The bla_OXA-1 gene encodes an oxacillin-hydrolyzing beta-lactamase of extended-spectrum beta-lactamase (ESBL)-producing microorganisms. The bla_OXA-1 gene is found in the resistomes of some Enterobacteriaceae, Morganellaceae, Pasteurellaceae, Moraxellaceae, Aeromonadaceae, Pseudomonadaceae, Yersiniaceae, and Vibrionaceae. Most ESBL detection methods, including those to detect OXA-1-producing microorganisms, are time-consuming, and require specialized equipment and qualified personnel. Here, we report a new CRISPR(Clustered Regularly Interspaced Short Palindromic Repeats)/Cas12a-based detection assay coupled with polymerase chain reaction (PCR) to sensitively detect OXA-1-bearing microorganisms. The PCR-coupled CRISPR/Cas12a-based fluorescence assay includes (i) a pre-amplification step and (ii) a nucleic acid detection step. The pre-amplification step is based on a commonly used PCR, and the detection step is based on the CRISPR/Cas12a property to nonspecifically hydrolyze single-stranded DNA fluorescent reporter molecules. The pre-amplification step takes 65 min, and the detection step is shortened and takes only 5 min. The developed assay can easily detect single (1.25) copies of the bla_OXA-1 gene in a reaction and is efficient not only in the detection of a bla_OXA-1 model matrix but also in the detection of bla_OXA-1-positive microorganisms. We hope that our assay has the potential to improve the monitoring of OXA-1-producing microorganisms and therefore contribute to mitigating the deadly global threat of antibiotic-resistant microorganisms.

Escherichia coli is a commensal and opportunistic bacterium widely distributed around the world in different niches including intestinal of humans and animals, and its extraordinary genome plasticity led to the emergence of pathogenic strains causing a wide range of diseases. E. coli is one of the monitored species in maternity hospitals, being the main etiological agent of urogenital infections, endometriosis, puerperal sepsis, and neonatal diseases. This study presents a comprehensive analysis of E. coli isolates obtained from the maternal birth canal of healthy puerperant women 3–4 days after labor. According to whole genome sequencing data, 31 sequence types and six phylogenetic groups characterized the collection containing 53 isolates. The majority of the isolates belonged to the B2 phylogroup. The data also includes phenotypic and genotypic antibiotic resistance profiles, virulence factors, and plasmid replicons. Phenotypic and genotypic antibiotic resistance testing did not demonstrate extensive drug resistance traits except for two multidrug-resistant E. coli isolates. The pathogenic factors revealed in silico were assessed with respect to CRISPR-element patterns. Multiparametric and correlation analyses were conducted to study the interrelation of different pathoadaptability factors, including antimicrobial resistance and virulence genomic determinants carried by the isolates under investigation. The data presented will serve as a valuable addition to further scientific investigations in the field of bacterial pathoadaptability, especially in studying the role of CRISPR/Cas systems in the E. coli genome plasticity and evolution.

The main challenge after engraftment of human tissues to mice is the development of graft-versus-host disease. It often occurs in an acute form, which reduces the time frame for observations. This is especially important to take into account when planning long-term studies of chronic diseases such as HIV infection. In addition, in mice, even with a similar genotype but different origin, the interaction between the graft and the recipient’s organism can manifest itself differently. We engrafted human immune cells in three different concentrations into immunodeficient NOD/Shi-Prkdc^scid Il2rg^em1/Cyagen mice. Then, the initial points of development of a severe graft-versus-host reaction and the maximum possible time window for humane observation were determined. The study included regular complete blood count and the monitoring of the dynamics of the concentration of human cells in the blood of mice. In addition, the effect of grafts on the activation of the recipient’s immune system was assessed. Finally, necropsy and histological and immunohistochemical examinations of the organs were performed to determine the localization of human cells. In this way, critical factors determining the success of human immune system reconstitution in mice were identified.

Graft failure and graft-versus-host disease are the main limiting factors for the engraftment of not only xenogeneic but also allogeneic grafts. Long-term engraftment of human immune cells in mice with severe combined immunodeficiency and IL2rg deletion is key to the success of long-term studies in modeling human immunodeficiency virus (HIV) infection. Human peripheral blood mononuclear cells were injected intravenously into males of one strain and females of two other strains of immunodeficient mice. Before the injection of human cells, mice of each strain were irradiated with three different doses. Interleukin-2 was administered to 50% of mice within 5 days of human cell injection. Its effect was assessed by monitoring graft-versus-host disease, survival, complete blood count (CBC) indices, quantification of human cells in whole blood, pathological changes, and immunohistochemical detection of human cells in tissues. Furthermore, the applicability of correlating immune-inflammation indices with mouse body weight (BW) dynamics, graft-versus-host disease (GVHD) progression, degrees of chimerism and human T-helper cells was determined for the first time. As a result of this work, a panel of biomarkers was developed for predicting the long-term welfare of mice with elements of the human immune system (HIS) in long-term studies, including HIV testing.

The CRISPR Cas9 system has revolutionized biomedical research by enabling precise genetic modifications in laboratory animals.