Exploring Homoeopathic Potential Against Typhoid: Baptisia tinctoria, Rhus toxicodendron, and Eupatorium perfoliatum in growth Inhibition of Salmonella typhi – In-vitro study

Exploring Homoeopathic Potential Against Typhoid: Baptisia tinctoria, Rhus toxicodendron, and Eupatorium perfoliatum in growth Inhibition of Salmonella typhi – In-vitro study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Exploring Homoeopathic Potential Against Typhoid: Baptisia tinctoria, Rhus toxicodendron, and Eupatorium perfoliatum in growth Inhibition of Salmonella typhi – In-vitro study P Bhuvanesh, K. Narayanasamy, G Sridevi, G Senthil Kumaran, J P Jerin This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4432701/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 3 You are reading this latest preprint version Abstract Typhoid fever, caused by Salmonella typhi, is a global health issue with high morbidity and mortality rates, especially in low-income regions. The rise of antibiotic-resistant strains necessitates new treatment strategies. This study explores the potential of homoeopathic medicines, known for their safety and non-induction of resistance, as an alternative treatment. Three homoeopathic medicines, Rhus toxicodendron, Eupatorium perfoliatum, and Baptisia tinctoria, were tested for their inhibitory effects on S. typhi using the agar disc diffusion method. The medicines, along with a control antibiotic, ceftriaxone, were applied to agar plates with S. typhi cultures. All tested homoeopathic medicines showed inhibitory effects against S. typhi. Rhus toxicodendron Q and Baptisia tinctoria 200 CH showed significant inhibition zones of 16 mm and 14 mm, respectively. Baptisia tinctoria 30 CH and Eupatorium perfoliatum 30 CH also showed inhibitory effects, while ceftriaxone exhibited the largest inhibition zone. This study highlights the need for innovative treatments against antibiotic-resistant infections. It confirms the antimicrobial potential of homoeopathic medicines against S. typhi, suggesting a possible solution for this infectious disease. Further research is needed to assess their clinical effectiveness in managing typhoid fever. Salmonella typhi antimicrobial activity agar disc diffusion method Baptisia tinctoria Rhus toxicodendron Eupatorium perfoliatum Typhoid fever Figures Figure 1 Figure 2 Figure 3 Figure 4 INTRODUCTION As per the World Health Organization (WHO), typhoid fever continues to impact between 11 to 20 million individuals annually and is accountable for 128,000 to 161,000 deaths each year. In urban locations of India, the rate of culture-confirmed typhoid fever was between 576 to 1173 cases per 100,000 child-years, whereas in rural areas, it was 35 cases [2] . When analyzing data from hospitals, the estimated occurrence ranged from 12 to 1622 cases for individuals aged 6 months to 14 years, and from 108 to 970 cases for those aged 15 years and older. The incidence rate for paratyphoid fever was 68 cases per 100,000 child-years. Typhoid fever, caused by the bacterium Salmonella typhi , is transmitted through person-to-person contact via contaminated food and water [8] . This transmission transpires through the fecal-oral route, implying that contaminated feces (and sometimes urine) may infiltrate water or food supplies, subsequently consumed by others causing infection. While infrequent in industrialized nations, it poses a significant threat in certain low-income countries [1, 16] . The symptoms of typhoid fever range from mild to severe, generally appearing one to three weeks after exposure to the bacteria. These symptoms encompass fever, headache, nausea, constipation or diarrhea, loss of appetite, and the presence of a rose-colored rash on the body. Symptoms of typhoid fever bear resemblance to other prevalent gastrointestinal illnesses. The only definitive method to ascertain a person's affliction with typhoid is by testing their blood or feces for Salmonella typhi . Salmonella typhi exclusively thrives in humans, with no animal reservoir. [8] Complications arising from typhoid fever encompass intestinal hemorrhage, intestinal perforation, encephalitis, metastatic abscess, and others. Paratyphoid and Typhoid conjugate vaccines are authorized in India. For the past 60-70 years, antibiotics have remained the sole mode of treatment since the discovery of Salmonella. Consequently, drug-resistant organisms have emerged, homoeopathic