Journal: Frontiers in Microbiology

Article Title: Genotypic and Phenotypic Characterization of Novel Sequence Types of Carbapenem-Resistant Acinetobacter baumannii, With Heterogeneous Resistance Determinants and Targeted Variations in Efflux Operons

doi: 10.3389/fmicb.2021.738371

Figure Lengend Snippet: Heat map represents antibiotic resistance of A. baumannii DR1, DR2, and AB067 strains. Antibiotic susceptibility testing was performed with different antibiotics in presence and absence of efflux pump inhibitor, (A) carbonyl cyanide m-chlorophenyl hydrazone (CCCP), and (B) reserpine against A. baumannii DR1, DR2, and AB067 strains. Data were tabulated and analyzed, and a heat map was constructed in GraphPad Prism software using mean zone of inhibition values of Kirby–Bauer assays to compare resistance and inhibition pattern among strains.

Article Snippet: The pmrAB locus was identified on (NOXP01000007.1, NOXO01000017.1, and LZOC01000005.1) in the genomes of strains DR1, DR2, and AB067, respectively, which exhibited > 99% identity with homologs from A. baumannii ATCC 17978.

Techniques: Construct, Software, Inhibition

Journal: Frontiers in Microbiology

Article Title: Genotypic and Phenotypic Characterization of Novel Sequence Types of Carbapenem-Resistant Acinetobacter baumannii, With Heterogeneous Resistance Determinants and Targeted Variations in Efflux Operons

doi: 10.3389/fmicb.2021.738371

Figure Lengend Snippet: Homology modeling of antibiotic resistance proteins in CRAB strains. (A) CarO from A. baumannii strains DR1, DR2, and AB067 strains were modeled using PDB 4fuv.1 as template in SWISS model ( https://swissmodel.expasy.org ) and visualized with Chimera 1.15 tool. (i) Superimposition of modeled monomeric CarO encoded by DR1, DR2, and AB067 strains were aligned with homo-dimeric CarO (PDB 4fuv.1). Homodimeric CarO (PDB 4fuv.1) (ii) , monomeric CarO from DR1 (iii) , DR2 (iv) , and AB067 (v) are displayed as modeled proteins. Mutations are highlighted in different colors. (B) Top view of overlapped monomeric Omp33–36 from DR1, DR2, and AB067 strains with A. baumannii Omp33 (PDB 6gie.1) as template (i) , panel (ii) A. baumannii Omp33 (PDB 6gie.1), (iii–v) corresponds to homology modeled monomeric Omp33–36 proteins of DR1, DR2, and AB967, respectively. (C) Side view of merged Omp33–36 modeled proteins from DR1, DR2, and AB067 strains ( i–v same as B ). Different strains displayed difference in loop regions. (D) Modeled AdeB multidrug resistance efflux pump from DR1, DR2, and AB067 strains were aligned with target template PDB 7kge.1 (i) and A. baumannii trimeric AdeB (PDB:7kge.1) (ii) , modeled AdeB from DR1 (iii) , DR2 (iv) , and AB067 (v) strains, respectively.

Article Snippet: The pmrAB locus was identified on (NOXP01000007.1, NOXO01000017.1, and LZOC01000005.1) in the genomes of strains DR1, DR2, and AB067, respectively, which exhibited > 99% identity with homologs from A. baumannii ATCC 17978.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Genotypic and Phenotypic Characterization of Novel Sequence Types of Carbapenem-Resistant Acinetobacter baumannii, With Heterogeneous Resistance Determinants and Targeted Variations in Efflux Operons

doi: 10.3389/fmicb.2021.738371

Figure Lengend Snippet: Comparative profiles of Biolog Phenotype MicroArray analysis of A. baumannii DR1, DR2, and AB067 strains. Phenotypic microarray analysis of A. baumannii DR1, DR2, and AB067 strains to determine metabolic utilization profiles using PM 09 to PM20 plates. Kinetic growth response curve for A. baumannii DR1 and DR2 (A) and A. baumannii DR1 and AB067 (B) strains in PM09 to PM20 plates (top panel left as PM09 to right bottom end panel as PM20). Growth kinetics of A. baumannii DR2 shown in green, compared with DR1 strain in red. Yellow indicates equivalent growth response or overlapping of DR1 and DR2 kinetic curves. Red kinetic response curves indicate a stronger response by DR1 strain, whereas green indicates a stronger growth response by DR2 strain. Heat maps of selected phenotype microarray profiles of A. baumannii DR1, DR2, and AB067 strains for antimicrobial compounds (C) , toxic (D) , and miscellaneous compounds (E) . Each dot represents Omnilog values representing all time points of bacterial growth. Omnilog values of 100 or more were considered for phenotype analysis. Gradient heat maps were generated among strains for each condition using Omnilog values in Morpheus online tool ( https://software.broadinstitute.org/morpheus ), which displayed blue color for relative lower growth, and red color represents high growth phenotype.

Article Snippet: The pmrAB locus was identified on (NOXP01000007.1, NOXO01000017.1, and LZOC01000005.1) in the genomes of strains DR1, DR2, and AB067, respectively, which exhibited > 99% identity with homologs from A. baumannii ATCC 17978.

