Biology subjects

Zhang, M.

Publications and source records attributed to Zhang, M..

At least 487 records · Page 27Linked to original sources

bioRxiv2022

ATM-Mediated Double-Strand Break Repair Is Required for Meiotic Genome Stability at High Temperature

In eukaryotes, the conserved kinase Ataxia Telangiectasia Mutated (ATM) negatively regulates DNA double-strand break (DSB) formation and plays a central role in DSB repair. Here, by using cytogenetic approaches, we demonstrate that ATM also plays an essential role in protecting meiotic chromosome integrity in Arabidopsis thaliana at extreme high temperature. We determined the chromosome localization patterns of DSB formation proteins SPO11-1 and DFO during prophase I, both of which were disturbed by heat stress. Evaluation of the number of RAD51, DMC1, SPO11-1 and DFO protein foci in meiocytes of Arabidopsis atm mutant clarifies that ATM does not mediate the heat-induced reduction in DSB formation. Interestingly, meiotic spread analysis showed that chromosome fragmentation level was significantly increased in atm but was lowered in the mre11 and mre11 atm mutants under high temperature, indicating that ATM-dependent meiotic chromosome integrity at high temperature relies on the functional MRE1-RAD50-NBS1 (MRN) complex. Moreover, contrary to the rad51 and mnd1 mutants, which exhibited enhanced meiotic chromosome integrity under heat stress, the rad51 atm and mnd1 atm mutants retained high levels of chromosome fragmentation at extreme high temperature. Furthermore, heat stress reduced chromosome fragmentation level in the syn1 and syn1 atm mutants. Collectively, these data suggest that ATM-mediated DSB repair is required for meiotic genome stability in plants at extreme high temperature, which possibly acts in a RAD51-independent manner and relies on functional chromosome axis.

plant biology↗

bioRxiv2022

Thalamic volume and functional connectivity are associated with nicotine dependence severity and craving

Tobacco smoking is associated with deleterious health outcomes. Most smokers want to quit smoking, yet relapse rates are high. Understanding neural differences associated with tobacco use may help generate novel treatment options. Several animal studies have recently highlighted the central role of the thalamus in substance use disorders, but this research focus has been understudied in human smokers. Here, we investigated associations between structural and functional magnetic resonance imaging measures of the thalamus and its subnuclei to distinct smoking characteristics. We acquired anatomical scans of 32 smokers as well as functional resting-state scans before and after a cue-reactivity task. Thalamic functional connectivity was associated with craving and dependence severity, whereas the volume of the thalamus was associated with dependence severity only. Craving, which fluctuates rapidly, was best characterized by differences in brain function, whereas the rather persistent syndrome of dependence severity was associated with both brain structural differences and function. Our study supports the notion that functional versus structural measures tend to be associated with behavioral measures that evolve at faster versus slower temporal scales, respectively. It confirms the importance of the thalamus to understand mechanisms of addiction and highlights it as a potential target for brain-based interventions to support smoking cessation, such as brain stimulation and neurofeedback.

neuroscience↗

bioRxiv2022

Pre- and post-task resting-state differs in clinical populations

Resting-state functional connectivity has generated great hopes as a potential brain biomarker for improving prevention, diagnosis, and treatment in psychiatry. This neuroimaging protocol can routinely be performed by patients and does not depend on the specificities of a task. Thus, it seems ideal for big data approaches that require aggregating data across multiple studies and sites. However, technical variability, diverging data analysis approaches, and differences in data acquisition protocols might introduce heterogeneity to the aggregated data. Besides these technical aspects, the psychological state of participants might also contribute to heterogeneity. In healthy participants, studies have shown that behavioral tasks can influence resting-state measures, but such effects have not yet been reported in clinical populations. Here, we fill this knowledge gap by comparing resting-state functional connectivity before and after clinically relevant tasks in two clinical conditions, namely substance use disorders and phobias. The tasks consisted of viewing craving-inducing and spider anxiety provoking pictures that are frequently used in cue-reactivity studies and exposure therapy. We found distinct pre- vs. post-task resting-state connectivity differences in each group, as well as decreased thalamo-cortical and increased intra-thalamic connectivity which might be associated with decreased vigilance in both groups. Notably, the pre- vs. post-task thalamus-amygdala connectivity change within a patient cohort seems more pronounced than the difference of that connection between the smoker vs. phobia clinical trait. Our results confirm that resting-state measures can be strongly influenced by changes in psychological states that need to be taken into account when pooling resting-state scans for clinical biomarker detection. This demands that resting-state datasets should include a complete description of the experimental design, especially when a task preceded data collection.

