Journal: PLoS medicine

Article Title: In celiac disease, a subset of autoantibodies against transglutaminase binds toll-like receptor 4 and induces activation of monocytes.

doi: 10.1371/journal.pmed.0030358

Figure Lengend Snippet: Figure 1. Celiac Peptide Is Recognized by Sera of Patients with Active Disease and Shares Homology with Microbial Antigens (A) The celiac peptide is recognized by serum IgA immunoglobulins of patients on GCD, but not by patients on GFD. Results are expressed as absorbance at 405 nm. (B) Sequence homology between the celiac peptide and infectious agents. The peptide sequence was compared with known microbial protein sequences using the BLASTP via the NCBI BLAST network service (: indicates identity and * indicates conservative substitutions). (C) Frequency of IgG antibodies directed against infectious agents in patients with active CD and in normal healthy controls. (D) Sera of patients with active CD contain IgA antibodies directed against the rotavirus major neutralizing protein VP-7. A rotavirus extract was probed with rabbit antiserum raised against a peptide (VIQVGGSNVLDI) of the VP-7 protein (Lane 1), with affinity-purified anti-celiac peptide antibodies (Lane 2), with antibodies affinity-purified against an irrelevant control peptide (Lane 3), with sera from adult patients with active disease (Lanes 4 and 5), with sera from the same patients on GFD (Lanes 7 and 8), and serum from a 1-y-old child with active CD (Lane 6) and on GFD (Lane 9). A peroxidase-labelled polyvalent anti-human Igs antibody (Lanes 2 and 3) and an anti-human IgA antibody (Lanes 4–9) were used for detection. DOI: 10.1371/journal.pmed.0030358.g001

Article Snippet: The anti-TLR4 monoclonal antibody clone HTA125, isotype IgG2b (Imgenex), was used for inhibition experiments.

Techniques: Sequencing, Affinity Purification, Control

Journal: PLoS medicine

Article Title: In celiac disease, a subset of autoantibodies against transglutaminase binds toll-like receptor 4 and induces activation of monocytes.

doi: 10.1371/journal.pmed.0030358

Figure Lengend Snippet: Figure 4. Purified Antibodies Directed against Celiac and VP-7 Peptides Bind Endomysial Structures Pooled affinity-purified antibodies against celiac (A and D) and VP-7 (B and E) peptides from ten patients bind endomysium. Purified antibodies against the irrelevant control peptide from five patients (C and F). (A–C) Slides stained with FITC conjugated anti-human IgA antibodies. (D–F) Slides stained with FITC conjugated anti-human IgG antibodies. DOI: 10.1371/journal.pmed.0030358.g004

Article Snippet: The anti-TLR4 monoclonal antibody clone HTA125, isotype IgG2b (Imgenex), was used for inhibition experiments.

Techniques: Purification, Affinity Purification, Control, Staining

Journal: PLoS medicine

Article Title: In celiac disease, a subset of autoantibodies against transglutaminase binds toll-like receptor 4 and induces activation of monocytes.

doi: 10.1371/journal.pmed.0030358

Figure Lengend Snippet: Figure 6. Pro-Inflammatory Cytokines Produced by Activated Monocytes Levels of IL-6 (A), IL-12 (B), and TNF-alpha (C) released in the supernatant by monocytes incubated with medium alone (Line 1), with antibodies directed against an irrelevant peptide (Line 2), with LPS (Line 3), with pooled Igs isolated from the 22 patients with active CD (Line 4), with pooled Igs isolated from the same patients on GFD (Line 5), with purified anti-celiac peptide antibodies obtained from ten patients (line 6), with purified anti-celiac peptide antibodies in the presence of an irrelevant mouse IgG2b antibody (20 lg/ml) (Line 7), and with purified anti-celiac peptide antibodies in the presence of the neutralizing mouse mono- clonal antibody anti-TLR4, clone HTA 125 (20 lg/ml) (Line 8). The y-axis represents the cytokine concentration expressed as pg/ml. Data represent the mean 6SD of three independently performed experi- ments. DOI: 10.1371/journal.pmed.0030358.g006

Article Snippet: The anti-TLR4 monoclonal antibody clone HTA125, isotype IgG2b (Imgenex), was used for inhibition experiments.

