Biology subjects

Kraft, L.

Publications and source records attributed to Kraft, L..

7 recordsLinked to original sources

bioRxiv2026

Ancient Reconstructed Proteins: A Framework for Resurrecting Protein Structures from Million-Year-Old Metagenomes

DNA sequences derived from ancient samples provide insights into human history, paleoenvironments, and evolutionary biology. Advances in laboratory techniques and computational tools have established ancient DNA research as a distinct field. However, current analyses focus mainly on the DNA level, while the protein space remains under-explored. Recent progress in the de novo assembly of ancient metagenomes and the availability of protein structure prediction tools, such as AlphaFold 2, enable the reconstruction of protein structures from these degraded sequences. Here, we present a computational framework to assemble contigs, evaluate their authenticity as ancient sequences, predict open reading frames, and fold ancient protein structures directly from highly damaged metagenomic data. Applying this pipeline to two-million-year-old datasets from the Kap Kobenhavn Formation, we successfully rescued ancient proteins involved in methane metabolism. By generating structural models with AlphaFold 2 and comparing them to modern predicted reference structures, we demonstrate that these ancient proteins can be reconstructed and aligned with high confidence. We showcase this by analyzing an archaeal V/A-type ATP synthase protein recovered from the 2M-year-old Greenlandic data. Ultimately, our work proves that ancient proteins can be reliably recovered from highly degraded palaeogenomic material, establishing a new computational avenue for evolutionary and biochemical research.

evolutionary biology↗

bioRxiv2025

Outbreak Investigation of Phytobacter diazotrophicus Using Fourier Transform Infrared Spectroscopy compared to cgMLST

IntroductionPhytobacter diazotrophicus is an emerging opportunistic pathogen associated with hospital outbreaks and a reservoir of antimicrobial resistance genes; however, its clonal epidemiology remains poorly understood. Accurate identification and rapid clonal typing are crucial for infection control. Core genome MLST (cgMLST) is the genomic gold standard; however, it is both costly and time-consuming. Fourier-transform infrared (FT-IR) spectroscopy using the IR Biotyper(R) emerges as a rapid and accessible typing alternative. ObjectiveThis study evaluated the performance of the IR Biotyper(R) in discriminating P. diazotrophicus outbreak isolates, compared to cgMLST. MethodologyEight epidemiologically relevant isolates, including two from a total parenteral nutrition (TPN) outbreak and four from a hemodialysis clinic, along with two outliers, were analyzed. Its identification was confirmed by API20E, MALDI-TOF MS, nifL gene qPCR, and WGS. Both cgMLST, based on 7,755 conserved genes, and the Fourier Transform Infrared (FT-IR) Spectroscopy by IR Biotyper(R) were used for clonal typing, with the spectral cutoff empirically determined based on known clonal isolates. ResultsBoth methodologies demonstrated remarkable agreement in detecting clonal clusters. The two TPN outbreak isolates showed 23 allele differences (ADs) by cgMLST and consistently clustered according to the IR Biotyper. Similarly, the four isolates from the outbreak at hemodialysis clinic formed homogeneous clusters by both methods (5-27 ADs by cgMLST). The outlier isolates were consistently discriminated. ConclusionFT-IR spectroscopy (IR Biotyper(R)) proved promising and complementary to cgMLST for typing P. diazotrophicus in outbreak scenarios, offering a rapid and cost-effective alternative for detecting clonal events. Its applicability in routine epidemiological surveillance is significant. Statement of ImportancePhytobacter diazotrophicus is an emerging opportunistic pathogen of increasing concern in clinical environments, associated with hospital outbreaks and carrying antimicrobial resistance genes. Rapid detection and precise clonal characterization are essential for infection control and epidemiological surveillance. However, these are challenging for less-understood species. Genomic methods, such as cgMLST, are the gold standard, but their cost and time limitations hinder routine application in outbreak response. This study evaluated the IR Biotyper system, a rapid and accessible phenotypic tool that uses Fourier Transform Infrared Spectroscopy, compared to cgMLST for clonal typing of P. diazotrophicus. Our findings demonstrate a remarkable agreement between the two methods in identifying clonal clusters. This validation suggests the potential of the IR Biotyper as an effective tool for real-time screening and active surveillance of P. diazotrophicus in clinical settings. This enables a more agile response to outbreaks and aids in managing antimicrobial resistance, reserving high-resolution genomic analyses for more in-depth investigations.

microbiology↗

bioRxiv2025

AdDeam: A Fast and Scalable Tool for Estimating and Clustering Reference-Level Damage Profiles

MotivationDNA damage patterns, such as increased frequencies of C[->]T and G[->]A substitutions at fragment ends, are widely used in ancient DNA studies to assess authenticity and detect contamination. In metagenomic studies, fragments can be mapped against multiple references or de novo assembled contigs to identify those likely to be ancient. Generating and comparing damage profiles, however, can be both tedious and time-consuming. Although tools exist for estimating damage in single reference genomes and metagenomic datasets, none efficiently cluster damage patterns. ResultsTo address this methodological gap, we developed AdDeam, a tool that combines rapid damage estimation with clustering for streamlined analyses and easy identification of potential contaminants or outliers. Our tool takes aligned aDNA fragments from various samples or contigs as input, computes damage patterns, clusters them and outputs representative damage profiles per cluster, a probability of each sample of pertaining to a cluster as well as a PCA of the damage patterns for each sample for fast visualisation. We evaluated AdDeam on both simulated and empirical datasets. AdDeam effectively distinguishes different damage levels, such as UDG-treated samples, sample-specific damages from specimens of different time periods, and can also distinguish between contigs containing modern or ancient fragments, providing a clear framework for aDNA authentication and facilitating large-scale analyses. Availability and ImplementationAdDeam is publicly available at https://github.com/LouisPwr/AdDeam and can also be installed via Bioconda. It is implemented in Python and C++. All analysis scripts and datasets are available at https://github.com/LouisPwr/AdDeamAnalysis and on Zenodo under: 10.5281/zenodo.15052427.