medicines do not cause drug resistance even when administered for long time. Hence our proposal aims to assess the In-vitro Efficacy of Homoeopathic Medicine against Salmonella typhi . The homeopathic remedies employed in this study encompass Baptisia tinctoria , Eupatorium perfoliatum , and Rhus toxicodendron in mother tincture, as well as potencies of 30 CH and 200 CH. OBJECTIVE: To determine the inhibitory effect of the various homoeopathic drugs against Salmonella typhi confirming the antimicrobial activity of potentized drugs in higher dilutions. MATERIALS AND METHODOLOGY Medicines: [4] As per Homoeopathic pharmacopoeia of India, standardised homoeopathic Medicines Baptisia tinctoria , Rhus toxicodendron and Eupatorium perfoliatum in different potencies like Q, 30 CH and 200 CH along with Ethanol of 90% v/v were procured from GMP certified Drug Manufacturing unit of Dr.Willmar Schwabe, India. Microorganism: The bacterial culture of S.Typhi (MTCC No: 3231) procured as lyophilized freeze-dried culture strain from the Microbial Type Culture Collection, Institute of Microbial Technology, Chandigarh, was used to evaluate the antimicrobial activity of homoeopathic drugs. Preparation of Agar medium: [8] The agar medium was prepared by dissolving 4.95 g of MacConkey agar in 100 ml of distilled water and dissolved completely with the help of a hot plate. Then the media was sterilised by autoclaving at 15 lb pressure (121 0 C) for 15 minutes. The sterilised agar was transferred into the petri plates (100 mm × 15 mm) and was allowed to solidify. These procedures were completed in laminar air flow under aseptic conditions. Preparation of bacterial culture: [8] The bacterial strain of Salmonella typhi (MTCC NO. 3231), lyophilized freeze-dried culture was aseptically opened in a laminar air flow and the suspension was made as per protocol. 0.5 ml of buffered peptone water was taken in the microfuge tube and freeze-dried culture was transferred into it and mixed well. The mixture was allowed for 20 minutes before it was transferred to solid media. Petri plates containing MacConkey agar medium (HiMedia, Mumbai, India, Catalogue No. M081) was incubated for 24 hours at 37 0 C to give white round colonies against a mild reddish pink background. Incubation period of Salmonella typhi is 24 hours. This culture was kept as Broth culture/stock culture. Bacterial cultures were repeatedly sub cultured using streaking methods and maintained to obtain pure isolation on the MacConkey medium for further drug sensitivity assay. Identification of Salmonella typhi: [8] Catalase Test: [8] 2 ml of hydrogen peroxide solution was taken into the test tube and by using a glass rod, samples of several colonies from the Broth Culture were immersed into the hydrogen peroxide solution. Formation of an immediate bubble confirmed the presence of Salmonella typhi . Preparation of disc for antimicrobial assay: Agar disc diffusion method was adopted for determining the antimicrobial activity of different homoeopathic medicines in various potencies. Whatman filter paper No.1 of size approximately 6 mm in diameter was used to prepare the standard discs and autoclaved at 121 0 C at 15 lbs for 15 minutes. These discs were soaked in Baptisia tinctoria , Rhus toxicodendron , Eupatorium perfoliatum under different potencies like Q, 30CH and 200CH, Ethanol of 90% v/v and Ceftriaxone for 30 to 45 minutes. after which they were taken out and allowed to dry under aseptic conditions. Determination of Antimicrobial Activity: [7, 8, 19] The antimicrobial activity of the selected homoeopathic medicines under different potencies were evaluated using the agar disc diffusion method by measuring the diameter of growth inhibition zones. Ceftriazone was used as a positive control and Ethanol of 90% v/v was used as a negative control. The discs were inverted and placed in the petri well plates in an incubator set to 37 0 C. After 24-hour growth, an inhibition zone around the discs was measured. The diameter of the growth inhibition zone around the discs was measured using a scale. Each petri well plate was divided into five compartments. Three compartments were allocated for different homoeopathic potencies and one for vehicle Ethanol (V) and one for Antibiotic (AB). RESULTS All the three homoeopathic medicine in various potencies exhibited the inhibitory action against Salmonella typhi ( Table 1 and Figure 