Type I nitroreductases of Escherichia coli

1981 ◽  
Vol 27 (1) ◽  
pp. 81-86 ◽  
Author(s):  
D. W. Bryant ◽  
D. R. McCalla ◽  
M. Leeksma ◽  
P. Laneuville
Keyword(s):  
Escherichia Coli ◽  
Reductase Activity ◽  
Deae Cellulose ◽  
Partial Purification ◽  
Type I ◽  
Activity Type ◽  
Purification Step ◽  
The Past ◽  
Type Ia ◽  
Resistant Strains

Analysis of partially purified crude extract of Escherichia coli K12 by chromatography and gel electrophoresis has resulted in the separation of three distinct activities which catalyse the reduction of nitrofurazone (semicarbazone of 5-nitro-2-furaldehyde) in the presence of oxygen (type I nitroreductases). The major enzymatic activity (type IA), which was dependent solely on NADPH as a cofactor, was absent from nitrofurazone-resistant strains NFR 402 and NFR 502, but present in SIL 41, a strain which is only marginally resistant to the nitrofuran. The remaining nitroreductase activities (IB1 and IB2) utilize either NADH or NADPH as a cofactor. These activities coelute from DEAE-cellulose at pH 7.2, but may be differentiated by their behaviour on CM-cellulose at pH 5.8. The reductase activity missing in SIL 41 was observed in extracts of strain NFR 402 but not NFR 502. This enzyme (IB1) though retained by DEAE-cellulose had no affinity for CM-cellulose. The only reductase present in extracts of NFR 502 (a nitrofuran-resistant strain selected after two mutational events) was type IB2. This activity, also detectable in SIL 41 and NFR 402, has not been mapped genetically. An interesting feature of the type IB2 enzyme is its apparent inactivation by MnCl2 which has been routinely used as a partial purification step in the past.

scholarly journals Characterization and antimicrobial resistance patterns of Escherichia coli isolated from feces of healthy broiler chickens

2016 ◽  
Vol 83 (0) ◽  
Author(s):  
Ariel Eurides Stella ◽  
Maria Cristina De Oliveira ◽  
Vera Lúcia Dias da Silva Fontana ◽  
Renato Paris Maluta ◽  
Clarissa Araújo Borges ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Broiler Chickens ◽  
Clinical Signs ◽  
Avian Pathogenic Escherichia Coli ◽  
E Coli ◽  
The Past ◽  
Pathogenic Escherichia Coli ◽  
Resistance Patterns ◽  
Resistant Strains ◽  
Polymerase Chain

ABSTRACT: Avian pathogenic Escherichia coli (APEC) strains are isolated from lesions of poultry presenting colibacillosis, which is a disease that causes either systemic or localized clinical signs. Such strains share many characteristics with E. coli strains that cause extra-intestinal illness in humans. There is not a consensus on how to define the APEC pathotype with regard to the presence of virulence traits. On the other hand, in the past few years, five minimal predictors for APEC detection were proposed. The E. coli isolates in this work were tested through polymerase chain reaction (PCR) to the five proposed minimal predictors and cva C. The strains presenting them were categorized as potential APEC. The APEC and non-APEC categories showed high resistance (> 50%) to cephalotin, erythromycin, streptomycin, sulphametoxazol/trimethoprim, ampicillin, and amoxicillin. Potential APEC strains were significantly more resistant to cephalotin (p < 0.05) and neomcycin (p < 0.01) than non-APEC. These latter were significantly more resistant to tetracycline (p < 0.01) than the potential APEC strains. These results demonstrate that feces of poultry present E. coli strains with resistant features, showing or not the potential of causing colibacillosis in poultry. Because APEC and extra-intestinal illness in humans may be similar, these resistant strains are of interest to public health.

scholarly journals The distribution of the DHFR genes in trimethoprim-resistant urinary tract isolates from Taiwan

1992 ◽  
Vol 109 (3) ◽  
pp. 453-462 ◽  
Author(s):  
Lin-Li Chang ◽  
Shui-Feng Chang ◽  
Teh-Yuan Chow ◽  
Wen-Jeng Wu ◽  
Jong-Chou Chang
Keyword(s):  
Escherichia Coli ◽  
Dihydrofolate Reductase ◽  
Type I ◽  
Type Ii ◽  
Colony Hybridization ◽  
Resistant Strains ◽  
High Level ◽  
Type V ◽  
Urinary Isolates ◽  
Level Resistance

