Journal of Applied Biological Sciences

PROTECTIVE EFFECT OF ALGERIAN SALVIA VERBENACA EXTRACT AGAINST INFLAMMATION AND OXIDATIVE STRESS

Lilia BOUSSOUF — 2023-01-29

This study aims to examine the anti-inflammatory and antioxidant effects of polyphenolic extract of Salvia verbenaca L. using in vivo and in vitro models. The in vivo anti-inflammatory activity was evaluated using carrageenan-induced mice paw edema method. The in vivo antioxidant effect of the extract was explored by measuring oxidative stress (MDA, CAT, GSH and SOD) parameters. The in vitro anti-inflammatory activity was performed using protein denaturation and membrane stabilization assays. DPPH and hydrogen peroxide tests were used for the in vitro antioxidant evaluation. The highest percentage of edema inhibition was 75.03% at the concentration of 500 mg/kg. Significant increases (p˂0.05) in the activities of CAT, SOD, GSH and significant decreases in the MDA level activity were showed. The extract was found to possess an in vitro anti‐inflammatory activity by inhibiting the heat induced protein denaturation and red blood cells membrane stabilization with the IC₅₀ values of 133.21±0.89 and 160.62±1.60 μg/mL, respectively. This study demonstrated that extract exhibited high free radical scavenging activity as showed by the low IC₅₀ values for DPPH (115.41±1.40 μg/mL) and for H₂O₂ (180.39±1.77 μg/mL) scavenging assays. Our findings suggest that Salvia verbenaca could be tested as a drug candidate against oxidative stress and inflammation diseases.

ANTIFUNGAL AND ANTIOXIDANT ACTIVITIES OF ACTINOBACTERIA ISOLATED FROM ALGERIAN DESERT SOILS

Habiba Belgacem — 2023-01-29

Actinomycetes are gram-positive filamentous bacteria that are thought to be the best candidates for producing biologically active secondary metabolites. Our research aims to identify antifungal and antioxidant activity, as well as purify and characterize the components involved in this phenomenon in isolated actinomycetes derived from the Algerian desert soil. Eighteen isolates were studied. The antagonism tests are performed by the agar cylinder technique against two gram-positive bacteria, Staphylococcus aureus and Listeria monocytogenes, and five fungi, which are known for their importance in human infectious pathologies. 14 isolates showed inhibitory activity against the two bacteria, and 15 isolates showed antifungal activity against the five fungal species: Fusarium graminearum, Aspergillus flavus, Aspergillus ochraceus, Penicillium expansum, and Aspergillus parasiticus. The results allowed us to choose two strains of Actinomycetes coded A10 and A7, both of which have strong inhibitory abilities. That was used for solid-state fermentation (AF). After 14 days of incubation at 28 °C, methanol is used for the extraction of the metabolites produced in the medium. The methanolic extract was effective against the five target fungi species targets even at low doses of 2 mg/ml. The ergosterol degradation test revealed that strain A7 generates an antifungal agent capable of changing membrane sterols, but strain A10 had no impact. A characterization of the inhibiting agent using chromatographic techniques was carried out. The antioxidant activity of the two crude extracts was measured using DPPH radical scavenging. The results demonstrate that the A10 strain extract has good activity, with a percentage inhibition of 58% and an IC50 test of 8.10. The study of the morphological, cultural, physiological, and biochemical characteristics of the two isolates, A7 and A10, allowed us to link them to the genus Streptomyces.

