Decolonizing the gut from multidrug-resistant bacteria: Current strategies and future perspectives

Abstract

The rise of multidrug-resistant organisms (MDROs) represents a serious global health crisis, with the gastrointestinal tract serving as a major reservoir for these pathogens. This review highlights the burden of gut colonization by MDROs, its role in spreading antimicrobial resistance, and explores current and emerging strategies for decolonization. Various non-antibiotic approaches such as probiotics, prebiotics, bacterial consortia, selective digestive decontamination, faecal microbiota transplantation, bacteriophage therapy, and Clustered Regularly Interspersed Short Palindromic Repeats—CRISPR-associated protein systems along with dietary interventions have been assessed for their potential to restore microbial balance and reduce MDRO carriage. While promising results have emerged from early studies and animal models, most interventions remain investigational. Rigorous clinical trials, standardized protocols, and safety assessments are essential before these approaches can be integrated into routine practice for MDRO management.

Key Words: Antimicrobial resistance; Fecal microbiota transplantation; Gut decolonisation; Multidrug-resistant organisms; Selective digestive decontamination

Core Tip: Asymptomatic colonization of the gastrointestinal tract by extended-spectrum β-lactamase or carbapenamase-producing enterobacterales poses a substantial risk of infections caused by these resistant bacteria. Furthermore, this colonization carries a significant risk of transmitting these organisms to other patients and the broader community. Emerging interventions such as faecal microbiota transplantation, phage therapy, engineered probiotics, and Clustered Regularly Interspersed Short Palindromic Repeats—CRISPR-associated protein systems offer new hope for precise and sustainable decolonization. Additionally, dietary interventions and immune modulation may serve as supportive strategies to enhance the resilience of the host microbiota. However, many of these approaches are still in the nascent stage, and their long-term efficacy, safety, and regulatory approval remain barriers to routine clinical application.

INTRODUCTION

Since the discovery of penicillin in the early 20^th century, antibiotics have revolutionized the healthcare system, offering a promising solution against deadly bacterial infections. However, antibiotics are frequently misused and have been prescribed inappropriately for non-infectious pathologies, viral infections or even in crop cultivation and animal husbandry to facilitate growth and prevent diseases in livestock. This unchecked use has generated an environment for antimicrobial resistance (AMR) to flourish. Over the last decade, healthcare systems have observed a steep increase in the number of multidrug-resistant (MDR) infections. Consequently, developing non-antibiotic innovations to curtail the dissemination of MDR strains while preserving the gut microbiota’s equilibrium has become an imperative necessity.

Asymptomatic colonization of the digestive tract by extended-spectrum β-lactamase (ESBL) or carbapenemase-producing enterobacterales (CRE), poses a significant risk factor for fatal infections caused by these resistant bacteria. Furthermore, it serves as a substantial source of transmission of these organisms to other patients and the broader community[1]. This constitutes a major public health threat, undermines the intended clinical utility of antibiotics, and compromises the management of complicated infectious diseases.

Different methods and interventions have been adopted worldwide for combating AMR to decrease the emergence and spread of MDR infections. An emerging role of the gut microbiome in preventing colonization by MDR organisms, which are typically the cause of subsequent invasive infections, has been observed. Considering these factors, interventions such as the use of probiotics, faecal microbiota transplantation (FMT), phage therapy, and bacterial consortia have recently gained prominence in research studies[2].

PROBLEM BURDEN

A systematic analysis published in 2019, assessing the global burden of bacterial AMR, concluded that AMR was associated with approximately 4.95 million deaths worldwide, with 1.27 million of these deaths directly attributable to bacterial AMR[3]. This puts AMR among the leading causes of death globally, surpassing diseases such as human immunodeficiency virus or acquired immunodeficiency syndrome and malaria. There are regional disparities and the prevalence of AMR is not uniformly distributed across the globe. Sub-Saharan Africa and South Asia experienced the highest impact, with 27.3 and 21.5 deaths per 100000 attributable to AMR, respectively. These regions face greater challenges due to limited healthcare infrastructure, inefficient infection control practices and suboptimal access to effective antibiotics[3].

