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Full Research Article

An Evaluation of Alternative Mushroom Extract: Chemical Composition, Phytonutrient Analysis and Antibacterial Activity of Mastitis Pathogens 

M
Maharach Matra1,*
0000-0003-2865-1836
B
Burarat Phesatcha2
0000-0001-6137-1372
1Division of Animal Science, Department of Agricultural Technology, Faculty of Technology, Mahasarakham University, Maha Sarakham 44150, Thailand.
2Department of Applied Biology, Faculty of Sciences and Liberal Arts, Rajamangala University of Technology Isan, Nakhon Ratchasima 30000, Thailand.

ABSTRACT

Background: Mastitis is an infection caused by pathogenic bacteria that often results in substantial damage to milk production in dairy cows. They are mostly caused by strains of Escherichia coli and Staphylococcus aureus. Bolete mushrooms are recognized as an essential source of phytonutrient compounds (PCs). They can act as natural antibacterial agents to inhibit pathogenic bacteria, especially those associated with mastitis.

Methods: The purpose of this study was to examine the nutritional composition of bolete mushroom extract (BME) and investigate its possible impacts on inhibition zones and antibacterial activity against S. aureus and E. coli. The microdilution method was implemented to evaluate the antibacterial activity.

Result: The findings indicate that PCs of total phenolic compounds (3.2 mg GAE/g DM) and total flavonoid compounds (0.2 mg QUE/g DM) were present in BME. The minimum bactericidal concentration (MBC), minimum inhibitory concentration (MIC) and inhibition zones were measured to evaluate antibacterial activity. These parameters were determined using spread plate assays. BME had the most effective antibacterial activity, specifically against E. coli and S. aureus, with the highest inhibition zones of 29.5 and 29.0 mm, respectively. The MIC value of E. coli and S. aureus was 25.0 and 12.5 mg/mL and the MBC value was 50.0 and 25.0 mg/mL treated by BME. In conclusion, BME demonstrates potential as a natural antibacterial agent. It offers a workable substitute for reducing antibiotic resistance and preventing bacterial infections.

INTRODUCTION

The production of milk is the defining characteristic of dairy farming, an agricultural sector. They function as a fundamental ingredient for dairy products (Britt et al., 2018). Mastitis is a disease of economic significance that consistently poses a challenge to the production sector (Paramasivam et al., 2023). One of the reproductive-related diseases of dairy cows is mastitis, a multifaceted inflammatory disease of the mammary glands (Haxhiaj et al., 2022). This disease is a highly intricate condition that is primarily caused by Staphylococcus aureus and Escherichia coli, among other bacterial pathogens (Alonso et al., 2020; Wang et al., 2026). Bacterial infection in mastitis instances may cause alveolar cell-integrity breakdown, sloughing, triggered apoptosis and an increase in poorly differentiated cells. Ruminant mammary glands’ milk production depends on the number and activity of milk-secreting cells, which can be influenced by inflammation levels (Wahyu and Sambodho, 2020). The primary therapy for mastitis often involves intramammary infusion of an intramuscular or intravenous administration of antibiotics (Tepeli, 2020). Currently, the field of antibacterial therapy has seen an increase in the use of natural compounds over antibiotics, as these alternatives offer enhanced safety for both consumers and animals (Boldbaatar et al., 2014). This investigation presents novel research evidence supporting the use of mushrooms as a natural and sustainable biotherapeutic substitute for traditional intramammary antibiotic therapies.
       
Various mushrooms contain various PC, especially phenolics and flavonoids. These compounds are secondary metabolites that play a role in the prevention and treatment of chronic diseases (Sánchez, 2017). One of the alternative mushrooms is the bolete mushroom (Thaeogyroporus porentosus), a significant species that has been researched for its beneficial PCs and is widely found in Thailand (Baowthongkum and Promboon, 2014). PCs are a predominantly underutilized resource for discovering novel antibacterial agents with distinct mechanisms of action (Newman and Cragg, 2020). Comprehensive studies have demonstrated the antibacterial efficacy of PCs against multidrug-resistant bacterial strains, emphasizing their potential as replacements or supplements to traditional antibiotics (Cushnie and Lamb, 2011). Accordingly, Arbab et al., (2022) demonstrated that the antibacterial susceptibility test of lemon balm extract demonstrated inhibition of E. coli and S. aureus. However, there has been no research conducted on the antibacterial activity of the PC-rich bolete mushroom extract, particularly in relation to the two primary species, E. coli and S. aureus.
       
