Fungicide resistance: the dose rate debate

It makes economic sense to use as little fungicide as possible on a crop. In many settings, it is common to apply less than the manufacturer's recommended dose. If sources of disease are scarce, or conditions are unsuitable for it to increase, the reduced control from a low dose may be adequate. In other cases, a big reduction in dose may cause little reduction in control, again permitting savings - especially for growers prepared to run a little risk. But the label recommendations for most fungicides state that to avoid resistance, a full dose must always be used. Are individual cost-savings therefore endangering everyone's access to an exceptionally useful tool? The emergence of fungicide resistance is evolution in action. In all cases, it involves the genetic replacement of the original susceptible population of the pathogen by a new population with genetically distinct biochemistry, which confers resistance. The resistant biochemistry originates in rare genetic mutations, so rare that initially the population is hardly altered. Replacement of susceptible forms by resistant ones happens because, with fungicide present, the resistant form multiplies more rapidly than the susceptible form. The key point to notice is that only the relative rates of multiplication of the resistant and susceptible types are involved in the evolution of resistance. The absolute rates are irrelevant.

Effects of fungicide dose and mixtures on selection for triazole resistance in Mycosphaerella graminicola under field conditions

The effects of varying doses of fungicides, alone or in mixtures, on selection for triazole resistance were examined under field conditions. Two experiments were conducted using the triazole fungicide fluquinconazole with the strobilurin fungicide azoxystrobin as a mixture partner. Inoculated wheat plots with a known ratio of more sensitive to less sensitive isolates of the leaf blotch fungus Mycosphaerella graminicola were sprayed with fungicide and sampled once symptoms had appeared. Selection for fluquinconazole resistance increased in proportion to the dose, up to one-half of the full dose (the maximum tested) in both experiments. At the higher doses of fluquinconazole, the addition of azoxystrobin was associated with a decrease in selection (nonsignificant in the first experiment) for triazole resistance. Control by low doses of fluquinconazole was increased by mixture with azoxystrobin, but at higher doses mixture with azoxystrobin sometimes decreased control, so that reduced selection was obtained at the cost of some reduction in control. The effects on resistance are not necessarily general consequences of mixing fungicides, and suggest that the properties of any specific mixture may need to be demonstrated experimentally. Selection was inversely related to control in the unmixed treatments in both experiments, but the relationship was weaker in the mixtures with azoxystrobin.

Changes in oestrogen receptor-alpha and -beta during progression to acquired resistance to tamoxifen and fulvestrant (Faslodex, ICI 182,780) in MCF7 human breast cancer cells

Cell culture models of antioestrogen resistance often involve applying selective pressures of oestrogen deprivation simultaneously with addition of tamoxifen or fulvestrant (Faslodex, ICI 182,780) which makes it difficult to distinguish events in development of antioestrogen resistance from those in loss of response to oestrogen or other components. We describe here time courses of loss of antioestrogen response using either oestrogen-maintained or oestrogen-deprived MCF7 cells in which the only alteration to the culture medium was addition of 10(-6) M tamoxifen or 10(-7) M fulvestrant. In both oestrogen-maintained and oestrogen-deprived models, loss of growth response to tamoxifen was not associated with loss of response to fulvestrant. However, loss of growth response to fulvestrant was associated in both models with concomitant loss of growth response to tamoxifen. Measurement of oestrogen receptor alpha (ER alpha) and oestrogen receptor beta (ER beta) mRNA by real-time RT-PCR together with ER alpha and ER beta protein by Western immunoblotting revealed substantial changes to ER alpha levels but very little alteration to ER beta levels following development of antioestrogen resistance. In oestrogen-maintained cells, tamoxifen resistance was associated with raised levels of ERa mRNA/protein. However by contrast, in oestrogen-deprived MCF7 cells, where oestrogen deprivation alone had already resulted in increased levels of ERa mRNA/protein, long-term tamoxifen exposure now reduced ER alpha levels. Whilst long-term exposure to fulvestrant reduced ERa. mRNA/protein levels in the oestrogen-maintained cells to a level barely detectable by Western immunoblotting and non-functional in inducing gene expression (ERE-LUC reporter or pS2), in oestrogen-deprived cells the reduction was much less substantial and these cells retained an oestrogen-induction of both the ERE-LUC reporter gene and the endogenous pS2 gene which could still be inhibited by antioestrogen. This demonstrates that whilst ER alpha can be abrogated by fulvestrant and increased by tamoxifen in some circumstances, this does not always hold true and mechanisms other than alteration to ER must be involved in the development of antioestrogen resistant growth. (c) 2006 Elsevier Ltd. All rights reserved.