The largest study conducted on the Indian sub-continent by Gandra et al[4], found an overall mortality of 13.1% with MDR infections. Gram-negative MDR infections had higher overall mortality rates (17.7%) compared to Gram-positive infections (10.8%), especially in the intensive care unit (ICU), where it was 26.9%. MDR and extensively drug-resistant (XDR) Escherichia coli (E. coli), XDR Klebsiella pneumoniae (K. pneumoniae), and MDR Acinetobacter baumannii shared the major percentage of mortality and increased the risk of death by 2–3 times compared to susceptible infections. Methicillin-resistant staphylococcus aureus (MRSA) infections exhibited greater mortality rates, particularly when resistance extended to aminoglycosides[4].

A retrospective cross-sectional study conducted in India, analyzed MDR organisms trends over a decade (2014–2023) and found a significant rise in MDR infections. Out of more than 1.3 million in-patients, 7311 cases were MDRs, with an overall incidence density rate (IDR) of 5.53 per 1000 patient days. CRE were the most common (42% of cases), followed by increasing rates of MRSA, vancomycin-resistant enterococcus (VRE), and MDR Pseudomonas aeruginosa and Acinetobacter spp. The overall MDR IDR nearly doubled from 4.20 to 8.77, indicating a rising trend in resistance[5].

Prevalence in the community

Although there is enough evidence on the burden of MDR bacteria in hospitalized settings and ICU environments, little is understood about their prevalence in the community. There is growing evidence that depicts a high level of carriage in healthy people without any signs of infection. Nevertheless, these colonized individuals may subsequently develop infections and, in turn, transmit them to the community (through various mechanisms such as fecal-oral route or surface contamination), particularly to the vulnerable population, whose numbers are surging globally[6]. The risk factors for acquiring MDR bacteria have been summarized in Table 1[6-8].

Table 1 Risk factors for acquiring multi-drug resistant bacteria.

Category	Risk factor
Healthcare-associated	Prolonged hospitalization (especially in ICU). Recent surgery or invasive procedures. Use of medical devices (catheters, ventilators, central lines). Residence in long-term care facilities. Frequent hospital admissions or outpatient visits. Hemodialysis or chronic outpatient treatments
Antibiotic exposure	Prolonged use of broad-spectrum antibiotics. Inappropriate or incomplete antibiotic courses. Over-the-counter or self-medicated antibiotic use
Patient-related	Immunocompromised status (e.g., cancer, HIV, transplants). Chronic illnesses (e.g., diabetes, COPD, renal failure). Extremes of age (infants and elderly). Malnutrition. Gut dysbiosis due to prolonged gastric acid suppression
Environmental/community	International travel to high MDR prevalence regions. Contact with infected or colonized individuals. Poor sanitation or overcrowded living conditions

ICU: Intensive care unit; HIV: Human immunodeficiency virus; COPD: Chronic obstructive pulmonary disease; MDR: Multi-drug resistant.

ROLE OF GUT MICROBIOME AND DIETARY INTERVENTIONS

The gut microbiome is a large ecological niche which is an intricately developed and highly organized ecosystem of microorganisms. It plays an important role in maintaining human health by aiding digestion, regulating immune cascades and controlling endocrine and neurological functions[2]. It is also a storehouse of notorious ABR genes which can be transmitted amongst the microbial species in the gut by methods of horizontal gene transfer such as conjugation, transformation and transduction[9].

Exposure to antibiotics causes further gut dysbiosis that disturbs the well-balanced microbial diversity, resulting in the selective expansion of enteric MDR organisms. Notably, gram-negative bacteria (GNB) like ESBL-E, CRE, VRE, pose the greatest threat and have been shortlisted on the World Health Organization critical priority list due to their growing prevalence and the inefficient treatment modalities available to address them[10].

Diets rich in fibre and fermented foods have been shown to boost microbial diversity, improve immune function and support the growth of beneficial gut bacteria (commensals). Components like polyphenols in plant-based foods have been found to have MDR organisms-suppressing antimicrobial effects. In contrast, diets high in processed foods, saturated fats, and sugars can disrupt the gut microbial balance, creating an environment conducive to the expansion of pathogenic bacteria and potentially promoting AMR[2,11]. It is unclear whether these effects are primarily due to dietary components themselves, antibiotic use in food animals, or other environmental factors. Future research should aim to separate these factors by using controlled dietary studies. These studies can help understand how specific nutrients or harmful additives and toxins affect the gut microbiota and the expression of AMR genes.