Thus, the objective of this article was to investigate the antibacterial activity of PC-rich bolete mushroom extract against E. coli and S. aureus, alongside establishing nutritional properties as a potential natural substrate for therapeutic application.
 

MATERIALS AND METHODS

Duration and site of research
 
This study was conducted between January and March 2026 at the Division of Animal Science, Faculty of Technology, Mahasarakham University, Thailand.
 
Sample preparation and chemical analysis
 
Fresh bolete mushroom was dehydrated at 60°C for 24 h to produce a dried sample, which was then ground through a 1 mm screen to achieve a powdered form. For the BME preparation, 20 g of powdered bolete mushrooms were extracted three times using a microwave-assisted extraction technique. The aqueous solution was passed through a Buchner funnel lined with Whatman No. 1 filter paper and then evaporated to extract the dry powder by using Freeze dryers (Labconco Corp., MO, USA) (Kanlayavattanakul et al., 2009).
       
The chemical evaluations of BME were conducted in accordance with AOAC (2012) and Van Soest  et al. (1991). Furthermore, the phytonutrients have been evaluated in BME. I. The modified Folin-Ciocalteu method was used to measure the total phenolic compounds (TPC) (Laosirisathian et al., 2020). II. A Two-fold serial reduction of standard quercetin in ethanol was used to create the standard calibration curve for measuring the total flavonoid compounds (TFC) (Sembiring et al., 2018).
 
Agar well diffusion technique
 
Preparation of BME for the stock testing by weighing (500 mg/mL) and dissolving extracts using 10% ethanol solvents. Antibacterial activity test against pathogenic bacteria using the agar well diffusion method (Bagul and Sivakumar, 2016; Balouiri et al., 2016). The pathogenic bacteria used in the test were S. aureus and E. coli. The test was performed by growing pathogenic bacteria in Mueller Hinton broth. Then the cultures were incubated for 16 h and by comparing the cultures with the McFarland standard #0.5, the concentration was adjusted to about 108 CFU/mL. Mueller Hinton agar was used to disseminate the bacteria. In order to create wells in the agar plate, a 6 mm diameter cork borer was utilized. 40 µL of each extract, which had a concentration of 500 mg/mL, was added to each well. Plates of agar were subjected to incubation at 37°C for 18 h. The diameter of the clear zone was measured in mm from two BME replicates for assessment. There was compared with a 10% ethanol solution as a negative control and a positive standard antibiotic using 25 mg/mL of norfloxacin.
 
Antibacterial activity
 
The MIC and MBC (measured in mg/mL) were determined by the microdilution technique using a 96-well microplate, as described by De Silva  et al. (2023). The pathogenic bacteria utilized were E. coli and S. aureus. All tested microbes were cultured in TSB and TSA media, incubated at 37°C for 16 h, achieving a final concentration of 108 CFU/mL. A microplate was utilized, containing 100 µL of filtered samples from BME or norfloxacin, employing a two-fold dilution technique for each sample. Approximately 20 µL of bacterial suspension were added, followed by 80 µL of media broth. The microplate was subjected to incubation at 37°C for a period of 24 h and bacterial growth was assessed on TSA media using the drop plate technique, with incubation under identical conditions. Following incubation, the growth of microorganisms on culture plates was assessed by the MIC, defined as the lowest level of the extract that inhibits visible growth of microbes after an overnight culture. The MBC value is defined as the minimum concentration of the extract capable of eliminating microorganisms, resulting in no growth on the culture agar media, as outlined by Balouiri et al., (2016). 
 
Statistical analysis
 
Descriptive statistics, such as percentage, standard deviation and mean, are used to analyze the study’s data. The program of Microsoft Excel (Microsoft Office LTSC Professional Plus 2021) was employed to indicate the table.

RESULTS AND DISCUSSION

Nutritional composition of bolete mushroom extract             
The proximate analysis of BME, which includes 99.4% of dry matter, 87.8% DM of organic matter, 2.1% DM of crude protein, 0.2% DM of neutral-detergent fiber and 0.1% DM of acid-detergent fiber, respectively. Importantly, BME contain PCs, particularly 3.2 mg GAE/g DM of TPC and 0.2 mg QUE/g DM of TFC, as provided in Table 1.

Table 1: Chemical composition and phytonutrient compounds of bolete mushroom extract.