neuroscience↗

bioRxiv2022

MalariaSED: a deep learning framework to decipher the regulatory contributions of noncoding variants in malaria parasites

Malaria remains one of the deadliest infectious diseases. Transcriptional regulation effects of noncoding variants in this unusual genome of malaria parasites remain elusive. We developed a sequence-based, ab initio deep learning framework, MalariaSED, for predicting chromatin profiles in malaria parasites. The MalariaSED performance was validated by published ChIP-qPCR and TF motifs results. Applying MalariaSED to [~]1.3 million variants shows that geographically differentiated noncoding variants are associated with parasite invasion and drug resistance. Further analysis reveals chromatin accessibility changes at Plasmodium falciparum rings are partly associated with artemisinin resistance. MalariaSED illuminates the potential functional roles of noncoding variants in malaria parasites.

bioinformatics↗

bioRxiv2022

SOGO-SOFI, light-modulated super-resolution optical fluctuation imaging using only 20 raw frames for high-fidelity reconstruction

Taking advantage of the stochastic photoswitching of genetically encodable reversibly photoswitchable fluorescent proteins (RSFPs), super-resolution optical fluctuation imaging (SOFI) and its variant photochromic stochastic optical fluctuation imaging (pcSOFI) are valuable tools for wide field super- resolution (SR) imaging. Live-cell (pc)SOFI, which requires a small number of original frames to reconstruct an SR image, is prone to structural discontinuity artifacts and low spatial resolution. Herein, we developed a repeated synchronized on- and gradually off-switching SOFI (SOGO-SOFI) that maximized the photoswitching frequency of RSFPs by light modulation and required only 20 frames for high-quality reconstruction. Live-cell SOGO-SOFI imaging of the endoplasmic reticulum (ER) exhibited 10 times higher temporal resolution (100 fps) and fewer artifacts than pcSOFI. Moreover, a combination of SOGO-SOFI with Airyscan further increased the image contrast and the resolution of Airyscan by a factor of 1.5 from 140 nm to 91 nm. The capabilities of SOGO-SOFI were further demonstrated by dual- color imaging of nucleolar proteins in mammalian cells and deep imaging of ER structures in thick brain slices (20.6 {micro}m).

biophysics↗

bioRxiv2022

Activation of ERK by altered RNA splicing in cancer

Many cancers carry change-of-function mutations affecting RNA splicing factors, however, less is known about the functional consequences of upregulated RNA splicing factors in cancer. Here, we demonstrate that SMNDC1, a poorly studied splicing factor, which we found to be upregulated in multiple carcinomas and associated with poor patient prognosis, promotes cell proliferation, clonal expansion, and tumor growth by promoting the retention of G-rich exons, which otherwise would be excluded or retained at a lower rate after RNA splicing in normal cells. Inclusion of exon 4 (E4) of MAPK3 (ERK1), which encodes both kinase phosphorylation sites (Thr202/Tyr204), was among the promoted exons by SMNDC1. Forced exclusion of MAPK3-E4 using anti-sense oligos inhibited the ERK1 phosphorylation, expression of target genes and decreased tumor cell growth. These data support that cancer cells exploit a "splicing switch" to promote ERK kinase activity and offer a druggable alternative to block oncogenic signaling and altered RNA splicing in cancer cells SIGNIFICANCEERK signaling promotes tumor growth and survival. Exon 4 of MAPK3 (ERK1) encodes the activation phosphorylation sites of ERK1 kinase. Aberrant RNA splicing induced by SMNDC1 in cancer cells increases the retention of exon 4 during mRNA splicing, unleashes the kinase activity. SMNDC1 potentializes as a cancer therapeutic target.

cancer biology↗

bioRxiv2022

Myoglobin promotes cardiomyocyte differentiation through oxidatively modulating the Hippo Kinase Pathway

