Genome-wide association study identifies genetic variation in neurocan as a susceptibility factor for bipolar disorder.

We conducted a genome-wide association study (GWAS) and a follow-up study of bipolar disorder (BD), a common neuropsychiatric disorder. In the GWAS, we investigated 499,494 autosomal and 12,484 X-chromosomal SNPs in 682 patients with BD and in 1300 controls. In the first follow-up step, we tested the most significant 48 SNPs in 1729 patients with BD and in 2313 controls. Eight SNPs showed nominally significant association with BD and were introduced to a meta-analysis of the GWAS and the first follow-up samples. Genetic variation in the neurocan gene (NCAN) showed genome-wide significant association with BD in 2411 patients and 3613 controls (rs1064395, p = 3.02 × 10(-8); odds ratio = 1.31). In a second follow-up step, we replicated this finding in independent samples of BD, totaling 6030 patients and 31,749 controls (p = 2.74 × 10(-4); odds ratio = 1.12). The combined analysis of all study samples yielded a p value of 2.14 × 10(-9) (odds ratio = 1.17). Our results provide evidence that rs1064395 is a common risk factor for BD. NCAN encodes neurocan, an extracellular matrix glycoprotein, which is thought to be involved in cell adhesion and migration. We found that expression in mice is localized within cortical and hippocampal areas. These areas are involved in cognition and emotion regulation and have previously been implicated in BD by neuropsychological, neuroimaging, and postmortem studies.

In vitro extracellular replication of Wolbachia endobacteria.

Obligate intracellular endobacteria of the genus Wolbachia are widespread in arthropods and several filarial nematodes. Control programs for vector-borne diseases (dengue, Zika, malaria) and anti-filarial therapy with antibiotics are based on this important endosymbiont. Investigating Wolbachia, however, is impeded by the need for host cells. In this study, the requirements for Wolbachia wAlbB growth in a host cell-free in vitro culture system were characterized via qPCRs. A cell lysate fraction from Aedes albopictus C6/36 insect cells containing cell membranes and medium with fetal bovine serum were identified as requisite for cell-free replication of Wolbachia. Supplementation with the membrane fraction of insect cell lysate increased extracellular Wolbachia replication by 4.2-fold. Replication rates in the insect cell-free culture were lower compared to Wolbachia grown inside insect cells. However, the endobacteria were able to replicate for up to 12 days and to infect uninfected C6/36 cells. Cell-free Wolbachia treated with the lipid II biosynthesis inhibitor fosfomycin had an enlarged phenotype, seen previously for intracellular Wolbachia in C6/36 cells, indicating that the bacteria were unable to divide. In conclusion, we have developed a cell-free culture system in which Wolbachia replicate for up to 12 days, providing an in vitro tool to elucidate the biology of these endobacteria, e.g., cell division by using compounds that may not enter the C6/36 cells. A better understanding of Wolbachia biology, and in particular host-symbiont interactions, is key to the use of Wolbachia in vector control programs and to future drug development against filarial diseases.

The ClpP peptidase of Wolbachia endobacteria is a novel target for drug development against filarial infections.

BACKGROUND: Filarial infections causing lymphatic filariasis or onchocerciasis (river blindness) can be treated with antibiotics (e.g. doxycycline) targeting the essential endosymbiotic Wolbachia bacteria. The depletion of Wolbachia inhibits worm development and causes worm death. Available antibiotics have restrictions for use in children and pregnant or breastfeeding women. Therefore, alternative antibiotics are needed that can be given to all members of the population and that are active with a shorter therapy time. Antibiotics of the acyldepsipeptide class have been shown to inhibit the growth of bacteria by overactivating the peptidase ClpP. The novel mode of action of this class of antibiotics could lead to faster killing of intracellular bacteria. OBJECTIVES: To characterize acyldepsipeptide activity against the Wolbachia ClpP. METHODS: The activity of acyldepsipeptides was investigated against Wolbachia in vitro in insect cells and also against worms in culture. In addition, structural effects were investigated by fluorescence microscopy and electron microscopy. The activity of ClpP was also investigated in vitro. RESULTS: We show that acyldepsipeptides are active against recombinant Wolbachia ClpP and endobacteria resident within insect cells in vitro, and some derivatives were also active against filarial worms in culture. As a consequence of treatment, the worms became immotile and died, the latter confirmed by a viability assay. CONCLUSIONS: The mode of action of the acyldepsipeptides in Wolbachia is the dysregulation of ClpP, causing the uncontrolled degradation of proteins, including the cell division protein FtsZ. Our results demonstrate that wolbachial ClpP is a target for further antifilarial antibiotic discovery.