STRATEGIES OF GUT DECOLONIZATION

Prebiotics and probiotics

Prebiotics are indigestible dietary fibres that selectively enhance the growth and function of beneficial gut bacteria. By supplying nutrients to these microbes, prebiotics support a balanced microbiome, which may help reduce the incidence of AMR[12]. Probiotics are live microorganisms which, when administered in adequate amounts, confer a health benefit on the host[13].

A systematic review of 29 randomized controlled trials involving 2871 participants evaluated probiotics vs placebo for removing AMR pathogens from the gut. The data revealed that pathogenic bacteria persisted in 22% of those treated with probiotics, compared to 30.8% in the placebo group. This resulted in a pooled odds ratio of 0.59 (95%CI: 0.43–0.81), demonstrating a clear benefit for probiotics (P = 0.0001). Outcomes depended significantly on the probiotic type (P < 0.018) and the pathogen targeted (P < 0.02), showing better results against enterobacterales than VRE[14].

Bacterial consortia, also called live biotherapeutic products, combine multiple strains with complementary antimicrobial effects, and have also proven effective at breaking down biofilms of MDR organisms and limiting their spread[15]. A 2023 study by Medlock et al[16] found that VE707, a consortium of 94 Live biotherapeutic strains, achieved a reduction exceeding 3-log10 in K. pneumoniae and E. coli colonization in mice (P = 0.002), suggesting possible future use in humans[16].

Selective digestive decontamination

Selective digestive decontamination (SDD) is a preventive antimicrobial approach aimed at clearing potentially harmful bacteria, such as ESBL-E and CRE, from the digestive tract while attempting to maintain the patient’s natural microbiota. It typically involves applying non-absorbable antibiotics to the oropharynx and gastrointestinal tract, often paired with intravenous antibiotics for broader coverage. The objective is to reduce resistant bacteria in these regions, thereby preventing infections and their transmission[17]. Although SDD has shown success in reducing ICU-acquired infections, standardized protocols are lacking and the regimens vary in antibiotic selection, administration methods, and the use of systemic agents.

A 16-year ecological study in Spain analyzed bacterial susceptibility in ICUs with and without SDD. Results indicated that ICUs not employing SDD had higher susceptibility rates for E. coli, Proteus mirabilis, and Enterococcus faecalis compared to those using SDD[18]. Another Spanish study employed SDD as a control measure for an MDR organisms outbreak in a 30-bed medical-surgical ICU. The study reported a decrease in the rates of colistin- and tobramycin-resistant colonization, and no significant increase in resistance over a period of 1000 days. This intervention also significantly lowered rates of ventilator-associated pneumonia and bloodstream infections[19].

A recently published meta-analysis by Zhang et al[20] indicated short-term advantages of SDD. ESBL-producing Enterobacterales were more responsive to antibiotic-based decolonization and SDD compared to CRE, which were harder to eradicate due to underlying resistance mechanisms. Results for CRE were more promising when combined with FMT. However, the long-term effects of these approaches, beyond one month, remained undefined[20].

A critical concern, however, is whether SDD might hasten resistance development in the gut microbiota. A South African study, for instance, attributed the rise of colistin-resistant K. pneumoniae in an ICU attributable to oral colistin used in an SDD regimen[21]. Additionally, SDD may disrupt anaerobic gut bacteria, markedly reducing populations of certain micro-organisms such as the Fecalibacterium prausnitzii group (valued for its anti-inflammatory properties), along with fusobacteria, proteobacteria, and actinobacteria[22,23].

In 2018, the European Society of Clinical Microbiology and Infectious Diseases-European Committee of Infection Control (ESCMID-EUCIC) reviewed 27 studies on decolonizing MDR GNB carriers and recommended against routine SDD for CRE, citing insufficient evidence[23]. These guidelines, based on pre-2018 data, may need re-evaluation given the changing prevalence of ESBL-E and CRE and the emergence of new research.

FMT

FMT refers to a process of transferring processed faeces from a healthy subject into the gastrointestinal tract of a patient to improve microbial diversity. This modality has already been recommended by the Infectious Diseases Society of America as well as ESCMID-EUCIC for treating recurrent Clostridioides difficile infection (CDI) not responding to appropriate antibiotics[24,25]. The donor stool must be thoroughly screened for microbes and ARGs, then rapidly processed and cryopreserved under strict cold chain protocols to preserve microbial viability—an effort that is both labour-intensive and costly. The modes of administration, which have been elaborated in Table 2, further affect the safety of the procedure and patient comfort[26,27].

