Preliminary insight on diarylpentanoids as potential antimalarials: In silico, in vitro pLDH and in vivo zebrafish toxicity assessment.

Malaria remains a major public health problem worldwide, including in Southeast Asia. Chemotherapeutic agents such as chloroquine (CQ) are effective, but problems with drug resistance and toxicity have necessitated a continuous search for new effective antimalarial agents. Here we report on a virtual screening of ∼300 diarylpentanoids and derivatives, in search of potential Plasmodium falciparum lactate dehydrogenase (PfLDH) inhibitors with acceptable drug-like properties. Several molecules with binding affinities comparable to CQ were chosen for in vitro validation of antimalarial efficacy. Among them, MS33A, MS33C and MS34C are the most promising against CQ-sensitive (3D7) with EC50 values of 1.6, 2.5 and 3.1 µM, respectively. Meanwhile, MS87 (EC50 of 1.85 µM) shown the most active against the CQ-resistant Gombak A strain, and MS33A and MS33C the most effective P. knowlesi inhibitors (EC50 of 3.6 and 5.1 µM, respectively). The in vitro cytotoxicity of selected diarylpentanoids (MS33A, MS33C, MS34C and MS87) was tested on Vero mammalian cells to evaluate parasite selectivity (SI), showing moderate to low cytotoxicity (CC50 > 82 µM). In addition, MS87 exhibited a high SI and the lowest resistance index (RI), suggesting that MS87 may exert effective parasite inhibition with low resistance potential in the CQ-resistant P. falciparum strain. Furthermore, the in vivo toxicity of the molecules on early embryonic development, the cardiovascular system, heart rate, motor activity and apoptosis were assessed in a zebrafish animal model. The overall results indicate the preliminary potential of diarylpentanoids, which need further investigation for their development as new antimalarial agents.

Impairments of cerebellar structure and function in a zebrafish KO of neuropsychiatric risk gene znf536.

Genetic variants in ZNF536 contribute to the risk for neuropsychiatric disorders such as schizophrenia, autism, and others. The role of this putative transcriptional repressor in brain development and function is, however, largely unknown. We generated znf536 knockout (KO) zebrafish and studied their behavior, brain anatomy, and brain function. Larval KO zebrafish showed a reduced ability to compete for food, resulting in decreased total body length and size. This phenotype can be rescued by segregating the homozygous KO larvae from their wild-type and heterozygous siblings, enabling studies of adult homozygous KO animals. In adult KO zebrafish, we observed significant reductions in anxiety-like behavior and social interaction. These znf536 KO zebrafish have decreased cerebellar volume, corresponding to decreased populations of specific neuronal cells, especially in the valvular cerebelli (Va). Finally, using a Tg[mbp:mgfp] line, we identified a previously undetected myelin structure located bilaterally within the Va, which also displayed a reduction in volume and disorganization in KO zebrafish. These findings indicate an important role for ZNF536 in brain development and implicate the cerebellum in the pathophysiology of neuropsychiatric disorders.

An indole-based near-infrared fluorescent "Turn-On" probe for H2O2: Selective detection and ultrasensitive imaging of zebrafish gallbladder.

Fluorescent probes play essential roles in medical imaging, where the researchers can select one of many molecules to use to help monitor the status of living systems under investigation. To date, a few scaffolds that allow the in vivo detection of H2O2 are available only. Herein, we provide a highly sensitive and selective near-infrared fluorescent probe that detects H2O2 based on the ICT sensing mechanism. We report the first indole-incorporated fluorescent probe Indo-H2O2 that allows H2O2 detection with a LOD of 25.2 nM featuring a boronate group conjugated to an indole scaffold; the boronate cleaves upon reaction with H2O2. A 5-membered malononitrile derivative was incorporated; Indo-H2O2 has near-infrared (NIR) properties and the reaction time is low (∼25 min) compared to other related probes. Indo-H2O2 was successfully employed in both endogenous and exogenous imaging trials of H2O2 in living cells. Indo-H2O2 also allows the real-time monitoring of H2O2in vivo. It preferentially accesses the gallbladder of zebrafish. Our findings support Indo-H2O2 as a highly sensitive fluorescent NIR probe for detecting H2O2, and an idea to incorporate a central indole unit in future fluorescent probe designs.

A Novel Anticancer Peptide Derived from Bryopsis plumosa Regulates Proliferation and Invasion in Non-Small Cell Lung Cancer Cells.

