Characterization of the brain virome in human immunodeficiency virus infection and substance use disorder.

Viruses can infect the brain in individuals with and without HIV-infection: however, the brain virome is poorly characterized. Metabolic alterations have been identified which predispose people to substance use disorder (SUD), but whether these could be triggered by viral infection of the brain is unknown. We used a target-enrichment, deep sequencing platform and bioinformatic pipeline named "ViroFind", for the unbiased characterization of DNA and RNA viruses in brain samples obtained from the National Neuro-AIDS Tissue Consortium. We analyzed fresh frozen post-mortem prefrontal cortex from 72 individuals without known viral infection of the brain, including 16 HIV+/SUD+, 20 HIV+/SUD-, 16 HIV-/SUD+, and 20 HIV-/SUD-. The average age was 52.3 y and 62.5% were males. We identified sequences from 26 viruses belonging to 11 viral taxa. These included viruses with and without known pathogenic potential or tropism to the nervous system, with sequence coverage ranging from 0.03 to 99.73% of the viral genomes. In SUD+ people, HIV-infection was associated with a higher total number of viruses, and HIV+/SUD+ compared to HIV-/SUD+ individuals had an increased frequency of Adenovirus (68.8 vs 0%; p<0.001) and Epstein-Barr virus (EBV) (43.8 vs 6.3%; p=0.037) as well as an increase in Torque Teno virus (TTV) burden. Conversely, in HIV+ people, SUD was associated with an increase in frequency of Hepatitis C virus, (25 in HIV+/SUD+ vs 0% in HIV+/SUD-; p=0.031). Finally, HIV+/SUD- compared to HIV-/SUD- individuals had an increased frequency of EBV (50 vs 0%; p<0.001) and an increase in TTV viral burden, but a decreased Adenovirus viral burden. These data demonstrate an unexpectedly high variety in the human brain virome, identifying targets for future research into the impact of these taxa on the central nervous system. ViroFind could become a valuable tool for monitoring viral dynamics in various compartments, monitoring outbreaks, and informing vaccine development.

Phlebitis in Takayasu arteritis: A case-based literature review.

Takayasu arteritis (TAK) is the main type of large vessel arteritis in young adults, which mainly affects the aorta and its main branches, leading to clinical manifestations such as syncope, intermittent limb claudication, hypertension, and abdominal pain. Among them, venous involvement is rarely reported. Here we show a case of TAK presenting as phlebitis. This was a 27-year-old woman, who initially admitted to our hospital with myalgia of the upper and lower extremities and night sweats. She was diagnosed as TAK according to the 1990 American College of Rheumatology TAK criteria. Surprisingly, vascular ultrasonography revealed wall thickening as indicated by macaroni sign of the multiple veins. TAK phlebitis appeared at the active phase, while disappearing rapidly at remission. Phlebitis might have a close relationship with disease activity. By retrospective study in our department, the estimated incidence rate of phlebitis might be 9.1% in TAK. With the literature review, it revealed that phlebitis might be an ignored manifestation in active TAK. However, due to the smaller sample size, it should be noted that a direct cause-effect relationship cannot be established.

A novel SPE-LC-MRM strategy for serum demethylzeylasteral quantitation developed with an 18O-labeled internal standard.