Requirement of lipid II biosynthesis for cell division in cell wall-less Wolbachia, endobacteria of arthropods and filarial nematodes.

Obligate Wolbachia endobacteria have a reduced genome and retained genes are hypothesized to be crucial for survival. Although intracellular bacteria do not need a stress-bearing peptidoglycan cell wall, Wolbachia encode proteins necessary to synthesize the peptidoglycan precursor lipid II. The activity of the enzymes catalyzing the last two steps of this pathway was previously shown, and Wolbachia are sensitive to inhibition of lipid II synthesis. A puzzling characteristic of Wolbachia is the lack of genes for l-amino acid racemases essential for lipid II synthesis. Transcription analysis showed the expression of a possible alternative racemase metC, and recombinant Wolbachia MetC indeed had racemase activity that may substitute for the absent l-Ala racemase. However, enzymes needed to form mature peptidoglycan are absent and the function of Wolbachia lipid II is unknown. Inhibition of lipid II biosynthesis resulted in enlargement of Wolbachia cells and redistribution of Wolbachia peptidoglycan-associated lipoprotein, demonstrating that lipid II is required for coordinated cell division and may interact with the lipoprotein. We conclude that lipid II is essential for Wolbachia cell division and that this function is potentially conserved in the Gram-negative bacteria.

Cannabis users differ from non-users on measures of personality and schizotypy.

Accumulating evidence indicates that cannabis use may be a risk factor for schizophrenia (SZ), and chronic cannabis users score higher than non-users on measures of schizotypal personality traits. The purpose of the present study was to investigate the relations between normal personality, schizotypy, and cannabis use. Sixty-two chronic cannabis users and 45 cannabis-naïve controls completed a measure of normal personality, the NEO-Five Factor Inventory (NEO-FFI), and two measures of schizotypy, the schizotypal personality questionnaire (SPQ) and perceptual aberration scale (PAS). Substance use was assessed using the SCID I alcohol/drug module and a locally developed drug use questionnaire. On the NEO-FFI, users scored higher than controls on openness, but lower on agreeableness and conscientiousness, and endorsed greater schizotypy on the SPQ and PAS. Higher neuroticism predicted greater schizotypy in both groups, and, higher Extraversion predicted lower negative-syndrome schizotypy among users. Finally, duration of cannabis use was positively correlated with scores on the SPQ and PAS among users, suggesting a relation between overall cannabis use chronicity and schizotypy. These data show that cannabis users differ from non-users on dimensions of normal personality and schizotypy, and provide further evidence that cannabis use is associated with increased levels of psychosis-related personality traits.

Variation in P2RX7 candidate gene (rs2230912) is not associated with bipolar I disorder and unipolar major depression in four European samples.

Two recent studies reported evidence for association between genetic variation of the positional candidate gene P2RX7 on chromosome 12q24 and bipolar I disorder (BPI) [Barden et al. (2006); Am J Med Genet Part B 141B:374-382; McQuillin et al. (2008); Mol Psychiatry 13:1-7] and one study found association with unipolar major depression (Mdd-UP) [Lucae et al. (2006); Hum Mol Genet 15:2438-2445]. In the present work, we aimed to replicate the SNP that showed the strongest association in the above-mentioned studies, namely rs2230912 (P2RX7-E13A) resulting in a change of the amino acid glutamine to arginine at position 460 (Gln460Arg), in four European bipolar I disorder samples from Germany, Poland, Romania, and Russia totaling 1,445 patients, in a German sample of recurrent Mdd-UP patients (N = 640), and a control sample of 2,006 subjects. We found no allelic or genotypic association between rs2230912 and BPI or Mdd-UP both in the national samples and in the combined European patient sample. Additional studies are needed to clarify the potential involvement of P2RX7 and of SNP rs2230912 in the etiology of major affective disorders.

Assessment of forebrain-dependent trace eyeblink conditioning in chronic cannabis users.

