Editorial – Avoiding Unethical Helicobacter pylori Clinical Trials: Susceptibility‐Based Studies and Probiotics as Adjuvants

As a general rule, any clinical study where the result is already known or when the investigator(s) compares an assigned treatment against another assigned treatment known to be ineffective in the study population (e.g., in a population with known clarithromycin resistance) is unethical. As susceptibility‐based therapy will always be superior to empiric therapy in any population with a prevalence of antimicrobial resistance >0%, any trial that randomizes susceptibility‐based therapy with empiric therapy would be unethical. The journal Helicobacter welcomes susceptibility or culture‐guided studies, studies of new therapies, and studies of adjuvants and probiotics. However, the journal will not accept for review any study we judge to be lacking clinical equipoise or which assign subjects to a treatment known to be ineffective, such as a susceptibility‐based clinical trial with an empiric therapy comparator. To assist authors, we provide examples and suggestions regarding trial design for comparative studies, for susceptibility‐based studies, and for studies testing adjuvants or probiotics.     

Re-Evaluation of the Critical Concentration for Ethambutol Antimicrobial Sensitivity Testing on the MGIT 960

The critical concentration (CC) for ethambutol testing on the Bactec MGIT 960 M. tuberculosis susceptibility testing has been questioned in recent publications. In this study, we correlate susceptibility results from the Bactec 460, MGIT 960 and embB gene sequencing to determine if the Bactec MGIT 960 adequately detects ethambutol resistance. We discovered discrepancies between the methods that highlight a need to re-evaluate ethambutol susceptibility testing recommendations, namely by considering lowering currently recommended CC on the MGIT 960. Further studies on the clinical significance of low-level ethambutol resistance are also required.     

Temperature checks the Red Queen? Resistance and virulence in a fluctuating environment

Numerous studies have revealed genetic variation in resistance and susceptibility in host–parasite interactions and therefore the potential for frequency‐dependent selection (Red Queen dynamics). Few studies, if any, have considered the abiotic environment as a mediating factor in these interactions. Using the pea aphid, Acyrthosiphon pisum, and its fungal pathogen, Erynia neoaphidis, as a model host–parasite system, we demonstrate how temperature can mediate the expression of genotypic variation for susceptibility and virulence. Whilst previous studies have revealed among‐clone variation in aphid resistance to this pathogen, we show that resistance rankings derived from assessments at one temperature, are not conserved across differing temperature regimes. We suggest that variation in environmental temperature, through its nonlinear impact on parasite virulence and host defence, may contribute to the general lack of evidence for frequency‐dependent selection in field systems.     

The Use of Machine Learning Methodologies to Analyse Antibiotic and Biocide Susceptibility in Staphylococcus aureus

Background

The rise of antibiotic resistance in pathogenic bacteria is a significant problem for the treatment of infectious diseases. Resistance is usually selected by the antibiotic itself; however, biocides might also co-select for resistance to antibiotics. Although resistance to biocides is poorly defined, different in vitro studies have shown that mutants presenting low susceptibility to biocides also have reduced susceptibility to antibiotics. However, studies with natural bacterial isolates are more limited and there are no clear conclusions as to whether the use of biocides results in the development of multidrug resistant bacteria.

Methods

The main goal is to perform an unbiased blind-based evaluation of the relationship between antibiotic and biocide reduced susceptibility in natural isolates of Staphylococcus aureus. One of the largest data sets ever studied comprising 1632 human clinical isolates of S. aureus originated worldwide was analysed. The phenotypic characterization of 13 antibiotics and 4 biocides was performed for all the strains. Complex links between reduced susceptibility to biocides and antibiotics are difficult to elucidate using the standard statistical approaches in phenotypic data. Therefore, machine learning techniques were applied to explore the data.

Results

In this pioneer study, we demonstrated that reduced susceptibility to two common biocides, chlorhexidine and benzalkonium chloride, which belong to different structural families, is associated to multidrug resistance. We have consistently found that a minimum inhibitory concentration greater than 2 mg/L for both biocides is related to antibiotic non-susceptibility in S. aureus.

