Enhancing the stability and therapeutic potential of the antimicrobial peptide Feleucin-K3 against Multidrug-Resistant a. Baumannii through rational utilization of a D-amino acid substitution strategy.

Antimicrobial peptides (AMPs), which have a low probability of developing resistance, are considered the most promising antimicrobial agents for combating antibiotic resistance. Feleucin-K3 is an amphiphilic cationic AMP that exhibits broad-spectrum antimicrobial activity. In our previous research, the first phenylalanine residue was identified as the critical position affecting its biological activity. Here, a series of Feleucin-K3 analogs containing hydrophobic D-amino acids were developed, leveraging the low sensitivity of proteases to unnatural amino acids and the regulatory effect of hydrophobicity on antimicrobial activity. Among them, K-1dF, which replaced the phenylalanine of Feleucin-K3 with its enantiomer (D-phenylalanine), exhibited potent antimicrobial activity with a therapeutic index of 46.97 and MICs between 4 to 8 µg/ml against both sensitive and multidrug-resistant Acinetobacter baumannii. The introduction of D-phenylalanine increased the salt tolerance and serum stability of Feleucin-K3. Moreover, K-1dF displayed a rapid bactericidal effect, a low propensity to develop resistance, and a synergistic effect when combined with antibiotics. More importantly, it exhibited considerable or superior efficacy to imipenem against pneumonia and skin abscess infection. In brief, the K-1dF obtained by simple and effective modification strategy has emerged as a promising candidate antimicrobial agent for tackling multidrug-resistant Acinetobacter baumannii infections.

Multidrug-resistant Pseudomonas aeruginosa is predisposed to lasR mutation through up-regulated activity of efflux pumps in non-cystic fibrosis bronchiectasis patients.

Background: Multidrug-resistant (MDR) Pseudomonas aeruginosa is a frequent opportunistic pathogen that causes significant mortality in patients with non-cystic fibrosis bronchiectasis (NCFB). Although the quorum sensing (QS) system is a potential target for treatment, lasR mutants that present with a QS-deficient phenotype have been frequently reported among clinical P. aeruginosa isolates. We aimed to investigate whether antibiotic resistance would select for lasR mutants during chronic P. aeruginosa lung infection and determine the mechanism underlying the phenomenon. Methods: We prospectively evaluated episodes of chronic P. aeruginosa lung infections in NCFB patients over a 2-year period at two centers of our institution. QS phenotypic assessments and whole-genome sequencing (WGS) of P. aeruginosa isolates were performed. Evolution experiments were conducted to confirm the emergence of lasR mutants in clinical MDR P. aeruginosa cultures. Results: We analyzed episodes of P. aeruginosa infection among 97 NCFB patients and found only prior carbapenem exposure independently predictive of the isolation of MDR P. aeruginosa strains. Compared with non-MDR isolates, MDR isolates presented significantly QS-deficient phenotypes, which could not be complemented by the exogenous addition of 3OC12-HSL. The paired isolates showed that their QS-phenotype deficiency occurred after MDR was developed. Whole-genome sequencing analysis revealed that lasR nonsynonymous mutations were significantly more frequent in MDR isolates, and positive correlations of mutation frequencies were observed between genes of lasR and negative-efflux-pump regulators (nalC and mexZ). The addition of the efflux pump inhibitor PAßN could not only promote QS phenotypes of these MDR isolates but also delay the early emergence of lasR mutants in evolution experiments. Conclusions: Our data indicated that MDR P. aeruginosa was predisposed to lasR mutation through the upregulated activity of efflux pumps. These findings suggest that anti-QS therapy combined with efflux pump inhibitors might be a potential strategy for NCFB patients in the challenge of MDR P. aeruginosa infections.

Overcoming the rising incidence and evolving mechanisms of antibiotic resistance by novel drug delivery approaches - An overview.

Antimicrobial resistance is a normal evolutionary process for microorganisms. Antibiotics exerted accelerated selective pressure that hasten bacterial resistance through mutation, and acquisition external genes. These genes often carry multiple antibiotic resistant determinants allowing the recipient microbe an instant "super-bug" status. The extent of Antimicrobial Resistance (AMR) has reached a level of global crisis, existing antimicrobials are no long effective in treating infections caused by AMR pathogens. The great majority of clinically available antimicrobial agents are administered through oral and intra-venous routes. Overcoming antibacterial resistance by novel drug delivery approach offered new hopes, particularly in the treatment of AMR pathogens in sites less assessible through systemic circulation such as the lung and skin. In the current review, we will revisit the mechanism and incidence of important AMR pathogens. Finally, we will discuss novel drug delivery approaches including novel local antibiotic delivery systems, hybrid antibiotics, and nanoparticle-based antibiotic delivery systems.