 
Bacterial inhibition zone
 
The inhibition zone of BME at 500 mg/mL against E. coli and S. aureus. This is compared to an antibiotic (norfloxacin) as a positive control and a 10% ethanol solution as a negative control using the agar well diffusion technique, showing a clear zone surrounding wells. The averages of the inhibition zone against E. coli and S. aureus were found in the range of 29.5-30.5 and 29.0-29.5 mm obtained from BME and norfloxacin, respectively (Fig 1 and Table 2).

Fig 1: Bacterial inhibition zones (mm) of bolete mushroom extracts in comparison to norfloxacin and 10% ethanol for the assessment of E. coli and S. aureus.



Table 2: Determination of diameter inhibition zone using bolete mushroom extract.


 
Antibacterial activity
 
The determination of the MIC and MBC of BME demonstrated inhibitory effects against E. coli and S. aureus. Especially, S. aureus had a high sensitivity, which showed the MBC value at 25.0 mg/mL and the MIC value of 12.5 mg/mL treated by BME, whilst the norfloxacin showed at 6.3 and 12.5 mg/mL, as shown in Table 3.

Table 3: Determination of antibacterial activity parameters using bolete mushroom extract.


       
Bovine mastitis is an infection of the mammary gland caused by microbes. Most mastitis is caused by opportunistic gram-negative bacterium E. coli. Due to immunosuppression during parturition and early lactation, cattle can develop intramammary infections. Infections can cause severe clinical mastitis and recurrence, but they usually last less than other pathogens and rarely require treatment (Blowey and Edmondson, 2010). Furthermore, clinical mastitis is primarily caused by S. aureus. This bacterium is a prevalent commensal in the nares and skin, but it can also function as an opportunistic pathogen, resulting in both superficial and invasive infections in ruminants (Foster and Geoghegan, 2015; Ali et al., 2026). The bacterial isolation findings indicated that S. aureus was present in 97.7% of cases, whereas E. coli was positive in 53.5% of instances. The impact of these two bacteria on mastitis formation was shown to be considerable in the examined cows (Arbab et al., 2022).
       
The current study enhances the data concerning the antimicrobial activity of mastitis pathogens derived from BME. The assessment of the BME revealed the inhibition of E. coli and S. aureus. This may be related to BME containing the PC particularly phenolics and flavonoids that can influence microbial pathology. These findings align with the research conducted by Lee et al., (2021); Laklai et al., (2025) and Hussien et al., (2026), which reported that various plant-based PC, exhibits promise as a natural antibacterial agent, particularly against E. coli and S. aureus. This presents a viable option for treating bacterial infections and mitigating antibiotic resistance. Importantly, phenolics and flavonoids eliminate pathogenic bacteria in two ways: i. by directly destroying them and damaging their cell membranes and ii. by indirectly influencing the host’s immune system and enhancing the gut ecosystem to prevent pathogen colonization (Wlodarska et al., 2015).
       
Nonetheless, it is crucial to acknowledge that the current research was performed under in vitro procedures. Despite BME exhibiting encouraging antibacterial activity against E. coli and S. aureus, these results may not accurately represent its efficacy under in vivo conditions. Consequently, additional in vivo investigations are necessary to assess the safety, appropriate concentration and possible therapeutic uses of BME in dairy cows suffering from mastitis. Such investigations are essential to confirm its potential as a natural alternative for the prevention and treatment of bovine mastitis.

CONCLUSION

The antibacterial activity of BME inhibited the mastitis-related pathogens E. coli and S. aureus. The extract exhibited robust bactericidal and inhibitory effects, as evidenced by the presence of large inhibition zones and the low MIC and MBC values. The results demonstrate that BME possesses considerable potential as a natural antibacterial agent for mastitis treatment. It has the potential to offer a complementary or alternative therapeutic approach to conventional antibiotics, potentially reducing the overuse of antibiotics and addressing the issue of antimicrobial resistance.

ACKNOWLEDGEMENT

This research project was financially supported by Mahasarakham University.
 
Disclaimers
 
The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
 
Informed consent
 
This study does not involve the use of animals.

CONFLICT OF INTEREST

The authors declare that there are no conflicts of interest regarding the publication of this article. No funding or sponsorship influenced the design of the study, data collection, analysis, decision to publish, or preparation of the manuscript.