BackgroundWhile cardiomyocytes undergo terminal differentiation postnatally and rarely re-enter the cell cycle, the endogenous mechanisms that propagate differentiation and prevent de-differentiation remain unclear. The monomeric heme protein myoglobin, which stores oxygen and regulates reactive oxygen/nitrogen species balance in the heart, increases in expression by over 50% during cardiomyocyte differentiation. Though myoglobin deletion without significant compensation is embryonic lethal in mice, a role for the protein in regulating cardiomyocyte differentiation has not been tested. We hypothesized that myoglobin expression is required for cardiomyocyte differentiation and the loss of myoglobin enables de-differentiation. MethodsMyoglobin was genetically silenced in HL-1, H9C2 cells, and neonatal rat ventricular cardiomyocytes (NRVM) to examine myoglobin-dependent effects on differentiation, proliferation, and Hippo pathway signaling. A zebrafish model of Mb depletion was made using CRISPR-Cas9 to test the effect of myoglobin depletion on cardiac regeneration after apical resection injury in vivo. ResultsMyoglobin deletion in cultured cell lines and NRVM decreased the gene expression of cardiomyocyte differentiation markers (troponin, myosin light chain, and myosin heavy chain), upregulated markers of dedifferentiation (runx1 and dab2) and stimulated cell proliferation. Mechanistically, we show that the heme prosthetic group of myoglobin catalyzes the oxidation of the Hippo pathway kinase LATS1, which activates the enzyme to phosphorylate the downstream Yes-associated protein (YAP) transcription factor, which prevents its transcriptional activity. Thus, the loss of myoglobin results in the de-phosphorylation and nuclear translocation of YAP, which propagates proliferation and fetal gene expression. In vivo, myoglobin-deficient zebrafish hearts recapitulated the increase in YAP signaling and showed accelerated regeneration at 20 days post apical injury. ConclusionWe a novel role for myoglobin as an endogenous driver of cardiomyocyte differentiation, and a regulator of the Hippo pathway. These findings suggest myoglobin as a potential target for strategies to enhance cardiac development and improve cardiac repair and regeneration.

cell biology↗

bioRxiv2022

Identification of transcriptional landscapes and functions of the inhibition of ARHGEF2 in hepatocellular carcinoma cells

The RHO guanine exchange factor ARHGEF2 has exchange activity toward RHOA, which is essential for the development of cancers such as liver cancer. However, the potential functions and mechanisms of ARHGEF2 in the progression of liver cancer are largely unknown. In this study, we identified the transcriptional landscapes of hepatocellular carcinoma cells treated with ARHGEF2 shRNAs. The gene enrichment assays such as KEGG and GO were used to further analyze the potential signaling pathways. Moreover, the PPI network and Reactome map were used to further identify the biological processes. The results showed that Alzheimers disease disease (AD) and Cushing syndrome (CS) are the major signaling pathways involved in the ARHGEF2-shRNAs treated hepatocellular carcinoma cells. We identified the top ten interactive genes including ICAM1, APOE, LDLR, NAT10, HSPA1A, EDN1, CACNA1C, KCNMA1, SNAI1, and ELN. Our study may provide novel mechanisms for the treatment of liver cancer by inhibiting ARHGEF2.

bioinformatics↗

bioRxiv2022

Apolipoprotein E genotype modulates longitudinal atrophy at the temporal lobe after mild traumatic brain injury

BackgroundsModerate or severe traumatic brain injury (TBI) is one of the strongest environmental risks for late-life dementia, especially Alzheimers disease (AD). However, the interaction between genetic factors and environmental exposure to mild TBI triggering neurodegenerative processes is still unclear. We used longitudinal imaging to differentiate spatial patterns of progressive brain volume loss in individual patients with and without carrying apolipoprotein E (APOE) {varepsilon}4 carriers. MethodsFor 59 patients with acute mild TBI (mTBI, age: 36.92 {+/-} 11.91 years; 14 APOE {varepsilon}4 carriers) and 48 matched healthy controls (HCs, age: 37.92 {+/-} 11.72 years; 10 APOE {varepsilon}4 carriers), longitudinal (6-12 months follow-up) brain structure was assessed using voxel-based morphometry on T1-weighted scans. Longitudinal volume changes in the temporal lobe were characterized by the Jacobian determinant (JD) metric, reflecting spatial warping between the baseline and follow-up scans. JD values were regionally calculated and correlated with neuropsychological measures. ResultsMTBI patients lost a mean of 0.27% (SD = 0.43) of GM and 0.27% (SD = 0.50) of WM volume in the temporal lobe over the 6-12 month follow-up. Patients with a high genetic risk for AD (APOE {varepsilon}4 allele carries) were associated with severer atrophy in the left superior temporal gyrus and middle temporal gyrus regions than the controls (P < 0.05). Furthermore, atrophy in these regions of gray matter could predict the performance change ratio of the language fluency in mTBI APOE {varepsilon}4 carriers ({beta} = 0.570, P < 0.05). While, compared with HC APOE {varepsilon}4 non-carriers, mTBI non-carriers exhibited volume loss in the medial temporal lobe. ConclusionsThis prospective study provided evidence that the APOE {varepsilon}4 allele interacting with mTBI increased the risk of AD phenotype development with language dysfunction. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/503827v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@39af96org.highwire.dtl.DTLVardef@136a0f7org.highwire.dtl.DTLVardef@181deb4org.highwire.dtl.DTLVardef@144349d_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2022

