Predictors of CD4:CD8 Ratio Normalization and Its Effect on Health Outcomes in the Era of Combination Antiretroviral Therapy

Background

HIV leads to CD4:CD8 ratio inversion as immune dysregulation progresses. We examined the predictors of CD4:CD8 normalization after combination antiretroviral therapy (cART) and determined whether normalization is associated with reduced progression to AIDS-defining illnesses (ADI) and death.

Methods

A Canadian cohort of HIV-positive adults with CD4:CD8<1.2 prior to starting cART from 2000–2010 were analyzed. Predictors of (1) reaching a CD4:CD8 ≥1.2 on two separate follow-up visits >30 days apart, and (2) ADI and death from all causes were assessed using adjusted proportional hazards models.

Results

4206 patients were studied for a median of 2.77 years and 306 (7.2%) normalized their CD4:CD8 ratio. Factors associated with achieving a normal CD4:CD8 ratio were: baseline CD4+ T-cells >350 cells/mm³, baseline CD8+ T-cells <500 cells/mm³, time-updated HIV RNA suppression, and not reporting sex with other men as a risk factor. There were 213 ADIs and 214 deaths in 13476 person-years of follow-up. Achieving a normal CD4:CD8 ratio was not associated with time to ADI/death.

Conclusions

In our study, few individuals normalized their CD4:CD8 ratios within the first few years of initiating modern cART. This large study showed no additional short-term predictive value of the CD4:CD8 ratio for clinical outcomes after accounting for other risk factors including age and HIV RNA.     

Habitat degradation impacts black howler monkey (Alouatta pigra) gastrointestinal microbiomes

The gastrointestinal (GI) microbiome contributes significantly to host nutrition and health. However, relationships involving GI microbes, their hosts and host macrohabitats remain to be established. Here, we define clear patterns of variation in the GI microbiomes of six groups of Mexican black howler monkeys (Alouatta pigra) occupying a gradation of habitats including a continuous evergreen rainforest, an evergreen rainforest fragment, a continuous semi-deciduous forest and captivity. High throughput microbial 16S ribosomal RNA gene sequencing indicated that diversity, richness and composition of howler GI microbiomes varied with host habitat in relation to diet. Howlers occupying suboptimal habitats consumed less diverse diets and correspondingly had less diverse gut microbiomes. Quantitative real-time PCR also revealed a reduction in the number of genes related to butyrate production and hydrogen metabolism in the microbiomes of howlers occupying suboptimal habitats, which may impact host health.     

A Multi-Omic Systems-Based Approach Reveals Metabolic Markers of Bacterial Vaginosis and Insight into the Disease

Background

Bacterial vaginosis (BV) is the most common vaginal disorder of reproductive-age women. Yet the cause of BV has not been established. To uncover key determinants of BV, we employed a multi-omic, systems-biology approach, including both deep 16S rRNA gene-based sequencing and metabolomics of lavage samples from 36 women. These women varied demographically, behaviorally, and in terms of health status and symptoms.

Principal Findings

16S rRNA gene-based community composition profiles reflected Nugent scores, but not Amsel criteria. In contrast, metabolomic profiles were markedly more concordant with Amsel criteria. Metabolomic profiles revealed two distinct symptomatic BV types (SBVI and SBVII) with similar characteristics that indicated disruption of epithelial integrity, but each type was correlated to the presence of different microbial taxa and metabolites, as well as to different host behaviors. The characteristic odor associated with BV was linked to increases in putrescine and cadaverine, which were both linked to Dialister spp. Additional correlations were seen with the presence of discharge, 2-methyl-2-hydroxybutanoic acid, and Mobiluncus spp., and with pain, diethylene glycol and Gardnerella spp.

Conclusions

The results not only provide useful diagnostic biomarkers, but also may ultimately provide much needed insight into the determinants of BV.     

