Evolution of MHC-G in humans and primates based on three new 3'UT polymorphisms

MHC-G is a class Ib (non-classical) major histocompatibility complex (MHC) whose functional and evolutionary characteristics are still under scrutiny. The study of noncoding sequences in the MHC genes may provide important phylogenetic information. In this work we have sequenced the MHC-G exon 8, which encodes for the 3'UT region, in different species of primates. It has been shown that: (1) a previously described 14 base pair (bp) deletion polymorphism is human-specific and the HLA-G alleles may be classified according to its absence or presence; (2) another newly described 3 bp deletion/insertion polymorphism is also human-specific; and (3) another newly described 51 bp deletion polymorphism is common to Pongidae and humans, but is not found in other primates belonging to the Cercopithecinae family. A hypothesis on the evolutionary pathway of this gene is put forward in the light of these findings.     

Evaluation of mutagenic activity in an extract of pepper tree stem bark (Schinus terebinthifolius Raddi)

An extract (decoction) from pepper tree stem bark (Schinus terebinthifolius Raddi) is widely used in Brazil as a topical antiinflammatory agent and to cicatrize wounds. The extract contains catechin, tannins, terpenes, flavonoids, and saponins; of these components, both mutagenic potential and antioxidant properties have been ascribed to flavonoids. The mutagenicity of some flavonoids is believed to be associated with the formation of reactive oxygen species and seems to depend on the number and position of hydroxyl groups. In the present study, we evaluated an extract of S. terebinthifolius in a series of cell-free and bacterial assays in order to determine its genotoxic potential. The extract was negative in a cell-free plasmid DNA test, indicating that it did not directly break DNA. Positive results, however, were obtained in the SOS chromotest, in a forward mutagenesis assay employing CC104 and CC104mutMmutY strains of Escherichia coli, and in the Salmonella reversion assay, using strains TA97, TA98, TA100, and TA102. All the bacterial tests were performed without exogenous metabolic activation due to the topical use of this preparation. The results indicate that pepper tree stem bark extract produces DNA damage and mutation in bacteria, and that oxidative damage may be responsible for the genotoxicity.     

Detection of Borrelia lonestari,putative agent of southern tick-associated rash illness,in white-tailed deer (Odocoileus virginianus) from the southeastern United States

To determine if white-tailed deer may serve as a reservoir host for Borrelia lonestari, we used a nested PCR for the Borrelia flagellin gene to evaluate blood samples collected from deer from eight southeastern states. Seven of 80 deer (8.7%) from 5 of 17 sites (29.4%) had sequence-confirmed evidence of a B. lonestari flagellin gene by PCR, indicating that deer are infected with B. lonestari or another closely related Borrelia species. Our findings expand the known geographic range of B. lonestari and provide the first evidence of this organism in a vertebrate other than humans.     

Is the outcome of in-vitro fertilization and embryo transfer treatment improved by spontaneous or surgical drainage of a hydrosalpinx?

A pilot study was designed to examine whether the outcome of embryo transfer in women with a hydrosalpinx might be improved by surgical drainage of the hydrosalpinx at the time of oocyte collection for in- vitro fertilization treatment. A comparative, controlled but retrospective analysis of the results was performed of all women with infective tubal damage aged <40 years old, who had ovulatory cycles, a normal uterus and a partner with normal spermatozoa. A standardized treatment regimen was used. A maximum of three embryos were transferred. Hydrosalpinx was defined by prior hysterosalpingography and/or laparoscopy with transcervical dye injection. A total of 237 embryo transfer cycles in women with hydrosalpinges (tubal distension not visible in 151, visible but not drained in 30 and drained in 56) were compared with 705 embryo transfer cycles in women with tubal disease but no hydrosalpinx. Results were analysed in the first three cycles but also separately in the first cycle to check for bias. Success rates were higher in the first cycle, but did not significantly influence overall differences. Implantation rates were significantly reduced overall in the hydrosalpinx group (8.0 versus 13.2% for controls; P < 0.001), being 8.3% (P < 0.01) in the subgroup without evident tubal distension and 7.5% (not significant) in the drained hydrosalpinx group. This study shows that tubal damage with distal occlusion is associated with a marked reduction in embryo implantation, even in the absence of obvious fluid distension. Surgical drainage of distended hydrosalpinges appears to offer no benefit.      

Identification and characterization of aquaporin-2 water channel mutations causing nephrogenic diabetes insipidus with partial vasopressin response

Congenital nephrogenic diabetes insipidus (NDI) is a rare disease caused most often by mutations in the vasopressin V2 receptor (AVPR2). We studied a family which included a female patient with NDI with symptoms dating from infancy. The patient responded to large doses of desmopressin (dDAVP) which decreased urine volume from 10 to 4 I/day. Neither the parents nor the three sisters were polyuric. The patient was found to be a compound heterozygote for two novel recessive point mutations in the aquaporin-2 (AQP2) gene: L22V in exon 1 and C181W in exon 3. Residue Cys181 in AQP2 is the site for inhibition of water permeation by mercurial compounds and is located near to the NPA motif conserved in all aquaporins. Osmotic water permeability (Pf) in Xenopus oocytes injected with cRNA encoding C181W-AQP2 was not increased over water control, while expression of L22V cRNA increased the Pf to approximately 60% of that for wild-type AQP2. Co-injection of the mutant cRNAs with the wild-type cRNA did not affect the function of the wild-type AQP2. Immunolocalization of AQP2-transfected CHO cells showed that the C181W mutant had an endoplasmic reticulum-like intracellular distribution, whereas L22V and wild-type AQP2 showed endosome and plasma membrane staining. Water permeability assays showed a high Pf in cells expressing wild-type and L22V AQP2. This study indicates that AQP2 mutations can confer partially responsive NDI.      

