Low Copy Research Articles

P169 Copy number variation of alpha defensin 1-3 is a predictive marker of therapeutic response in ulcerative colitis

Abstract Background Alpha-defensin 1-3 (DEFA1A3) is an antimicrobial peptide found in phagocytic cells such as neutrophils, functioning as an effector factor in innate immunity. Reports suggest that copy number variation (CNV) of the DEFA1A3 gene is associated with the severity of ulcerative colitis (UC). However, the impact of DEFA1A3 on the pathogenesis of UC, and treatment response has not been fully investigated. Therefore, we aimed to evaluate the relationship between DEFA1A3-CNV and UC treatment response. Methods This was a retrospective and two medical centers study. The study included UC patients with DEFA1A3-CNV measurements, who attended our department and related facilities between 2020 and 2022. Clinical background and white blood cell count and serological marker were collected from the medical records of included patients. DEFA1A3-CNV was measured using droplet digital PCR after DNA extraction from peripheral blood mononuclear cells. Mayo score was used to determine the response to treatment and severity of UC. Results In total, 197 UC patients were included in the study. The median DEFA1A3 copy number was 7.36, and there was a significant correlation between DEFA1A3 copy number and Mayo score. The high DEFA1A3 copy number group had a significantly higher severity of UC during the observation period, compared with the low copy number group. In the analysis of the relationship between prednisolone/immunosuppressive therapy and DEFA1A3-CNV, the DEFA1A3 copy number was significantly higher in the non-remission group than in the remission group following prednisolone treatment. However, there was no correlation between other immunosuppressive therapies and DEFA1A3 copy number. Conclusion DEFA1A3 copy number is a predictive marker of disease severity and prednisolone treatment response.

The use of Anchored Hybrid Enrichment data to resolve higher-level phylogenetic relationships: A proof-of-concept applied to Asterales (Eudicotyledoneae; Angiosperms)

Anchored Hybrid Enrichment (AHE) is a tool for capturing orthologous regions of the nuclear genome shared in low or single copy across lineages. Despite the increasing number of studies using this method, its usefulness to estimate relationships at deeper taxonomic levels in plants has not been fully explored. Here we present a proof of concept about the performance of nuclear loci obtained with AHE to infer phylogenetic relationships and explore the use of gene sampling schemes to estimate divergence times in Asterales. We recovered low-copy nuclear loci using the AHE method from herbarium material and silica-preserved samples. Maximum likelihood, Bayesian inference, and coalescence approaches were used to reconstruct phylogenomic relationships. Dating analyses were conducted under a multispecies coalescent approach by jointly inferring species tree and divergence times with random gene sampling schemes and multiple calibrations. We recovered 403 low-copy nuclear loci for 63 species representing nine out of eleven families of Asterales. Phylogenetic hypotheses were congruent among the applied methods and previously published results. Analyses with concatenated datasets were strongly supported, but coalescence-based analyses showed low support for the phylogenetic position of families Argophyllaceae and Alseuosmiaceae. Estimated family ages were congruent among gene sampling schemes, with the mean age for Asterales around 130 Myr. Our study documents the usefulness of AHE for resolving phylogenetic relationships at deep phylogenetic levels in Asterales. Observed phylogenetic inconsistencies were possibly due to the non-inclusion of families Phellinceae and Pentaphragmataceae. Random gene sampling schemes produced consistent age estimates with coalescence and species tree relaxed clock approaches.