Natural bioactive compounds (NBCs) are widely used in clinical treatment. For example, Tripterygium wilfordii Hook f. is commonly known in China as Lei-Gong-Teng which means thunder god vine. This herb is widely distributed in Eastern and Southern China, Korea, and Japan. The natural bioactive compounds of this herb can be extracted and made into tripterygium glycoside tablets. It is one of the most commonly used and effective traditional Chinese herbal medicines against rheumatoid arthritis (RA), nephrotic syndrome (NS), autoimmune hepatis (AIH), and so on. However, many NBCs are difficult to reliably quantify in the serum due to the effects of matrix and RSD. In addition, the targeted compound's internal standard (IS) is rarely sold due to the complex isotope internal standard synthesis pathway. In this study, a new quantitation method for 18O labeling combined with off-line SPE was formulated. We contrasted the recoveries and matrix effects of various separation methods in order to choose the best method. Furthermore, we optimized the conditions for SPE loading and washing. An isotopic internal standard was prepared by the 16O/18O exchanging reaction in order to eliminate the matrix effects. The method's accuracy and precision met the requirements for method validation. The recovery of this method was close to 60%. The relative standard deviation (RSD) of the high-concentration sample was 2%, and the limit of detection (LOD) was 1 ng/mL. This method could be used to analyze the clinical serum concentration of demethylzeylasteral. Sixty samples were collected from 10 patients with diabetes nephropathy. The quantitation results of demethylzeylasteral in patients' serum obtained using this method exhibited a correlation between therapeutic drug monitoring (TDM) and decreased urinary protein. This work may have broad implications for the study of drug metabolism in vivo and the clinical application of low-abundance and difficult-to-quantify NBCs.

Characterizing Epstein-Barr virus infection of the central nervous system in Zambian adults living with HIV.

The significance of Epstein-Barr virus (EBV) detection in the cerebrospinal spinal fluid (CSF) in people living with HIV (PLWH) is not entirely understood. The detection of EBV DNA may represent active central nervous system (CNS) infection, reactivation in the setting of another CNS pathogen or due to impaired immunity, or detection of quiescent virus. We screened 470 adult PLWH in Zambia with neurological symptoms for the presence of EBV DNA in the CSF. We performed quantitative EBV PCR on the CSF and blood. We then performed quantitative EBV DNA PCR on the blood of controls with documented HIV viral suppression without CNS symptoms. The prevalence of EBV DNA in the CSF of patients with CNS symptoms was 28.9% (136/470). EBV DNA positivity was associated with younger age, shorter duration of HIV diagnosis, lower CSF glucose levels, higher CSF protein and white blood cell levels, and a positive CSF Mycobacterium tuberculosis result. The median EBV DNA load was 8000 cps/mL in both the CSF and blood with a range of 2000-2,753,000 cps/mL in the CSF and 1000 to 1,871,000 cps/mL in the blood. Molecular screening of CSF for other possible causes of infection identified Mycobacterium tuberculosis in 30.1% and cytomegalovirus (CMV) in 10.5% of samples. EBV DNA load in the blood and CSF was not associated with mortality. Our results suggest that even though EBV DNA was commonly detected in the CSF of our population, it appears to have limited clinical significance regardless of EBV DNA load.

A live-cell image-based machine learning strategy for reducing variability in PSC differentiation systems.

The differentiation of pluripotent stem cells (PSCs) into diverse functional cell types provides a promising solution to support drug discovery, disease modeling, and regenerative medicine. However, functional cell differentiation is currently limited by the substantial line-to-line and batch-to-batch variabilities, which severely impede the progress of scientific research and the manufacturing of cell products. For instance, PSC-to-cardiomyocyte (CM) differentiation is vulnerable to inappropriate doses of CHIR99021 (CHIR) that are applied in the initial stage of mesoderm differentiation. Here, by harnessing live-cell bright-field imaging and machine learning (ML), we realize real-time cell recognition in the entire differentiation process, e.g., CMs, cardiac progenitor cells (CPCs), PSC clones, and even misdifferentiated cells. This enables non-invasive prediction of differentiation efficiency, purification of ML-recognized CMs and CPCs for reducing cell contamination, early assessment of the CHIR dose for correcting the misdifferentiation trajectory, and evaluation of initial PSC colonies for controlling the start point of differentiation, all of which provide a more invulnerable differentiation method with resistance to variability. Moreover, with the established ML models as a readout for the chemical screen, we identify a CDK8 inhibitor that can further improve the cell resistance to the overdose of CHIR. Together, this study indicates that artificial intelligence is able to guide and iteratively optimize PSC differentiation to achieve consistently high efficiency across cell lines and batches, providing a better understanding and rational modulation of the differentiation process for functional cell manufacturing in biomedical applications.

Ocean acidification drives gut microbiome changes linked to species-specific immune defence.