Techniques: Produced, Incubation, Isolation, Purification, Concentration Assay

Journal: eBioMedicine

Article Title: Global emergence of Carbapenem-resistant Hypervirulent Klebsiella pneumoniae driven by an IncFII K34 KPC-2 plasmid

doi: 10.1016/j.ebiom.2025.105627

Figure Lengend Snippet: Genomic features of four KL1-ST23 carbapenem-resistant hypervirulent Klebsiella pneumoniae .

Article Snippet: Among the 174 KPC-encoding genomes out of the 414 global CR-hvKp genomes, IncFII K34 KPC plasmid (24%, 42/174) was the most prevalent IncF KPC plasmid, followed by F33 (9%, 16/174) and F35 (8%, 14/174) plasmids ( g).

Techniques: Plasmid Preparation, CRISPR

Journal: eBioMedicine

Article Title: Global emergence of Carbapenem-resistant Hypervirulent Klebsiella pneumoniae driven by an IncFII K34 KPC-2 plasmid

doi: 10.1016/j.ebiom.2025.105627

Figure Lengend Snippet: Analysis of IncFII K1-34 alleles and global epidemiology of CR-hvKp . a . The classification tree for different IncFII K1-34 alleles was constructed using the Neighbour-Joining method in MEGA and was rooted at the midpoint. Sequences from the copA region of the IncFII K replicon were used in the tree construction. b . The alignment of the IncFII K34 allele with 33 known IncFII K alleles is shown. Dots indicate nucleotides identical to the consensus sequence, and dashes in grey represent gaps. c . The proportion of plasmids harbouring carbapenemase-encoding genes in each IncFII K plasmid type was calculated using data from PLSDB v. 2021_06_23_v2. d . The global distribution of 2,168 hvKp and CR-hvKp genomes with country information available from the NCBI GenBank is illustrated. Countries with more than 20 reported hvKp and CR-hvKp genomes were presented, and the predominant carbapenemase type within the country is highlighted on the top of the bar. e . The percentages of ST23, ST86, and ST65 among 2,343 hvKp and CR-hvKp strains are presented. f. The percentages of carbapenemases found among 414 CR-hvKp strains are detailed. g. The distribution of different IncF plasmid replicons among 174 KPC-encoding CR-hvKp strains is depicted.

Article Snippet: Among the 174 KPC-encoding genomes out of the 414 global CR-hvKp genomes, IncFII K34 KPC plasmid (24%, 42/174) was the most prevalent IncF KPC plasmid, followed by F33 (9%, 16/174) and F35 (8%, 14/174) plasmids ( g).

Techniques: Construct, Sequencing, Plasmid Preparation

Journal: eBioMedicine

Article Title: Global emergence of Carbapenem-resistant Hypervirulent Klebsiella pneumoniae driven by an IncFII K34 KPC-2 plasmid

doi: 10.1016/j.ebiom.2025.105627

Figure Lengend Snippet: Comparative analysis of KPC plasmids in CR-hvKp clinical isolates and phylogenetic analysis of ST23 hvKp and CR-hvKp strains . a . Circular comparison of the three IncFII K34 plasmids (pKPC-H39, pKPC-DD02357 and pKPC-DD01665) in CR-hvKp with pDD02172-2 (CP087613), p1140-KPC (CP047689) and pKP18069-KPC (CP059890). b . Genetic elements of bla KPC-2 on IncFII K34 plasmid. c . Circular comparison of the IncN plasmid (pKPC-DD02201) in CR-hvKp with pNB5_KPC-2 (CP092655), pSZN_KPC (MH917123) and pECN580 (KF914891). d . Genetic elements of bla KPC-2 on IncN plasmid. Open reading frames (ORFs) are indicated by arrows and coloured based on predicted gene function. The replication-associated genes are denoted as blue arrows. Resistance genes are indicated by red arrows, while insertion sequences are shown by yellow arrows. Genes in the conjugation module are shown by green arrows. Other genes are indicated by grey arrows. e . Time-based phylogenetic analysis of 342 ST23 hvKp and CR-hvKp strains. Colours in columns illustrate region of origin, BAPS cluster, host, source of isolation, K locus type, ICE type, carbapenemase type and presence of replicons. Selected divergence time and 95% CIs are shown at the nodes. All the representative genomes are complete, and strains with red dots on the graph are hvKp and with blue dots are CR-hvKp. The nine CR-hvKp strains harbouring the IncFII K34 bla KPC-2 plasmids are indicated by black arrows.

