Establishment and clinical applications of a portable system for capturing influenza viruses released through coughing.

Coughing plays an important role in influenza transmission; however, there is insufficient information regarding the viral load in cough because of the lack of convenient and reliable collection methods. We developed a portable airborne particle-collection system to measure the viral load; it is equipped with an air sampler to draw air and pass it through a gelatin membrane filter connected to a cone-shaped, megaphone-like device to guide the cough airflow to the membrane. The membrane was dissolved in a medium, and the viral load was measured using quantitative real-time reverse transcriptase-polymerase chain reaction and a plaque assay. The approximate viral recovery rate of this system was 10% in simulation experiments to collect and quantify the viral particles aerosolized by a nebulizer. Using this system, cough samples were collected from 56 influenza A patients. The total viral detection rate was 41% (23/56), and the viral loads varied significantly (from <10, less than the detection limit, to 2240 viral gene copies/cough). Viable viruses were detected from 3 samples with ≤18 plaque forming units per cough sample. The virus detection rates were similar among different groups of patients infected with different viral subtypes and during different influenza seasons. Among patients who did not receive antiviral treatment, viruses were detected in one of six cases in the vaccinated group and four of six cases in the unvaccinated group. We found cases with high viral titers in throat swabs or oral secretions but very low or undetectable in coughs and vice versa suggesting other possible anatomical sites where the viruses might be mixed into the cough. Our system is easy to operate, appropriate for bedside use, and is useful for comparing the viral load in cough samples from influenza patients under various conditions and settings. However, further large-scale studies are warranted to validate our results.

Detection of respiratory viruses in nasopharyngeal secretions and middle ear fluid from children with acute otitis media.

CONCLUSIONS: Our results suggest that various respiratory viruses contribute to the pathogenesis of acute otitis media (AOM). OBJECTIVE: AOM is one of the most common complications of viral upper respiratory tract infections in children. Recently, the importance of respiratory viruses has been stressed as causative agents of AOM. SUBJECTS AND METHODS: A total of 1092 children < or =10 years old (average age 1.38 years) diagnosed as having AOM between 2002 and 2004 were studied. Bacterial and viral cultures of both nasopharyngeal secretions (NPS) and middle ear fluid (MEF) were performed for all 1092 children. Body temperature, changes of the tympanic membrane, and the number of days from the onset of illness were analyzed. RESULTS: Respiratory viruses were detected in 360 of 1092 NPS specimens, including 157 isolates of respiratory syncytial virus and 88 of influenza virus. Among 1092 MEF specimens, 102 were virus-positive, including 43 for respiratory syncytial virus and 29 for influenza virus. In 75 children, respiratory viruses were only detected in MEF. The viral detection rate was higher in children with fever at an early stage of their illness. The tympanic membrane changes associated with viral infection tended to be less severe, while changes were more severe in cases with bacterial infection, especially co-infection with bacteria and viruses.

Application of the Relapse Risk Scale to alcohol-dependent individuals in Japan: comparison with stimulant abusers.

OBJECTIVE: To develop and validate the Alcohol Relapse Risk Scale (ARRS) for Japanese alcohol-dependent individuals and to compare the features of relapse risk for alcohol-dependent individuals with those for stimulant abusers. METHODS: The ARRS is a multidimensional self-rating scale consisting of 32 items based on the Stimulant Relapse Risk Scale (SRRS). Two hundred eighteen inpatients and outpatients with a history of alcohol dependence (181 males and 36 females) were recruited, provided informed consent, and were administered the ARRS. The Visual Analog Scale (VAS) for alcohol craving, current state of drinking, and data on relapse within 1 month after the rating were used for validation. RESULTS: Exploratory factor analysis highlighted five factors: stimulus-induced vulnerability (SV), emotionality problems (EP), compulsivity for alcohol (CA), lack of negative expectancy for alcohol (NE), and positive expectancy for alcohol (PE). Cronbach's alpha coefficient for each of the subscales ranged from .55 to .90 and was .90 for the total ARRS, indicating their adequate internal consistency. SV, EP, CA, PE, and total ARRS were significantly correlated with the VAS and current drinking state, supporting their concurrent validity. SV and total ARRS were significantly correlated with relapse, suggesting that the ARRS is useful for predicting relapse risk in alcohol-dependent individuals, similar to the SRRS for stimulant abusers. Compared with stimulant abusers, alcohol-dependent individuals tended to express their desires related to relapse more honestly on the scales. CONCLUSIONS: The ARRS has multidimensional psychometric properties that are useful for assessing the various aspects of alcohol relapse risk.

