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Meloidogyne spp. are the most devastating plant‐parasitic nematodes affecting tomato worldwide. Although resistant cultivars and rootstocks are used, selection for virulence occurs in the pathogen. Consequently, using other resistance sources, such as Solanumtorvum, could improve resistance durability. Several experiments in microplots and plastic greenhouses were carried out to determine the potential use of S. torvum as a tomato rootstock to protect against M. incognita and M. javanica. In microplots, the relationship between nematode density at transplanting (Pi) and multiplication rate did not differ between Meloidogyne species in either ungrafted or grafted tomato. However, maximum multiplication rate and maximum density on grafted tomato were 1.27% and 2.93% those on ungrafted, respectively. The grafted tomato plants yielded between 2.9 and 7.5 more times than the ungrafted plants at Pi ≥ 100 eggs + J2s per 100 cm3 of soil, but no differences were observed in plastic greenhouse where a large amount of scion‐rooting occurred. In microplots, the quality of the tomato fruits of ungrafted and grafted plants was affected by the Pi. In parallel, some pot experiments were conducted on S. torvum and susceptible eggplant to determine the putative selection for nematode virulence to S. torvum and the nematode fitness cost. These showed that the nematode subpopulations infected and reproduced less on S. torvum than on eggplant. However, the female fertility was only reduced after development of three or four subpopulations on S. torvum. Finally, a histopathological study showed that nematode infection and development in S. torvum was delayed compared to eggplant. [ABSTRACT FROM AUTHOR]

Bacterial conjugation, wherein DNA is transferred between cells through direct contact, is highly prevalent in complex microbial communities and is responsible for spreading myriad genes related to human and environmental health. Despite their importance, much remains unknown regarding the mechanisms driving the spread and persistence of these plasmids in situ. Studies have demonstrated that transferring, acquiring, and maintaining a plasmid imposes a significant metabolic burden on the host. Simultaneously, emerging evidence suggests that the presence of a conjugative plasmid can also provide both obvious and unexpected benefits to their host and local community. Combined, this highlights a continuous cost‐benefit tradeoff at the population level, likely contributing to overall plasmid abundance and long‐term persistence. Yet, while the metabolic burdens of plasmid conjugation, and their causes, are widely studied, their attendant potential advantages are less clear. Here, we summarize current perspectives on conjugative plasmids’ metabolic burden and then highlight the lesser‐appreciated yet critical benefits that plasmid‐mediated metabolic burdens may provide. We argue that this largely unexplored tradeoff is critical to both a fundamental theory of microbial populations and engineering applications and therefore warrants further detailed study. [ABSTRACT FROM AUTHOR]

Simple Summary: As an invasive pest, the fall webworm has caused huge economic losses in agricultural and forestry production, and the development of resistance in the fall webworm to Cry1Ab toxin is an important issue in plant protection. Our study shows that there is no fitness cost for the resistance of the fall webworm to Cry1A toxin, which comes from autosomal recessive inheritance and does not lead to cross-resistance to Cry2Ab toxin. According to the results of this study, transgenic poplars expressing multiple insecticidal proteins should be used directly and shelters should be established to control the fall webworm more effectively and delay the development of resistance. Transgenic poplars have been used to control quarantine pests worldwide, such as the fall webworm (Hyphantria cunea, FW). However, the studies on the resistance mechanism of FW to Cry toxins are limited. This study obtained an FW strain with 45-fold resistance to Cry1Ab toxin by continuous screening in the laboratory. The resistance to Cry1Ab was autosomally completely recessive, and it had approximately 40-fold cross-resistance to Cry1Ac but no cross-resistance to Cry2Ab toxin. Analysis of indoor population life tables for the resistant and susceptible strains found no significant difference in the net reproduction rate (R0) between the two strains. The resistant strain had an insignificantly longer larval development duration and a comparable pupation rate, pupal weight, emergence rate, eggs laidper female, and egg hatching rate compared to the susceptible strain. This indicated that there was no fitness cost for Cry1Ab resistance. Our study helps in evaluating the risk of resistance to Cry1Ab toxin and the mechanisms of resistance to Cry1Ab toxin in FW. [ABSTRACT FROM AUTHOR]

Life‐history traits and population size of organisms may be affected by fitness costs associated with insecticide resistance. In this study, we examined the insecticide resistance and life‐history traits of Paederus fuscipes (Curtis), a common generalist predator, in field populations from Taiwan (high‐latitude region; 22° N–25° N) and Malaysia (low‐latitude region; 3° N–5° N). The results showed that Malaysian populations had higher imidacloprid resistance compared with Taiwanese populations. The fitness costs associated with imidacloprid resistance resulted in reduced fecundity, longer adult pre‐oviposition period, shorter egg length, and lighter pupal weight in the Malaysian populations, in contrast to the Taiwanese populations. However, the immature developmental time of the Malaysian populations was relatively short compared with the Taiwanese populations. The longevity of the Taiwanese populations was significantly shorter compared with the Malaysian populations. The fecundity of the adult populations increased in inverse proportion to longevity. The intrinsic rate of increase was significantly lower in the Malaysian populations compared with the Taiwanese populations. This regional‐scale study provided insights into the differences in life‐history traits and population growth of P. fuscipes as a result of fitness cost. However, it is worth noting that despite the high intrinsic rate of increase observed in the Taiwanese populations, among other factors, cannibalism may serve as a density‐dependent compensatory mechanism in regulating population size. [ABSTRACT FROM AUTHOR]

Background: The pink bollworm, Pectinophora gossypiella (Saunders), is a devastating global pest of cotton that has caused substantial economic damage to Bt Bollgard-II® cotton plants in recent years due to the evolution of Bt resistance. The associated fitness cost is assumed to be one of the factors delaying the development of resistance against Bt transgenic crops. Hence, the present study was undertaken to assess the biological performance of pink bollworms by comparing the life history and demographic parameters of a resistant (Field-R) and susceptible (Lab-S) population. Results: Prolonged larval duration (23.40 days in Field-R vs 18.80 days in Lab-S population), total life cycle (male = 50.00 vs 42.80 days; female = 53.60 vs 46.20 days), reduced fecundity (100.60 vs 154.20 eggs/female) and fertility (88.00 vs 138.00 fertile eggs/female) was observed. The demographic parameters indicated a significant reduction in the net reproductive rate (184.27 vs 276.72), innate capacity for increase in number (0.11 vs 0.15), finite rate of increase in number (1.12 vs 1.16 female progenies produced/female/day), weekly multiplication rate (2.16 vs 2.86), potential fecundity (545.06 vs 634.11 eggs), number of hypothetical F2 females (33 955.65 vs 76 572.41), but longer mean length of generation (47.54 vs 37.74 days) and population doubling time (6.30 vs 4.62 days) in Field-R compared with Lab-S population. A stage-specific life table demonstrated the differences in survival rates between susceptible and resistant populations at various life stages, with the resistant population having higher generation mortality (0.22 vs 0.19). Conclusions: The study confirms the involvement of fitness costs associated with Bt resistance in P. gossypiella. Despite reduced reproductive fitness, the resistant population tried prolonging the larval stage as a compensatory mechanism to repair the damaged host tissues due to Bt intoxication and for accumulation of enough nutrient reserves for normal pupation and adult emergence. Presence of a high proportion of double Bt-resistant larvae in the field coupled with continued noncompliance with refug planting certainly favours the flaring up of this monophagous pest despite the observed fitness costs. The resistance cannot be effectively reversed unless suitable alternative management strategies are deployed. [ABSTRACT FROM AUTHOR]

BACKGROUND: Elucidating fitness cost associated with field‐evolved insect resistance to insecticide is of particular importance to current sustainable pest control. The global pest whitefly Bemisia tabaci has developed resistance to many members of neonicotinoids, but little is known about whitefly resistance to neonicotinoid nitenpyram and its associated fitness cost. Using insecticide bioassay and life‐table approach, this study aims to investigate nitenpyram resistance status in field‐collected whitefly populations, and to explore whether such resistance is accompanied by a fitness cost. RESULTS: The bioassay results revealed that 14 of 29 whitefly populations displayed moderate to extremely high resistance to nitenpyram, demonstrating a widespread field‐evolved resistance to nitenpyram. This field‐evolved resistance in the whitefly has increased gradually over the past 3 years from 2021 to 2023. Further life‐table study showed that two resistant whitefly populations exhibited longer developmental time, shorter lifespans of adult, and lower fecundity compared with the most susceptible population. The relative fitness cost of the two resistant populations was calculated as 0.69 and 0.56 by using net productive rate R0, which suggests that nitenpyram resistance comes with fitness cost in the whitefly, especially on reproduction. CONCLUSION: Overall, these results represent field‐evolved high resistance to nitenpyram in the whitefly. The existing fitness costs associated with nitenpyram resistance are helpful to propose a suitable strategy for sustainable control of whiteflies by rotation or mixture of insecticide with different modes of action. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

