Hwanghudson4803

mics and maintenance of the naive humoral immune response to SARS-CoV-2 is of great importance. In addition, long-term responses after asymptomatic infection are not well-characterized, given the challenges in identifying such cases. Here, we investigated the longitudinal humoral profile in a well-characterized cohort of young adults with documented asymptomatic or mildly symptomatic SARS-CoV-2 infection. By analyzing samples collected preinfection, early after infection and during late convalescence, we found that, while neutralizing activity decreased over time, high levels of serum S2 IgG and Antibody-Dependent Monocyte Phagocytosis (ADMP) activity were maintained up to 8.5 months after infection. This suggests that a subset of antibodies with specific functions could contribute to long-term protection against SARS-CoV-2 in convalescent unvaccinated individuals.Within epithelial cells, Pseudomonas aeruginosa depends on its type III secretion system (T3SS) to escape vacuoles and replicate rapidly in the cytosol. Previously, it was assumed that intracellular subpopulations remaining T3SS-negative (and therefore in vacuoles) were destined for degradation in lysosomes, supported by data showing vacuole acidification. Here, we report in both corneal and bronchial human epithelial cells that vacuole-associated bacteria can persist, sometimes in the same cells as cytosolic bacteria. Using a combination of phase-contrast, confocal, and correlative light-electron microscopy (CLEM), we also found they can demonstrate biofilm-associated markers cdrA and cyclic-di-GMP (c-di-GMP). Vacuolar-associated bacteria, but not their cytosolic counterparts, tolerated the cell-permeable antibiotic ofloxacin. Surprisingly, use of mutants showed that both persistence in vacuoles and ofloxacin tolerance were independent of the biofilm-associated protein CdrA or exopolysaccharides (Psl, Pel, apulations with T3SS-negative members enabling persistence while rapid replication is accomplished by more vulnerable T3SS-positive siblings. Intracellular P. aeruginosa persisting and tolerating antibiotics independently of the T3SS or biofilm-associated factors could present additional challenges to development of more effective therapeutics.Hydrocephalus, the leading indication for childhood neurosurgery worldwide, is particularly prevalent in low- and middle-income countries. Hydrocephalus preceded by an infection, or postinfectious hydrocephalus, accounts for up to 60% of hydrocephalus in these areas. Since many children with hydrocephalus suffer poor long-term outcomes despite surgical intervention, prevention of hydrocephalus remains paramount. Our previous studies implicated a novel bacterial pathogen, Paenibacillus thiaminolyticus, as a causal agent of neonatal sepsis and postinfectious hydrocephalus in Uganda. Here, we report the isolation of three P. thiaminolyticus strains, Mbale, Mbale2, and Mbale3, from patients with postinfectious hydrocephalus. We constructed complete genome assemblies of the clinical isolates as well as the nonpathogenic P. thiaminolyticus reference strain and performed comparative genomic and proteomic analyses to identify potential virulence factors. All three isolates carry a unique beta-lactamase gene, and two rt. Whole-genome sequencing, RNA sequencing, and proteomics of clinical isolates and a reference strain in combination with CRISPR editing identified type IV pili as a critical virulence factor for P. thiaminolyticus infection. Acquisition of a type IV pilus-encoding mobile genetic element critically contributed to converting a nonpathogenic strain of P. thiaminolyticus into a pathogen capable of causing devastating diseases. Given the widespread presence of type IV pilus in pathogens, the presence of the type IV pilus operon could serve as a diagnostic and therapeutic target in P. thiaminolyticus and related bacteria.Genome editing technology is a powerful tool for programming microbial cell factories. However, rat APOBEC1-derived cytosine base editor (CBE) that converts C•G to T•A at target genes induced DNA off-targets, regardless of single-guide RNA (sgRNA) sequences. Although the high efficiencies of the bacterial CBEs have been developed, a risk of unidentified off-targets impeded genome editing for microbial cell factories. To address the issues, we demonstrate the genome engineering of Corynebacterium glutamicum as a GC-rich model industrial bacterium by generating premature termination codons (PTCs) in desired genes using high-fidelity cytosine base editors (CBEs). Through this CBE-STOP approach of introducing specific cytosine conversions, we constructed several single-gene-inactivated strains for three genes (ldh, idsA, and pyc) with high base editing efficiencies of average 95.6% (n = 45, C6 position) and the highest success rate of up to 100% for PTCs and ultimately developed a strain with five genes (ldh, actcrobial cell factories. To address the issues, we identified the DNA off-targets for single and multiple genome engineering of the industrial bacterium Corynebacterium glutamicum using whole-genome sequencing. Further, we developed the high-fidelity (HF)-CBE with significantly reduced off-targets with comparable efficiency and precision. We believe that our DNA off-target analysis and the HF-CBE can promote CRISPR-assisted genome engineering over conventional gene manipulation tools by providing a markerless genetic tool without need for a foreign DNA donor.Nicotinamide adenine dinucleotide (NAD) is a key metabolite synthesised from vitamin B3 or tryptophan. Disruption of genes encoding NAD synthesis enzymes reduces NAD levels and causes congenital NAD deficiency disorder (CNDD), characterised by multiple congenital