Proline-106 EPSPS Mutation Imparting Glyphosate Resistance in Goosegrass (Eleusine indica) Emerges in South America

Weed Science ◽  
2018 ◽  
Vol 67 (1) ◽  
pp. 48-56 ◽  
Author(s):  
Hudson K. Takano ◽  
Rafael R. Mendes ◽  
Leonardo B. Scoz ◽  
Ramiro F. Lopez Ovejero ◽  
Jamil Constantin ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Substitution ◽  
Rapid Detection ◽  
Resistance Mechanism ◽  
Fold Increase ◽  
In Planta ◽  
Eleusine Indica ◽  
Target Site Resistance ◽  
Resistance Trait

AbstractGlyphosate-resistant (GR) goosegrass [Eleusine indica(L.) Gaertn.] was recently identified in Brazil, but its resistance mechanism was unknown. This study elucidated the resistance mechanism in this species and developed a molecular marker for rapid detection of this target-site resistance trait. The resistance factor for the resistant biotype was 4.4-fold compared with the glyphosate-susceptible (GS) in greenhouse dose–response experiments. This was accompanied by a similar (4-fold) difference in the levels of in vitro andin plantashikimate accumulation in these biotypes. However, there was no difference in uptake, translocation, or metabolism of glyphosate between the GS and GR biotypes. Moreover, both biotypes showed similar values for 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) copy number and transcription. Sequencing of a 330-bp fragment of theEPSPSgene identified a single-nucleotide polymorphism that led to a Pro-106-Ser amino acid substitution in the enzyme from the GR biotype. This mutation imparted a 3.8-fold increase in the amount of glyphosate required to inhibit 50% of EPSPS activity, confirming the role of this amino acid substitution in resistance to glyphosate. A quantitative PCR–based genotyping assay was developed for the rapid detection of resistant plants containing this Pro-106-Ser mutation.

scholarly journals Alterations in Penicillin-Binding Protein 1A Confer Resistance to β-Lactam Antibiotics in Helicobacter pylori

2002 ◽  
Vol 46 (7) ◽  
pp. 2229-2233 ◽  
Author(s):  
M. M. Gerrits ◽  
D. Schuijffel ◽  
A. A. van Zwet ◽  
E. J. Kuipers ◽  
C. M. J. E. Vandenbroucke-Grauls ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Amino Acid ◽  
Amino Acid Substitution ◽  
Binding Protein ◽  
Resistance Mechanism ◽  
Fold Increase ◽  
Single Amino Acid ◽  
Penicillin Binding Protein ◽  
H Pylori ◽  
High Level

ABSTRACT Most Helicobacter pylori strains are susceptible to amoxicillin, an important component of combination therapies for H. pylori eradication. The isolation and initial characterization of the first reported stable amoxicillin-resistant clinical H. pylori isolate (the Hardenberg strain) have been published previously, but the underlying resistance mechanism was not described. Here we present evidence that the β-lactam resistance of the Hardenberg strain results from a single amino acid substitution in HP0597, a penicillin-binding protein 1A (PBP1A) homolog of Escherichia coli. Replacement of the wild-type HP0597 (pbp1A) gene of the amoxicillin-sensitive (Amxs) H. pylori strain 1061 by the Hardenberg pbp1A gene resulted in a 100-fold increase in the MIC of amoxicillin. Sequence analysis of pbp1A of the Hardenberg strain, the Amxs H. pylori strain 1061, and four amoxicillin-resistant (Amxr) 1061 transformants revealed a few amino acid substitutions, of which only a single Ser414→Arg substitution was involved in amoxicillin resistance. Although we cannot exclude that mutations in other genes are required for high-level amoxicillin resistance of the Hardenberg strain, this amino acid substitution in PBP1A resulted in an increased MIC of amoxicillin that was almost identical to that for the original Hardenberg strain.

