Constitutive Resistance Research Articles

Tri‐Trophic Interactions of Cabbage Cultivars, a Generalist Aphid, Myzus persicae (Sulzer) and the Braconid Diaeretiella rapae McIntosh

ABSTRACTThe tri‐trophic interactions among five cabbage (Brassica oleracea var. capitata) cultivars—Copenhagen Market (CM), Red Acre (RA), Vertus Savoy (VS), Sugarloaf (SL) and Danish Ballhead (DBH)—a generalist aphid, Myzus persicae (Sulzer) and its parasitoid, Diaeretiella rapae McIntosh, were assessed. Copenhagen Market, known for its reported aphid resistance, recorded the fewest aphids (9.11 per plant; F = 6.50; df = 4, p < 0.01), followed closely by Red Acre (11.83 per plant), which was statistically similar. Upon aphid attack, all cultivars produced plant volatiles that influenced the attraction of the parasitoid, D. rapae. Copenhagen Market consistently showed a higher response across cultivar combinations, attracting 62.59%–67.50% of parasitoids. It also recorded a higher parasitisation rate (39.9%; F = 5.94; df = 4/20, p < 0.01) in a multiple‐choice test. The results indicate that cabbage cultivars employ both constitutive and induced resistance, as aphid‐resistant cultivars were also found to be more attractive to parasitoids. In addition to plant odour cues, factors such as plant colour, texture and host aphid kairomones can influence the orientation and parasitisation by D. rapae.

Design and structure-activity relationships of ether-linked alkylides: Hybrids of 3-O-descladinosyl macrolides and quinolone motifs

Ketolides (3-keto) such as TE-802 and acylides (3-O-acyl) like TEA0929 are ineffective against constitutively resistant pathogens harboring erythromycin ribosomal methylation (erm) genes. Following our previous work on alkylides (3-O-alkyl), we explored the structure–activity relationships of hybrids combining (R/S) 3-descladinosyl erythromycin with 6/7-quinolone motifs, featuring extended ether-linked spacers, with a focus on their efficacy against pathogens bearing constitutive erm gene resistance. Optimized compounds 17a and 31f not only reinstated efficacy against inducibly resistant pathogens but also demonstrated significantly augmented activities against constitutively resistant strains of Streptococcus pneumoniae and Streptococcus pyogenes, which are typically refractory to existing C-3 modified macrolides. Notably, hybrid 31f (coded ZN-51) represented a pioneering class of agents distinguished by its dual modes of action, with ribosomes as the primary target and topoisomerases as the secondary target. As a novel chemotype of macrolide-quinolone hybrids, alkylide 31f is a valuable addition to our armamentarium against macrolide-resistant bacteria.

Quantitative evaluation of oncogenic KRAS signaling and therapeutic effects of molecular-targeted drugs in living cells by single-molecule imaging.

76 Background: Oncogenic KRAS mutations pose challenges due to their constitutive activation and resistance to drugs in colorectal cancer. However, several aspects remain unresolved. Single-molecule imaging has revealed distinct behaviors of KRAS. Inactive KRAS wild-type (WT) molecules exhibit continuous lateral diffusion on the plasma membrane, while activated or oncogenic KRAS molecules display slower diffusion and transient trapping due to interactions with signaling molecules. These observations suggest that the overall signal intensity of KRAS within a cell is finely regulated by integrating short-term pulse signals during transient trapping. Methods: We conducted single-molecule observations of KRAS (WT, G13D, and G12D/C/V) in living colon cancer cells, both with and without molecular-targeted drugs, before and after EGF stimulation. We quantitatively analyzed their diffusional behavior to evaluate KRAS signaling and the therapeutic effects of molecular-targeted drugs. Results: The temporal fraction of trapped KRAS WT was highest at 2 minutes (6.4%) and decreased to pre-activation level (3.0%) at 5 minutes after EGF stimulation. Although the temporal fractions of trapped KRAS mutants were higher than those of KRAS WT both before stimulation (5%) and after stimulation (10%), their diffusional behaviors were mutation-specific. The temporal fraction of trapped KRAS G13D increased sharply, and those of KRAS G12D and G12C increased at a moderate rate, while that of KRAS G12V did not elevate after EGF stimulation. Cetuximab treatment significantly reduced the temporal fraction of trapped KRAS WT after stimulation, but only moderately decreased those of KRAS mutants. Notably, transient trappings of KRAS G12D and G12C were abrogated by KRAS inhibitors, and the combined therapy of KRAS inhibitors and cetuximab demonstrated additive effects. Conclusions: Our findings indicate that the mutation-specific diffusional behavior of oncogenic KRAS can be quantitatively assessed by single-molecule imaging. This method serves as a powerful tool to uncover the mechanisms underlying oncogenic KRAS signaling and evaluate the therapeutic efficacy of molecular-targeted drugs in living cells.

