1.    Introduction

A substance's antimicrobial activity refers to its capacity to eradicate or prevent the development of microorganisms. Health care, food production, and environmental cleanliness are just a few of the scenarios where antimicrobials can be used [1-4]. Antimicrobials come in a variety of forms, such as antibiotics, antiseptics, disinfectants, and sanitizers [5–7]. Antibiotics are used to treat infections caused by bacteria. To treat wounds and stop infection, antiseptics are utilized. To eliminate bacteria from inanimate surfaces, disinfectants are utilized. Sanitizers are used to lower the number of bacteria on inanimate surfaces to a safe level [8].

Due primarily to the rise of microbial resistance, many antimicrobial medicines have lost their efficacy in treating infectious diseases, creating serious problems for today's healthcare systems around the globe. It is highly beneficial to search for bioactive chemicals capable of treating pathogenic bacteria that have developed resistance to current medications. There is rising interest in discovering novel antimicrobial agents in the environment, particularly in bacteria, fungi, and plants. The majority of novel medicine compounds come from natural products, especially those derived from microorganisms and plants [9–12].

Mushrooms produce several classes of bioactive chemicals, including terpenoids, flavonoids, tannins, alkaloids, and polysaccharides. Mushrooms are an abundant and, for the most part, unexplored source of bioactive chemicals [13,14]. Several cell components and secondary metabolites have been extracted and identified from the fruiting bodies, and all of them contain bioactive chemicals. Mushrooms, both their fruiting bodies and their mycelium, provide health benefits such as immunostimulatory, antimicrobial, and antioxidative qualities. Potential therapeutic values could be achieved through the synergistic impact of these drugs [15–17]. The importance of mushrooms in the lives of the tribal people of Chhattisgarh is evident from the fact that they have a rich knowledge of mushrooms and their uses. They have developed traditional methods of collecting, processing, and using mushrooms. This knowledge is passed down from generation to generation. The tribal people of Chhattisgarh are also at the forefront of research on mushrooms. They are working with scientists to develop new ways to use mushrooms for food, medicine, and income [18]. In this panorama, we have collected three medicinally important mushrooms from the Achanakmar Biosphere Reserve area to evaluate their antimicrobial activity. Brief reviews of the selected species are presented in the Table 1.

2.    Literature Review

Mushrooms are rich in bioactive compounds such as terpenoids, flavonoids, tannins, alkaloids, and polysaccharides, which exhibit antimicrobial, immunostimulatory, and antioxidant properties [13,14]. These compounds are increasingly explored as alternatives to synthetic antibiotics due to rising drug resistance.

  Inonotus obliquus (Chaga mushroom)

Traditionally used for gastrointestinal disorders, inflammation, and tuberculosis [19]. Ethanol and aqueous extracts have shown strong antibacterial activity against Pseudomonas aeruginosa.

Pleurotus ostreatus (Oyster mushroom)

Widely cultivated and consumed; recognized for nutritional and medicinal properties [20]. Exhibits antibacterial and antioxidant activity; supplementation of substrates enhances bioactive compounds [7]. Reported inhibitory effects against Staphylococcus aureus and Candida albicans [2].

Tremetes versicolor (Turkey tail mushroom)

·         Known for immune system activation and detoxification [21]. Methanol and ethanol extracts demonstrated antibacterial activity against Staphylococcus aureus and Salmonella Enteritidis [3, 4]. Also studied for antifungal activity and potential anticancer properties [22].

Global context of mushroom research

Mushrooms are considered promising therapeutic resources, with potential applications in clinical medicine [23,24]. Secondary metabolites from fungi are abundant and diverse, making them strong candidates for drug discovery [12]. Ethnobotanical knowledge, especially from tribal communities, provides valuable insights into traditional uses and guides modern research [18].

3. Material and Methods

3.1   Sample collection and identification

The Achanakmar Biosphere Reserve in Chhattisgarh, India, provided the three wild mushrooms that were harvested. Their identification was made by comparing their physical traits to those in relevant literature from the Department of Botany of the Dr. C. V. Raman University, Kota, Bilaspur, Chhattisgarh, India.

3.2 Preparation of Extract

Using the maceration process and solvents with deionized water, mushrooms were extracted [15,16]. In this case, 1 g of powdered mushroom was combined with 10 ml of water solvent and shaken for 72 hours using an incubator shaker at 150 rpm. The extracts were centrifuged at 3000 rpm for 15 minutes, filtered using Whatman No. 1 filter paper, and then dried and evaporated using a rotary evaporator at 50 °C. The extracts were dried by freeze-drying and stored in a deep freezer set at -80°C. They were then placed in an amber-colored vial and placed in the refrigerator at 4 °C for later examination.