DETERMINATION OF ANTIBIOTIC RESISTANCE PROFILES IN ACINETOBACTER SPECIES ISOLATED FROM TANK MILK

Cansu Önlen Güneri — 2023-01-29

The aim of this study was to determine the occurrence of Acinetobacter spp. in tank milk samples (TMS) in Hatay province and to analyze their antimicrobial susceptibility profile. In this study, a total of 60 TMS were collected between April and December 2016. Acinetobacter spp. was identified by using Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) mass spectrometry (MS). The identified Acinetobacter isolates were tested to determine the antimicrobial resistance profiles towards 23 antibiotics using the disc diffusion method. A total of 13 Acinetobacter isolates were obtained from TMS (21.6%). Species distribution were as follow; Acinetobacter baumannii (n=9), Acinetobacter ursingii (n=3) and Acinetobacter iwoffii (n=1). Resistance to aztreonam, chloramphenicol, amoxicillin and clavulanate, cefoxitin, cefpodoxime gentamicin, cefuroxime, florphenicol was 53.8%, 15.3% 15.3%, 15.3%, 15.3%, 7.7%, 7.7%, and 7.7%, respectively. However, none of the isolates was resistant to nalidixic acid, ciprofloxacin, enrofloxacin, levofloxacin, norfloxacin, penicillin, tetracycline, sulfamethoxazole/trimethoprim, ampicillin, imipenem, colistin, cephalothin, and ceftazidime. As a result, the TMS were found to be contaminated with different species of Acinetobacter. Another important finding of this study is the antimicrobial resistance characteristics of these strains, which might pose a public health problem. Therefore, it may be said that TMS would be better to be monitored for this potentially pathogenic bacterium.

ASSESSMENT OF VOLATILE FATTY ACIDS AND METHANE PRODUCTION IN VITRO OF OPUNTIA FICUS-INDICA CLADODES IN AN ALGERIAN ARID AREA

Samir Medjekal — 2023-01-29

The main objective of the present study was to evaluate the volatile fatty acids (VFAs) and methane (CH₄) production of cactus Opuntia Ficus-Indica (O. ficus indica) Cladodes, Medicago sativa L. (M. sativa) and barley straw (H. vulgare) using chemical composition and in vitro gas production. There were significant (p<0.05) differences among studied samples in all measured nutrients except dry matter (DM) value. O. ficus indica showed particularly the highest Ash content (284.07 g/kg DM). The crude protein (CP) of the studied feedstuff ranged from 51.41 to 156.09 g/kg DM and it was especially high within M. sativa and low in H. vulgare. No significant difference was observed in pH values, total VFAs, propionate concentrations and CH₄production (ml/100 gas) among plants species studied. The lowest Acetate concentration was observed in H. vulgare (711.17 mmol/mol) and M. sativa (715.33 mmol/mol), respectively (p < 0.05). The maximum in vitro gas production was recorded with M. sativa and H. vulgare. Based on the results, it can be concluded that O. ficus indica could replace common plants species, especially in the arid and semi-arid areas in which livestock production frequently suffers from low efficiency and big losses. Furthermore, these plantations in agroforestry development could consolidate the protection of the environment against erosion, allowing the fixation of the soil and its protection against water erosion.

COMPARATIVE EVALUATION OF MULTIDRUG RESISTANCE AND APOPTOSIS IN DIFFERENT VARIANTS OF HEPATOMA CELLS

Suman Jain — 2023-01-29

Multidrug Resistance is a major obstacle in cancer chemotherapy. The current study was designed to evaluate the multidrug resistance and apoptotic properties of Hepatoma cells. Antiproliferative effect of five anti-cancer drugs: three of bacterial origin namely, puromycin, actinomycin D, doxorubicin and two plant derived drugs viz., colchicine and vinblastine were assessed on heat- resistant variants of dexamethasone–resistant and the dexamethasone- sensitive variants of hepatoma cell lines. These variants showed increased drug resistance to the different anticancer drugs used, i.e., they became moderately multidrug-resistant. The severely heat-treated H56 cells became moderately resistant only to certain drugs. All the experimental variants of Hepatoma cells overexpressed functional P-glycoprotein, a drug-resistant associated marker protein, which attributes to the resistance shown by these cells. The anticancer drugs were capable of inducing apoptosis in both H56 and clone 2 cells and it was found that H56 cells were more prone to undergo apoptosis as compared to clone 2 cells. Furthermore, preliminary immunocytochemical studies revealed that there was a significant difference in expression of p53, a tumor suppressor gene, in H56 and clone 2 cells where H56 cells showed very strong staining for p53 thereby justifying their proneness to apoptosis.

