Relationship between the expression level of miRNA-4485 and the severity of depressive symptoms in major depressive disorder patients

Background and objectives: Despite the growing pieces of evidence on the relationship between the altered expression level of miRNAs and major depressive disorder (MDD), few studies have focused on the relationship between the altered expression of miRNAs and the severity of depressive symptoms. This study aimed to investigate the relationship between the expression level of miRNA-4485 and the severity of depressive symptoms in major depressive disorder (MDD) patients.MethodsEighty MDD patients without antidepressants and 45 healthy controls were placed and tested for the expression level of miRNA-4485 using quantitative RT‒PCR. At the same time, the Hamilton Depression Scale (HAMD) was used to assess depression symptoms for MDD patients. Twenty-nine out of 80 MDD patients were selected for miRNA expression level testing and symptomatology assessments before and after three weeks of treatment.ResultsThe expression level of miRNA-4485 in the MDD group was significantly overexpressed compared to that in healthy controls (P < 0.05), and the expression level of miRNA-4485 in the higher HAMD group was also much higher than that in the lower HAMD group and healthy controls (P < 0.05). The expression level of miRNA-4485 in MDD patients was negatively correlated with HAMD total score, anxiety/somatization, and bodyweight factor score (P < 0.05), accounting for 9.4%, 12.4% and 5.7%, respectively. MiRNA-4485 significantly predicted MDD and the severity of depressive symptoms (P < 0.05). Compared with that before treatment, the expression level of miRNA-4485 was significantly downregulated after treatment, while the patient's depressive symptoms were improved (p < 0.05). The improvement in depressive symptoms was positively correlated with the downregulation of miRNA-4485, which could significantly predict the effects of antidepressant treatment on MDD (P < 0.05). (AU)

Rapid Detection of Alternaria Species Involved in Pear Black Spot Using Loop-Mediated Isothermal Amplification.

Alternaria species are the most important fungal pathogens that attack various crops as well as fruit trees such as pear and cause black spot disease. Here, a loop-mediated isothermal amplification (LAMP) assay is developed for the detection of Alternaria species. A. alternata cytochrome b (cyt-b) gene was used to design two pairs of primers and amplified a 229-bp segment of Aacyt-b gene. The results showed that LAMP assay is faster and simpler than polymerase chain reaction (PCR). LAMP assay is highly sensitive method for the detection of about 1 pg of genomic DNA of A. alternata by using optimized concentration of MgCl2 (4 mM) in final LAMP reaction. In contrast, the limit of detection was 1 ng of target DNA via conventional PCR. Among the genomic DNA of 46 fungal species, only the tubes containing DNA of Alternaria spp. except A. porri, A. solani, and A. infectoria changed color from orange to yellowish green with SYBR Green I including the main pathogens of pear black spot. The yellowish green color was indicative of DNA amplification. Moreover, LAMP assay was used for testing infected tissues among 22 healthy and diseased pear tissues; the orange color changed to yellowish green for infected tissues only. Altogether, we conclude that cyt-b gene can be used for the detection of Alternaria spp. via LAMP assay, which is involved in pear black spot disease.

Rapid Detection of Ustilaginoidea virens from Rice using Loop-Mediated Isothermal Amplification Assay.

Ustilaginoidea virens is an important fungus that causes rice false smut disease. This disease significantly reduces both grain yield and quality. Various methods have been developed for the detection of U. virens but most of these methods need sophisticated equipment such as a thermal cycler. Here, we present a loop-mediated isothermal amplification (LAMP) assay for the specific detection of U. virens. This assay used a specific region of the UvG-ß1 gene (212-bp region) to design six LAMP primers. The LAMP assay was optimized by the combination of rapidity, simplicity, and high sensitivity for the detection of about 1 pg of target genomic DNA in the reaction whereas, with polymerase chain reaction (PCR), there was no amplification of DNA with concentrations less than 1 ng. Among the genomic DNA of 22 fungus species and two strains of U. virens, only the tube containing the DNA of U. virens changed to yellowish green with SYBR Green I. The color change was indicative of DNA amplification. No DNA was amplified from either the other 22 fungus species or the negative control. Moreover, 20 spikelets and 22 rice seed samples were used for the detection of rice false smut via LAMP. The results were comparable with conventional PCR. We conclude that gene UvG-ß1 coupled with LAMP assay, can be used for the detection and identification of U. virens gene via LAMP.