A novel polymorphism of human gene has an amino acid substitution (V365M) that decreases enzymatic activity in vitro and in vivo

2004 ◽  
Vol 76 (6) ◽  
pp. 519-527 ◽  
Author(s):  
T FUKAMI ◽  
M NAKAJIMA ◽  
R YOSHIDA ◽  
Y TSUCHIYA ◽  
Y FUJIKI ◽  
...  
Keyword(s):  
Amino Acid ◽  
Enzymatic Activity ◽  
Amino Acid Substitution ◽  
Human Gene

Release of somatostatin from rodent antral mucosae maintained in organ culture

1981 ◽  
Vol 241 (1) ◽  
pp. G59-G66
Author(s):  
L. M. Lichtenberger ◽  
L. S. Shaw ◽  
R. B. Bailey
Keyword(s):  
Amino Acid ◽  
Bovine Serum Albumin ◽  
Organ Culture ◽  
Fold Increase ◽  
Mean Value ◽  
Hormone Release ◽  
Fourfold Increase ◽  
Somatostatin Secretion ◽  
Mucosal Explants

Somatostatin secretion was investigated from rodent antral mucosal biopsies maintained in organ culture. Under control conditions, the somatostatin secretory output was fairly constant, with each biopsy releasing a mean value of 29.1 +/- 7.5 pg of hormone over a 15-min period. Incubation of the mucosa in medium containing 5% peptone directly induced a 20-fold increase in medium somatostatin levels to 622.5 +/- 15.5 pg/ml. In contrast, a peptone dialysate was a weaker stimulant of hormone release, inducing a three- to fourfold increase in medium somatostatin concentration. Additionally, it was determined that a peptic hydrolysate of bovine serum albumin was a significantly more potent in vitro stimulant of somatostatin secretion than the intact undigested molecule. These results suggest that the amino acid and peptide constituents of these proteinaceous substances are the major stimulants of somatostatin release in vitro. It is presently uncertain whether this direct peptone-induced release of somatostatin from antral mucosal explants can account for the previously reported [Am. J. Physiol. 235 (Endocrinol. Metab. Gastrointest. Physiol. 4): E410-E415, 1978] gastrin secretory "off response" to the same agent, because addition of exogenous somatostatin to the medium failed to block this peptone-induced gastrin secretory response.

scholarly journals Fluoroquinolone resistance in Clostridium difficile isolates from a prospective study of C. difficile infections in Europe

2008 ◽  
Vol 57 (6) ◽  
pp. 784-789 ◽  
Author(s):  
Patrizia Spigaglia ◽  
Fabrizio Barbanti ◽  
Paola Mastrantonio ◽  
Jon S. Brazier ◽  
Frédéric Barbut ◽  
...  
Keyword(s):  
Amino Acid ◽  
Clostridium Difficile ◽  
Prospective Study ◽  
Amino Acid Substitution ◽  
Amino Acid Change ◽  
Fluoroquinolone Resistance ◽  
Genes Encoding ◽  
High Level ◽  
A Prospective Study