The rice stem borer (RSB) Chilo suppressalis is a devastating rice pest with resistance to a number of insecticides. Recently, the new meta-diamide insecticide cyproflanilide has been considered an effective insecticide to control RSB. However, its resistance risk has not been reported. In the present study, we aimed to assess the resistance risk and evaluate the fitness cost after the RSB was exposed to cyproflanilide. After five generations of selection, the resistance level of RSB increased by 1.5-fold. When h2 was 0.125, a 10-fold resistance increase in the LD50 values was expected in fourteen and thirty-one generations at the selection intensity of 90% and 50%, respectively. The selected population (RSB-SEL) had significant differences in the developmental duration of eggs, 1st, 2nd, 3rd, and 6th instar larvae, and female pupae compared to the unselected population (RSB-UNSEL). Besides, the adult longevity was shortened, and the average pupal weight of males, the emergence rate, the sex ratio, the oviposition, the mean fecundity, and the full life cycle rate were decreased in RSB-SEL. The intrinsic rate of increase (r), the net reproductive rate (R0), and the finite rate of increase (λ) of RSB-SEL were significantly lower than those of RSB-UNSEL, while the mean generation time (T) of RSB-SEL was significantly longer than that of RSB-UNSEL. Based on the results of the prediction of the generations required for a 10-fold resistance increase in the LD50, a potential risk of resistance development exists in RSB after continuous and excessive use of cyproflanilide. These results will be useful in designing the dose of cyproflanilide to control C. suppressalis in field. [ABSTRACT FROM AUTHOR]

Abstract Background The pink bollworm, Pectinophora gossypiella (Saunders), is a devastating global pest of cotton that has caused substantial economic damage to Bt Bollgard-II® cotton plants in recent years due to the evolution of Bt resistance. The associated fitness cost is assumed to be one of the factors delaying the development of resistance against Bt transgenic crops. Hence, the present study was undertaken to assess the biological performance of pink bollworms by comparing the life history and demographic parameters of a resistant (Field-R) and susceptible (Lab-S) population. Results Prolonged larval duration (23.40 days in Field-R vs 18.80 days in Lab-S population), total life cycle (male = 50.00 vs 42.80 days; female = 53.60 vs 46.20 days), reduced fecundity (100.60 vs 154.20 eggs/female) and fertility (88.00 vs 138.00 fertile eggs/female) was observed. The demographic parameters indicated a significant reduction in the net reproductive rate (184.27 vs 276.72), innate capacity for increase in number (0.11 vs 0.15), finite rate of increase in number (1.12 vs 1.16 female progenies produced/female/day), weekly multiplication rate (2.16 vs 2.86), potential fecundity (545.06 vs 634.11 eggs), number of hypothetical F2 females (33 955.65 vs 76 572.41), but longer mean length of generation (47.54 vs 37.74 days) and population doubling time (6.30 vs 4.62 days) in Field-R compared with Lab-S population. A stage-specific life table demonstrated the differences in survival rates between susceptible and resistant populations at various life stages, with the resistant population having higher generation mortality (0.22 vs 0.19). Conclusions The study confirms the involvement of fitness costs associated with Bt resistance in P. gossypiella. Despite reduced reproductive fitness, the resistant population tried prolonging the larval stage as a compensatory mechanism to repair the damaged host tissues due to Bt intoxication and for accumulation of enough nutrient reserves for normal pupation and adult emergence. Presence of a high proportion of double Bt-resistant larvae in the field coupled with continued noncompliance with refug planting certainly favours the flaring up of this monophagous pest despite the observed fitness costs. The resistance cannot be effectively reversed unless suitable alternative management strategies are deployed.

Objective: The aim of the present study is to explore the impact of the tet(A) type I variant (tetA-v1) on its fitness effect in Klebsiella pneumoniae. Methods: Clinical K. pneumoniae strains were utilized as parental strains to generate strains carrying only the plasmid vector (pBBR1MCS-5) or the tetA-v1 recombinant plasmid (ptetA-v1). Antimicrobial susceptibility testing was conducted to estimate the contribution of tetA-v1 to drug resistance. Plasmid stability was evaluated by serial passage over 10 consecutive days in the absence of tigecycline. Biological fitness was examined through growth curve analysis, in vitro competition assays and a neutropenic mouse thigh infection model. Results: A 2–4-fold increase in tigecycline MIC was observed following the acquisition of tetA-v1. Without tigecycline treatment, the stability of ptetA-v1 plasmids has been decreasing since day 1. The ptetA-v1 plasmid in Kp89, Kp91, and Kp93 exhibited a decrease of about 20% compared to the pBBR1MCS-5 plasmid. The acquisition of the tetA-v1 gene could inhibit the growth ability of K. pneumoniae strains both in vitro and in vivo. tetA-v1 gene imposed a fitness cost in K. pneumoniae, particularly in the CRKP strain Kp51, with a W value of approximately 0.56. Conclusion: The presence of tetA-v1 is associated with a significant fitness cost in K. pneumoniae in the absence of tigecycline, both in vitro and in vivo.

Abstract Background This study analyzed the genetic traits and fitness costs of vancomycin-resistant Enterococcus faecium (VREfm) blood isolates carrying Tn1546-type transposons harboring the vanA operon. Methods All E. faecium blood isolates were collected from eight general hospitals in South Korea during one-year study period. Antimicrobial susceptibility testing and vanA and vanB PCR were performed. Growth rates of E. faecium isolates were determined. The vanA-positive isolates were subjected to whole genome sequencing and conjugation experiments. Results Among 308 E. faecium isolates, 132 (42.9%) were positive for vanA. All Tn1546-type transposons harboring the vanA operon located on the plasmids, but on the chromosome in seven isolates. The plasmids harboring the vanA operon were grouped into four types; two types of circular, nonconjugative plasmids (Type A, n = 50; Type B, n = 46), and two types of putative linear, conjugative plasmids (Type C, n = 16; Type D, n = 5). Growth rates of vanA-positive E. faecium isolates were significantly lower than those of vanA-negative isolates (P

Plasmids incur a fitness cost that has the potential to restrict the dissemination of resistance in bacterial pathogens. However, bacteria can overcome this disadvantage by compensatory evolution to maintain their resistance. Compensatory evolution can occur via both chromosomes and plasmids, but there are a few reviews regarding this topic, and most of them focus on plasmids. In this review, we provide a comprehensive overview of the currently reported mechanisms underlying compensatory evolution on chromosomes and plasmids, elucidate key targets regulating plasmid fitness cost, and discuss future challenges in this field. We found that compensatory evolution on chromosomes primarily arises from mutations in transcriptional regulatory factors, whereas compensatory evolution of plasmids predominantly involves three pathways: plasmid copy number regulation, conjugation transfer efficiency, and expression of antimicrobial resistance (AMR) genes. Furthermore, the importance of reasonable selection of research subjects and effective integration of diverse advanced research methods is also emphasized in our future study on compensatory mechanisms. Overall, this review establishes a theoretical framework that aims to provide innovative ideas for minimizing the emergence and spread of AMR genes. [ABSTRACT FROM AUTHOR]

One of the longstanding puzzles of antimicrobial resistance is why the frequency of resistance persists at intermediate levels. Theoretical explanations for the lack of fixation of resistance include cryptic costs of resistance or negative frequency-dependence but are seldom explored experimentally. β -lactamases, which detoxify penicillin-related antibiotics, have well-characterized frequency-dependent dynamics driven by cheating and cooperation. However, bacterial physiology determines whether β -lactamases are cooperative, and we know little about the sociality or fitness of β -lactamase producers in infections. Moreover, media-based experiments constrain how we measure fitness and ignore important parameters such as infectivity and transmission among hosts. Here, we investigated the fitness effects of broad-spectrum AmpC β -lactamases in Enterobacter cloacae in broth, biofilms, and gut infections in a model insect. We quantified frequency- and dose-dependent fitness using cefotaxime, a third-generation cephalosporin. We predicted that infection dynamics would be similar to those observed in biofilms, with social protection extending over a wide dose range. We found evidence for the sociality of β -lactamases in all contexts with negative frequency-dependent selection, ensuring the persistence of wild-type bacteria, although cooperation was less prevalent in biofilms, contrary to predictions. While competitive fitness in gut infections and broth had similar dynamics, incorporating infectivity into measurements of fitness in infections significantly affected conclusions. Resistant bacteria had reduced infectivity, which limited the fitness benefits of resistance to infections challenged with low antibiotic doses and low initial frequencies of resistance. The fitness of resistant bacteria in more physiologically tolerant states (in biofilms, in infections) could be constrained by the presence of wild-type bacteria, high antibiotic doses, and limited availability of β -lactamases. One conclusion is that increased tolerance of β -lactams does not necessarily increase selection pressure for resistance. Overall, both cryptic fitness costs and frequency dependence curtailed the fitness benefits of resistance in this study. [ABSTRACT FROM AUTHOR]