The discovery of new highly effective anticancer drugs with few side effects is a challenge for drug development research. Natural or synthetic anticancer peptides (ACPs) represent a new generation of anticancer agents with high selectivity and specificity. The rapid emergence of chemoradiation-resistant lung cancer has necessitated the discovery of novel anticancer agents as alternatives to conventional therapeutics. In this study, we synthesized a peptide containing 22 amino acids and characterized it as a novel ACP (MP06) derived from green sea algae, Bryopsis plumosa. Using the ACP database, MP06 was predicted to possess an alpha-helical secondary structure and functionality. The anti-proliferative and apoptotic effects of the MP06, determined using the cytotoxicity assay and Annexin V/propidium iodide staining kit, were significantly higher in non-small-cell lung cancer (NSCLC) cells than in non-cancerous lung cells. We confirmed that MP06 suppressed cellular migration and invasion and inhibited the expression of N-cadherin and vimentin, the markers of epithelial-mesenchymal transition. Moreover, MP06 effectively reduced the metastasis of tumor xenografts in zebrafish embryos. In conclusion, we suggest considering MP06 as a novel candidate for the development of new anticancer drugs functioning via the ERK signaling pathway.

Insights into the recognition mechanism in the UBR box of UBR4 for its specific substrates.

The N-end rule pathway is a proteolytic system involving the destabilization of N-terminal amino acids, known as N-degrons, which are recognized by N-recognins. Dysregulation of the N-end rule pathway results in the accumulation of undesired proteins, causing various diseases. The E3 ligases of the UBR subfamily recognize and degrade N-degrons through the ubiquitin-proteasome system. Herein, we investigated UBR4, which has a distinct mechanism for recognizing type-2 N-degrons. Structural analysis revealed that the UBR box of UBR4 differs from other UBR boxes in the N-degron binding sites. It recognizes type-2 N-terminal amino acids containing an aromatic ring and type-1 N-terminal arginine through two phenylalanines on its hydrophobic surface. We also characterized the binding mechanism for the second ligand residue. This is the report on the structural basis underlying the recognition of type-2 N-degrons by the UBR box with implications for understanding the N-end rule pathway.

Adverse effects of bifenthrin exposure on neurobehavior and neurodevelopment in a zebrafish embryo/larvae model.

Bifenthrin, a third-generation synthetic pyrethroid, is widely used as an agricultural insecticide. However, it can flow into surface and groundwater, leading to adverse consequences such as immunotoxicity, hepatotoxicity, hormone dysregulation, or neurotoxicity. Nevertheless, the entire range of its neurotoxic consequences, particularly in aquatic organisms, remains unclear. In this study, we conducted an extensive examination of how exposure to bifenthrin affects the behavior and nervous system function of aquatic vertebrates, using a zebrafish model and multiple-layered assays. We exposed wild-type and transgenic lines [tg(elavl3:eGFP) and tg(mbp:mGFP)] to bifenthrin from <3 h post-fertilization (hpf) to 120 hpf. Our findings indicate that bifenthrin exposure concentrations of 103.9 and 362.1 µg/L significantly affects the tail-coiling response at 24 hpf and the touch-evoked responses at 72 hpf. Moreover, it has a significant effect on various aspects of behavior such as body contact, distance between subjects, distance moved, and turn angle. We attribute these effects to changes in acetylcholinesterase and dopamine levels, which decrease in a concentration-dependent manner. Furthermore, neuroimaging revealed neurogenesis defects, e.g., shortened brain and axon widths, and demyelination of oligodendrocytes and Schwann cells. Additionally, the transcription of genes related to neurodevelopment (e.g., gap43, manf, gfap, nestin, sox2) were significantly upregulated and neurotransmitters (e.g., nlgn1, drd1, slc6a4a, ache) was significantly downregulated. In summary, our data shows that bifenthrin exposure has detrimental effects on neurodevelopmental and neurotransmission systems in the zebrafish embryo/larvae model.

Highly sensitive and rapid detection of hypochlorous acid in aqueous media and its bioimaging in live cells and zebrafish using an ESIPT-driven mycophenolic acid-based fluorescent probe.

