Genetic Platforms Research Articles

The expression of integron arrays is shaped by the translation rate of cassettes

Integrons are key elements in the rise and spread of multidrug resistance in Gram-negative bacteria. These genetic platforms capture cassettes containing promoterless genes and stockpile them in arrays of variable length. In the current integron model, expression of cassettes is granted by the Pc promoter in the platform and is assumed to decrease as a function of its distance. Here we explored this model using a large collection of 136 antibiotic resistance cassettes and show the effect of distance is in fact negligible. Instead, cassettes have a strong impact in the expression of downstream genes because their translation rate affects the stability of the whole polycistronic mRNA molecule. Hence, cassettes with reduced translation rates decrease the expression and resistance phenotype of cassettes downstream. Our data puts forward an integron model in which expression is contingent on the translation of cassettes upstream, rather than on the distance to the Pc.

Bacteriocin distribution patterns in Enterococcus faecium and Enterococcus lactis: bioinformatic analysis using a tailored genomics framework.

This work significantly expands the knowledge on the understudied bacteriocin diversity in opportunistic enterococci, revealing their contribution in the adaptation to different environments. It underscores the importance of placing increased emphasis on genetic platforms carrying bacteriocins as well as on cryptic plasmids that often exclusively harbor bacteriocins since bacteriocin production can significantly contribute to plasmid maintenance, potentially facilitating their stable transmission across generations. Further characterization of strain-level bacteriocin landscapes could inform strategies to combat high-risk clones. Overall, these insights provide a framework for unraveling the therapeutic and biotechnological potential of bacteriocins.

Biomarkers in breast cancer 2024: an updated consensus statement by the Spanish Society of Medical Oncology and the Spanish Society of Pathology.

This revised consensus statement of the Spanish Society of Medical Oncology (SEOM) and the Spanish Society of Pathological Anatomy (SEAP) updates the recommendations for biomarkers use in the diagnosis and treatment of breast cancer that we first published in 2018. The expert group recommends determining in early breast cancer the estrogen receptor (ER), progesterone receptor (PR), Ki-67, and Human Epidermal growth factor Receptor 2 (HER2), as well as BReast CAncer (BRCA) genes in high-risk HER2-negative breast cancer, to assist prognosis and help in indicating the therapeutic options, including hormone therapy, chemotherapy, anti-HER2 therapy, and other targeted therapies. One of the four available genetic prognostic platforms (Oncotype DX®, MammaPrint®, Prosigna®, or EndoPredict®) may be used in ER-positive patients with early breast cancer to establish a prognostic category and help decide with the patient whether adjuvant treatment may be limited to hormonal therapy. In second-line advanced breast cancer, in addition, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) and estrogen receptor 1 (ESR1) should be tested in hormone-sensitive cases, BRCA gene mutations in HER2-negative cancers, and in triple-negative breast cancer (TNBC), programmed cell death-1 ligand (PD-L1). Newer biomarkers and technologies, including tumor-infiltrating lymphocytes (TILs), homologous recombination deficiency (HRD) testing, serine/threonine kinase (AKT) pathway activation, and next-generation sequencing (NGS), are at this point investigational.

Tissue culture and Agrobacterium-mediated genetic transformation of the oil crop sunflower

Sunflower is one of the four major oil crops in the world. ‘Zaoaidatou’ (ZADT), the main variety of oil sunflower in the northwest of China, has a short growth cycle, high yield, and high resistance to abiotic stress. However, the ability to tolerate adervesity is limited. Therefore, in this study, we used the retention line of backbone parent ZADT as material to establish its tissue culture and genetic transformation system for new variety cultivating to enhance resistance and yields by molecular breeding. The combination of 0.05 mg/L IAA and 2 mg/L KT in MS was more suitable for direct induction of adventitious buds with cotyledon nodes and the addition of 0.9 mg/L IBA to MS was for adventitious rooting. On this basis, an efficient Agrobacterium tumefaciens-mediated genetic transformation system for ZADT was developed by the screening of kanamycin and optimization of transformation conditions. The rate of positive seedlings reached 8.0%, as determined by polymerase chain reaction (PCR), under the condition of 45 mg/L kanamycin, bacterial density of OD600 0.8, infection time of 30 min, and co-cultivation of three days. These efficient regeneration and genetic transformation platforms are very useful for accelerating the molecular breeding process on sunflower.