Background: The increased incidence of infections due to multidrug-resistant Gram-negative bacteria has led to the renewed interest in the use of 'forgotten' antibiotics such as colistin. In this work, we studied the chromosomal colistin resistance mechanisms among laboratory-induced colistin-resistant Escherichia coli isolates. Methods: Three colistin-susceptible (ColS) clinical isolates of E. coli assigning to ST131, ST405, and ST361 were exposed to successively increasing concentrations of colistin. The nucleotide sequences of pmrA, pmrB, pmrD, phoP, phoQ, and mgrB genes were determined. The fitness burden associated with colistin resistance acquisition was determined by measuring the in vitro growth rate. Results: Colistin resistance induction resulted in 16–64 times increase in colistin MICs in mutants (n = 8) compared with parental isolates. Analysis of chromosomal genes in colistin-resistant mutants compared with those of ColS ancestors revealed genetic alterations confined to PmrAB two-component system and included PmrA G53R/R81S/L105P and PmrB E121K/E121A/A159P/A159V/G302E changes. The PmrB E121 was found as a critical position for colistin resistance development being altered in three mutants with different ancestors. The acquired colistin-resistance phenotype was stable following 10 consecutive passages in the absence of selective pressure of colistin and it did not alter the susceptibility of mutants to other antimicrobial agents. All mutants exhibited growth rates similar to their respective ColS ancestors, except for one isolate, which revealed a significant growth defect. Conclusion: Our results revealed that colistin resistance in E. coli was more related to PmrAB alterations, which did not impose a fitness cost in most cases. [ABSTRACT FROM AUTHOR]

Antimicrobial resistance (AMR) is a significant global health threat, with multidrug-resistant (MDR) bacterial clones becoming a major concern. Polymyxins, especially colistin, have reemerged as last-resort treatments for MDR Gram-negative infections. However, colistin use in livestock has spread mobile colistin resistance (mcr) genes, notably mcr-1, impacting human health. In consequence, its livestock use was banned in 2017, originating a natural experiment to study bacterial adaptation. The aim of this work was to analyse the changes in the mcr-1 genetic background after colistin restriction across the world. This study analyses 3163 Escherichia coli genomes with the mcr-1 gene from human and livestock hosts, mainly from Asia (n = 2621) and Europe (n = 359). Genetic characterisation identifies IncI2 (40.4%), IncX4 (26.7%), and multidrug-resistant IncHI2 (18.8%) as the most common plasmids carrying mcr-1. There were differences in plasmids between continents, with IncX4 (56.6%) being the most common in Europe, while IncI2 (44.8%) was predominant in Asia. Promoter variants related to reduced fitness costs and ISApl1 showed a distinct pattern of association that appears to be associated with adaptation to colistin restriction, which differed between continents. Thus, after the colistin ban, Europe saw a shift to specialised mcr-1 plasmids as IncX4, while ISApl1 decreased in Asia due to changes in the prevalence of the distinct promoter variants. These analyses illustrate the evolution of mcr-1 adaptation following colistin use restrictions and the need for region-specific strategies against AMR following colistin restrictions. [ABSTRACT FROM AUTHOR]

This article explores the relationship between antibiotic resistance mutations and their fitness benefits in different contexts. It emphasizes that the expression of traits is influenced by both genetic factors and the environment. The study specifically focuses on rifampicin-resistant Escherichia coli mutants and shows that the level of resistance to rifampicin can vary depending on the conditions of the experiment. The authors caution against making broad generalizations about antibiotic resistance and stress the importance of considering the environment when studying and addressing antimicrobial resistance. The article includes a list of references covering various aspects of antibiotic resistance, including the evolution of antibiotic treatment for tuberculosis, the role of genotype-by-environment interactions in shaping resistance, the limitations of certain metrics for measuring antibiotic response, and the significance of understanding the infectious microenvironment. These references provide diverse perspectives on antibiotic resistance and highlight the need for further research in this field. [Extracted from the article]

Introduction: The escalating prevalence of bacterial resistance, particularly multidrug-resistant bacteria like Acinetobacter baumannii, has become a significant global public health concern. The CRISPR-Cas system, a crucial defense mechanism in bacteria against foreign genetic elements, provides a competitive advantage. Type I-Fb and Type I-Fa are two subtypes of CRISPRCas systems that were found in A. baumannii, and the I-Fb CRISPR-Cas system regulates antibiotic resistance in A. baumannii. However, it is noteworthy that a majority of clinical isolates of A. baumannii lack or have incomplete CRISPRCas systems and most of them are multidrug-resistant. In light of this, our study aimed to examine the impact of antibiotic pressure on the fitness cost of the I-Fb CRISPR-Cas system in A. baumannii. Methods and Results: In the study, we conducted in vitro competition experiments to investigate the influence of sub-minimum inhibitory concentration (sub-MIC) on the CRISPR-Cas systems' fitness cost in A. baumannii. We found that the fitness cost of the CRISPR-Cas system was increased under sub-MIC conditions. The expression of CRISPR-Cas-related genes was decreased, while the conjugation frequency was increased in AB43 under sub-MIC conditions. Through metabolomic analysis, we identified that sub-MIC conditions primarily affected energy metabolism pathways. In particular, we observed increased carbon metabolism, nitrogen metabolism, and intracellular ATP. Notably, the CRISPR-Cas system demonstrated resistance to the efflux pump-mediated resistance. Furthermore, the expression of efflux pump-related genes was increased under sub-MIC conditions. Conclusion: Our findings suggest that the I-Fb CRISPR-Cas system confers a significant competitive advantage in A. baumanni. However, under sub-MIC conditions, its function and the ability to inhibit the energy required for efflux pumps are reduced, resulting in an increased fitness cost and loss of competitive advantage. [ABSTRACT FROM AUTHOR]

Background: This study analyzed the genetic traits and fitness costs of vancomycin-resistant Enterococcus faecium (VREfm) blood isolates carrying Tn1546-type transposons harboring the vanA operon. Methods: All E. faecium blood isolates were collected from eight general hospitals in South Korea during one-year study period. Antimicrobial susceptibility testing and vanA and vanB PCR were performed. Growth rates of E. faecium isolates were determined. The vanA-positive isolates were subjected to whole genome sequencing and conjugation experiments. Results: Among 308 E. faecium isolates, 132 (42.9%) were positive for vanA. All Tn1546-type transposons harboring the vanA operon located on the plasmids, but on the chromosome in seven isolates. The plasmids harboring the vanA operon were grouped into four types; two types of circular, nonconjugative plasmids (Type A, n = 50; Type B, n = 46), and two types of putative linear, conjugative plasmids (Type C, n = 16; Type D, n = 5). Growth rates of vanA-positive E. faecium isolates were significantly lower than those of vanA-negative isolates (P < 0.001), and reduction in growth rate under vancomycin pressure was significantly larger in isolates harboring putative linear plasmids than in those harboring circular plasmids (P = 0.020). Conclusions: The possession of vanA operon was costly to bacterial hosts in antimicrobial-free environment, which provide evidence for the importance of reducing vancomycin pressure for prevention of VREfm dissemination. Fitness burden to bacterial hosts was varied by type and size of the vanA operon-harboring plasmid. [ABSTRACT FROM AUTHOR]

Background: The resistance of Aspergillus flavus to the azole antifungal drugs is an emerging problem. Mutations in the molecular targets of the azole antifungals ‐ CYP 51 A, B and C ‐ are possible mechanisms of resistance, but data to confirm this hypothesis are scarce. In addition, the behaviour of resistant strains in vitro and in vivo is not yet understood. Objectives: This study had 3 objectives. The first was to compare the sequences of CYP51 A, B and C in resistant and susceptible strains of A. flavus. The second was to look for the existence of a fitness cost associated with resistance. The third was to evaluate the activity of voriconazole and posaconazole on resistant strains in the Galleria mellonella model. Methods: The CYP51 A, B and C sequences of seven resistant strains with those of four susceptible strains are compared. Fitness costs were assessed by growing the strains in RPMI medium and testing their virulence in G. mellonella larvae. In addition, G. mellonella larvae infected with strains of A. flavus were treated with voriconazole and posaconazole. Results: In the CYP51A sequences, we found the A91T, C708T and A1296T nucleotide substitutions only in the resistant strains. The resistant strains showed a fitness cost with reduced in vitro growth and reduced virulence in G. mellonella. In vivo resistance to posaconazole is confirmed in a strain with the highest MIC for this antifungal agent. Conclusions: These results allow to conclude that some substitutions in CYP51 genes, in particular CYP51A, contribute to resistance to azole drugs in A. flavus. The study of the relationship between drug dosage and treatment duration with resistance and the reduction of fitness costs in resistant strains is a major perspective of this study. This work could help to establish recommendations for the treatment of infections with resistant strains of A. flavus. [ABSTRACT FROM AUTHOR]

