Flavonoids interfere with biofilm formation by targeting diguanylate cyclases in multidrug resistant Vibrio cholerae – Nature

Report on the Anti-Biofilm Efficacy of Flavonoids Against Multidrug-Resistant Vibrio cholerae and Alignment with Sustainable Development Goals

Introduction

The persistence of cholera, particularly in the Global South, presents a significant challenge to global public health, directly impacting the achievement of Sustainable Development Goal 3 (Good Health and Well-being). The rise of multidrug-resistant (MDR) strains of Vibrio cholerae has led to increased treatment failures, undermining efforts to reduce mortality from communicable diseases. The pathogen’s ability to form biofilms is a key survival mechanism, contributing to both environmental persistence and antibiotic tolerance. This persistence in aquatic environments also poses a threat to Sustainable Development Goal 6 (Clean Water and Sanitation).

This report details an investigation into alternative therapeutic strategies that target V. cholerae biofilms. The study explores the anti-biofilm efficacy of two naturally derived flavonoids, baicalein and fisetin. By focusing on plant-based compounds, this research aligns with principles of Sustainable Development Goal 12 (Responsible Consumption and Production), promoting sustainable alternatives to conventional antibiotics. The primary objective was to assess the capacity of these flavonoids to inhibit and eradicate biofilms and to elucidate the underlying molecular mechanisms, specifically their interference with the cyclic-di-GMP (c-di-GMP) signaling pathway, a critical regulator of biofilm formation.

Key Findings on Anti-Biofilm Efficacy

Inhibition and Eradication of Biofilms

The study evaluated the efficacy of baicalein and fisetin against high biofilm-forming, multidrug-resistant V. cholerae strains. The results demonstrated significant anti-biofilm activity for both compounds, with fisetin showing superior potency.

• Minimum Biofilm Inhibitory Concentration (MBIC): The concentration required to inhibit biofilm formation by at least 50% was determined.
  • Baicalein: 40 µg/mL
  • Fisetin: 30 µg/mL
• Minimum Biofilm Eradication Concentration (MBEC): The concentration required to eradicate 50% of a pre-formed biofilm was also established.
  • Baicalein: 70 µg/mL
  • Fisetin: 50 µg/mL

These findings indicate that both flavonoids can not only prevent the formation of biofilms but also disrupt established ones, a critical requirement for treating chronic infections and decontaminating surfaces, thereby supporting both SDG 3 and SDG 6.

Impact on Biofilm Structure and Adhesion Factors

Treatment with flavonoids resulted in a significant reduction of key factors that contribute to biofilm integrity and bacterial adhesion.

1. Reduction of Extracellular Polymeric Substances (EPS): Both carbohydrate and protein content within the biofilm matrix were significantly reduced following treatment, leading to a weaker, less stable biofilm structure. This was visually confirmed by Atomic Force Microscopy (AFM), which showed a substantial decrease in biofilm thickness.
2. Inhibition of Adhesion Properties: The flavonoids effectively diminished accessory adhesion factors.
   • Auto-aggregation: The ability of bacterial cells to clump together was reduced.
   • Cell Surface Hydrophobicity: Adhesion to hydrocarbon surfaces was significantly decreased, particularly with fisetin treatment. This suggests potential applications as surface disinfectants, contributing to SDG 6.
3. Suppression of Indole Production: Indole, a signaling molecule that promotes biofilm formation, was substantially decreased upon flavonoid treatment.

Molecular Mechanisms of Action

Interference with c-di-GMP Signaling Pathway

The study provides novel insights into the mechanism by which these flavonoids exert their anti-biofilm effects, demonstrating their ability to target the c-di-GMP signaling pathway. This pathway is a central regulator of biofilm formation in many bacteria.

