DNA mismatch and damage patterns revealed by single-molecule sequencing.

Mutations accumulate in the genome of every cell of the body throughout life, causing cancer and other diseases1,2. Most mutations begin as nucleotide mismatches or damage in one of the two strands of the DNA before becoming double-strand mutations if unrepaired or misrepaired3,4. However, current DNA-sequencing technologies cannot accurately resolve these initial single-strand events. Here we develop a single-molecule, long-read sequencing method (Hairpin Duplex Enhanced Fidelity sequencing (HiDEF-seq)) that achieves single-molecule fidelity for base substitutions when present in either one or both DNA strands. HiDEF-seq also detects cytosine deamination-a common type of DNA damage-with single-molecule fidelity. We profiled 134 samples from diverse tissues, including from individuals with cancer predisposition syndromes, and derive from them single-strand mismatch and damage signatures. We find correspondences between these single-strand signatures and known double-strand mutational signatures, which resolves the identity of the initiating lesions. Tumours deficient in both mismatch repair and replicative polymerase proofreading show distinct single-strand mismatch patterns compared to samples that are deficient in only polymerase proofreading. We also define a single-strand damage signature for APOBEC3A. In the mitochondrial genome, our findings support a mutagenic mechanism occurring primarily during replication. As double-strand DNA mutations are only the end point of the mutation process, our approach to detect the initiating single-strand events at single-molecule resolution will enable studies of how mutations arise in a variety of contexts, especially in cancer and ageing.

Cortex-dependent corrections as the tongue reaches for and misses targets.

Precise tongue control is necessary for drinking, eating and vocalizing1-3. However, because tongue movements are fast and difficult to resolve, neural control of lingual kinematics remains poorly understood. Here we combine kilohertz-frame-rate imaging and a deep-learning-based neural network to resolve 3D tongue kinematics in mice drinking from a water spout. Successful licks required corrective submovements that-similar to online corrections during primate reaches4-11-occurred after the tongue missed unseen, distant or displaced targets. Photoinhibition of anterolateral motor cortex impaired corrections, which resulted in hypometric licks that missed the spout. Neural activity in anterolateral motor cortex reflected upcoming, ongoing and past corrective submovements, as well as errors in predicted spout contact. Although less than a tenth of a second in duration, a single mouse lick exhibits the hallmarks of online motor control associated with a primate reach, including cortex-dependent corrections after misses.

Envafolimab combined with chemotherapy in the treatment of combined small cell lung cancer: A case report.

BACKGROUND: Combined small cell lung cancer (C-SCLC) is a special subtype of small cell lung cancer that is relatively rare, aggressive, and prone to early metastasis and has a poor prognosis. Currently, there are limited studies on C-SCLC, and there is no uniform standard treatment, especially for extensive C-SCLC, which still faces great challenges. In recent years, the development and progress of immunotherapy have provided more possibilities for the treatment of C-SCLC. We used immunotherapy combined with first-line chemotherapy to treat extensive-stage C-SCLC to explore its antitumor activity and safety. CASE SUMMARY: We report a case of C-SCLC that presented early with adrenal, rib, and mediastinal lymph node metastases. The patient received carboplatin and etoposide with concurrent initiation of envafolimab. After 6 cycles of chemotherapy, the lung lesion was significantly reduced, and the comprehensive efficacy evaluation showed a partial response. No serious drug-related adverse events occurred during the treatment, and the drug regimen was well tolerated. CONCLUSION: Envafolimab combined with carboplatin and etoposide in the treatment of extensive-stage C-SCLC has preliminary antitumor activity and good safety and tolerability.

Critical respiratory failure due to pregnancy complicated by COVID-19 and bacterial coinfection: A case report.

BACKGROUND: In the past 3 years, the global pandemic of coronavirus disease 2019 (COVID-19) has posed a great threat to human life and safety. Among the causes of death in COVID-19 patients, combined or secondary bacterial infection is an important factor. As a special group, pregnant women experience varying degrees of change in their immune status, cardiopulmonary function, and anatomical structure during pregnancy, which puts them at higher risk of contracting COVID-19. COVID-19 infection during pregnancy is associated with increased adverse events such as hospitalisation, admission to the intensive care unit, and mechanical ventilation. Therefore, pregnancy combined with coinfection of COVID-19 and bacteria often leads to critical respiratory failure, posing severe challenges in the diagnosis and treatment process. CASE SUMMARY: We report a case of COVID-19 complicated with Staphylococcus aureus (S. aureus) coinfection in a pregnant woman at 34 wk of gestation. Her rapid progression of pulmonary lesions caused severe respiratory failure, and she received non-invasive ventilator-assisted respiratory treatment. Subsequently, we delivered a foetus via emergency caesarean section after accelerating the maturity of the foetal pulmonary system, and the respiratory condition of the puerperant woman significantly improved after the delivery of the foetus. Lavage fluid was taken under tracheoscopy to quickly search for pathogens by the metagenomic next-generation sequencing (mNGS), and both COVID-19 and S. aureus were detected. After targeted anti-infective treatment, the maternal condition gradually improved, and the patient was discharged from the hospital. CONCLUSION: The coinfection of pregnancy with COVID-19 and bacteria often leads to critical respiratory failure, which is a great challenge in the process of diagnosis and treatment. It is crucial to choose the right time to deliver the foetus and to quickly find pathogens by mNGS.