2 ). Among these, Rhus toxicodendron Q and Baptisia tinctoria 200 CH exhibited maximum inhibition zone of diameter of 16 mm and 14 mm respectively followed by Baptisia tinctoria 30 CH and Eupatorium perfoliatum 30 CH showing the least zone of inhibition of 11 mm and 10.9 mm respectively. However the antibiotic positive control ceftriaxone showed the maximum zone of inhibition. Table 1 : Zone of Inhibition of Homoeopathic Medicines in different potencies Drugs Mother Tincture 30 CH 200 CH Control (Ceftriaxone) Vehicle (Ethanol 90% v/v) Baptisia tinctoria 12 mm 11 mm 14 mm 12 mm 7 mm Rhus toxicodendron 16 mm 12 mm 13 mm 19 mm 7.1 mm Eupatorium perfoliatum 13 mm 10.9 mm 12 mm 18 mm 6.9 mm DISCUSSION From the sources of various homoeopathic Materia Medica, Baptisia tinctoria in low dilutions produces a form of antibodies to bac.typhosus, viz, the agglutinins thus it raises the bodily natural resistances to the invasion of the bacillary intoxication , which produces the typhoid syndrome [ 4 ] . Rhus toxicodendron indicated therapeutically for typhoid fever [ 4 ] . From the sources of drug proving and clinical verification by pioneers the antibacterial activity has been recorded even before the antimicrobial era. This study reiterates the fact and scientifically confirms the antimicrobial activity of Baptisia tinctoria and Rhus toxicodendron . Eupatorium perfoliatum is a well-known antiviral remedy [ 4 ] , however it exhibits antimicrobial activities. CONCLUSION In recent days, repeated use of antibiotics in frequent intervals has caused drug resistance in patients, which poses a serious threat to the community. This resistance has pushed them to discover new generations of antibiotics year after year. Homoeopathic medicines do not cause any resistance and do not produce undesirable side-effects as it is being dispensed from low attenuation to ultra-high dilutions since the Hahnemannian era onwards. This evidence based in-vitro study contributes to the scientific and literature fact that homoeopathic medicine has definite antimicrobial activity against Salmonella typhi . Declarations CONFLICT OF INTEREST: None Author Contribution Dr.K.Narayanasamy - Conceptualization, Material method, Manuscript preparation Manuscript editing, software ,Manuscript review.Dr.P. Bhuvanesh of intellectual Concepts, investigation Design, content, Manuscript reviewG. Sridevi-content ,writing, material and methods, Supervision Senthilkumaran.G - Manuscript review, Result, Software JP.Jerin- discussion,Manuscript review, Manuscript editing Acknowledgement We thank our colleagues at The Tamilnadu Dr. MGR Medical University, Tamilnadu, India, for their support, and the reviewers of BMC Microbiology for their valuable feedback. References Adak S, Chakraborty D, Maji HS, Basu S, Roy P, Mitra S, et al. Comparison of the antimicrobial activity of the phyto-constituents obtained from the stem bark and leaf extracts of Phyllanthus emblica L. against different strains of Staphylococcus aureus and Salmonella typhi . Rese Jour Pharmacol Pharmacod. 2018;10:53. https://doi.org/10.5958/2321-5836.2018.00009.5 . admin. In Vitro Antibacterial Activity of Homoeopathy Medicines. Homeopathy Resource by HomeobookCom 2012. https://www.homeobook.com/in-vitro-antibacterial-activity-of-homoeopathy-medicines/ (accessed August 27, 2023). An in-vitro study on antibacterial activity of homoeopathic medicines-Silicea. Sulphur and Mercurius solubilis against Pseudomonas aeruginosa. Ijpr 2020;12. https://doi.org/10.31838/ijpr/2020.12.03.095 . Boericke W. New manual of homoeopathic materia medica and repertory: including Indian drugs, nosodes, uncommon remedies, relationship, sides of the body & drug affinities. Augmented ed., based on 9th ed., reprint ed. New Delhi: B. Jain Publishers; 2005. Christy S, Nivedhitha MS. Antimicrobial Efficacy of Azadirachta indica against Streptococcus mutans -An In vitro Study. Asian Jour Pharm and Technol. 2019;9:149. https://doi.org/10.5958/2231-5713.2019.00025.4 . Gnanavel S, Bharathidasan R, Panneerselvam RmahalingamPM. Antimicrobial Activity of Strychnos nux-vomica Linn and Cassia angustifolia Linn. Asian J Pharm Technol. 2012;2:08–11. Gupta G, Srivastava A, Gupta N, Gupta G, Mishra S. Anti-candidal activity of homoeopathic drugs: An in-vitro evaluation. Indian J Res Homoeopathy. 