Journal of BioScience and Biotechnology

Frugal method for extracting DNA from saliva suitable for low-budget labs and usable in PCR assays

2024-05-08T08:49:34+00:00

Blood is the preferred source of DNA as it yields large quantities of high-quality DNA. However, there are challenges associated with this process, including difficulty in finding small and thin veins, rigid veins due to aging, and individuals with phlebotomy aversion, especially children. Therefore, obtaining DNA from saliva is considered a suitable alternative that requires minimal resources and is usable in DNA assays without the need for a phlebotomist, at a lower cost compared to most DNA extraction kits that require expensive materials. The suggested frugal method relies on providing a new solution for extracting DNA from human saliva without the need for enzymes or high grade reagents or columns, which makes suitable for low budget laboratories in developing countries to manufacture it locally, it uses a phase separation with technical skills to extract the DNA from the saliva achieving safety by not using hazardous reagents, and it is a sustainable eco-friendly method that gives decent yield of good quality DNA in ~20 minutes to be used in PCR applications with a cost that is estimated to be 10 times cheaper than current used methods.

Exploring the coordination, anti-oxidant, and bacterial behavior of a new azo ligand derived from 4,5-dimethylimidazole with copper and zinc divalent ions

2024-09-24T07:07:10+00:00

New azo ligand synthesized from 4, 5-dimethylimidazole, focusing on its coordination properties with copper and zinc divalent ions. The experimental findings confirmed the formation of octahedral geometrical complexes, where the coordination occurs through one nitrogen atom of azo and an N3 atom of imidazole. Evidence from spectroscopic techniques (13-CNMR, Mass, FT-IR, and UV-Vis), conductivity measurements, and magnetic susceptibility supported the structural integrity of the complexes. Biological evaluation showed that the ligand and its complexes possess notable antibacterial properties, particularly the zinc complex, which exhibited stronger inhibitory effects against both Staphylococcus and E. coli bacteria. Antioxidant tests using the DPPH assay indicated that the ligand effectively scavenges free radicals, achieving 70% inhibition at 25 ppm, making it a promising candidate when compared to ascorbic acid. The results underscore the potential of this ligand and its complexes for applications in medicinal chemistry, especially in the development of antimicrobial and antioxidant agents.

Response surface methodological approach for optimizing the enzyme activity and enzymatically mediated bioprecipitation of heavy metals by alkaline phosphatase

2024-07-30T11:50:20+00:00

This study investigates using an alkaline phosphatase enzyme isolated from Bacillus cereus to decontaminate heavy metals. The experiments were performed with several process parameters, including substrate concentration, pH, and temperature. To optimize the best experimental conditions, they were estimated by using a central composite experimental design combined with response surface methodology (RSM). Variables were concentration of substrate ((p-NPP 14 to 17 mM), pH 8 to 10.5, and temperature (35 to 45^oC). Statistical analysis of variance (ANOVA) was performed to classify the competence of the developed model and revealed a good understanding between the experimental data and the proposed model. The highest enzymatic activity 25.73 units/ml was identified by the RSM with the following optimal set of parameters: concentration of substrate 15.5 mM, pH 9.25, and temperature 34oC. The accuracy of the predicted model optimum parameters was confirmed by experimenting under the same parameters. It was found that the experimental enzyme activity efficiency under optimum conditions was very close (less than a 3% error) to the model-predicted value. The removal efficiency of each heavy metal was found to be in the following order: Hg >Pb>As.