Macrolones target bacterial ribosomes and DNA gyrase and can evade resistance mechanisms.

Growing resistance toward ribosome-targeting macrolide antibiotics has limited their clinical utility and urged the search for superior compounds. Macrolones are synthetic macrolide derivatives with a quinolone side chain, structurally similar to DNA topoisomerase-targeting fluoroquinolones. While macrolones show enhanced activity, their modes of action have remained unknown. Here, we present the first structures of ribosome-bound macrolones, showing that the macrolide part occupies the macrolide-binding site in the ribosomal exit tunnel, whereas the quinolone moiety establishes new interactions with the tunnel. Macrolones efficiently inhibit both the ribosome and DNA topoisomerase in vitro. However, in the cell, they target either the ribosome or DNA gyrase or concurrently both of them. In contrast to macrolide or fluoroquinolone antibiotics alone, dual-targeting macrolones are less prone to select resistant bacteria carrying target-site mutations or to activate inducible macrolide resistance genes. Furthermore, because some macrolones engage Erm-modified ribosomes, they retain activity even against strains with constitutive erm resistance genes.

Patterns of constitutive and induced herbivore defence are complex, but share a common genetic basis in annual and perennial monkeyflower

Abstract Despite multiple ecological and evolutionary hypotheses that predict patterns of phenotypic relationships between plant growth, reproduction and constitutive and/or induced resistance to herbivores, these hypotheses do not make any predictions about the underlying molecular genetic mechanisms that mediate these relationships. We investigated how divergent plant life‐history strategies in the yellow monkeyflower and a life‐history altering locus, DIV1, influence plasticity of phytochemical herbivory resistance traits in response to attack by two herbivore species with different diet breadth. Life‐history strategy (annual vs. perennial) and the DIV1 locus significantly influenced levels of constitutive herbivory resistance, as well as resistance induction following both generalist and specialist herbivory. Perennial plants had higher total levels of univariate constitutive and induced defence than annuals, regardless of herbivore type. Annuals induced less in response to generalist herbivory than did perennials, while induction response was equivalent across the ecotypes for specialist herbivory. The effects of the DIV1 locus on levels of constitutive and induced defence were dependent on genetic background, the annual versus perennial haplotype of DIV1 and herbivore identity. The patterns of univariate induction due to DIV1 were non‐additive and did not always match expectations based on patterns of divergence for annual/perennial parents. For example, perennial plants had higher levels of constitutive and induced defence than did annuals, but when the annual DIV1 was present in the perennial genetic background induction response to herbivory was higher than for the perennial parent lines. Patterns for multivariate defence arsenals generally echoed those of univariate, with annual and perennial monkeyflowers and those with alternative versions of DIV1 differing significantly in constitutive and induced resistance. Like univariate resistance, induced multivariate defence arsenals were affected by herbivore identity. Our results highlight the complexity of the genetic mechanisms underlying plastic response to herbivory. While a genetic locus underlying substantial phenotypic variation in life‐history strategy and constitutive defence also influences defence plasticity, the induction response also depends on genetic background. This result demonstrates the potential for some degree of evolutionary independence between constitutive and induced defence, or induced defence and life‐history strategy, in monkeyflowers. Read the free Plain Language Summary for this article on the Journal blog.

Molecular Mechanisms of Bacterial Resistance to Antimicrobial Peptides in the Modern Era: An Updated Review.