Sex hormone control of left ventricular structure/function: mechanistic insights using echocardiography, expression, and DNA methylation analyses in adult mice

Calcium flux into and out of the sarco(endo)plasmic reticulum is vitally important to cardiac function because the cycle of calcium entry and exit controls contraction and relaxation. Putative estrogen and androgen consensus binding sites near to a CpG island are present in the cardiac calsequestrin 2 (CSQ2) promoter. Cardiomyocytes express sex hormone receptors and respond to sex hormones. We hypothesized that sex hormones control CSQ2 expression in cardiomyocytes and so affect cardiac structure/function. Echocardiographic analysis of male and female C57bl6n mice identified thinner walled and lighter hearts in females and significant concentric remodeling after long-term gonadectomy. CSQ2 and sodium-calcium exchanger-1 (NCX1) expression was significantly increased in female compared with male hearts and decreased postovariectomy. NCX1, but not CSQ2, expression was increased postcastration. CSQ2 expression was reduced when H9c2 cells were cultured in hormone-deficient media; increased when estrogen receptor-α (ERα), estrogen receptor-β (ERβ), or androgen agonists were added; and increased in hearts from ERβ-deficient mice. CSQ2 expression was reduced in mice fed a diet low in the methyl donor folic acid and in cells treated with 5-azadeoxycytidine suggesting an involvement of DNA methylation. DNA methylation in CpG in the CSQ2 CpG island was significantly different in males and females and was additionally changed postgonadectomy. Expression of DNA methyltransferases 1, 3a, and 3b was unchanged. These studies strongly link sex hormone-directed changes in CSQ2 expression to DNA methylation with changed expression correlated with altered left ventricular structure and function.     

Aminopeptidase-resistant peptides are targeted to lysosomes and subsequently degraded

Most cytoplasmic and nuclear proteins are degraded via the ubiquitin-proteasome system into peptides, which are subsequently hydrolyzed by downstream aminopeptidases. Inefficient degradation can lead to accumulation of protein fragments, and subsequent aggregation and toxicity. Whereas the role of the proteasome and the effect of its impairment on aggregation have been intensively studied, little is known about how cells deal with peptides that show resistance to degradation by aminopeptidases. Here, we introduced peptidase-resistant peptides into living cells and show that these peptides rapidly and irreversibly accumulate into puncta in the perinuclear region of the cell. Accumulation appears to be independent of peptide sequence but is less efficient for longer peptides. The puncta colocalize with autophagosomal and lysosomal markers, suggesting that these peptides end up within lysosomes via macroautophagy. Surprisingly, the peptides still accumulate within lysosomes when macroautophagy is impaired, suggesting a trafficking route independent of macroautophagy. Upon lysosomal uptake, peptides are degraded, suggesting that cells can clear peptidase-resistant proteasomal products by an alternative pathway, which targets them to lysosomes.     

A Highlights from MBoC Selection: Nebulin binding impedes mutant desmin filament assembly

Desmin intermediate filaments (DIFs) form an intricate meshwork that organizes myofibers within striated muscle cells. The mechanisms that regulate the association of desmin to sarcomeres and their role in desminopathy are incompletely understood. Here we compare the effect nebulin binding has on the assembly kinetics of desmin and three desminopathy-causing mutant desmin variants carrying mutations in the head, rod, or tail domains of desmin (S46F, E245D, and T453I). These mutants were chosen because the mutated residues are located within the nebulin-binding regions of desmin. We discovered that, although nebulin M160–164 bound to both desmin tetrameric complexes and mature filaments, all three mutants exhibited significantly delayed filament assembly kinetics when bound to nebulin. Correspondingly, all three mutants displayed enhanced binding affinities and capacities for nebulin relative to wild-type desmin. Electron micrographs showed that nebulin associates with elongated normal and mutant DIFs assembled in vitro. Moreover, we measured significantly delayed dynamics for the mutant desmin E245D relative to wild-type desmin in fluorescence recovery after photobleaching in live-cell imaging experiments. We propose a mechanism by which mutant desmin slows desmin remodeling in myocytes by retaining nebulin near the Z-discs. On the basis of these data, we suggest that for some filament-forming desmin mutants, the molecular etiology of desminopathy results from subtle deficiencies in their association with nebulin, a major actin-binding filament protein of striated muscle.     

Diversity and enzymatic potentialities of <Emphasis Type="Italic">Bacillus</Emphasis> sp. strains isolated from a polluted freshwater ecosystem in Cuba

Genotypic and phenotypic characterization of Bacillus spp. from polluted freshwater has been poorly addressed. The objective of this research was to determine the diversity and enzymatic potentialities of Bacillus spp. strains isolated from the Almendares River. Bacilli strains from a polluted river were characterized by considering the production of extracellular enzymes using API ZYM. 14 strains were selected and identified using 16S rRNA, gyrB and aroE genes. Genotypic diversity of the Bacillus spp. strains was evaluated using pulsed field gel electrophoresis. Furthermore, the presence of genetic determinants of potential virulence toxins of the Bacillus cereus group and proteinaceous crystal inclusions of Bacillus thuringiensis was determined. 10 strains were identified as B. thuringiensis, two as Bacillus megaterium, one as Bacillus pumilus and one as Bacillus subtilis. Most strains produced proteases, amylases, phosphatases, esterases, aminopeptidases and glucanases, which reflect the abundance of biopolymeric matter in Almendares River. Comparison of the typing results revealed a spatio-temporal distribution among B. thuringiensis strains along the river. The results of the present study highlight the genotypic and phenotypic diversity of Bacillus spp. strains from a polluted river, which contributes to the knowledge of genetic diversity of Bacilli from tropical polluted freshwater ecosystems.     