Microbial biofilms on macroalgae harbour diverse integron gene cassettes.

Integrons are genetic platforms that capture, rearrange and express mobile modules called gene cassettes. The best characterized gene cassettes encode antibiotic resistance, but the function of most integron gene cassettes remains unknown. Functional predictions suggest that many gene cassettes could encode proteins that facilitate interactions with other cells and with the extracellular environment. Because cell interactions are essential for biofilm stability, we sequenced gene cassettes from biofilms growing on the surface of the marine macroalgae Ulva australis and Sargassum linearifolium. Algal samples were obtained from coastal rock platforms around Sydney, Australia, using seawater as a control. We demonstrated that integrons in microbial biofilms did not sample genes randomly from the surrounding seawater, but harboured specific functions that potentially provided an adaptive advantage to both the bacterial cells in biofilm communities and their macroalgal host. Further, integron gene cassettes had a well-defined spatial distribution, suggesting that each bacterial biofilm acquired these genetic elements via sampling from a large but localized pool of gene cassettes. These findings suggest two forms of filtering: a selective acquisition of different integron-containing bacterial species into the distinct biofilms on Ulva and Sargassum surfaces, and a selective retention of unique populations of gene cassettes at each sampling location.

VIM-type metallo-β-lactamase (MBL)-encoding genomic islands in Pseudomonas spp. in Poland: predominance of clc-like integrative and conjugative elements (ICEs).

To characterize VIM-type metallo-β-lactamase (MBL)-encoding genomic islands (GIs) in Pseudomonas aeruginosa and P. putida group isolates from Polish hospitals from 2001-2015/16. Twelve P. aeruginosa and 20 P. putida group isolates producing VIM-like MBLs were selected from a large collection of these based on epidemiological and typing data. The organisms represented all major epidemic genotypes of these species spread in Poland with chromosomally located blaVIM gene-carrying integrons. The previously determined short-read sequences were complemented by long-read sequencing in this study. The comparative structural analysis of the GIs used a variety of bioinformatic tools. Thirty different GIs with blaVIM integrons were identified in the 32 isolates, of which 24 GIs from 26 isolates were integrative and conjugative elements (ICEs) of the clc family. These in turn were dominated by 21 variants of the GI2/ICE6441 subfamily with a total of 19 VIM integrons, each inserted in the same position within the ICE's Tn21-like transposon Tn4380. The three other ICEs formed a novel ICE6705 subfamily, lacking Tn4380 and having different VIM integrons located in another site of the elements. The remaining six non-ICE GIs represented miscellaneous structures. The presence of various integrons in the same ICE sublineage, and of the same integron in different GIs, indicated circulation and recombination of the integron-carrying genetic platforms across Pseudomonas species/genotypes. Despite the general diversity of the blaVIM-carrying GIs in Pseudomonas spp. in Poland, a clear predominance of broadly spread and rapidly evolving clc-type ICEs was documented, confirming their significant role in antimicrobial resistance epidemiology.

Identification of promoter activity in gene-less cassettes fromVibrionaceae superintegrons.

Integrons are genetic platforms that acquire new genes encoded in integron cassettes (ICs), building arrays of adaptive functions. ICs generally encode promoterless genes, whose expression relies on the platform-associated Pc promoter, with the cassette array functioning as an operon-like structure regulated by the distance to the Pc. This is relevant in large sedentary chromosomal integrons (SCIs) carrying hundreds of ICs, like those in Vibrio species. We selected 29 gene-less cassettes in four Vibrio SCIs, and explored whether their function could be related to the transcription regulation of adjacent ICs. We show that most gene-less cassettes have promoter activity on the sense strand, enhancing the expression of downstream cassettes. Additionally, we identified the transcription start sites of gene-less ICs through 5'-RACE. Accordingly, we found that most of the superintegron in Vibrio cholerae is not silent. These promoter cassettes can trigger the expression of a silent dfrB9 cassette downstream, increasing trimethoprim resistance>512-fold in V. cholerae and Escherichia coli. Furthermore, one cassette with an antisense promoter can reduce trimethoprim resistance when cloned downstream. Our findings highlight the regulatory role of gene-less cassettes in the expression of adjacent cassettes, emphasizing their significance in SCIs and their clinical importance if captured by mobile integrons.