2015;9:79. https://doi.org/10.4103/0974-7168.159522 . Kanungo R. Ananthanarayan and Paniker’s Textbook of Microbiology. Universities; 2018. Khan MY, Gupta P, Singh VK, Yadav S, Verma VK. Cymbopogon Citrates Oil Showing Antimicrobial Activity against Microbes of Environmental, Clinical and Food Origin. Asian J Pharm Technol. 2013;3:67–72. Kumar JS. A Descriptive study to assess the knowledge regarding Enteric fever in children among mothers in a selected Hospital, Ludhiana, Punjab. Asian J Nurs Educ Res. 2020;10:413–8. https://doi.org/10.5958/2349-2996.2020.00088.9 . Malathi R, Cholarajan A, Karpagam K, Jaya KR, Muthukumaran P. Antimicrobial Studies on Selected Medicinal Plants (Coleus amboinicus, Phyla nodiflora and Vitex negundo). Asian J Pharm Technol. 2011;1:53–5. Maneesh K, Vijayabhaskar K, Firdouse H, Srinivasa Rao P, Prajwitha M, Swetha S. Evaluation of Antimicrobial of P. vesicularis, Streptococcus faecalis, Aeromonas hydrophilia, Salmonela typhae, Stphylococcus cohni, Serratia ficaria and E. coli. of crude and n-butanol fraction fruit latex of Carica papaya L. (Caricaceae). AJPR. 2021;92–4. https://doi.org/10.52711/2231-5691.2021.00017 . Mercado CX, Kluthe BG. Fungicidal effects of homoeopathic medicines versus allopathic ketoconazole in Candida albicans. Indian J Res Homoeopathy 2021;15. https://doi.org/10.53945/2320-7094.1027 . Mohite S, Shah R, Patel N. Antimicrobial activity of leaves extracts of Passiflora foetida . Asia Journ Resear Pharmac Scie. 2018;8:17. https://doi.org/10.5958/2231-5659.2018.00004.8 . Mohite SA, Shah RR, Patel NR. Antimicrobial Activity of Leaves extracts of Jatropha curcas . Asian Jour Pharmac Rese. 2018;8:17. https://doi.org/10.5958/2231-5691.2018.00004.7 . NEJM Journal Watch. Summaries of and commentary on original medical and scientific articles from key medical journals n.d. https://www.jwatch.org/na56020/2023/04/28/typhoid-and-paratyphoid-fever-india-burdens-remain-high (accessed August 27, 2023). Mohan PnK, Saravanan S, Dutta J. Microwave Assisted Synthesis, Characterization and Antimicrobial Activity of Some Schiff Bases of 2- Amino (4- Chloro Phenyl) Thiazoles. Asian J Res Chem. 2010;3:751–4. Pareek S, Jadhav AB. In-Vitro Evaluation of Antibacterial Activity of Homoeopathic Preparations on Klebsiella pneumoniae. Int J Health Sci Res. 2020;10:176–80. Prajapati S, Mahima S, Kumar A, Gupta P, Dwivedi B, Arya B, et al. Antimicrobial activity of different homoeopathic drugs and their potencies against Aspergillus niger In vitro. Indian J Res Homoeopathy. 2019;13:150–8. https://doi.org/10.4103/ijrh.ijrh_46_18 . Priyanka N, Brahmeswari P, Babu GR, Sowjanya M, Ramayyappa M. Evaluation of Anti-bacterial and Anti-inflammatory Activities of Ethanolic Extract of Hibiscus hirtus Linn. Leaves AJPR 2022:5–10. https://doi.org/10.52711/2231-5691.2022.00002 . Rao N, Mittal S, Sudhanshu, Menghani E. Assessment of Phytochemical Screening, Antioxidant and Antibacterial Potential of the Methanolic Extract of Ricinus communis l. Asian J Pharm Technol. 2013;3:20–5. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editor assigned by journal 06 Jun, 2024 Submission checks completed at journal 06 Jun, 2024 First submitted to journal 16 May, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4432701","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":311495015,"identity":"cfc6f750-effc-4f87-b083-dcc2c48ea3ed","order_by":0,"name":"P Bhuvanesh","email":"","orcid":"","institution":"Sri Sairam Homoeopathy Medical College and Research Center","correspondingAuthor":false,"prefix":"","firstName":"P","middleName":"","lastName":"Bhuvanesh","suffix":""},{"id":311495016,"identity":"6238ba34-ab37-46fe-9f88-0604b08f6038","order_by":1,"name":"K. Narayanasamy","email":"data:image/png;base64,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","orcid":"","institution":"The Tamilnadu Dr.M.G.R Medical University","correspondingAuthor":true,"prefix":"","firstName":"K.","middleName":"","lastName":"Narayanasamy","suffix":""},{"id":311495017,"identity":"cf50400c-f322-4d3b-b13f-5c03b6fc739f","order_by":2,"name":"G Sridevi","email":"","orcid":"","institution":"UG Scholar, Sri Sairam Homoeopathy Medical College and Research 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In urban locations of India, the rate of culture-confirmed typhoid fever was between 576 to 1173 cases per 100,000 child-years, whereas in rural areas, it was 35 cases\u003csup\u003e[2]\u003c/sup\u003e. When analyzing data from hospitals, the estimated occurrence ranged from 12 to 1622 cases for individuals aged 6 months to 14 years, and from 108 to 970 cases for those aged 15 years and older. The incidence rate for paratyphoid fever was 68 cases per 100,000 child-years.