Glutamate cysteine ligase modifier (GCLM) subunit gene is not associated with methamphetamine-use disorder or schizophrenia in the Japanese population.

A recent study showed a significant association between schizophrenia in European samples and the glutamate cysteine ligase modifier (GCLM) subunit gene, which is the key glutathione (GSH)-synthesizing enzyme. Since the symptoms of methamphetamine (METH)-induced psychosis are similar to those of schizophrenia, the GCLM gene is thought to be a good candidate gene for METH-use disorder or related disorders. To evaluate the association between the GCLM gene and METH-use disorder and schizophrenia, we conducted a case-control study of Japanese subjects (METH-use disorder, 185 cases; schizophrenia, 742 cases; and controls, 819). Four SNPs (2 SNPs from an original report and JSNP database, and 2 "tagging SNPs" from HapMap database) in the GCLM gene were examined in this association analysis; one SNP showed an association with both METH-use disorder and METH-induced psychosis. After Bonferroni's correction for multiple testing, however, this significance disappeared. No significant association was found with schizophrenia. Our findings suggest that a common genetic variation in the GCLM gene might not contribute to the risk of METH-use disorder and schizophrenia in the Japanese population.

Alpha4 and beta2 subunits of neuronal nicotinic acetylcholine receptor genes are not associated with methamphetamine-use disorder in the Japanese population.

The mesolimbic system is thought to be involved in the reinforcing action of many addictive drugs and the release of dopamine modulated by neuronal nicotine cholinergic receptors (nAChRs). Several investigations suggested that nAChRs on dopaminergic terminals play an important role in the development of some long-lasting adaptations associated with drug abuse. A majority of high-affinity nicotine binding sites in the brain have been showed in heteropentameric alpha4 (alpha4) and beta2 subunit (beta2) of nAChRs. Therefore, we conducted a genetic association analysis of the alpha4 gene (CHRNA4) and beta2 gene (CHRNB2) with methamphetamine (METH)-use disorder (191 cases and 753 controls). We first evaluated the linkage disequilibrium (LD) structure of these genes and selected 7 and 5 tagging SNPs (tag SNPs) on CHRNA4 and CHRNB2, respectively. Some tag SNPs were significantly associated with total METH-use disorder and METH-induced psychosis; however, these associations were no longer statistically significant after Bonferroni's correction for multiple testing. In conclusion, our results suggest that neither CHRNA4 nor CHRNB2 plays a major role in Japanese METH-use disorder.

Prostate apoptosis response 4 gene is not associated with methamphetamine-use disorder in the Japanese population.

Abnormal intracellular signaling molecules in dopamine signal transduction are thought to be associated with the pathophysiology of methamphetamine (METH)-use disorder. A recent study reported that a new intracellular protein, prostate apoptosis response 4 (Par-4), plays a critical role in dopamine 2 receptor signaling. We therefore analyzed the association between the Par-4 gene (PAWR) and METH-use disorder in a Japanese population (191 patients with METH-use disorder and 466 healthy controls). Using the recommended "gene-based" association analysis, we selected five tagging SNPs in PAWR from the HapMap database. No significant allele/genotype-wise or haplotype-wise association was found between PAWR and METH-use disorder. These results suggest that PAWR does not play a major role in METH-use disorders in the Japanese population.

Reduced CYP2D6 activity is a negative risk factor for methamphetamine dependence.

Because methamphetamine (METH) is metabolized by CYP2D6 at the first step of hydroxylation and demethylation, it is possible that functional variants of CYP2D6 alter susceptibility to methamphetamine-induced dependence. We genotyped CYP2D6*1, *4, *5, *10, and *14 for 202 patients with METH dependence and 337 controls in a Japanese population and found a significant association of the CYP2D6 gene with METH dependence (p=0.0299). The patients had fewer *10 and *14 alleles, which are hypofunction alleles, than the controls. CYP2D6 genotypes were divided into three phenotypes: extensive metabolizers, intermediate metabolizers, and poor metabolizers. There was no poor metabolizer among our Japanese subjects, and intermediate metabolizers of CYP2D6 were significantly fewer in methamphetamine-dependent subjects than in controls (p=0.0212), with an odds ratio of 0.62 (95% confidence interval: 0.51-0.76). The present study demonstrated that reduced CYP2D6 activity was a negative risk factor for methamphetamine dependence.