REFERENCES


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    Full Research Article

    An Evaluation of Alternative Mushroom Extract: Chemical Composition, Phytonutrient Analysis and Antibacterial Activity of Mastitis Pathogens 

    M
    Maharach Matra1,*
    0000-0003-2865-1836
    B
    Burarat Phesatcha2
    0000-0001-6137-1372
    1Division of Animal Science, Department of Agricultural Technology, Faculty of Technology, Mahasarakham University, Maha Sarakham 44150, Thailand.
    2Department of Applied Biology, Faculty of Sciences and Liberal Arts, Rajamangala University of Technology Isan, Nakhon Ratchasima 30000, Thailand.

    ABSTRACT

    Background: Mastitis is an infection caused by pathogenic bacteria that often results in substantial damage to milk production in dairy cows. They are mostly caused by strains of Escherichia coli and Staphylococcus aureus. Bolete mushrooms are recognized as an essential source of phytonutrient compounds (PCs). They can act as natural antibacterial agents to inhibit pathogenic bacteria, especially those associated with mastitis.

    Methods: The purpose of this study was to examine the nutritional composition of bolete mushroom extract (BME) and investigate its possible impacts on inhibition zones and antibacterial activity against S. aureus and E. coli. The microdilution method was implemented to evaluate the antibacterial activity.

    Result: The findings indicate that PCs of total phenolic compounds (3.2 mg GAE/g DM) and total flavonoid compounds (0.2 mg QUE/g DM) were present in BME. The minimum bactericidal concentration (MBC), minimum inhibitory concentration (MIC) and inhibition zones were measured to evaluate antibacterial activity. These parameters were determined using spread plate assays. BME had the most effective antibacterial activity, specifically against E. coli and S. aureus, with the highest inhibition zones of 29.5 and 29.0 mm, respectively. The MIC value of E. coli and S. aureus was 25.0 and 12.5 mg/mL and the MBC value was 50.0 and 25.0 mg/mL treated by BME. In conclusion, BME demonstrates potential as a natural antibacterial agent. It offers a workable substitute for reducing antibiotic resistance and preventing bacterial infections.

    INTRODUCTION

    The production of milk is the defining characteristic of dairy farming, an agricultural sector. They function as a fundamental ingredient for dairy products (Britt et al., 2018). Mastitis is a disease of economic significance that consistently poses a challenge to the production sector (Paramasivam et al., 2023). One of the reproductive-related diseases of dairy cows is mastitis, a multifaceted inflammatory disease of the mammary glands (Haxhiaj et al., 2022). This disease is a highly intricate condition that is primarily caused by Staphylococcus aureus and Escherichia coli, among other bacterial pathogens (Alonso et al., 2020; Wang et al., 2026). Bacterial infection in mastitis instances may cause alveolar cell-integrity breakdown, sloughing, triggered apoptosis and an increase in poorly differentiated cells. Ruminant mammary glands’ milk production depends on the number and activity of milk-secreting cells, which can be influenced by inflammation levels (Wahyu and Sambodho, 2020). The primary therapy for mastitis often involves intramammary infusion of an intramuscular or intravenous administration of antibiotics (Tepeli, 2020). Currently, the field of antibacterial therapy has seen an increase in the use of natural compounds over antibiotics, as these alternatives offer enhanced safety for both consumers and animals (Boldbaatar et al., 2014). This investigation presents novel research evidence supporting the use of mushrooms as a natural and sustainable biotherapeutic substitute for traditional intramammary antibiotic therapies.
           
    Various mushrooms contain various PC, especially phenolics and flavonoids. These compounds are secondary metabolites that play a role in the prevention and treatment of chronic diseases (Sánchez, 2017). One of the alternative mushrooms is the bolete mushroom (Thaeogyroporus porentosus), a significant species that has been researched for its beneficial PCs and is widely found in Thailand (Baowthongkum and Promboon, 2014). PCs are a predominantly underutilized resource for discovering novel antibacterial agents with distinct mechanisms of action (Newman and Cragg, 2020). Comprehensive studies have demonstrated the antibacterial efficacy of PCs against multidrug-resistant bacterial strains, emphasizing their potential as replacements or supplements to traditional antibiotics (Cushnie and Lamb, 2011). Accordingly, Arbab et al., (2022) demonstrated that the antibacterial susceptibility test of lemon balm extract demonstrated inhibition of E. coli and S. aureus. However, there has been no research conducted on the antibacterial activity of the PC-rich bolete mushroom extract, particularly in relation to the two primary species, E. coli and S. aureus.
           