While CB1 knockout mice exhibit striking impairments on a cerebellar-dependent task called delay eyeblink conditioning (dEBC), these animals demonstrate intact forebrain-dependent trace EBC (tEBC). Although heavy human cannabis users also show impaired delay EBC, their performance on tEBC is currently unknown. Therefore, 13 heavy cannabis users and 13 cannabis naive controls completed a tEBC procedure. The cannabis group exhibited similar rates of conditioned responding compared to controls in the acquisition and extinction phase. Consistent with reports of overt attentional abnormalities, the cannabis group exhibited decreased N100 ERP amplitudes to the tone CS that were unrelated to mean levels of conditioning across blocks during the acquisition phase. The lack of a significant effect of heavy cannabis use on tEBC reported here, combined with the previous report of impaired dEBC in such users, mirrors the findings observed in CB1 knockout mice, and suggests that the cannabinoid system differentially mediates forebrain- and cerebellar-dependent learning processes in both humans and animals.

Enhancing the effectiveness of biological control programs of invasive species through a more comprehensive pest management approach.

Invasive species are one of the greatest economic and ecological threats to agriculture and natural areas in the US and the world. Among the available management tools, biological control provides one of the most economical and long-term effective strategies for managing widespread and damaging invasive species populations of nearly all taxa. However, integrating biological control programs in a more complete integrated pest management approach that utilizes increased information and communication, post-release monitoring, adaptive management practices, long-term stewardship strategies, and new and innovative ecological and genetic technologies can greatly improve the effectiveness of biological control. In addition, expanding partnerships among relevant national, regional, and local agencies, as well as academic scientists and land managers, offers far greater opportunities for long-term success in the suppression of established invasive species. In this paper we direct our recommendations to federal agencies that oversee, fund, conduct research, and develop classical biological control programs for invasive species. By incorporating these recommendations into adaptive management strategies, private and public land managers will have far greater opportunities for long-term success in suppression of established invasive species. © 2016 Society of Chemical Industry.

Comparison of Certification and Recertification Examinee Performance on Multiple-Choice Items in Forensic Psychiatry.

Research on the association between age and performance on tests of medical knowledge has generally shown an inverse relationship, which is of concern because of the positive association between measures of knowledge and measures of clinical performance. Because the certification and maintenance of certification (MOC) examinations in the subspecialty of forensic psychiatry draw on a common item bank, performance of the two groups of examinees on the same items could be compared. In addition, the relationship between age and test performance was analyzed. Performance on items administered to certification and MOC examinees did not differ significantly, and the mean amount of time spent on each item was similar for the two groups. Although the majority (five of eight) of the correlations between age and test score on the certification and MOC examinations were negative, only three were significant, and the amount of variance explained by age was small. In addition, examination performance for those younger than 50 was similar to those 60 and older, and diplomates recertifying for the second time outperformed those doing so for the first time. These results indicate that in this subspecialty, there is no clear evidence of an age-related decline in knowledge as assessed by multiple-choice items.

A magnetic resonance imaging-safe method for the study of human eyeblink conditioning.

Eyeblink conditioning (EBC) is a widely used translational probe of cerebellar function in both humans and non-human animals. Decades of animal research have identified the cerebellum as critical for EBC. While there is evidence for the involvement of the cerebellum in human EBC, the neural circuitry of EBC in healthy humans has yet to be fully elucidated. The purpose of this study was to design and validate a highly customisable system for EBC stimulus presentation and response recording using infrared (IR) reflectance suitable for use in magnetic resonance imaging (MRI) environments; in this way, the neural activity of EBC could be investigated using fMRI in humans. Four participants underwent delay EBC and simultaneous fMRI. The results indicate (1) a high signal-to-noise ratio in the IR reflectance data that effectively quantifies the eyeblink morphology and timing and (2) evidence of conditioning in the fMRI environment. The quality of the data, the feasibility of conducting EBC experiments in the fMRI environment, and the customisability of the current system to fit a variety of EBC experimental design parameters are discussed.

Striatal D(2)/D(3) receptor availability is inversely correlated with cannabis consumption in chronic marijuana users.