Development of a nested-PCR assay for the rapid detection of Pilidiella granati in pomegranate fruit.

Pilidiella granati, a causal agent of twig blight and crown rot of pomegranate, is an emerging threat that may cause severe risk to the pomegranate industry in the future. Development of a rapid assay for the timely and accurate detection of P. granati will be helpful in the active surveillance and management of the disease caused by this pathogen. In this study, a nested PCR method was established for the detection of P. granati. Comparative analysis of genetic diversity within 5.8S rDNA internal transcribed spacer (ITS) sequences of P. granati and 21 other selected fungal species was performed to design species-specific primers (S1 and S2). This primer pair successfully amplified a 450 bp product exclusively from the genomic DNA of P. granati. The developed method can detect 10 pg genomic DNA of the pathogen in about 6 h. This technique was successfully applied to detect the natural infection of P. granati in the pomegranate fruit. The designed protocol is rapid and precise with a high degree of sensitivity.

Simple and rapid detection of Tilletia horrida causing rice kernel smut in rice seeds.

A simple and rapid method for the detection of Tilletia horrida, the causal agent of rice kernel smut, in rice seeds is developed based on specific polymerase chain reaction (PCR). To design the specific primers for the detection of T. horrida, partial sequences of internal transcribed spacer (ITS) DNA region of T. horrida, T. controversa, T. walkeri, T. ehrhartae, T. indica and T. caries were analyzed and compared. A 503-bp fragment was amplified with the designed primers from the T. horrida genomic DNA. However, no PCR product was obtained from the DNA of other five Tilletia species and 22 fungal plant pathogens tested in the present work indicating the specificity of the primers for the detection of T. horrida. The PCR was performed by directly using the spores, isolated from the 21 different rice seed samples, as template DNA. The T. horrida was detected in 6 of the samples, indicating that 28.6% of the rice samples were contaminated with the kernel smut pathogen. This simple PCR based diagnostic assay can be applied for the direct and rapid detection and identification of T. horrida to screen large numbers of rice seed samples.

Genotypes and Phenotypic Characterization of Field Fusarium asiaticum Isolates Resistant to Carbendazim in Anhui Province of China.

Fusarium asiaticum is a causal agent of Fusarium head blight (FHB) of wheat in the southern part of China. Carbendazim has been extensively used for controlling FHB for more than 30 years, leading to the widespread carbendazim-resistant isolates in all major wheat-producing provinces in China, especially in Anhui Province. F. asiaticum isolates were collected throughout Anhui Province between 2010 and 2012 to monitor their sensitivity to carbendazim. In total, 74 of 899 single-spore isolates F. asiaticum were found to be resistant to carbendazim. Resistant isolates were collected from all of the sampled sites except Hefei of Anhui Province. The overall frequency of carbendazim resistance was shown to be 8.2%. Of the 74 isolates, 1, 68, and 5 had low resistance (LR), moderate resistance (MR) ,and high resistance (HR), respectively, to carbendazim. Five types of point mutations (F167Y, E198L, E198K, F200Y, and E198Q) in the ß2-tubulin gene conferring resistance to carbendazim were detected in the field-resistant isolates with frequencies of 89.2, 2.7, 4.1, 2.7, and 1.4%, respectively. The point mutations at codon 167, 198, or 200 of the ß2-tubulin gene were correlated with different levels of carbendazim resistance. Some of the sensitive and resistant isolates appeared to possess different biological characteristics; however, these might not be due to resistance. Because carbendazim resistance was generally widespread throughout Anhui Province, the sensitivity of F. asiaticum populations to carbendazim should be constantly monitored for the development of carbendazim resistance in natural populations.