Characterization and evaluation of antioxidant properties of leaf extracts from Rhododendron arboreum

2024-10-18T17:39:32+00:00

Phytochemicals have been known to suppress the microbial activity and oxidative stress, which indicates the significance of plant material as anti-microbial and antioxidant agent. In this light the current study is worked out to screen the phytochemicals from the Rhododendron arboretum leaf extract and characterize it using HPLC. The alkaloids and flavonoids are the major secondary metabolites found during screening. Further, anti-oxidant properties of leaf extract of R. arboreum prepared in different solvent (methanol, ethyl acetate, petroleum ether, and chloroform) were analyzed using DPPH and FRAP assay. Observation of the present study indicates that methanolic extracts was found to show significant anti-oxidant activity with the IC₅₀ value of 241.2 µg/ml and 163.6 µg/ml in DPPH and FRAP assay respectively. In conclusion, the leaf extracts of R. arboretum can be one of the better natural source of antioxidant source and could be used for therapeutic purpose in the treatments of different diseases.

Analgesic effect of ethylacetate fraction of the methanol leaves extract of Hannoa klaineana in rats

2024-09-20T06:27:00+00:00

Pain remains a major health, social, and economic problem worldwide. Hannoa klaineana Pierre & Engl. is a medicinal plant found in many African countries and used for the treatment of many diseases including pain-associated disorders. This study was conducted to evaluate the analgesic effect of the ethylacetate fraction of methanol leaf extract of Hannoa klaineana in rats. The analgesic effect of the ethylacetate fraction of methanol leaves extract of Hannoa klaineana (200, 400, and 600 mg/kg b.wt) was evaluated using acetic acid-induced writhing, tail flick (immersion), and hot plate model. In the acetic acid-induced writhing test, the extract (200, 400, and 600 mg/kg) demonstrated a significant (p < 0.05) decrease in the number of writhes with maximum percentage inhibition (75.61%) at 600 mg/kg dose of the extract. In tail flick and hot plate tests, the extract (200, 400, and 600 mg/kg b.wt) exhibited a significant (p < 0.05) increase in rats’ response with a steady increase in reaction time. Findings from this study show that ethylacetate fraction of the methanol leaves extract of Hannoa klaineana possessed analgesic activity which provided justification for the local use of the plant in the treatment of pain.

Cloning, characterization and expression pattern of the ovarian cytochrome P450 Cyp19a1a gene in gonadal developmental period of cobaltcap silverside Hypoatherina tsurugae

2024-08-07T15:53:18+00:00

The upregulation of the cyp19a1a transcription factor is required for granulosa cell differentiation and ovarian maintenance. 1630 bp of cyp19a1 mRNA transcript of Hypoatherina tsurugae was cloned and sequenced. It consists of an open reading frame (ORF) of 1551 bp that encodes a 517 aa protein, found to be identical to the sequence of other fish species. A phylogenetic tree was constructed by comparing the mRNA sequence of 41 different fishes across various taxa available in the NCBI database and using an outgroup as Acipenser sinensis. The tree shows a high homology of cyp19a1a from H. tsurugae with cyp19a1a of Maelanotaenia boesemani, the two forming a single clade. The qRT expression of cyp19a1a was studied in both amhy+ (male) and amhy- (female) individuals. In amhy- (female) individuals, the expression was begins from 0 wah and peak at 6 wah then sharply decreases whereas in amhy+ (male) individuals expression was very low and it is in the baseline. The histological sections of gonads were studied in different stages of biweekly collected larvae during the sex determination/differentiation period and it showed that differentiation of gonads male/female was decided at 6 wah. In this stage, the primary oocytes are recognized. These findings add to the knowledge for a better understanding of molecular mechanisms of sex determination and differentiation period in fishes.

Optimization and validation of a real-time PCR method for the simultaneous detection of Lactococcus garvieae and Streptococcus agalactiae in fish

2024-09-24T06:51:19+00:00

Background: Lactococcus garvieae and Streptococcus agalactiae infections contribute to heavy losses in aquaculture farms worldwide. Currently, available pathogen diagnostic tools use biochemical and microbiological methods beleaguered by very low accuracy, reproducibility and specificity.

Aim: To optimize and validate a rapid, sensitive and specific real-time PCR (qPCR) method for detecting L. garvieae and S. agalactiae in fish.

