Specific Primer Design for Acetylcholinesterase-1 (ace-1) as a Putative Insecticide Resistance Gene for Aedes aegypti

Dengue fever caused by Aedes aegypti mosquito is the main vector of dengue hemorrhagic fever (DHF) in urban areas. This study aims to determine differences in density of Aedes aegypti mosquitoes based on the floor of the building, because the number of dengue fever in Bukittinggi is quite high. This research was conducted in May-June 2019 in the STIKes Fort De Kock Bukittinggi building. This research is experimental with the independent variable building floor density and the dependent variable is the density of the aedes aegypti mosquito seen from the ovitrap index. The object of the experiment was ovitrap, the number of ovitraps used was 54 ovitraps. The results of this study to determine differences in density of Aedes aegypti mosquitoes based on the floor of the building at various concentrations were tested using ANOVA. The results showed that of 54 ovites there were 86 Aedes aegypti mosquito eggs. Based on the ANOVA test results with an ? value of 5% or 0.05, p value = 0.524 (p??) is obtained, which means that H? is accepted. So there is a minimum floor that does not have a significant difference in the density of Aedes aegypti mosquito eggs. It can be concluded that there is no difference based on the floor with the density of Aedes aegypti mosquito eggs including, with an ? value of 5% or 0.05, a p value = 0.524 (p??), which means that H? is accepted. So there is a minimum floor that does not have a significant difference in the density of Aedes aegypti mosquito eggs. Keywords :Floor,ovitrap,Aedes aegypti mosquito

AIMS: To develop a DNA based plate hybridisation assay for the detection of polymerase chain reaction (PCR) products amplified from Aspergillus fumigatus DNA; and to determine the sensitivity of this...

Human chromosome band 16q24 commonly undergoes loss of heterozygosity (LOH) in human hepatocellular carcinoma (HCC). To further localize the region of deletion on 16q24 and to evaluate the genetic role of 17-beta-HSD, which is near 16q24, in HCC, we examined the pattern of loss of heterozygosity in 88 HCC patients. DNAs from 88 pairs of HCCs and corresponding non-tumor parts were prepared. Loss of heterozygosity on chromosomes 16q24 was investigated by 11 sets of microsatellite markers. Mutation analysis of type II 17-beta-HSD was performed by automatic sequencing. LOH on 16q24 for at least 1 locus was found in 43 of the 88 tumor DNAs (49%). Three non-overlapping regions of frequent LOH were defined in these 43 tumors with partial deletions. The first region was between D16S516 loci and D16S507, encompassed by a 1-cM region, defined by the D16S504. The second region was defined by the 17HSDB2 locus between D16S505 and D16S422, encompassed approximately by a 1-cM region. The third region was between D16S520 and D16S413, defined by D16S3048, encompassed approximately by a 4-cM region. Homozygous deletions of any exons in 17HSDB2 gene were identified in 7 of 27 cases (26%). Automated sequencing analysis of 17HSDB2 failed to demonstrate mutations in any of these specimens. Our data suggest that the 17HSDB2 locus is a frequent target of deletion in HCC but the inactivation of 17HSDB2 may not involve sequence mutations. Furthermore, the presence of the other 2 frequent LOH regions suggest that the putative tumor suppressor genes at these locations might be involved in the development of HCC.

Aedes aegypti L. Mosquito is the only vector of Dengue Hemorrhagic Fever (DHF). The chemical-based control agents have a negative effect if used for a long time. Therefore, this research focused on finding the natural agent that may act as an adulticide, like Bintaro (Cerbera manghas L.). Bintaro leaves contain active compounds such as cerberin (one of the alkaloids), flavonoids, tannins, steroids, and saponins that have a mortality effect on mosquitoes. The study was to determine the effectiveness of Bintaro leaves extract as an adulticide against Aedes aegypti mosquitoes. This experimental research uses the posttest-only control group design and uses Ae. Aegypti mosquito aged 3-5 days as the sample. There were seven different concentrations of Bintaro leaves extract as a treatment, namely 300 ppm, 350 ppm, 400 ppm, 450 ppm, 500 ppm, 550 ppm, and 600 ppm. The method used in this research is using a 250 ml Duran bottle. The results of the study showed that the Bintaro leaves extract at a concentration of 600 ppm was more effective against the mortality of Ae. Aegypti, but not significantly different from the 400 ppm, 450 ppm, 500 ppm, and 550 ppm treatments. And the LC50 probit regression analysis was 455.602 ppm, and the LC90 value was 1735.956 ppm.