Ocean acidification (OA) has important effects on the intrinsic phenotypic characteristics of many marine organisms. Concomitantly, OA can alter the extended phenotypes of these organisms by perturbing the structure and function of their associated microbiomes. It is unclear, however, the extent to which interactions between these levels of phenotypic change can modulate the capacity for resilience to OA. Here, we explored this theoretical framework assessing the influence of OA on intrinsic (immunological responses and energy reserve) and extrinsic (gut microbiome) phenotypic characteristics and the survival of important calcifiers, the edible oysters Crassostrea angulata and C. hongkongensis. After one-month exposure to experimental OA (pH 7.4) and control (pH 8.0) conditions, we found species-specific responses characterised by elevated stress (hemocyte apoptosis) and decreased survival in the coastal species (C. angulata) compared with the estuarine species (C. hongkongensis). Phagocytosis of hemocytes was not affected by OA but in vitro bacterial clearance capability decreased in both species. Gut microbial diversity decreased in C. angulata but not in C. hongkongensis. Overall, C. hongkongensis was capable of maintaining the homeostasis of the immune system and energy supply under OA. In contrast, C. angulata's immune function was suppressed, and the energy reserve was imbalanced, which might be attributed to the declined microbial diversity and the functional loss of essential bacteria in the guts. This study highlights a species-specific response to OA determined by genetic background and local adaptation, shedding light on the understanding of host-microbiota-environment interactions in future coastal acidification.

Directional fabrication and dissolution of larval and juvenile oyster shells under ocean acidification.

Biomineralization is one of the key biochemical processes in calcifying bivalve species such as oysters that is affected by ocean acidification (OA). Larval life stages of oysters are made of aragonite crystals whereas the adults are made of calcite and/or aragonite. Though both calcite and aragonite are crystal polymorphs of calcium carbonate, they have different mechanical properties and hence it is important to study the micro and nano structure of different life stages of oyster shells under OA to understand the mechanisms by which OA affects biomineralization ontogeny. Here, we have studied the larval and juvenile life stages of an economically and ecologically important estuarine oyster species, Crassostrea hongkongensis, under OA with focus over shell fabrication under OA (pHNBS 7.4). We also look at the effect of parental exposure to OA on larvae and juvenile microstructure. The micro and nanostructure characterization reveals directional fabrication of oyster shells, with more organized structure as biomineralization progresses. Under OA, both the larval and juvenile stages show directional dissolution, i.e. the earlier formed shell layers undergo dissolution at first, owing to longer exposure time. Despite dissolution, the micro and nanostructure of the shell remains unaffected under OA, irrespective of parental exposure history.

Wild oyster population resistance to ocean acidification adversely affected by bacterial infection.

The carbon dioxide induced ocean acidification (OA) process is well known to have profound effects on physiology, survival and immune responses in marine organisms, and particularly calcifiers including edible oysters. At the same time, some wild populations could develop a complex and sophisticated immune system to cope with multiple biotic and abiotic stresses, such as bacterial infections and OA, over the long period of coevolution with the environment. However, it is unclear how immunological responses and the underlying mechanisms are altered under the combined effect of OA and bacterial infection, especially in the ecologically and economically important edible oysters. Here, we collected the wild population of oyster species Crassostrea hongkongensis (the Hong Kong oyster) from their native estuarine area and carried out a bacterial challenge with the worldwide pervasive pathogen of human foodborne disease, Vibrio parahaemolyticus, to investigate the host immune responses and molecular mechanisms under the high-CO2 and low pH-driven OA conditions. The wild population had a high immune resistance to OA, but the resistance is compromised under the combined effect of OA and bacterial infection both in vivo or in vitro. We classified all transcriptomic genes based on expression profiles and functional pathways and identified the specifically switched on and off genes and pathways under combined effect. These genes and pathways were mainly involved in multiple immunological processes including pathogen recognition, immune signal transduction and effectors. This work would help understand how the immunological function and mechanism response to bacterial infection in wild populations and predict the dynamic distribution of human health-related pathogens to reduce the risk of foodborne disease under the future climate change scenario.