SOME INDICATORS OF DAMAGES AND RESPONSES OF XANTHORIA PARIETINA (L.) Th. Fr. TO FLUORIDE AND LEAD INDUCED-STRESS

Ouahiba Benhamada — 2023-01-29

One of the major problems of current time is air pollution, the assessment of air quality through the use of bioindicators is a major concern at the moment. Our work aims to study the response of Xanthoria parietina (L.) Th. Fr. to fluoride- and lead-induced stress as an example of the phytotoxic air pollutants. For this purpose, lichen thalli have been treated with sodium fluoride (NaF) and lead nitrates (Pb(NO₃)₂) solutions at 0, 0.5, 1.0, 5.0 and 10 mM, for time scale of 0, 24, 48 and 96 h. Lipid peroxidation measured by Malondialdehyde (MDA) and chlorophyll degradation measured by optical density OD₄₃₅/OD₄₁₅ ratio are used as results of stress induced by fluoride and lead, and the accumulation of proline and soluble sugars are measured as indicators of responses used by X. parietina. Based on the obtained results, it was noted that lipid peroxidation increased correlating with increasing concentrations of NaF and Pb(NO₃)₂ (r=0.773, p=0.000712*** and r 0.865, p=0.000031***, respectively), however, chlorosis and proline increased correlating with increasing exposure time of NaF (r=- 0.737, p=0.0011** and r=0.783, p=0.00032***, respectively) and Pb(NO₃)₂ (r=-0.926, p<0.0001*** and r=0.811, p=0.00013***, respectively), whereas soluble sugar contents increased according to increasing concentrations of NaF (r=0.678, p=0.0010***) and according to increasing exposure time of Pb(NO₃)₂ (r=0.780, p=0.00036***). Although lead was significantly more toxic than fluoride (p=0.02*), X. parietina offers a very high sensitivity to fluoride, which allowed us to conclude that the toxicity of fluorine is comparable to that of lead.

INVESTIGATION OF THE CYTOTOXIC EFFECTS OF LAVANDULA STOECHAS ESSENTIAL OILS IN PANCREATIC DUCTAL ADENOCARCINOMA

Özge Kalhan — 2023-01-29

Pancreatic cancer is one of the highest mortality types in the world and it is reported that it ranks 4th in the latest data. In recent years, studies on the treatment of the disease has focused on active substances derived from natural products. Lavender plant is known to have many biological activities including cytotoxic effects in many cancer studies. The aim of this study is to investigate the cytotoxic efficacy of essential oils of Lavandula stoechas (L. stoechas) extracted by water distillation in pancreatic ductal adenocarcinoma. Pancreatic adenocarcinoma cells (AsPC-1) were produced on high glucose DMEM medium and separated into control and L. stoechas essential oil applied group (LS). L. stoechas essential oil was obtained by water distillation method and applied to cells at different concentrations (0.1, 0.25, 0.5 and 1 µg/ml) for 24 hours. Cell viability tests were performed by MTT method and effective concentration was determined. Expression levels of EGFR and K-ras oncogenes were investigated by real-time quantitative polymerase chain reaction from mRNA samples. Viability tests showed that LS has the effective concentration of 1 µg/ml concentration. In gene expression analysis, it was found that effective concentration of LS downregulates the expression of both EGFR and Kras oncogenes. In the light of all these data, it was concluded that essential oil components of L. stoechas could be considered as a potential agent in the regression of PDAK pathogenesis. In order to be able to say this subject in full, in vivo and further studies are needed.

HEALTH RISK ASSESSMENT OF HEAVY METALS IN GOAT’S MILK FOR ADULTS GROUP IN JIJEL PROVINCE (ALGERIA)