The Epidemiology and Pathogenesis and Treatment of Pseudomonas aeruginosa Infections: An Update.

Pseudomonas aeruginosa is a Gram-negative bacterial pathogen that is a common cause of nosocomial infections, particularly pneumonia, infection in immunocompromised hosts, and in those with structural lung disease such as cystic fibrosis. Epidemiological studies have identified increasing trends of antimicrobial resistance, including multi-drug resistant (MDR) isolates in recent years. P. aeruginosa has several virulence mechanisms that increase its ability to cause severe infections, such as secreted toxins, quorum sensing and biofilm formation. Management of P. aeruginosa infections focuses on prevention when possible, obtaining cultures, and prompt initiation of antimicrobial therapy, occasionally with combination therapy depending on the clinical scenario to ensure activity against P. aeruginosa. Newer anti-pseudomonal antibiotics are available and are increasingly being used in the management of MDR P. aeruginosa.

Antibiotic Susceptibility of Staphylococcus aureus Plankton and Biofilm Forms Isolated in Implant-Associated Infection.

Comparison of activity of antibiotics against Staphylococcus aureus strains in their plankton form and in biofilms of varying maturity showed that the concentrations of antibiotics causing death of 90% S. aureus strains (MIC90) in their plankton form was 0.07-2.80 µg/ml and significantly (p<0.05) differed from MIC90 concentrations (3-245 µg/ml) for the strains in 24 and 48 h biofilms. MIC90 value was affected by the degree of biofilm maturity: microbial resistance in 48-h biofilms for all analyzed antibiotics was significantly (p<0.05) higher than in 24-h biofilms.

Nanomedicine to fight infectious disease.

The ubiquity and potency of antibiotics may give the false impression that infection is a solved problem. Unfortunately, even bacterial infections, the target of antibiotics, remain a major cause of illness and death. Several major unmet needs persist: biofilms, such as those on implanted hardware, largely resist antibiotics; the inflammatory host response to infection often produces more damage than the infection itself; and systemic antibiotics often decimate the gut microbiome, which can predispose to additional infections and even predispose to non-infectious diseases. Additionally, there is an increasing threat from multi-drug resistant microorganisms, though market forces may continue to inhibit innovation in this realm. These numerous unmet infection-related needs provide attractive goals for innovation of targeted drug delivery technologies, especially those of nanomedicine. Here we review several of those innovations in pre-clinical development, the two such therapies which have made it to clinical use, and the opportunities for further technology development for treating infections.

Phloretin potentiates polymyxin E activity against gram-negative bacteria.

AIMS: The spread of plasmid-mediated polymyxin resistance has jeopardized the use of polymyxin, the last defender that combats infections caused by multidrug-resistant (MDR) gram-negative pathogens. MAIN METHODS: In this study, phloretin, as a monomeric compound extracted from natural plants, showed a good synergistic effect with polymyxin E against gram-negative bacteria, as evaluated by minimal inhibit concentration (MIC) assay and a series of assays, including growth curve, time-killing, and Western blot assays. A model of mice infected by Salmonella sp. stain HYM2 was established to further identify the synergistic effect of phloretin with polymyxin E. KEY FINDINGS: The results suggested that phloretin had the potential ability to recover the antibacterial sensitivity of polymyxin E from 64 µg/mL to no more than 2 µg/mL in E. coli ZJ478 or in Salmonella sp. stain HYM2 with a 32-fold decrease. A series of strains, including mcr-1-positive and mcr-1-negative strains, were treated with a combination of phloretin and polymyxin E, and the fractional inhibitory concentration (FIC) values were all found to be below 0.5. However, the combination of phloretin and polymyxin E did not lead to bacterial resistance. In vivo, the survival rate of infected mice reached nearly 80% with the combination treatment, and the cecal colony value also decreased significantly. SIGNIFICANCE: All the above results indicated that phloretin is a potential polymyxin potentiator to combat gram-negative stains.

Plant-derived nanotherapeutic systems to counter the overgrowing threat of resistant microbes and biofilms.