A medium-density genotyping platform for cultivated strawberry using DArTag technology.

Genomic prediction in breeding populations containing hundreds to thousands of parents and seedlings is prohibitively expensive with current high-density genetic marker platforms designed for strawberry. We developed mid-density panels of molecular inversion probes (MIPs) to be deployed with the "DArTag" marker platform to provide a low-cost, high-throughput genotyping solution for strawberry genomic prediction. In total, 7742 target single nucleotide polymorphism (SNP) regions were used to generate MIP assays that were tested with a screening panel of 376 octoploid Fragaria accessions. We evaluated the performance of DArTag assays based on genotype segregation, amplicon coverage, and their ability to produce subgenome-specific amplicon alignments to the FaRR1 assembly and subsequent alignment-based variant calls with strong concordance to DArT's alignment-free, count-based genotype reports. We used a combination of marker performance metrics and physical distribution in the FaRR1 assembly to select 3K and 5K production panels for genotyping of large strawberry populations. We show that the 3K and 5K DArTag panels are able to target and amplify homologous alleles within subgenomic sequences with low-amplification bias between reference and alternate alleles, supporting accurate genotype calling while producing marker genotypes that can be treated as functionally diploid for quantitative genetic analysis. The 3K and 5K target SNPs show high levels of polymorphism in diverse F. × ananassa germplasm and UC Davis cultivars, with mean pairwise diversity (π) estimates of 0.40 and 0.32 and mean heterozygous genotype frequencies of 0.35 and 0.33,respectively.

Efficient and stable CRISPR/Cas9-mediated genome-editing of human type 2 innate lymphoid cells.

Innate lymphoid cells (ILCs) are a family of innate lymphocytes with important roles in immune response coordination and maintenance of tissue homeostasis. The ILC family includes group 1 (ILC1s), group 2 (ILC2s) and group 3 (ILC3s) 'helper' ILCs, as well as cytotoxic Natural Killer (NK) cells. Study of helper ILCs in humans presents several challenges, including their low proportions in peripheral blood or needing access to rare samples to study tissue resident ILC populations. In addition, the lack of established protocols harnessing genetic manipulation platforms has limited the ability to explore molecular mechanism regulating human helper ILC biology. CRISPR/Cas9 is an efficient genome editing tool that enables the knockout of genes of interest, and is commonly used to study molecular regulation of many immune cell types. Here, we developed methods to efficiently knockout genes of interest in human ILC2s. We discuss challenges and lessons learned from our CRISPR/Cas9 gene editing optimizations using a nucleofection transfection approach and test a range of conditions and nucleofection settings to obtain a protocol that achieves effective and stable gene knockout while maintaining optimal cell viability. Using IL-4 as a representative target, we compare different ribonucleoprotein configurations, as well as assess effects of length of time in culture and other parameters that impact CRISPR/Cas9 transfection efficiency. Collectively, we detail a CRISPR/Cas9 protocol for efficient genetic knockout to aid in studying molecular mechanism regulating human ILC2s.

Transfection Models to Investigate Plasmodium vivax-Type Dormant Liver Stage Parasites.

Plasmodium vivax causes the second highest number of malaria morbidity and mortality cases in humans. Several biological traits of this parasite species, including the formation of dormant stages (hypnozoites) that persist inside the liver for prolonged periods of time, present an obstacle for intervention measures and create a barrier for the elimination of malaria. Research into the biology of hypnozoites requires efficient systems for parasite transmission, liver stage cultivation and genetic modification. However, P. vivax research is hampered by the lack of an in vitro blood stage culture system, rendering it reliant on in vivo-derived, mainly patient, material for transmission and liver stage culture. This has also resulted in limited capability for genetic modification, creating a bottleneck in investigations into the mechanisms underlying the persistence of the parasite inside the liver. This bottleneck can be overcome through optimal use of the closely related and experimentally more amenable nonhuman primate (NHP) parasite, Plasmodium cynomolgi, as a model system. In this review, we discuss the genetic modification tools and liver stage cultivation platforms available for studying P. vivax persistent stages and highlight how their combined use may advance our understanding of hypnozoite biology.

