Ultrahigh-throughput screening (uHTS) techniques can identify unique functionality from millions of variants. To mimic the natural selection mechanisms that occur by compartmentalization in vivo, we ...developed a technique based on single-cell encapsulation in droplets of a monodisperse microfluidic double water-in-oil-in-water emulsion (MDE). Biocompatible MDE enables in-droplet cultivation of different living species. The combination of droplet-generating machinery with FACS followed by next-generation sequencing and liquid chromatography-mass spectrometry analysis of the secretomes of encapsulated organisms yielded detailed genotype/phenotype descriptions. This platform was probed with uHTS for biocatalysts anchored to yeast with enrichment close to the theoretically calculated limit and cell-to-cell interactions. MDE–FACS allowed the identification of human butyrylcholinesterase mutants that undergo self-reactivation after inhibition by the organophosphorus agent paraoxon. The versatility of the platform allowed the identification of bacteria, including slow-growing oral microbiota species that suppress the growth of a common pathogen, Staphylococcus aureus, and predicted which genera were associated with inhibitory activity.
Adoptive cell transfer (ACT) is a promising approach to cancer immunotherapy, but its efficiency fundamentally depends on the extent of tumor-specific T cell enrichment within the graft. This can be ...estimated via activation with identifiable neoantigens, tumor-associated antigens (TAAs), or living or lysed tumor cells, but these approaches remain laborious, time-consuming, and functionally limited, hampering clinical development of ACT. Here, we demonstrate that homology cluster analysis of T cell receptor (TCR) repertoires efficiently identifies tumor-reactive TCRs allowing to: (1) detect their presence within the pool of tumor-infiltrating lymphocytes (TILs); (2) optimize TIL culturing conditions, with IL-2
/IL-21/anti-PD-1 combination showing increased efficiency; (3) investigate surface marker-based enrichment for tumor-targeting T cells in freshly isolated TILs (enrichment confirmed for CD4
and CD8
PD-1
/CD39
subsets), or re-stimulated TILs (informs on enrichment in 4-1BB-sorted cells). We believe that this approach to the rapid assessment of tumor-specific TCR enrichment should accelerate T cell therapy development.
Since periodontitis and type 2 diabetes mellitus are complex diseases, a thorough understanding of their pathogenesis requires knowing the relationship of these pathologies with other disorders and ...environmental factors. In this study, the representability of the subgingival periodontal microbiome of 46 subjects was studied by 16S rRNA gene sequencing and shotgun sequencing of pooled samples. We examined 15 patients with chronic periodontitis (CP), 15 patients with chronic periodontitis associated with type 2 diabetes mellitus (CPT2DM), and 16 healthy subjects (Control). The severity of generalized chronic periodontitis in both periodontitis groups of patients (CP and CPT2DM) was moderate (stage II). The male to female ratios were approximately equal in each group (22 males and 24 females); the average age of the subjects was 53.9 ± 7.3 and 54.3 ± 7.2 years, respectively. The presence of overweight patients (Body Mass Index (BMI) 30-34.9 kg/m
) and patients with class 1-2 obesity (BMI 35-45.9 kg/m
) was significantly higher in the CPT2DM group than in patients having only chronic periodontitis or in the Control group. However, there was no statistically significant difference in all clinical indices between the CP and CPT2DM groups. An analysis of the metagenomic data revealed that the alpha diversity in the CPT2DM group was increased compared to that in the CP and Control groups. The microbiome biomarkers associated with experimental groups were evaluated. In both groups of patients with periodontitis, the relative abundance of
was increased compared to that in the Control group. The CPT2DM group was characterized by a lower relative abundance of
/
and a higher abundance of
compared to those in the CP and Control groups. Furthermore, the CP and CPT2DM groups differed in terms of the relative abundance of
(which was decreased in the CPT2DM group compared to CP) and
(which was increased in the CPT2DM group compared to CP). In addition, differences in bacterial content were identified by a combination of shotgun sequencing of pooled samples and genome-resolved metagenomics. The results indicate that there are subgingival microbiome-specific features in patients with chronic periodontitis associated with type 2 diabetes mellitus.