Nassima Balli — 2023-01-29

The continuous urbanization and industrialization in many parts of the world and Algeria have led to high levels of heavy metal contamination in the soil, groundwater, and food chain products. Given the importance of milk and dairy products in the human diet, the aim of this study is to estimate the concentration of heavy metals (lead, cadmium, zinc, and copper) in goat’s milk and to assess the potential health risks of those metals on adult consumers. To do so, goat milk was collected from three regions (El-Milia, Texenna, and Djimla) of Jijel, Algeria. Three samples were taken from each region and mixed to create a final sample that was pooled from each region. Then, Flame Atomic Absorption Spectrometry (FAAS) was used to determine the concentration of heavy metals. On the other hand, an online survey was carried out to estimate the daily intake of goat milk. The non-carcinogenic and carcinogenic health risk values were calculated. Our experiment demonstrated that the concentration of lead, cadmium, and copper significantly exceed the acceptable limit values; whereas, the zinc concentration was within the acceptable limit. The highest lead concentration was found in the El-milia region (0.42 mg/kg). The concentration gradient of average cadmium is as follows: C Djimla>C El-milia>C Texenna. They range from (0.0359±0.00247) mg/kg, (0.0155±0.00214) mg/kg, (0.00397±0.000742) mg /kg respectively, while the Texenna region has the highest copper concentration. Daily estimated intake (EDI) of all metals did not exceed the limit value. In addition, the target hazard quotient (THQ) of all metals and hazard index (HI) of the mixture of metals were both below the acceptable limit, representing no carcinogenic risk to the residents. Contrariwise, it was found that the carcinogenic risk (CR) for Cd was higher than the acceptable level (1×10⁶), as was the cumulative carcinogenic risk for Cd and Pb. This study will be quite helpful for both inhabitants in taking protective measures and government officials in reducing heavy metal contamination.

ANALYSIS OF PHYSICOCHEMICAL, NUTRITIONAL, PHYTOCHEMICAL PROPERTIES AND ANTIOXIDANT CAPACITY OF THREE STRAWBERRY CULTIVARS "FRAGACIA ×ANANASA DUCH." FROM EASTERN OF ALGERIA AND CARACTERIZATION OF STRAWBERRY JAM

Amina Bouchefra — 2023-01-29

In eastern Algeria, the predominant strawberry cultivars (Fragaria×ananassa Duch.) include Savana, Sabrina, and Fortuna. The aim of this study is to evaluate the physicochemical and antioxidant properties of three strawberry cultivars, as well as to characterize two types of strawberry jam. The values of compounds such as caliber, titratable acidity, total soluble solids, moisture, ash, protein, fat, carbohydrate, total phenolic and anthocyanin content, DPPH radical scavenging activity, total sugars and sensory analysis of the jam were determined. The titratable acidity and total soluble solids values demonstrated a significant variation (p˂0.05) throughout all strawberry cultivars. The protein and lipid content of the three strawberry cultivars was less than 1%. The strawberry cultivars contain significant levels of phenolic compounds. Strawberry cultivars have a DPPH free radical scavenging activity ranging from 40.84 to 45.25%. The strawberry jam with ginger (J1) had the highest ratings for all sensory qualities, making it the greatest jam in the judges' view. These findings give useful information on the phytochemical and nutritional quality of Algerian strawberries grown in the eastern region of the country and may be utilized to enhance the nutritional content of a functional food in a healthy diet.

FIELD TRIAL OF PGPR, Bacillus megaterium E-U2-1, ON SOME VEGETABLE SPECIES

Emel Ünlü — 2023-01-29

Excessive and unconscious use of chemical fertilizers and pesticides in crop production causes deterioration of soil health, environmental pollution, and an increase in the population of pathogens and pests in the soil. There are many toxic and dangerous chemicals in agricultural ecosystems, and they get into plants, soil, groundwater, and food. As an alternative way to solve this problem, it is very important to use beneficial microorganisms, to use biological agents instead of chemicals in agriculture, and to promote the resistance to biotic and abiotic stress conditions and to increase the yield of plants. Microbial fertilizers, which are becoming increasingly widespread in plant production, are produced from formulations containing Bacillus megaterium, especially from Bacillus group bacteria. In this study, Bacillus megaterium, E-U2-1 (OL673801 EU. U21-NCBI accession code) strain carrying phytase, phosphatase, siderophore, and indole acetic acid encoding genes was used for the trials. A field trial was carried out with some vegetable species (pepper, eggplant, cucumber, watermelon, melon, and snake cucumber) with high economic value. In the study, 5 ml, 10 ml, and 15 ml of bacterial suspensions with 3x107 cfu were applied per plant from the soil. It was determined that it increased vegetative growth and shortened the flowering period in all species. The effect of the bacterial application on the important agronomic properties of vegetables was investigated, and it was determined that especially 10 ml and 15 ml applications had a positive effect on yield and quality parameters. A significant increase was recorded vegetatively, and while flowering was observed in the earlier period, it was determined that the number of flowers increased considerably. Accordingly, the fruit setting rate increased.

