scholarly journals Antimicrobial Resistance and Sensitivity of Phytophthora agathidicida

2021 ◽  
Author(s):  
Kaitlyn Daley
Keyword(s):  
Natural Products ◽  
New Zealand ◽  
Antimicrobial Resistance ◽  
Plant Pathogens ◽  
Keystone Species ◽  
Resistance Mutations ◽  
E Coli ◽  
Oxysterol Binding Protein ◽  
Significant Difference

Members of the oomycete genus Phytophthora are highly infectious plant pathogens. P. agathidicida affects the New Zealand native keystone species Agathis australis(kauri) and is the cause of kauri dieback. The complex oomycete lifecycle makes Phytophthora infections hard to manage. The current management of kauri dieback has been limited and antimicrobial resistance is a concern. Phosphite agrichemical preparations are commonly used in the control of Phytophthora diseases, including kauri dieback. However, phosphite is not the only option; the agrichemicals oxathiapiprolin, and the plant-derived natural products polygodial and falcarindiol, have also been shown to have activity against P. agathidicida. The overall goal of this thesis was to further explore aspects of sensitivity and resistance of P. agathidicidatowards these four compounds.In New Zealand, there are three commercially available phosphite preparations, Agri-Fos 600, Phosgard, and Foschek. All previous studies have used Agri-Fos 600, so the first aim was to determine whether the particular formulation altered anti-oomycete activity. No significant difference was found between the 50% inhibitory concentrations (EC50 values) for the three formulations. Interestingly, however, formulating polygodial and falcarindiol with the surfactants and other non-phosphite ingredients of Foschek led to a significant increase in their inhibitory effects. The second aim of this thesis was to implement a serial passaging protocol for P. agathidicida and attempt to isolate mutants with increased resistance to phosphite, polygodial or falcarindiol. Serial passaging was carried out on amended agar plated with increasing concentrations of each chemical. However, even after 7 passages, over 16-18 weeks of growth, no mutants with increased resistance were isolated. This could be due to the complicated modes of action of the polygodial, falcarindioland phosphite, which makes it likely that several specific mutations are required to effect resistance. <br>IIOxathiapiprolin is a highly potent, new anti-oomycete agrichemical. It targets the Phytophthora oxysterol binding protein (OSBP) related protein (ORP1). Mutations in this protein are known to give oxathiapiprolin resistance in other species of Phytophthora; however, the P. agathidicida protein (PaORP1) has never been studied. In this work, the gene for PaORP1 was partially sequenced from five P. agathidicida isolates. None contained any of the known resistance mutations. A new protocol for expressing PaORP1 in E. coli and purifying it using immobilised metal affinity chromatography was also developed. After optimisation, this protocol yielded up to 30 mg of purified protein per litre of E. coli culture and is the first successful example of heterologously expressing and purifying any P. agathidicida protein. In future, this will allow the biomolecular interaction between PaORP1 and oxathiapiprolin to be studied in more detail. Overall, the work presented in this thesis assessed commercial formulations of phosphite, established a directed evolution protocol for studying resistance in P. agathidicida, and reported the first in vitro characterisation of a P. agathidicidaprotein. This research suggests that commercial formulation of plant-derived natural products may be a powerful new approach for combatting kauri dieback and, promisingly, also suggests that the risk of developing resistance to these compounds might be low.

scholarly journals Antimicrobial Resistance and Sensitivity of Phytophthora agathidicida

2021 ◽  
Author(s):  
Kaitlyn Daley
Keyword(s):  
Natural Products ◽  
New Zealand ◽  
Antimicrobial Resistance ◽  
Plant Pathogens ◽  
Keystone Species ◽  
Resistance Mutations ◽  
E Coli ◽  
Oxysterol Binding Protein ◽  
Significant Difference