Epigenetic-associated phenotypic plasticity of the ocean acidification-acclimated edible oyster in the mariculture environment.

For marine invertebrates with a pelagic-benthic life cycle, larval exposure to ocean acidification (OA) can affect adult performance in response to another environmental stressor. This carry-over effect has the potential to alter phenotypic traits. However, the molecular mechanisms that mediate "OA"-triggered carry-over effects have not been explored despite such information being key to improving species fitness and management strategies for aquafarming. This study integrated the genome-wide DNA methylome and transcriptome to examine epigenetic modification-mediated carry-over OA impacts on phenotypic traits of the ecologically and commercially important oyster species Crassostrea hongkongensis under field conditions. Larvae of C. hongkongensis were exposed to control pH 8.0 and low pH 7.4 conditions, mimicking near future OA scenario in their habitat, before being outplanted as post-metamorphic juveniles at two mariculture field sites with contrasting environmental stressors for 9 months. The larval carry-over OA effect was found to have persistent impacts on the growth and survival trade-off traits on the outplanted juveniles, although the beneficial or adverse effect depended on the environmental conditions at the outplanted sites. Site-specific plasticity was demonstrated with a diverse DNA methylation-associated gene expression profile, with signal transduction and the endocrine system being the most common and highly enriched functions. Highly methylated exons prevailed in the key genes related to general metabolic and endocytic responses and these genes are evolutionarily conserved in various marine invertebrates in response to OA. These results suggest that oysters with prior larval exposure history to OA had the ability to trigger rapid local adaptive responses via epigenetic modification to cope with multiple stressors in the field.

Epstein-Barr Virus Detection in the Central Nervous System of HIV-Infected Patients.

Simply detecting Epstein-Barr virus deoxyribonucleic acid (EBV-DNA) is insufficient to diagnose EBV-associated diseases. The current literature around EBV-DNA detection from cerebrospinal fluid (CSF) in human immunodeficiency virus (HIV)-positive non-lymphoma patients was systematically reviewed and a meta-analysis reporting the estimated pooled prevalence in this population when PCR methods are employed, targeting different sequence segments within the EBV genome, was conducted. Using a combination of three key concepts-Epstein-Barr virus detection, central nervous system disease, and human cerebrospinal fluid-and their MeSH terms, the PubMed database was searched. A total of 273 papers reporting the detection of EBV in CNS were screened, of which 13 met the inclusion criteria. The meta-analysis revealed a pooled prevalence of EBV-DNA in CSF of 20% (CI: 12-31%). The highest pooled prevalence was from studies conducted on the African population at 39% (CI: 27-51%). The investigation of the presence of EBV-DNA in the CSF was also very varied, with several gene targets used. While most patients from the articles included in this review and meta-analysis were symptomatic of CNS disorders, the pathogenicity of EBV in non-lymphoma HIV patients when detected in CSF has still not been determined. The presence of EBV-DNA in the CNS remains a concern, and further research is warranted to understand its significance in causing CNS disorders.

Lupus nephritis: new progress in diagnosis and treatment.

Systemic lupus erythematosus (SLE) is a chronic multifactorial autoimmune disease that affects many organs, including the kidney. Lupus nephritis (LN) is a common manifestation characterized by heterogeneous clinical and histopathological findings, and often associates with poor prognosis. The diagnosis and treatment of LN is challenging, depending largely on renal biopsy, and there is no reliable non-invasive LN biomarker. Up to now, the complete remission rate of LN is only 20%∼30% after receiving six months of standard treatment, which is far from satisfactory. Moreover, adverse reactions to immunosuppressants, especially glucocorticoids, further compromise the prognosis of LN. Biological reagents targetting autoimmune responses and inflammatory pathways, bring hope to the treatment of intractable lupus. The European Renal Association-European Dialysis and Transplant Association (EULAR/ERA-EDTA) and KDIGO (Kidney Disease: Improving Global Outcomes) have been working on and launched the recommendations for the management of LN. In this review, we update our knowledge in the pathogenesis, diagnosis, and management of LN and prospect for the future potential targets in the management of LN.