    Thus, the objective of this article was to investigate the antibacterial activity of PC-rich bolete mushroom extract against E. coli and S. aureus, alongside establishing nutritional properties as a potential natural substrate for therapeutic application.
     

    MATERIALS AND METHODS

    Duration and site of research
     
    This study was conducted between January and March 2026 at the Division of Animal Science, Faculty of Technology, Mahasarakham University, Thailand.
     
    Sample preparation and chemical analysis
     
    Fresh bolete mushroom was dehydrated at 60°C for 24 h to produce a dried sample, which was then ground through a 1 mm screen to achieve a powdered form. For the BME preparation, 20 g of powdered bolete mushrooms were extracted three times using a microwave-assisted extraction technique. The aqueous solution was passed through a Buchner funnel lined with Whatman No. 1 filter paper and then evaporated to extract the dry powder by using Freeze dryers (Labconco Corp., MO, USA) (Kanlayavattanakul et al., 2009).
           
    The chemical evaluations of BME were conducted in accordance with AOAC (2012) and Van Soest  et al. (1991). Furthermore, the phytonutrients have been evaluated in BME. I. The modified Folin-Ciocalteu method was used to measure the total phenolic compounds (TPC) (Laosirisathian et al., 2020). II. A Two-fold serial reduction of standard quercetin in ethanol was used to create the standard calibration curve for measuring the total flavonoid compounds (TFC) (Sembiring et al., 2018).
     
    Agar well diffusion technique
     
    Preparation of BME for the stock testing by weighing (500 mg/mL) and dissolving extracts using 10% ethanol solvents. Antibacterial activity test against pathogenic bacteria using the agar well diffusion method (Bagul and Sivakumar, 2016; Balouiri et al., 2016). The pathogenic bacteria used in the test were S. aureus and E. coli. The test was performed by growing pathogenic bacteria in Mueller Hinton broth. Then the cultures were incubated for 16 h and by comparing the cultures with the McFarland standard #0.5, the concentration was adjusted to about 108 CFU/mL. Mueller Hinton agar was used to disseminate the bacteria. In order to create wells in the agar plate, a 6 mm diameter cork borer was utilized. 40 µL of each extract, which had a concentration of 500 mg/mL, was added to each well. Plates of agar were subjected to incubation at 37°C for 18 h. The diameter of the clear zone was measured in mm from two BME replicates for assessment. There was compared with a 10% ethanol solution as a negative control and a positive standard antibiotic using 25 mg/mL of norfloxacin.
     
    Antibacterial activity
     
    The MIC and MBC (measured in mg/mL) were determined by the microdilution technique using a 96-well microplate, as described by De Silva  et al. (2023). The pathogenic bacteria utilized were E. coli and S. aureus. All tested microbes were cultured in TSB and TSA media, incubated at 37°C for 16 h, achieving a final concentration of 108 CFU/mL. A microplate was utilized, containing 100 µL of filtered samples from BME or norfloxacin, employing a two-fold dilution technique for each sample. Approximately 20 µL of bacterial suspension were added, followed by 80 µL of media broth. The microplate was subjected to incubation at 37°C for a period of 24 h and bacterial growth was assessed on TSA media using the drop plate technique, with incubation under identical conditions. Following incubation, the growth of microorganisms on culture plates was assessed by the MIC, defined as the lowest level of the extract that inhibits visible growth of microbes after an overnight culture. The MBC value is defined as the minimum concentration of the extract capable of eliminating microorganisms, resulting in no growth on the culture agar media, as outlined by Balouiri et al., (2016). 
     
    Statistical analysis
     
    Descriptive statistics, such as percentage, standard deviation and mean, are used to analyze the study’s data. The program of Microsoft Excel (Microsoft Office LTSC Professional Plus 2021) was employed to indicate the table.

    RESULTS AND DISCUSSION

    Nutritional composition of bolete mushroom extract             
    The proximate analysis of BME, which includes 99.4% of dry matter, 87.8% DM of organic matter, 2.1% DM of crude protein, 0.2% DM of neutral-detergent fiber and 0.1% DM of acid-detergent fiber, respectively. Importantly, BME contain PCs, particularly 3.2 mg GAE/g DM of TPC and 0.2 mg QUE/g DM of TFC, as provided in Table 1.

    Table 1: Chemical composition and phytonutrient compounds of bolete mushroom extract.