Members of the oomycete genus Phytophthora are highly infectious plant pathogens. P. agathidicida affects the New Zealand native keystone species Agathis australis(kauri) and is the cause of kauri dieback. The complex oomycete lifecycle makes Phytophthora infections hard to manage. The current management of kauri dieback has been limited and antimicrobial resistance is a concern. Phosphite agrichemical preparations are commonly used in the control of Phytophthora diseases, including kauri dieback. However, phosphite is not the only option; the agrichemicals oxathiapiprolin, and the plant-derived natural products polygodial and falcarindiol, have also been shown to have activity against P. agathidicida. The overall goal of this thesis was to further explore aspects of sensitivity and resistance of P. agathidicidatowards these four compounds.In New Zealand, there are three commercially available phosphite preparations, Agri-Fos 600, Phosgard, and Foschek. All previous studies have used Agri-Fos 600, so the first aim was to determine whether the particular formulation altered anti-oomycete activity. No significant difference was found between the 50% inhibitory concentrations (EC50 values) for the three formulations. Interestingly, however, formulating polygodial and falcarindiol with the surfactants and other non-phosphite ingredients of Foschek led to a significant increase in their inhibitory effects. The second aim of this thesis was to implement a serial passaging protocol for P. agathidicida and attempt to isolate mutants with increased resistance to phosphite, polygodial or falcarindiol. Serial passaging was carried out on amended agar plated with increasing concentrations of each chemical. However, even after 7 passages, over 16-18 weeks of growth, no mutants with increased resistance were isolated. This could be due to the complicated modes of action of the polygodial, falcarindioland phosphite, which makes it likely that several specific mutations are required to effect resistance. <br>IIOxathiapiprolin is a highly potent, new anti-oomycete agrichemical. It targets the Phytophthora oxysterol binding protein (OSBP) related protein (ORP1). Mutations in this protein are known to give oxathiapiprolin resistance in other species of Phytophthora; however, the P. agathidicida protein (PaORP1) has never been studied. In this work, the gene for PaORP1 was partially sequenced from five P. agathidicida isolates. None contained any of the known resistance mutations. A new protocol for expressing PaORP1 in E. coli and purifying it using immobilised metal affinity chromatography was also developed. After optimisation, this protocol yielded up to 30 mg of purified protein per litre of E. coli culture and is the first successful example of heterologously expressing and purifying any P. agathidicida protein. In future, this will allow the biomolecular interaction between PaORP1 and oxathiapiprolin to be studied in more detail. Overall, the work presented in this thesis assessed commercial formulations of phosphite, established a directed evolution protocol for studying resistance in P. agathidicida, and reported the first in vitro characterisation of a P. agathidicidaprotein. This research suggests that commercial formulation of plant-derived natural products may be a powerful new approach for combatting kauri dieback and, promisingly, also suggests that the risk of developing resistance to these compounds might be low.

scholarly journals Biological Cost of Single and Multiple Norfloxacin Resistance Mutations in Escherichia coli Implicated in Urinary Tract Infections

2005 ◽  
Vol 49 (6) ◽  
pp. 2343-2351 ◽  
Author(s):  
Patricia Komp Lindgren ◽  
Linda L. Marcusson ◽  
Dorthe Sandvang ◽  
Niels Frimodt-Møller ◽  
Diarmaid Hughes
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Infection Model ◽  
Resistance Mutations ◽  
Topoisomerase Iv ◽  
E Coli ◽  
Tract Infections ◽  
Subsequent Selection

ABSTRACT Resistance to fluoroquinolones in urinary tract infection (UTIs) caused by Escherichia coli is associated with multiple mutations, typically those that alter DNA gyrase and DNA topoisomerase IV and those that regulate AcrAB-TolC-mediated efflux. We asked whether a fitness cost is associated with the accumulation of these multiple mutations. Mutants of the susceptible E. coli UTI isolate Nu14 were selected through three to five successive steps with norfloxacin. Each selection was performed with the MIC of the selected strain. After each selection the MIC was measured; and the regions of gyrA, gyrB, parC, and parE, previously associated with resistance mutations, and all of marOR and acrR were sequenced. The first selection step yielded mutations in gyrA, gyrB, and marOR. Subsequent selection steps yielded mutations in gyrA, parE, and marOR but not in gyrB, parC, or acrR. Resistance-associated mutations were identified in almost all isolates after selection steps 1 and 2 but in less than 50% of isolates after subsequent selection steps. Selected strains were competed in vitro, in urine, and in a mouse UTI infection model against the starting strain, Nu14. First-step mutations were not associated with significant fitness costs. However, the accumulation of three or more resistance-associated mutations was usually associated with a large reduction in biological fitness, both in vitro and in vivo. Interestingly, in some lineages a partial restoration of fitness was associated with the accumulation of additional mutations in late selection steps. We suggest that the relative biological costs of multiple mutations may influence the evolution of E. coli strains that develop resistance to fluoroquinolones.