Com probe implemented STexS II greatly enhances specificity in SARS-CoV-2 variant detection

The initial introduction of utilizing double helix structural oligonucleotides known as SNP typing with excellent specificity (STexS) in a standard PCR greatly improved the detection of single nucleotide polymorphisms (SNP) by enhancing amplification rates of primer-matching strands and interrupting mismatched strands by constant instability of kinetics regarding alignment attaching and detaching. The model was beneficial overall in detecting SNP variants consisting of large amounts of wildtype strands such as EGFR mutation genotyping for early detection of non-small cell lung cancer. While the STexS PCR is advantageous in detecting SNPs and biomarkers, limitations were yet observed. Despite the ability to detect variants 10 times more effective than a typical amplification-refractory mutation system PCR, it could only perform optimally in DNA concentrations around 101 ~ 105. To further enhance STexS specificity to perform detecting viral-RNA variants such as the infamous SARS-CoV-2, a novel improvement of the regular TaqMan Probe using Com-probes to inhibit high copy wild targets and amplify low copy mutant targets. By introducing the novel STexS II, omicron variants of SARS-CoV-2 were able to be successfully detected in high concentrations of normal genes.

Correction: Revised molecular phylogeny, global biogeography, and diversification of palms subfamily Coryphoideae (Arecaceae) based on low copy nuclear and plastid regions

Correction: Revised molecular phylogeny, global biogeography, and diversification of palms subfamily Coryphoideae (Arecaceae) based on low copy nuclear and plastid regions

The development of compulsive coping behaviour is associated with a downregulation of Arc in a Locus Coeruleus neuronal ensemble

Some compulsive disorders have been considered to stem from the loss of control over coping strategies, such as displacement. However, the cellular mechanisms involved in the acquisition of coping behaviours and their subsequent compulsive manifestation in vulnerable individuals have not been elucidated. Considering the role of the locus coeruleus (LC) noradrenaline-dependent system in stress and related excessive behaviours, we hypothesised that neuroplastic changes in the LC may be associated with the acquisition of an adjunctive polydipsic water drinking, a prototypical displacement behaviour, and the ensuing development of compulsion in vulnerable individuals. Thus, male Sprague Dawley rats were characterised for their tendency, or not, to develop compulsive polydipsic drinking in a schedule-induced polydipsia (SIP) procedure before their fresh brains were harvested. A new quantification tool for RNAscope assays revealed that the development of compulsive adjunctive behaviour was associated with a low mRNA copy number of the plasticity marker Arc in the LC which appeared to be driven by specific adaptations in an ensemble of tyrosine hydroxylase (TH)+, zif268− neurons. This ensemble was specifically engaged by the expression of compulsive adjunctive behaviour, not by stress, because its functional recruitment was not observed in individuals that no longer had access to the water bottle before sacrifice, while it consistently correlated with the levels of polydipsic water drinking only when it had become compulsive. Together these findings suggest that downregulation of Arc mRNA levels in a population of a TH+/zif268− LC neurons represents a signature of the tendency to develop compulsive coping behaviours.

Low copy numbers for mitochondrial DNA moderates the strength of nuclear-cytoplasmic incompatibility in plants.

Plant cells contain only small amounts of mitochondrial DNA (mtDNA), with the genomic information shared among multiple mitochondria. The biological relevance and molecular mechanism underlying this hallmark of plant cells has been unclear. Here, we report that Arabidopsis thaliana plants exhibited significantly reduced growth and mitochondrial dysfunction when the mtDNA copy number was increased to the degree that each mitochondrion possessed DNA. The amounts of mitochondrion-encoded transcripts increased several fold in the presence of elevated mtDNA levels. However, the efficiency of RNA editing decreased with this excess of mitochondrion-encoded transcripts, resulting in impaired assembly of mitochondrial complexes containing mtDNA-encoded subunits, such as respiratory complexes I and IV. These observations indicate the occurrence of nuclear-mitochondrial incompatibility in the cells with increased amounts of mtDNA and provide an initial answer to the fundamental question of why plant cells have much lower mtDNA levels than animal cells. We propose that keeping mtDNA levels low moderates nuclear-mitochondrial incompatibility and that this may be a crucial factor driving plant cells to restrict the copy numbers of mtDNA.

Development and Preliminary Application of a Droplet Digital PCR Assay for Quantifying the Oncolytic Herpes Simplex Virus Type 1 in the Clinical-Grade Production.