ABSTRACT The dissemination of antibiotic resistance genes (ARGs) through plasmids is a major mechanism for the development of bacterial antimicrobial resistance. The adaptation and evolution mechanisms of multidrug-resistant (MDR) plasmids with their hosts are not fully understood. Herein, we conducted experimental evolution of a 244 kb MDR plasmid (pJXP9) under various conditions including no antibiotics and mono- or combinational drug treatments of colistin (CS), cefotaxime (CTX), and ciprofloxacin (CIP). Our results showed that long-term with or without positive selections for pJXP9, spanning approximately 600 generations, led to modifications of the plasmid-encoded MDR and conjugative transfer regions. These modifications could mitigate the fitness cost of plasmid carriage and enhance plasmid maintenance. The extent of plasmid modifications and the evolution of plasmid-encoded antibiotic resistance depended on treatment type, particularly the drug class and duration of exposure. Interestingly, prolonged exposure to mono- and combinational drugs of CS and CIP resulted in a substantial loss of the plasmid-encoded MDR region and antibiotic resistance, comparable to the selection condition without antibiotic. By contrast, combinational treatment with CTX contributed to the maintenance of the MDR region over a long period of time. Furthermore, drug selection was able to maintain and even amplify the corresponding plasmid-encoded ARGs, with co-selection of ARGs in the adjacent regions. In addition, parallel mutations in chromosomal arcA were also found to be associated with pJXP9 plasmid carriage among endpoint-evolved clones from diverse treatments. Meanwhile, arcA deletion improved the persistence of pJXP9 plasmid without drugs. Overall, our findings indicated that plasmid-borne MDR region deletion and chromosomal arcA inactivation mutation jointly contributed to co-adaptation and co-evolution between MDR IncHI2 plasmid and Salmonella Typhimurium under different drug selection pressure.IMPORTANCEThe plasmid-mediated dissemination of antibiotic resistance genes has become a significant concern for human health, even though the carriage of multidrug-resistant (MDR) plasmids is frequently associated with fitness costs for the bacterial host. However, the mechanisms by which MDR plasmids and bacterial pairs evolve plasmid-mediated antibiotic resistance in the presence of antibiotic selections are not fully understood. Herein, we conducted an experimental evolution of a large multidrug-resistant plasmid in a Salmonella enterica Typhimurium host under single and combinatorial drug selection pressures. Our results show the adaptive evolution of plasmid-encoded antibiotic resistance through alterations of the MDR region in the plasmid, in particular substantial loss of the MDR region, in response to different positive selections, especially mono- and combinational drugs of colistin and ciprofloxacin. In addition, strong parallel mutations in chromosomal arcA were associated with pJXP9 carriage in Salmonella Typhimurium from diverse treatments. Our results thus highlight promoting the loss of the plasmid’s MDR region could offer an alternative approach for combating plasmid-encoded antibiotic resistance.

The rice stem borer (RSB), Chilo suppressalis, a notorious rice pest in China, has evolved a high resistance level to commonly used insecticides. Tetraniliprole, a new anthranilic diamide insecticide, effectively controls multiple pests, including RSB. However, the potential resistance risk of RSB to tetraniliprole is still unknown. In this study, the tetraniliprole‐selection (Tet‐R) strain was obtained through 10 continuous generations of selection with tetraniliprole 30% lethal concentration (LC30). The realized heritability (h2) of the Tet‐R strain was 0.387, indicating that resistance of RSB to tetraniliprole developed rapidly under the continuous selection of tetraniliprole. The Tet‐R strain had a high fitness cost (relative fitness = 0.53). We established the susceptibility baseline of RSB to tetraniliprole (lethal concentration at LC50 = 0.727 mg/L) and investigated the resistance level of 6 field populations to tetraniliprole. All tested strains that had resistance to chlorantraniliprole exhibited moderate‐ to high‐level resistance to tetraniliprole (resistance ratio = 27.7−806.8). Detection of ryanodine receptor (RyR) mutations showed that the Y4667C, Y4667D, I4758M, and Y4891F mutations were present in tested RSB field populations. RyR mutations were responsible for the cross‐resistance between tetraniliprole and chlorantraniliprole. Further, the clustered regularly interspaced palindromic repeats (CRISPR) / CRISPR‐associated protein 9‐mediated genome‐modified flies were used to study the contribution of RyR mutations to tetraniliprole resistance. The order of contribution of a single RyR mutation to tetraniliprole resistance was Y4667D > G4915E > Y4667C ≈ I4758M > Y4891F. In addition, the I4758M and Y4667C double mutations conferred higher tetraniliprole resistance than single Y4667C mutations. These results can guide resistance management practices for diamides in RSB and other arthropods. [ABSTRACT FROM AUTHOR]

Integrative and conjugative elements (ICEs) are important vectors of lateral gene transfer and contribute to the evolution of bacterial pathogens. However, studies on the transfer among species and the physiological consequences of ICEs are rare. The objective of this study was to investigate the cross-species transferability of newly identified erm(B)-carried ICE in Streptococcus anginosus San95 and its physiological consequences after transfer. The erm(B)-carried ICE, characterized by a triple serine integrase module, integrated into hsdM genes, thus designated ICESan95_hsdM. Analysis of ICESan95_hsdM revealed 32 additional ICESan95-like ICEs in the available NCBI genome (n = 24) and sequence of clinical isolates (n = 8). Polymerase chain reaction (PCR) was used to evaluate the 467 clinical isolates, of which 84 were positive for core genes (integrase, relaxase, and T4SS genes) of ICESan95_hsdM. Cross-species transfer experiments demonstrated that ICESan95_hsdM could transfer from S. anginosus to different streptococcal and enterococcal recipients. Growth and competitive culture assays showed acquisition of ICESan95_hsdM incurred no fitness cost. Our work discovered a group of ICEs in Streptococci and Enterococci. For the first time, we demonstrated the broad cross-species transferability to different species or genera of ICEs with no fitness cost that enables commensal S. anginosus to deliver antimicrobial resistance genes to other streptococci and enterococci. [ABSTRACT FROM AUTHOR]

Eravacycline (ERV) has emerged as a therapeutic option for the treatment of carbapenem-resistant pathogens. However, the advent of heteroresistance (HR) to ERV poses a challenge to these therapeutic strategies. This study aimed to investigate ERV HR prevalence among common clinical isolates and further characterize ERV HR in carbapenem-resistant Klebsiella pneumoniae (CRKP). A total of 280 clinical pathogens from two centers were selected for HR and analyzed using population analysis profiling (PAP) and modified E-tests. The PAP assay revealed an overall ERV HR prevalence of 0.7% (2/280), with intermediate heterogeneity observed in 24.3% (68/280) of strains. The proportion of heteroresistant strains was 18.3% according to modified E-test results. A time-killing assay demonstrated that CRKP CFU increased significantly after 10 h of ERV treatment, contributing to the reduced bactericidal effect of ERV in vitro. Interestingly, dual treatment with ERV and polymyxin B effectively inhibited the total CFU, simultaneously reducing the required polymyxin B concentration. Furthermore, fitness cost measurements revealed a growth trade-off in CRKP upon acquiring drug resistance, highlighting fitness costs as crucial factors in the emergence of ERV HR in CRKP. Overall, the findings of the current study suggest that ERV HR in clinical strains presents a potential obstacle in its clinical application. [ABSTRACT FROM AUTHOR]

