30 was 69% and 79% (astemizole), 64% and 71% (azatadine), 69% and 64% (mequitazine) and 61% and 64% (terfenadine), respectively. The difference in the mean size of the wheal in mm2 for house dust, Dermatophagoides pteronyssinus and farinae was statistically significant (p less than 0.001) except when comparing the results obtained for mequitazine on days -1 and +15. This was also the case on days -1 and +30 for terfenadine. The values of the serum IgE and specific IgE did not attain statistical significance (p greater than 0.05) in the various tests carried out. After incubation with H1 receptor antagonists an inhibition curve of the histamine release in the presence of Dermatophagoides pteronyssinus was obtained. The inhibition of this curve on day -1 was higher for terfenadine and mequitazine than for azatadine and astemizole.(ABSTRACT TRUNCATED AT 250 WORDS)

Parietaria judaica and dermo-respiratory allergy in Barcelona: a study of 250 cases.

The wide geographical distribution of parietaria in southern and western Europe is responsible for the high incidence of pollinic hypersensitivity. Our centre detected the existence of hypersensitivity to parietaria judaica in 250 patients (12.15%) out of a group of 2.057 patients. Exclusive consideration of the pollinic patients in this group 980/2,057 showed that the parietaria is the spore responsible for 25.5% of these patients. The most common symptomatology was rhinoconjunctivitis (41.2%) and the least frequent, the dermic picture (1.2%). The diagnostic methodology of these 250 patients included: anamnesis, rhinoscopy, rhinomanometry, skin tests, respiratory function, immunoglobulin levels (IgG, IgA, IgM, IgE), specific IgE and histamine release test. All patients were subjected to immunotherapy using parietaria judaica extracts. The overall evolution following immunotherapy was favourable in more than 80% of the patients, which confirms the suitability of the pareitaria judaica extract in the diagnosis and treatment of this pollinosis.