\u003c/p\u003e\n\u003cp\u003eTyphoid fever, caused by the bacterium \u003cem\u003eSalmonella typhi\u003c/em\u003e, is transmitted through person-to-person contact via contaminated food and water \u003csup\u003e[8]\u003c/sup\u003e. This transmission transpires through the fecal-oral route, implying that contaminated feces (and sometimes urine) may infiltrate water or food supplies, subsequently consumed by others causing infection. While infrequent in industrialized nations, it poses a significant threat in certain low-income countries \u003csup\u003e[1, 16]\u003c/sup\u003e. The symptoms of typhoid fever range from mild to severe, generally appearing one to three weeks after exposure to the bacteria. These symptoms encompass fever, headache, nausea, constipation or diarrhea, loss of appetite, and the presence of a rose-colored rash on the body. Symptoms of typhoid fever bear resemblance to other prevalent gastrointestinal illnesses. The only definitive method to ascertain a person\u0026apos;s affliction with typhoid is by testing their blood or feces for \u003cem\u003eSalmonella typhi\u003c/em\u003e. \u003cem\u003eSalmonella typhi\u003c/em\u003e exclusively thrives in humans, with no animal reservoir.\u003csup\u003e\u0026nbsp;[8]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003eComplications arising from typhoid fever encompass intestinal hemorrhage, intestinal perforation, encephalitis, metastatic abscess, and others. Paratyphoid and Typhoid conjugate vaccines are authorized in India. For the past 60-70 years, antibiotics have remained the sole mode of treatment since the discovery of Salmonella. Consequently, drug-resistant organisms have emerged, homoeopathic medicines do not cause drug resistance even when administered for long time. \u0026nbsp;Hence our proposal aims to assess the In-vitro Efficacy of Homoeopathic Medicine against \u003cem\u003eSalmonella typhi\u003c/em\u003e. The homeopathic remedies employed in this study encompass \u003cem\u003eBaptisia tinctoria\u003c/em\u003e, \u003cem\u003eEupatorium perfoliatum\u003c/em\u003e, and \u003cem\u003eRhus toxicodendron\u003c/em\u003e in mother tincture, as well as potencies of 30 CH and 200 CH.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eOBJECTIVE:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo determine the inhibitory effect of the various homoeopathic drugs against \u003cem\u003eSalmonella typhi\u003c/em\u003e confirming the antimicrobial activity of potentized drugs in higher dilutions.\u003c/p\u003e"},{"header":"MATERIALS AND METHODOLOGY","content":"\u003cp\u003e\u003cstrong\u003eMedicines:\u003c/strong\u003e\u003csup\u003e\u0026nbsp;[4]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003eAs per Homoeopathic pharmacopoeia of India, standardised homoeopathic Medicines \u003cem\u003eBaptisia tinctoria\u003c/em\u003e, \u003cem\u003eRhus toxicodendron\u003c/em\u003e and \u003cem\u003eEupatorium perfoliatum\u003c/em\u003e in different potencies like Q, 30 CH and 200 CH along with Ethanol of 90% v/v were procured from GMP certified Drug Manufacturing unit of Dr.Willmar Schwabe, India.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMicroorganism:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe bacterial culture of \u003cem\u003eS.Typhi\u003c/em\u003e (MTCC No: 3231) procured as lyophilized freeze-dried culture strain from the Microbial Type Culture Collection, Institute of Microbial Technology, Chandigarh, was used to evaluate the antimicrobial activity of homoeopathic drugs.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePreparation of Agar medium:\u003c/strong\u003e\u003csup\u003e\u0026nbsp;[8]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003eThe agar medium was prepared by dissolving 4.95 g of MacConkey agar in 100 ml of distilled water and dissolved completely with the help of a hot plate. \u0026nbsp; Then the media was sterilised by autoclaving at 15 lb pressure (121\u003csup\u003e0\u003c/sup\u003eC) for 15 minutes. The sterilised agar was transferred into the petri plates (100 mm \u0026times; 15 mm) and was allowed to solidify. These procedures were completed in laminar air flow under aseptic conditions.