Heavy oil exposure suppresses antiviral activities in Japanese flounder Paralichthys olivaceus infected with viral hemorrhagic septicemia virus (VHSV).

A combined treatment of heavy oil (HO) exposure and virus infection induces increased mortality in Japanese flounder (Paralichthys olivaceus). In this study, we addressed how HO exposure affects the immune system, especially antiviral activities, in Japanese flounder. The fish were infected with viral hemorrhagic septicemia virus (VHSV), followed by exposure to HO. We analyzed virus titers in the heart and mRNA expression in the kidney of surviving fish. The virus titers in fish exposed to heavy oil were higher than the threshold for onset. The results suggest that HO exposure may allow the replication of VHSV, leading to higher mortality in the co-treated group. Gene-expression profiling demonstrated that the expression of antiviral-activity-related genes, such as those for interferon and apoptosis induction, were lower in the co-treated group than in the group with VHSV infection only. These results helped explain the high virus titers in fish treated with both stressors. Thus, interferon production in the virus-infected cells and apoptosis induction by natural killer cells worked normally in the VHSV-infected fish without HO exposure, but these antiviral activities were slightly suppressed by HO exposure, possibly leading to extensive viral replication in the host cells and the occurrence of VHS.

Sex-specific immunomodulatory action of the environmentalestrogen 17α-ethynylestradiol alongside with reproductive impairment in fish.

Estrogenic endocrine disrupting chemicals (EEDCs) are present ubiquitously in sediments and aquatic ecosystems worldwide. The detrimental impact of EEDCs on the reproduction of wildlife is widely recognized. Increasing evidence shows the immunosuppressive effects of EEDCs in vertebrates. Yet, no studies have considered concomitantly EEDC-induced impacts on reproductive impairment and immune suppression in vivo, which are deemed essential for risk assessment and environmental monitoring. In this study, EE2 was used as a representative EEDC, for parallel evaluation of EEDC-induced immune suppression (immune marker gene expression, leukocyte numbers, host resistance assay, and immune competence index) and reproductive impairment (estrogen responsive gene expression, fecundity, fertilization success, hatching success, and reproductive competence index) in an established fish model (marine medaka Oryzias melastigma), considering sex-specific induction and adaptation and recovery responses under different EE2 exposure scenarios. The findings in marine medaka reveal distinct sex differences in the EE2-mediated biological responses. For female fish, low concentration of exogenous EE2 (33 ng/L) could induce hormesis (immune enhancement), enable adaptation (restored reproduction) and even boost fish resistance to bacterial challenge after abatement of EE2. However, a prolonged exposure to high levels of EE2 (113 ng/L) not only impaired F0 immune function, but also perturbed females recovering from reproductive impairment, resulting in a persistent impact on the F1 generation output. Thus, for female fish, the exposure concentration of EE2 is more critical than the dose of EE2 in determining the impacts of EE2 on immune function and reproduction. Conversely, male fish are far more sensitive than females to the presence of low levels of exogenous EE2 in water and the EE2-mediated biological impacts are clearly dose-dependent. It is also evident in male fish that direct contact of EE2 is essential to sustain impairments of immune competence and reproductive output as well as deregulation of immune function genes in vivo. The immunomodulatory pathways altered by EE2 were deciphered for male and female fish, separately. Downregulation of hepatic tlr3 and c3 (in female) and tlr3, tlr5 and c3 (in male) may be indicative of impaired fish immune competence. Taken together, impaired immune competence in the EE2-exposed fish poses an immediate thread on the survival of F0 population. Impaired reproduction in the EE2-exposed fish can directly affect F1 output. Parallel evaluation of immune competence and reproduction are important considerations when assessing the risk of sublethal levels of EE2/EEDCs in aquatic environments.

Analysis of genes encoding high-antigenicity polypeptides in three serotypes of Miamiensis avidus.