Methods: Pairs of Streptococcus-specific (IGS-s/IGS-a) and Lactococcus-specific (CAU12F/CAU15R) primers were tested for specificity and sensitivity in the qPCR. qPCR was carried out at different temperatures and primer concentrations. The optimal conditions were determined to be the temperature and primer concentration with the lowest C_T values.

Results: For both primer sets, the optimal annealing temperature was 60^oC, and the optimal primer concentration was 500 nM. The detection limit for L. garvieae was at dilution factor 10^-3, with a mean C_T value of 25.0, for S. agalactiae, 10^-4 with a mean C_T value of 29.8. The PCR efficiencies were 97% for L. garvieae and 91% for S. agalactiae, with linear slopes (R² = 0.999). The assay demonstrated high repeatability and reproducibility.

Conclusion: The optimum conditions established for the qPCR method enable rapid, highly sensitive and specific diagnosis of L. garvieae and S. agalactiae infection in fish.

Phytochemical screening and anti-Helicobacter pylori activity of Bryophyllum pinnatum, Ocimum gratissimum and Vernonia amygdalina

2024-09-19T07:35:48+00:00

Helicobacter pylori, a common gastrointestinal bacterial pathogenic isolate infects 50% and 90% of the global and developing nations population respectively. This study aimed at evaluating the bioactive components and therapeutic potential of Bryophyllum pinnatum, Ocimum gratissimum and Vernonia amygdalina plants extracts against Helicobacter pylori infection using standard physicochemical, in-vitro and in-vivo microbiological methods. Five (5) stool samples were collected from patients who presented with symptoms of gastrointestinal distress and diagnosed of ulcer at the University of Benin Teaching Hospital, for the isolation of Helicobacter pylori. Phytochemical screening of ethanol extract of the test plants revealed the presence of bioactive constituents such as flavonoids, tannins, cardiac glycosides, saponin, steroids, phenols, alkaloids and terpenoids. The minimum inhibitory concentration (MIC) of the plant’s extracts were determined at concentrations of 125, 250, 500, and 1000 μg/ml. While the different plants extract demonstrated a better anti-Helicobacter pylori activity as well as MIC when compared to the commonly used antibiotic amoxicillin, and other conventional antibiotics, the anti-Helicobacter pylori activity and MIC of Vernonia amygdalina was higher, followed by Bryophyllum pinnatum and Ocimum gratissimum. The in-vivo study as carried out using Wistar albino rats demonstrated the promising therapeutic effect of the plants extract against Helicobacter pylori infection. This study therefore suggests that Vernonia amygdalina, Bryophyllum pinnatum and Ocimum gratissimum extracts possess anti-Helicobacter pylori properties, offering safe, effective, and cost-effective treatment options for the treatment of ulcer, caused by Helicobacter pylori.

Synergistic effect of meropenem and vancomycin antibiotics with Ecballium elaterium and Senna italica extracts

2024-08-07T12:56:54+00:00

The aim of this study is to determine whether the use of plant extracts like Ecballium elaterium and Senna italica leaf aqueous extracts helps support the activity of meropenem and vancomycin antibiotics against Staphylococcus aureus and Staphylococcus epidermis bacteria. The minimum inhibitory concentration (MIC) and the fractional inhibitory concentration (FIC) were used to find out the antibacterial effect of E. elaterium and S. italica extracts in combination with meropenem and vancomycin antibiotics by employing microbroth dilution assay and checkerboard assay. According to the obtained results, all combinations showed a synergistic effect against S. aureus except vancomycin and S. italica combinations which exhibited an additive effect. Moreover, all combinations exhibited an additive effect against S. epidermis except vancomycin and E. elatenium, which showed a synergistic effect. In conclusion, E. elaterium and S. italica leaf aqueous extracts have great potential as sources of antibacterial compounds against microorganisms and they can be used to treat infectious diseases caused by antibiotic-resistant microorganisms.