Table 2 Different approaches for faecal microbiota transplantation.

Approach	Method	Pros	Cons
Colonoscopy	Delivery of fecal material into the colon via a colonoscope	High success rate, allows direct placement in the colon	Invasive, requires bowel preparation, potential complications like perforation
Nasogastric/nasoenteric tube	Tube insertion through the nose into the stomach or small intestine for faecal infusion	Non-surgical, effective for small intestine delivery	Risk of aspiration, discomfort, nausea and vomiting
Capsule delivery	Freeze-dried faecal material in capsules taken orally	Non-invasive, convenient, avoids procedural risks	Requires multiple capsules, potential for reduced efficacy in some cases
Enema	Faecal material mixed with solution and introduced via the rectum	Simple, can be done at home, avoids invasive procedures	Lower retention time, may require multiple doses
Rectal infusion	Controlled delivery of faecal material into the rectum	Less invasive than colonoscopy, localized delivery	Requires professional administration, may not reach upper colon effectively

The findings of a study by Millan et al[28] indicated that patients who had recurrent CDI had a larger number of ABR genes, including efflux pumps, beta-lactam and fluoroquinolone inactivation, compared to healthy adults. They also had a higher prevalence of Proteobacteria, E. coli and K. pneumoniae. On the positive side, the study also concluded the effectiveness of FMT in eradicating antibiotic-resistant pathogens and eliminating ABR genes[28]. However, resistance genes can also be transferred to the recipient from FMT donor stool, making the careful selection of healthy donors essential and necessitating further standardization[29].

Singh et al[30], reported the successful decolonisation of MDR organisms by FMT[30]. Another study by Dinh and colleagues also reported the benefits of FMT in restoring the gut microbiota and controlling antibiotic-resistant pathogens, specifically CRE and VRE[31]. A study on New Delhi metallo- β-lactamase-producing K. pneumoniae by Seong et al[32], found accelerated decolonization of these pathogens by FMT with effects seen for 205 days for healthy controls, but only 42 days for patients who underwent FMT (P = 0.007)[32]. More recently, Davido et al[33] in 2024 published a study reporting no statistically significant difference in the rate of complete carbapenemase-producing enterobacterales (CPE) decolonization between FMT-treated patients vs the control group. However, microbiota analysis revealed that responders exhibited significantly higher bacterial species richness and diversity after FMT compared to non-responders. Specific bacterial taxa were also more abundant in responders. Additionally, responders had a relatively lower abundance of CPE species before FMT than non-responders. These findings suggest that while FMT alone may not completely decolonize CPE, the presence of certain bacterial taxa and overall microbial diversity may somewhat contribute to successful decolonization[33].

Although an increasing number of studies and case reports support the use of FMT for MDR decolonization, clear clinical guidelines and standardized protocols are still lacking. Key questions, including optimal donor selection, dosing frequency, long-term safety, and the durability of decolonization remain unanswered. Establishing evidence-based protocols is essential to ensure the safe and effective use of FMT in managing MDR-GNB[14].

Bacteriophages

Bacteriophages, or phages are highly specific viruses that can target a single species or even a specific strain of bacteria. These entities were discovered long back by Frederick Twort and Félix d'Hérelle in 1915 and 1917, respectively. Their specificity arises from the phage’s ability to bind to unique receptors on the surface of bacterial cells, triggering a sequence of events that ultimately leads to bacterial destruction[34]. After attaching to its target bacteria, it transfers its genetic material to the bacteria leading to a process that makes multiple copies of phage particles. During the lytic cycle, this bacterial cell then ruptures, releasing new phages capable of infecting additional bacteria. Hence, the phages can continue multiplying and self-amplifying at the infection site[35].

Unlike antibiotics, which typically have broad-spectrum activity and can eliminate a wide array of bacterial species, phages act with more precision, targeting only their specific bacterial hosts (Table 3)[36]. Their host specificity, along with the unique ability to co-evolve with bacterial populations, makes phages a promising and adaptable weapon in addressing antibiotic-resistant infections[37].