Techniques: Microarray, Generated, Software

Journal: Frontiers in Microbiology

Article Title: Genotypic and Phenotypic Characterization of Novel Sequence Types of Carbapenem-Resistant Acinetobacter baumannii, With Heterogeneous Resistance Determinants and Targeted Variations in Efflux Operons

doi: 10.3389/fmicb.2021.738371

Figure Lengend Snippet: Determination of MIC for polymyxin and growth inactivation assays. (A) Growth of A. baumannii DR1, DR2, and AB067 strains was evaluated in presence of different polymyxin concentrations. Optical density of growth was plotted against a range of polymyxin concentrations to determine growth and minimum inhibitory concentrations of polymyxin. (B) Resazurin microtiter assay was performed to evaluate cell viability of A. baumannii DR1, DR2, and AB067 strains in presence of various concentrations of polymyxin (PMB). After period of incubation for 15 h at 37°C, PrestoBlue, a resazurin-based cell viability reagent (Invitrogen), was added to wells containing A. baumannii DR1, DR2, and AB067 strains and control (only media) and further incubated as per manufacturer’s protocol. After period of incubation, color change from natural resazurin blue to pink (reduced form) in wells was scored to determine MIC; no color change indicates no bacterial growth. (C) Growth inhibition assay was performed in presence and absence of efflux inhibitors verapamil (VER), CCCP, and reserpine (RES) (each of 5 μg/ml) with polymyxin in DR1 (i) , DR2 (ii) , and AB067 (iii) strains. (D) Growth inhibition assay was performed in presence of tetracycline as substrate for A. baumannii strains DR1 (i) , DR2 (ii) , and AB067 (iii) , tabulated and analyzed using GraphPad Prism software.

Article Snippet: The pmrAB locus was identified on (NOXP01000007.1, NOXO01000017.1, and LZOC01000005.1) in the genomes of strains DR1, DR2, and AB067, respectively, which exhibited > 99% identity with homologs from A. baumannii ATCC 17978.

Techniques: Incubation, Growth Inhibition Assay, Software

Journal: Frontiers in Microbiology

Article Title: Genotypic and Phenotypic Characterization of Novel Sequence Types of Carbapenem-Resistant Acinetobacter baumannii, With Heterogeneous Resistance Determinants and Targeted Variations in Efflux Operons

doi: 10.3389/fmicb.2021.738371

Figure Lengend Snippet: Relative expression levels of antibiotic-resistant determinants; efflux pumps, outer membrane protein, and porins in CRAB strains. A. baumannii DR1 strain was used as a reference control.

Article Snippet: The pmrAB locus was identified on (NOXP01000007.1, NOXO01000017.1, and LZOC01000005.1) in the genomes of strains DR1, DR2, and AB067, respectively, which exhibited > 99% identity with homologs from A. baumannii ATCC 17978.

Techniques: Expressing

Journal: Frontiers in Microbiology

Article Title: Genotypic and Phenotypic Characterization of Novel Sequence Types of Carbapenem-Resistant Acinetobacter baumannii, With Heterogeneous Resistance Determinants and Targeted Variations in Efflux Operons

doi: 10.3389/fmicb.2021.738371

Figure Lengend Snippet: (A) Linear genome map of A. baumannii strains DR1, DR2, and AB067 with other completely sequenced strains. Different strains used are A. baumannii strain 1461963 (JEWQ01), 1293320 (JFEE01), PR333 (NGDW01), OIFC111 (AMFY01), AB900 (ABXK01), AB900 (JAAVKC01), PR389 (NGEW01), PR373 (NGDA01), ARLG1941 (NGIN01), J9 (CP041587), DR1 (NOXP01), J15 (VKGL01), MRSN31461 (VHFT01), ABBL071 (LLGB01), PM2696 (JAAGSK01), VB25836 (RHLW01), BA18786 (JAAEGP01), AB_026 (RIAH01), AB067 (LZOC01), GML_KP7_Col_R_AB_TR (QHHE01), 4300STDY7045687 (UFIZ01), SGH0905 (PYEH01), 554L (NKXQ01), Aci00835 (VAFQ01), KUFAR39 (NEPK01), 461 (LCTE01), SP2107 (JAAGSY01), DR2 (NOXO01), SP2181 (JAAGTC01), XH731 (CP021321), AB252 (LXNE01), KT722 (RXIN01), GE_2017_62 (WIHB01), abaum000 (UWVS01), ACICU (CP031380), AYE (CU459141), SDF (CU468230), ATCC19606 (CZWC01), and ATCC_17978 (CP000521). Accession numbers are mentioned in parentheses for each genome. (B) Average nucleotide identity (ANI) analysis of A. baumannii strains DR1, DR2, and AB067 with other sequenced strains; values clearly indicate nucleotide level genome similarity in MDR strains used in this study belong to international clone 2. (C) Phylogenomic tree inferred based on single-nucleotide polymorphisms (SNP) in A. baumannii strains using maximum likelihood method. Unrooted phylogenomic tree was analyzed, and strains A. baumannii DR1 belong to ST128 (ST49) group, whereas both A. baumannii strains DR2 and AB067 that belonged to ST1816 (ST195) were distinctly clustered.

Article Snippet: The pmrAB locus was identified on (NOXP01000007.1, NOXO01000017.1, and LZOC01000005.1) in the genomes of strains DR1, DR2, and AB067, respectively, which exhibited > 99% identity with homologs from A. baumannii ATCC 17978.

Techniques:

Journal:

Article Title: Class II MHC antigen presentation defect in neonatal monocytes is not correlated with decreased MHC-II expression

doi: 10.1016/j.cellimm.2007.01.003

Figure Lengend Snippet: HLA-DR1 neonatal monocytes isolated from frozen PBMCs are deficient at MHC-II antigen processing and presentation of protein antigen and presentation of exogenous peptide

Article Snippet: Using our new strategy, antigen processing and presentation function was analyzed in negatively selected monocytes derived from frozen PBMC samples that expressed HLA-DR1 (allele DRB1*0101, ) or HLA-DR3 (allele DRB1*0301, ).

Techniques: Isolation