• Downregulation of Diguanylate Cyclases (DGCs): Gene expression analysis revealed that both baicalein and fisetin significantly downregulated the expression of key DGCs responsible for synthesizing c-di-GMP. The targeted genes included:
  • cdgA
  • cdgH
  • cdgK
  • cdgL
  • cdgM
  • vpvC
• Suppression of Biofilm Regulators: Consequently, the expression of major biofilm regulators that are activated by c-di-GMP was also reduced.
  • vpsR
  • vpsT
  • aphA

Fisetin demonstrated a more pronounced downregulatory effect on these genes compared to baicalein, correlating with its superior anti-biofilm performance.

In Silico Docking and Simulation

Molecular docking and dynamic simulation studies were conducted to validate the experimental findings. The results confirmed that both flavonoids could bind to the active site of a DGC protein. Notably, fisetin exhibited a higher binding affinity and formed a more stable complex with the DGC active site. This enhanced interaction, potentially due to the specific arrangement of hydroxyl groups in its structure, provides a molecular explanation for its heightened efficacy in suppressing biofilm formation.

Conclusion and Implications for Sustainable Development Goals

Summary of Report

This investigation successfully demonstrates that the flavonoids baicalein and fisetin possess potent anti-biofilm properties against multidrug-resistant Vibrio cholerae. Fisetin was identified as the more effective agent. The primary mechanism of action involves the downregulation of diguanylate cyclases (DGCs), leading to the disruption of the c-di-GMP signaling cascade that governs biofilm formation. The compounds were also found to be biocompatible with human cells at effective concentrations.

Contribution to Sustainable Development Goals

• SDG 3 (Good Health and Well-being): By identifying a novel strategy to combat cholera and circumvent antibiotic resistance, this research contributes directly to the global effort to end epidemics of communicable diseases. These findings support the WHO’s goal of reducing cholera-associated mortality by 90% by 2030.
• SDG 6 (Clean Water and Sanitation): The ability of these flavonoids to disrupt biofilms and reduce bacterial adhesion on surfaces suggests their potential use as disinfectants for water storage and food processing equipment. This application would help control the environmental reservoirs of V. cholerae, contributing to safer water and sanitation systems.
• SDG 12 (Responsible Consumption and Production): The exploration of naturally derived, plant-based compounds as therapeutic agents represents a sustainable approach to drug discovery. It promotes the use of renewable resources and offers an alternative to the production of synthetic antibiotics, aligning with the goal of sustainable production patterns.

In conclusion, the study presents compelling evidence for the use of flavonoids as a viable alternative strategy for cholera management, with significant positive implications for global health and sustainable development.

SDGs, Targets, and Indicators Analysis

1. Which SDGs are addressed or connected to the issues highlighted in the article?

The article primarily addresses issues related to global health, disease control, and scientific innovation, which directly connect to several Sustainable Development Goals (SDGs). The analysis identifies the following relevant SDGs:

• SDG 3: Good Health and Well-being: This is the most prominent SDG, as the article focuses on cholera, a severe communicable disease, the growing problem of antibiotic resistance, and the development of new therapeutic strategies to improve health outcomes.
• SDG 6: Clean Water and Sanitation: The article indirectly connects to this goal by highlighting challenges in cholera treatment. It mentions that “during severe outbreaks, sourcing contamination-free water to be used for rehydration therapy pose a significant challenge,” linking the spread and management of cholera to the availability of clean water.
• SDG 9: Industry, Innovation, and Infrastructure: The core of the article is a scientific study exploring innovative solutions to a health crisis. It details research into flavonoids as alternative anti-biofilm agents to combat multidrug-resistant Vibrio cholerae, which aligns with the goal of promoting scientific research and innovation.

2. What specific targets under those SDGs can be identified based on the article’s content?

Based on the article’s discussion of cholera epidemics, antibiotic resistance, and the need for new treatments, the following specific targets can be identified:

1. Under SDG 3 (Good Health and Well-being):
   • Target 3.3: “By 2030, end the epidemics of AIDS, tuberculosis, malaria and neglected tropical diseases and combat hepatitis, water-borne diseases and other communicable diseases.”
     