Oncolytic herpes simplex virus (oHSV) is a type of virus that selectively targets and kills cancer cells, leaving normal cells unharmed. Accurate viral titer is of great importance for the production and application of oHSV products. Droplet digital PCR (ddPCR) is known for having good reproducibility, not requiring a standard curve, not being affected by inhibitors, and being precise even in the detection of low copies. In the present study, we developed a droplet digital PCR assay for the quantification of HSV-1 and applied it in the oHSV production. The established ddPCR showed good specificity, linearity, a low limit of quantification, great reproducibility, and accuracy. The quantification result was well-associated with that of plaque assay and CCID50. Amplification of the purified virus without DNA extraction by ddPCR presented similar results to that from the extracted DNA, confirming the good resistance against PCR inhibitors. With the ddPCR, viral titer could be monitored in real time during the production of oHSV; the optimal harvest time was determined for the best virus yield in each batch. The ddPCR can be used as a useful tool for the quantification of oHSV and greatly facilitate the manufacturing process of oHSV products.

Antigen-specific IgG subclass composition in recipient mice can indicate the degree of red blood cell alloimmunization as well as discern between primary and secondary immunization.

Despite the vast antigen disparity between donor and recipient red blood cells (RBCs), only 2%-6% of transfusion patients mount an alloantibody response. Recently, RBC antigen density has been proposed as one of the factors that can influence alloimmunization, however, there has been no characterization of the role of antigen density along with RBC dose in primary and secondary immunization. To generate RBCs that express distinct antigen copy numbers, different quantities of hen egg lysozyme (HEL) were coupled to murine RBCs. The HEL-RBCs were subsequently transfused into recipient mice at different RBC doses and their HEL-specific IgM, IgG, and IgG subclass response was evaluated. Productive immune responses could be generated through a high copy number antigen transfused at low RBC doses or a low copy number transfused at high RBC doses. Further, primary but submaximal humoral immunization predominantly induced the IgG2b and IgG3 subclasses. In contrast, a maximal primary immunization or a secondary immunization induced all four IgG subclasses. Our results confirm the existence of an antigen threshold for productive immune responses but indicate that a high antigen copy number alone might not be enough to induce a response, but rather a combination of both antigen copy number and cell dosage may determine the outcome of immunization. Further, this study provides a proof of concept that the IgG subclass composition can be an indicator of the level of RBC alloimmunization as well as discern between primary and secondary immunization at least in this murine model.

A deep learning based method for automatic analysis of high-throughput droplet digital PCR images.

Droplet digital PCR (ddPCR) is a technique for absolute quantification of nucleic acid molecules and is widely used in biomedical research and clinical diagnosis. ddPCR partitions the reaction solution containing target molecules into a large number of independent microdroplets for amplification and performs quantitative analysis of target molecules by calculating the proportion of positive droplets by the principle of Poisson distribution. Accurate recognition of positive droplets in ddPCR images is of great importance to guarantee the accuracy of target nucleic acid quantitative analysis. However, hand-designed operators are sensitive to interference and have disadvantages such as low contrast, uneven illumination, low sample copy number, and noise, and their accuracy and robustness still need to be improved. Herein, we developed a deep learning-based high-throughput ddPCR droplet detection framework for robust and accurate ddPCR image analysis, and the experimental results show that our method achieves excellent performance in the recognition of positive droplets (99.71%) within a limited time. By combining the Hough transform and a convolutional neural network (CNN), our novel method can automatically filter out invalid droplets that are difficult to be identified by local or global encoding methods and realize high-precision localization and classification of droplets in ddPCR images under variable exposure, contrast, and uneven illumination conditions without the need for image pre-processing and normalization processes.

Mitochondrial DNA Isolation from Plants.