Since antiquity, the survival of human civilization has always been threatened by the microbial infections. An alarming surge in the resistant microbial strains against the conventional drugs is quite evident in the preceding years. Furthermore, failure of currently available regimens of antibiotics has been highlighted by the emerging threat of biofilms in the community and hospital settings. Biofilms are complex dynamic composites rich in extracellular polysaccharides and DNA, supporting plethora of symbiotic microbial life forms, that can grow on both living and non-living surfaces. These enforced structures are impervious to the drugs and lead to spread of recurrent and non-treatable infections. There is a strong realization among the scientists and healthcare providers to work out alternative strategies to combat the issue of drug resistance and biofilms. Plants are a traditional but rich source of effective antimicrobials with wider spectrum due to presence of multiple constituents in perfect synergy. Other than the biocompatibility and the safety profile, these phytochemicals have been repeatedly proven to overcome the non-responsiveness of resistant microbes and films via multiple pathways such as blocking the efflux pumps, better penetration across the cell membranes or biofilms, and anti-adhesive properties. However, the unfavorable physicochemical attributes and stability issues of these phytochemicals have hampered their commercialization. These issues of the phytochemicals can be solved by designing suitably constructed nanoscaled structures. Nanosized systems can not only improve the physicochemical features of the encapsulated payloads but can also enhance their pharmacokinetic and therapeutic profile. This review encompasses why and how various types of phytochemicals and their nanosized preparations counter the microbial resistance and the biofouling. We believe that phytochemical in tandem with nanotechnological innovations can be employed to defeat the microbial resistance and biofilms. This review will help in better understanding of the challenges associated with developing such platforms and their future prospects.

Upward trends in new, rifampicin-resistant and concurrent extrapulmonary tuberculosis cases in northern Guizhou Province of China.

Similar to global trends, the incidence rate of tuberculosis (TB) in China declined from 2000 to 2018. In this study, we aimed to evaluate TB trends in northern Guizhou Province and identify risk factors associated with rifampicin-resistant (RR) and concurrent extrapulmonary TB (EPTB). We analyzed data of TB patients hospitalized in Affiliated Hospital of Zunyi Medical University from 2011 to 2018, and assessed correlations between demographic characteristics of patients and RR-TB as well as concurrent EPTB. Our results showed that numbers of new, retreated, RR-TB and concurrent EPTB cases increased gradually from 2011 to 2018. Retreated patients had the highest odds of RR-TB but a lower likelihood of concurrent EPTB compared to new patients. Patients between 21 and 40 years of age had a higher likelihood of RR-TB compared to those 20 years and younger. Female patients and patients from Bijie city as well as the Miao ethnic minority had higher odds of concurrent EPTB. In summary, our data demonstrate upward trends in new, rifampicin-resistant and concurrent extrapulmonary TB cases in northern Guizhou Province of China, which should not be overlooked especially during and post the COVID-19 pandemic because TB is a greater long-term global health threat than COVID-19.

Elucidating the mechanism by which synthetic helper peptides sensitize Pseudomonas aeruginosa to multiple antibiotics.

The emergence and rapid spread of multi-drug resistant (MDR) bacteria pose a serious threat to the global healthcare. There is an urgent need for new antibacterial substances or new treatment strategies to deal with the infections by MDR bacterial pathogens, especially the Gram-negative pathogens. In this study, we show that a number of synthetic cationic peptides display strong synergistic antimicrobial effects with multiple antibiotics against the Gram-negative pathogen Pseudomonas aeruginosa. We found that an all-D amino acid containing peptide called D-11 increases membrane permeability by attaching to LPS and membrane phospholipids, thereby facilitating the uptake of antibiotics. Subsequently, the peptide can dissipate the proton motive force (PMF) (reducing ATP production and inhibiting the activity of efflux pumps), impairs the respiration chain, promotes the production of reactive oxygen species (ROS) in bacterial cells and induces intracellular antibiotics accumulation, ultimately resulting in cell death. By using a P. aeruginosa abscess infection model, we demonstrate enhanced therapeutic efficacies of the combination of D-11 with various antibiotics. In addition, we found that the combination of D-11 and azithromycin enhanced the inhibition of biofilm formation and the elimination of established biofilms. Our study provides a realistic treatment option for combining close-to-nature synthetic peptide adjuvants with existing antibiotics to combat infections caused by P. aeruginosa.

Host Colonization as a Major Evolutionary Force Favoring the Diversity and the Emergence of the Worldwide Multidrug-Resistant Escherichia coli ST131.