Spodoptera littoralis (Boisd.) (Lepidoptera: Noctuidae) is a major economic pest attacking a variety of crops in Egypt and other Mediterranean countries. S. littoralis has developed resistance to both traditional and novel insecticides. The current study investigated S. littoralis resistance to indoxacarb regarding inheritance mode, realized heritability (h 2), and fitness costs. An indoxacarb-resistant strain (Indoxa-SEL) was obtained by selecting a field strain with indoxacarb. Indoxa-SEL strain outperformed the susceptible one (Indoxa-S) by 29.77-fold after 16 consecutive generations of selection. Based on the LC50 values of the progenies of reciprocal crosses F1 (R♂ × S♀) and F1ʹ (R♀ × S♂), S. littoralis resistance to indoxacarb was found to be autosomal and partially recessive. Chi-square tests for goodness-of-fit between observed and expected mortalities of self-bred F1 and resistant strain reciprocal crosses revealed that the resistance was controlled by multiple genes. The resistant strain had a relative fitness of 0.80, with significantly increased total preovipositional period of females, egg, larvae, pupae, preadult, adult, and total longevity period. The estimated realized heritability value in the Indoxa-SEL strain was 0.21. The current study will contribute to sustaining indoxacarb efficacy and designing effective resistance management programs against S. littoralis. [ABSTRACT FROM AUTHOR]

ABSTRACT The widespread sulfonamide resistance genes sul1, sul2, and sul3 in food and gut bacteria have attracted considerable attention. In this study, we assessed the in vivo fitness of sul gene-dependent sulfonamide-resistant Escherichia coli, using a murine model. High fitness costs were incurred for sul1 and sul3 gene-dependent E. coli strains in vivo. A fitness advantage was found in three of the eight mice after intragastric administration of sul2 gene-dependent E. coli strains. We isolated three compensatory mutant strains (CMSs) independently from three mice that outcompeted the parent strain P2 in vivo. Whole-genome sequencing revealed seven identical single nucleotide polymorphism (SNP) mutations in the three CMSs compared with strain P2, an additional SNP mutation in strain S2-2, and two additional SNP mutations in strain S2-3. Furthermore, tandem mass tag-based quantitative proteomic analysis revealed abundant differentially expressed proteins (DEPs) in the CMSs compared with P2. Of these, seven key fitness-related DEPs distributed in two-component systems, galactose and tryptophan metabolism pathways, were verified using parallel reaction monitoring analysis. The DEPs in the CMSs influenced bacterial motility, environmental stress tolerance, colonization ability, carbohydrate utilization, cell morphology maintenance, and chemotaxis to restore fitness costs and adapt to the mammalian gut environment.IMPORTANCESulfonamides are traditional synthetic antimicrobial agents used in clinical and veterinary medical settings. Their long-term excessive overuse has resulted in widespread microbial resistance, limiting their application for medical interventions. Resistance to sulfonamides is primarily conferred by the alternative genes sul1, sul2, and sul3 encoding dihydropteroate synthase in bacteria. Studying the potential fitness cost of these sul genes is crucial for understanding the evolution and transmission of sulfonamide-resistant bacteria. In vitro studies have been conducted on the fitness cost of sul genes in bacteria. In this study, we provide critical insights into bacterial adaptation and transmission using an in vivo approach.

Abstract Plasmids incur a fitness cost that has the potential to restrict the dissemination of resistance in bacterial pathogens. However, bacteria can overcome this disadvantage by compensatory evolution to maintain their resistance. Compensatory evolution can occur via both chromosomes and plasmids, but there are a few reviews regarding this topic, and most of them focus on plasmids. In this review, we provide a comprehensive overview of the currently reported mechanisms underlying compensatory evolution on chromosomes and plasmids, elucidate key targets regulating plasmid fitness cost, and discuss future challenges in this field. We found that compensatory evolution on chromosomes primarily arises from mutations in transcriptional regulatory factors, whereas compensatory evolution of plasmids predominantly involves three pathways: plasmid copy number regulation, conjugation transfer efficiency, and expression of antimicrobial resistance (AMR) genes. Furthermore, the importance of reasonable selection of research subjects and effective integration of diverse advanced research methods is also emphasized in our future study on compensatory mechanisms. Overall, this review establishes a theoretical framework that aims to provide innovative ideas for minimizing the emergence and spread of AMR genes.

Persistent microbial contamination commonly occurs in the environment. However, the characteristics and associated risks remain largely unknown. The coexistence of virulence factor genes (VFGs) and “last-resort” antibiotic resistance genes (LARGs) on human bacterial pathogens (HBPs) are notorious, creating ecological concerns and health risks. Herein, we explored the pathogenicity and antibiotic resistance levels of LARG-harboring HBPs in agricultural soils. Our findings revealed a high distribution level of VFGs and LARGs in soils (an absolute abundance up to 4.7 × 107 gene copies/g soil) by quantitative PCR (qPCR). Furthermore, most isolated LARG-harboring HBPs exhibited a 100 % lethality rate to Galleria mellonella. LARG-carrying plasmids had a low fitness cost to their host bacteria, implying the high adaptation of these plasmids within the HBPs. Most importantly, multiple LARG and VFG plasmid fusion and core genetic arrangements suggested that these LARG/VFG-linked plasmids endowed the stable and persistent horizontal spread of these genes in and/or cross the species and environments. This study not only unveiled high risk, multisource, compliance and stability aspects of environmentally persistent microbial contamination but also illuminated the importance of linking the phenotype-genotype-niche colonization of environmental microbial contamination within “One Health” framework.

The developing resistance of Venturia inaequalis to toxicants commonly used in systemic fungicides against apple scab has reduced their effectiveness, causing substantial fruit loss in orchards. To improve the situation and manage the resistance, a thorough analysis of the fitness potential among different pathogen biotypes, particularly those resistant to fungicides, is needed. In this study, the mycelial growth of V. inaequalis isolates with baseline sensitivity and resistance to one or more fungicides was assessed in vitro at four temperatures (6, 18, 27, and 30°C) and three agar concentrations in the nutrient medium (2, 4, and 6% m/V). Except for the mycelial growth at 27°C, the indicators of fitness predicted in vitro did not differ significantly between the V. inaequalis isolates with multiple resistance to fungicides and the biotypes with baseline sensitivity.

Objectives: This study aimed to delineate the ability of a plasmid, pS130–4, which harboured both hypervirulence and multidrug resistance genes, to disseminate within Klebsiella pneumoniae, as well as its potential formation mechanism. Methods: We employed whole-genome sequencing to decipher the genetic architecture of pS130–4. Its capability to conjugate and transfer was assessed through a series of experiments, including plasmid stability, competitive growth, and growth curve analysis. Its expression stability was further evaluated using drug sensitivity, larval survival, and biofilm formation tests. Results: pS130–4 contained four intact modules typical of self-transmissible plasmids. BLAST analysis revealed a sequence identity exceeding 90% with other plasmids from a variety of hosts, suggesting its broad prevalence. Our findings indicated the plasmid's formation resulted from IS26-mediated recombination, leading us to propose a model detailing the creation of this conjugative fusion plasmid housing both blaKPC-2 and hypervirulence genes. Our conjugation experiments established that pS130–4, when present in the clinical strain S130, was self-transmissible with an estimated efficiency between 10−5 and 10−4. Remarkably, pS130–4 showcased a 90% retention rate and did not impede the growth of host bacteria. Galleria mellonella larval infection assay demonstrated that S130 had pronounced toxicity when juxtaposed with high-virulence control strain NTUH-K2044 and low-toxicity control strain ATCC700603. Furthermore, pS130–4′s virulence remained intact postconjugation. Conclusion: A fusion plasmid, encompassing both hypervirulence and multidrug resistance genes, was viable within K. pneumoniae ST11-KL64 and incurred minimal fitness costs. These insights underscored the criticality of rigorous monitoring to pre-empt the escalation and distribution of this formidable super-plasmid.

Transgenic poplars have been used to control quarantine pests worldwide, such as the fall webworm (Hyphantria cunea, FW). However, the studies on the resistance mechanism of FW to Cry toxins are limited. This study obtained an FW strain with 45-fold resistance to Cry1Ab toxin by continuous screening in the laboratory. The resistance to Cry1Ab was autosomally completely recessive, and it had approximately 40-fold cross-resistance to Cry1Ac but no cross-resistance to Cry2Ab toxin. Analysis of indoor population life tables for the resistant and susceptible strains found no significant difference in the net reproduction rate (R0) between the two strains. The resistant strain had an insignificantly longer larval development duration and a comparable pupation rate, pupal weight, emergence rate, eggs laidper female, and egg hatching rate compared to the susceptible strain. This indicated that there was no fitness cost for Cry1Ab resistance. Our study helps in evaluating the risk of resistance to Cry1Ab toxin and the mechanisms of resistance to Cry1Ab toxin in FW.