Gene-nutrient interactions in the metabolic syndrome: single nucleotide polymorphisms in ADIPOQ and ADIPOR1 interact with plasma saturated fatty acids to modulate insulin resistance

Background: Progression of the metabolic syndrome (MetS) is determined by genetic and environmental factors. Gene-environment interactions may be important in modulating the susceptibility to the development of MetS traits. Objective: Gene-nutrient interactions were examined in MetS subjects to determine interactions between single nucleotide polymorphisms (SNPs) in the adiponectin gene (ADIPOQ) and its receptors (ADIPOR1 and ADIPOR2) and plasma fatty acid composition and their effects on MetS characteristics. Design: Plasma fatty acid composition, insulin sensitivity, plasma adiponectin and lipid concentrations, and ADIPOQ, ADIPOR1, and ADIPOR2 SNP genotypes were determined in a cross-sectional analysis of 451 subjects with the MetS who participated in the LIPGENE (Diet, Genomics, and the Metabolic Syndrome: an Integrated Nutrition, Agro-food, Social, and Economic Analysis) dietary intervention study and were repeated in 1754 subjects from the LIPGENE-SU.VI.MAX (SUpplementation en VItamines et Mineraux AntioXydants) case-control study (http://www.ucd.ie/lipgene). Results: Single SNP effects were detected in the cohort. Triacylglycerols, nonesterified fatty acids, and waist circumference were significantly different between genotypes for 2 SNPs (rs266729 in ADIPOQ and rs10920533 in ADIPOR1). Minor allele homozygotes for both of these SNPs were identified as having degrees of insulin resistance, as measured by the homeostasis model assessment of insulin resistance, that were highly responsive to differences in plasma saturated fatty acids (SFAs). The SFA-dependent association between ADIPOR1 rs10920533 and insulin resistance was replicated in cases with MetS from a separate independent study, which was an association not present in controls. Conclusions: A reduction in plasma SFAs could be expected to lower insulin resistance in MetS subjects who are minor allele carriers of rs266729 in ADIPOQ and rs10920533 in ADIPOR1. Personalized dietary advice to decrease SFA consumption in these individuals may be recommended as a possible therapeutic measure to improve insulin sensitivity. This trial was registered at clinicaltrials.

The evolutionary ecology of resistance to parasitoids by Drosophila

Parasitoids are the most important natural enemies of many insect species. Larvae of many Drosophila species can defend themselves against attack by parasitoids through a cellular immune response called encapsulation. The paper reviews recent studies of the evolutionary biology and ecological genetics of resistance in Drosophila, concentrating on D. melanogaster. The physiological basis of encapsulation, and the genes known to interfere with resistance are briefly summarized. Evidence for within- and between-population genetic variation in resistance from isofemale line, artificial selection and classical genetic studies are reviewed. There is now firm evidence that resistance is costly to Drosophila, and the nature of this cost is discussed, and the possibility that it may involve a reduction in metabolic rate considered. Comparative data on encapsulation and metabolic rates across seven Drosophila species provides support for this hypothesis. Finally, the possible population and community ecological consequences of evolution in the levels of host resistance are examined.