The emergence of multidrug-resistant Escherichia coli ST131 is a major worldwide public health problem in humans. According to the "one health" approach, this study investigated animal reservoirs of ST131, their relationships with human strains, and the genetic features associated with host colonization. High-quality genomes originating from human, avian, and canine hosts were classified on the basis of their accessory gene content using pangenomic. Pangenomic clusters and subclusters were specifically and significantly associated with hosts. The functions of clustering accessory genes were mainly enriched in functions involved in DNA acquisition, interactions, and virulence (e.g., pathogenesis, response to biotic stimulus and interaction between organisms). Accordingly, networks of cooccurrent host interaction factors were significantly associated with the pangenomic clusters and the originating hosts. The avian strains exhibited a specific content in virulence factors. Rarely found in humans, they corresponded to pathovars responsible for severe human infections. An emerging subcluster significantly associated with both human and canine hosts was evidenced. This ability to significantly colonize canine hosts in addition to humans was associated with a specific content in virulence factors (VFs) and metabolic functions encoded by a new pathogenicity island in ST131 and an improved fitness that is probably involved in its emergence. Overall, VF content, unlike the determinants of antimicrobial resistance, appeared as a key actor of bacterial host adaptation. The host dimension emerges as a major driver of genetic evolution that shapes ST131 genome, enhances its diversity, and favors its dissemination. IMPORTANCE Until now, there has been no indication that the evolutionary dynamics of Escherichia coli ST131 may reflect independent and host-specific adaptation of this lineage outside humans. In contrast, the limited number of ST131 reports in animals supported the common view that it rather reflects a spillover of the human sector. This study uncovered a link between host, ST131 population structure, and virulence factor content which appeared to reflect adaptation to hosts. This study helps to better understand the reservoir of ST131, the putative transmission flux, associated risks and the evolutionary dynamics of this bacterial population and highlights a paradigm in which host colonization stands as a key ecological force of the ST131 evolution.

An insight into the implementation of the global action plan on antimicrobial resistance in the WHO African region: A roadmap for action.

INTRODUCTION: The Global Action Plan (GAP) on antimicrobial resistance (AMR) delivers a 'One Health' strategy for the development of the national action plan. It encourages the optimal use of antimicrobials and strengthens the evidence base through surveillance and research. METHODS: This study evaluated the current status of implementation of the GAP on AMR in World Health Organization (WHO) African countries via a retrospective, cross-sectional analysis of routinely collected data on AMR. A SWOT analysis was used to identify the strengths, weaknesses, opportunities and threats involved in the GAP implementation. A roadmap for action was proposed. RESULTS: The overall mean GAP performance score across all the countries that were assessed was 32% ± SD12 (95% CI 27-36%). The mean thematic scores were 59% ± 12 (53-65%) for multi-sector and One Health collaboration; 50% ± 22 (42-58%) for developing national AMR action plans; 38% ± 12 (33-42%) for awareness and training; 18% ± 13 (13-23%) for surveillance; 33% ± 13 (29-38%) for infection prevention and control; and 28% ± 23 (20-37%) for optimal use of antimicrobial medicines in human, animal and plant health. The difference in GAP performance scores between African sub-regions and between income categories was not statistically significant (P > 0.05). While seven countries exhibited strengths in two themes, 25 countries exhibited weaknesses across all themes. Six threats and six opportunities were identified to inform a practical roadmap for AMR action. CONCLUSION: The findings from this study indicate that the overall GAP implementation on AMR in the WHO African region is inadequate. Some thematic GAP scores appeared to be relatively good, but on closer inspection, individual indicators revealed a lack of progress and implementation, requiring action.

Prevalence of CTX-M ß-Lactamases Producing Multidrug Resistant Escherichia coli and Klebsiella pneumoniae among Patients Attending Bir Hospital, Nepal.

The emergence of multidrug resistant (MDR) bacteria which is attributable to extended spectrum ß-lactamases (ESBLs) production of CTX-M types is an obvious problem worldwide. This study is aimed at determining the prevalence of CTX-M ß-lactamases producing multidrug resistant Escherichia coli and Klebsiella pneumoniae among patients attending Bir Hospital. A cross-sectional study was conducted between April and September 2019 at Bir Hospital, Kathmandu, and Department of Microbiology, National College, Kathmandu, Nepal. A total of 5,690 different clinical specimens were subjected to cultural, microscopic, and biochemical analyses for the identification of the isolates. Antimicrobial susceptibility testing of the isolates was done, and MDR isolates were selected and processed for further ESBL confirmation by the combination disks method. All confirmed ESBL isolates were screened for CTX-M type ß-lactamases (bla CTX-M) by PCR. Of the total 345 isolates (227 Escherichia coli and 118 Klebsiella pneumoniae), 232 were MDR. All 232 (67.24%) MDR isolates were suspected as ESBL producers on the screening test. However, on the phenotypic test, 135 (58.18%) of total MDR bacteria were confirmed as ESBL producers with the highest proportion in K. pneumoniae (59.37%). The major source of ESBL producers was urine. ESBL producing isolates were mostly identified from outpatients and patients belonging to age group 41-60. Gentamicin was found to be effective against ESBL producers. The prevalence of bla CTX-M was (89.62%) with the highest frequency for E. coli (93.81%). High prevalence of ESBL of CTX-M types among MDR E. coli and K. pneumoniae was detected from clinical specimens of patients in Bir Hospital. This study warrants the need for the judicious use of antibiotics as well as emphasize the use of modern diagnostic tools for the early detection of MDR and ESBL producers to curb the emergence and spread of MDR and ESBL producing bacteria in the clinical settings of Nepal.