The glycine transporter 1 gene (GLYT1) is associated with methamphetamine-use disorder.

Glycine transporter (GlyT)-1 plays a pivotal role in maintaining the glycine level at the glutamatergic synapse. Glycine is an allosteric agonist of N-methyl-D-aspartate (NMDA) receptors. Because activation of NMDA receptors is an essential step for induction of methamphetamine dependence and psychosis, differences in the functioning of GlyT-1 due to genetic variants of the GlyT-1 gene (GLYT1) may influence susceptibility. A case-control genetic association study of the GLYT1 gene examined 204 patients with methamphetamine-use disorder and 210 healthy controls. We examined three single nucleotide polymorphisms (SNPs), SNP1, IVS3 + 411C > T, rs2486001; SNP2, 1056G > A, rs2248829; and SNP3, IVS11 + 22G > A, rs2248632, of the GLYT1 gene and found that SNP1 showed a significant association in both genotype (P = 0.0086) and allele (P = 0.0019) with methamphetamine-use disorder. The T-G haplotype at SNP1 and SNP2 was a significant risk factor for the disorder (P = 0.000039, odds ratio: 2.04). The present findings indicate that genetic variation of the GLYT1 gene may contribute to individual vulnerability to methamphetamine dependence and psychosis.

The dysbindin gene (DTNBP1) is associated with methamphetamine psychosis.

BACKGROUND: The dysbindin (DTNBP1 [dystrobrevin-binding protein 1]) gene has repeatedly been shown to be associated with schizophrenia across diverse populations. One study also showed that risk haplotypes were shared with a bipolar disorder subgroup with psychotic episodes, but not with all cases. DTNBP1 may confer susceptibility to psychotic symptoms in various psychiatric disorders besides schizophrenia. METHODS: Methamphetamine psychosis, the psychotic symptoms of which are close to those observed in schizophrenia, was investigated through a case (n = 197)-control (n = 243) association analyses of DTNBP1. RESULTS: DTNBP1 showed significant associations with methamphetamine psychosis at polymorphisms of P1635 (rs3213207, p = .00003) and SNPA (rs2619538, p = .049) and the three-locus haplotype of P1655 (rs2619539)-P1635-SNPA (permutation p = .0005). The C-A-A haplotype, which was identical to the protective haplotype previously reported for schizophrenia and psychotic bipolar disorders, was a protective factor (p = .0013, odds ratio [OR] = .62, 95% confidence interval [CI] .51-.77) for methamphetamine psychosis. The C-G-T haplotype was a risk for methamphetamine psychosis (p = .0012, OR = 14.9, 95% CI 3.5-64.2). CONCLUSIONS: Our genetic evidence suggests that DTNBP1 is involved in psychotic liability not only for schizophrenia but also for other psychotic disorders, including substance-induced psychosis.

Acute otitis media associated with cytomegalovirus infection in infants and children.

OBJECTIVE: Acute otitis media (AOM) is one of the most common complications of viral respiratory tract infections in children, but the role of each virus is still to be elucidated. We analyzed AOM associated with infection by cytomegalovirus (CMV), which is known as one of the major causes of viral respiratory tract infection. METHODS: Four hundred and ninety-five children (292 boys and 203 girls) diagnosed as having AOM in 2002 were studied. All of the children were under 6 years old, with the average age being 1.31+/-1.36 years. Bacterial and viral culture of both nasopharyngeal secretions (NPS) and middle ear fluid (MEF) was performed in all 495 children. The levels of glutamyl pyruvic transaminase (GPT) and the serum IgM antibody for CMV were measured. CMV infection was defined on the basis of isolation of this virus by culture and/or positive anti-CMV IgM antibody. NPS and MEF specimens of the subjects diagnosed as having CMV infection were tested for the virus by nested PCR. RESULTS: Twelve of the 495 children were found to have CMV infection. They included 6 boys and 6 girls aged from 3 to 25 months, with the average age being 11+/-7 months. Among 10 children in whom CMV infection was diagnosed by viral culture, CMV was isolated from NPS alone in nine cases and from both NPS and MEF in one case. Nested PCR was performed in all 12 subjects diagnosed as having CMV infection, and all NPS samples were positive, as were 8 MEF samples. We obtained serum samples from 205 children under 2 years of age, including 9 with CMV infection. The mean serum GPT level of 124 children in whom no viruses were detected was 20.7+/-14.4 IU/L. While, the serum GPT levels of 9 children with CMV infection ranged from 10 to 280 IU/L with the average titer being 78.4+/-81.9 IU/L, and the GPT levels of the children with CMV infection were significantly higher than those of the children in whom no viruses were detected (p<0.05). CONCLUSION: Our results suggested that CMV is a causative pathogen of AOM, and that CMV infection should be suspected in patients with AOM and liver dysfunction.