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePreparation of bacterial culture:\u003c/strong\u003e\u003csup\u003e\u0026nbsp;[8]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003eThe bacterial strain of \u003cem\u003eSalmonella typhi\u003c/em\u003e (MTCC NO. 3231), lyophilized freeze-dried culture was aseptically opened in a laminar air flow and the suspension was made as per protocol. 0.5 ml of buffered peptone water was taken in the microfuge tube and freeze-dried culture was transferred into it and mixed well. \u0026nbsp;The mixture was allowed for 20 minutes before it was transferred to solid media. Petri plates containing MacConkey agar medium (HiMedia, Mumbai, India, Catalogue No. M081) was incubated for 24 hours at 37\u003csup\u003e0\u003c/sup\u003eC to give white round colonies against a mild reddish pink background. Incubation period of \u003cem\u003eSalmonella typhi\u003c/em\u003e is 24 hours. This culture was kept as Broth culture/stock culture. Bacterial cultures were repeatedly sub cultured using streaking methods and maintained to obtain pure isolation on the MacConkey medium for further drug sensitivity assay.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003c/strong\u003e\u003cstrong\u003eIdentification of Salmonella typhi:\u003c/strong\u003e\u003csup\u003e\u0026nbsp;[8]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCatalase Test:\u003c/strong\u003e\u003csup\u003e\u0026nbsp;[8]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003e2 ml of hydrogen peroxide solution was taken into the test tube and by using a glass rod, samples of several colonies from the Broth Culture were immersed into the hydrogen peroxide solution. \u0026nbsp;Formation of an immediate bubble confirmed the presence of \u003cem\u003eSalmonella typhi\u003c/em\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePreparation of disc for antimicrobial assay:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAgar disc diffusion method was adopted for determining the antimicrobial activity of different homoeopathic medicines in various potencies. Whatman filter paper No.1 of size approximately 6 mm in diameter was used to prepare the standard discs and autoclaved at 121\u003csup\u003e0\u003c/sup\u003eC at 15 lbs for 15 minutes. These discs were soaked in Baptisia\u003cem\u003e\u0026nbsp;tinctoria\u003c/em\u003e, \u003cem\u003eRhus toxicodendron\u003c/em\u003e, \u003cem\u003eEupatorium perfoliatum\u003c/em\u003e under different potencies like Q, 30CH and 200CH, Ethanol of 90% v/v and Ceftriaxone for 30 to 45 minutes. after which they were taken out and allowed to dry under aseptic conditions.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDetermination of Antimicrobial Activity:\u003c/strong\u003e\u003csup\u003e\u0026nbsp;[7, 8, 19]\u003c/sup\u003e\u003c/p\u003e\n\u003cp\u003eThe antimicrobial activity of the selected homoeopathic medicines under different potencies were evaluated using the agar disc diffusion method by measuring the diameter of growth inhibition zones. Ceftriazone was used as a positive control \u0026nbsp;and Ethanol of 90% v/v was used as a negative control. The discs were inverted and placed in the petri well plates in an incubator set to 37\u003csup\u003e0\u003c/sup\u003eC. After 24-hour growth, an inhibition zone around the discs was measured. The diameter of the growth inhibition zone around the discs was measured using a scale. Each petri well plate was divided into five compartments. Three compartments were allocated for different homoeopathic potencies and one for vehicle Ethanol (V) and one for Antibiotic (AB).\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003eAll the three homoeopathic medicine in various potencies exhibited the inhibitory action against \u003cem\u003eSalmonella typhi\u003c/em\u003e (\u003cem\u003eTable 1 and Figure 2\u003c/em\u003e). Among these, \u003cem\u003eRhus toxicodendron\u003c/em\u003e Q and \u003cem\u003eBaptisia tinctoria\u003c/em\u003e 200 CH exhibited maximum inhibition zone of diameter of 16 mm and 14 mm respectively followed by \u003cem\u003eBaptisia tinctoria\u003c/em\u003e 30 CH and \u003cem\u003eEupatorium perfoliatum\u003c/em\u003e 30 CH showing the least zone of inhibition of 11 mm and 10.9 mm respectively. However the antibiotic positive control ceftriaxone showed the maximum zone of inhibition.