BACKGROUND: Although the incidence of cannabis abuse/dependence in Americans is rising, the neurobiology of cannabis addiction is not well understood. Imaging studies have demonstrated deficits in striatal D(2)/D(3) receptor availability in several substance-dependent populations. However, this has not been studied in currently using chronic cannabis users. OBJECTIVE: The purpose of this study was to compare striatal D(2)/D(3) receptor availability between currently using chronic cannabis users and healthy controls. METHODS: Eighteen right-handed males age 18-34 were studied. Ten subjects were chronic cannabis users; eight were demographically matched controls. Subjects underwent a [(11)C]raclopride (RAC) PET scan. Striatal RAC binding potential (BP(ND)) was calculated on a voxel-wise basis. Prior to scanning, urine samples were obtained from cannabis users for quantification of urine Δ-9-tetrahydrocannabinol (THC) and THC metabolites (11-nor-Δ-9-THC-9-carboxylic acid; THC-COOH and 11-hydroxy-THC;OH-THC). RESULTS: There were no differences in D(2)/D(3) receptor availability between cannabis users and controls. Voxel-wise analyses revealed that RAC BP(ND) values were negatively associated with both urine levels of cannabis metabolites and self-report of recent cannabis consumption. CONCLUSIONS: In this sample, current cannabis use was not associated with deficits in striatal D(2)/D(3) receptor availability. There was an inverse relationship between chronic cannabis use and striatal RAC BP(ND). Additional studies are needed to identify the neurochemical consequences of chronic cannabis use on the dopamine system.

Examining the effects of former cannabis use on cerebellum-dependent eyeblink conditioning in humans.

RATIONALE: Previous work in humans has shown that chronic cannabis users exhibit disruptions in classical eyeblink conditioning (EBC), a form of associative learning that is known to be dependent on the cerebellum. Based upon previous work in animals, it was hypothesized that these learning deficits were related to cannabinoid receptor (CB1R) downregulation. However, it remains unclear whether there is a recovery of cerebellum-dependent learning after the cessation of cannabis use. METHODS: Therefore, former cannabis users (n=10), current cannabis users (n=10), and cannabis-naïve controls (n=10), all free of DSM-IV Axis-I or -II disorders, were evaluated. A standard delay EBC procedure was utilized in which paired presentations of a conditioned stimulus (CS; e.g., tone) and a co-terminating unconditioned stimulus (US; e.g., ocular airpuff) were administered, thus eliciting a conditioned eyeblink response (CR). The primary dependent measures were percentage of CRs and CR latency across conditioning blocks. RESULTS: Similar to prior studies, current cannabis users exhibited marked impairments in both the acquisition and timing of CRs compared to controls. Although former cannabis users showed intact CR acquisition compared to controls, they exhibited significantly impaired (shorter) CR latencies. In both cannabis groups, UR amplitude did not differ from controls, indicating normal US processing. CONCLUSIONS: These data suggest that a recovery of function has occurred for the learning of the CS-US association, while the accurate timing of the CR shows lasting impairments. Taken together, these results suggest that heavy cannabis use can disrupt timing-related synaptic plasticity within the cerebellum, even after the cessation of cannabis use.

The effect of chronic cannabinoids on broadband EEG neural oscillations in humans.

Animal and cellular work has shown that central cannabinoid-1 receptors modulate neural oscillations in the gamma range (40 Hz), which may be important for normal perceptual and cognitive processes. In order to assess the effect of cannabinoids on broadband-frequency neural oscillations in humans, the current study examined the effect of chronic cannabis use on auditory steady-state responses (ASSRs) utilizing electroencephalography (EEG). Passive ASSRs were assessed using varying rates of binaural stimulation (auditory click-trains; 10-50 Hz in increments of 5 Hz; 80 dB SPL) in carefully screened cannabis users and controls. Chronic cannabis users (n=22; 12 h abstinence before study; positive 11-nor-9-carboxy-delta-9-tetrahydrocannabinol urine levels) and cannabis naïve controls (n=24) were evaluated. Time X frequency analyses on EEG data were performed using Fourier-based mean trial power (MTP) and phase-locking (inter-trial coherence; ITC). Transient ERPs to stimulus onset (auditory N100 components) were also evaluated. As predicted, a decrease in spectral power (MTP) at 40 Hz was observed in the cannabis group (p<0.018). No effects on phase-locking (ITC) or the N100 were observed. Further, within the cannabis group, lower 40 Hz power correlated with an earlier age of onset of cannabis use (p<0.04). These data suggest that chronic exposure to exogenous cannabinoids can alter the ability to generate neural oscillations, particularly in the gamma range. This is consistent with preclinical animal and cellular data, which may have implications for understanding the short- and long-term psychopharmacological effects of cannabis.

Structural network topology revealed by white matter tractography in cannabis users: a graph theoretical analysis.