The ciliate Miamiensis avidus causes scuticociliatosis in Japanese flounder Paralichthys olivaceus. We previously reported three serotypes of this ciliate distinguishable by serotype-specific antigenic polypeptides (serotype I, 30kDa; serotype II, 38kDa; serotype III, 34kDa). In this study, we determined the localization site of the serotype-specific polypeptides in the ciliate and determined the genes encoding the polypeptides, using the isolates IyoI (serotype I), Nakajima (serotype II), and Mie0301 (serotype III). SDS-PAGE and immunoblot analysis of cilia, membrane proteins, and cytoskeletal elements of the ciliates revealed that the polypeptides were abundant in the former two. Scanning electron microscopy of ciliates immobilized by homologous antiserum showed morphological changes in the cilia. These evidences suggested that the polypeptides were ciliary membrane immobilization antigens. The ciliary genes identified showed low identity scores-<51.5% between serotypes. To differentiate the serotypes, we designed serotype-specific PCR primer sets based on the DNA sequences. The PCR-based serotyping results were completely consistent with conventional serotyping methods (immobilization assay and immunoblot analysis). Twenty of 21 isolates were classified as either serotype I or II, and one isolate was undistinguishable. The combination of species-specific PCR previously reported and three serotype-specific PCR could be useful for identifying, serotyping, and surveillance for occurrences of new serotypes of M. avidus.

Adenosine A1 receptors measured with 11 C-MPDX PET in early Parkinson's disease.

Adenosine A1 receptors (A1 Rs) interact negatively with dopamine D1 receptors (D1 Rs) in neurons of the basal ganglia's direct pathway, while adenosine A2A receptors (A2A Rs) negatively interact with dopamine D2 receptors (D2 Rs) in indirect-pathway neurons. The aim of this study was to investigate the cerebral density of A1 Rs in Parkinson's disease (PD) in its early stages, using PET scans with the radioligand 8-dicyclopropylmethyl-1-11 C-methyl-3-propylxanthine (11 C-MPDX). We studied 10 drug-naïve patients with early PD. Each patient was also examined for dopamine transporters (DATs) and D2 Rs by PET using 11 C-2-ß-carbomethoxy-3-ß-(4-fluorophenyl)-tropane (11 C-CFT) and 11 C-raclopride (11 C-RAC), respectively. Ten elderly, healthy volunteers were recruited as controls for 11 C-MPDX PET scanning and eight elderly volunteers were recruited as controls for 11 C-CFT and 11 C-RAC PET scanning. The PET scans revealed a decrease in the uptake ratio index (URI) of 11 C-CFT and an increase in the URI of 11 C-RAC in patients. In the temporal lobe, the binding potential for 11 C-MPDX was higher in the patient group than in healthy subjects, but not in the other regions examined, including the striatum. In patients, we observed motor-symptom asymmetry and a relationship between parkinsonism and the striatal density of DATs, but not A1 R density. In the putamen of early PD, asymmetrical down-regulation of A2A Rs is likely a compensatory mechanism in response to a decrease in dopamine. However, our study suggests that A1 Rs are unaltered in the putamen of early PD.

Encystment and excystment of kinetoplastid Azumiobodo hoyamushi, causal agent of soft tunic syndrome in ascidian aquaculture.

Soft tunic syndrome in the edible ascidian Halocynthia roretzi is caused by the kinetoplastid flagellate Azumiobodo hoyamushi, which was found to assume a fusiform cell form with 2 flagella in axenic, pure culture. When the flagellate form was incubated in sterilized artificial seawater (pH 8.4), some of the cells became cyst-like and adhered to the bottom of the culture plate. The cyst-like forms were spherical or cuboidal, and each had 2 flagella encapsulated in its cytoplasm. Encystment was also induced in culture medium alkalified to the pH of seawater (8.4) but not in unmodified (pH 7.2) or acidified media (pH 6.4). More than 95% of the cyst-like cells converted to the flagellate form within 1 d following transfer to seawater containing ascidian tunic extracts from host ascidians. The cyst-like cells were able to survive in seawater with no added nutrients for up to 2 wk at 20°C and for a few months at 5 to 15°C. The survival period in seawater depended on temperature: some cyst-like cells survived 3 mo at 10°C, and ca. 95% of these converted to flagellate forms in seawater containing tunic extracts. Thus, A. hoyamushi is able to persist under adverse conditions in a cyst-like form able to adhere to organic and inorganic substrata for protracted periods of time.

Simple solution for preventing cerebrospinal fluid loss and brain shift during multitrack deep brain stimulation surgery in the semisupine position: polyethylene glycol hydrogel dural sealant capping: rapid communication.