ANALYSIS AND IDENTIFICATION OF MICROBIAL SPECIES ASSOCIATED WITH THE EGYPTIAN DATE FRUITS DURING POSTHARVEST STORAGE

Mahmoud M. Abo-El-Saad — 2023-01-29

Date fruits are consumed in many countries globally and are mostly loaded with a mixture of microbes. As a result, various bacterial and fungal species attack them, thus causing their spoilage at the ripening, storage, and processing stages. The present study investigates the current spoilage bacteria and fungi of twelve postharvest date fruits collected from different Egyptian geographical regions (Aswan, Siwa, and El-Wady El-Gadeed). Isolation and identification of date-borne microbes based on morphological and molecular identification were studied. The microscopic examination of infected date samples showed various fungal structures (e.g., hyphae, spores, conidial head, etc.). In addition, all fungal structures observed in the infected specimens belong to the fungus Aspergillus niger. After six days of incubation on a PDA medium, the total viable counts of fungi in twelve date varieties samples ranged from 1500 to 80000 (CFU/g). Five fungal isolates were isolated (Aspergillus niger F4, A. terreus F1, Penicillium sp. F3, Rhizopus sp. F2, and Circinella sp. F5) from the different samples. A. niger F4 (GenBank accession number MW811390) was the most abundant in all samples. Similarly, the total viable bacteria counts in twelve date varieties samples ranged from 4500 to 75000 (CFU/g) after 48 h incubation on nutrient agar (NA) medium. Four morphologically different bacterial phenotypes (B1, B2, B3, and B4) were isolated and purified on the NA plates. Pseudomonas aeruginosa B1 (GenBank accession number MW811391) was the most abundant bacterial isolate in all samples. Aflatoxins were also analyzed using HPLC/MS/MS in all date varieties. The results showed that aflatoxins G2, G1, and B2 were not detected in all date varieties. However, aflatoxin B1 was detected in two varieties, Partamoda and Frehi.

THE EFFECT OF ENCEPHALITOZOON INTESTINALIS ON OXIDATIVE STRESS AND CYTOKINE LEVELS IN U937 CELLS: AN IN VITRO STUDY

Meryem Şentürk — 2023-01-29

This in vitro study was performed to investigate the changes in nitric oxide (NO), malondialdehyde (MDA), total antioxidant capacity (TAC), tumor necrosis factor-α (TNF-α) and interleukin-10 (IL-10) levels in human monocytic (U937) cells infected with Encephalitozoon intestinalis. E. intestinalis was first cultured in the African green monkey renal epithelial (Vero) cells to obtain sufficient amount of parasite for the study. U937 human macrophage cell line and E. intestinalis 50506 (ATCC) strain were used in the study. U937 macrophages were infected with 5x10⁶ spores after stimulation with Phorbol 12-Myristate 13-Acetate (PMA) and uninfected U937 cells were used as control. Culture media were collected at the 6^th, 12^th, and 24^th hour after infection to determine NO, MDA, TAC, TNF-α and IL-10 levels. NO levels significantly increased at the 6^th, 12^th and 24^th hour. There was no significant difference in MDA and TAC levels at the 6^th and 12^th hour, but a significant increase in MDA, and a significant decrease in TAC was observed at the 24^th hour. TNF-α levels did not differ in all sampling times, and IL-10 level decreased only at the 6^th hour. In conclusion, E. intestinalis caused oxidative stress by increasing the levels of oxidants (NO and MDA) and by suppressing TAC level without no significant changes in cytokine levels in U937 cell line.