Blinded rebiopsy and analysis of noneuploid embryos with 2 distinct preimplantation genetic testing platforms for aneuploidy

To determine how often a noneuploid result from a single trophectoderm (TE) biopsy tested with the next-generation sequencing (NGS)-based preimplantation genetic testing for aneuploidy (PGT-A) is concordant with rebiopsies tested with a single-nucleotide polymorphism (SNP) array-based PGT-A platform. Blinded prospective cohort study. University-affiliated fertility center. One hundred blastocysts were chosen from donated samples; on TE biopsy with NGS-based PGT-A, 40 had at least one whole chromosome full copy number aneuploidy alone, 20 had a single whole chromosome intermediate copy number ("whole chromosome mosaic"), 20 had a single full segmental aneuploidy (segA), and 20 had a single segmental intermediate copy number ("segmental mosaic"). Four rebiopsies were collected from each embryo: 3 TE biopsies and the remaining embryo. Each rebiopsy was randomized, blinded, and assessed with an SNP array-based PGT-A platform that combines copy number and allele ratio analyses, without mosaicism reporting. Concordance between the NGS result and rebiopsy results and within each embryo's blinded rebiopsy results. Next-generation sequencing-diagnosed whole chromosome aneuploidy (WCA) was reconfirmed in 95% (95% confidence interval [CI], 83%-99%) of embryos; 2 embryos with NGS-diagnosed WCA were called euploid on all conclusive rebiopsies. Among embryos with NGS-diagnosed whole chromosome mosaicism, 35% (95% CI, 15%-59%) were called euploid and 15% (95% CI, 3%-38%) were called whole chromosome aneuploid on all conclusive rebiopsies. A total of 30% (95% CI, 12%-54%) of embryos with NGS-diagnosed segA and 65% (95% CI, 41%-85%) of embryos with NGS-diagnosed segmental mosaicism were called euploid on all conclusive rebiopsies. In total, 13% (95% CI, 6%-25%) of embryos with NGS-diagnosed full copy number aneuploidy and 50% (95% CI, 34%-66%) of embryos with NGS-diagnosed mosaicism had uniformly euploid SNP results. Conversely, all embryos with at least one noneuploid SNP result (n = 72) either had SNP-diagnosed aneuploidy on another rebiopsy from the same embryo or NGS-diagnosed aneuploidy/mosaicism involving the same chromosome. Next-generation sequencing-diagnosed WCA is highly concordant with rebiopsies tested with an SNP array-based PGT-A; however, whole chromosome mosaicism, segA, and segmental mosaicism are less concordant, reinforcing that embryos with these results may have reproductive potential and be suitable for transfer.

Detection and genetic characterization of blaESBL-carrying plasmids of cloacal Escherichia coli isolates from white stork nestlings (Ciconia ciconia) in Spain

This study aimed to characterize Escherichia coli isolates from cloacal samples of white stork nestlings, with a special focus on extended-spectrum β-lactamases (ESBLs)-producing E. coli isolates and their plasmid content. Cloacal samples of 88 animals were seeded on MacConkey-agar and chromogenic-ESBL plates to recover E. coli and ESBL-producing E. coli. Antimicrobial susceptibility was screened using the disc diffusion method, and the genotypic characterization was performed by polymerase chain reaction (PCR) and subsequent sequencing. S1 nuclease Pulsed-Field-Gel-Electrophoresis (PFGE), Southern blotting, and conjugation essays were performed on ESBL-producing E. coli, as well as whole-genome sequencing by short- and long-reads. The four blaESBL-carrying plasmids were completely sequenced. A total of 113 non-ESBL-producing E. coli isolates were collected on antibiotic-free MacConkey-agar, of which 27 (23.9%) showed a multidrug-resistance (MDR) phenotype, mainly associated with β-lactam-phenicol-sulfonamide resistance (blaTEM/cmlA/floR/sul1/sul2/sul3). Moreover, four white stork nestlings carried ESBL-producing E. coli (4.5%) with the following characteristics: blaSHV-12/ST38-D, blaSHV-12/ST58-B1, blaCTX-M-1/ST162-B1, and blaCTX-M-32/ST155-B1. Whole-genome sequencing followed by Southern blot hybridizations on S1-PFGE gels in ESBL-positive isolates proved that the blaCTX-M-1 gene and one of the blaSHV-12 genes were carried by IncI1/pST3 plasmids, while the second blaSHV-12 gene and the blaCTX-M-32 gene were located on IncF plasmids. The two blaSHV-12 genes and the two blaCTX-M genes had similar but non-identical close genetic environments, as all four genes were flanked by a variety of insertion sequences. The role played by several genetic platforms in the mobility of ESBL genes allows for interchangeability on a remarkably small scale (gene-plasmid-clones), which may support the spread of ESBL genes.