scholarly journals Lactic Acid Bacteria (Lactobacillus bulgaricus and Streptococcus thermophillus) in Yogurt Inhibit the Growth of Escherichia coli, Salmonella typhimurium, and Shigella sp. In Vitro

2021 ◽  
Vol 31 (4) ◽  
pp. 2
Author(s):  
IDSAP Peramiarti
Keyword(s):  
Escherichia Coli ◽  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Salmonella Typhimurium ◽  
Pathogenic Bacteria ◽  
Test Method ◽  
P Value ◽  
E Coli ◽  
Significant Difference

Diarrhea is defecation with a frequency more often than usual (three times or more) a day (10 mL/kg/day) with a soft or liquid consistency, even in the form of water alone. Pathogenic bacteria, such as Escherichia coli, Salmonella typhimurium, and Shigella sp., play a role in many cases, to which antibiotics are prescribed as the first-line therapy. However, since antibiotic resistance cases are often found, preventive therapies are needed, such as consuming yogurt, which is produced through a fermentation process by lactic acid bacteria (LAB). This research aimed to determine the activity of lactic acid bacteria (Liactobacillus bulgaricus and Streptococcus thermophilus) in yogurt in inhibiting the growth of the pathogenic bacteria E. coli, S. typhimurium, and Shigella sp. The research applied in vitro with the liquid dilution test method and the true experimental design research method with post-test-only and control group design. The design was used to see the inhibitory effect of yogurt LAB on the growth of E. coli, S. typhimurium, and Shigell sp. to compare the effect of several different yogurt concentrations, namely 20%, 40%, 60%, and 80%. The results of the Least Significance Different analysis showed that there was a significant difference between yogurt with a concentration of 0% and that with various concentrations in inhibiting the growth of E. coli, S. typhimurium, and Shigella sp. with a p-value of &lt;0.05. Whereas, there was no significant difference in the various concentrations of yogurt in inhibiting the growth of the three kinds of bacteria with a p-value of &gt; 0.05.<p class="Default" align="center"> </p>

scholarly journals Effect of Long-Term Starvation on the Survival, Recovery, and Carbon Utilization Profiles of a Bovine Escherichia coli O157:H7 Isolate from New Zealand

2014 ◽  
Vol 80 (14) ◽  
pp. 4383-4390 ◽  
Author(s):  
Ron N. Xavier ◽  
Hugh W. Morgan ◽  
Ian R. McDonald ◽  
Helen Withers
Keyword(s):  
Escherichia Coli ◽  
New Zealand ◽  
Membrane Integrity ◽  
Nutrient Level ◽  
Carbon Utilization ◽  
Content Type ◽  
E Coli ◽  
Significant Difference ◽  
Carbon Utilization Profiles ◽  
Phosphate Buffered Saline

ABSTRACTThe ability to maintain a dual lifestyle of colonizing the ruminant gut and surviving in nonhost environments once shed is key to the success ofEscherichia coliO157:H7 as a zoonotic pathogen. Both physical and biological conditions encountered by the bacteria are likely to change during the transition between host and nonhost environments. In this study, carbon starvation at suboptimal temperatures in nonhost environments was simulated by starving a New Zealand bovineE. coliO157:H7 isolate in phosphate-buffered saline at 4 and 15°C for 84 days. Recovery of starved cells on media with different nutrient availabilities was monitored under aerobic and anaerobic conditions. We found that the New Zealand bovineE. coliO157:H7 isolate was able to maintain membrane integrity and viability over 84 days and that the level of recovery depended on the nutrient level of the recovery medium as well as the starvation temperature. In addition, a significant difference in carbon utilization was observed between starved and nonstarved cells.