A genetic and clinical study of individuals with nonsyndromic retinopathy consequent upon sequence variants in HGSNAT,the gene associated with Sanfilippo C mucopolysaccharidosis

Pathogenic variants in the gene HGSNAT (heparan‐α‐glucosaminide N‐acetyltransferase) have been reported to underlie two distinct recessive conditions, depending on the specific genotype, mucopolysaccharidosis type IIIC (MPSIIIC)—a severe childhood‐onset lysosomal storage disorder, and adult‐onset nonsyndromic retinitis pigmentosa (RP). Here we describe the largest cohort to‐date of HGSNAT‐associated nonsyndromic RP patients, and describe their retinal phenotype, leukocyte enzymatic activity, and likely pathogenic genotypes. We identified biallelic HGSNAT variants in 17 individuals (15 families) as the likely cause of their RP. None showed any other symptoms of MPSIIIC. All had a mild but significant reduction of HGSNAT enzyme activity in leukocytes. The retinal condition was generally of late‐onset, showing progressive degeneration of a concentric area of paramacular retina, with preservation but reduced electroretinogram responses. Symptoms, electrophysiology, and imaging suggest the rod photoreceptor to be the cell initially compromised. HGSNAT enzymatic testing was useful in resolving diagnostic dilemmas in compatible patients. We identified seven novel sequence variants [p.(Arg239Cys); p.(Ser296Leu); p.(Phe428Cys); p.(Gly248Ala); p.(Gly418Arg), c.1543‐2A>C; c.1708delA], three of which were considered to be retina‐disease‐specific alleles. The most prevalent retina‐disease‐specific allele p.(Ala615Thr) was observed heterozygously or homozygously in 8 and 5 individuals respectively (7 and 4 families). Two siblings in one family, while identical for the HGSNAT locus, but discordant for retinal disease, suggest the influence of trans‐acting genetic or environmental modifying factors.     

The role of size, sequence and haplotype in the stability of FRAXA and FRAXE alleles during transmission

Factors involved in the stability of trinucleotide repeats during transmission were studied in 139 families in which a full mutation, premutation or intermediate allele at either FRAXA or FRAXE was segregating. The transmission of alleles at FRAXA, FRAXE and four microsatellite loci were recorded for all individuals. Instability within the minimal and common ranges (0-40 repeats for FRAXA, 0-30 repeats for FRAXE) was extremely rare; only one example was observed, an increased in size at FRAXA from 29 to 39 repeats. Four FRAXA and three FRAXE alleles in the intermediate range (41-60) repeats for FRAXA, 31-60 for FRAXE) were unstably transmitted. Instability was more frequent for FRAXA intermediate alleles that had a tract of pure CGG greater than 37 although instability only occurred in two of 13 such transmissions: the changes observed were limited to only one or two repeats. Premutation FRAXA alleles over 100 repeats expanded to a full mutation during female transmission in 100% of cases, in agreement with other published series. There was no clear correlation between haplotype and probability of expansion of FRAXA premutations. Instability at FRAXA or FRAXE was more often observed in conjunction with a second instability at an independent locus suggesting genomic instability as a possible mechanism by which at least some FRAXA and FRAXE mutations arise.      

IMXQB-80: A Quillaja brasiliensis saponin-based nanoadjuvant enhances Zika virus specific immune responses in mice

Vaccine adjuvants are compounds that enhance/prolong the immune response to a co-administered antigen. Saponins have been widely used as adjuvants for many years in several vaccines – especially for intracellular pathogens – including the recent and somewhat revolutionary malaria and shingles vaccines. In view of the immunoadjuvant potential of Q. brasiliensis saponins, the present study aimed to characterize the QB-80 saponin-rich fraction and a nanoadjuvant prepared with QB-80 and lipids (IMXQB-80). In addition, the performance of such adjuvants was examined in experimental inactivated vaccines against Zika virus (ZIKV). Analysis of QB-80 by DI-ESI-ToF by negative ion electrospray revealed over 29 saponins that could be assigned to known structures existing in their congener Q. saponaria, including the well-studied QS-21 and QS-7. The QB-80 saponins were a micrOTOF able to self-assembly with lipids in ISCOM-like nanoparticles with diameters of approximately 43 nm, here named IMXQB-80. Toxicity assays revealed that QB-80 saponins did present some haemolytical and cytotoxic potentials; however, these were abrogated in IMXQB-80 nanoparticles. Regarding the adjuvant activity, QB-80 and IMXQB-80 significantly enhanced serum levels of anti-Zika virus IgG and subtypes (IgG1, IgG2b, IgG2c) as well as neutralized antibodies when compared to an unadjuvanted vaccine. Furthermore, the nanoadjuvant IMXQB-80 was as effective as QB-80 in stimulating immune responses, yet requiring fourfold less saponins to induce the equivalent stimuli, and with less toxicity. These findings reveal that the saponin fraction QB-80, and particularly the IMXQB-80 nanoadjuvant, are safe and capable of potentializing immune responses when used as adjuvants in experimental ZIKV vaccines.