A near complete genome assembly of chia assists in identification of key fatty acid desaturases in developing seeds

Chia is an annual crop whose seeds have the highest content of -linolenic acid (ALA) of any plant species. We generated a high-quality assembly of the chia genome using circular consensus sequencing of PacBio. The assembled six chromosomes are composed of 21 contigs and have a total length of 361.7 Mb. Genome annotation revealed a 53.5% repeat content and 35,850 protein-coding genes. Chia shared a common ancestor with Salvia splendens ~6.1 million years ago. Utilizing the reference genome and two transcriptome datasets, we identified candidate fatty acid desaturases responsible for ALA biosynthesis during chia seed development. Because the seed of S. splendens contains significantly lower proportion of ALA but similar total contents of unsaturated fatty acids, we suggest that strong expression of two ShFAD3 genes are critical for the high ALA content of chia seeds. This genome assembly will serve as a valuable resource for breeding, comparative genomics, and functional genomics studies of chia.

genomics↗

bioRxiv2022

The topology of time: transition networks in simulated and real neural dynamics

Characterizing large-scale dynamic organization of the brain relies on both data-driven and mechanistic modeling, which demands a low vs. high level of prior knowledge and assumptions about how constituents of the brain interact. However, the conceptual translation between the two is not straightforward. The present work aims to provide a bridge between data-driven and mechanistic modeling. We conceptualize brain dynamics as a complex landscape that is continuously modulated by internal and external changes. The modulation can induce transitions between one stable brain state (attractor) to another. Here, we provide a novel method - Temporal Mapper - built upon established tools from the field of Topological Data Analysis to retrieve the network of attractor transitions from time-series data alone. For theoretical validation, we use a biophysical network model to induce transitions in a controlled manner, which provides simulated time series equipped with a ground-truth attractor transition network. Our approach reconstructs the ground-truth transition network from simulated time-series data better than existing time-varying approaches. For empirical relevance, we apply our approach to fMRI data gathered during a continuous multitask experiment. We found that occupancy of the high-degree nodes and cycles of the transition network was significantly associated with subjects behavioral performance. Taken together, we provide an important first step towards integrating data-driven and mechanistic modeling of brain dynamics.

neuroscience↗

bioRxiv2022

Biological condensates form percolated networks with molecular motion properties distinctly different from dilute solutions

Formation of membraneless organelles or biological condensates via phase separation hugely expands cellular organelle repertoire. Biological condensates are dense and viscoelastic soft matters instead of canonical dilute solutions. Unlike discoveries of numerous different biological condensates to date, mechanistic understanding of biological condensates remains scarce. In this study, we developed an adaptive single molecule imaging method that allows simultaneous tracking of individual molecules and their motion trajectories in both condensed and dilute phases of various biological condensates. The method enables quantitative measurements of phase boundary, motion behavior and speed of molecules in both condensed and dilute phases as well as the scale and speed of molecular exchanges between the two phases. Surprisingly, molecules in the condensed phase do not undergo uniform Brownian motion, but instead constantly switch between a confined state and a random motion state. The confinement is consistent with formation of large molecular networks (i.e., percolation) specifically in the condensed phase. Thus, molecules in biological condensates behave distinctly different from those in dilute solutions. This finding is of fundamental importance for understanding molecular mechanisms and cellular functions of biological condensates in general.