Antimicrobial activity of recombinant E. coli expressed chicken AvBD-2 and its mRNA expression in Indian native Aseel and Kadaknath chicken breeds

2017 ◽  
Author(s):  
T. R. Kannaki ◽  
M. R. Reddy ◽  
P. C. Verma
Keyword(s):  
Antimicrobial Activity ◽  
Mrna Expression ◽  
Affinity Column ◽  
White Leghorn ◽  
Coding Region ◽  
Recombinant Peptide ◽  
E Coli ◽  
Significant Difference ◽  
Chicken Breeds

Avian b defensins (AvBD) are antimicrobial peptides that play a crucial role in the innate immune response in chickens. In the present study, chicken AvBD2 gene was cloned, expressed in E. coli system and the in vitro antimicrobial activity of recombinant peptide was evaluated. The entire mature peptide region of chicken AvBD2 region was amplified and cloned in pUC29 cloning vector. Further, the coding region was sub cloned in pET-28A expression vector. After transformation in E. coli cells, the peptide synthesis was induced and recombinant protein (7.7 kDa) was purified by using Ni-NTA affinity column. The recombinant chicken AvBD2 showed antibacterial activity against S. Pullorum. The minimum bactericidal concentration (MBC) of recombinant chicken AvBD2 evaluated by micro-broth dilution assay was 35 µg/ ml. We also quantified the expression of AvBD2 transcript expression in day-old spleen tissue of Indian native chicken breeds (Aseel and Kadaknath) and White Leghorn. Measurable AvBD2 mRNA expression was found in the spleen of all three breeds. However, no significant difference was found in AvBD2 gene expression between native chickens and White Leghorn.

scholarly journals Antimicrobial Drug Resistance Genes Do Not Convey a Secondary Fitness Advantage to Calf-Adapted Escherichia coli

2006 ◽  
Vol 72 (1) ◽  
pp. 443-448 ◽  
Author(s):  
Artashes R. Khachatryan ◽  
Dale D. Hancock ◽  
Thomas E. Besser ◽  
Douglas R. Call
Keyword(s):  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Null Mutant ◽  
Antimicrobial Drug Resistance ◽  
E Coli ◽  
Fitness Advantage ◽  
Antimicrobial Resistance Genes ◽  
Mutant Strains

ABSTRACT Maintenance of antimicrobial drug resistance in bacteria can be influenced by factors unrelated to direct selection pressure such as close linkage to other selectively advantageous genes and secondary advantage conveyed by antimicrobial resistance genes in the absence of drug selection. Our previous trials at a dairy showed that the maintenance of the antimicrobial resistance genes is not influenced by specific antimicrobial selection and that the most prevalent antimicrobial resistance phenotype of Escherichia coli is specifically selected for in young calves. In this paper we examine the role of secondary advantages conveyed by antimicrobial resistance genes. We tested antimicrobial-susceptible null mutant strains for their ability to compete with their progenitor strains in vitro and in vivo. The null mutant strains were generated by selection for spontaneous loss of resistance genes in broth supplemented with fusaric acid or nickel chloride. On average, the null mutant strains were as competitive as the progenitor strains in vitro and in newborn calves (in vivo). Inoculation of newborn calves at the dairy with antimicrobial-susceptible strains of E. coli did not impact the prevalence of antimicrobial-resistant E. coli. Our results demonstrate that the antimicrobial resistance genes are not responsible for the greater fitness advantage of antimicrobial-resistant E. coli in calves, but the farm environment and the diet clearly exert critical selective pressures responsible for the maintenance of antimicrobial resistance genes. Our current hypothesis is that the antimicrobial resistance genes are linked to other genes responsible for differential fitness in dairy calves.

scholarly journals The Streptomycin-Sulfadiazine-Tetracycline Antimicrobial Resistance Element of Calf-Adapted Escherichia coli Is Widely Distributed among Isolates from Washington State Cattle

2007 ◽  
Vol 74 (2) ◽  
pp. 391-395 ◽  
Author(s):  
Artashes R. Khachatryan ◽  
Thomas E. Besser ◽  
Douglas R. Call
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Resistance ◽  
Selective Advantage ◽  
Washington State ◽  
Resistance Pattern ◽  
Genetic Element ◽  
Dairy Calf ◽  
Genetic Traits ◽  
E Coli