Emergence of NDM-1 and KPC-3 carbapenemases in Kluyvera cryocrescens: Investigating genetic heterogeneity and acquisition routes of blaNDM-1 in Enterobacterales species in Portugal

A higher diversity of species, clones and genes have been increasingly implicated in carbapenemases spread, though the mobile genetic elements responsible for their acquisition and dispersion at local and global levels are less explored, particularly in species other than K. pneumoniae or E. coli. We aim to explain the emergence of NDM-1 and KPC-3 carbapenemases in a Kluyvera cryocrescens isolate, and to shed light on the heterogeneity of genetic platforms and acquisition routes of blaNDM-1 in diverse Enterobacterales species in Portugal. A KPC-3 and NDM-1-producing K. cryocrescens colonizing a hospitalized patient in 2019 was characterized by whole-genome sequencing and antibiotic resistance profiling following standard methods. Conjugative transfer of carbapenemases genes was assessed by filter mating. Plasmids were reconstructed with in silico and in vitro approaches. blaNDM-1 genetic context was compared with that of diverse NDM-1-producing Enterobacterales species, sporadically described in Portugal. K. cryocrescens K629 showed a multidrug resistance profile. Resistance gene blaKPC-3 was harboured by a Tn4401d transposon within a worldwide-spread IncN-ST15 plasmid (pKLU-KPC3), whereas blaNDM-1 was located in a Tn3000 within a non-typeable mosaic plasmid (pKLU-NDM1). The heterogeneous blaNDM-1 genetic platforms and variable plasmid backbones identified in various Enterobacterales species suggested multiple introductions of blaNDM-1, mediated by different insertion sequences. We report the convergence of KPC-3 and NDM-1 in K. cryocrescens and the variable dissemination modes of these carbapenemases in different Enterobacterales species, underlining the need to track down genetic platforms responsible for carbapenemases diffusion.

Rodent models to study type 1 and type 2 diabetes induced human diabetic nephropathy.

Diabetic nephropathy (DN), an outcome of prolonged diabetes, has affected millions of people worldwide and every year the incidence and prevalence increase substantially. The symptoms may start with mild manifestations of the disease such as increased albuminuria, serum creatinine levels, thickening of glomerular basement membrane, expansion of mesangial matrix to severe pathological symptoms such as glomerular lesions and tubulointerstitial fibrosis which may further proceed to cardiovascular dysfunction or end-stage renal disease. Numerous therapeutic interventions are being explored for the management of DN, however, these interventions do not completely halt the progression of this disease and hence animal models are being explored to identify critical genetic and molecular parameters which could help in tackling the disease. Rodent models which mostly include mice and rats are commonly used experimental animals which provide a wide range of advantages in understanding the onset and progression of disease in humans and also their response to a wide range of interventions helps in the development of effective therapeutics. Rodent models of type 1 and type 2 diabetes induced DN have been developed utilizing different platforms and interventions during the last few decades some of which mimic various stages of diabetes ranging from early to later stages. However, a rodent model which replicates all the features of human DN is still lacking. This review tries to evaluate the rodent models that are currently available and understand their features and limitations which may help in further development of more robust models of human DN. Using these rodent models can help to understand different aspects of human DN although further research is required to develop more robust models utilizing diverse genetic platforms which may, in turn, assist in developing effective interventions to target the disease at different levels.