Excessive production of potent biological oxidants such as HOCl has been implicated in numerous diseases. Thus, it is crucial to develop highly specific and precise methods to detect HOCl in living systems, preferably with molecules that can show a distinct therapeutic effect. Our study introduces the synthesis and application of a highly sensitive fluorescence "turn-on" probe, Myco-OCl, based on the mycophenolic acid scaffold with exceptional water solubility. The ESIPT-driven mechanism enables Myco-OCl to specifically and rapidly detect (<5 s) HOCl with an impressive Stokes shift of 105 nm (λex = 417 nm, λem = 522 nm) and a sub-nanomolar (97.3 nM) detection limit with the detection range of 0 to 50 µM. The potential of Myco-OCl as an excellent biosensor is evident from its successful application for live cell imaging of exogenous and endogenous HOCl. In addition, Myco-OCl enabled us to detect HOCl in a zebrafish inflammatory animal model. These underscore the great potential of Myco-OCl for detecting HOCl in diverse physiological systems. Our findings thus offer a highly promising tool for detecting HOCl in living organisms.

Statistically unbiased prediction enables accurate denoising of voltage imaging data.

Here we report SUPPORT (statistically unbiased prediction utilizing spatiotemporal information in imaging data), a self-supervised learning method for removing Poisson-Gaussian noise in voltage imaging data. SUPPORT is based on the insight that a pixel value in voltage imaging data is highly dependent on its spatiotemporal neighboring pixels, even when its temporally adjacent frames alone do not provide useful information for statistical prediction. Such dependency is captured and used by a convolutional neural network with a spatiotemporal blind spot to accurately denoise voltage imaging data in which the existence of the action potential in a time frame cannot be inferred by the information in other frames. Through simulations and experiments, we show that SUPPORT enables precise denoising of voltage imaging data and other types of microscopy image while preserving the underlying dynamics within the scene.

A cryptic microdeletion del(12)(p11.21p11.23) within an unbalanced translocation t(7;12)(q21.13;q23.1) implicates new candidate loci for intellectual disability and Kallmann syndrome.

In a patient diagnosed with both Kallmann syndrome (KS) and intellectual disability (ID), who carried an apparently balanced translocation t(7;12)(q22;q24)dn, array comparative genomic hybridization (aCGH) disclosed a cryptic heterozygous 4.7 Mb deletion del(12)(p11.21p11.23), unrelated to the translocation breakpoint. This novel discovery prompted us to consider the possibility that the combination of KS and neurological disorder in this patient could be attributed to gene(s) within this specific deletion at 12p11.21-12p11.23, rather than disrupted or dysregulated genes at the translocation breakpoints. To further support this hypothesis, we expanded our study by screening five candidate genes at both breakpoints of the chromosomal translocation in a cohort of 48 KS patients. However, no mutations were found, thus reinforcing our supposition. In order to delve deeper into the characterization of the 12p11.21-12p11.23 region, we enlisted six additional patients with small copy number variations (CNVs) and analyzed eight individuals carrying small CNVs in this region from the DECIPHER database. Our investigation utilized a combination of complementary approaches. Firstly, we conducted a comprehensive phenotypic-genotypic comparison of reported CNV cases. Additionally, we reviewed knockout animal models that exhibit phenotypic similarities to human conditions. Moreover, we analyzed reported variants in candidate genes and explored their association with corresponding phenotypes. Lastly, we examined the interacting genes associated with these phenotypes to gain further insights. As a result, we identified a dozen candidate genes: TSPAN11 as a potential KS candidate gene, TM7SF3, STK38L, ARNTL2, ERGIC2, TMTC1, DENND5B, and ETFBKMT as candidate genes for the neurodevelopmental disorder, and INTS13, REP15, PPFIBP1, and FAR2 as candidate genes for KS with ID. Notably, the high-level expression pattern of these genes in relevant human tissues further supported their candidacy. Based on our findings, we propose that dosage alterations of these candidate genes may contribute to sexual and/or cognitive impairments observed in patients with KS and/or ID. However, the confirmation of their causal roles necessitates further identification of point mutations in these candidate genes through next-generation sequencing.

Interpretation of the effects of anthropogenic chlorine on nitrate formation over northeast Asia during KORUS-AQ 2016.