Here we present the complete genome sequence of Bacteroides fragilis isolate BOB25. It is an enterotoxigenic isolate that was obtained from a stool sample of a patient with dysbiosis.
Viliuisk encephalomyelitis (VE) is a rare endemic neurodegenerative disease occurring in the Yakut population of Northeastern Siberia. The main clinical features of VE are spasticity, dysarthria, ...dementia, central paresis and paralysis, and cortical atrophy observed via MRI. Many hypotheses have been proposed regarding its etiology, including infectious agents, genetics, environmental factors, and immunopathology. Each of these hypotheses has been supported to some extent by epidemiological and experimental data. Nevertheless, none of them has been decisively proven. Gut microbiome is one of the factors that might be involved in VE pathogenesis. Here we performed a pilot survey of the stool microbiomes of Yakut subjects with VE (n = 6) and without VE (n = 11). 16S rRNA sequencing showed that in comparison with the control group, the Yakuts with VE had increased proportions of Methanobrevibacter and Christensenella, which are reported to be linked to body mass index, metabolism, dietary habits and potentially to neurodegenerative disorders. The identified associations suggest that the microbiome may be involved in VE. Overall, the Yakut microbiome was quite specific in comparison with other populations, such as metropolitan Russians and native inhabitants of the Canadian Arctic. Describing the gut microbiome of indigenous human populations will help to elucidate the impact of dietary and environmental factors on microbial community structure and identify risks linked to the lifestyles of such groups as well as endemic diseases.
Type 2 diabetes (T2D) is a serious disease. The gut microbiota (GM) has recently been identified as a new potential risk factor in addition to well-known diabetes risk factors. To investigate the GM ...composition in association with the dietary patterns in patients with different glucose tolerance, we analyzed 92 patients: with normal glucose tolerance (n=48), prediabetes (preD, n=24), and T2D (n=20). Metagenomic analysis was performed using 16S rRNA sequencing. The diet has been studied by a frequency method with a quantitative evaluation of food intake using a computer program. Microbiota in the samples was predominantly represented by Firmicutes, in a less degree by Bacteroidetes. Blautia was a dominant genus in all samples. The representation of Blautia, Serratia was lower in preD than in T2D patients, and even lower in those with normal glucose tolerance. After the clustering of the samples into groups according to the percentage of protein, fat, carbohydrates in the diet, the representation of the Bacteroides turned to be lower and Prevotella abundance turned to be higher in carbohydrate cluster. There were more patients with insulin resistance, T2D in the fat–protein cluster. Using the Calinski–Harabasz index identified the samples with more similar diets. It was discovered that half of the patients with a high-fat diet had normal tolerance, the others had T2D. The regression analysis showed that these T2D patients also had a higher representation of Blautia. Our study provides the further evidence concerning the structural modulation of the GM in the T2DM pathogenesis depending on the dietary patterns.
What strategies do bacteria employ for adaptation to their hosts and are these strategies different for varied hosts? To date, many studies on the interaction of the bacterium and its host have been ...published. However, global changes in the bacterial cell in the process of invasion and persistence, remain poorly understood. In this study, we demonstrated phase transition of the avian pathogen Mycoplasma gallisepticum upon invasion of the various types of eukaryotic cells (human, chicken, and mouse) which was stable during several passages after isolation of intracellular clones and recultivation in a culture medium. It was shown that this phase transition is manifested in changes at the proteomic, genomic and metabolomic levels. Eukaryotic cells induced similar proteome reorganization of M. gallisepticum during infection, despite different origins of the host cell lines. Proteomic changes affected a broad range of processes including metabolism, translation and oxidative stress response. We determined that the activation of glycerol utilization, overproduction of hydrogen peroxide and the upregulation of the SpxA regulatory protein occurred during intracellular infection. We propose SpxA as an important regulator for the adaptation of M. gallisepticum to an intracellular environment.
Introduction.