Abundant diversity of accessory genetic elements and associated antimicrobial resistance genes in pseudomonas aeruginosa isolates from a single Chinese hospital

ObjectivesPseudomonas aeruginosa has intrinsic antibiotic resistance and the strong ability to acquire additional resistance genes. However, a limited number of investigations provide detailed modular structure dissection and evolutionary analysis of accessory genetic elements (AGEs) and associated resistance genes (ARGs) in P. aeruginosa isolates. The objective of this study is to reveal the prevalence and transmission characteristics of ARGs by epidemiological investigation and bioinformatics analysis of AGEs of P. aeruginosa isolates taken from a Chinese hospital.MethodsDraft-genome sequencing was conducted for P. aeruginosa clinical isolates (n = 48) collected from a single Chinese hospital between 2019 and 2021. The clones of P. aeruginosa isolates, type 3 secretion system (T3SS)-related virulotypes, and the resistance spectrum were identified using multilocus sequence typing (MLST), polymerase chain reaction (PCR), and antimicrobial susceptibility tests. In addition, 17 of the 48 isolates were fully sequenced. An extensive modular structure dissection and genetic comparison was applied to AGEs of the 17 sequenced P. aeruginosa isolates.ResultsFrom the draft-genome sequencing, 13 STs were identified, showing high genetic diversity. BLAST search and PCR detection of T3SS genes (exoT, exoY, exoS, and exoU) revealed that the exoS+/exoU- virulotype dominated. At least 69 kinds of acquired ARGs, involved in resistance to 10 different categories of antimicrobials, were identified in the 48 P. aeruginosa isolates. Detailed genetic dissection and sequence comparisons were applied to 25 AGEs from the 17 isolates, together with five additional prototype AGEs from GenBank. These 30 AGEs were classified into five groups -- integrative and conjugative elements (ICEs), unit transposons, IncpPBL16 plasmids, Incp60512−IMP plasmids, and IncpPA7790 plasmids.ConclusionThis study provides a broad-scale and deeper genomics understanding of P. aeruginosa isolates taken from a single Chinese hospital. The isolates collected are characterized by high genetic diversity, high virulence, and multiple drug resistance. The AGEs in P. aeruginosa chromosomes and plasmids, as important genetic platforms for the spread of ARGs, contribute to enhancing the adaptability of P. aeruginosa in hospital settings.

Diversity of genetic platforms harboring the blaPER-2 gene in Enterobacterales and insights into the role of ISPa12 in its mobilization and dissemination

The production of PER-like extended-spectrum β-lactamases has recently been associated with reduced susceptibility to the last resort drugs aztreonam/avibactam and cefiderocol. PER-2 has been mainly confined to Argentina and neighboring countries. Until now, only three plasmids harboring blaPER-2 genes have been characterized but very little is known about the involvement of different plasmid groups in its dissemination. The diversity of genetic platforms associated with blaPER-2 genes from a collection of PER-producing Enterobacterales was analysed by describing the close environment and the plasmid backbones. Full sequences of 11 plasmids were obtained by short read (Illumina) and long read (Oxford Nanopore or PacBio) sequencing technologies. De novo assemblies, annotation and sequence analysis were performed by Unicycler, Prokka and BLAST. Plasmid analysis revealed that the blaPER-2 gene is encoded on plasmids of different incompatibility groups (A, C, FIB, HI1B, N2), indicating that this gene may have been disseminated through a variety of plasmids. Comparison with the few publicly available nucleotide sequences describing the blaPER-2 genetic environment, including those from the environmental species Pararheinheimera spp. (considered as the progenitor of blaPER genes), indicates a role of ISPa12 in blaPER-2 gene mobilization from the chromosome of Pararheinheimera spp. Also, the blaPER-2 gene was carried by a novel ISPa12-composite transposon, Tn7390. In addition, its association with ISKox2-like elements in the close genetic environment in all plasmids analysed suggests a role of these insertion sequence elements in further dissemination of blaPER-2 genes.

Advances in point-of-care genetic testing for personalized medicine applications.