Endocannabinoid receptors modulate synaptic plasticity in the brain and may therefore impact cortical connectivity not only during development but also in response to substance abuse in later life. Such alterations may not be evident in volumetric measures utilized in brain imaging, but could affect the local and global organization of brain networks. To test this hypothesis, we used a novel computational approach to estimate network measures of structural brain connectivity derived from diffusion tensor imaging (DTI) and white matter tractography. Twelve adult cannabis (CB) users and 13 healthy subjects were evaluated using a graph theoretic analysis of both global and local brain network properties. Structural brain networks in both CB subjects and controls exhibited robust small-world network attributes in both groups. However, CB subjects showed significantly decreased global network efficiency and significantly increased clustering coefficients (degree to which nodes tend to cluster around individual nodes). CB subjects also exhibited altered patterns of local network organization in the cingulate region. Among all subjects, schizotypal and impulsive personality characteristics correlated with global efficiency but not with the clustering coefficient. Our data indicate that structural brain networks in CB subjects are less efficiently integrated and exhibit altered regional connectivity. These differences in network properties may reflect physiological processes secondary to substance abuse-induced synaptic plasticity, or differences in brain organization that increase vulnerability to substance use.

SUMO1 as a candidate gene for non-syndromic cleft lip with or without cleft palate: no evidence for the involvement of common or rare variants in Central European patients.

OBJECTIVE: Studies in mice and humans have suggested that SUMO1, which codes for the small ubiquitin-related modifier 1 (SUMO1), is a promising candidate gene for non-syndromic cleft lip with or without cleft palate (NSCL/P). To investigate the possible involvement of this gene in NSCL/P patients from Central Europe, we performed: (i) a case control association study, and (ii) a resequencing study. METHODS: Genotyping and the subsequent single marker and haplotype association analyses were performed for 413 NSCL/P patients and 412 controls. A total of 17 tagging single-nucleotide polymorphisms (SNPs) were used. In the resequencing study, the complete coding region and splice sites were sequenced in 65 index patients from multiply affected families. RESULTS: One of the 17 tested SNPs (rs16838917) had a borderline significant P-value of 0.0416 in the single-marker association analysis. However, this result did not withstand correction for multiple testing (P(corr)=0.707). No association was observed for any haplotypic marker combination. Sequencing failed to identify any novel rare sequence variants. CONCLUSIONS: The results of the present study do not support the hypothesis that common or rare variants in SUMO1 play a significant role in the development of NSCL/P in Central-European patients. However, smaller effects of common variants or the presence of rare high penetrance mutations in other non-investigated familial cases cannot be excluded. Further analysis of SUMO1 in independent samples from Central European and other populations is therefore warranted.

Investigation of the tryptophan hydroxylase 2 gene in bipolar I disorder in the Romanian population.

OBJECTIVE: Since the discovery of the tryptophan hydroxylase 2 gene (TPH2) several studies reported the association of TPH2 genetic variation with bipolar I (BPI) disorder. Our first objective was to replicate the recently described association of a rare functional single nucleotide polymorphism (SNP) (rs17110563) and of a haplotype covering the 5' region of TPH2 with BPI in a sample from the Romanian population. The second objective was to investigate the influence of the phenotypic traits 'age-of-onset' , 'family history', and 'parent-of-origin', defined according to clinical criteria, on the degree of association between TPH2 and BPI. METHOD: Sixteen TPH2 SNPs were genotyped in a Romanian sample of 198 BPI patients and 180 controls screened for psychiatric disorders. Statistical analysis of the data was performed with Haploview v.3.32 and FAMHAP. RESULTS: The functional SNP rs17110563 (encoding a Pro206Ser substitution) was present in one Romanian BPI patient and absent in controls. SNPs located in the 5'-region (rs11178997, rs11178998, rs7954758) that had earlier been found to be significantly associated with BPI in a German sample were not associated with BPI in the overall Romanian sample at the single-marker level, but gave evidence for association in a subgroup of patients with paternal transmission of the disease at the haplotypic level. Further evidence of association was identified between haplotypes located in the 3'-region of TPH2 and BPI in the overall sample as well as in the subgroups of familial cases, the patient group with paternal transmission, and the patient group with age of onset below or equal to 25 years. CONCLUSION: These data provide further support for the involvement of genetic variation in TPH2 in the etiology of BPI.