The Weather Research and Forecasting-Community Multiscale Air Quality (WRF-CMAQ) model, implemented with anthropogenic chlorine (Cl) emissions, was evaluated against ground and NASA DC-8 aircraft measurements during the Korea-United States Air Quality (KORUS-AQ) 2016 campaign. The latest anthropogenic Cl emissions, including gaseous HCl and particulate chloride (pCl-) emissions from the Anthropogenic Chlorine Emissions Inventory of China (ACEIC-2014) (over China) and a global emissions inventory (Zhang et al., 2022) (over outer China), were used to examine the impacts of Cl emissions and the role of nitryl chloride (ClNO2) chemistry in N2O5 heterogeneous reactions on secondary nitrate (NO3-) formation across the Korean Peninsula. The model results against aircraft measurements clearly showed significant Cl- underestimations due mainly to the high gas-particle (G/P) partitioning ratios at aircraft measurement altitudes such as 700-850 hPa, but the ClNO2 simulations were reasonable. Several simulations of CMAQ-based sensitivity experiments against ground measurements indicated that although addition of Cl emission did not significantly alter NO3- formation, the activated ClNO2 chemistry with Cl emissions showed the best model performance with the reduced normalized mean bias (NMB) of 18.7 % compared to a value of 21.1 % for the Cl emissions-free case. In our model evaluation, ClNO2 accumulated during the night but quickly produced Cl radical due to ClNO2 photolysis at sunrise, which modulated other oxidation radicals (e.g., ozone [O3] and hydrogen oxide radicals [HOx]) in the early morning. In the morning hours (0800-1000 LST), the HOx were the dominant oxidants, contributing 86.6 % of the total oxidation capacity (sum of major oxidants such as O3 and HOx species), while oxidability was enhanced by up to ∼6.4 % (increase in 1 h HOx average of 2.89 × 106 molecules·cm-3) in the early morning mainly due to the changes in OH (+7.2 %), hydroperoxyl radical (HO2)(+10.0 %), and O3 (+4.2 %) over the Seoul Metropolitan Area, during the KORUS-AQ campaign. Our results improve understanding of the atmospheric changes in the PM2.5 formation pathway caused by ClNO2 chemistry and Cl emissions over northeast Asia.

IFT46 gene promoter-driven ciliopathy disease model in zebrafish.

Ciliopathies are human genetic disorders caused by abnormal formation and dysfunction of cellular cilia. Cilia are microtubule-based organelles that project into the extracellular space and transduce molecular and chemical signals from the extracellular environment or neighboring cells. Intraflagellar transport (IFT) proteins are required for the assembly and maintenance of cilia by transporting proteins along the axoneme which consists of complexes A and B. IFT46, a core IFT-B protein complex, is required for cilium formation and maintenance during vertebrate embryonic development. Here, we introduce transgenic zebrafish lines under the control of ciliated cell-specific IFT46 promoter to recapitulate human ciliopathy-like phenotypes. We generated a Tg(IFT46:GAL4-VP16) line to temporo-spatially control the expression of effectors including fluorescent reporters or nitroreductase based on the GAL4/UAS system, which expresses GAL4-VP16 chimeric transcription factors in most ciliated tissues during embryonic development. To analyze the function of IFT46-expressing ciliated cells during zebrafish development, we generated the Tg(IFT46:GAL4-VP16;UAS;nfsb-mCherry) line, a ciliated cell-specific injury model induced by nitroreductase (NTR)/metrodinazole (MTZ). Conditionally, controlled ablation of ciliated cells in transgenic animals exhibited ciliopathy-like phenotypes including cystic kidneys and pericardial and periorbital edema. Altogether, we established a zebrafish NTR/MTZ-mediated ciliated cell injury model that recapitulates ciliopathy-like phenotypes and may be a vertebrate animal model to further investigate the etiology and therapeutic approaches to human ciliopathies.

Zebrafish: A Powerful Model for Genetics and Genomics.

Our understanding of fundamental biological mechanisms and the pathogenesis of human diseases has been greatly improved by studying the genetics and genomics of zebrafish [...].

"Lighting up" fluoride: cellular imaging and zebrafish model interrogations using a simple ESIPT-based mycophenolic acid precursor-based probe.

The discovery and implementation of media that derive from bioinspired designs and bear optical readouts featuring large Stokes shifts are of continued interest to a wide variety of researchers and clinicians. Myco-F, a novel mycophenolic acid precursor-based probe features a cleavable tert-butyldimethylsiloxy group to allow for fluoride detection. Myco-F exhibits high selectivity and specificity towards F- (Stokes shift = 120 nm). All measurements were performed in complete aqueous media (LOD=0.38 µM). Myco-F enables detection of fluoride ions in living HEK293 cells and localizes in the eye region (among other regions) of the zebrafish. DFT calculations support the proposed ESIPT working photomechanism.