Antimicrobial resistance (AMR) poses a serious global health concern, resulting in a significant number of deaths annually due to infections that are resistant to treatment. Amidst this crisis, antimicrobial peptides (AMPs) have emerged as promising alternatives to conventional antibiotics (ATBs). These cationic peptides, naturally produced by all kingdoms of life, play a crucial role in the innate immune system of multicellular organisms and in bacterial interspecies competition by exhibiting broad-spectrum activity against bacteria, fungi, viruses, and parasites. AMPs target bacterial pathogens through multiple mechanisms, most importantly by disrupting their membranes, leading to cell lysis. However, bacterial resistance to host AMPs has emerged due to a slow co-evolutionary process between microorganisms and their hosts. Alarmingly, the development of resistance to last-resort AMPs in the treatment of MDR infections, such as colistin, is attributed to the misuse of this peptide and the high rate of horizontal genetic transfer of the corresponding resistance genes. AMP-resistant bacteria employ diverse mechanisms, including but not limited to proteolytic degradation, extracellular trapping and inactivation, active efflux, as well as complex modifications in bacterial cell wall and membrane structures. This review comprehensively examines all constitutive and inducible molecular resistance mechanisms to AMPs supported by experimental evidence described to date in bacterial pathogens. We also explore the specificity of these mechanisms toward structurally diverse AMPs to broaden and enhance their potential in developing and applying them as therapeutics for MDR bacteria. Additionally, we provide insights into the significance of AMP resistance within the context of host-pathogen interactions.

Trends of Antibacterial Interactions in Multidrug – Resistant Isolates: Exploring Resistance Phenotypes in the Ndé Division, West-Cameroon

Background: Reliable pieces of information concerning bacterial-antibiotic interactions are key assets for therapeutic management of bacterial diseases.
 Objective: The present study aimed at detecting phenotypic characteristics of bacterial resistance in multidrug-resistant isolates recovered from clinical specimens at the “Université des Montagnes” Teaching Hospital.
 Methods: The total of 226 isolates (142 Gram-negative rods and 84 Gram-positive cocci) were subjected to phenotypic screening of resistance mechanisms. All procedural steps were conducted according to standard protocols on bacterial susceptibility to antibiotics.
 Results: Primary pieces of information revealed high rates of resistant isolates, especially with beta-lactams and Trimethoprim/Sulfamethoxazole while Nitrofurantoin and Imipenem were most effective. Amongst Gram-negative rods, 56% expressed one enzymatic resistance mechanism and 12% expressed two against beta-lactams. Also, with extended spectrum beta-lactamases, high level cephalosporinases and inducible cephalosporinases most commonly observed. About 62% and 14% of Gram-positive cocci expressed constitutive and Clindamycin-inducible resistance, respectively. Decreased susceptibility to Ceftriaxone and Penicillin G was also recorded in suspect mutant isolates selected by these antibiotics. Potential synergetic and other antagonistic interactions were evenly detected.
 Conclusion: Overall, the data could represent reliable clue for advocacy about personalized combination therapy, then capacity building for routine affordable susceptibility tests in caretaking.

Prevalence of inducible clindamycin resistance amongst methicillin resistant and sensitive strains of Staphylococcus aureus from a tertiary care hospital, West Bengal, India.

Background: Staphylococcus aureus is a prevalent pathogen causing both nosocomial and community-acquired infections worldwide. The emergence of methicillin-resistant Staphylococcus aureus (MRSA) due to the acquisition of mecA and mecC genes poses a significant clinical challenge. The injudicious use of clindamycin for treating MRSA has led to the development of clindamycin resistance. This study aimed to determine the prevalence of inducible clindamycin resistance (iMLSB resistance phenotype) in Staphylococcus aureus isolates, employing the D test according to CLSI guidelines, particularly focusing on erythromycin-resistant strains. Methodology: A total of 147 Staphylococcus aureus isolates were subjected to antibiotic susceptibility testing using the Kirby Bauer disc diffusion method. The D test was employed to identify inducible clindamycin resistance. Results: The study revealed that 34% of isolates exhibited inducible clindamycin resistance, 40% demonstrated constitutive resistance, and the remaining 26% exhibited the MS phenotype. Notably, inducible clindamycin resistance was more prevalent in MRSA (40%) compared to MSSA (22%). Conclusion: The findings underscore the importance of incorporating the D test as a mandatory procedure in standard disc diffusion testing to accurately identify inducible clindamycin resistance. This knowledge is crucial for guiding appropriate antibiotic therapy in the face of increasing resistance patterns.

Histological, chemical and gene expression differences between western redcedar seedlings resistant and susceptible to cedar leaf blight.