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003cstrong\u003e: Zone of Inhibition of Homoeopathic Medicines in different potencies\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"24.23756019261637%\"\u003e\n \u003cp\u003e\u003cstrong\u003eDrugs\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e\u003cstrong\u003eMother Tincture\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e\u003cstrong\u003e30 CH\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e\u003cstrong\u003e200 CH\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.088282504012842%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(Ceftriaxone)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e\u003cstrong\u003eVehicle\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(Ethanol 90% v/v)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"24.23756019261637%\"\u003e\n \u003cp\u003e\u003cem\u003eBaptisia tinctoria\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e12 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e11 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e14 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.088282504012842%\"\u003e\n \u003cp\u003e12 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e7 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"24.23756019261637%\"\u003e\n \u003cp\u003e\u003cem\u003eRhus toxicodendron\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e16 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e12 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e13 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.088282504012842%\"\u003e\n \u003cp\u003e19 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e7.1 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"24.23756019261637%\"\u003e\n \u003cp\u003e\u003cem\u003eEupatorium perfoliatum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e13 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e10.9 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.593900481540931%\"\u003e\n \u003cp\u003e12 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.088282504012842%\"\u003e\n \u003cp\u003e18 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.74317817014446%\"\u003e\n \u003cp\u003e6.9 mm\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eFrom the sources of various homoeopathic Materia Medica, \u003cem\u003eBaptisia tinctoria\u003c/em\u003e in low dilutions produces a form of antibodies to \u003cem\u003ebac.typhosus, viz, the agglutinins thus it raises the bodily natural resistances to the invasion of the bacillary intoxication\u003c/em\u003e, which produces the typhoid syndrome\u003csup\u003e[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]\u003c/sup\u003e. \u003cem\u003eRhus toxicodendron\u003c/em\u003e indicated therapeutically for typhoid fever\u003csup\u003e[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]\u003c/sup\u003e. From the sources of drug proving and clinical verification by pioneers the antibacterial activity has been recorded even before the antimicrobial era. This study reiterates the fact and scientifically confirms the antimicrobial activity of \u003cem\u003eBaptisia tinctoria\u003c/em\u003e and \u003cem\u003eRhus toxicodendron\u003c/em\u003e. \u003cem\u003eEupatorium perfoliatum\u003c/em\u003e is a well-known antiviral remedy\u003csup\u003e[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]\u003c/sup\u003e, however it exhibits antimicrobial activities.\u003c/p\u003e"},{"header":"CONCLUSION","content":"\u003cp\u003eIn recent days, repeated use of antibiotics in frequent intervals has caused drug resistance in patients, which poses a serious threat to the community. This resistance has pushed them to discover new generations of antibiotics year after year. Homoeopathic medicines do not cause any resistance and do not produce undesirable side-effects as it is being dispensed from low attenuation to ultra-high dilutions since the Hahnemannian era onwards. This evidence based in-vitro study contributes to the scientific and literature fact that homoeopathic medicine has definite antimicrobial activity against \u003cem\u003eSalmonella typhi\u003c/em\u003e.