Conclusions

Two important results emerged from our work, one methodological and one other with relevance in the field of antibiotic resistance. We could not conclude on whether the use of antibiotics selects for biocide resistance or vice versa. However, the observation of association between multiple resistance and two biocides commonly used may be of concern for the treatment of infectious diseases in the future.     

Behaviour of Apricot Cultivars Against Hop Stunt Viroid

The apricot species is susceptible to infection by different viruses and viroids including the Hop stunt viroid (HSVd), widely distributed around the world and described as the causal agent associated with fruit degeneration disease in apricot. However, to date, there are no ‘ad hoc’ studies about the resistance or susceptibility of apricot cultivars to this viroid. In this study, we tested the resistance/susceptibility to HSVd of 26 Mediterranean and North American apricot cultivars in controlled greenhouse conditions. All apricot cultivars assayed were infected by HSVd, showing a different range of susceptibility. As no sources of resistance among the apricot cultivars evaluated have been detected yet, it is necessary to continue the search for such sources to be included in apricot breeding programmes. In addition, the high level of susceptibility found highlights the importance of identifying this viroid in routine tests performed by nurseries and plant protection services.     

Susceptibility of fruit from diverse apple and crabapple germplasm to attack by plum curculio (Coleoptera: Curculionidae)

Plum curculio, Conotrachelus nenuphar (Herbst) (Coleoptera: Curculionidae), is an important apple, Malus domestica Borkh., pest that significantly hinders sustainable apple production in eastern North America. The potential for host plant resistance to plum curculio among apple germplasm has never been rigorously evaluated. Thus, studies were conducted to assess the susceptibility of a number of exotic and domestic Malus accessions housed at the USDA Plant Genetic Resources Unit (PGRU) "core" collection in Geneva, NY. Contrary to earlier published reports and promising data from a field assessment in 2005, these results suggest that there is probably little potential for genetic resistance to plum curculio among the Malus germplasm collection evaluated. More specifically, four Malus hybrid selections that have previously been released with claims of plum curculio resistance were shown to be susceptible to plum curculio attack. Because there are additional accessions housed at PGRU outside of the core collection that are currently classified as resistant, further studies are necessary to evaluate the true resistance qualities of these releases. It is also important to clarify such discrepancies in both the USDA online Germplasm Resources Information Network and in the horticultural literature. Although other Malus species exhibited some variability in fruit susceptibility, none could be classified as being truly resistant to plum curculio attack by any definition that would have relevance to commercial production and sale of apples.     

Antimicrobial susceptibility changes and T-OMP shifts in pyrithione-passaged planktonic cultures of Pseudomonas aeruginosa PAO1

AIMS: The aim of this study was to determine whether passaging Pseudomonas aeruginosa PAO1 with sub-MICs of the pyrithione biocides results in both the induction of decreased susceptibility towards these antimicrobials and associated outer membrane profile changes. METHODS AND RESULTS: Previous work by this group has shown that it is possible to induce susceptibility changes towards the isothiazolone biocides in Ps. aeruginosa PAO1 by successive passages in the presence of increasing sub-MICs of biocide. This procedure was accompanied by the loss of a 35 kDa outer membrane protein, T-OMP. In this experiment, this process was repeated with the biocides sodium pyrithione (NaPT), zinc pyrithione (ZnPT) and cetrimide. The pattern of susceptibility was similar to that observed with the isothiazolone biocides. Upon removal of biocide, the observed MIC did not return to the original pre-exposure value. The onset and development of resistance was accompanied by the loss of T-OMP from outer membrane profiles, which suggests that this is a non-specific membrane channel whose production within the cell is sensitive to biocide presence. The T-OMP reappeared when the cells were passaged in the absence of pyrithione. Cross-resistance studies indicated that induced resistance to one biocide yields partial resistance towards other members of the group and the positive control. CONCLUSIONS: These results indicate that the pyrithione biocides have similar susceptibility profiles in Ps. aeruginosa to those exhibited by the isothiazolones, but that the acquired changes in susceptibility to the pyrithiones is largely irreversible. SIGNIFICANCE AND IMPACT OF THE STUDY: This study indicates that acquired susceptibility changes towards sub-MICs of selected biocides are multifactorial in nature.     