Single-strand mismatch and damage patterns revealed by single-molecule DNA sequencing.

Mutations accumulate in the genome of every cell of the body throughout life, causing cancer and other genetic diseases1-4. Almost all of these mosaic mutations begin as nucleotide mismatches or damage in only one of the two strands of the DNA prior to becoming double-strand mutations if unrepaired or misrepaired5. However, current DNA sequencing technologies cannot resolve these initial single-strand events. Here, we developed a single-molecule, long-read sequencing method that achieves single-molecule fidelity for single-base substitutions when present in either one or both strands of the DNA. It also detects single-strand cytosine deamination events, a common type of DNA damage. We profiled 110 samples from diverse tissues, including from individuals with cancer-predisposition syndromes, and define the first single-strand mismatch and damage signatures. We find correspondences between these single-strand signatures and known double-strand mutational signatures, which resolves the identity of the initiating lesions. Tumors deficient in both mismatch repair and replicative polymerase proofreading show distinct single-strand mismatch patterns compared to samples deficient in only polymerase proofreading. In the mitochondrial genome, our findings support a mutagenic mechanism occurring primarily during replication. Since the double-strand DNA mutations interrogated by prior studies are only the endpoint of the mutation process, our approach to detect the initiating single-strand events at single-molecule resolution will enable new studies of how mutations arise in a variety of contexts, especially in cancer and aging.

Single duplex DNA sequencing with CODEC detects mutations with high sensitivity.

Detecting mutations from single DNA molecules is crucial in many fields but challenging. Next-generation sequencing (NGS) affords tremendous throughput but cannot directly sequence double-stranded DNA molecules ('single duplexes') to discern the true mutations on both strands. Here we present Concatenating Original Duplex for Error Correction (CODEC), which confers single duplex resolution to NGS. CODEC affords 1,000-fold higher accuracy than NGS, using up to 100-fold fewer reads than duplex sequencing. CODEC revealed mutation frequencies of 2.72 × 10-8 in sperm of a 39-year-old individual, and somatic mutations acquired with age in blood cells. CODEC detected genome-wide, clonal hematopoiesis mutations from single DNA molecules, single mutated duplexes from tumor genomes and liquid biopsies, microsatellite instability with 10-fold greater sensitivity and mutational signatures, and specific tumor mutations with up to 100-fold fewer reads. CODEC enables more precise genetic testing and reveals biologically significant mutations, which are commonly obscured by NGS errors.

Vedolizumab-associated diffuse interstitial lung disease in patients with ulcerative colitis: A case report.

BACKGROUND: Vedolizumab, a newer class of integrin antagonist biological agents, has been applied to treat patients with moderate-to-severe Crohn's disease (CD) and ulcerative colitis (UC), especially for patients who are refractory to traditional therapies and tumor necrosis factor antagonists. However, some rare but life-threatening adverse effects warrant pharmacovigilance. We describe the first fatal case of vedolizumab-associated severe diffuse interstitial lung disease in China. CASE SUMMARY: We present a case of new-onset diffuse parenchymal lung disease developing under treatment with vedolizumab in a patient with UC. After two doses of vedolizumab, he developed persistent fever and progressively worsening dyspnea. Extensive workups, including bronchoalveolar lavage, transbronchial lung biopsy and metagenomic next-generation sequencing, identified no infectious causes, and other potential causes (such as tumors and cardiogenic pulmonary edema) were also excluded. As a result, a diagnosis of vedolizumab-related interstitial lung disease was established. Unfortunately, although corticosteroids and empiric antibiotics were administered, the patient eventually died of respiratory failure. CONCLUSION: Vedolizumab-related interstitial lung disease in patients with UC is rare but potentially lethal. Gastroenterologists and pulmonologists should be aware of vedolizumab-related adverse drug reactions.

Judd-Ofelt spectroscopic properties of Er3+-doped NaLa(WO4)2 polycrystalline powder.