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eCONFLICT OF INTEREST:\u003c/h2\u003e \u003cp\u003eNone\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eDr.K.Narayanasamy - Conceptualization, Material method, Manuscript preparation Manuscript editing, software ,Manuscript review.Dr.P. Bhuvanesh of intellectual Concepts, investigation Design, content, Manuscript reviewG. Sridevi-content ,writing, material and methods, Supervision Senthilkumaran.G - Manuscript review, Result, Software JP.Jerin- discussion,Manuscript review, Manuscript editing\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eWe thank our colleagues at The Tamilnadu Dr. MGR Medical University, Tamilnadu, India, for their support, and the reviewers of BMC Microbiology for their valuable feedback.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAdak S, Chakraborty D, Maji HS, Basu S, Roy P, Mitra S, et al. Comparison of the antimicrobial activity of the phyto-constituents obtained from the stem bark and leaf extracts of \u003cem\u003ePhyllanthus emblica\u003c/em\u003e L. against different strains of \u003cem\u003eStaphylococcus aureus\u003c/em\u003e and \u003cem\u003eSalmonella typhi\u003c/em\u003e. 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Asian J Pharm Technol. 2013;3:20\u0026ndash;5.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-microbiology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mcro","sideBox":"Learn more about [BMC Microbiology](http://bmcmicrobiol.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/mcro","title":"BMC Microbiology","twitterHandle":"#bmcmicrobiology","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Salmonella typhi, antimicrobial activity, agar disc diffusion method, Baptisia tinctoria, Rhus toxicodendron, Eupatorium perfoliatum, Typhoid fever","lastPublishedDoi":"10.21203/rs.3.rs-4432701/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4432701/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eTyphoid fever, caused by Salmonella typhi, is a global health issue with high morbidity and mortality rates, especially in low-income regions. The rise of antibiotic-resistant strains necessitates new treatment strategies. This study explores the potential of homoeopathic medicines, known for their safety and non-induction of resistance, as an alternative treatment.\u003c/p\u003e \u003cp\u003eThree homoeopathic medicines, Rhus toxicodendron, Eupatorium perfoliatum, and Baptisia tinctoria, were tested for their inhibitory effects on S. typhi using the agar disc diffusion method. The medicines, along with a control antibiotic, ceftriaxone, were applied to agar plates with S. typhi cultures.\u003c/p\u003e \u003cp\u003eAll tested homoeopathic medicines showed inhibitory effects against S. typhi. Rhus toxicodendron Q and Baptisia tinctoria 200 CH showed significant inhibition zones of 16 mm and 14 mm, respectively. Baptisia tinctoria 30 CH and Eupatorium perfoliatum 30 CH also showed inhibitory effects, while ceftriaxone exhibited the largest inhibition zone.\u003c/p\u003e \u003cp\u003eThis study highlights the need for innovative treatments against antibiotic-resistant infections. It confirms the antimicrobial potential of homoeopathic medicines against S. typhi, suggesting a possible solution for this infectious disease. Further research is needed to assess their clinical effectiveness in managing typhoid fever.\u003c/p\u003e","manuscriptTitle":"Exploring Homoeopathic Potential Against Typhoid: Baptisia tinctoria, Rhus toxicodendron, and Eupatorium perfoliatum in growth Inhibition of Salmonella typhi – In-vitro study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-06-20 08:00:56","doi":"10.21203/rs.3.rs-4432701/v1","editorialEvents":[{"type":"communityComments","content":1},{"type":"editorAssigned","content":"","date":"2024-06-06T19:10:26+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-06-06T17:57:47+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Microbiology","date":"2024-05-16T18:01:11+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-microbiology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mcro","sideBox":"Learn more about [BMC Microbiology](http://bmcmicrobiol.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/mcro","title":"BMC Microbiology","twitterHandle":"#bmcmicrobiology","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"f953b5e7-7bf2-4fe2-9d5c-94fc167c06f6","owner":[],"postedDate":"June 20th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2024-06-20T08:00:56+00:00","versionOfRecord":[],"versionCreatedAt":"2024-06-20 08:00:56","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4432701","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4432701","identity":"rs-4432701","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}