Monitoring western corn rootworm (Coleoptera: Chrysomelidae) susceptibility to carbaryl and cucurbitacin baits in the areawide management pilot program

Areawide pest management involves the uniform application of a pest control strategy over wide geographic areas. Therefore, these programs are likely to impose intense selective pressures, and the risk for resistance development among pest species for which areawide management programs are implemented is likely to be high. Pilot studies for areawide management of western corn rootworm, Diabrotica virgifera virgifera LeConte, were conducted from 1996 to 2002 at four different sites across the Corn Belt. This program used cucurbitacin baits to deliver high doses of a traditional neurotoxic insecticide (carbaryl) to individual insects while reducing the overall rate of insecticide use. Because of the concern and potential for resistance evolution, annual assessments of susceptibility to the active ingredient carbaryl were conducted both within the managed area as well as from untreated control areas. Significantly reduced susceptibility to carbaryl based on survival at a diagnostic concentration was detected in three of the four management sites (Kansas, Iowa, and Illinois/Indiana), whereas susceptibility of beetles collected outside the managed areas remained unchanged. Additionally, significantly reduced responsiveness to cucurbitacin baits was observed in beetles collected from the managed area relative to the control area at the same three sites. These results suggest strongly that areawide management has the potential to select for resistance and that a strategy for managing resistance and reducing selective pressure should be proactively implemented.     

The genetic basis of quantitative variation in susceptibility of Arabidopsis thaliana to Pseudomonas syringae (Pst DC3000): evidence for a new genetic factor of large effect

* Pathogens represent an important threat to plant communities and agriculture, and can shape many aspects of plant evolution. Natural variation in plant disease susceptibility is typically quantitative, yet studies on the molecular basis of disease resistance have focused mainly on qualitative variation. * Here we investigated the genetic architecture of quantitative susceptibility to the bacterium Pseudomonas syringae by performing a quantitative trait locus (QTL) analysis on the F2 progeny of two natural accessions of Arabidopsis thaliana under two nutrient treatments. * We found that a single QTL explains most of the variation (77%) in susceptibility between accessions Columbia (Col-0) and San Feliu-2 (Sf-2), and its effect is independent of nutrients. The Sf-2 allele at this QTL is dominant and can reduce the bacterial population size by 31-fold, much like a classical resistance (R) gene. However, minor QTLs, whose effects are altered by nutrient treatment, were also detected. * Surprisingly, we found that none of the QTLs for susceptibility had any effect on fruit production, suggesting that the use of resistance genes for crop improvement and evolutionary analysis of plant-pathogen interactions requires caution.     

Naturally occurring drug resistance plasmids in hospital strains of Staphylococcus aureus

A genetic analysis of multiply inorganic salts and antibiotic--resistant strains of Staphylococcus aureus was performed. Experiments designed to show reversion of organisms to antibiotic and inorganic salt susceptibility, as well as studies on the influence of ultraviolet irradiation of phage on the transduction frequencies of the resistance markers, indicated that determinants of chloramphenicol, tetracycline, aminoglycoside antibiotics, inorganic salts, and penicillin resistance in hospital strain are present on separate plasmids. Transduced by us plasmids pN742 and pN794 determined resistance to neomycin, kanamycin, paromomycin, lividomycin and streptomycin.     