In vivo whole-brain imaging of zebrafish larvae using three-dimensional fluorescence microscopy.

As a vertebrate model animal, larval zebrafish are widely used in neuroscience and provide a unique opportunity to monitor whole-brain activity at the cellular resolution. Here, we provide an optimized protocol for performing whole-brain imaging of larval zebrafish using three-dimensional fluorescence microscopy, including sample preparation and immobilization, sample embedding, image acquisition, and visualization after imaging. The current protocol enables in vivo imaging of the structure and neuronal activity of a larval zebrafish brain at a cellular resolution for over 1 h using confocal microscopy and custom-designed fluorescence microscopy. The critical steps in the protocol are also discussed, including sample mounting and positioning, preventing bubble formation and dust in the agarose gel, and avoiding motion in images caused by incomplete solidification of the agarose gel and paralyzation of the fish. The protocol has been validated and confirmed in multiple settings. This protocol can be easily adapted for imaging other organs of a larval zebrafish.

Structure-Function of the Human WAC Protein in GABAergic Neurons: Towards an Understanding of Autosomal Dominant DeSanto-Shinawi Syndrome.

Dysfunction of the WW domain-containing adaptor with coiled-coil, WAC, gene underlies a rare autosomal dominant disorder, DeSanto-Shinawi syndrome (DESSH). DESSH is associated with facial dysmorphia, hypotonia, and cognitive alterations, including attention deficit hyperactivity disorder and autism. How the WAC protein localizes and functions in neural cells is critical to understanding its role during development. To understand the genotype-phenotype role of WAC, we developed a knowledgebase of WAC expression, evolution, human genomics, and structural/motif analysis combined with human protein domain deletions to assess how conserved domains guide cellular distribution. Then, we assessed localization in a cell type implicated in DESSH, cortical GABAergic neurons. WAC contains conserved charged amino acids, phosphorylation signals, and enriched nuclear motifs, suggesting a role in cellular signaling and gene transcription. Human DESSH variants are found within these regions. We also discovered and tested a nuclear localization domain that impacts the cellular distribution of the protein. These data provide new insights into the potential roles of this critical developmental gene, establishing a platform to assess further translational studies, including the screening of missense genetic variants in WAC. Moreover, these studies are essential for understanding the role of human WAC variants in more diverse neurological phenotypes, including autism spectrum disorder.

Embryonic mercury exposure in zebrafish: Alteration of metabolites and gene expression, related to visual and behavioral impairments.

The widespread presence of mercury, a heavy metal found in the environment and used in numerous industries and domestic, raises concerns about its potential impact on human health. Nevertheless, the adverse effects of this environmental toxicant at low concentrations are often underestimated. There are emerging studies showing that accumulation of mercury in the eye may contribute to visual impairment and a comorbidity between autism spectrum disorders (ASD) trait and visual impairment. However, the underlying mechanism of visual impairment in humans and rodents is challenging. In response to this issue, zebrafish larvae with a cone-dominated retinal visual system were exposed to 100 nM mercury chloride (HgCl2), according to our previous study, followed by light-dark stimulation, a social assay, and color preference to examine the functionality of the visual system in relation to ASD-like behavior. Exposure of embryos to HgCl2 from gastrulation to hatching increased locomotor activity in the dark, reduced shoaling and exploratory behavior, and impaired color preference. Defects in microridges as the first barrier may serve as primary tools for HgCl2 toxicity affecting vision. Depletion of polyunsaturated fatty acids (PUFAs), linoleic acid, arachidonic acid (ARA), alpha-linoleic acid, docosahexaenoic acid (DHA), stearic acid, L-phenylalanine, isoleucine, L-lysine, and N-acetylputrescine, along with the increase of gamma-aminobutyric acid (GABA), sphingosine-1-phosphate, and citrulline assayed by liquid chromatography-mass spectrometry (LC-MS) suggest that these metabolites serve as biomarkers of retinal impairments that affect vision and behavior. Although suppression of adsl, shank3a, tsc1b, and nrxn1a gene expression was observed, among these tsc1b showed more positive correlation with ASD. Collectively, these results contribute new insights into the possible mechanism of mercury toxicity give rise to visual, cognitive, and social deficits in zebrafish.

A microdeletion del(12)(p11.21p11.23) with a cryptic unbalanced translocation t(7;12)(q21.13;q23.1) implicates new candidate loci for intellectual disability and Kallmann syndrome.