Prevalence and risk factors for faecal carriage of multidrug resistant Escherichia coli among slaughterhouse workers.

The increasing occurrence of antimicrobial-resistant Escherichia coli in human and animal population has become a global public health problem that requires immediate intervention. We aimed to investigate prevalence and risk factors for faecal carriage of drug-resistant E. coli among slaughterhouse workers. We conducted this cross-sectional study among 118 apparently healthy workers in the largest slaughterhouses in Abuja and Lagos from July to December 2020. E. coli was isolated from stool samples of slaughterhouse workers and antimicrobial susceptibility testing performed using the Kirby-Bauer disk diffusion method. Multi-drug resistance (MDR) was defined as resistance to three or more classes of antibiotics. Majority were males: 88.1% (n = 104), aged > 41 years: 28.8% (n = 34), married: 70.3% (n = 83), and were butchers: 53.4% (n = 63). Prevalence of MDR E. coli was 50% (n = 59), highest among butchers compared to slaughterhouse cleaners. Of 75 E. coli isolates identified, 25.3% (n = 19) were ESBL producers; 78.7% (n = 59) were MDR. Keeping animals (p = 0.01); eating at the slaughterhouse (p = 0.03) and collecting waste (p = 0.02) remained independent risk factors for acquiring MDR E. coli. Prevalence of resistant E. coli was highest among butchers and associated with keeping animals at home, eating at work, and waste-collection. Hand-hygiene and responsible use of antibiotics among slaughterhouse workers should be encouraged.

Roles of Multidrug Resistance Protein 4 in Microbial Infections and Inflammatory Diseases.

Numerous studies have reported the emergence of antimicrobial resistance during the treatment of common infections. Multidrug resistance (MDR) leads to failure of antimicrobial treatment, prolonged illness, and increased morbidity and mortality. Overexpression of multidrug resistance proteins (MRPs) as drug efflux pumps are one of the main contributions of MDR, especially multidrug resistance protein 4 (MRP4/ABCC4) in the development of antimicrobial resistance. The molecular mechanism of antimicrobial resistance is still under investigation. Various intervention strategies have been developed for overcoming MDR, but the effect is limited. Suppression of MRP4 may be an attractive therapeutic approach for addressing drug resistance. However, there are few reports on the involvement of MRP4 in antimicrobial resistance and inflammatory diseases. In this review, we introduced the function and regulation of MRP4, and then summarized the roles of MRP4 in microbial infections and inflammatory diseases as well as polymorphisms in the gene encoding this transporter. Further studies should be conducted on drug therapy targeting MRP4 to improve the efficacy of antimicrobial therapy. This review can provide useful information on MRP4 for overcoming antimicrobial resistance and anti-inflammatory therapy.

Genotypic and phenotypic profiles of antibiotic-resistant bacteria isolated from hospitalised patients in Bangladesh.