森林生态系统中植食性昆虫与寄主的互作机制、假说与证据

围绕"多样性稳定性"假说、"联合抗性假说"、"生长势假说"、"胁迫假说"、以及下调、上调和推拉等机制与假说提出的背景与实验验证的证据,力图辨析其概念以及它们之间的相互关系。作者认为,多样性-稳定性机制关注森林生态系统的功能,是基于群落甚至景观层次。多样性条件下的联合抗性机制和联合易感性应属于稳定性中的抵抗力范畴。联合抗性机制的主要基础是基于资源集中假说和天敌假说,这些观点在种群层次上更易理解;上调力和下调力机制是以食物网底部的资源与顶端的天敌来探讨这种互作关系。因此,资源集中与上调力有着对应关系,而天敌假说只是下调力机制中的一个层面而已。植物生长势假说和植物胁迫假说力图从植物个体或种的群体的生长状态出发解析植食性动物的对寄主的选择趋势。上述有关植食性昆虫与寄主互作的机制、假说与证据是基于不同的层面提出的,因而在解析研究目标时,由于基本面的差异有可能会得出不同的结论。以近年来的研究进展和研究成果为依据有针对性地阐述这些理论对森林有害生物生态调控技术的指导作用,其中,联合抗性和联合易感性理论对指导森林有害生物生态控制具有更直接的指导作用。进一步提出了相应的亟待解决的科学问题。     

Usefulness of marker‐assisted selection to improve maize for increased resistance to Sesamia nonagrioides attack with no detrimental effect on yield

Two decades of investigations on maize resistance to Mediterranean corn borer (Sesamia nonagrioides Lefebvre; MCB) have shown that breeding for increased resistance to stem tunnelling by MCB often resulted in reduced yield because significant genetic correlation between both traits exists in some backgrounds. Unlike phenotypic selection, marker‐assisted selection (MAS) could differentiate markers linked only to one trait from those linked simultaneously to yield potential and susceptibility to the pest. In the current study, the suitability of MAS for improving resistance to stem tunnelling without adverse effects on yield has been tested. The unfavourable genetic relationship between yield potential and susceptibility could be overcome using MAS. Gains obtained using MAS were weak, because genetic variance explained by the quantitative trait loci (QTL) was low but results encourage us to persevere in using marker information for simultaneous improvement of resistance and yield especially if genome‐wide approaches are applied. Approaches to detect QTL are widely used, but studies on the suitability of markers linked to QTL for performing MAS have been mostly neglected.     

Mutational fitness landscape and drug resistance

Robust technology has been developed to systematically quantify fitness landscapes that provide valuable opportunities to improve our understanding of drug resistance and define new avenues to develop drugs with reduced resistance susceptibility. We outline the critical importance of drug resistance studies and the potential for fitness landscape approaches to contribute to this effort. We describe the major technical advancements in mutational scanning, which is the primary approach used to quantify protein fitness landscapes. There are many complex steps to consider in planning and executing mutational scanning projects including developing a selection scheme, generating mutant libraries, tracking the frequency of variants using next-generation sequencing, and processing and interpreting the data. Key experimental parameters impacting each of these steps are discussed to aid in planning fitness landscape studies. There is a strong need for improved understanding of drug resistance, and fitness landscapes provide a promising new approach.     

Evolutionary determinants of genetic variation in susceptibility to infectious diseases in humans

Although genetic variation among humans in their susceptibility to infectious diseases has long been appreciated, little focus has been devoted to identifying patterns in levels of variation in susceptibility to different diseases. Levels of genetic variation in susceptibility associated with 40 human infectious diseases were assessed by a survey of studies on both pedigree-based quantitative variation, as well as studies on different classes of marker alleles. These estimates were correlated with pathogen traits, epidemiological characteristics, and effectiveness of the human immune response. The strongest predictors of levels of genetic variation in susceptibility were disease characteristics negatively associated with immune effectiveness. High levels of genetic variation were associated with diseases with long infectious periods and for which vaccine development attempts have been unsuccessful. These findings are consistent with predictions based on theoretical models incorporating fitness costs associated with the different types of resistance mechanisms. An appreciation of these observed patterns will be a valuable tool in directing future research given that genetic variation in disease susceptibility has large implications for vaccine development and epidemiology.     