For most eukaryotes, sequencing and assembly of the mitochondrial DNA (mtDNA) is possible by starting the analysis from total cellular DNA, but the exploration of the mtDNA of plants is more challenging because of the low copy number, limited sequence conservation, and complex structure of the mtDNA. The very large size of the nuclear genome of many plant species and the very high ploidy of the plastidial genome further complicate the analysis, sequencing, and assembly of plant mitochondrial genomes. An enrichment of mtDNA is therefore necessary. For this, plant mitochondria are purified prior to mtDNA extraction and purification. The relative enrichment in mtDNA can be assessed by qPCR, while the absolute enrichment can be deduced from the proportion of NGS reads mapping to each of the three genomes of the plant cell. Here we present methods for mitochondrial purification and mtDNA extraction applied to different plant species and tissues, and compare the mtDNA enrichment obtained with the different procedures.

Efficient Quantification of Extrinsic Fluctuations via Stochastic Simulations.

At the molecular level, all the biological processes are exposed to fluctuations emanating from various sources in and around the cellular system. Often these fluctuations dictate the outcome of a cell-fate decision-making event. Thus, having an accurate estimate of these fluctuations for any biological network is extremely important. There are well-established theoretical and numerical methods to quantify the intrinsic fluctuation present within a biological network arising due to the low copy numbers of cellular components. Unfortunately, the extrinsic fluctuations arising due to cell division events, epigenetic regulation, etc. have received very little attention. However, recent studies demonstrate that these extrinsic fluctuations significantly affect the transcriptional heterogeneity of certain important genes. Herein, we propose a new stochastic simulation algorithm to efficiently estimate these extrinsic fluctuations for experimentally constructed bidirectional transcriptional reporter systems along with the intrinsic variability. We use the Nanog transcriptional regulatory network and its variants to illustrate our numerical method. Our method reconciled experimental observations related to Nanog transcription, made exciting predictions, and can be applied to quantify intrinsic and extrinsic fluctuations for any similar transcriptional regulatory network.

Live-cell super-resolution imaging unconventional dynamics and assemblies of nuclear pore complexes.

Super-resolution microscopy has promoted the development of cell biology, but imaging proteins with low copy numbers in cellular structures remains challenging. The limited number of designated proteins within nuclear pore complexes (NPCs) impedes continuous observation in live cells, although they are often used as a standard for evaluating various SR methods. To address this issue, we tagged POM121 with Halo-SiR and imaged it using structured illumination microscopy with sparse deconvolution (Sparse-SIM). Remarkably, POM121-SiR exhibited more than six-fold fluorescence intensity and four-fold enhanced contrast compared to the same protein labeled with tandem-linked mCherry, while showing negligible photo-bleaching during SR imaging for 200 frames. Using this technique, we discovered various types of NPCs, including ring-like and cluster-like structures, and observed dynamic remodeling along with the sequential appearance of different Nup compositions. Overall, Halo-SiR with Sparse-SIM is a potent tool for extended SR imaging of dynamic structures of NPCs in live cells, and it may also help visualize proteins with limited numbers in general.

Enrichment of Membrane Proteins for Downstream Analysis Using Styrene Maleic Acid Lipid Particles (SMALPs) Extraction.

Integral membrane proteins are an important class of cellular proteins. These take part in key cellular processes such as signaling transducing receptors to transporters, many operating within the plasma membrane. More than half of the FDA-approved protein-targeting drugs operate via interaction with proteins that contain at least one membrane-spanning region, yet the characterization and study of their native interactions with therapeutic agents remains a significant challenge. This challenge is due in part to such proteins often being present in small quantities within a cell. Effective solubilization of membrane proteins is also problematic, with the detergents typically employed in solubilizing membranes leading to a loss of functional activity and key interacting partners. In recent years, alternative methods to extract membrane proteins within their native lipid environment have been investigated, with the aim of producing functional nanodiscs, maintaining protein-protein and protein-lipid interactions. A promising approach involves extracting membrane proteins in the form of styrene maleic acid lipid particles (SMALPs) that allow the retention of their native conformation. This extraction method offers many advantages for further protein analysis and allows the study of the protein interactions with other molecules, such as drugs. Here, we describe a protocol for efficient SMALP extraction of functionally active membrane protein complexes within nanodiscs. We showcase the method on the isolation of a low copy number plasma membrane receptor complex, the nicotinic acetylcholine receptor (nAChR), from adult Drosophila melanogaster heads. We demonstrate that these nanodiscs can be used to study native receptor-ligand interactions. This protocol can be applied across many biological scenarios to extract the native conformations of low copy number integral membrane proteins.