NEUTROPHIL-INFILTRATING REGULATORY POTENTIAL OF Lactobacillus pentosus 124-2 IN ACUTE INFLAMMATORY MODEL

Babayemi Olawale Oladejo — 2023-01-29

Neutrophils are often responsible for pathological inflammation, as well as other immunological responses. Controlling their activity is crucial to maintaining host integrity, particularly preventing against chronic inflammation. Inflammatory oedema was induced in paw of male Wistar rats using 1% iota-carrageenan. Control group was neither induced nor treated with lactic acid bacteria (LAB), while negative control group was induced without treatment. Experimental group was orogastrically treated with Lactobacillus pentosus 124-2 (5x10⁷ CFU/mL), while positive control group received diclofenac sodium treatment (150 mg/kg body weight) following carrageenan administration. Biochemical assays including myeloperoxidase (MPO), malondialdehyde (MDA), and nitric oxide (NO) were performed on the blood samples at t=1, 4, 24, and 72 hours, and tissue histology was done using standard methods to monitor neutrophil infiltration. LAB treated group showed marked decrease in NO and MPO activity at 4 hours following inducement as well as increase in MDA activity till 72 hours (p<0.05). Neutrophil infiltration was markedly controlled in the LAB treated group between 4 to 72 hours and compared favorably with the diclofenac sodium. This inflammatory model established that administered L. pentosus 124-2 inhibited neutrophil infiltration and activation resulting in the significant MPO reduction, while also regulating NO production.

IN VITRO EFFICACY OF FIVE ESSENTIAL OILS ON THE MYCELIAL GROWTH OF ASPERGILLUS NIGER ISOLATES FROM ONION BULBS

Abdou Rasmane Ouedraogo — 2023-01-29

Onion is a vegetable produced in practically all regions of Burkina Faso. Many efforts are being made to increase national production, but in storage, the loss due to black rot limits the disponibility of local bulbs during the year. The essential oils of Ocimum gratissimum, Ocimum basilicum, Cymbopogon citratus, Cymbopogon Gigantes, and Lippia multiflora, known to have antifungal properties, were added to a PDA medium and assessed in vitro at 500, 1000, 1500 and 2000 ppm concentrations on mycelial growth of isolates of Aspergillus niger. After the 4- and 7-day incubation, there was a highly significant difference between the essential oils at concentrations of 500 and 1000 ppm (p<0.001), and Ocimum gratissimum is considered the best essential oil with an inhibition percentage varying between 47.3 and 89.5%. However, all essential oils showed 100% total inhibition at 1500 ppm and the essential oils of Ocimum gratissimum and Cymbopogon giganteus demonstrated fungicidal effects at concentrations of 1500 and 2000 ppm, respectively. The intense antifungal activity of Ocimum gratissimum oil is related to the presence of thymol, a potent antioxidant responsible for the fungal activity. The activity of Ocimum gratissimum oil must also be tested in vivo to develop biopesticide formulations for the control of onion spoilage fungi.

CYTOTOXIC, APOPTOTIC AND NECROTIC EFFECTS OF INULA VISCOSA: AN IN VITRO STUDY ON DIFFERENT CELL LINES

Abdullah Melekoglu — 2023-01-29

This study aimed to evaluate the cytotoxic, apoptotic and necrotic effects of Inula viscosa on mouse fibroblast cell line (L929), human breast cancer cell line (MCF7) and human lung adenocarcinoma cell line (A549). The WST-1 (4-[3-(4-Iodophenyl) -2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate, BioVision) test was used to determine cytotoxicity. Apoptotic and necrotic rates were determined by double staining technique, Propidium iodide and Hoechst 33342.The water extract of Inula viscosa extract caused cytotoxicity in a dose dependent manner on L929, MCF7, and A549 cell lines with IC₅₀ values of 0.217 mg/ml, 0.102 mg/ml, and 0.103 mg/ml, respectively. At all doses, the cell viability of L929 fibroblast cell line was significantly the highest compared to MCF7 and A549 cell lines (p= 0.0001), except for doses of 0.5mg/ml and 0.015625mg/ml. Both apoptotic and necrotic rates changed in a dose dependent manner in all three cell lines and cell death occurred through apoptosis, but mainly necrosis. Except in few concentrations, MCF7 cell line seems to be the most sensitive to Inula viscosa extract with the highest apoptotic and necrotic rates (p=0.0001) while L929 fibroblasts seem to the least sensitive with the least apoptotic and necrotic rates (p=0.0001). Further studies should focus on chemical constituents of Inula viscosa raised in different region of the territory and additional mechanistic studies to reveal exact mechanism of cell death that may provide additional treatment modalities in cancer cases.