ABSTRACT Association of specific antimicrobial resistance patterns with unrelated selective traits has long been implicated in the maintenance of antimicrobial resistance in a population. Previously we demonstrated that Escherichia coli strains with a specific resistance pattern (resistant to streptomycin, sulfadiazine, and tetracycline [SSuT]) have a selective advantage in dairy calf intestinal environments and in the presence of a milk supplement commonly fed to the calves. In the present study we identified the sequence of the genetic element that confers the SSuT phenotype and show that this element is present in a genetically diverse group of E. coli isolates, as assessed by macrorestriction digestion and pulsed-field gel electrophoresis. This element was also found in E. coli isolates from 18 different cattle farms in Washington State. Using in vitro competition experiments we further demonstrated that SSuT strains from 17 of 18 farms were able to outcompete pansusceptible strains. In a separate set of experiments, we were able to transfer the antimicrobial resistance phenotype by electroporation to a laboratory strain of E. coli (DH10B), making that new strain more competitive during in vitro competition with the parental DH10B strain. These data indicate that a relatively large genetic element conferring the SSuT phenotype is widely distributed in E. coli from cattle in Washington State. Furthermore, our results indicate that this element is responsible for maintenance of these traits owing to linkage to genetic traits that confer a selective advantage in the intestinal lumens of dairy calves.

scholarly journals Antimicrobial Resistance in Fecal Escherichia coli Isolates from Healthy Urban Children of Two Age Groups in Relation to Their Antibiotic Therapy

2011 ◽  
Vol 55 (6) ◽  
pp. 3005-3007 ◽  
Author(s):  
Ivan Literak ◽  
Radim Petro ◽  
Monika Dolejska ◽  
Erika Gruberova ◽  
Hana Dobiasova ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Czech Republic ◽  
Antimicrobial Resistance ◽  
Age Groups ◽  
Antibiotic Resistant ◽  
The Czech Republic ◽  
Content Type ◽  
E Coli ◽  
Significant Difference ◽  
Two Age Groups

ABSTRACTThe study was performed in the Czech Republic during 2007 to 2009. OfEscherichia coliisolates from 275 children aged 6 weeks, 36% (n= 177) were resistant to 1 to 7 antibiotics. Of isolates from 253 children aged 6 to 17 years, 24% (n= 205) were resistant to 1 to 5 antibiotics. There was no significant difference in the prevalences of antibiotic-resistantE. coliisolates between these groups of children, even though the consumptions of antibiotics were quite different.

scholarly journals Phytochemical Screening, Mathematical Analysis and Antimicrobial Activity of Methanolic Seed Extract of Hunteria Umbellata

2020 ◽  
pp. 1-17
Author(s):  
Oluwaseun Raphael Aderele ◽  
Adekunle Kareem Rasaq ◽  
Johnson Oshiobugie Momoh
Keyword(s):  
Antimicrobial Activity ◽  
Seed Extract ◽  
T Test ◽  
Diffusion Method ◽  
Phytochemical Analysis ◽  
In Vitro Antimicrobial Activity ◽  
E Coli ◽  
Significant Difference ◽  
Agar Well Diffusion Method