Integrated Use of Pyraclostrobin and Epoxiconazole for the Control of Fusarium Head Blight of Wheat in Anhui Province of China.

Fusarium asiaticum and F. graminearum are the primary causal agents of Fusarium head blight (FHB) of wheat in China. Carbendazim (a benzimadazole fungicide, MBC), has been extensively used for the control of FHB, resulting in severe MBC resistance in China. This article presents the baseline sensitivity of F. asiaticum and F. graminearum isolates from Anhui Province of China to fungicides pyraclostrobin (a quinone outside inhibitor) and epoxiconazole (a sterol demethylation inhibitor). In the presence of salicylhydroxamic acid, the 50% effective concentration (EC50) values for pyraclostrobin in inhibiting mycelial growth of the 126 F. asiaticum isolates and 63 F. graminearum isolates were 0.012 to 0.135 µg/ml and 0.010 to 0.105 µg/ml, and the EC50 values for pyraclostrobin in inhibiting conidium germination of the F. asiaticum and F. graminearum populations were 0.047 to 0.291 and 0.042 to 0.255 µg/ml, respectively. The EC50 values for epoxiconazole in inhibiting mycelial growth of the F. asiaticum and F. graminearum populations were 0.12 to 0.95 and 0.16 to 0.93 µg/ml, respectively. All of the baseline sensitivity curves were unimodal. This study also suggested that there was no cross-resistance between MBC and pyraclostrobin or epoxiconazole. In the protective and curative tests, pyraclostrobin and epoxiconazole applied at 200 and 300 µg/ml exhibited over 75% protective and curative control efficacy in all treatments. In field trials, both pyraclostrobin and epoxiconazole at 225 g a.i./ha provided over 80% efficacy in 2010 and 2011 at both sites where MBC resistance occurred, suggesting excellent activity against FHB. Interestingly, integrated use of pyraclostrobin + epoxiconazole applied at 150 + 150 g a.i./ha provided over 85% efficacy at both sites in 2010 and 2011. Pyraclostrobin and epoxiconazole should be good alternatives to MBC for the control of FHB, and integrated use of these two fungicides might achieve greater efficacy.

[3T3L1 adipocytes induce rat beta-cell dysfunction].

OBJECTIVE: To investigate the integrated effects of adipocytes on rat beta-cells, differentiated 3T3L1 adipocytes and rat islet cells co-culture system was established. METHODS: There were two groups: control group (SD rat islet cells) and co-culture group (islet cells and 3T3L1 adipocytes coculture system). Islet cells were obtained for determination of (1) insulin secretion and insulin content; (2) mRNA expressions of GLUT2, GCK and Kir6.2; (3) protein expressions of IR-beta, IRS-1 and their tyrosine phosphorylation level. RESULTS: (1) At low glucose, insulin secretion of co-culture group increased compared with that of control group (0.79 +/- 0.35) ng x h(-1) x ml(-1) islet vs. (0.38 +/- 0.09) ng x h(-1) x ml(-1) x islet, P = 0.028. At high glucose, insulin secretion of those two groups was almost at the same level (P = 0.760). Compared with control group (2.84 +/- 0.92), stimulation index (SI, insulin release at high glucose/ low glucose) of co-culture system decreased to (1.57 +/- 0.61, P = 0.04). And the insulin content of the both groups was almost at the same level (P = 0.102). (2) The mRNA of GCK, GLUT2 and Kir6.2 in co-culture group downregulated to (0.27 +/- 0.11, P = 0.01), (0.34 +/- 0.24, P = 0.009) and (0.41 +/- 0.09, P = 0.003) compared with control group (mRNA = 1). (3) The protein levels of IR-beta, IRS-1 and their tyrosine phosphorylation decreased in co-culture system. CONCLUSIONS: 3T3L1 adipocytes are involved in beta-cell dysfunction, which may facilitate the development of type 2 diabetes. The effects may be mediated by multiple pathways, which include downregulation of GSIS related gene expressions and suppression of islet cell insulin signaling.