SUMMARYBetween July 1987 and June 1989, 1054 urinary isolates of enterobacteria from Kaohsiung, Taiwan were studied for their trimethoprim resistance. Trimethoprim resistance was defined as MIC greater than 4 μg/ml and high-level resistance by MIC greater than 1000 μg/ml. The incidence of trimethoprim resistance increased from 33·6% in 1987 to 42·1% in 1989. Among the resistant strains studied, 90% were resistant to high levels of trimethoprim. An increase in the proportion of resistant strains (33·9–46·3%) exhibiting high-level non-transferable trimethoprim resistance was noted. The distribution of the dihydrofolate reductase (DHFR) genes by colony hybridization in 374 trimethoprim-resistant isolates revealed the presence of type I and type V DHFR genes in most of these isolates (45·4% and 10·4% respectively). Type I was predominant inEscherichia coliwhereas type V was frequently seen inEnterobacterspp. None showed homology with the type II and type III DHFR probe DNA. In addition, transposon Tn7 was present in 7·8% of 374 trimethoprim-resistant enterobacteria.

scholarly journals The diversity of alleles at the hsd locus in natural populations of Escherichia coli.

Genetics ◽  
1995 ◽  
Vol 140 (4) ◽  
pp. 1187-1197
Author(s):  
V A Barcus ◽  
A J Titheradge ◽  
N E Murray
Keyword(s):  
Escherichia Coli ◽  
Natural Populations ◽  
Enteric Bacteria ◽  
Type I ◽  
Bacterial Strains ◽  
E Coli ◽  
Alternative Families ◽  
Type Ia ◽  
Natural Isolates ◽  
Id Genes

Abstract In enteric bacteria three discrete families of type I restriction and modification systems (IA, IB and ID) are encoded by alleles of the serB-linked hsd locus. Probes specific for each of the three families were used to monitor the distribution of related systems in 37 of the 72 wild-type Escherichia coli strains comprising the ECOR collection. All 25 members of group A in this collection were screened; 12 were probe-positive, nine have hsd genes in the IA family, two in the IB and one in the ID. Twelve strains, representing all groups other than A, were screened; five were probe-positive, one has hsd genes in the IA family, one in the IB and three in the ID. The type ID genes are the first representatives of this family in E. coli, the probe-negative strains could have alternative families of hsd genes. The type IA and IB systems added at least five new specificities to the five already identified in natural isolates of E. coli. The distribution of alleles is inconsistent with the dendrogram of the bacterial strains derived from other criteria. This discrepancy and the dissimilar coding sequences of allelic hsd genes both imply lateral transfer of hsd genes.

scholarly journals Purification and properties of nitrite reductase from Escherichia coli K12

1978 ◽  
Vol 175 (2) ◽  
pp. 483-493 ◽  
Author(s):  
K J Coleman ◽  
A Cornish-Bowden ◽  
J A Cole
Keyword(s):  
Escherichia Coli ◽  
Nitrite Reductase ◽  
Specific Activity ◽  
Reductase Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Deae Cellulose ◽  
Sedimentation Coefficient ◽  
Coli Strain ◽  
Molecular Weights

NADH-nitrite oxidoreductase (EC 1.6.4) was purified to better than 95% homogeneity from batch cultures of Escherichia coli strain OR75Ch15, which is partially constitutive for nitrite reductase synthesis. Yields of purified enzyme were low, mainly because of a large loss of activity during chromatography on DEAE-cellulose. The quantitative separation of cytochrome c-552 from nitrite reductase activity resulted in an increase in the specific activity of the enzyme: this cytochrome is not therefore an integral part of nitrite reductase. The subunit molecular weights of nitrite reductase and of a haemoprotein contaminant, as determined by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, were 88000 and 80000 respectively. The sedimentation coefficient was calculated to be in the range 8.5-9.5S, consistent with a mol.wt. of 190000. It is suggested therefore that the native enzyme is a dimer with two identical or similar-sized subunits. Purest samples contained 0.4 mol of flavin/mol of enzyme, but no detectable haem. Catalytic activity was totally inhibited by 20 micron-p-chloromercuribenzoate and 1 mM-cyanide, slightly inhibited by 1 micron-sulphite and 10mM-arsenite, but insensitive to 1 mM-2,2′-bipyridine, 4mM-1,10-phenanthroline and 10mM-NaN3. Three molecules of NADH were oxidized for each NO2-ion reduced: the product of the reaction is therefore assumed to be NH4+. The specific activity of hydroxylamine reductase increased at each step in the purification of nitrite reductase, and the elution profiles for these two activities during chromatography on DEAE-Sephadex were coincident. It is likely that a single enzyme is responsible for both activities.