Development and validation of the Stimulant Relapse Risk Scale for drug abusers in Japan.

OBJECTIVE: To develop and validate a multidimensional measure of relapse risk for stimulants in Japanese drug abusers. METHODS: A Stimulant Relapse Risk Scale (SRRS) was developed based on the Marijuana Craving Questionnaire and a discussion among three psychiatrists. We created 48 items after confirming the items including a variety of relapse risk, such as craving (expectancy, compulsivity, etc.) and emotionality problems. One hundred inpatients and outpatients with a history of stimulant abuse (71 males and 29 females) were recruited with informed consent, and were administered the SRRS. The Visual Analogue Scale for drug craving (VAS), Addiction Severity Index for Japanese (ASI-J), and data on relapse within 3 and 6 months after the rating were used for the validation. RESULTS: Exploratory factor analysis highlighted five factors: anxiety and intention to use drug (AI), emotionality problems (EP), compulsivity for drug use (CD), positive expectancies and lack of control over drug (PL), and lack of negative expectancy for drug use (NE). These accounted for 48.3% of the total variance. Thirty of the 43 items were classified into the five subscales. Cronbach's alpha coefficient for each subscale ranged from .55 to .82, and was .86 for the total SRRS, indicating their adequate internal consistency. AI, CD, PL, and total SRRS were significantly correlated with the drug-use composite score of the ASI-J, supporting their concurrent validity. AI, PL, NE, and total SRRS were significantly correlated with relapse, implying their predictive validity. CONCLUSIONS: The SRRS has multidimensional psychometric properties useful for assessing the various aspects of stimulant relapse risk.

Reliability and validity of the Japanese version of the Addiction Severity Index (ASI-J).

The Addiction Severity Index (ASI) is a frequently used clinical and research instrument that collects data from substance abusers in seven problem areas: medical, employment, alcohol, drug use, legal, family-social functioning, and psychiatric status. In each area, the ASI provides a composite score and severity rating that estimate the seriousness of the problem and the client's need for treatment. In the present study, we investigated the reliability and validity of the Japanese version of the ASI (ASI-J). One hundred and eleven subjects with a history of drug abuse were interviewed with a test battery including the ASI with informed consent. This revealed that: (a) the problem areas were independent of each other, underscoring the need for multidimensional assessment, (b) the inter-rater correlation of severity ratings in each area ranged from 0.68 to 0.99, and Cronbach's alpha coefficient for the items used for the composite score in each area ranged from 0.57 to 0.86, indicating their reliability with the exception of the drug and employment areas, and (c) several composite scores were significantly correlated with the drug craving levels assessed by a visual analogue scale, the abstinence period, mental health, and/or relapse, supporting their concurrent and predictive validity. These results suggest that the ASI-J has acceptable reliability and validity.

[Treatment of substance dependence by a bio-cognitive model based on behavioral pharmacology].

We have introduced cognitive behavior therapy (CBT) into the treatment of substance dependence patients, which involves disease education and focused group therapy to obtain insight into the taking behavior and to establish concrete countermeasures to prevent relapse. We have created a bio-cognitive model based on biological aspects to explain the pathology of substance dependence. 'Dependence' is a term in behavioral pharmacology defined as reinforced drug seeking and taking behavior. Changes in taking behavior are thought to occur due to the repetition of the reinforcement action of psychoactive substances in the reward system of the brain. Therefore, when intake desire is strong, it is hard for patients to control themselves, and there is a feature of difficulties considering the process of thinking in CBT. In other words, when craving becomes strong, a chain of behavior happens spontaneously, without schema, involving automatic thoughts. We think that the improvement of protracted withdrawal syndrome (PWS) and entire frontal lobe function are important in learning to discern distortion of cognition. When PWS is improved, a conflict is easy to bring about in the process of drug seeking and taking behavior. And, it is easy to execute avoidance plans (coping skills) which are established to cope with craving in advance. We think that a goal for treatment is to discern drug seeking and taking behavior with natural emotion. The recovery of PWS and frontal lobe dysfunction takes a long time with a serious dependence, so we must perform repetition of CBT. As the treatment introduction of involuntary admission cases is adequate or cases of 1 to 3 months of admission treatment based on voluntary admission are hard to treat, treatment to obtain insights into patients while carrying out repeated CBT using a bio-cognitive model and to improve PWS could be a possibility as one treatment for the pathology of diversified substance dependence.