Detection of miRNAs.

MicroRNAs (miRNAs) are small, noncoding RNAs that regulate gene expression. They play an important role in many biological processes including human diseases. However, miRNAs are challenging to detect due to their short sequence length and low copy number. A number of conventional (e.g., Northern blot, microarray, and RT-qPCR) and emerging (e.g., nanostructured materials and electrochemical methods) techniques have been developed to detect miRNA, each with their own strengths and weaknesses. Some of these techniques have been combined to detect miRNAs as disease biomarkers in point-of-care (POC) settings. Nonetheless, there is still potential for further innovation to facilitate the detection of miRNAs.

A novel narnavirus is widespread in Saccharomyces cerevisiae and impacts multiple host phenotypes.

RNA viruses are a widespread, biologically diverse group that includes the narnaviridiae, a family of unencapsidated RNA viruses containing a single ORF that encodes an RNA-dependent RNA polymerase. In the yeast Saccharomyces cerevisiae, the 20S and 23S RNA viruses are well-studied members of the narnaviridiae, which are present at low intracellular copy numbers, unless induced by stress or unfavorable growth conditions (Wickneret al. 2013), and are not known to affect host fitness. In this study, we describe a new Saccharomyces cerevisiae narnavirus that we designate as N1199. We show that N1199 is uniquely present as a double-stranded RNA at a high level relative to other known members of this family in one strain background, YJM1199, and is present as a single-stranded RNA at lower levels in 98 of the remaining 100-genomes strains. Further, we see a strong association between the presence of high level N1199 and host phenotype defects, including greatly reduced sporulation efficiency and growth on multiple carbon sources. Finally, we describe associations between N1199 abundance and host phenotype defects, including autophagy.

Using Whole Slide Gray Value Map to Predict HER2 Expression and FISH Status in Breast Cancer.

Accurate detection of HER2 expression through immunohistochemistry (IHC) is of great clinical significance in the treatment of breast cancer. However, manual interpretation of HER2 is challenging, due to the interobserver variability among pathologists. We sought to explore a deep learning method to predict HER2 expression level and gene status based on a Whole Slide Image (WSI) of the HER2 IHC section. When applied to 228 invasive breast carcinoma of no special type (IBC-NST) DAB-stained slides, our GrayMap+ convolutional neural network (CNN) model accurately classified HER2 IHC level with mean accuracy 0.952 ± 0.029 and predicted HER2 FISH status with mean accuracy 0.921 ± 0.029. Our result also demonstrated strong consistency in HER2 expression score between our system and experienced pathologists (intraclass correlation coefficient (ICC) = 0.903, Cohen's κ = 0.875). The discordant cases were found to be largely caused by high intra-tumor staining heterogeneity in the HER2 IHC group and low copy number in the HER2 FISH group.