The European Study Group on Clostridium difficile (ESGCD) conducted a prospective study in 2005 to monitor and characterize C. difficile strains circulating in European hospitals, collecting 411 isolates. Eighty-three of these isolates, showing resistance or intermediate resistance to moxifloxacin (MX), were selected for this study to assess susceptibility to other fluoroquinolones (FQs) and to analyse the gyr genes, encoding the DNA gyrase subunits GyrA and GyrB. Twenty MX-susceptible isolates from the surveillance study were included for comparison. Overall, one amino acid substitution in GyrA (Thr82 to Ile) and four different substitutions in GyrB (Ser416 to Ala, Asp426 to Asn, Asp426 to Val and Arg447 to Lys) were identified. A high level of resistance (MIC ≥32 μg ml−1) to MX, ciprofloxacin (CI), gatifloxacin (GA) and levofloxacin (LE) was found in 68 isolates showing the amino acid substitution Thr82 to Ile in GyrA, in eight isolates with the substitutions Thr82 to Ile in GyrA and Ser416 to Ala in GyrB, in two isolates showing the substitution Asp426 to Asn in GyrB and in one isolate with Asp426 to Val in GyrB. The remaining four isolates showed high MICs for CI and LE, but different MIC levels for MX and GA. In particular, intermediate levels of resistance to MX were shown by two isolates, one with the substitution Thr82 to Ile in GyrA, and one showing Asp426 to Asn in GyrB. The substitution Arg447 to Lys in GyrB was found in two strains resistant to MX, CI and LE but susceptible to GA. No substitutions in GyrA were found in the FQ-susceptible strains, whereas two strains showed the amino acid change Ser416 to Ala in GyrB. Thr82 to Ile was the most frequent amino acid change identified in the C. difficile isolates examined. In contrast to previous observations, 10 % of the isolates showed this substitution in association with Ser416 to Ala in GyrB. The other amino acid changes found were characteristic of a few strains belonging to certain types and/or countries. Two new substitutions for C. difficile, Ser416 to Ala and Arg447 to Lys, were found in GyrB. Whereas the former does not seem to have a key role in resistance, since it was also detected in susceptible strains, the latter substitution occurred in the same position where other amino acid variations take place in resistant Escherichia coli and other C. difficile strains. A large number of C. difficile isolates now show an alarming pattern of resistance to the majority of FQs currently used in hospitals and outpatient settings, therefore judicious use of these antibiotics and continuous monitoring of in vitro resistance are necessary.

scholarly journals Amino Acid Substitutions in GyrA of Burkholderia glumae Are Implicated in Not Only Oxolinic Acid Resistance but Also Fitness on Rice Plants

2006 ◽  
Vol 73 (4) ◽  
pp. 1114-1119 ◽  
Author(s):  
Yukiko Maeda ◽  
Akinori Kiba ◽  
Kouhei Ohnishi ◽  
Yasufumi Hikichi
Keyword(s):  
Amino Acid ◽  
Amino Acid Substitution ◽  
Oxolinic Acid ◽  
The Other ◽  
Amino Acid Substitutions ◽  
Burkholderia Glumae ◽  
Field Isolates ◽  
Rice Plants ◽  
Resistant Mutants

ABSTRACT Oxolinic acid (OA) resistance in field isolates of Burkholderia glumae, a causal agent of bacterial grain rot, is dependent on an amino acid substitution at position 83 in GyrA (GyrA83). In the present study, among spontaneous in vitro mutants from the OA-sensitive B. glumae strain Pg-10, we selected OA-resistant mutants that emerged at a rate of 5.7 � 10−10. Nucleotide sequence analysis of the quinolone resistance-determining region in GyrA showed that Gly81Cys, Gly81Asp, Asp82Gly, Ser83Arg, Asp87Gly, and Asp87Asn are observed in these OA-resistant mutants. The introduction of each amino acid substitution into Pg-10 resulted in OA resistance, similar to what was observed for mutants with the responsible amino acid substitution. In vitro growth of recombinants with Asp82Gly was delayed significantly compared to that of Pg-10; however, that of the other recombinants did not differ significantly. The inoculation of each recombinant into rice spikelets did not result in disease. In inoculated rice spikelets, recombinants with Ser83Arg grew less than Pg-10 during flowering, and growth of the other recombinants was reduced significantly. On the other hand, the reduced growth of recombinants with Ser83Arg in spikelets was compensated for under OA treatment, resulting in disease. These results suggest that amino acid substitutions in GyrA of B. glumae are implicated in not only OA resistance but also fitness on rice plants. Therefore, GyrA83 substitution is thought to be responsible for OA resistance in B. glumae field isolates.

scholarly journals Resistance to ACCase inhibitors in Eleusine indica from Brazil involves a target site mutation