Studies have shown evidence of production of nitric oxide (NO) in adipose tissue, as well as inhibition of lipolysis by NO. We have analyzed nitric oxide synthase (NOS) expression in subcutaneous adipose tissue from 13 nonobese and 18 obese male subjects. Using a competitive reverse transcription polymerase chain reaction method, endothelial (eNOS) and inducible (iNOS), but not neuronal (nNOS), nitric oxide synthase mRNA expression was detected in isolated fat cells and pieces of adipose tissue. Tissue mRNA levels for eNOS were 3,814 +/- 825 and 5,956 +/- 476 amol/mg RNA (P = 0.043), and for iNOS 306 +/- 38 and 332 +/- 48 amol/mg RNA, for nonobese and obese individuals, respectively. Western blotting revealed similar eNOS protein levels in isolated fat cells and adipose tissue pieces. Protein levels for eNOS in nonobese and obese individuals, respectively, were (in optical density [OD] units per mm(2) per 100 microgram of total protein) 0.11 +/- 0.08 and 2.80 +/- 1.30 (P = 0.043). iNOS protein was detectable, but not measurable, at low levels in a subset of obese patients (3 of 10). iNOS protein levels could not be detected in nonobese individuals. Hormone-sensitive lipase (HSL), the key regulating enzyme in lipolysis, is reduced in obesity. The expression of HSL protein in subcutaneous adipose tissue was studied in the same subset of patients; in agreement with previous results, HSL levels were reduced in obese subjects: 4.64 +/- 1.10 and 1.27 +/- 0.35 (P = 0.012) in nonobese and obese subjects, respectively. In conclusion, this study shows that eNOS and iNOS, but not nNOS, are present in human subcutaneous adipose tissue. Gene expression and protein levels of eNOS are increased, whereas HSL protein levels are decreased in obesity. It is speculated that increased NO production, preferably by eNOS, and decreased HSL levels may cause decreased subcutaneous adipose tissue lipolysis in obesity. synthases in subcutaneous adipose tissue of nonobese and obese humans.

Antibiotic resistance and the golden age of microbiology

Frequent T and B Cell Oligoclones in Histologically and Immunophenotypically Characterized Angioimmunoblastic Lymphadenopathy

Betaine, Dimethyl Sulfoxide, and 7-Deaza-dGTP, a Powerful Mixture for Amplification of GC-Rich DNA Sequences

The fungus Erysiphe lagerstroemiae is commonly known as the powdery mildew pathogen in crape myrtle (Lagerstroemiae indica) in the United States, and Erysiphe australiana is the powdery mildew pathogen reported in Japan, China, and Australia. The teleomorph often used to identify powdery mildew fungi rarely develops in crape myrtle, and in our observations, ascocarps never formed. Our study showed that the crape myrtle pathogen overwintered as mycelia on dormant buds. The internal transcribed spacer (ITS) regions of rDNA and the intervening 5.8S rRNA gene were amplified using standard polymerase chain reaction (PCR) protocols and the universal primer pairs ITS1 and ITS4. PCR products were analyzed by electrophoresis in a 1.5% agarose gel and sequenced, and the ITS PCR product was 666 bp from ITS1/ITS4 and 704 bp from ITS1-F/ITS4. BLAST analysis of the sequence of the PCR products showed identical similarity with E. australiana reported in Japan, China, and Australia. Comparison of ITS sequences with information in the GenBank on other powdery mildew fungi showed a closest alignment (93% similarity) to Erysiphe juglandis that infects walnut. Specific primers for E. australiana were developed and evaluated for use as diagnostic tools. Out of 12 specific primer pairs evaluated, four primer pairs and four double primer pairs were highly specific to E. australiana and did not amplify Erysiphe pulchra of dogwood, Erysiphe syringae of common lilac, Erysiphe circinata of maple, or Phyllactinia guttata of oak. The E. australiana-specific primers amplified 16 samples of crape myrtle powdery mildew collected from diverse locations in mid-Tennessee. These results clearly showed that the crape myrtle powdery mildew in mid-Tennessee was caused by E. australiana. Specific primers reported in this article provide a diagnostic tool and may be used to confirm the identity of crape myrtle powdery mildew pathogen in other areas in the United States and wherever the disease occurs.