bioinformatics↗

bioRxiv2024

CarpeDeam: A De Novo Metagenome Assemblerfor Heavily Damaged Ancient Datasets

De novo assembly of ancient metagenomic datasets is a challenging task. Ultra-short fragment size and characteristic postmortem damage patterns of sequenced ancient DNA molecules leave current tools ill-equipped for ideal assembly. We present CarpeDeam, a novel damage-aware de novo assembler designed specifically for ancient metagenomic samples. Utilizing maximum-likelihood frameworks that integrate sample-specific damage patterns, CarpeDeam demonstrates improved recovery of longer continuous sequences and protein sequences in many simulated and empirical datasets compared to existing assemblers. As a pioneering ancient metagenome assembler, CarpeDeam opens the door for new opportunities in functional and taxonomic analyses of ancient microbial communities.

genomics↗

bioRxiv2023

Benchmarking software tools for trimming adapters and merging next-generation sequencing data for ancient DNA

Ancient DNA is highly degraded, resulting in very short sequences. Reads generated with modern high-throughput sequencing machines are generally longer than ancient DNA molecules, therefore the reads often contain some portion of the sequencing adaptors. It is crucial to remove those adaptors, as they can interfere with downstream analysis. Furthermore, overlapping portions when DNA has been read forward and backward (paired-end) can be merged to correct sequencing errors and improve read quality. Several tools have been developed for adapter trimming and read merging, however, no one has attempted to evaluate their accuracy and evaluate their potential impact on downstream analyses. Through the simulation of sequencing data, seven commonly used tools were analyzed in their ability to reconstruct ancient DNA sequences through read merging. The analyzed tools exhibit notable differences in their abilities to correct sequence errors and identify the correct read overlap, but the most substantial difference is observed in their ability to calculate quality scores for merged bases. Selecting the most appropriate tool for a given project depends on several factors, although some tools such as fastp have some shortcomings, whereas others like leeHom outperform the other tools in most aspects. While the choice of tool did not result in a measurable difference when analyzing population genetics using principal component analysis, it is important to note that downstream analyses that rely on quality scores can be significantly impacted by the choice of tool.

bioinformatics↗

bioRxiv2021

LY-CoV1404 potently neutralizes SARS-CoV-2 variants

SARS-CoV-2 neutralizing monoclonal antibodies (mAbs) can reduce the risk of hospitalization when administered early during COVID-19 disease. However, the emergence of variants of concern has negatively impacted the therapeutic use of some authorized mAbs. Using a high throughput B-cell screening pipeline, we isolated a highly potent SARS-CoV-2 spike glycoprotein receptor binding domain (RBD)-specific antibody called LY-CoV1404 (also known as bebtelovimab). LY-CoV1404 potently neutralizes authentic SARS-CoV-2 virus, including the prototype, B.1.1.7, B.1.351 and B.1.617.2). In pseudovirus neutralization studies, LY-CoV1404 retains potent neutralizing activity against numerous variants including B.1.1.7, B.1.351, B.1.617.2, B.1.427/B.1.429, P.1, B.1.526, B.1.1.529, and the BA.2 subvariant and retains binding to spike proteins with a variety of underlying RBD mutations including K417N, L452R, E484K, and N501Y. Structural analysis reveals that the contact residues of the LY-CoV1404 epitope are highly conserved with the exception of N439 and N501. Notably, the binding and neutralizing activity of LY-CoV1404 is unaffected by the most common mutations at these positions (N439K and N501Y). The breadth of reactivity to amino acid substitutions present among current VOC together with broad and potent neutralizing activity and the relatively conserved epitope suggest that LY-CoV1404 has the potential to be an effective therapeutic agent to treat all known variants causing COVID-19. In BriefLY-CoV1404 is a potent SARS-CoV-2-binding antibody that neutralizes all known variants of concern and whose epitope is rarely mutated. HighlightsO_LILY-CoV1404 potently neutralizes SARS-CoV-2 authentic virus and known variants of concern including the B.1.1.529 (Omicron), the BA.2 Omicron subvariant, and B.1.617.2 (Delta) variants C_LIO_LINo loss of potency against currently circulating variants C_LIO_LIBinding epitope on RBD of SARS-CoV-2 is rarely mutated in GISAID database C_LIO_LIBreadth of neutralizing activity and potency supports clinical development C_LI

immunology↗

bioRxiv2020

LY-CoV555, a rapidly isolated potent neutralizing antibody, provides protection in a non-human primate model of SARS-CoV-2 infection

SARS-CoV-2 poses a public health threat for which therapeutic agents are urgently needed. Herein, we report that high-throughput microfluidic screening of antigen-specific B-cells led to the identification of LY-CoV555, a potent anti-spike neutralizing antibody from a convalescent COVID-19 patient. Biochemical, structural, and functional characterization revealed high-affinity binding to the receptor-binding domain, ACE2 binding inhibition, and potent neutralizing activity. In a rhesus macaque challenge model, prophylaxis doses as low as 2.5 mg/kg reduced viral replication in the upper and lower respiratory tract. These data demonstrate that high-throughput screening can lead to the identification of a potent antiviral antibody that protects against SARS-CoV-2 infection. One Sentence SummaryLY-CoV555, an anti-spike antibody derived from a convalescent COVID-19 patient, potently neutralizes SARS-CoV-2 and protects the upper and lower airways of non-human primates against SARS-CoV-2 infection.

immunology↗