Er3+-doped NaLa(WO4)2 is a promising phosphor material for applications in many fields including the ratiometric thermometry based on thermal effect of fluorescence intensity ratio (FIR) of green fluorescence of Er3+, which is directly correlated with Judd-Ofelt parameters Ωi (i = 2, 4, 6). Present paper reports synthesis and Judd-Ofelt spectroscopic properties of Er3+-doped NaLa(WO4)2 µm-sized phosphor. The phosphor was synthesized by solid-state chemical reaction and characterized by X-ray diffraction and Raman scattering techniques. The results show that the phosphor is dominated by NaLa(WO4)2 crystalline phase and has a maximum phonon energy 927 cm-1. Judd-Ofelt analysis was done for the phosphor using a safe, reliable method based on diffuse reflection spectrum and Er3+ 1.5 µm fluorescence lifetime. First, diffuse reflection spectrum of the phosphor was measured and relative absorption spectrum was calibrated from it using Kubelka-Munk theory. Second, Er3+ 1.5 µm fluorescence lifetime of the phosphor was measured and absorption cross-section spectrum was obtained based on the assumption that the 1.5 µm emission has 100% quantum efficiency. Finally, based on the absorption cross-section spectrum, standard Judd-Ofelt analysis was carried out to extract the Ωi. Radiative rate, fluorescent branch ratio and radiative lifetime of some transitions have been obtained from the known Ωi values. In addition, FIR proportional factor was evaluated in terms of Ωi and compared with those values of other materials. The result shows that the phosphor has a better prospect for the application in ratiometric thermometry.

Error evaluation of Judd-Ofelt spectroscopic analysis.

Judd-Ofelt (J-O) spectroscopic knowledge of a rare-earth-doped luminescent material is crucial to its application. Although a large number of papers with regard to the J-O study of various rare-earth-doped luminescent materials have been reported each year, few papers presented the errors of the J-O intensity parameters Ωi (i = 2, 4, 6) and radiative probabilities evaluated from them. Present study focuses on the error evaluation of the J-O parameters and radiative probabilities. An error theory is established for the J-O analysis and radiative probability. Two error analysis methods based on root mean square of the difference either between measured and calculated oscillator strengths (δfrms) or between measured and calculated line strengths (δSrms) are studied. Explicit error expressions are presented for the J-O parameters and radiative probability. The validity of the theory is verified by applying it to widely studied Er3+, Tm3+, Ho3+ and Nd3+ ions that are doped into four single-crystals (LiNbO3, SrGdGa3O7, LiYF4 and YVO4) and a glass. The two methods are identical in nature and give similar results of errors of Ωi.

Effects of heat-moisture, autoclaving, and microwave treatments on physicochemical properties of proso millet starch.

Proso millet starch was modified by heat-moisture treatment (HMT), autoclaving treatment (AT), and microwave treatment (MT). The effects of these treatments on the starch physicochemical, structural, and molecular properties were investigated. The amylose and resistant starch contents were increased by AT and MT, but only slightly by HMT. HMT and AT significantly increased the water-holding capacity, to 172.66% and 191.63%, respectively. X-ray diffractometry showed that the relative crystallinity of the HMT sample decreased by 20.88%, and the crystalline peaks disappeared from the AT and MT sample patterns. The thermal treatments decreased the proso millet starch molecular weight to 1.769 × 106, 7.886 × 105, and 3.411 × 104 g/mol, respectively. The thermal enthalpy decreased significantly in HMT. Modification significantly changed the pasting profiles of the native proso millet starch, and the peak viscosity, setback, and breakdown values decreased. These results clarify the mechanism of starch changes caused by thermal treatment.

Sextuple ratiometric thermometry based on 980-nm-upconverted green fluorescence of Er3+ ions in submicron crystals.

980-nm-upconverted 530 and 550 nm Er3+ green fluorescence spectra of Er3+/Yb3+-codoped NaGd(WO4)2 submicron crystals were measured in the temperature range of 298-383 K. A sextuple ratiometric thermometry is proposed. It is established on the basis of six schemes of fluorescence intensity ratio (FIR) that considers three component peaks of the 530 nm emission band and two component peaks of the 550 nm emission band, which involve electronic transitions between two Stark sublevels of Er3+. The study shows that the phosphor shows strong green fluorescence, which is verified by measured quantum yield, and thermally stable spectral structure desired for the sextuple ratiometric thermometry. All of the six FIR schemes display highly efficient sensing performances with slightly different thermal sensitivities. Each scheme gives a temperature value and the six schemes give an averaged result. In parallel, we have also carried out an ex vivo experimental study on the temperature characteristics of the green fluorescence of the phosphor. Almost same results have been obtained, verifying biological applicability of the phosphor. The ex vivo experimental results also show that the sextuple thermometry increases considerably the accuracy and reliability of temperature measurement in comparison with the conventional intensity integration method.