3.3  Antimicrobial assay

3.3.1 Test organism preparation

As test organisms, we selected to use one gram-negative strain (Pseudomonas aeruginosa, MTCC 3541), one gram-positive strain (Staphylococcus aureus, MTCC 96), and one fungal strain (Candida albicans, MTCC 854).

Table 1. A brief review on the Folk practices of selected mushrooms

3.4  Antibacterial activity

The zone inhibition method, also known as the Kirby-Bauer method described [25,26] was utilized in order to evaluate the antibacterial activity. The Mueller-Hilton Agar (MHA) plates were inoculated by distributing 100 μl of bacterial cultures, (adjusted to 0.5 McFarland units, approximately 1.5 x 10⁸ CFU/mL), and then adding discs containing 10 μl of different concentrations, ranging from 0 to 50 mg/ml for P. aeruginosa and 0 to 100mg/ml for Staphylococcus aureus. The plates were then incubated at 37 degrees Celsius for 24 hours. Each disk in every plate had been loaded with just the solvent, which served as the vehicle control (DMSO). The other disc in each plate contained 10 micrograms of ciprofloxacin, which was used as the positive control. Bacterial cultural were grown on plates that were heated to 37 degrees Celsius for a period of 24 hours.

3.5  Antifungal activity

The Zone Inhibition Method Kirby-Bauer method was used to assess the antifungal activity. The Sabouraud dextrose agar (SDA) plates were inoculated by spreading 100 µl of C. albicans fungal culture (adjusted to 0.5 McFarland units) and then adding the discs containing 10 µl of various concentrations (0 to 500 mg/ml). As a vehicle control, one disc in each plate was loaded solely with solvent, and Amphotericin B discs (50 g) were used as a positive control. C. albicans plates were incubated for a full day at 37 °C.

3.6  Statistical Analysis

The distance between distinct zones that had been constructed around the disc was measured and recorded in the instance of testing the antibacterial potency of extracts. The data were reported as the mean with the standard error for each experiment, which was carried out in triplicate. SPSS was used to analyze the data, and MS Excel was used to make the graphs.

4  . Results and Discussion

All of the mushrooms employed in this investigation were discovered to have antagonistic effects on the tested microbes to varying degrees (Fig. 1). The distinct zone of inhibition that the bacteria and fungi around the tested mushroom extracts produced served as proof of this. Inonotus obliquus and Tremetes versicolor demonstrated the greatest in vitro antibacterial activity (9.67±0.57 mm and 9.00±0.57 mm) against P. aeruginosa (Table 2, Fig. 1). Pleurotus ostreatus, with a 7.67±0.57 mm zone of inhibition, was next in line. From the ethanol and water extracts of Inonotus obliquus, Pleurotus ostreatus, and Tremetes versicolor, respectively, (1,22) achieved comparable data in the case of P. aeruginosa. In the case of Staphylococcus aureus, Tremetes versicolor and Pleurotus ostreatus generated inhibitory zones measuring 7.67±0.57 mm and 7.33±1.16 mm, respectively (Fig. 1). Similar findings were also reported for Tremetes versicolor and Pleurotus ostreatus, respectively. The lowest zone of inhibition (6.50±0.71 mm) is found in the Inonotus obliquus extract, which is consistent with research [27]. The aqueous extracts of Inonotus obliquus and Tremetes versicolor were shown to have zones of inhibition for Candida albicans of 8.00±0.00 mm from the same concentration (312.5 µg/disc), which are statistically superior to the extract of Pleurotus ostreatus (7.33 mm, Fig. 1).

Fig. 1. Antimicrobial activity of three selected mushrooms in against of three microbs (ABC for Pseudomonas aeruginosa; DEF for Staphylococcus aureus; and GHI for Candida albicans): ADG for Tremetes versicolor; BEH for Pleurotus ostreatus; CFI for Inonotus obliquus.

The prevalence of multidrug-resistant organisms is currently on the rise, which poses a threat to the efficacy of treatment for an expanding variety of infectious diseases. As a direct result of this, there is an immediate need for the discovery of new medications that are effective against the bacteria that are already antibiotic-resistant [28]. It has been demonstrated that different species of fungi are excellent potential sources of bioactive chemicals that have significant medicinal effects. In addition to that, they are the most abundant sources of secondary metabolites. The three different types of mushrooms that are discussed in this article have demonstrated potentially useful antimicrobial properties against the organisms that were tested.

Table 2. Antimicrobial activity of aqueous extracts of selected three wild mushrooms collected from Achanakmar Biosphere region