The Type VI Secretion System (T6SS) is a widespread and highly effective mechanism of microbial warfare; it confers the ability to efficiently kill susceptible cells within close proximity. Due to its large physical size, complexity, and ballistic basis for intoxication, it has widely been assumed to incur significant growth costs in the absence of improved competitive outcomes. In this study, we precisely examine the fitness costs of constitutive T6SS firing in the bacterium Vibrio cholerae. We find that, contrary to expectations, constitutive expression of the T6SS has a negligible impact on growth, reducing growth fitness by 0.025 ± 0.5% (95% CI) relative to a T6SS− control. Mathematical modeling of microbial populations demonstrates that, due to clonal interference, constitutive expression of the T6SS will often be neutral, with little impact on evolutionary outcomes. Our findings underscore the importance of precisely measuring the fitness costs of microbial social behaviors and help explain the prevalence of the T6SS across Gram‐negative bacteria. [ABSTRACT FROM AUTHOR]

Resistance evolution during exposure to non-lethal levels of antibiotics is influenced by various stress responses of bacteria which are known to affect growth rate. Here, we aim to disentangle how the interplay between resistance development and associated fitness costs is affected by stress responses. We performed de novo resistance evolution of wild-type strains and single-gene knockout strains in stress response pathways using four different antibiotics. Throughout resistance development, the increase in minimum inhibitory concentration (MIC) is accompanied by a gradual decrease in growth rate, most pronounced in amoxicillin or kanamycin. By measuring biomass yield on glucose and whole-genome sequences at intermediate and final time points, we identified two patterns of how the stress responses affect the correlation between MIC and growth rate. First, single-gene knockout E. coli strains associated with reactive oxygen species (ROS) acquire resistance faster, and mutations related to antibiotic permeability and pumping out occur earlier. This increases the metabolic burden of resistant bacteria. Second, the ΔrelA knockout strain, which has reduced (p)ppGpp synthesis, is restricted in its stringent response, leading to diminished growth rates. The ROS-related mutagenesis and the stringent response increase metabolic burdens during resistance development, causing lower growth rates and higher fitness costs. [ABSTRACT FROM AUTHOR]

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BACKGROUND: Understanding the trade‐offs between insecticide resistance and the associated fitness is of particular importance to sustainable pest control. One of the most devastating pest worldwide, the whitefly Bemisia tabaci, has developed resistance to various insecticides, especially the neonicotinoid group. Although neonicotinoid resistance often is conferred by P450s‐mediated metabolic resistance, the relationship between such resistance and the associated fitness phenotype remains largely elusive. By gene cloning, quantitative reverse transcription (qRT)‐PCR, RNA interference (RNAi), transgenic Drosophila melanogaster, metabolism capacity in vitro and 'two sex‐age stage' life table study, this study aims to explore the molecular role of a P450 gene CYP4CS5 in neonicotinoid resistance and to investigate whether such resistance mechanism carries fitness costs in the whitefly. RESULTS: Our bioassay tests showed that a total of 13 field‐collected populations of B. tabaci MED biotype displayed low‐to‐moderate resistance to thiamethoxam and clothianidin. Compared to the laboratory susceptible strain, we then found that an important P450 CYP4CS5 was remarkably upregulated in the field resistant populations. Such overexpression of CYP4CS5 had a good match with the resistance level among the whitefly samples. Further exposure to the two neonicotinoids resulted in an increase in CYP4CS5 expression. These results implicate that overexpression of CYP4CS5 is closely correlated with thiamethoxam and clothianidin resistance. RNAi knockdown of CYP4CS5 increased mortality of the resistant and susceptible populations after treatment with thiamethoxam and clothianidin in bioassay, but obtained an opposite result when using a transgenic line of D. melanogaster expressing CYP4CS5. Metabolic assays in vitro revealed that CYP4CS5 exhibited certain capacity of metabolizing thiamethoxam and clothianidin. These in vivo and in vitro assays indicate an essential role of CYP4CS5 in conferring thiamethoxam and clothianidin resistance in whitefly. Additionally, our life‐table analysis demonstrate that the field resistant whitefly exhibited a prolonged development time, shortened longevity and reduced fecundity compared to the susceptible, suggesting an existing fitness cost as a result of the resistance. CONCLUSION: Collectively, in addition to the important role of CYP4CS5 in conferring thiamethoxam and clothianidin resistance, this resistance mechanism is associated with fitness costs in the whitefly. These findings not only contribute to the development of neonicotinoids resistance management strategies, but also provide a new target for sustainable whitefly control. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Alpha-cypermethrin interacts with the sodium channel and causes nerve blockage in insects. It is used to manage Aedes aegypti (Linnaeus) (Diptera: Culicidae), a primary vector of dengue worldwide. It not only affects both target and non-target organisms, but overuse of this insecticide increases the chances of resistance development in insect pests. In this study, resistance development, biological parameters, and stability of alpha-cypermethrin resistance were studied in a laboratory-selected strain of Ae. aegypti. The alpha-cypermethrin selected strain (Alpha Sel) developed an 11.86-fold resistance level after 12 rounds of alpha-cypermethrin selection compared to the unselected strain (Unsel). In biological parameters, Alpha Sel and Cross1 (Unsel ♂ and Alpha Sel♀) had shorter larval durations compared to Unsel and Cross2 (Unsel ♀ and Alpha Sel ♂) populations. The pupal duration of Alpha Sel and both crosses was shorter than that in the Unsel strain. The relative fitness of Alpha Sel, Cross1, and Cross2 was significantly less than that of the Unsel strain. These results indicate that alpha-cypermethrin resistance comes with fitness costs. Moreover, the frequency of alpha-cypermethrin resistance decreased when the Alpha Sel population was reared without further selection pressure for four generations. So, resistance was unstable and reversed when insecticide pressure ceased. We concluded that the judicious and rotational use of different insecticides with different modes of action and the adoption of other IPM-recommended practices would suppress resistance development for more extended periods in Ae. aegypti. [ABSTRACT FROM AUTHOR]

Quantifying the level of seed germiabiliy of herbicide-resistant (R) and susceptible (S) weeds is useful for understanding the evolutionary development of herbicide resistance, but also for implementing herbicide-resistance management strategies. Germination is a crucial aspect in the life phase of weeds. Phalaris brachystachys biotypes resistant to ACCase-inhibiting herbicides have been confirmed in wheat fields in Iran. This study aimed to investigate the germination behaviour of ACCase- resistant and susceptible subpopulations P. brachystachys under different environmental factors. An analysis of the seed germination of P. brachystachys sub populations at the seed stage was therefore conducted. The resistant (R) and susceptible (S) P. brachystachys subpopulation germination traits were tested in different temperature, salinity stress, drought, and burial depth conditions. All tests were carried out with five replications in a completely randomized design. The highest germination percentage in both subpopulations occurred at a temperature range of 20 to 25 °C. At 35 °C, no germination occurred. In terms of estimated cardinal temperatures, no differences were observed between S and R subpopulations. Seeds containing the ACCase Ile-1781-Thr mutation germinated better under salt and osmotic stress. There were no fitness differences in pH conditions. The percentage and rate of emergence of the R subpopulation were more than S subpopulation at different burial depths. The seedling emergence reached a maximum of 50% in the S subpopulation at a depth of 6.65 cm and in the R subpopulation at depths of 9.81 cm, respectively. Significant differences in seed germination were found between herbicide-resistant and susceptible sub population, and the pleiotropic effect of resistant alleles on germination and seed emergence under different environmental conditions was demonstrated. Results suggested that deep tillage operations and the delayed sowing of autumn-sown crops could control resistant populations that emerge more rapidly than S population; therefore, the prevalence in the R population may decrease. [ABSTRACT FROM AUTHOR]

Multidrug-resistant yeast Candida auris is a serious threat to public health with documented survival in various hospital niches. The dynamics of this survival benefit and its trade off with drug resistance are still unknown for this pathogen. In this study we investigate the oxidative stress response (OSR) in fluconazole-resistant C. auris and compare its relative fitness with fluconazole-susceptible strains. A total of 351 C. auris clinical isolates (61 fluconazole-susceptible and 290 fluconazole-resistant) were screened for stress tolerance by spot assay and 95.08 % fluconazole-susceptible isolates were hyper-resistant to oxidative stress while majority (94.5 %) fluconazole-resistant isolates had lower oxidative tolerance. Expression of Hog1 and Cta1 gene transcript levels and cellular catalase levels were significantly higher in fluconazole-susceptible isolates and a corresponding higher intracellular reactive oxygen species level (iROS) was accumulated in the fluconazole-resistant isolates. Biofilm formation and cell viability under oxidative stress revealed higher biofilm formation and better viability in fluconazole-susceptible isolates. Fluconazole-resistant isolates had higher basal cell wall chitin. On comparison of virulence, the % cytotoxicity in A549 cell line was higher in fluconazole-susceptible isolates and the median survival of the infected larvae in G. mellonella infection model was higher in fluconazole-resistant (5; IQR:4.5–5 days) vs. fluconazole-susceptible C. auris (2; IQR:1.5–2.5 days). All organisms evolve with changes in their environmental conditions, to ensure an optimal balance between proliferation and survival. Development of tolerance to a certain kind of stress example antifungal exposure in yeast can leads to a compensatory decrease in tolerance for other stresses. This study provides useful insights into the comparative fitness and antifungal susceptibility trade off in C. auris. We report a negative association between H2O2 tolerance and fluconazole susceptibility. Using in-vitro cell cytotoxicity and in-vivo survival assays we also demonstrate the higher virulence potential of fluconazole-susceptible C. auris isolates corroborating the negative correlation between susceptibility and pathogen survival or virulence. These findings could also be translated to clinical practice by investigating the possibility of using molecules targeting stress response and fitness regulating pathways for management of this serious infection.