One-Pot Enzymatic Synthesis and Biological Evaluation of Ganglioside GM3 Derivatives as Potential Cancer Immunotherapeutics.

Cancer is a leading cause of death worldwide. Recent research studies have revealed that GM3 derivatives have considerable promise as potential therapeutic agents for cancer. To discover novel GM3 derivatives as potential antitumor agents, a one-pot enzymatic synthesis was established, yielding 14 GM3 derivatives in high total yields (22-41%). Subsequently, the inhibitory activities of GM3 derivatives were assessed by wound-healing assays and Transwell assays and tumor-bearing animal models. Among all the GM3 derivatives, N-12 showed excellent migration and invasion inhibitory effects in cells and marked antitumor activity in C57BL/6 mice. The subsequent analysis of cancer tissues and serum samples revealed that N-12 induces tumor inhibition, which was closely related to immune response. Taken together, N-12 can be further developed as an effective therapeutic for the treatment of cancer. An RNA-sequencing (RNA-seq) analysis was then performed and indicated that the antitumor mechanism of N-12 involved focal adhesion and ECM-receptor interaction signaling pathways.

LTA4H Prevalence and Mortality in Adult Zambians with Tuberculous Meningitis.

We conducted a prospective cohort study to determine the prevalence of leukotriene A4 hydrolase (LTA4H) polymorphisms in Zambian adults with tuberculous meningitis (TBM) and its association with mortality. We completed genotype testing on 101 definite cases of TBM and 119 consecutive non-TBM controls. The distribution of genotypes among TBM patients was as follows: C/C (0.83), C/T (0.14), T/T (0.03). There was no significant difference in genotype distribution between TBM and non-TBM patients. We found no relationship between LTA4H polymorphism and survival. Prospective studies are needed to determine the benefit of adjuvant steroids in TBM based upon population LTA4H genotype. ANN NEUROL 2021;90:994-998.

Teriflunomide Inhibits JCPyV Infection and Spread in Glial Cells and Choroid Plexus Epithelial Cells.

Several classes of immunomodulators are used for treating relapsing-remitting multiple sclerosis (RRMS). Most of these disease-modifying therapies, except teriflunomide, carry the risk of progressive multifocal leukoencephalopathy (PML), a severely debilitating, often fatal virus-induced demyelinating disease. Because teriflunomide has been shown to have antiviral activity against DNA viruses, we investigated whether treatment of cells with teriflunomide inhibits infection and spread of JC polyomavirus (JCPyV), the causative agent of PML. Treatment of choroid plexus epithelial cells and astrocytes with teriflunomide reduced JCPyV infection and spread. We also used droplet digital PCR to quantify JCPyV DNA associated with extracellular vesicles isolated from RRMS patients. We detected JCPyV DNA in all patients with confirmed PML diagnosis (n = 2), and in six natalizumab-treated (n = 12), two teriflunomide-treated (n = 7), and two nonimmunomodulated (n = 2) patients. Of the 21 patients, 12 (57%) had detectable JCPyV in either plasma or serum. CSF was uniformly negative for JCPyV. Isolation of extracellular vesicles did not increase the level of detection of JCPyV DNA versus bulk unprocessed biofluid. Overall, our study demonstrated an effect of teriflunomide inhibiting JCPyV infection and spread in glial and choroid plexus epithelial cells. Larger studies using patient samples are needed to correlate these in vitro findings with patient data.

Design, synthesis and antitumor activities of thiazole-containing mitochondrial targeting agents.