Table 3 Comparison between phage therapy and antibiotics.

Feature	Phage therapy	Antibiotics
Mechanism of action	Targets and infects specific bacteria, leading to their lysis	Interferes with essential bacterial processes like cell wall or protein synthesis
Host specificity	Highly specific—usually affects only certain strains and species	Broad spectrum—may act on various bacterial species
Resistance development	Bacteria can develop resistance, but phages may co-evolve	Resistance is a growing issue, and development of new antibiotics is slow
Impact on microbiota	Minimal disruption to beneficial microbiota	Can disrupt gut flora, leading to dysbiosis or secondary infections like Clostridioides difficile infection
Replication in host	Multiplies at the infection site if host bacteria are present	Does not self-replicate; efficacy depends on dosage
Immunogenicity	May trigger immune response, especially with repeated use	Less likely to elicit strong immune reactions
Production and customization	Can be tailored to target specific pathogens	Mass-produced with fixed formulations
Environmental impact	Generally considered eco-friendly	Overuse can contribute to antibiotic resistance and gut dysbiosis

Phage therapy has demonstrated even greater therapeutic potential when used in conjugation with antibiotics, especially in managing complex persistent infections[38]. A major challenge in treating bacterial infections is the formation of biofilm. Phages have the unique ability to penetrate and disrupt these biofilms, helping to decrease bacterial load and making the remaining bacteria more vulnerable to antibiotics. This synergistic interaction has been supported by multiple studies[39,40]. For example, in cases of chronic respiratory infections caused by Pseudomonas aeruginosa (known for its resilient biofilm structures), combined phage-antibiotic therapy has resulted in marked improvements in patient outcomes[41,42]. Such a synergistic approach is especially beneficial in difficult-to-treat infections involving high bacterial loads or biofilm-associated resistance.

While phage therapy offers a promising role in combating bacterial infections, it is not completely immune to the problem of resistance. Similar to how bacteria can become resistant to antibiotics, they can also evolve defence mechanisms against phages. Mechanism of this resistance is the alterations in the bacterial surface receptors that phages use for attachment and entry, or the activation of innate defence systems like Clustered Regularly Interspersed Short Palindromic Repeats—CRISPR-associated protein (CRISPR-Cas), which enables bacteria to recognize and neutralize phage DNA[43].

However, unlike antibiotic resistance, resistance to phages can sometimes be temporary or reversible. One effective approach to counteract phage resistance is the use of phage cocktails, combinations of multiple phages that target the same bacterial species but use different receptors or pathways. This cocktail attack makes it harder for bacteria to develop resistance to all phages simultaneously. Another emerging tactic involves the genetic modification of phages, allowing them to adapt by changing their binding sites or bypassing bacterial defence systems. These engineered phages can potentially infect bacterial strains that have developed resistance to natural phages. While these strategies show considerable promise, they are still under development and face multiple challenges like regulatory hurdles and issues with production (Table 4)[36,45,46]. Further, research is required to optimize their safety and effectiveness in clinical settings[44].

Table 4 Challenges and potential solutions in phage therapy.

Challenge	Description	Potential solutions
Narrow host range	Phages are highly specific, targeting only a limited range of bacterial strains	Develop phage cocktails targeting multiple strains
Bacterial resistance development	Bacteria may evolve resistance to phages, just like with antibiotics	Using phage combinations or cocktails; engineer phages to overcome resistance
Immunogenicity of phages	The human immune system may neutralize phages, limiting their effectiveness	Use encapsulation techniques (e.g., liposomes) to shield phages; select less immunogenic phages
Phage clearance by organs	The liver and spleen may rapidly clear phages from the bloodstream	Modifications in phages to evade immune detection; optimize dosing regimens to maintain therapeutic levels
Horizontal gene transfer risk	Temperate phages can transfer harmful genes (e.g., toxin or resistance genes) between bacteria	Prefer strictly lytic phages over temperate ones; genetically screen and engineer phages for safety
Storage and stability	Phages can lose viability due to improper storage conditions	Optimize formulations and storage conditions (e.g., lyophilization, buffer systems) for stability
Lack of standardized regulations	Absence of global guidelines makes approval and clinical use challenging	Developing common regulatory frameworks; global collaborations on phage therapy policies
Limited clinical trials	Few large-scale, randomized controlled trials exist to validate efficacy and safety	Encourage funding and support for rigorous clinical studies to build evidence for medical approval