This study evaluated preliminary findings on the efficacy of polyethylene glycol (PEG) hydrogel dural sealant capping for the prevention of cerebrospinal fluid (CSF) leakage and pneumocephalus during deep brain stimulation (DBS) surgery in the semisupine position. Group A consisted of 5 patients who underwent bilateral subthalamic nucleus (STN)-DBS surgery without PEG hydrogel dural sealant capping. Group B consisted of 5 patients who underwent bilateral STN-DBS surgery with PEG hydrogel dural sealant capping. The immediate postoperative intracranial air volume was measured in all patients and compared between the 2 groups using the Welch test. Adverse effects were also examined in both groups. The intracranial air volume in Group A was 32.3 ± 12.3 ml (range 19.1-42.5 ml), whereas that in Group B was 1.3 ± 1.5 ml (range 0.0-3.5 ml), showing a significant difference (p < 0.005). No hemorrhage or venous air embolisms were observed in either group. The effect of brain shift was discriminated by STN recordings in Group B. These preliminary findings indicate that PEG hydrogel dural sealant capping may reduce adverse effects related to CSF leakage and brain shift during DBS surgery.

Effect of heavy oil exposure on antibacterial activity and expression of immune-related genes in Japanese flounder Paralichthys olivaceus.

Heavy oil (HO) pollution is one of the most important environmental issues globally. However, little is known about the immunotoxicity of HO in fish. We therefore investigated the effects of HO exposure on immunocompetence and expression of immune-related genes in Japanese flounder, Paralichthys olivaceus. To test immunocompetency, serum collected from the fish was mixed with Edwardsiella tarda, plated, and the resultant numbers of bacterial colonies were counted. Plates with serum from HO-exposed fish (5 d postexposure [dpe]) had significantly higher numbers of colonies than those of the untreated control group, suggesting that HO exposure suppresses immunocompetency. Downregulation of the immunoglobulin light chain (IgM) gene in HO-exposed fish at 5 dpe was detected by real-time polymerase chain reaction. These results suggest that IgM-mediated immunity is suppressed by HO exposure. We measured polycyclic aromatic hydrocarbon (PAH) concentrations in the liver of the fish. Low molecular weight PAHs were found to be taken up at high concentrations in fish liver; therefore, they are likely the cause of immune suppression in the fish.

A time-course study of immune response in Japanese flounder Paralichthys olivaceus exposed to heavy oil.

PURPOSE: The immunotoxicities of oil and its components on fish immunities have been investigated, but there is little literature on the recovery of the fish from the immune suppression. Therefore, the recovery of Japanese flounder Paralichthys olivaceus from an immunosuppressive effect due to heavy oil (HO) exposure was investigated in this study. METHODS: Fish were exposed to HO at a concentration of 0.385 g/L for 2 days, while control fish received no exposure. Seven fish were sampled at 0, 3, 7, and 14 days post-exposure. The respiratory rate was measured everyday as an indicator of the acute effect of HO exposure. Fish serum was collected and used for antibacterial activity assay against Edwardsiella tarda. Expression changes of respiratory and immune-related genes were evaluated by real-time PCR. RESULTS AND DISCUSSION: The respiratory rate was significantly increased in the HO-exposed group until 4 days post-exposure. A respiratory-related gene, ß-hemoglobin, was also significantly downregulated in the spleen both at 0 and 7 days post-exposure and kidney at 3 days post-exposure in HO-exposed fish. Immunotoxicity, including suppression of antibacterial activities and downregulation of the IgM gene, was observed in HO-exposed fish until 3 days post-exposure, but not after that time. From these results, we conclude that the fish likely return to normal status around 1 week.

Heavy oil fraction induces the dysplastic sperm in male mouse.

Heavy oil is one of the most serious pollutants in marine ecosystem. The poisonous influences of the chemical substances contained in heavy oil on many kinds of marine organisms are widely studied. However, the influence of the chemical compounds in heavy oil on our health has not been cleared yet. In order to reveal the poisonous influences of these chemical compounds on mammalian reproductive system, water-soluble fraction (WSF) extracted from heavy oil was administrated to mice for 2 weeks. WSF-administrated mice were crossed with either WSF- or distilled water-administrated group for mating experiment. When WSF-administrated male mice were used as a father, it reduced not only mating ratio, but also neonatal male ratio. The numbers of sperms of WSF-administrated male mice were decreased. In addition, abnormality of sperms such as bent or twisted tail was increased approximately 6-fold by WSF intake. The level of testosterone in serum from WSF-administrated mice was lower than that from control mice. Testosterone is the most important for the spermatogenesis in vertebrate. It is supposed from these findings, the decrease in the number of sperms may relate with the reduction of sex hormone level in serum. It is suggested from these results that the chemical substances in WSF affected the sperm function in reproductive system of male mice.