ABSTRACT Reduced susceptibility to ART, the first-line treatment against malaria, is common in South East Asia (SEA). It is associated with point mutations, mostly in kelch13 (k13) but also in other genes, like ubp1. K13 and its compartment neighbors (KICs), including UBP1, are involved in endocytosis of host cell cytosol. We tested 135 mutations in KICs but none conferred ART resistance. Double mutations of k13C580Y with k13R539T or k13C580Y with ubp1R3138H, did also not increase resistance. In contrast, k13C580Y parasites subjected to consecutive RSAs did, but the k13 sequence was not altered. Using isogenic parasites with different k13 mutations, we found correlations between K13 protein amount, resistance, and fitness cost. Titration of K13 and KIC7 indicated that the cellular levels of these proteins determined resistance through the rate of endocytosis. While fitness cost of k13 mutations correlated with ART resistance, ubp1R3138H caused a disproportionately higher fitness cost.IMPORTANCEParasites with lowered sensitivity to artemisinin-based drugs are becoming widespread. However, even in these “resistant” parasites not all parasites survive treatment. We found that the proportion of surviving parasites correlates with the fitness cost of resistance-inducing mutations which might indicate that the growth disadvantages prevents resistance levels where all parasites survive treatment. We also found that combining two common resistance mutations did not increase resistance levels. However, selection through repeated ART-exposure did, even-though the known resistance genes, including k13, were not further altered, suggesting other causes of increased resistance. We also observed a disproportionally high fitness cost of a resistance mutation in resistance gene ubp1. Such high fitness costs may explain why mutations in ubp1 and other genes functioning in the same pathway as k13 are rare. This highlights that k13 mutations are unique in their ability to cause resistance at a comparably low fitness cost.

ABSTRACTAzoles are the primary antifungal drugs used to treat infections caused by Aspergillus fumigatus. However, the emergence of azole resistance in A. fumigatus has become a global health concern despite the low proportion of resistant isolates in natural populations. In bacteria, antibiotic resistance incurs a fitness cost that renders strains less competitive in the absence of antibiotics. Consequently, fitness cost is a key determinant of the spread of resistant mutations. However, the cost of azole resistance and its underlying causes in A. fumigatus remain poorly understood. In this observation, we revealed that the 10 out of 15 screened azole-resistant isolates, which possessed the most common azole-targeted cyp51A mutations, particularly the presence of tandem repeats in the promoter region, exhibit fitness cost when competing with the susceptible isolates in azole-free environments. These results suggest that fitness cost may significantly influence the dynamics of azole resistance, which ultimately contributes to the low prevalence of azole-resistant A. fumigatus isolates in the environment and clinic. By constructing in situ cyp51A mutations in a parental azole-susceptible strain and reintroducing the wild-type cyp51A gene into the azole-resistant strains, we demonstrated that fitness cost is not directly dependent on cyp51A mutations but is instead associated with the evolution of variable mutations related to conidial germination or other unknown development-related processes. Importantly, our observations unexpectedly revealed that some azole-resistant isolates showed no detectable fitness cost, and some even exhibited significantly increased competitive fitness in azole-free environments, highlighting the potential risk associated with the prevalence of these isolates.IMPORTANCEAzole resistance in the human fungal pathogen Aspergillus fumigatus presents a global public health challenge. Understanding the epidemic trends and evolutionary patterns of azole resistance is critical to prevent and control the spread of azole-resistant isolates. The primary cause is the mutation of the drug target 14α-sterol-demethylase Cyp51A, yet its impact on competitive ability remains uncertain. Our competition assays revealed a diverse range of fitness outcomes for environmental and clinical cyp51A-mutated isolates. We have shown that this fitness cost is not reliant on cyp51A mutations but might be linked to unknown mutations induced by stress conditions. Among these isolates, the majority displayed fitness costs, while a few displayed enhanced competitive ability, which may have a potential risk of spread and the need to closely monitor these isolates. Our observation reveals the variation in fitness costs among azole-resistant isolates of A. fumigatus, highlighting the significant role of fitness cost in the spread of resistant strains.

The rice stem borer (RSB) Chilo suppressalis is a devastating rice pest with resistance to a number of insecticides. Recently, the new meta-diamide insecticide cyproflanilide has been considered an effective insecticide to control RSB. However, its resistance risk has not been reported. In the present study, we aimed to assess the resistance risk and evaluate the fitness cost after the RSB was exposed to cyproflanilide. After five generations of selection, the resistance level of RSB increased by 1.5-fold. When h2 was 0.125, a 10-fold resistance increase in the LD50 values was expected in fourteen and thirty-one generations at the selection intensity of 90% and 50%, respectively. The selected population (RSB-SEL) had significant differences in the developmental duration of eggs, 1st, 2nd, 3rd, and 6th instar larvae, and female pupae compared to the unselected population (RSB-UNSEL). Besides, the adult longevity was shortened, and the average pupal weight of males, the emergence rate, the sex ratio, the oviposition, the mean fecundity, and the full life cycle rate were decreased in RSB-SEL. The intrinsic rate of increase (r), the net reproductive rate (R0), and the finite rate of increase (λ) of RSB-SEL were significantly lower than those of RSB-UNSEL, while the mean generation time (T) of RSB-SEL was significantly longer than that of RSB-UNSEL. Based on the results of the prediction of the generations required for a 10-fold resistance increase in the LD50, a potential risk of resistance development exists in RSB after continuous and excessive use of cyproflanilide. These results will be useful in designing the dose of cyproflanilide to control C. suppressalis in field.

Antimicrobial resistance (AMR) is a significant global health threat, with multidrug-resistant (MDR) bacterial clones becoming a major concern. Polymyxins, especially colistin, have reemerged as last-resort treatments for MDR Gram-negative infections. However, colistin use in livestock has spread mobile colistin resistance (mcr) genes, notably mcr-1, impacting human health. In consequence, its livestock use was banned in 2017, originating a natural experiment to study bacterial adaptation. The aim of this work was to analyse the changes in the mcr-1 genetic background after colistin restriction across the world. This study analyses 3163 Escherichia coli genomes with the mcr-1 gene from human and livestock hosts, mainly from Asia (n = 2621) and Europe (n = 359). Genetic characterisation identifies IncI2 (40.4%), IncX4 (26.7%), and multidrug-resistant IncHI2 (18.8%) as the most common plasmids carrying mcr-1. There were differences in plasmids between continents, with IncX4 (56.6%) being the most common in Europe, while IncI2 (44.8%) was predominant in Asia. Promoter variants related to reduced fitness costs and ISApl1 showed a distinct pattern of association that appears to be associated with adaptation to colistin restriction, which differed between continents. Thus, after the colistin ban, Europe saw a shift to specialised mcr-1 plasmids as IncX4, while ISApl1 decreased in Asia due to changes in the prevalence of the distinct promoter variants. These analyses illustrate the evolution of mcr-1 adaptation following colistin use restrictions and the need for region-specific strategies against AMR following colistin restrictions.

ABSTRACT: Objectives: We isolated a strain of Enterobacter hormaechei, ECC2783, co-harbouring blaNDM-1, blaKPC-2 and mcr-9 plasmids from a bloodstream infection and investigated its biological features. Methods: The presence of carbapenemase genes and mcr-9 was confirmed by polymerase chain reaction amplification. Whole genome sequencing and genomic analysis were performed on ECC2783. Experiments assessing the conjugation and stability of plasmids carrying the carbapenemase gene were performed. We also performed a colistin resistance induction experiment and studied the fitness cost of transconjugants. Results: ECC2783 has an extensive drug resistance phenotype. Multilocus sequence typing analysis results showed that ECC2783 belongs to sequence type 93. Bioinformatics analysis confirmed that ECC2783 has four plasmids, of which pECC2783_a, carrying mcr-9, is the IncHI2 type, and pECC2783_c, carrying blaNDM-1, is the IncX3 type. pECC2783_d, carrying blaKPC-2, is an unclassified type. We successfully obtained two transconjugants (J53/ECC2783_1, carrying blaNDM-1, and J53/ECC2783_2, carrying blaKPC-2 and blaNDM-1). There was no statistically significant difference in the relative growth rate between J53 and J53/ECC2783_2. Conclusion: For the first time, we isolated carbapenem-resistant E. hormaechei plasmids co-harbouring blaNDM-1, blaKPC-2, and mcr-9 from a patient with a blood stream infection. This isolate has a survival advantage in a hospital environment, and its clinical monitoring should be strengthened.