cell biology↗

bioRxiv2022

Brassinosteroids signaling component SlBZR1 promotes fruit ripening in tomato

Fruit ripening evolved to be attractive to frugivores that derive energy and nutrition from the fruits in exchange for assisting seed dispersal, which is accompanied by the dramatically change of fruit characteristics, including color, aroma, and texture. The plant hormone ethylene plays a key role in climacteric fruit ripening, while the role of other phytohormones as well as their cross talk with ethylene in modulating fruit ripening remains elusive. Here, we report growth-promoting phytohormone brassinosteroids promote fruit ripening in tomato through regulation of ethylene biosynthesis. Exogenous BR treatment and the increase of endogenous BR content in SlCYP90B3-OE promoted ethylene production and fruit ripening. SlBZR1, a central component and positive regulator of BR signaling pathway, promotes ethylene production and carotenoid accumulation through direct transcriptional regulation of SlACO1, SlACO3 and SlPSY1. Furthermore, SlBIN2, a negative regulator of BR signaling upstream of SlBZR1, decreases ethylene production and carotenoid accumulation. Together, our results demonstrate that BR signaling integrates ethylene and carotenoid biosynthetic pathway to regulate fruit ripening.

plant biology↗

bioRxiv2022

Noninvasive cancer detection by extracting and integrating multi-modal data from whole-methylome sequencing of plasma cell-free DNA

AbstractPlasma cell-free DNA (cfDNA) methylation and fragmentation signatures have been shown to be valid biomarkers for blood-based cancer detection. However, conventional methylation sequencing assays are inapplicable for fragmentomic profiling due to bisulfite-induced DNA damage. Here using enzymatic conversion-based low-pass whole-methylome sequencing (WMS), we developed a novel approach to comprehensively interrogate the genome-wide plasma methylation, fragmentation, and copy number profiles for sensitive and noninvasive multi-cancer detection. With plasma WMS data from a clinical cohort comprising 497 healthy controls and 780 patients with both early- and advanced-stage cancers of the breast, colorectum, esophagus, stomach, liver, lung, or pancreas, genomic features including methylation, fragmentation size, copy number alteration, and fragment end motif were extracted individually and subsequently integrated to develop an ensemble cancer classifier, called THEMIS, using machine learning algorithms. THEMIS outperformed individual biomarkers for differentiating cancer patients of all seven types from healthy individuals and achieved a combined area under the curve value of 0.971 in the independent test cohort, translating to a sensitivity of 86% and early-stage (I and II) sensitivity of 77% at 99% specificity. In addition, we built a cancer signal origin classifier with true-positive cancer samples at 100% specificity based on methylation and fragmentation profiling of tissue-specific accessible regulatory elements, which localized cancer-like signal to a limited number of clinically informative sites with 66% accuracy. Overall, this proof-of-concept work demonstrates the feasibility of extracting and integrating multi-modal biomarkers from a single WMS run for noninvasive detection and localization of common cancers across stages.

cancer biology↗

bioRxiv2022

α1 adrenergic receptor - PKC - Pyk2 - Src signaling boosts L-type Ca2+ channel Cav1.2 activity and long-term potentiation in rodents

The cellular mechanisms mediating norepinephrine functions in brain to result in behaviors are unknown. We identified the L-type Ca2+ channel (LTCC) CaV1.2 as a principal target for Gq- coupled 1-adrenergic receptors (ARs). 1AR signaling increased LTCC activity in hippocampal neurons. This regulation required PKC-mediated activation of the tyrosine kinases Pyk2 and, downstream, Src. Pyk2 and Src were associated with CaV1.2. In model neuroendocrine PC12 cells, stimulation of PKC induced tyrosine phosphorylation of CaV1.2, a modification abrogated by inhibition of Pyk2 and Src. Upregulation of LTCC activity by 1AR and formation of a signaling complex with PKC, Pyk2, and Src suggests that CaV1.2 is a central conduit for signaling by norepinephrine. Indeed, a form of hippocampal LTP in young mice requires both the LTCC and 1AR stimulation. Inhibition of Pyk2 and Src blocked this LTP, indicating that enhancement of CaV1.2 activity via 1AR - Pyk2 - Src signaling regulates synaptic strength.

neuroscience↗

bioRxiv2022

Visualization of a Limonene Synthesis Metabolon inside Living Bacteria by Hyperspectral SRS Microscopy