Effect of heavy oil on the development of the nervous system of floating and sinking teleost eggs.

Heavy oil (HO) on the sea surface penetrates into fish eggs and prevents the normal morphogenesis. To identify the toxicological effects of HO in the context of the egg types, we performed exposure experiments using floating eggs and sinking eggs. In the course of development, HO-exposed embryos of floating eggs showed abnormal morphology, whereas early larva of the sinking eggs had almost normal morphology. However, the developing peripheral nervous system of sinking eggs showed abnormal projections. These findings suggest that HO exposed fishes have problems in the developing neurons, although they have no morphological malformations. Through these observations, we conclude that HO is strongly toxic to floating eggs in the morphogenesis, and also affect the neuron development in both floating and sinking eggs.

Adenosine A(2A) receptors measured with [C]TMSX PET in the striata of Parkinson's disease patients.

Adenosine A(2A) receptors (A2ARs) are thought to interact negatively with the dopamine D(2) receptor (D2R), so selective A2AR antagonists have attracted attention as novel treatments for Parkinson's disease (PD). However, no information about the receptor in living patients with PD is available. The purpose of this study was to investigate the relationship between A2ARs and the dopaminergic system in the striata of drug-naïve PD patients and PD patients with dyskinesia, and alteration of these receptors after antiparkinsonian therapy. We measured binding ability of striatal A2ARs using positron emission tomography (PET) with [7-methyl-(11)C]-(E)-8-(3,4,5-trimethoxystyryl)-1,3,7-trimethylxanthine ([(11)C]TMSX) in nine drug-naïve patients with PD, seven PD patients with mild dyskinesia and six elderly control subjects using PET. The patients and eight normal control subjects were also examined for binding ability of dopamine transporters and D2Rs. Seven of the drug-naïve patients underwent a second series of PET scans following therapy. We found that the distribution volume ratio of A2ARs in the putamen were larger in the dyskinesic patients than in the control subjects (p<0.05, Tukey-Kramer post hoc test). In the drug-naïve patients, the binding ability of the A2ARs in the putamen, but not in the head of caudate nucleus, was significantly lower on the more affected side than on the less affected side (p<0.05, paired t-test). In addition, the A2ARs were significantly increased after antiparkinsonian therapy in the bilateral putamen of the drug-naïve patients (p<0.05, paired t-test) but not in the bilateral head of caudate nucleus. Our study demonstrated that the A2ARs in the putamen were increased in the PD patients with dyskinesia, and also suggest that the A2ARs in the putamen compensate for the asymmetrical decrease of dopamine in drug-naïve PD patients and that antiparkinsonian therapy increases the A2ARs in the putamen. The A2ARs may play an important role in regulation of parkinsonism in PD.

Disruption of Sema3A expression causes abnormal neural projection in heavy oil exposed Japanese flounder larvae.

It has been well known that oil spills cause serious problems in the aquatic organisms. In particular, some species of teleosts, which develop on the sea surface thought to be affected by heavy oil (HO). During the embryogenesis, the nervous system is constructed. Therefore, it is important to study the toxicological effects of HO on the developing neurons. We exposed HO to eggs of Japanese flounder (Paralichthys olivaceus) and investigated the neural disorder. In larvae exposed by HO at the concentration of 8.75 mg/L, the facial and lateral line nerves partially entered into the incorrect region and the bundle was defasciculated. Furthermore, in the HO-exposed larvae, Sema3A, a kind of axon guidance molecule, was broadly expressed in second pharyngeal arch, a target region of facial nerve. Taken together, we suggested the possibility that the abnormal expression of Sema3A affected by HO exposure causes disruption of facial nerve scaffolding.

Heavy oil exposure induces high moralities in virus carrier Japanese flounder Paralichthys olivaceus.

