A multifaceted interplay between virulence, drug resistance, and the phylogeographic landscape of Mycobacterium tuberculosis.

Latin-American Mediterranean (LAM) family is one of the most significant and global genotypes of Mycobacterium tuberculosis. Here, we used the murine model to study the virulence and lethality of the genetically and epidemiologically distinct LAM strains. The pathobiological characteristics of the four LAM strains (three drug resistant and one drug susceptible) and the susceptible reference strain H37Rv were studied in the C57BL/6 mouse model. The whole-genome sequencing was performed using the HiSeq Illumina platform, followed by bioinformatics and phylogenetic analysis. The susceptible strain H37Rv showed the highest virulence. Drug-susceptible LAM strain (spoligotype SIT264) was more virulent than three multidrug-resistant (MDR) strains (SIT252, SIT254, and SIT266). All three MDR isolates were low lethal, while the susceptible isolate and H37Rv were moderately/highly lethal. Putting the genomic, phenotypic, and virulence features of the LAM strains/spoligotypes in the context of their dynamic phylogeography over 20 years reveals three types of relationships between virulence, resistance, and transmission. First, the most virulent and more lethal drug-susceptible SIT264 increased its circulation in parts of Russia. Second, moderately virulent and pre-XDR SIT266 was prevalent in Belarus and continues to be visible in North-West Russia. Third, the low virulent and MDR strain SIT252 previously considered as emerging has disappeared from the population. These findings suggest that strain virulence impacts the transmission, irrespective of drug resistance properties. The increasing circulation of susceptible but more virulent and lethal strains implies that personalized TB treatment should consider not only resistance but also the virulence of the infecting M. tuberculosis strains. IMPORTANCE The study is multidisciplinary and investigates the epidemically/clinically important and global lineage of Mycobacterium tuberculosis, named Latin-American-Mediterranean (LAM), yet insufficiently studied with regard to its pathobiology. We studied different LAM strains (epidemic vs endemic and resistant vs susceptible) in the murine model and using whole-genome analysis. We also collected long-term, 20-year data on their prevalence in Eurasia. The findings are both expected and unexpected. (i) We observe that a drug-susceptible but highly virulent strain increased its prevalence. (ii) By contrast, the multidrug-resistant (MDR) but low-virulent, low-lethal strain (that we considered as emerging 15 years ago) has almost disappeared. (iii) Finally, an intermediate case is the MDR strain with moderate virulence that continues to circulate. We conclude that (i) the former and latter strains are the most hazardous and require close epidemiological monitoring, and (ii) personalized TB treatment should consider not only drug resistance but also the virulence of the infecting strains and development of anti-virulence drugs is warranted.

The Use of Mesenchymal Stem Cells in the Complex Treatment of Kidney Tuberculosis (Experimental Study).

In recent years, the application of mesenchymal stem cells (MSCs) has been recognized as a promising method for treatment of different diseases associated with inflammation and sclerosis, which include nephrotuberculosis. The aim of our study is to investigate the effectiveness of MSCs in the complex therapy of experimental rabbit kidney tuberculosis and to evaluate the effect of cell therapy on the reparative processes. Methods: To simulate kidney tuberculosis, a suspension of the standard strain Mycobacterium tuberculosis H37Rv (106 CFU) was used, which was injected into the cortical layer of the lower pole parenchyma of the left kidney under ultrasound control in rabbits. Anti-tuberculosis therapy (aTBT) was started on the 18th day after infection. MSCs (5 × 107 cells) were transplanted intravenously after the start of aTBT. Results: 2.5 months after infection, all animals showed renal failure. Conducted aTBT significantly reduced the level of albumin, ceruloplasmin, elastase and the severity of disorders in the proteinase/inhibitor system and increased the productive nature of inflammation. A month after MSC transplantation, the level of inflammatory reaction activity proteins decreased, the area of specific and destructive inflammation in kidneys decreased and the formation of mature connective tissue was noted, which indicates the reparative reaction activation.

Pulmonary alveolar proteinosis: a long way to correct diagnosis: problems of diagnostics and therapy in routine practice.

Pulmonary Alveolar Proteinosis is a rare disease with unknown etiology that is due to an abnormal surfactant metabolism. Retrospective analysis of a consecutive series of 68 patients with PAP who were seen at Pulmonology Clinic of Pavlov State Medical University from 1977-2013 was performed. The mean time since first abnormalities were found until diagnosis establishment was 34 months. During that time, most patients were treated with antibiotics, tuberculostatics, glucocorticoids, and immunosupressants. The most effective PAP treatment method was found to be whole lung lavage procedure: 82% of patients showed improvement after its implementation. Delayed diagnosis and incorrect administration of antibiotics, tuberculostatics etc. reduce the probability of a long symptom-free period after lung lavage and spontaneous resolution of the disease.

Certain surfactants significantly enhance the activity of antibiotics in the mouse model of MTB and drug resistant MTB infection and effectively remove the bacteria from a pulmonary cavity in human ex-vivo study.

Surfactants have the potential to overcome natural resistance of MTB to antibiotics which is mediated by barriers that impede the penetration of drugs to their targets. A major component of this barrier is trehalose dimycolate (TDM) which surrounds the bacteria with a thick lipid shield. In this study dodecyl maltoside (DDM) was evaluated for this purpose. This surfactant is an excellent cellular permeabilizing agent with associated low toxicity. The administration of the surfactant as an aerosol into the lungs of the infected mice achieved a 5-10 times enhancement of the isoniazid (INH) treatment gauged by the reduction of the colony forming units. This study also established proof of principle that surfactants alone applied as an aerosol can reduce the bacteria count in lungs infected with MTB. The potential of the surfactant in the therapy of human cavitary TB was also investigated using a surgically removed lung from a patient with extreme drug resistant MTB (XDR-TB). A cavity in this lung was flushed with DDM solution ex-vivo. The procedure readily removed the bacteria, excessive amounts of TDM and necrotic tissue from the cavity. These studies demonstrate that DDM can disrupt the layers of TDM and free embedded MTB and, consequently, surfactants have promise as a proficient modality for the treatment of pulmonary MTB.

Morphological manifestations of the atypical mycobacteriosis caused by nontuberculous mycobacteria in the HIV infected patients.

Infection with atypical mycobacteria (MAC) is a well-known complication of AIDS that typically occurs only in people with advanced immunodeficiency. We studied tissues from 13 patients with HIV and atypical mycobacterial infection who died in St Petersburg Russia from 2009-2012. Three patterns of disease were identified that suggest effects of host resistance. The first pattern was in people paucibacillary disease. They had positive blood cultures and histologic changes consistent with mycobacterial infection, but no stainable acid fast bacilli (AFB). The second group had disseminated infection in many organs including the lungs with extensive necrosis with many AFB. Finally, the third group had massive infection of many organs, but not the lungs, and only minimal necrosis. These observations suggest significant heterogeneity in atypical mycobacterial infections.