OBJECTIVES: Characterisation of resistance phenotype and genotype is crucial to understanding the burden and transmission of antimicrobial resistance (AMR). This study aims to determine the spectrum of AMR and associated genes encoding aminoglycoside, macrolide and ß-lactam classes of antimicrobials in bacteria isolated from hospitalised patients in Bangladesh. METHODS: 430 bacterial isolates from patients with respiratory, intestinal, wound infections and typhoid fever, presenting to clinical care from 2015 to 2019, were examined. They included Escherichia coli (n = 85); Staphylococcus aureus (n = 84); Salmonella typhi (n = 82); Klebsiella pneumoniae (n = 42); Streptococcus pneumoniae (n = 36); coagulase-negative staphylococci (n = 28); Enterococcus faecalis (n = 27); Pseudomonas aeruginosa (n = 26); and Acinetobacter baumannii (n = 20). Reconfirmation of these clinical isolates and antimicrobial susceptibility tests was performed. PCR amplification using resistance gene-specific primers was done, and the amplified products were confirmed by Sanger sequencing. RESULTS: 53% of isolates were multidrug-resistant (MDR), including 97% of Escherichia coli. There was a year-wise gradual increase in MDR isolates from 2015 to 2018, and there was an almost twofold increase in the number of MDR strains isolated in 2019 (P = 0.00058). Among the 5 extended-spectrum ß-lactamases investigated, CTX-M-1 was the most prevalent (63%) followed by NDM-1 (22%); Escherichia coli was the major reservoir of these genes. The ermB (55%) and aac(6')-Ib (35%) genes were the most frequently detected macrolide and aminoglycoside resistance genes, respectively. CONCLUSION: MDR pathogens are highly prevalent in hospital settings of Bangladesh.

The Challenge of Global Emergence of Novel Colistin-Resistant Escherichia coli ST131.

Escherichia coli ST131 is one of the high-risk multidrug-resistant clones with a global distribution and the ability to persist and colonize in a variety of niches. Carbapenemase-producing E. coli ST131 strains with the ability to resist last-line antibiotics (i.e., colistin) have been recently considered a significant public health. Colistin is widely used in veterinary medicine and therefore, colistin-resistant bacteria can be transmitted from livestock to humans through food. There are several mechanisms of resistance to colistin, which include chromosomal mutations and plasmid-transmitted mcr genes. E. coli ST131 is a great model organism to investigate the emergence of superbugs. This microorganism has the ability to cause intestinal and extraintestinal infections, and its accurate identification as well as its antibiotic resistance patterns are vitally important for a successful treatment strategy. Therefore, further studies are required to understand the evolution of this resistant organism for drug design, controlling the evolution of other nascent emerging pathogens, and developing antibiotic stewardship programs. In this review, we will discuss the importance of E. coli ST131, the mechanisms of resistance to colistin as the last-resort antibiotic against resistant Gram-negative bacteria, reports from different regions regarding E. coli ST131 resistance to colistin, and the most recent therapeutic approaches against colistin-resistance bacteria.

Drug repurposing for next-generation combination therapies against multidrug-resistant bacteria.

Antimicrobial resistance has been a global health challenge that threatens our ability to control and treat life-threatening bacterial infections. Despite ongoing efforts to identify new drugs or alternatives to antibiotics, no new classes of antibiotic or their alternatives have been clinically approved in the last three decades. A combination of antibiotics and non-antibiotic compounds that could inhibit bacterial resistance determinants or enhance antibiotic activity offers a sustainable and effective strategy to confront multidrug-resistant bacteria. In this review, we provide a brief overview of the co-evolution of antibiotic discovery and the development of bacterial resistance. We summarize drug-drug interactions and uncover the art of repurposing non-antibiotic drugs as potential antibiotic adjuvants, including discussing classification and mechanisms of action, as well as reporting novel screening platforms. A pathogen-by-pathogen approach is then proposed to highlight the critical value of drug repurposing and its therapeutic potential. Finally, general advantages, challenges and development trends of drug combination strategy are discussed.

Biofilm formation in the lung contributes to virulence and drug tolerance of Mycobacterium tuberculosis.

Tuberculosis is a chronic disease that displays several features commonly associated with biofilm-associated infections: immune system evasion, antibiotic treatment failures, and recurrence of infection. However, although Mycobacterium tuberculosis (Mtb) can form cellulose-containing biofilms in vitro, it remains unclear whether biofilms are formed during infection in vivo. Here, we demonstrate the formation of Mtb biofilms in animal models of infection and in patients, and that biofilm formation can contribute to drug tolerance. First, we show that cellulose is also a structural component of the extracellular matrix of in vitro biofilms of fast and slow-growing nontuberculous mycobacteria. Then, we use cellulose as a biomarker to detect Mtb biofilms in the lungs of experimentally infected mice and non-human primates, as well as in lung tissue sections obtained from patients with tuberculosis. Mtb strains defective in biofilm formation are attenuated for survival in mice, suggesting that biofilms protect bacilli from the host immune system. Furthermore, the administration of nebulized cellulase enhances the antimycobacterial activity of isoniazid and rifampicin in infected mice, supporting a role for biofilms in phenotypic drug tolerance. Our findings thus indicate that Mtb biofilms are relevant to human tuberculosis.