Mechanisms involved in effects of antimicrobial peptides, bactenectins ChBac3.4, ChBac5, and mini-ChBac7.5Nb, on bacterial cells

2017 ◽  
pp. 43-50
Author(s):  
О.В. Шамова ◽  
М.С. Жаркова ◽  
П.М. Копейкин ◽  
Д.С. Орлов ◽  
Е.А. Корнева
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Activity ◽  
Innate Immunity ◽  
Infectious Diseases ◽  
Metabolic Activity ◽  
Capra Hircus ◽  
Bacterial Cells ◽  
Antibiotic Resistant ◽  
Resistant Strains

Антимикробные пептиды (АМП) системы врожденного иммунитета - соединения, играющие важную роль в патогенезе инфекционных заболеваний, так как обладают свойством инактивировать широкий спектр патогенных бактерий, обеспечивая противомикробную защиту живых организмов. В настоящее время АМП рассматриваются как потенциальные соединения-корректоры инфекционной патологии, вызываемой антибиотикорезистентными бактериями (АБР). Цель данной работы состояла в изученим механизмов антибактериального действия трех пептидов, принадлежащих к семейству бактенецинов - ChBac3.4, ChBac5 и mini-ChBac7.5Nb. Эти химически синтезированные пептиды являются аналогами природных пролин-богатых АМП, обнаруженных в лейкоцитах домашней козы Capra hircus и проявляющих высокую антимикробную активность, в том числе и в отношении грамотрицательных АБР. Методы. Минимальные ингибирующие и минимальные бактерицидные концентрации пептидов (МИК и МБК) определяли методом серийных разведений в жидкой питательной среде с последующим высевом на плотную питательную среду. Эффекты пептидов на проницаемость цитоплазматической мембраны бактерий для хромогенного маркера исследовали с использованием генетически модифицированного штамма Escherichia coli ML35p. Действие бактенецинов на метаболическую активность бактерий изучали с применением маркера резазурина. Результаты. Показано, что все исследованные пептиды проявляют высокую антимикробную активность в отношении Escherichia coli ML35p и антибиотикоустойчивых штаммов Escherichia coli ESBL и Acinetobacter baumannii in vitro, но их действие на бактериальные клетки разное. Использован комплекс методик, позволяющих наблюдать в режиме реального времени динамику действия бактенецинов в различных концентрациях (включая их МИК и МБК) на барьерную функцию цитоплазматической мембраны и на интенсивность метаболизма бактериальных клеток, что дало возможность выявить различия в характере воздействия бактенецинов, отличающихся по структуре молекулы, на исследуемые микроорганизмы. Установлено, что действие каждого из трех исследованных бактенецинов в бактерицидных концентрациях отличается по эффективности нарушения целостности бактериальных мембран и в скорости подавления метаболизма клеток. Заключение. Полученная информация дополнит существующие фундаментальные представления о механизмах действия пролин-богатых пептидов врожденного иммунитета, а также послужит основой для биотехнологических исследований, направленных на разработку на базе этих соединений новых антибиотических препаратов для коррекции инфекционных заболеваний, вызываемых АБР и являющимися причинами тяжелых внутрибольничных инфекций. Antimicrobial peptides (AMPs) of the innate immunity are compounds that play an important role in pathogenesis of infectious diseases due to their ability to inactivate a broad array of pathogenic bacteria, thereby providing anti-microbial host defense. AMPs are currently considered promising compounds for treatment of infectious diseases caused by antibiotic-resistant bacteria. The aim of this study was to investigate molecular mechanisms of the antibacterial action of three peptides from the bactenecin family, ChBac3.4, ChBac5, and mini-ChBac7.5Nb. These chemically synthesized peptides are analogues of natural proline-rich AMPs previously discovered by the authors of the present study in leukocytes of the domestic goat, Capra hircus. These peptides exhibit a high antimicrobial activity, in particular, against antibiotic-resistant gram-negative bacteria. Methods. Minimum inhibitory and minimum bactericidal concentrations of the peptides (MIC and MBC) were determined using the broth microdilution assay followed by subculturing on agar plates. Effects of the AMPs on bacterial cytoplasmic membrane permeability for a chromogenic marker were explored using a genetically modified strain, Escherichia coli ML35p. The effect of bactenecins on bacterial metabolic activity was studied using a resazurin marker. Results. All the studied peptides showed a high in vitro antimicrobial activity against Escherichia coli ML35p and antibiotic-resistant strains, Escherichia coli ESBL and Acinetobacter baumannii, but differed in features of their action on bacterial cells. The used combination of techniques allowed the real-time monitoring of effects of bactenecin at different concentrations (including their MIC and MBC) on the cell membrane barrier function and metabolic activity of bacteria. The differences in effects of these three structurally different bactenecins on the studied microorganisms implied that these peptides at bactericidal concentrations differed in their capability for disintegrating bacterial cell membranes and rate of inhibiting bacterial metabolism. Conclusion. The obtained information will supplement the existing basic concepts on mechanisms involved in effects of proline-rich peptides of the innate immunity. This information will also stimulate biotechnological research aimed at development of new antibiotics for treatment of infectious diseases, such as severe in-hospital infections, caused by antibiotic-resistant strains.