The relationship between chemical exposure and disease outbreak in fish has not been fully defined due to the limitations of experimental systems (model fish and pathogens). Therefore, we constructed a system using the Japanese flounder, Paralichthys olivaceus, and viral haemorrhagic septicemia virus (VHSV), and evaluated it by heavy oil (HO) exposure. The fish were exposed to HO at 0.3, 0.03, 0.003, and 0 g/L following VHSV infection at doses of 10(2.5) or 10(3.5) tissue culture infectious dose (TCID)50/fish. As a result, groups given the dual stressors showed more than 90% mortality. Although VHSV infection at 10(2.5) and 10(3.5) TCID50/fish without HO exposure also induced high mortality, at 68.8% and 81.3%, respectively, HO exposure induced faster and higher mortality in the virus carrier fish, indicating that chemical stressors raise the risk of disease outbreak in fish. The experimental system established in this study could be useful for chemical risk assessment.

Nervous system disruption and concomitant behavioral abnormality in early hatched pufferfish larvae exposed to heavy oil.

BACKGROUND AND OBJECTIVE: Spills of heavy oil (HO) over the oceans have been proven to have an adverse effect on marine life. It has been hypothesized that exposure of early larvae of sinking eggs to HO leads largely to normal morphology, whereas abnormal organization of the developing neural scaffold is likely to be found. HO-induced disruption of the nervous system, which controls animal behavior, may in turn cause abnormalities in the swimming behavior of hatched larvae. To clarify the toxicological effects of HO, we performed exposure experiments and morphological and behavioral analyses in pufferfish (Takifugu rubripes) larvae. EXPERIMENTAL PROCEDURE: Fertilized eggs of pufferfish were exposed to 50 mg/L of HO for 8 days and transferred to fresh seawater before hatching. The hatched larvae were observed for their swimming behavior, morphological appearance, and construction of muscles and nervous system. RESULTS AND DISCUSSION: In HO-exposed larvae, we did not detect any anomaly of body morphology. However, they showed an abnormal swimming pattern and disorganized midbrain, a higher center controlling movement. Our results suggest that HO-exposed fishes suffer developmental disorder of the brain that triggers an abnormal swimming behavior and that HO may be selectively toxic to the brain and cause physical disability throughout the life span of these fishes.

Severe abnormalities in the reproductive organs of mice caused by chemical substances contained in heavy oil.

It is well known that heavy oil pollution results in various negative impacts on the marine environment. Although there is a low possibility of direct exposure to heavy oil, the chemical substances contained in heavy oil may be released into the environment and accumulated by marine organisms which in turn can be taken by humans via the food chain. In this study, we examined the biological risk of heavy oil extract using the common mouse, whose genetic backgrounds and immune system are well known and relatively homologous to humans. Water-soluble fraction (WSF) was extracted from heavy oil with water and the extract orally administrated to female or male mice for 7 days. In the WSF administrated group, cystoma-like formation was observed in the ovary in approximately 80% of female mice. On the other hand, we found that the prostate gland size in male mice was markedly reduced in comparison with male control mice. Continuous administration of WSF for 28 days resulted in continued hypertrophy of the cystoma around the ovary and atrophy in the prostate gland. In addition, it was revealed that chemical substances within WSF have estrogenic activity. A major component of heavy oil, polycyclic aromatic hydrocarbons (PAHs), is known to present estrogenic activity. It is likely that the cystoma-like formation in female mice and atrophy of prostate gland in male resulted of estrogenic substances present in the WSF which might be the PAHs.

Toxicogenomic analysis of immune system-related genes in Japanese flounder (Paralichthys olivaceus) exposed to heavy oil.

Heavy oil contamination is one of the most important environmental issues. Toxicities of polycyclic aromatic hydrocarbons (PAHs), including immune toxicities, are well characterized, however, the immune toxic effects of heavy oil, as a complex mixture of PAHs, have not been investigated. In the present study, we selected Japanese flounder (Paralichthys olivaceus) as a model organism, and observed alteration of immune function by the exposure to heavy oil. To analyze the expression profiles of immune system-related genes, we selected 309 cDNAs from our flounder EST library, and spotted them on a glass slide. Using this cDNA array, alteration of gene expression profiles was analyzed in the kidneys of flounders exposed to heavy oil. Six Japanese flounders (mean body weight: 197 g) were acclimated to laboratory conditions at 19-20 degrees C. Three fish were exposed to heavy oil C (bunker C) at a concentration of 3.8 g/L for 3 days, and the others were kept in seawater without heavy oil and used as the control. After the exposure period, the fish were transferred into control seawater and maintained for 4 days, and then they were dissected and their kidneys were removed. Total RNA was extracted from the kidney samples to use in gene expression analyses. The microarray detected alteration of immune system-related genes in the kidneys of heavy oil-exposed flounders, including down-regulation of immunoglobulin light chain, CD45, major histocompatibility complex class II antigens and macrophage colony-stimulating factor precursor, and up-regulation of interleukin-8 and lysozyme. These results suggest that pathogen resistance may be weakened in heavy oil-exposed fish, causing a subsequent bacterial infection, and then proinflammatory genes may be induced as a defensive response against the infection. Additionally, we found candidate genes for use as biomarkers of heavy oil exposure, such as N-myc downstream regulated gene 1 and heat shock cognate 71 kDa proteins.