2012 ◽  
Vol 30 (3) ◽  
pp. 675-681 ◽  
Author(s):  
M.D. Osuna ◽  
I.C.G.R. Goulart ◽  
R.A. Vidal ◽  
A. Kalsing ◽  
J.P. Ruiz Santaella ◽  
...  
Keyword(s):  
Amino Acid Position ◽  
Target Site ◽  
In Vitro Assays ◽  
Site Mutation ◽  
Eleusine Indica ◽  
Herbicide Resistant ◽  
Frequent Use ◽  
Target Site Resistance ◽  
Grass Weed

Eleusine indica (goosegrass) is a diploid grass weed which has developed resistance to ACCase inhibitors during the last ten years due to the intensive and frequent use of sethoxydim to control grass weeds in soybean crops in Brazil. Plant dose-response assays confirmed the resistant behaviour of one biotype obtaining high resistance factor values: 143 (fenoxaprop), 126 (haloxyfop), 84 (sethoxydim) to 58 (fluazifop). ACCase in vitro assays indicated a target site resistance as the main cause of reduced susceptibility to ACCase inhibitors. PCR-generated fragments of the ACCase CT domain of the resistant and sensitive reference biotype were sequenced and compared. A point mutation was detected within the triplet of aspartate at the amino acid position 2078 (referred to EMBL accession no. AJ310767) and resulted in the triplet of glycine. These results constitute the first report on a target site mutation for a Brazilian herbicide resistant grass weed.

scholarly journals Selection of the same mutation in the U69 protein kinase gene of human herpesvirus-6 after prolonged exposure to ganciclovir in vitro and in vivo

2001 ◽  
Vol 82 (11) ◽  
pp. 2767-2776 ◽  
Author(s):  
Chaysavanh Manichanh ◽  
Camille Olivier-Aubron ◽  
Jean-Pierre Lagarde ◽  
Jean-Thierry Aubin ◽  
Phillipe Bossi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Kinase ◽  
Amino Acid Substitution ◽  
Consensus Sequence ◽  
Human Herpesvirus ◽  
Wild Type ◽  
Human Herpesvirus 6 ◽  
Genotypic Analysis ◽  
Herpesvirus 6

After serial passage in the presence of increasing concentrations of ganciclovir (GCV) in vitro, a human herpesvirus-6 (HHV-6) mutant exhibiting a decreased sensitivity to the drug was isolated. Analysis of drug susceptibility showed that the IC50 of this mutant was 24-, 52- and 3-fold higher than that of the wild-type (wt) IC50 in the case of GCV, cidofovir and foscarnet, respectively. Genotypic analysis showed two single nucleotide changes as compared to the wild-type: an A→G substitution of the U69 protein kinase (PK) gene resulted in an M318V amino acid substitution and the other change, located in the C-terminal part of the U38 gene, resulted in an A961V amino acid substitution within the DNA polymerase. The M318V change was located within the consensus sequence DISPMN of the putative catalytic domain VI of the PK. This change was homologous to the M460V and M460I changes that had been reported previously within the consensus sequence DITPMN of the human cytomegalovirus (HCMV) UL97 PK and associated with the resistance of HCMV to GCV. The M318V change was also detected by PCR in HHV-6-infected PBMCs from an AIDS patient who had been treated with GCV for a long period of time and exhibited a clinically GCV-resistant HCMV infection. These findings provide strong circumstantial evidence that the M318V change of the PK gene is associated with resistance to GCV and raise the question of cross resistance to this drug among different betaherpesviruses.

scholarly journals In Vitro Activities and Inoculum Effects of Ceftazidime-Avibactam and Aztreonam-Avibactam against Carbapenem-Resistant Enterobacterales Isolates from South Korea

Antibiotics ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 912
Author(s):  
Taeeun Kim ◽  
Seung Cheol Lee ◽  
Moonsuk Bae ◽  
Heungsup Sung ◽  
Mi-Na Kim ◽  
...  
Keyword(s):  
Resistance Mechanism ◽  
Fold Increase ◽  
Multidrug Resistant ◽  
Minimal Inhibitory Concentrations ◽  
E Coli ◽  
Carbapenem Resistant ◽  
High Inoculum ◽  
Inoculum Effect