The presence of a germinal BRCA mutation occurs in 3–4% of all breast cancer (BC) patients with various biological subtypes, but significantly with a high frequency in patients with a ...triple negative biological subtype (in 10–20% of cases). For the treatment of patients with HER2-negative metastatic breast cancer associated with gBRCA mutation, the effectiveness of biologically targeted drugs from the group of PARP inhibitors (olaparib and talazoparib) has been proven.
P
urpose.
Comparison of the results of our experience with the use of talazoparib in patients with HER2-gBRCA+ + mBC with the data of the EMBRACA registration study.
Mat
erials and
methods.
As part of the multicenter compassionate use program (CUP) with the support of Pfizer, 24 patients with HER2-negative metastatic gBRCA-associated mutation metastatic breast cancer (HER2-gBRCA+ breast cancer) received biologically targeted therapy with the PARP inhibitor talazoparib at a standard oral dose of 1 mg per day for vital indications . The average age of patients with HER2-gBRCAm+ breast cancer was 50 years (29–90 years).
Results.
Objective response (OR) was registered in 29% of cases, disease control (OR+stabilization) – in 71% of cases. The median progression-free survival (PFS) was 6.5 months (95% CI 3–10). Objective response, disease control, and median PFS were evaluated depending on the biological subtype, the number of lines of previous therapy, and the presence of platinum-containing agents in the anamnesis.
Objective response and disease control were evaluated depending on the biological subtype: in patients with ER+HER2-mBC versus patients with triple negative subtype, OR was 33% vs 22%, and disease control was 83% vs 61%, respectively. In the presence of < 3 vs ≥ 3 lines of therapy for metastatic disease in the anamnesis, OR was 31% vs 12.5%, disease control – 75% vs 50% of cases, respectively. In the presence or absence of platinum-containing agents in the anamnesis, OR was observed in 22% vs 33% of cases, and disease control – 67% vs 67%, respectively.
In patients with the luminal subtype versus patients with the triple negative subtype, the PFS was 9 months vs 5 months, respectively (HR = 0.705; 95% CI 0.231–2.147; p = 0.5208). Median PFS in the presence of <3 vs ≥3 lines of therapy for metastatic disease in the anamnesis was 9 months vs 4 months, respectively (HR = 4,216; 95% CI 1,334–13,327; p = 0.0056). In the presence or absence of platinum-containing agents in previous lines of therapy 5 months vs 9.5 months, respectively (HR =1.484; 95% CI 0.48–4.582; p = 0.4750).
During the treatment with talazoparib adverse events of the 3rd-4th grades were observed in 5 patients (20,8%). These include moderate and severe anemia in 3 patients (12.5%), thrombocytopenia in 1 patient (4%), and neutropenia in 1 patient (4%). The majority of patients (79,5%), which received talazoparib, did not require dose adjustment. The need to reduce the dose to 0.75 mg was noted in 3 patients (12.5%), to 0.5 mg – in 2 patients (8%). Hemotransfusion was performed in 3 patients. For effective therapy safety management regular monitoring of blood parameters is necessary.
Conclusion.
Thus, targeted therapy with talazoparib is an effective treatment option for HER2-gBRCA+ mBC.
The genus
relates to Gram-positive bacteria that lack a cell wall and are capable to cause chronic disease in humans and animals. Among the agents of infection and disease in domestic poultry and ...wild birds,
is the most important mycoplasma species, causing considerable losses in the poultry industry. In the present paper, we provide data on adaptation of
to the eukaryotic host cells on the genomic level. The major changes were predominantly localized in the VlhA-hemagglutinin genes which are important components of pathogenesis. The ability of mycoplasmas to change dramatically the repertoire of surface antigens and to vary the immunogenicity of these components allows them to remain undetected by the immune system of the host. The data presented in this article are related to the article entitled "Phase Transition of the Bacterium upon Invasion of a Host Cell as a Mechanism of Adaptation: a
Model." (Matyushkina et al., 2016) 1. Data posted in repository https://www.ncbi.nlm.nih.gov/bioproject/315515. Bioproject ID: PRJNA315515.
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