     
    Bacterial inhibition zone
     
    The inhibition zone of BME at 500 mg/mL against E. coli and S. aureus. This is compared to an antibiotic (norfloxacin) as a positive control and a 10% ethanol solution as a negative control using the agar well diffusion technique, showing a clear zone surrounding wells. The averages of the inhibition zone against E. coli and S. aureus were found in the range of 29.5-30.5 and 29.0-29.5 mm obtained from BME and norfloxacin, respectively (Fig 1 and Table 2).

    Fig 1: Bacterial inhibition zones (mm) of bolete mushroom extracts in comparison to norfloxacin and 10% ethanol for the assessment of E. coli and S. aureus.



    Table 2: Determination of diameter inhibition zone using bolete mushroom extract.


     
    Antibacterial activity
     
    The determination of the MIC and MBC of BME demonstrated inhibitory effects against E. coli and S. aureus. Especially, S. aureus had a high sensitivity, which showed the MBC value at 25.0 mg/mL and the MIC value of 12.5 mg/mL treated by BME, whilst the norfloxacin showed at 6.3 and 12.5 mg/mL, as shown in Table 3.

    Table 3: Determination of antibacterial activity parameters using bolete mushroom extract.


           
    Bovine mastitis is an infection of the mammary gland caused by microbes. Most mastitis is caused by opportunistic gram-negative bacterium E. coli. Due to immunosuppression during parturition and early lactation, cattle can develop intramammary infections. Infections can cause severe clinical mastitis and recurrence, but they usually last less than other pathogens and rarely require treatment (Blowey and Edmondson, 2010). Furthermore, clinical mastitis is primarily caused by S. aureus. This bacterium is a prevalent commensal in the nares and skin, but it can also function as an opportunistic pathogen, resulting in both superficial and invasive infections in ruminants (Foster and Geoghegan, 2015; Ali et al., 2026). The bacterial isolation findings indicated that S. aureus was present in 97.7% of cases, whereas E. coli was positive in 53.5% of instances. The impact of these two bacteria on mastitis formation was shown to be considerable in the examined cows (Arbab et al., 2022).
           
    The current study enhances the data concerning the antimicrobial activity of mastitis pathogens derived from BME. The assessment of the BME revealed the inhibition of E. coli and S. aureus. This may be related to BME containing the PC particularly phenolics and flavonoids that can influence microbial pathology. These findings align with the research conducted by Lee et al., (2021); Laklai et al., (2025) and Hussien et al., (2026), which reported that various plant-based PC, exhibits promise as a natural antibacterial agent, particularly against E. coli and S. aureus. This presents a viable option for treating bacterial infections and mitigating antibiotic resistance. Importantly, phenolics and flavonoids eliminate pathogenic bacteria in two ways: i. by directly destroying them and damaging their cell membranes and ii. by indirectly influencing the host’s immune system and enhancing the gut ecosystem to prevent pathogen colonization (Wlodarska et al., 2015).
           
    Nonetheless, it is crucial to acknowledge that the current research was performed under in vitro procedures. Despite BME exhibiting encouraging antibacterial activity against E. coli and S. aureus, these results may not accurately represent its efficacy under in vivo conditions. Consequently, additional in vivo investigations are necessary to assess the safety, appropriate concentration and possible therapeutic uses of BME in dairy cows suffering from mastitis. Such investigations are essential to confirm its potential as a natural alternative for the prevention and treatment of bovine mastitis.

    CONCLUSION

    The antibacterial activity of BME inhibited the mastitis-related pathogens E. coli and S. aureus. The extract exhibited robust bactericidal and inhibitory effects, as evidenced by the presence of large inhibition zones and the low MIC and MBC values. The results demonstrate that BME possesses considerable potential as a natural antibacterial agent for mastitis treatment. It has the potential to offer a complementary or alternative therapeutic approach to conventional antibiotics, potentially reducing the overuse of antibiotics and addressing the issue of antimicrobial resistance.

    ACKNOWLEDGEMENT

    This research project was financially supported by Mahasarakham University.
     
    Disclaimers
     
    The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
     
    Informed consent
     
    This study does not involve the use of animals.

    CONFLICT OF INTEREST

    The authors declare that there are no conflicts of interest regarding the publication of this article. No funding or sponsorship influenced the design of the study, data collection, analysis, decision to publish, or preparation of the manuscript.

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