CRISPR-Cas systems

The CRISPR-Cas system is a natural bacterial immune mechanism, that provides adaptive defence against invasion of viruses (bacteriophages) and plasmids. It is gradually emerging as a promising tool in the fight against MDR organisms[47]. One of its most remarkable features is the ability to selectively target and destroy specific DNA sequences, including those responsible for antibiotic resistance. CRISPR-Cas systems can recognize and cut resistance genes, such as mecA, or vanA and selectively eliminate resistant bacteria or deactivate the resistance genes, which then re-sensitizes these pathogens to previously resistant antibiotics. CRISPR can also target plasmids carrying resistance genes and block their spread among bacterial populations, hence disrupting the horizontal gene transfer that amplifies resistance[48]. Another major advantage of CRISPR-based antimicrobials is their narrow spectrum of activity, which allows them to preserve the beneficial components of the microbiome, reducing the risk of dysbiosis or opportunistic infections such as Clostridioides difficile.

Delivery methods currently under investigation include bacteriophage-based systems (phagemids), conjugative plasmids, and nanoparticles. Despite its promise, the use of CRISPR in clinical settings is still in the early stages. However, with continued innovation, CRISPR-Cas technology could revolutionize our approach to treating antibiotic-resistant infections by offering targeted, potent, and microbiome-sparing therapeutic options[36] (Table 5).

Table 5 Summary of the methods for gut decolonization.

Method	Mechanism/strategy	Current status/potential	Limitations
Synbiotics (Prebiotics + Probiotics)	Combination approach to feed beneficial bacteria and inhibit pathogen growth	Evidence for Clostridioides difficile; under evaluation for broader MDRO decolonization	Strain/pathogen-specific efficacy; low to moderate benefits only; inconsistent results across trials
Live biotherapeutic products	Consortia of beneficial bacteria designed to displace pathogens and modulate immunity	Examples include VE707; under investigation for MDROs	Mostly preclinical; unclear long-term effects; regulatory challenges for approval
Selective digestive decontamination	Use of non-absorbable oral and systemic antibiotics to decolonize gut MDROs	Used in ICU settings; variable evidence; some success in ESBL-E decolonization	Promotes resistance (e.g., colistin-resistant strains); disrupts microbiota; lacks standardized protocols
Fecal microbiota transplantation	May restore healthy microbiota to compete MDROs	Successful in small studies; ongoing trials for CRE, VRE. Requires standardization and safety screening	Risk of transferring ARGs; labor-intensive donor screening; cold-chain dependency
Bacteriophage therapy	Phages specifically target and lyse pathogenic strains without affecting commensals	Promising; several preclinical and early clinical studies underway. Challenges include phage resistance and regulatory issues	Narrow host range; potential phage resistance; immunogenicity; storage and regulatory hurdles
CRISPR-Cas system	Use of gene-editing tools to selectively target and eliminate resistance genes	Under experiment, promising specificity and minimal off-target effects	Delivery method challenges; early-stage development; ethical and safety concerns

MDRO: Multi-drug resistant organisms; ICU: Intensive care unit; ESBL: Extended spectrum beta-lactamases; CRE: Carbapenem resistant enterobacteriaceae; VRE: Vancomycin resistant enterococcus; ARG: Antibiotic resistant.

CONCLUSION

As the burden of AMR continues to grow, innovative microbiota-targeted strategies have become increasingly critical in the fight against MDR bacteria colonization. Evidence supports the role of gut dysbiosis in fostering resistant strains and dietary interventions may serve as supportive strategies to enhance the resilience of the host microbiota. The future lies in emerging interventions such as phage therapy, engineered probiotics, and CRISPR-Cas systems that offer new hope for precise and sustainable decolonization. However, many of these approaches are still in the initial stages of development, and their long-term efficacy, safety, and regulatory approval remain areas of ongoing research. In the future, an integrated and multi-step approach that combines microbiome engineering, personalized therapeutics, and public health measures may provide the most effective strategy for reducing the prevalence of MDROs and preventing the development of resistant infections. To realize this potential, there is a need for investment in translational research, the development of standardized clinical protocols, and the establishment of clear regulatory pathways to guide implementation in routine care.