In this study, a novel batch of thiazole-containing mitochondrial targeting agents were designed and synthesized. Four kinds of mitochondrial targeting moieties and six kinds of linkers were designed. Their structures were confirmed by NMR and HR-MS. The screening of antiproliferative activity revealed that most compounds displayed cytotoxicity on HeLa cancer cell. In particular, D1 has an IC50 value of 35.32 µmol·L-1 against HeLa cell. In addition, cellular respiratory activities were also tested on HeLa cancer cells. D1 had a basal oxygen consumption rate of 8.84 pmol·s-1·mL-1. Also, D1 inhibited the mitochondrial respiration of HeLa cell significantly at 5 µmol·L-1, as well as a complete inhibitory of oxygen consumption for cellular ATP coupling. Furthermore, the pKa, logP, and logD under different pH conditions of all the compounds were calculated by the ACD/Percepta-PhysChem Suite, and the results manifested the correlation between physicochemical properties and chemical activity of compounds. The results identify D1 as a promising mitochondria inhibitor and anticancer agent with appropriate physicochemical properties.

Biological evaluation of mitochondria targeting small molecules as potent anticancer drugs.

Cancer therapy targets specific metabolic pathways or a single gene. This may result in low therapeutic effects due to drug selectivity and drug resistance. Recent studies revealed that the mitochondrial membrane potential and transmembrane permeability of cancerous mitochondria are differed from normal mitochondria. Thus, chemotherapy targeting cancerous mitochondria could be an innovative and competent strategy for cancer therapy. Previously, our work with a novel group of mitochondria targeting small molecules presented promising inhibitory capability toward various cancer cell lines and suppressed adenosine triphosphate (ATP) generation. Therefore, it is critical to understand the anticancer effect and targeting mechanism of these small molecules. This study investigated the inhibitory activity of mitochondria targeting small molecules with human cervical cancer cells - HeLa to further explore their therapeutic potential. HeLa cells were exposed to 10 µM of synthesized compounds and presented elevation in intracellular reactive oxygen species (ROS) level, impaired mitochondrial membrane potential and upregulation of apoptosis as well as necrosis. In vivo, HeLa cell tumor-bearing BALB/c nude mice were treated with mitochondria targeting small molecules for 12 days consecutively. Throughout this chemotherapy study, no deleterious side effects nor the appearance of toxicity was observed. Furthermore, mitochondria targeting small molecules treated groups exhibited significant down-regulation with both tumor volume and tumor weight compared to the Doxorubicin (DOX) treated group. Thus, inhibition of mitochondrial ATP synthesis, activation of intracellular ROS production, down-regulation of mitochondrial membrane potential and upregulation of apoptosis and necrosis rates are the indications of cancer therapy. In this work, we examined the anticancer capability of four mitochondria targeting small molecules in vitro and in vivo, and demonstrated a novel therapeutic approach in cancer therapy with tremendous potential.

Design, synthesis and biological evaluation of novel thiazole-derivatives as mitochondrial targeting inhibitors of cancer cells.

Mitochondria are pivotal energy production sources for cells to maintain necessary metabolism activities. Targeting dysfunctional mitochondrial features has been a hotspot for mitochondrial-related disease researches. Investigation with cancerous mitochondrial metabolism is a continuing concern within tumor therapy. Herein, we set out to assess the anti-cancer activities of a novel family of TPP-thiazole derivatives based on our earlier research on mitochondrial targeting agents. Specifically, we designed and synthesized a series of TPP-thiazole derivatives and revealed by the MTT assay that most synthesized compounds effectively inhibited three cancer cell lines (HeLa, PC3 and MCF-7). After structure modifications, we explored the SAR relationships and identified the most promising compound R13 (IC50 of 5.52 µM) for further investigation. In the meantime, we performed ATP production assay to assess the selected compounds inhibitory effect on HeLa cells energy production. The results displayed the test compounds significantly restrained ATP production of cancer cells. Overall, we have designed and synthesized a series of compounds which exhibited significant cytotoxicity against cancer cells and effectively inhibited mitochondrial energy production.

Transgenerational responses to seawater pH in the edible oyster, with implications for the mariculture of the species under future ocean acidification.