The timing of induced resistance and induced susceptibility in the soybean-Mexican bean beetle system

Induced plant responses to herbivory have been demonstrated in many systems. It has been suggested that the timing of these responses may influence the impact of induced resistance on herbivore populations, and may affect the evolution of induced defenses. This study used a bioassay to characterize the time course of systemic induced responses to Mexican bean beetle herbivory in four genotypes of soybeans. The results suggest that the time course of induced responses in this system is more complex than most previous studies have indicated. Herbivory provoked both rapid induced resistance and subsequent induced susceptibility to beetle feeding. All four genotypes of soybean induced significant resistance to beetle damage (beetles preferred undamaged to damaged plants) by 3 days after damage. By 15 days after damage, this resistance had decayed (beetles showed no preference for undamaged over damaged plants), and by 20 days after damage, all four genotypes exhibited significant induced susceptibility (beetles preferred previously damaged plants over undamaged plants). The magnitude of induced resistance in each genotype correlated strongly with the magnitude of induced susceptibility in that genotype. Received: 28 September 1997 / Accepted: 1 December 1997     

A review on biocide reduced susceptibility due to plasmid-borne antiseptic-resistant genes—special notes on pharmaceutical environmental isolates

Biocides (antiseptics and disinfectants) are widely used in hospitals and pharmaceutical industries for contamination control. The emergence of reduced susceptibility to biocides is the major concern and this is caused by various factors, among which plasmid-mediated resistance is common. Many publications describe the antibiotic resistance and mechanisms in a clinical setting. However, there are only limited studies available worldwide addressing the molecular mechanisms of biocide resistance in the pharmaceutical sector. In addition, there is a considerable lack of scientific reports regarding minimum inhibitory concentration (MIC) values of typical biocides against pharmaceutical cleanroom environmental isolates. This review analyses the plasmid-mediated resistance in typical pharmaceutical micro-organisms and prevalence of biocide-resistant genes among common clinical and pharmaceutical isolates. This review discusses the MIC values of biocides in pharmaceutical environmental isolates, indicating the importance of the correlation between the presence or absence of biocide-resistant genes and reduced susceptibility of MIC values. This review recommends that pharmaceutical organizations adopt policies and test methodologies to examine the MICs of common cleanroom biocides against the most common types of cleanroom environmental isolates.     

Genotypic vs. condition effects on parasite-driven rare advantage

Models and empirical studies of coevolution assume host resistance and parasite infectivity are genetically based. However, nongenetic physiological or environmental influences could alter host susceptibility even when the relationship is genetically based. In this experiment we examined the influence of host genotype, host condition at the time of infection (age and reproductive status), and their interaction on resistance of the freshwater snail Potamopyrgus antipodarum) to its dominant trematode parasite (Microphallus sp.). We used a laboratory infection experiment of a clonal snail population to determine the susceptibility of juveniles, brooding adult females, and nonbrooding adult females. We found a significant effect of both life-history state and clonal genotype on the prevalence of infection. However, the relative susceptibility of different clonal genotypes was not altered by condition; genotypes that were rare in the natural population were less infected than those that were common for each life-history state. These results suggest that although host condition affects susceptibility, it does not disrupt the specificity of the match between parasites and common clonal genotypes. Hence these findings support the Red Queen hypothesis for the maintenance of sex under genetically based host-parasite interactions.     

Seasonal fluctuation in susceptibility to insecticides within natural populations of Drosophila melanogaster. II. Features of genetic variation in susceptibility to organophosphate insecticides within natural populations of D. melanogaster