Genetic predisposition to type 2 diabetes is associated with impaired insulin secretion but does not modify insulin resistance or secretion in response to an intervention to lower dietary saturated fat

Genome-wide association studies have identified SNPs reproducibly associated with type 2 diabetes (T2D). We examined the effect of genetic predisposition to T2D on insulin sensitivity and secretion using detailed phenotyping in overweight individuals with no diagnosis of T2D. Furthermore, we investigated whether this genetic predisposition modifies the responses in beta-cell function and insulin sensitivity to a 24-week dietary intervention. We genotyped 25 T2D-associated SNPs in 377 white participants from the RISCK study. Participants underwent an IVGTT prior to and following a dietary intervention that aimed to lower saturated fat intake by replacement with monounsaturated fat or carbohydrate. We composed a genetic predisposition score (T2D-GPS) by summing the T2D risk-increasing alleles of the 25 SNPs and tested for association with insulin secretion and sensitivity at baseline, and with the change in response to the dietary intervention. At baseline, a higher T2D-GPS was associated with lower acute insulin secretion (AIRg 4% lower/risk allele, P = 0.006) and lower insulin secretion for a given level of insulin sensitivity, assessed by the disposition index (DI 5% lower/risk allele, P = 0.002), but not with insulin sensitivity (Si). T2D-GPS did not modify changes in insulin secretion, insulin sensitivity or the disposition index in response to the dietary interventions to lower saturated fat. Participants genetically predisposed to T2D have an impaired ability to compensate for peripheral insulin resistance with insulin secretion at baseline, but this does not modify the response to a reduction in dietary saturated fat through iso-energetic replacement with carbohydrate or monounsaturated fat.

Characterisation of the intracellular-glutamate decarboxylase system (GAD): analysis of its function, transcription and role in the acid resistance of various strains of Listeria monocytogenes

The glutamate decarboxylase (GAD) system is important for the acid resistance of Listeria monocytogenes. We previously showed that under acidic conditions, glutamate (Glt)/γ-aminobutyrate (GABA) antiport is impaired in minimal media but not in rich ones, like brain heart infusion. Here we demonstrate that this behavior is more complex and it is subject to strain and medium variation. Despite the impaired Glt/GABA antiport, cells accumulate intracellular GABA (GABA(i)) as a standard response against acid in any medium, and this occurs in all strains tested. Since these systems can occur independently of one another, we refer to them as the extracellular (GAD(e)) and intracellular (GAD(i)) systems. We show here that GAD(i) contributes to acid resistance since in a ΔgadD1D2 mutant, reduced GABA(i) accumulation coincided with a 3.2-log-unit reduction in survival at pH 3.0 compared to that of wild-type strain LO28. Among 20 different strains, the GAD(i) system was found to remove 23.11% ± 18.87% of the protons removed by the overall GAD system. Furthermore, the GAD(i) system is activated at milder pH values (4.5 to 5.0) than the GAD(e) system (pH 4.0 to 4.5), suggesting that GAD(i) is the more responsive of the two and the first line of defense against acid. Through functional genomics, we found a major role for GadD2 in the function of GAD(i), while that of GadD1 was minor. Furthermore, the transcription of the gad genes in three common reference strains (10403S, LO28, and EGD-e) during an acid challenge correlated well with their relative acid sensitivity. No transcriptional upregulation of the gadT2D2 operon, which is the most important component of the GAD system, was observed, while gadD3 transcription was the highest among all gad genes in all strains. In this study, we present a revised model for the function of the GAD system and highlight the important role of GAD(i) in the acid resistance of L. monocytogenes.