Ceftazidime-avibactam (CAZ-AVI) and aztreonam-avibactam (AZT-AVI) are novel antibiotic combinations active against multidrug-resistant Gram-negative pathogens. This study aimed to evaluate their in vitro activities and inoculum effects in carbapenem-resistant Enterobacterales (CRE), including carbapenemase-producing (CP)-CRE and non-CP-CRE. A total of 81 independent clinical isolates of carbapenem-resistant Escherichia coli and Klebsiella pneumoniae were collected. CAZ-AVI and AZT-AVI minimal inhibitory concentrations (MICs) were evaluated by broth microdilution using standard and high inocula. The inoculum effect was defined as an ≥8-fold increase in MIC with high inoculum. Phenotypic determination of β-lactam resistance mechanism and PCR for carbapenemase genes were performed. Of the 81 CRE isolates, 35 (43%) were CP-CRE. Overall, 73% of the isolates were susceptible to CAZ-AVI, and 95% had low AZT-AVI MICs (≤8 µg/mL). The MIC50/MIC90s of CAZ-AVI and AZT-AVI were 4/≥512 µg/mL and 0.5/4 µg/mL, respectively. CAZ-AVI was more active against non-CP-CRE than against CP-CRE (susceptibility 80% vs. 63%, p = 0.08; MIC50/MIC90, 2/16 μg/mL vs. 4/≥512 μg/mL), whereas AZT-AVI was more active against CP-CRE (MIC50/MIC90, 0.25/1 μg/mL vs. 0.5/8 μg/mL). All four isolates with high AZT-AVI MIC (≥16 μg/mL) were resistant to CAZ-AVI, but only 18% (4/22) of CAZ-AVI-resistant isolates had high AZT-AVI MIC. The rates of the inoculum effect for CAZ-AVI and AZT-AVI were 18% and 47%, respectively (p < 0.001). Interestingly, the frequency of the AZT-AVI inoculum effect was higher in K. pneumoniae than E. coli (64% vs. 8%, p < 0.001). AZT-AVI is more active against CRE than CAZ-AVI, even in CP-CRE and CAZ-AVI-resistant isolates. The presence of a substantial inoculum effect may contribute to clinical failure in high-inoculum infections treated with AZT-AVI.

USING RIGIDITY ANALYSIS TO PROBE MUTATION-INDUCED STRUCTURAL CHANGES IN PROTEINS

2012 ◽  
Vol 10 (03) ◽  
pp. 1242010 ◽  
Author(s):  
FILIP JAGODZINSKI ◽  
JEANNE HARDY ◽  
ILEANA STREINU
Keyword(s):  
Amino Acid ◽  
Amino Acid Substitution ◽  
Structural Changes ◽  
Protein Structures ◽  
Data Bank ◽  
Accessible Surface Area ◽  
Solvent Accessible Surface Area ◽  
Single Amino Acid ◽  
Fast Evaluation

Predicting the effect of a single amino acid substitution on the stability of a protein structure is a fundamental task in macromolecular modeling. It has relevance to drug design and understanding of disease-causing protein variants. We present KINARI-Mutagen, a web server for performing in silico mutation experiments on protein structures from the Protein Data Bank. Our rigidity-theoretical approach permits fast evaluation of the effects of mutations that may not be easy to perform in vitro, because it is not always possible to express a protein with a specific amino acid substitution. We use KINARI-Mutagen to identify critical residues, and we show that our predictions correlate with destabilizing mutations to glycine. In two in-depth case studies we show that the mutated residues identified by KINARI-Mutagen as critical correlate with experimental data, and would not have been identified by other methods such as Solvent Accessible Surface Area measurements or residue ranking by contributions to stabilizing interactions. We also generate 48 mutants for 14 proteins, and compare our rigidity-based results against experimental mutation stability data. KINARI-Mutagen is available at http://kinari.cs.umass.edu .

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