Western redcedar (Thuja plicata) is an important species in the Cupressaceae both at economic and cultural levels in the Pacific Northwest of North America. In adult trees, the species produces one of the most weathering-resistant heartwoods among conifers, making it one of the preferred species for outdoor applications. However, young T. plicata plants are susceptible to infection with cedar leaf blight (Didymascella thujina), an important foliar pathogen that can be devastating in nurseries and small-spaced plantations. Despite that, variability in the resistance against D. thujina in T. plicata has been documented, and such variability can be used to breed T. plicata for resistance against the pathogen. This investigation aimed to discern the phenotypic and gene expression differences between resistant and susceptible T. plicata seedlings to shed light on the potential constitutive resistance mechanisms against cedar leaf blight in western redcedar. The study consisted of two parts. First, the histological differences between four resistant and four susceptible families that were never infected with the pathogen were investigated. And second, the differences between one resistant and one susceptible family that were infected and not infected with the pathogen were analyzed at the chemical (C, N, mineral nutrients, lignin, fiber, starch, and terpenes) and gene expression (RNA-Seq) levels. The histological part showed that T. plicata seedlings resistant to D. thujina had constitutively thicker cuticles and lower stomatal densities than susceptible plants. The chemical analyses revealed that, regardless of their infection status, resistant plants had higher foliar concentrations of sabinene and α-thujene, and higher levels of expression of transcripts that code for leucine-rich repeat receptor-like protein kinases and for bark storage proteins. The data collected in this study shows that constitutive differences at the phenotypic (histological and chemical) and gene expression level exist between T. plicata seedlings susceptible and resistant to D. thujina. Such differences have potential use for marker-assisted selection and breeding for resistance against cedar leaf blight in western redcedar in the future.

Clindamycin Resistance Detection among MRSA at a Tertiary Care Hospital: A Cross-sectional Study

Introduction: Antimicrobial resistance is on the rise, posing a global threat to the success of modern surgical procedures. A common and notorious superbug causing community and hospital-acquired infections is Methicillin-resistant Staphylococcus aureus (MRSA). Its potential for easy and rapid spread leads to increased mortality in hospitals. Clindamycin, a relatively inexpensive and effective drug, is empirically prescribed for the treatment of MRSA infections. Due to the risk of misidentifying clindamycin resistance as susceptibility, leading to treatment failure, its judicious use after D-test is advocated. Aim: To isolate Staphylococcus aureus (S.aureus) from various clinical specimens and to identify MRSA using cefoxitin disc (30 μg), further determining the incidence of inducible clindamycin resistance among MRSA by the disc diffusion method (D-test). Materials and Methods: The present cross-sectional study was conducted in the Department of Microbiology, Government Mohan Kumaramangalam Medical College Hospital (GMKMCH), Salem, Tamil Nadu, India, from June 2023 to August 2023. Study was conducted using 185 S.aureus isolates collected from samples received from patients of all genders treated at the present study hospital. Various clinical samples such as pus, sputum, blood, urine and fluids collected from patients treated at the study Institute Salem during the study period were included. A cefoxitin disc (30 μg) was used to detect MRSA among the 185 S.aureus isolates by disc diffusion method as per Clinical and Laboratory Standards Institute (CLSI) guidelines, which were further tested for clindamycin resistance using clindamycin (2 μg) with erythromycin (15 μg) discs by the D-test. The data of the study were analysed using Statistical Package for the Social Sciences (SPSS) software version 21.0. Further descriptive statistics and a chi-square test with a level of significance, p-value ≤0.005 (statistically significant), were used. Results: Among 185 S.aureus isolates, 130 (70.27%) were Methicillin-sensitive S.aureus (MSSA) and 55 (29.7%) were MRSA. Inducible clindamycin resistance was observed in 22 (40%) isolates, while 17 (30.9%) isolates showed constitutive resistance, 9 (16.36%) showed the MS phenotype, and the remaining 7 (12.72%) showed the susceptible phenotype. Among the clindamycin resistance patterns in MRSA, inducible clindamycin resistance was reported predominantly. Conclusion: The majority of S.aureus was isolated from pus samples, highlighting its importance as a pyogenic microorganism. The current study records a high rate of MRSA resistance among S.aureus isolates. The present study reports a higher rate of inducible clindamycin resistance among MRSA isolates when compared to other phenotypes of clindamycin resistance. Therefore, routine D-testing must be implemented to identify inducible resistance to clindamycin in MRSA to avoid treatment failure.