In an apparently balanced translocation t(7;12)(q22;q24)dn exhibiting both Kallmann syndrome (KS) and intellectual disability (ID), we detected a cryptic heterozygous 4.7 Mb del(12)(p11.21p11.23) unrelated to the translocation breakpoint. This new finding raised the possibility that KS combined with neurological disorder in this patient could be caused by gene(s) within this deletion at 12p11.21-12p11.23 instead of disrupted or dysregulated genes at the genomic breakpoints. Screening of five candidate genes at both breakpoints in 48 KS patients we recruited found no mutation, corroborating our supposition. To substantiate this hypothesis further, we recruited six additional subjects with small CNVs and analyzed eight individuals carrying small CNVs in this region from DECIPHER to dissect 12p11.21-12p11.23. We used multiple complementary approaches including a phenotypic-genotypic comparison of reported cases, a review of knockout animal models recapitulating the human phenotypes, and analyses of reported variants in the interacting genes with corresponding phenotypes. The results identified one potential KS candidate gene ( TSPAN11 ), seven candidate genes for the neurodevelopmental disorder ( TM7SF3 , STK38L , ARNTL2 , ERGIC2 , TMTC1 , DENND5B , and ETFBKMT ), and four candidate genes for KS with ID ( INTS13 , REP15 , PPFIBP1 , and FAR2 ). The high-level expression pattern in the relevant human tissues further suggested the candidacy of these genes. We propose that the dosage alterations of the candidate genes may contribute to sexual and/or cognitive impairment in patients with KS and/or ID. Further identification of point mutations through next generation sequencing will be necessary to confirm their causal roles.

Eye movement defects in KO zebrafish reveals SRPK3 as a causative gene for an X-linked intellectual disability.

Intellectual disability (ID) is a common neurodevelopmental disorder characterized by significantly impaired intellectual and adaptive functioning. X-linked ID (XLID) disorders, caused by defects in genes on the X chromosome, affect 1.7 out of 1,000 males. Employing exome sequencing, we identified three missense mutations (c.475C>G; p.H159D, c.1373C>A; p.T458N, and c.1585G>A; p.E529K) in the SRPK3 gene in seven XLID patients from three independent families. Clinical features common to the patients are intellectual disability, agenesis of the corpus callosum, abnormal smooth pursuit eye movement, and ataxia. SRPK proteins are known to be involved in mRNA processing and, recently, synaptic vesicle and neurotransmitter release. In order to validate SRPK3 as a novel XLID gene, we established a knockout (KO) model of the SRPK3 orthologue in zebrafish. In day 5 of larval stage, KO zebrafish showed significant defects in spontaneous eye movement and swim bladder inflation. In adult KO zebrafish, we found agenesis of cerebellar structures and impairments in social interaction. These results suggest an important role of SRPK3 in eye movements, which might reflect learning problems, intellectual disability, and other psychiatric disorders.

Survival Factor A (SvfA) Contributes to Aspergillus nidulans Pathogenicity.

Survival factor A (SvfA) in Aspergillus nidulans plays multiple roles in growth and developmental processes. It is a candidate for a novel VeA-dependent protein involved in sexual development. VeA is a key developmental regulator in Aspergillus species that can interact with other velvet-family proteins and enter into the nucleus to function as a transcription factor. In yeast and fungi, SvfA-homologous proteins are required for survival under oxidative and cold-stress conditions. To assess the role of SvfA in virulence in A. nidulans, cell wall components, biofilm formation, and protease activity were evaluated in a svfA-gene-deletion or an AfsvfA-overexpressing strain. The svfA-deletion strain showed decreased production of ß-1,3-glucan in conidia, a cell wall pathogen-associated molecular pattern, with a decrease in gene expression for chitin synthases and ß-1,3-glucan synthase. The ability to form biofilms and produce proteases was reduced in the svfA-deletion strain. We hypothesized that the svfA-deletion strain was less virulent than the wild-type strain; therefore, we performed in vitro phagocytosis assays using alveolar macrophages and analyzed in vivo survival using two vertebrate animal models. While phagocytosis was reduced in mouse alveolar macrophages challenged with conidia from the svfA-deletion strain, the killing rate showed a significant increase with increased extracellular signal-regulated kinase ERK activation. The svfA-deletion conidia infection reduced host mortality in both T-cell-deficient zebrafish and chronic granulomatous disease mouse models. Taken together, these results indicate that SvfA plays a significant role in the pathogenicity of A. nidulans.