Metabolons consisting of cellular structure elements and sequential metabolic enzymes are expected to be involved in diverse biological processes. However, direct visualization of metabolons in prokaryotic cells is still challenging. In this study, we report direct visualization of concentrated subcellular regions of limonene synthesis inside single engineered Escherichia coli by using hyperspectral stimulated Raman scattering (hSRS) microscopy. Equipped with spectral unmixing, hSRS imaging provides a reliable method to quantify intracellular limonene content. In E. coli strains with a complete limonene synthesis pathway, intracellular limonene is found locally concentrated and colocalized with proteins. Furthermore, dual-modality SRS and two-photon fluorescence imaging showed colocalization of limonene and GFP-fused limonene synthase. Significance StatementMonitoring biosynthesis activity at the single-cell level is key to metabolic engineering but is still difficult to achieve in a label-free manner. Using hyperspectral stimulated Raman scattering imaging in the 670-900 cm-1 region, we visualized localized limonene synthesis inside engineered E. coli. The colocalization of limonene and GFP-fused limonene synthase was confirmed by co-registered stimulated Raman scattering and two-photon fluorescence images. Our finding suggests a limonene synthesis metabolon with a polar distribution inside the cells. This finding expands our knowledge of de novo limonene biosynthesis in engineered bacteria and highlights the potential of SRS chemical imaging in metabolic engineering research.

biophysics↗

bioRxiv2022

A prebiotic diet modulates microglial states and motor deficits in α-synuclein overexpressing mice

Parkinsons disease (PD) is a movement disorder characterized by neuroinflammation, -synuclein pathology, and neurodegeneration. Most cases of PD are non-hereditary, suggesting a strong role for environmental factors, and it has been speculated that disease may originate in peripheral tissues such as the gastrointestinal (GI) tract before affecting the brain. The gut microbiome is altered in PD and may impact motor and GI symptoms as indicated by animal studies, though mechanisms of gut-brain interactions remain incompletely defined. Intestinal bacteria ferment dietary fibers into short-chain fatty acids, with fecal levels of these molecules differing between PD and healthy controls and in mouse models. Among other effects, dietary microbial metabolites can modulate activation of microglia, brain-resident immune cells implicated in PD. We therefore investigated whether a fiber-rich diet influences microglial function in -synuclein overexpressing (ASO) mice, a preclinical model with PD-like symptoms and pathology. Feeding a prebiotic high-fiber diet attenuates motor deficits and reduces -synuclein aggregation in the substantia nigra of mice. Concomitantly, the gut microbiome of ASO mice adopts a profile correlated with health upon prebiotic treatment, which also reduces microglial activation. Single-cell RNA-seq analysis of microglia from the substantia nigra and striatum uncovers increased pro-inflammatory signaling and reduced homeostatic responses in ASO mice compared to wild-type counterparts on standard diets. However, prebiotic feeding reverses pathogenic microglial states in ASO mice and promotes expansion of protective disease-associated macrophage (DAM) subsets of microglia. Notably, depletion of microglia using a CSF1R inhibitor eliminates the beneficial effects of prebiotics by restoring motor deficits to ASO mice despite feeding a prebiotic diet. These studies uncover a novel microglia-dependent interaction between diet and motor symptoms in mice, findings that may have implications for neuroinflammation and PD.

physiology↗

bioRxiv2022

Development of zebrafish (Danio rerio) in vivo model for Borrelia burgdorferi infection

Borrelia burgdorferi is the spirochetal bacterium that causes Lyme disease. Despite the fact that antimicrobial sensitivity of B. burgdorferi has been widely studied, there is still a need to develop an affordable, practical, high-throughput in vivo model which can be used to find effective antibiotic therapies, especially for the recently discovered persister and biofilm forms. Recent studies showed that Zebrafish (Danio rerio) could offer a novel, high-throughput, affordable model in antibiotic therapies for various infections agents can be studied. Therefore, in this study, we developed a straightforward, standardized infection procedure and tested it for zebrafish survival rate, morphological or behavioral changes post-infection as well as providing evidence that B. burgdorferi persists in zebrafish using polymerase chain reaction (PCR) and immunohistochemical (IHC) techniques. Morphological and physiological examination showed a significant size difference between control and infected zebrafish at 2 wpi, and infected zebrafish exhibited slower heart rates through 72 hpi. Furthermore, our results showed B. burgdorferi DNA can be detected and active replication of the B. burgdorferi 16S rRNA gene can be confirmed through 10 days post-infection via PCR and Reverse Transcription PCR respectively. Fluorescent microscopy and immunohistochemical staining revealed spirochetes present in the eyes, gills, heart, liver, tail, and hindbrain tissues though 72 hpi as well as the stomach and digestive tract at 2 weeks post-infection, respectively. These findings demonstrate that zebrafish could serve as a promising animal model to study the mechanism of B. burgdorferi infection as well as in vivo antibiotic sensitivity.

microbiology↗