Effect Of Negative Temperature Shock On The Primary Rezistance Of Maize Hybrids

The paper presents the results of assessing the resistance of 14 maize hybrids to a negative temperature shock (NTS) of -4°C for 16 hours. The germination and growth seedlings parameters of maize hybrids were studied. Germination of maize seeds at optimal temperature of 26°C for 5 days was used as a control, and pretreatment of maize seeds with NTS followed by germination and subsequent growth at 26°C for 5 days was considered experimental. Pretreatment of maize seeds with NTS made it possible to identify and evaluate the primary resistance of each hybrid to stress, and according to the degree of constitutive frost resistance of each hybrid, the distribution and classification of the studied hybrids into 3 zones was carried out, including hybrids with high, medium and low frost resistance. Also, the effect of NTS on the morpho-physiological traits of the maize seedlings are shown.

Characterization of biofilm formation in uropathogenic Staphylococcus aureus and their association with antibiotic resistance

IntroductionThe increase in antibiotic resistance and biofilm formation in Staphylococcus aureus is a serious public health problem, complicating the treatment of infections caused by these bacteria, including urinary tract infections (UTIs). This research aimed to assess the correlation between antibiotic resistance and the capacity for biofilm formation in S. aureus strains isolated from patients with UTIs. MethodsThe antibiotic susceptibility testing was conducted using disc diffusion and broth micro-dilution method. The blaZ and mecA genes were investigated in all penicillin and cefoxitin-resistant S. aureus strains, using PCR. Biofilm formation was investigated using the Congo red agar and microtitre plate methods. All S. aureus strains were screened for genes encoding adhesion and biofilm formation (spa, fnb AB, clfA, bap) and intercellular adhesin (icaADB and icaR). ResultsAmong 20 strains of S. aureus, 55% were found to be multidrug-resistant (MDR) and methicillin-resistant (MRSA). All strains were biofilm producers, including 55% moderate biofilm producers and 35% strong biofilm producers. A significant correlation (p-value <0.01) was observed between MDR and non-MDR strains in terms of biofilm production. All strains were sensitive to linezolid, and 20% exhibited the constitutive clindamycin resistance phenotype. The spa, fnbB, and clfA genes were present in all strains followed by the bap (95%) and fnbA (90%). The icaA (90%) and icaB (60%) genes were most detected. The coexistence of icaA, icaADB, and icaAD genes was significantly associated with MRSA strains. In contrast, the icaR gene was significantly detectable in the methicillin-susceptible group of strains. ConclusionThis study uncovered a significant association between the formation of biofilms by uropathogenic S. aureus and antibiotic resistance, highlighting a noteworthy concern for public health.

Aphid Resistance Evaluation and Constitutive Resistance Analysis of Eighteen Lilies.

Lilies (Lilium spp.) are famous bulb flowers worldwide, with high ornamental value. Aphid damage has seriously constrained the development of the lily industry. In this study, the aphid resistance of 16 lily cultivars and 2 wild lily species was characterized in the field and greenhouse. Leaf color parameters, stomatal density and size, thickness of leaf layers, leaf waxy content, and leaf water content were determined to explore the constitutive resistance of lilies. The results show that there was a significant positive correlation between the number of aphids in the field and in the greenhouse (p ≤ 0.05, r = 0.47). This indicated that the level of aphid infestation in both the field and the greenhouse is generally consistent across different types of lily plants. Among these 18 lilies, 'Palazzo', 'Nymph', 'Cameleon' and L. lancifolium were resistant to A. gossypii, while 'Black Beauty' and 'Magnefique' had poor resistance. The correlation analysis results showed that the number of aphids was negatively correlated with leaf abaxial surface a*, stomatal size, water content, and thickness of leaf palisade tissue and positively correlated with leaf distal axial surface b*, C*, and waxy content. Among them, the correlation between the number of aphids and the thickness of leaf palisade tissue reached a significant level (p ≤ 0.05, r = -0.521). This indicated that the thickness of the palisade tissue of lily leaves might be an important factor influencing the proliferation of aphids. This study not only screened out aphid-resistant lilies but also established a crucial research foundation for the targeted breeding and molecular breeding of lilies with aphid resistance.