Genetic diversity of transmission-blocking vaccine candidate antigens Pvs25 and Pvs28 in Plasmodium vivax isolates from China

BackgroundTransmission-blocking vaccines (TBVs) target the sexual stages of malaria parasites to reduce or interrupt the transmission cycle in human and mosquito populations. The genetic diversity of TBVs candidate antigens, Pvs25 and Pvs28, in Plasmodium vivax could provide evidence for the development of TBVs.MethodsDry blood spots from P. vivax patients were collected from Dandong, Suining, Hainan, Nyingchi, Tengchong, and Yingjiang in China. The pvs25 and pvs28 genes were amplified and sequenced. The genetic diversity of pvs25 and pvs28 were analyzed using DNASTAR, MEGA6, and DnaSP 5.0 programs.ResultsA total of 377 samples were collected, among which 324 and 272 samples were successfully amplified in the pvs25 and pvs28 genes, respectively. Eight haplotypes were identified in Pvs25, for which the predominant mutation was I130T with 100% prevalence. A variety of 22 haplotypes in Pvs28 were identified. The number of GSGGE/D repeats of Pvs28 was a range of 4–8, among which, high (7–8) and low (4–5) copy numbers of tandem repeats were found in haplotypes H2 and H17, respectively. The nucleotide diversity of pvs28 (π = 0.00305 ± 0.00061) was slightly higher than that of pvs25 (π = 0.00146 ± 0.00007), thus they were not significantly different (P > 0.05). The Tajima's D value of pvs25 was positive whereas pvs28 was negative, which indicated that both genes were affected by natural selection.ConclusionThe genetic diversity of pvs25 and pvs28 genes in China was relatively limited, which provided valuable information for TBVs design and optimization.

Revised molecular phylogeny, global biogeography, and diversification of palms subfamily Coryphoideae (Arecaceae) based on low copy nuclear and plastid regions.

Coryphoideae are palmate-leaved palms from the family Arecaceae consisting of 46 genera representing 421 species. Although several phylogenetic analyses based on different genomic regions have been carried out on Coryphoideae, a fully resolved molecular phylogenetic tree has not been reported yet. To achieve this, we applied two phylogenetic reconstruction methods: Maximum Likelihood and Bayesian Inference, using amplified sampling by retrieving chloroplast and nuclear DNA sequences from NCBI and adding newly produced sequences from Indian accession into the dataset. The same dataset (chloroplast + nuclear DNA sequences) was used to estimate divergence times and the evolutionary history of Coryphoideae with a Bayesian uncorrelated, lognormal relaxed-clock approach and a Statistical Divergence-Vicariance Analysis method,respectively. The phylogenetic analyses based on a combined chloroplast and nuclear DNA sequence dataset showed well-resolved relationships within the subfamily. Both phylogenetic trees divide Coryphoideae into two main groups: CSPT (Crysophileae, Sabaleae, Phoeniceae, and Trachycarpeae) and the Syncarpous group. These main groups are segregated into eight tribes (Trachycarpeae, Phoeniceae, Sabaleae, Crysophileae, Borasseae, Corypheae, Caryoteae, and Chuniophoeniceae) and four subtribes (Rhapidine, Livistoninae, Hyphaeninae, and Lataniinae) with strong support-values. Most previously unresolved and doubtful relationships within tribes Trachycarpeae and Crysophilieae are now resolved and well-supported. The reconstructed phylogenetic trees support all previous systematic revisions of the subfamily. All Indian sampled species of Arenga, Bentinckia, Hyphaene, and Trachycarpus show close relation with their respective congeneric species. Molecular dating results and integration of biogeography suggest that Coryphoideae originated in Laurasia at ~95.12Ma and then diverged into the tropical and subtropical regions of the whole world. This study offers the correct combination of nuclear and plastid regions to test the current and future systematic revisions.

525. A Universal ‘Day Zero’ Infectious Disease Testing Strategy Exploiting Crispr-Based Sample Depletion And Metagenomic Sequencing