     Explanation: The article is centered on cholera, a major water-borne communicable disease. It explicitly mentions the global effort to control it by stating, “the present work would help to add newer insights into cholera management strategies and contribute towards the target set by WHO for reducing cholera associated mortality upto 90% by 2030.” This directly aligns with the ambition of ending epidemics of communicable diseases.
   • Target 3.d: “Strengthen the capacity of all countries, in particular developing countries, for early warning, risk reduction and management of national and global health risks.”
     
     Explanation: The article identifies the “rise of antibiotic resistance” and the emergence of “multidrug-resistant (MDR) and extensive drug-resistant (XDR) V. cholerae strains” as a “substantial public health issue” and a global health risk. The research into alternative treatments like flavonoids is a direct effort to strengthen the capacity to manage this risk, for which current antibiotic therapies are failing.
2. Under SDG 6 (Clean Water and Sanitation):
   • Target 6.1: “By 2030, achieve universal and equitable access to safe and affordable drinking water for all.”
     
     Explanation: While the study’s focus is not on water infrastructure, it highlights the critical role of clean water in managing cholera. The statement that “sourcing contamination-free water to be used for rehydration therapy pose a significant challenge” underscores that the lack of safely managed drinking water (the focus of Target 6.1) exacerbates the health crisis during a cholera outbreak.
3. Under SDG 9 (Industry, Innovation, and Infrastructure):
   • Target 9.5: “Enhance scientific research, upgrade the technological capabilities of industrial sectors in all countries, in particular developing countries… encouraging innovation…”
     
     Explanation: The entire article is a testament to this target. It is a scientific research paper that seeks to find an innovative solution (using flavonoids baicalein and fisetin) to the problem of antibiotic resistance in cholera treatment. The study employs advanced methods like molecular docking and gene expression analysis to “unveil the potential of flavones to interact with DGCs and subsequently modulate c-di-GMP signalling pathways,” which is a clear example of enhancing scientific research to address a critical development challenge.

3. Are there any indicators mentioned or implied in the article that can be used to measure progress towards the identified targets?

The article mentions and implies several quantitative and qualitative indicators that can be used to measure progress towards the identified targets.

• For Target 3.3 (End epidemics of communicable diseases):
  • Mortality Rate from Cholera: The article explicitly mentions the WHO’s goal of “reducing cholera associated mortality upto 90% by 2030.” It also cites a report that a “126% increased rate of mortality associated with cholera was witnessed in 2024.” This directly implies that the mortality rate is a key indicator for tracking progress.
  • Incidence of Cholera: The article provides specific data on the number of cholera cases, stating that in 2023, “the 7th cholera pandemic continued to surge with 535,321 cases, up from 472,697 in 2022.” The number of new cases per year is a direct indicator of the scale of the epidemic.
• For Target 3.d (Strengthen capacity to manage health risks):
  • Prevalence of Antibiotic Resistance: The article’s premise is the “Rise of antibiotic resistance” and the emergence of “multidrug-resistant (MDR) and extensive drug-resistant (XDR) V. cholerae strains.” Tracking the prevalence of these strains is an indicator of the growing health risk that needs management.
  • Development of Alternative Therapeutic Agents: The study itself, focusing on the efficacy of flavonoids, serves as an indicator. The measurement of “Minimum biofilm inhibitory concentrations (MBIC)” and “Minimum biofilm eradication concentrations (MBEC)” for baicalein and fisetin are specific, measurable outcomes that demonstrate progress in developing new tools to manage drug-resistant infections.
• For Target 9.5 (Enhance scientific research and innovation):
  • Investment in and Publication of Scientific Research: The existence of this detailed scientific paper is an indicator of ongoing research and innovation in the field. The depth of the study, from in vitro experiments to in silico molecular dynamics simulations, represents the kind of advanced scientific research this target aims to promote.

4. Table of SDGs, Targets, and Indicators

Source: nature.com