Evaluation of tRNA intergenic length polymorphism (tDNA-pCR) for the differentiation of the members of the Burkholderia cepacia complex

Nowadays, tentative identification of B. cepacia complex bacteria in routine diagnostic laboratories is based on a combination of selective media, conventional biochemical reactions, commercial test systems and PCR-based assays. Some of these assays have the capacity to discriminate reliably among several members of the B. cepacia complex, however one single method differentiating all B. cepacia-like organisms is not available. In this study, the applicability of tDNA-PCR for the differentiation and rapid identification of the different members of the B. cepacia complex was evaluated. For B. gladioli and most of the B. cepacia genomovars, differentiable patterns were obtained. For some of the members of the B. cepacia complex however, the tDNA-PCR patterns were very similar and sometimes multiple patterns existed within in a single genomovar. No distinction could be made between the tDNA-PCR patterns of B. vietnamiensis and B. pyrrocinia and of B. cepacia genomovars I and VIII respectively. We could conclude that, although tDNA-PCR is not sufficient as a single method to identify all the members of the B. cepacia complex unambiguously or to replace the currently used methods, it is a very fast and easily applicable method that could be a very useful tool for the differentiation and identification of B. cepacia-like organisms.     

Characterisation of wild legume nodulating bacteria (LNB) in the infra-arid zone of Tunisia

We report on the isolation and the characterization of nitrogen-fixing root nodule bacteria isolated from natural legumes in a region of South Tunisia corresponding to the infra-arid climatic zone. A collection of 60 new bacterial root nodule isolates were obtained from 19 legume species belonging to the genera Acacia, Anthyllis, Argyrolobium, Astragalus, Calycotome, Coronilla, Ebenus, Genista, Hedysarum, Hippocrepis, Lathyrus, Lotus, Medicago, Ononis. The isolates were characterised by (1) comparative 16S ARDRA using 7 enzymes, (2) total cell protein SDS-PAGE analysis and (3) 16S rDNA sequencing. The results show that these isolates are diverse and belong to the genera Rhizobium, Sinorhizobium, Mesorhizobium and Bradyrhizobium. Bradyrhizobium were further characterised by 16S-23S rDNA IGS sequencing. Surprisingly strains nodulating Astragalus cruciatus, Lotus creticus and Anthyllis henoniana were identified as Rhizobium galegae, a species recorded only as endosymbiont of Galega officinalis and G. orientalis in northern regions so far.     

The HTT CAG-Expansion Mutation Determines Age at Death but Not Disease Duration in Huntington Disease

Huntington disease (HD) is caused by an expanded HTT CAG repeat that leads in a length-dependent, completely dominant manner to onset of a characteristic movement disorder. HD also displays early mortality, so we tested whether the expanded CAG repeat exerts a dominant influence on age at death and on the duration of clinical disease. We found that, as with clinical onset, HD age at death is determined by expanded CAG-repeat length and has no contribution from the normal CAG allele. Surprisingly, disease duration is independent of the mutation’s length. It is also unaffected by a strong genetic modifier of HD motor onset. These findings suggest two parsimonious alternatives. (1) HD pathogenesis is driven by mutant huntingtin, but before or near motor onset, sufficient CAG-driven damage occurs to permit CAG-independent processes and then lead to eventual death. In this scenario, some pathological changes and their clinical correlates could still worsen in a CAG-driven manner after disease onset, but these CAG-related progressive changes do not themselves determine duration. Alternatively, (2) HD pathogenesis is driven by mutant huntingtin acting in a CAG-dependent manner with different time courses in multiple cell types, and the cellular targets that lead to motor onset and death are different and independent. In this scenario, processes driven by HTT CAG length lead directly to death but not via the striatal pathology associated with motor manifestations. Each scenario has important ramifications for the design and testing of potential therapeutics, especially those aimed at preventing or delaying characteristic motor manifestations.