scholarly journals The Use of Lavender (Lavandula angustifolia) Essential Oil as an Additive to Drinking Water for Broiler Chickens and Its In Vitro Reaction with Enrofloxacin

Animals ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1535
Author(s):  
M. Adaszyńska-Skwirzyńska ◽  
D. Szczerbińska ◽  
S. Zych
Keyword(s):  
Escherichia Coli ◽  
Drinking Water ◽  
Body Weight ◽  
Essential Oil ◽  
Water Consumption ◽  
Broiler Chickens ◽  
Lavender Oil ◽  
Resistant Strains ◽  
Synergistic Reaction

Biological activity of lavender essential oil is a property that can potentially find an application in poultry nutrition. Nowadays, the use of bioactive compounds is encouraged in many areas of industry and agriculture, since these substances have similar properties as withdrawn antibiotic growth promoters. Additionally, antibiotic resistance bacteria are one of the most important current threats to animal health. The purpose of the study was to determine the influence of lavender essential oil on the production parameters and blood parameters in broiler chickens and to assess the lavender oil’s in vitro reaction in a combination with enrofloxacin towards Escherichia coli. One-day-old non-sexed chicks (Ross 308) were divided into three experimental groups, each consisting of 100 individuals (five replicate of 20 boiler chicken each). The chickens in the control group received drinking water with no addition of lavender essential oil. In the experimental groups, lavender oil was added to the drinking water at a concentration of 0.4 mL/L, in the LEO1–42 from 1 to 42 days of age and the LEO22–42 group from the 22 to 42 days of age. The chickens’ body weight, feed consumption, water consumption, deaths and elimination due to health reasons were determined in the experiment. On day 42 of the chickens’ lives, blood samples were collected based on which selected parameters were identified. An in vitro experiment of lavender oil in combination with enrofloxacin was investigated with a checkerboard method. The results of the experiment showed the antimicrobial and antioxidant activity of lavender essential oil and its positive effect on the production results of broiler chickens. The study results proved that the addition of lavender oil positively impacted the chickens’ final body weight and feed conversion ratio (p < 0.01). No differences were observed between the groups for water consumption, death rate and the examined biochemical and immunological blood serum indices. Lavender essential oil was demonstrated to increase the blood serum’s total antioxidant status. A synergistic reaction in vitro was observed for lavender oil combined with enrofloxacin against resistant strains of Escherichia coli. Based on our study, a health-promoting effect of adding LEO to water for broiler chickens was found. Moreover, in vitro studies indicate a significant effect of lavender essential oil on the inhibition of the resistant strains of Escherichia coli growth and synergistic reaction with enrofloxacin.

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