Modification of enrofloxacin treatment regimens for poultry experimentally infected with Salmonella enterica serovar Typhimurium DT104 to minimize selection of resistance

We hypothesized that higher doses of fluoroquinolones for a shorter duration could maintain efficacy (as measured by reduction in bacterial count) while reducing selection in chickens of bacteria with reduced susceptibility. Chicks were infected with Salmonella enterica serovar Typhimurium DT104 and treated 1 week later with enrofloxacin at the recommended dose for 5 days (water dose adjusted to give 10 mg/kg of body weight of birds or equivalence, i.e., water at 50 ppm) or at 2.5 or 5 times the recommended dose for 2 days or 1 day, respectively. The dose was delivered continuously (ppm) or pulsed in the water (mg/kg) or by gavage (mg/kg). In vitro in sera, increasing concentrations of 0.5 to 8 mu g/ml enrofloxacin correlated with increased activity. In vivo, the efficacy of the 1-day treatment was significantly less than that of the 2- and 5-day treatments. The 2-day treatments showed efficacy similar to that of the 5-day treatment in all but one repeat treatment group and significantly (P < 0.01) reduced the Salmonella counts. Dosing at 2.5x the recommended dose and pulsed dosing both increased the peak antibiotic concentrations in cecal contents, liver, lung, and sera as determined by high-pressure liquid chromatography. There was limited evidence that shorter treatment regimens (in particular the 1-day regimen) selected for fewer strains with reduced susceptibility. In conclusion, the 2-day treatment would overall require a shorter withholding time than the 5-day treatment and, in view of the increased peak antibiotic concentrations, may give rise to improved efficacy, in particular for treating respiratory and systemic infections. However, it would be necessary to validate the 2-day regimen in a field situation and in particular against respiratory and systemic infections to validate or refute this hypothesis.

Modelling the resistance of Enterprise Architecture adoption: linking strategic level of Enterprise Architecture to organisational changes and change resistance

During the last few years Enterprise Architecture (EA) has received increasing attention among industry and academia. By adopting EA, organisations may gain a number of benefits such as better decision making,increased revenues and cost reduction, and alignment of business and IT. However, EA adoption has been found to be difficult. In this paper a model to explain resistance during EA adoption process (REAP) is introduced and validated. The model reveals relationships between strategic level of EA, resulting organisational changes, and sources of resistance. By utilising REAP model, organisations may anticipate and prepare for the organisational change resistance during EA adoption.

Fungicide-resistance management tactics: impacts on Zymoseptoria tritici populations

Azoles and Succinate Dehydrogenase Inhibitors (SDHIs) are the main fungicides available for septoria tritici blotch control, causal agent Zymoseptoria tritici. Decline in azole sensitivity, in combination with European legislation, poses a threat to wheat production in Ireland. Azole fungicides select CYP51 mutations differentially; it was hypothesised that using combinations of azoles could be an effective anti-resistance tool. Naturally inoculated field experiments were carried out in order to understand the impacts of using combinations of azoles, epoxiconazole and metconazole, on azole sensitivity. Approximately 3700 isolates were isolated and their sensitivity to both azoles analysed. Findings showed that limiting the number of applications, by alternating each fungicide, slowed selection for reduced azole sensitivity. Limiting azole use by reducing doses did not reduce selection for decreased azole sensitivity. Although not complete, cross-resistance was observed between the two azoles, which will lead to general reduction in azole sensitivity. A sub-selection of isolates from each treatment at each location were analysed for changes in the CYP51 gene. Sequence analysis identified 49 combinations of mutations in the CYP51 gene, and three different inserts in the CYP51 promoter. Intragenic recombination also featured in these populations. Baseline studies of five new SDHIs were carried out on 209 naturally infected, non-SDHI-treated isolates. With the exception of fluopyram, cross-resistance was apparent between the SDHIs. Analysis of 2300 isolates found that when compared to the solo products, mixing the SDHI isopyrazam and the azole epoxiconazole increased epoxiconazole sensitivity, but had no apparent effect on isopyrazam sensitivity. SDHI resistance-conferring mutations were absent in the baseline and experimental isolates. As long as azoles are used, Z. tritici populations will continue to evolve towards resistance. Combining different modes-of-action, SDHIs and multi-sites, with azoles will relieve some of that selective pressure. To get the best out of available fungicides, they should be used in combination with host resistance and good crop management practices.