A missense variation in human casein kinase I epsilon gene that induces functional alteration and shows an inverse association with circadian rhythm sleep disorders.

Recent studies have shown that functional variations in clock genes, which generate circadian rhythms through interactive positive/negative feedback loops, contribute to the development of circadian rhythm sleep disorders in humans. Another potential candidate for rhythm disorder susceptibility is casein kinase I epsilon (CKIepsilon), which phosphorylates clock proteins and plays a pivotal role in the circadian clock. To determine whether variations in CKIepsilon induce vulnerability to human circadian rhythm sleep disorders, such as delayed sleep phase syndrome (DSPS) and non-24-h sleep-wake syndrome (N-24), we analyzed all of the coding exons of the human CKIepsilon gene. One of the variants identified encoded an amino-acid substitution S408N, eliminating one of the putative autophosphorylation sites in the carboxyl-terminal extension of CKIepsilon. The N408 allele was less common in both DSPS (p = 0.028) and N-24 patients (p = 0.035) compared to controls. When DSPS and N-24 subjects were combined, based on an a priori prediction of a common mechanism underlying both DSPS and N-24, the inverse association between the N408 allele and rhythm disorders was highly significant (p = 0.0067, odds ratio = 0.42, 95% confidence interval: 0.22-0.79). In vitro kinase assay revealed that CKIepsilon with the S408N variation was approximately 1.8-fold more active than wild-type CKIepsilon. These results indicate that the N408 allele in CKIepsilon plays a protective role in the development of DSPS and N-24 through alteration of the enzyme activity.

Deactivation by benzodiazepine of the basal forebrain and amygdala in normal humans during sleep: a placebo-controlled [15O]H2O PET study.

OBJECTIVE: The authors' goal was to identify differences in regional brain activity between physiological and benzodiazepine-induced sleep to clarify the brain structures involved in the drug's hypnotic effect. METHOD: Using positron emission tomography, they compared regional cerebral blood flow during non-REM sleep in nine volunteers treated with placebo or triazolam, a short-acting benzodiazepine, in a double-blind, crossover design. RESULTS: Blood flow in the basal forebrain and amygdaloid complexes was lower during non-REM sleep when subjects were given triazolam than when they were given placebo. CONCLUSIONS: The hypnotic effect of the benzodiazepines may be mediated mainly by deactivation of the forebrain control system for wakefulness and also by the anxiolytic effect induced by deactivation of the emotional center.

Sleep and circadian rhythm disturbances in patients with delayed sleep phase syndrome.

STUDY OBJECTIVES: The objective of this study was to clarify sleep characteristics and pathophysiology in patients with delayed sleep phase syndrome (DSPS), which is a major circadian rhythm sleep disorder subtype. DESIGN: Polysomnography was performed for 2 consecutive nights and core body temperature was sampled for 7 consecutive days, including the polysomnography study period, in all subjects. Findings were compared and statistically analyzed between patients with DSPS and matched controls. SETTING: Sleep disorders unit in National Center Hospital. PARTICIPANTS: 11 DSPS patients and 11 age-matched healthy volunteers. INTERVENTIONS: N/A. MEASUREMENTS AND RESULTS: Sleep latency, total sleep time, wakefulness after sleep-onset, and the amount and percentage of Stage 1 sleep were greater in DSPS patients than in volunteers. Sleep efficiency and the amount and percentage of slow wave sleep were lower in DSPS patients than in volunteers. Compared with the healthy volunteers, DSPS patients showed a decreased number and different temporal distribution of high-voltage and low-frequency delta waves. The time of minimum body temperature appeared earlier in the sleep phase for the patients than for the volunteers. Significant correlation was found between the amount of slow wave sleep and the time from sleep onset to minimum body temperature and between the amount and percentage of slow wave sleep and time from minimum body temperature to sleep offset. CONCLUSIONS: Disturbances were found in the sleep structure of patients with DSPS, and these disturbances were related to the discrepancy between patients and controls in the phase relationship difference between sleep and core body temperature rhythms.