malformations. SLC6A19 (encoding B0AT1, a neutral amino acid transporter), represents the main transporter for free tryptophan in the intestine and kidney. Here, we tested whether Slc6a19 heterozygosity in mice limits the tryptophan available for NAD synthesis during pregnancy and causes adverse pregnancy outcomes. Pregnant Slc6a19+/- mice were fed diets depleted of vitamin B3, so that tryptophan was the source of NAD during gestation. This perturbed the NAD metabolome in pregnant Slc6a19+/- females, resulting in reduced NAD levels and increased rates of embryo loss. Surviving embryos were small and exhibited specific combinations of CNDD-associated malformations. Our results show that genes not directly involved in NAD synthesis can affect NAD metabolism and cause CNDD. They also suggest that human female carriers of a SLC6A19 loss-of-function allele might be susceptible to adverse pregnancy outcomes unless sufficient NAD precursor amounts are available during gestation. This article has an associated First Person interview with the first author of the paper.First Person is a series of interviews with the first authors of a selection of papers published in Disease Models & Mechanisms, helping researchers promote themselves alongside their papers. Hartmut Cuny is first author on ' Maternal heterozygosity of Slc6a19 causes metabolic perturbation and congenital NAD deficiency disorder in mice', published in DMM. Hartmut is a senior postdoc in the lab of Prof. Sally Dunwoodie at Victor Chang Cardiac Research Institute, Sydney, Australia, investigating the genetic and environmental causes of congenital malformations.The incidence of tickborne infections in the United States has risen significantly. Automation is needed for the increasing demand for testing. The Panther Fusion (Fusion) has an Open Access functionality to perform lab developed tests (LDTs) on a fully automated system. Our laboratory adapted two LDTs on Fusion; a multiplex real-time PCR for Anaplasma phagocytophilum and Ehrlichia chaffeensis (AP/EC) and a Babesia microti (BM) PCR. Limits of detection (LODs) were performed with target region plasmid panels spiked into whole blood. The LODs for AP, BM, and EC on the Fusion were 11, 17, and 10 copies/reaction, respectively. The performance of AP/EC was evaluated with 80 whole blood specimens, including 50 specimens previously positive for AP by our test of record (TOR) and 30 specimens (including 20 AP positive) spiked with EC plasmid. AP was detected in 49 out of 50 positive specimens and EC was detected in all 30 spiked specimens. BM PCR on Fusion was evaluated with 75 whole blood samples, including 16 speci specimens previously tested for Anaplasma phagocytophilum, Ehrlichia chaffeensis, and Babesia microti, while WB spiked with DNA from plasmid clones of the target regions were used for analytical sensitivity. We demonstrated that the Panther Fusion assays performed similar to the manual PCR tests used clinically in our laboratory and that automation of these tests had no adverse effect on the performance.The complete remineralization of organic matter in anoxic environments relies on communities of microorganisms that ferment organic acids and alcohols to CH4. This is accomplished through syntrophic association of H2 or formate producing bacteria and methanogenic archaea, where exchange of these intermediates enables growth of both organisms. While these communities are essential to Earth's carbon cycle, our understanding of the dynamics of H2 or formate exchanged is limited. Here, we establish a model partnership between Syntrophotalea carbinolica and Methanococcus maripaludis. Through sequencing a transposon mutant library of M. maripaludis grown with ethanol oxidizing S. PD0325901 supplier carbinolica, we found that genes encoding the F420-dependent formate dehydrogenase (Fdh) and F420-dependent methylene-tetrahydromethanopterin dehydrogenase (Mtd) are important for growth. Competitive growth of M. maripaludis mutants defective in either H2 or formate metabolism verified that, across multiple substrates, interspecies formate mutant strains of M. maripaludis that are defective for either H2 or formate metabolism, we determined that interspecies formate exchange drives syntrophic growth of these organisms. Together, these results advance our understanding of the degradation of organic matter in anoxic environments.Pbam-32 is a new carbon allotrope formed by hybrid sp3 atoms. It is promising for application as a super-hard material owing to its remarkable Young's modulus and Vicker's hardness. However, thermodynamic properties related to the stability, production and practical performance of Pbam-32 are still unclear, so further study is required to clarify these details. With the help of ab initio simulations, we systematically explore the structure, atomic environment, phonon dispersion, entropy and enthalpy changes of Pbam-32 compared with diamond and graphite. As an allotrope consisting of five-membered and seven-membered rings, Pbam-32 is a meta-stable structure without imaginary frequencies in phonon dispersion. An acoustic band gap between 39.58 THz and 42.57 THz indicates potential applications of Pbam-32 as a terahertz phononic crystal. The entropy and enthalpy of Pbam-32 are similar to those of diamond. Besides, Pbam-32 exhibits an excellent ability to resist pressure. Unfortunately, the free energy of Pbam-32 is higher than that of diamond over a wide range of pressure and temperature conditions, which means that Pbam-32 is absent in the phase diagram of the three allotropes. Therefore, the production of Pbam-32 must be kinetically controlled. Our results are helpful for understanding the thermodynamic properties of Pbam-32.