Tuberculosis has been reaffirmed as the infectious disease causing most deaths in the world. Co-infection with HIV and the increase in multi-drug resistant Mycobacterium tuberculosis strains complicate treatment and increases mortality rates, making the development of new drugs an urgent priority. In this study we have identified a promising candidate by screening antimicrobial peptides for their capacity to inhibit mycobacterial growth. This non-toxic peptide, NZX, is capable of inhibiting both clinical strains of M. tuberculosis and an MDR strain at therapeutic concentrations. The therapeutic potential of NZX is further supported in vivo where NZX significantly lowered the bacterial load with only five days of treatment, comparable to rifampicin treatment over the same period. NZX possesses intracellular inhibitory capacity and co-localizes with intracellular bacteria in infected murine lungs. In conclusion, the data presented strongly supports the therapeutic potential of NZX in future anti-TB treatment.

Background Intracellular delivery of antimicrobial agents by nanoparticles, such as mesoporous silica particles (MSPs), offers an interesting strategy to treat intracellular infections. In tuberculosis (TB), Mycobacterium tuberculosis avoids components of the immune system by residing primarily inside alveolar macrophages, which are the desired target for TB therapy. Methods and findings We have previously identified a peptide, called NZX, capable of inhibiting both clinical and multi-drug resistant strains of M . tuberculosis at therapeutic concentrations. In this study we analysed the potential of MSPs containing NZX for the treatment of tuberculosis. The MSPs released functional NZX gradually into simulated lung fluid and the peptide filled MSPs were easily taken up by primary macrophages. In an intracellular infection model, the peptide containing particles showed increased mycobacterial killing compared to free peptide. The therapeutic potential of peptide containing MSPs was investigated in a murine infection model, showing that MSPs preserved the effect to eliminate M . tuberculosis in vivo . Conclusions In this study we found that loading the antimicrobial peptide NZX into MSPs increased the inhibition of intracellular mycobacteria in primary macrophages and preserved the ability to eliminate M . tuberculosis in vivo in a murine model. Our studies provide evidence for the feasibility of using MSPs for treatment of tuberculosis.

Epidemiological contact tracing complemented with genotyping of clinical Mycobacterium tuberculosis isolates is important for understanding disease transmission. In Sweden, tuberculosis (TB) is mostly reported in migrant and homeless where epidemiologic contact tracing could pose a problem. This study compared epidemiologic linking with genotyping in a low burden country. Mycobacterium tuberculosis isolates (n = 93) collected at Scania University Hospital in Southern Sweden were analysed with the standard genotyping method mycobacterial interspersed repetitive units-variable number tandem repeats (MIRU-VNTR) and the results were compared with whole genome sequencing (WGS). Using a maximum of twelve single nucleotide polymorphisms (SNPs) as the upper threshold of genomic relatedness noted among hosts, we identified 18 clusters with WGS comprising 52 patients with overall pairwise genetic maximum distances ranging from zero to nine SNPs. MIRU-VNTR and WGS clustered the same isolates, although the distribution differed depending on MIRU-VNTR limitations. Both genotyping techniques identified clusters where epidemiologic linking was insufficient, although WGS had higher correlation with epidemiologic data. To summarize, WGS provided better resolution of transmission than MIRU-VNTR in a setting with low TB incidence. WGS predicted epidemiologic links better which could consolidate and correct the epidemiologically linked cases, avoiding thus false clustering.

Mycobacterium bovis bacille Calmette-Guérin (BCG) is currently the only approved vaccine against tuberculosis (TB). BCG mimics M. tuberculosis (Mtb) in its persistence in the body and is used as a benchmark to compare new vaccine candidates. BCG was originally designed for mucosal vaccination, but comprehensive knowledge about its interaction with epithelium is currently lacking. We used primary airway epithelial cells (AECs) and a murine model to investigate the initial events of mucosal BCG interactions. Furthermore, we analysed the impact of the G-protein-coupled receptors (GPCRs), CXCR1 and CXCR2, in this process, as these receptors were previously shown to be important during TB infection. BCG infection of AECs induced GPCR-dependent Rac1 up-regulation, resulting in actin redistribution. Thealtered distribution of the actin cytoskeleton involved the MAPK signalling pathway. Blocking of the CXCR1 or CXCR2 prior to infection decreased Rac1 expression, and increased epithelial transcriptional activity and epithelial cytokine production. BCG infection did not result in epithelial cell death as measured by p53 phosphorylation and annexin. This study demonstrated that BCG infection of AECs manipulated the GPCRs to suppress epithelial signalling pathways. Future vaccine strategies could thus be improved by targeting GPCRs.

Much of the pronounced host inflammatory response that occurs in tuberculosis (TB) is related to failed immunity against the invading pathogen. The G-protein coupled receptors CXCR1 and CXCR2 are implicated in important signal transduction pathways in lung inflammatory responses. We investigated the expression and function of these receptors in a simple whole blood model from 24 patients with pulmonary TB and in subjects with latent TB infection (LTBI). Healthy controls were recruited from close contacts to the pulmonary index patients. We found that pulmonary TB patients had significantly increased CXCR1 expression on blood cells compared to LTBI subjects and controls (p < 0.001). In contrast, LTBI subjects had a significant increase in CXCR2 expression compared to pulmonary TB patients (p < 0.001) and controls (p < 0.01). Leukocyte function, measured as oxidative capacity, was decreased in pulmonary TB patients compared to LTBI and controls (p < 0.001) and correlated with the increased CXCR1 expression. Leukocyte recruitment, measured as the expression of microRNA-223 was increased in pulmonary TB patients compared to LTBI (p < 0.05). We found that variations in receptor expression are linked to disease progression and affect the immune response against Mycobacterium tuberculosis (Mtb).