Aim: The study evaluates the in-vitro antimicrobial activity of Hunteria umbellata against Escherichia coli, Staphylococcus aureus and Streptococcus sp. Place and Duration of Study: The study was carried out for three months in 2019 in Biochemistry Laboratory, Department of Chemical Sciences (Biochemistry unit), School of Pure and Applied Sciences, Lagos State Polytechnic, Ikorodu, Lagos- Nigeria. Methodology: The qualitative and GC-MS analysis of Hunteria umbellata methanolic seed extract were determined using standard procedure. The antimicrobial activity was evaluated by the disc diffusion method and agar well diffusion method. The experimental data was resampled 1000 times to allow for higher degrees of freedom in carrying out t-test to test for the difference of the effect of in-vitro antimicrobial activity of H. umbellata against E. coli, S. aureus and Streptococcus sp using mathematical software R language (3.6.1 version). Line plots, histogram and t-test are used to explain the effect of antimicrobial activity of H. umbellate on the selected bacteria. MIC and MBC were determined using standard methods. Results: The Phytochemical analysis of methanolic seed extract of Hunteria umbellata showed the presence of secondary metabolites like saponins, tannins, flavonoids, steroids, phenol among others. GC-MS assay of the H. umbellata seed extract revealed the presence of eight different compounds. Agar well diffusion method was characterized by inhibition zones of 18.36±0.87, 19.13±1.03 and 21.62±2.53 mm for E.coli, S. aureus and Streptococcus sp respectively at 300 mg/ml-1 and 21.70± 1.60, 23.83± 2.64 and 28.57± 1.52 for E.coli, S. aureus and Streptococcus sp respectively at 500 mg/ml. The results of the analysis show that there is a significant difference between the effects of in-vitro antimicrobial activity of H. umbellate on 3001 and 500 mg/ml on each bacteria tested at 5% level of significance. E.coli, S. aureus and Streptococcus sp were tested against 12 standard antimicrobial agents, of which six was sensitive and another six was resistance to E .coli, seven was sensitive, and five was resistance to S. aureus while four was resistance and eight sensitive to Streptococcus sp. The minimum inhibitory concentration (MIC) for E.coli, S. aureus, and  Streptococcus sp were 250, 125 and 31.25 mgml-1 while their minimum bactericidal concentration (MBC) were 500, 250 and 125 respectively. MIC and MBC tests showed that H. umbellata methanolic seed extract had noticeable bactericidal effects with MBC/MIC values ranging between 2 to 4. The extract has strong potency against these microorganisms with Streptococcus sp being the most susceptible. Conclusions: Hunteria umbellata has potential as natural therapeutic agents against E. coli, S. aureus and Streptococcus sp and they may prevent pathogenic diseases.

scholarly journals In vitro Evaluationof the Antimi- crobial Activity of a Topical Skin Preparation Containing 0.1% Polyhexanide vs a Topical Skin Prepa-ration Containing 1% Silver Sulfadiazine

2021 ◽  
Vol 6 (2) ◽  
pp. 1-7
Author(s):  
Barbara Maglione ◽  
Keyword(s):  
Antibacterial Activity ◽  
Positive Control ◽  
Negative Control ◽  
Silver Sulfadiazine ◽  
Bacterial Strains ◽  
Skin Preparation ◽  
E Coli ◽  
Topical Cream ◽  
Significant Difference

Aim: The effective in vitro antibacterial activity on Staphylococcus aureus (S.aureus), Pseudomonas aeruginosa (P.aeruginosa), Klebsiella pneumoniae (K.pneumoniae),Escherichia coli (E.Coli) and the combination of S.aureus and K. pneumonia of a topical cream based on 0.1% polyhexanidewas compared to a topical cream based on 1% silver sulfadiazine.A topical cream containing 0,1% gentamicin was used as a positive control and a white blank topical cream was used as negative control. Materials and Methods: The in vitro antibacterial activities were determined by agar well-diffusion assay. Two-way Analysis of Variance (ANOVA) was used to test, by calculation of P-values, for significant antiseptic activity in bacteria treated with 0.1% polyhexanide topical cream compared to 1% silver sulfadiazine and to the negative and positive controls. Results: Among the derivatives tested, all the active topical creams analyzed were able to reduce microbial strains. The topical cream based on 0.1% polyhexanide showed a significantly higher antibacterial efficacy in comparison to the topical cream based on 1% silver sulfadiazine on S. aureus and K. pneumonia and on the combination of S. aureus and K. pneumoniae,while no significant difference was detected between the antibacterial activity of the two topical creams against P. aeruginosa and E. coli. Conclusion: These results provide a further insight into the antibacterial activity of polyhexanide and its non-inferiority compared to silver sulfadiazine towards certain bacterial strains (P. aeruginosa and E. coli) and superiority towards other (S. aureus and K. pneumoniae)and support the use of 0.1% Polyhexanide topical preparation for the treatment of wounds that are infected or at risk of infection.

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