Effects of heavy oil in the developing spotted halibut, Verasper variegatus.

It is well known that heavy oil (HO) on the sea surface causes serious problems in the aquatic environment. In particular, some species of teleosts which develop on the sea surface are thought to be affected by the HO which flows out from tankers or coastal industry. However, the toxicological effects of HO are not fully understood. We performed exposure experiments using the Pleuronectiformean fish, spotted halibut (Verasper variegatus), which is an important fishery resource in Japan. In course of the development, HO-exposed embryos showed remarkable delay in developmental processes including somite formation. We further observed abnormal development of the head morphology. Notably, treated embryos had relatively small eyes and craniofacial structures. These findings strongly suggest that HO seriously affects the cell proliferation and differentiation of the embryo. In addition, HO-exposed embryos showed abnormal neuronal development. We also performed the exposure in the larval stage. Treatment of post-hatching larvae with HO resulted in significantly greater mortality compared with controls. Through these observations, we finally conclude that HO is strongly toxic to halibut in their early life stages.

Does heavy oil pollution induce bacterial diseases in Japanese flounder Paralichthys olivaceus?

As basic research for the effect of heavy oil on the fish immune system, in this study, the number of leukocyte was counted in Japanese flounder Paralichthys olivaceus, after exposure to heavy oil at a concentration of 30 g/8L for 3 days. To compare the numbers of bacteria in the skin mucus between oil-exposed and control fish, viable bacteria were enumerated by counting colony forming unit (CFU). Compared with 5.79+/-1.88 x 10(7)leukocytes/mL in the controls, the exposed fish demonstrated higher counts, averaging 1.45+/-0.45 x 10(8)cells/mL. The bacterial numbers of control fish were 4.27+/-3.68 x 10(4)CFU/g, whereas they were 4.58+/-1.63 x 10(5)CFU/g in the exposed fish. The results suggest that immune suppression of the fish occurred due to heavy oil stressor, and bacteria could invade in the mucus, resulting in the increasing leukocyte number to prevent infectious disease.

Validation of the 7-Minute Screen for the detection of early-stage Alzheimer's disease.

BACKGROUND/AIMS: The 7-Minute Screen (7MS) is a screening battery to identify individuals with a high probability of Alzheimer's disease (AD). The 7MS consists of four subtests (Temporal Orientation, Enhanced Cued Recall, Clock Drawing, and Verbal Fluency), each measuring a different aspect of cognition. The present study is designed to examine the predictive validity of the 7MS to distinguish between patients with early-stage AD and healthy control subjects. METHODS: Sixty-three patients who were diagnosed as having probable or possible AD and 91 community-dwelling elderly individuals of comparable age, sex distribution, and education were administered the 7MS (paper-and-pencil version) and other screening batteries. All patients were rated Clinical Dementia Rating (CDR) 0.5-1, categorized as early-stage AD, and all participants obtained a score of less than 6 for the Geriatric Depression Scale (15-item version). RESULTS: Mean scores for patients and controls on all the four subtests were significantly different. When using the total score of the 7MS, which was calculated by the original logistic regression formula based on all of the four subtests, the sensitivity for early-stage AD was 90.5% with a specificity of 92.3%. Correlation analysis indicated high concurrent validity between the 7MS and existing standard cognitive screening batteries (e.g., MMSE, HDS-R). In correlation analysis and multiple regression analysis, demographic effects (age, sex, and education) were not significantly associated with the total score of the 7MS in controls. CONCLUSIONS: The results showed that the 7MS had a high level of sensitivity and specificity. We also found that the 7MS was not affected by demographic characteristics. These findings demonstrated that the 7MS is a useful screening tool for discriminating patients with early-stage AD from intact individuals.