Article Snippet: Among the 174 KPC-encoding genomes out of the 414 global CR-hvKp genomes, IncFII K34 KPC plasmid (24%, 42/174) was the most prevalent IncF KPC plasmid, followed by F33 (9%, 16/174) and F35 (8%, 14/174) plasmids ( g).

Techniques: Comparison, Plasmid Preparation, Conjugation Assay, Isolation

Journal: eBioMedicine

Article Title: Global emergence of Carbapenem-resistant Hypervirulent Klebsiella pneumoniae driven by an IncFII K34 KPC-2 plasmid

doi: 10.1016/j.ebiom.2025.105627

Figure Lengend Snippet: Conjugation assays of bla KPC-2 bearing plasmids and RNA-seq analysis of transconjugants . Conjugation frequencies of KPC-2 plasmids a from CR-hvKp or CRKp isolates to E. coli J53, and b from E. coli J53 to K. pneumoniae NTUH-K2044. c . Conjugation frequencies of virulence plasmids from K. pneumoniae NTUH-K2044 to E. coli J53. Three biologic repeats were performed for each strain. Conjugation frequencies are calculated by dividing the number of transconjugants (CFU/mL) by the number of recipients (CFU/mL) and indicated by mean ± SD. P values were calculated from student t-tests, ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. d . The schematic diagram of the conjugation experiments. The numbers on the arrows are conjugation frequencies. The drugs on the bottom were used for selection in each conjugation experiment. MEM is meropenem, NaN 3 is sodium azide and K 2 TeO 3 is potassium tellurite. Only plasmids were shown in the figure. Plasmid profiles were from the results of Nanopore sequencing. The plasmid profile of HS11286 were from the GenBank. e. Heatmap of the RNA-seq analysis of NTUH-K2044 conjugated with IncFII K34 and IncFII K2 KPC-2 plasmids. Three biologic repeats were performed for each strain. f. Volcano plot of DEGs between NTUH-K2044 conjugated with IncFII K34 and IncFII K2 plasmids. g . Heatmap of expression of conjugation-associated genes.

Article Snippet: Among the 174 KPC-encoding genomes out of the 414 global CR-hvKp genomes, IncFII K34 KPC plasmid (24%, 42/174) was the most prevalent IncF KPC plasmid, followed by F33 (9%, 16/174) and F35 (8%, 14/174) plasmids ( g).

Techniques: Conjugation Assay, RNA Sequencing, Selection, Plasmid Preparation, Nanopore Sequencing, Expressing

Journal: eBioMedicine

Article Title: Global emergence of Carbapenem-resistant Hypervirulent Klebsiella pneumoniae driven by an IncFII K34 KPC-2 plasmid

doi: 10.1016/j.ebiom.2025.105627

Figure Lengend Snippet: The competition experiments of NTUH-K2044 conjugated with IncFII K34 and IncFII K2 bla KPC-2 bearing plasmids . Growth curves of isolates cultured with no meropenem a , 0.5 μg/mL meropenem b , 1 μg/mL meropenem c and 2 μg/mL meropenem d . e . The competition between isolates under conditions with and without 1 μg/mL meropenem. NTUH-K2044::pKPC-H39 and NTUH-K2044::pKPC-HS11286 were used in the competition assays. The copies of bla KPC-2 f and pKPC-H39, pKPC-DD02357 and pKPC-HS11286 in NTUH-K2044 g , respectively. Transcriptional levels of bla KPC-2 in qRT-PCR h and RNA-seq i . The transcriptional levels of bla KPC-2 in qRT-PCR were normalized to the endogenous reference gene rrsE . The normalized read counts of bla KPC-2 in RNA-seq were from the DESeq analysis results. The MICs of NTUH-K2044::pKPC-H39 and NTUH-K2044:: pKPC-DD02357 harbouring IncFII K34 KPC-2 plasmids to meropenem were 8 μg/mL, while NTUH-K2044::pKPC-HS11286 with the IncFII K2 KPC-2 plasmid were 4 μg/mL. Three biologic repeats were performed for each strain. P values were calculated from student t-tests, ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001.

Article Snippet: Among the 174 KPC-encoding genomes out of the 414 global CR-hvKp genomes, IncFII K34 KPC plasmid (24%, 42/174) was the most prevalent IncF KPC plasmid, followed by F33 (9%, 16/174) and F35 (8%, 14/174) plasmids ( g).

Techniques: Cell Culture, Quantitative RT-PCR, RNA Sequencing, Plasmid Preparation