Breakthroughs within the fields of genomics and bioinformatics have enabled the identification of numerous genetic biomarkers that reflect an individual's disease susceptibility, disease progression, and therapy responsiveness. The personalized medicine paradigm capitalizes on these breakthroughs by utilizing an individual's genetic profile to guide treatment selection, dosing, and preventative care. However, integration of personalized medicine into routine clinical practice has been limited-in part-by a dearth of widely deployable, timely, and cost-effective genetic analysis tools. Fortunately, the last several decades have been characterized by tremendous progress with respect to the development of molecular point-of-care tests (POCTs). Advances in microfluidic technologies, accompanied by improvements and innovations in amplification methods, have opened new doors to health monitoring at the point-of-care. While many of these technologies were developed with rapid infectious disease diagnostics in mind, they are well-suited for deployment as genetic testing platforms for personalized medicine applications. In the coming years, we expect that these innovations in molecular POCT technology will play a critical role in enabling widespread adoption of personalized medicine methods. In this work, we review the current and emerging generations of point-of-care molecular testing platforms and assess their applicability toward accelerating the personalized medicine paradigm.

Worldwide Dissemination of blaKPC Gene by Novel Mobilization Platforms in Pseudomonas aeruginosa: A Systematic Review.

The dissemination of blaKPC-harboring Pseudomonas aeruginosa (KPC-Pa) is considered a serious public health problem. This study provides an overview of the epidemiology of these isolates to try to elucidate novel mobilization platforms that could contribute to their worldwide spread. A systematic review in PubMed and EMBASE was performed to find articles published up to June 2022. In addition, a search algorithm using NCBI databases was developed to identify sequences that contain possible mobilization platforms. After that, the sequences were filtered and pair-aligned to describe the blaKPC genetic environment. We found 691 KPC-Pa isolates belonging to 41 different sequence types and recovered from 14 countries. Although the blaKPC gene is still mobilized by the transposon Tn4401, the non-Tn4401 elements (NTEKPC) were the most frequent. Our analysis allowed us to identify 25 different NTEKPC, mainly belonging to the NTEKPC-I, and a new type (proposed as IVa) was also observed. This is the first systematic review that consolidates information about the behavior of the blaKPC acquisition in P. aeruginosa and the genetic platforms implied in its successful worldwide spread. Our results show high NTEKPC prevalence in P. aeruginosa and an accelerated dynamic of unrelated clones. All information collected in this review was used to build an interactive online map.

Investigating the significance of segmental aneuploidy findings in preimplantation embryos

Segmental aneuploidies (SAs) are structural imbalances, namely, gains or losses, involving a chromosomal segment. Most preimplantation genetic testing platforms can detect segmental imbalances greater than 5-10 Mb, either full or mosaic; however, questions remain about clinical significance. An in-depth review was performed to determine the accuracy, frequency, and types of SAs detected in preimplantation embryos. A comprehensive search of the literature revealed an incidence of approximately 8.15% in preimplantation embryos, compared with a prevalence of 3.55% in prenatal diagnosis samples. Several studies have used rebiopsy analysis to validate the accuracy and reproducibility of such findings in blastocyst-stage embryos. A comparison of these studies yielded a mean confirmation rate of SAs slightly higher than 30%. This result could be attributed to their mitotic origin as well as to the technical limitations of preimplantation genetic testing. In addition, the few available studies in which embryos with a segmental finding were transferred in utero are analyzed to discuss the reproductive competence of such embryos. Except for 1 study, all outcomes were described for segmental embryos in a mosaic state. As a result, there is still insufficient evidence to provide accurate information about the effect of segmental imbalances on embryonic reproductive competence and to determine gestational and newborn risks.

Pharmacologic Management of Monogenic and Very Early Onset Inflammatory Bowel Diseases.

Inflammatory bowel disease (IBD) is treated with a variety of immunomodulating and immunosuppressive therapies; however, for the majority of cases, these therapies are not targeted for specific disease phenotypes. Monogenic IBD with causative genetic defect is the exception and represents a disease cohort where precision therapeutics can be applied. With the advent of rapid genetic sequencing platforms, these monogenic immunodeficiencies that cause inflammatory bowel disease are increasingly being identified. This subpopulation of IBD called very early onset inflammatory bowel disease (VEO-IBD) is defined by an age of onset of less than six years of age. Twenty percent of VEO-IBDs have an identifiable monogenic defect. The culprit genes are often involved in pro-inflammatory immune pathways, which represent potential avenues for targeted pharmacologic treatments. This review will provide an overview of the current state of disease-specific targeted therapies, as well as empiric treatment for undifferentiated causes of VEO-IBD.