Direct sequencing of polymerase chain reaction (PCR) products without cloning is a rapid and efficient way of sequence analysis. Prior to direct sequencing, it is necessary to purify the PCR products from excess primers, nucleotides and enzymes that could interfere with the sequencing reaction. There are several PCR product purification methods such as spin column-based purification, enzymatic purification, ethanol precipitation and gel extraction. In this study, it is aimed to evaluate the efficiency of ethanol-ammonium acetate (EtOH-NH4Ac) and polyethylene glycol (PEG) precipitation methods for purification of PCR products prior to the dye terminator cycle sequencing. A 741 bp region of the Toll-Like Receptor (TLR) 4 gene was amplified from bovine genomic DNA using PCR. After analyzing the PCR product using agarose gel electrophoresis, it was purified with one of the following methods: a) PEG precipitation, b) EtOH-NH4Ac precipitation and c) ExoSAP-IT PCR product cleanup reagent. ExoSAP-IT reagent was used as a standard PCR product cleanup protocol. Sanger sequencing of PCR samples purified with different purification methods was performed on a Beckman Coulter CEQ8800 Genetic Analysis System. The sequence data were analyzed using Sequencing Analysis software implemented within the system. The quality check and alignment of sequences were performed using BioEdit software. The sequencing results of PCR products purified with different purification methods were compared with each other. It was found that PCR products purified with both purification methods provided good-quality sequencing templates like that of purified with ExoSAP-IT reagent.

Aedes aegypti is the primary vector of dengue hemorrhagic fever (DHF), which remains a major public health concern in Indonesia. Continuous use of synthetic insecticides has led to the emergence of resistance andadverse environmental impacts. This study aimed to evaluate the biopotential of ethanol extract from Altingia excelsa (rasamala) leaves as a botanical insecticide against Aedes aegypti mosquitoes. The experiment was conducted by exposing female Aedes aegypti mosquitoes aged 3–5 days to vapor from rasamala leaf extract at threeconcentrations (25%, 50%, and 75%) using an electric liquid vaporizer device. Mortality percentages were recorded at 8, 16, and 24 hours and subsequently analyzed using statistical and probit methods. The results demonstrated that rasamala leaf extract possesses insecticidal activity. However, the LC₅₀ value of 182% indicated low efficacy, as it exceeded the highest concentration tested. Moreover, the LT₅₀ value could not be reliably determined within the observed time frame. These findings suggest that while Altingia excelsa extract exhibits insecticidal potential, further optimization of its formulation or an increase in concentration is required to achieve greater effectiveness.

Culture of rat pancreatic islets with interleukin-1 (IL-1) results in up-regulation of the inducible isoform of nitric oxide synthase and overproduction of nitric oxide (NO). This is associated with reversible inhibition of both glucose-induced insulin secretion and islet glucose oxidation, and these effects are prevented by the inducible nitric oxide synthase inhibitor NG-monomethylarginine. IL-1 also induces accumulation of nonesterified arachidonic acid in islets by an NO-dependent mechanism, and one potential explanation for that effect would involve an IL-1-induced enhancement of islet glycolytic flux. We have therefore examined effects of IL-1 on islet glycolytic utilization of glucose and find that culture of islets with IL-1 in medium containing 5.5 mM glucose results in suppression of islet glucose utilization subsequently measured at glucose concentrations between 6 and 18 mM. The IL-1-induced suppression of islet glucose utilization is associated with a decline in islet glucokinase mRNA content, as determined by competitive reverse transcriptase-polymerase chain reaction, and in glucokinase protein synthesis, as determined by immuoprecipitation experiments, and all of these effects are prevented by NG-monomethylarginine. These findings suggest that IL-1 can down-regulate islet glucokinase, which is the primary component of the islet glucose-sensor apparatus, by an NO-dependent mechanism. Because reductions in islet glucokinase levels are known to cause a form of type II diabetes mellitus, these observations raise the possibility that factors which increase islet NO levels might contribute to development of glucose intolerance.

Large Pathogenic Expansions in the SCA2 and SCA7 Genes Can Be Detected by Fluorescent Repeat-Primed Polymerase Chain Reaction Assay

Development and application of DNA analytical methods for the detection of GMOs in food

We have surveyed the tandemly repeated genes encoding U2 snRNA in a diverse panel of humans. We found only two polymorphisms within the U2 repeat unit: a SacI polymorphism (alleles SacI+ or SacI-) and a CT microsatellite polymorphism (alleles CT+ or CT-). Surprisingly, individual U2 tandem arrays are entirely SacI+ or SacI-, and entirely CT+ or CT-, although the SacI and CT alleles can occur in any combination. We also found that polymorphisms in the left and right junction regions flanking the tandem array fall into only two haplotypes (JL+ and JL-, JR+ and JR-). Most surprisingly, JL+ is always associated with JR+, and JL- with JR-. Thus individual U2 arrays do not exchange flanking markers, despite independent assortment and subsequent homogenization of the SacI and CT alleles within the U2 repeat units. We propose that the primary driving force for concerted evolution of the tandem U2 genes is intrachromosomal homogenization; interchromosomal genetic exchanges are much rarer, and reciprocal nonsister chromatid exchange apparently does not occur. Thus concerted evolution of the U2 tandem array occurs in situ along a chromosome lineage, and linkage disequilibrium between sequences flanking the U2 array may persist for long periods of time.