The majority of common edible oysters are projected to grow more slowly and have smaller impaired shells because of anthropogenic CO2-induced reductions in seawater carbonate ion concentration and pH, a process called ocean acidification (OA). Recent evidence has shown that OA has carryover effects, for example, larvae exposed to OA will also exhibit either positive or negative effects after metamorphosis. This study examined the hidden carryover effects of OA exposure during parental and larval stages on post-metamorphic traits of the commercially important oyster species Crassostrea hongkongensis. Adults of C. hongkongensis were exposed to control pH (pHNBS 8.0) and OA-induced low pH (pHNBS 7.4) conditions. Their larval offspring were then exposed to the same aquarium conditions before being out-planted as post-metamorphic juveniles at a mariculture site for 10 months. Initially, larval offspring were resilient to low pH with or without parental exposure. The larvae exposed to low pH had significantly faster development and higher percentage of settlement success compared to control groups. The out-planted juveniles with parental exposure had improved survival and growth compared to juveniles without parental exposure, regardless of the larval exposure history. This implies that transgenerational effects due to parental exposure not only persists but also have a greater influence than the within-generational effects of larval exposure. Our results shed light on the importance of linking the various life history stages when assessing the OA-induced carryover capacity of C. hongkongensis in the natural environment. Understanding these linked relationships helps us better predict the species rapid adaptation responses in the face of changing coastal conditions due to OA.

Estimating Feature-Label Dependence Using Gini Distance Statistics.

Identifying statistical dependence between the features and the label is a fundamental problem in supervised learning. This paper presents a framework for estimating dependence between numerical features and a categorical label using generalized Gini distance, an energy distance in reproducing kernel Hilbert spaces (RKHS). Two Gini distance based dependence measures are explored: Gini distance covariance and Gini distance correlation. Unlike Pearson covariance and correlation, which do not characterize independence, the above Gini distance based measures define dependence as well as independence of random variables. The test statistics are simple to calculate and do not require probability density estimation. Uniform convergence bounds and asymptotic bounds are derived for the test statistics. Comparisons with distance covariance statistics are provided. It is shown that Gini distance statistics converge faster than distance covariance statistics in the uniform convergence bounds, hence tighter upper bounds on both Type I and Type II errors. Moreover, the probability of Gini distance covariance statistic under-performing the distance covariance statistic in Type II error decreases to 0 exponentially with the increase of the sample size. Extensive experimental results are presented to demonstrate the performance of the proposed method.

DNA methylation changes in response to ocean acidification at the time of larval metamorphosis in the edible oyster, Crassostrea hongkongensis.

Unprecedented rate of increased CO2 level in the ocean and the subsequent changes in carbonate system including decreased pH, known as ocean acidification (OA), is predicted to disrupt not only the calcification process but also several other physiological and developmental processes in a variety of marine organisms, including edible oysters. Nonetheless, not all species are vulnerable to those OA threats, e.g. some species may be able to cope with OA stress using environmentally induced modifications on gene and protein expressions. For example, external environmental stressors including OA can influence the addition and removal of methyl groups through epigenetic modification (e.g. DNA methylation) process to turn gene expression "on or off" as part of a rapid adaptive mechanism to cope with OA. In this study, we tested the above hypothesis through testing the effect of OA, using decreased pH 7.4 as proxy, on DNA methylation pattern of an endemic and a commercially important estuary oyster species, Crassostrea hongkongensis at the time of larval habitat selection and metamorphosis. Larval growth rate did not differ between control pH 8.1 and treatment pH 7.4. The metamorphosis rate of the pediveliger larvae was higher at pH 7.4 than those in control pH 8.1, however over one-third of the larvae raised at pH 7.4 failed to attach on optimal substrate as defined by biofilm presence. During larval development, a total of 130 genes were differentially methylated across the two treatments. The differential methylation in the larval genes may have partially accounted for the higher metamorphosis success rate under decreased pH 7.4 but with poor substratum selection ability. Differentially methylated loci were concentrated in the exon regions and appear to be associated with cytoskeletal and signal transduction, oxidative stress, metabolic processes, and larval metamorphosis, which implies the high potential of C. hongkongensis larvae to acclimate and adapt through non-genetic ways to OA threats within a single generation.