Abstract The Type VI Secretion System (T6SS) is a widespread and highly effective mechanism of microbial warfare; it confers the ability to efficiently kill susceptible cells within close proximity. Due to its large physical size, complexity, and ballistic basis for intoxication, it has widely been assumed to incur significant growth costs in the absence of improved competitive outcomes. In this study, we precisely examine the fitness costs of constitutive T6SS firing in the bacterium Vibrio cholerae. We find that, contrary to expectations, constitutive expression of the T6SS has a negligible impact on growth, reducing growth fitness by 0.025 ± 0.5% (95% CI) relative to a T6SS− control. Mathematical modeling of microbial populations demonstrates that, due to clonal interference, constitutive expression of the T6SS will often be neutral, with little impact on evolutionary outcomes. Our findings underscore the importance of precisely measuring the fitness costs of microbial social behaviors and help explain the prevalence of the T6SS across Gram‐negative bacteria.

Aedes mosquitoes are well-known vectors of arthropod-borne viruses (arboviruses). Mosquitoes are more frequently infected with insect-specific viruses (ISVs) that cannot infect vertebrates. Some ISVs interfere with arbovirus replication in mosquito vectors, which has gained attention for potential use against arbovirus transmission. Cell-fusing agent virus (CFAV), a widespread ISV, can reduce arbovirus dissemination in Ae. aegypti. However, vectorial capacity is largely governed by other parameters than pathogen load, including mosquito survival and biting behaviour. Understanding how ISVs impact these mosquito fitness-related traits is critical to assess the potential risk of using ISVs as biological agents. Here, we examined the effects of CFAV infection on Ae. aegypti mosquito fitness. We found no significant reduction in mosquito survival, blood-feeding behaviour and reproduction, suggesting that Ae. aegypti is tolerant to CFAV. The only detectable effect was a slight increase in human attraction of CFAV-infected females in one out of eight trials. Viral tolerance is beneficial for introducing CFAV into natural mosquito populations, whereas the potential increase in biting activity must be further investigated. Our results provide the first insight into the link between ISVs and Aedes mosquito fitness and highlight the importance of considering all aspects of vectorial capacity for arbovirus control using ISVs.

Annual indoxacarb resistance in Helicoverpa armigera (Hübner) populations collected from various crops in Australia was monitored between 2013 and 2023. Resistance frequency determined by F2 screening using a predetermined discriminating dose of indoxacarb, was lowest in the 2013–2014 and 2015–2016 seasons at 0.0164 and 0.0246, respectively. Resistance then increased significantly to a ten-year high of 0.0869 in 2018–2019 but declined to 0.0557 in 2019–2020 during a severe drought, remaining relatively stable thereafter to 2023. Indoxacarb resistance was first detected in H. armigera collected from maize in the Gwydir valley, New South Wales, in 2013 (strain GY7-39). In 2017, a second indoxacarb resistant H. armigera strain (UN1U3-10) was isolated from a population collected in chickpeas in the Liverpool Plains, New South Wales. Indoxacarb resistance of this strain was characterized to evaluate its potential to compromise the ongoing effectiveness of insecticide resistance management strategies in Australian farming systems. Survival at the discriminating dose of indoxacarb in UN1U3-10 was 28.9, 52.6, 86.7, and 92.9% in the F2, F3, F4, and F5, respectively. Following introgression with a susceptible strain and reselection with the discriminating dose of indoxacarb, the resistance ratio of UN1U3-10 was approximately 800-fold. Resistance was autosomal, incompletely dominant and conferred by more than 1 locus. While indoxacarb resistance in UN1U3-10 did not confer to emamectin benzoate or spinetoram and there was no evidence of major cross-resistance to the Bt toxins Cry1A, Cry2A or Vip3A, there was 5-fold reduced sensitivity to chlorantraniliprole. Indoxacarb resistance was suppressed by approximately 10-fold by PBO with no synergism by TPP or DEM, suggesting the involvement of cytochrome P450 enzymes. A stability analysis indicated a fitness cost may be associated with the genes that confer resistance in the UN1U3-10 strain. The potential risk for diverse indoxacarb resistance in the Australian H. armigera population is discussed. [ABSTRACT FROM AUTHOR]

Introduction: Understanding fitness costs associated with fungicide resistance is critical to improve resistance management strategies. E198A in b-tubulin confers resistance to the fungicide thiophanate-methyl and has been widely reported in several plant pathogens including Colletotrichum siamense. Method: To better understand potential fitness costs associated with the resistance, a ribonucleoprotein (RNP) complex mediated CRISPR/Cas9 system was used to create a point mutation (E198A) through homology directed repair (HDR) in each of the sensitive (E198) C. siamense isolates collected from strawberries, raspberries, and peaches. The RNP complex was delivered into fungal protoplasts using polyethylene glycol-mediated (PEG) transfection. Results: The transformation efficiency, the proportion of transformants of sensitive parental isolates containing the E198A mutation, averaged 72%. No off-target mutations were observed when sequences similar to the b-tubulin target region with a maximum of four mismatch sites were analyzed, suggesting that the CRISPR/Cas9 system used in this study was highly specific for genome editing in C. siamense.Of the 41 comparisons of fitness between mutant and wild type isolates through in vitro and detached fruit assays, mutant isolates appeared to be as fit (24 of 41 comparisons), if not more fit than wild-type isolates (10 of 41 comparisons). Discussion: The use of CRISPR/Cas9 to evaluate fitness costs associated with point mutations in this study represents a novel and useful method, since wildtype and mutant isolates were genetically identical except for the target mutation. [ABSTRACT FROM AUTHOR]

In recent decades, phage therapy has been overshadowed by the widespread use of antibiotics in Western countries. However, it has been revitalized as a powerful approach due to the increasing prevalence of antimicrobial-resistant bacteria. Although bacterial resistance to phages has been reported in clinical cases, recent studies on the fitness trade-offs between phage and antibiotic resistance have revealed new avenues in the field of phage therapy. This strategy aims to restore the antibiotic susceptibility of antimicrobial-resistant bacteria, even if phage-resistant variants develop. Here, we summarize the basic virological properties of phages and their applications within the context of antimicrobial resistance. In addition, we review the occurrence of phage resistance in clinical cases, and examine fitness trade-offs between phage and antibiotic sensitivity, exploring the potential of an evolutionary fitness cost as a countermeasure against phage resistance in therapy. Finally, we discuss future strategies and directions for phage-based therapy from the aspect of fitness trade-offs. This approach is expected to provide robust options when combined with antibiotics in this era of phage 're'-discovery. [ABSTRACT FROM AUTHOR]

The emergence of phage-resistant bacterial strains is one of the biggest challenges for phage therapy. However, the emerging phage-resistant bacteria are often accompanied by adaptive trade-offs, which supports a therapeutic strategy called "phage steering". The key to phage steering is to guide the bacterial population toward an evolutionary direction that is favorable for treatment. Thus, it is important to systematically investigate the impacts of phages targeting different bacterial receptors on the fitness of the bacterial population. Herein, we employed 20 different phages to impose strong evolutionary pressure on the host Pseudomonas aeruginosa PAO1 and examined the genetic and phenotypic responses of their phage-resistant mutants. Among these strains with impaired adsorptions, four types of mutations associated with bacterial receptors were identified, namely, lipopolysaccharides (LPSs), type IV pili (T4Ps), outer membrane proteins (OMPs), and exopolysaccharides (EPSs). PAO1, responding to LPS- and EPS-dependent phage infections, mostly showed significant growth impairment and virulence attenuation. Most mutants with T4P-related mutations exhibited a significant decrease in motility and biofilm formation ability, while the mutants with OMP-related mutations required the lowest fitness cost out of the bacterial populations. Apart from fitness costs, PAO1 strains might lose their resistance to antibiotics when counteracting with phages, such as the presence of large-fragment mutants in this study, which may inspire the usage of phage–antibiotic combination strategies. This work provides methods that leverage the merits of phage resistance relative to obtaining therapeutically beneficial outcomes with respect to phage-steering strategies. [ABSTRACT FROM AUTHOR]