To elucidate genetic variation in susceptibility to organophosphate insecticides within natural populations of Drosophila melanogaster, we conducted an analysis of variance for mortality data sets of isofemale lines (10-286 lines) used in the previous studies. Susceptibility of isofemale lines to the three organophosphate insecticides was continuously distributed within each natural population, ranging from susceptible to resistant. Analysis of variance showed highly significant variation among isofemale lines in susceptibility to each insecticide for each natural population. Significant genetic variances in susceptibility to the three chemicals were estimated for the Katsunuma population; 0.0529-0.2722 for malathion, 0.0492-0.1603 for prothiophos, and 0.0469-0.1696 for fenitrothion. Contrary to the consistent seasonal tendency towards an increase in mean susceptibility in the fall, reported in the previous study, genetic variances in susceptibility to the three organophosphates did not change significantly in 1997 but tended to increase by 2- to 5-times in 1998. We tested whether both the observed situations, maintenance and increase in genetic variance in organophosphate resistance, can be generated under circumstances in which the levels of resistance to the three organophosphates tended to decrease, by conducting a simulation analysis, based on the hypothesis that resistant genotypes have lower fitnesses than susceptible ones under the density-independent condition. The simulation analysis generally explained the pattern in the mean susceptibility and genetic variances in susceptibility to the three organophosphates, observed in the Katsunuma population of D. melanogaster. It was suggested that the differences in the frequencies of resistance genes in the summer population could affect the patterns in genetic variance in organophosphate resistance in the fall population.     

Quantitative resistance against Bemisia tabaci in Solanum pennellii: Genetics and metabolomics

The whitefly Bemisia tabaci is a serious threat in tomato cultivation worldwide as all varieties grown today are highly susceptible to this devastating herbivorous insect.Many accessions of the tomato wild relative Solanum pennellii show a high resistance towards B. tabaci. A mapping approach was used to elucidate the genetic background of whiteflyresistance related traits and associated biochemical traits in this species. Minor quantitative trait loci(QTLs) for whitefly adult survival(AS) and oviposition rate(OR) were identified and some were confirmed in an F2BC1 population, where they showed increased percentages of explained variance(more than 30%). Bulked segregant analyses on pools of whiteflyresistant and-susceptible F2 plants enabled the identification of metabolites that correlate either with resistance or susceptibility. Genetic mapping of these metabolites showed that a large number of them co-localize with whiteflyresistance QTLs. Some of these whitefly-resistance QTLs are hotspots for metabolite QTLs. Although a large number of metabolite QTLs correlated to whitefly resistance or susceptibility, most of them are yet unknown compounds and further studies are needed to identify the metabolic pathways and genes involved. The results indicate a direct genetic correlation between biochemical-based resistance characteristics and reduced whitefly incidence in S. pennellii.     

Susceptibility of antibiotic-resistant cocci to biocides

Biocide resistance has hitherto been a poorly studied subject, possibly due to the belief that such resistance was rare and clinically insignificant. Various recent findings, however, have underlined the importance of biocide resistance as a clinically relevant phenomenon. Outbreaks of biocide-resistant organisms in hospitals have been described and the genetic mechanism for resistance to quaternary ammonium compounds (QACs) in Staphylococcus aureus has now been elucidated. Mycobacteria resistant to commonly used endoscope disinfectants are now commonly reported and have caused numerous adverse clinical events. Cross-resistance between triclosan and antituberculous drugs has been demonstrated in other strains of mycobacteria. This is related to a common mechanism of action. The work presented here describes studies into the biocide resistance of antibiotic-resistant cocci and attempts to create biocide-resistant strains in vitro. Strains of staphylococci (including methicillin-resistant Staph. aureus (MRSA)) and enterococci (including vancomycin-resistant enterococci (VRE)) had their susceptibility to biocides assayed using broth macro dilution methods and resistant strains were selected by serial subculture on biocide-containing media. Mutants were created with relative ease; for instance, triclosan minimal bactericidal concentrations (MBCs) increased from 0.002 to 3.12 mg l(-1). Some strains of MRSA which have intermediate resistance to glycopeptides were demonstrated to have decreased susceptibility to some biocides. Biocide resistance amongst enterococci was demonstrated although there was no clear correlation between biocide and antibiotic resistance. The exact mechanisms of resistance in these strains are still being studied but it is clear that biocide resistance is an important clinical phenomenon.