Abstract Background The lack of preparedness for detecting the highly infectious SARS-CoV-2 pathogen — the pathogen responsible for the COVID-19 disease — caused enormous harm to the public health, the economy and society as a whole. It took ∼60 days for the first RT-PCR tests for SARS-CoV-2 infection developed by the United States Centers for Disease Control (CDC) to be made available. It then took >270 days to deploy 800,000 of these tests at a time when the estimated actual testing needs required over 6 million tests per day. Testing was therefore limited to only individuals with symptoms or individuals in close contact with confirmed positive cases. Testing strategies that can be deployed on a population scale at ‘day zero’ (i.e., at the time of the first reported case) are needed. Next Generation Sequencing (NGS) has such day zero capabilities with the potential to enable feasible and broad large-scale testing strategies, however it has limited detection sensitivity for low copy numbers of pathogens which may be present. Here we demonstrate that using CRISPR-Cas9 to remove abundant sequences that do not contribute to pathogen detection, NGS detection sensitivity is equivalent to RT-PCR. In addition, we show that this assay can be used for variant strain typing, co-infection detection, and individual human host response assessment – all in a single workflow using existing open-source analysis pipelines. This NGS workflow is pathogen agnostic, and therefore has the potential to radically transform how both very large-scale pandemic response and focused clinical infectious disease testing are pursued in the future. microbial composition at two test sites Detection Sensitivity Methods Covid positive samples with RT-PCR Ct values from 16-39 were processed through the CRISRP enhanced mNGS pipeline. Data processing workflow Results Sn/Sp compared to RT-PCR was 97%/100%. Strain calling concordance compared to amplicon sequencing was 100%. Co-infections from Covid positive samples were identified with high confidence. Host response signatures match the published literature. Conclusion Applying CRISPR enhanced metagenomic NGS at Day Zero of the next pandemic can mitigate the time gap in developing approved diagnostics at population scale and potentially save lives. Disclosures All Authors: No reported disclosures.

1653. Transcriptional Variation of Adhesion and Biofilm Genes in CTX-M-15-Expressing Klebsiella pneumoniae

Abstract Background Klebsiella pneumoniae can produce a number of β-lactamases, including the cefotaximase, CTX-M-15. While classically associated with ST131 Escherichia coli, CTX-M-15 is associated with increased virulence in K. pneumoniae. Clinical isolates of K. pneumoniae have been shown to produce high levels of CTX-M-15 RNA. But, the overall transcriptomic changes due to CTX-M-15 expression are unknown. The purpose of this study was to determine if acquisition of CTX-M-15 altered the global transcriptome of K. pneumoniae in addition to decreasing β-lactam susceptibility. Methods The structural gene and native promoter of CTX-M-15 were cloned into a low copy number vector and transformed into Kp 23, a clinical isolate from the 1970s with no known acquired β-lactam resistance genes. CTX-M-15 production was confirmed by qRT PCR and western blot. MICs to cefotaxime, ceftazidime, cefepime, ceftolozane/tazobactam, ceftazidime/avibactam, meropenem, meropenem-vaborbactam, and imipenem-relebactam were determined by E-test according to CLSI guidelines. RNA-Seq analysis and bioinformatics were performed by MiGS. Results CTX-M-15 carriage led to non-susceptible MICs to cefotaxime, ceftazidime, and cefepime. CTX-M-15 production was associated with statistically significant upregulation of 204 genes and downregulation of 231 genes. Among these, the biofilm genes mrkA and two copies of mrkB were downregulated by 7.5, 4 and 3.5-fold, respectively. Additionally, ecpA, ecpB, ecpC, and ecpR – components of the E. coli Common Pilus (ECP) – were upregulated by 3, 2, 2, and 3-fold, respectively. Conclusion High-level expression of CTX-15 β-lactamase has a significant impact on the transcriptome of K. pneumoniae. These changes include a down regulation of genes associated with biofilm formation with a concomitant increase in the expression of genes associated with adherence. These data indicate that overexpression of CTX-M-15 and perhaps other β-lactamases may help promote the pathogenicity of K. pneumoniae or influence its ability to colonize the tissue it infects. This may be important during urinary tract infections when an increase in adhesion by CTX-M-15 producing isolates may be required over biofilm formation to establish an active infection. Disclosures Nancy D. Hanson, PhD, Merck: Grant/Research Support.