Targeting the aryl hydrocarbon receptor (AhR) with BAY 2416964: a selective small molecule inhibitor for cancer immunotherapy

BackgroundThe metabolism of tryptophan to kynurenines (KYN) by indoleamine-2,3-dioxygenase or tryptophan-2,3-dioxygenase is a key pathway of constitutive and adaptive tumor immune resistance. The immunosuppressive effects of KYN in the tumor...

Estrogen receptor α isoforms generated by alternative use of cryptic exons.

Estrogen receptor α (ERα) regulates several physiological functions. In pathophysiological conditions, ERα is involved in the development and progression of estrogen-sensitive tumors. The ERα gene contains multiple 5'-untranslated exons and eight conventional coding exons and presents multiple isoforms generated by alternative promoter usage and alternative splicing. This gene also possesses non-conventional exons in the 3'- and intronic regions, and alternative use of cryptic exons contributes to further diversity of ERα mRNAs and proteins. Recently, the genomic organization of ERα genes and the splicing profiles of their transcripts were comparatively analyzed in humans, mice, and rats, and multiple ERα isoforms with distinct structures and functions were identified. These transcripts contain cryptic sequences that encode insertion-containing or truncated ERα proteins. In particular, alternative cryptic exons with in-frame stop codons yield transcripts encoding C-terminally-truncated ERα proteins. The C-terminally-truncated ERα isoforms lack part or all of the ligand-binding domain but possess an isoform-specific sequence. Some of these isoforms exhibit constitutive transactivation and resistance to estrogen receptor antagonists. Although numerous studies have reported conflicting results regarding their functions, the critical determinant for their gain-of-function has been identified structurally. Here we review recent progress in ERα variant research concerning the genomic organization of ERα genes, splicing profiles of ERα transcripts, and transactivation properties of ERα isoforms.

Antixenosis in Constitutive Resistance in Maize Genotypes to the Stink Bug Diceraeus melacanthus

Corn is one of the most important agricultural crops in the world, however, it can be affected by numerous phytophagous insects that causing economic losses in production. Diceraeus melacanthus belongs to the complex of pests that attack maize. The objective of this work was to evaluate 17 maize genotypes regarding the effects of antixenosis resistance to the stink bug D. melacanthus. The experiments were carried out in a greenhouse with maize plants in stage V2 and adult stink bugs. Bioassays of attractiveness and food preference were carried out, in addition to evaluating physical and morphological factors of the plants, such as tissue hardness, number of punctures and colorimetric factors. Genotypes 30A37, IAC 8390, Defender, NS 77 and Supremo Tg were the ones that expressed the greatest antixenotic effects in tests free choice and no-choice, among these Defender and Supremo Tg due to possible morphological causes such as plant tissue hardness. Although the IAC 8390 genotype also presents great potential for resistance, it was not possible to attribute its causes, so more studies should be conducted to evaluate the possible chemical constituents that give them this characteristic.

OR11-03 A Novel Mutation In The Ligand-binding Domain Of The NR3C1 Gene Associated With Fluctuating Resistance To Glucocorticoids

Abstract Disclosure: M. Laulhé: None. E. Kuhn: None. J. Bouligand: None. L. Amazit: None. J. Perrot: None. P. Kamenicky: None. M. Lombes: None. J. Fagart: None. S. Viengchareun: None. L. Martinerie: None. Introduction: Constitutive glucocorticoid resistance is associated with mutations in the NR3C1 gene, encoding the Glucocorticoid Receptor (GR). We identified a novel heterozygous point mutation causing fluctuating resistance to glucocorticoid depending on ligand concentrations. Initially, the propositus, a 45-year-old man, was diagnosed with a right adrenal incidentaloma and an ACTH-independent biological hypercortisolism. Shortly after adrenal surgery, plasma cortisol increased again (612 nmol/L) associated with inadequate ACTH levels (122 pg/mL) without any evidence of pituitary adenoma or ectopic ACTH-secreting tumor, suggesting a glucocorticoid resistance syndrome. Methods: We sequenced NR3C1 gene and performed in vitro analysis to assess contribution of this variant to the patient’s phenotype and how it affects biological activity of GR. Results: Exon sequencing of the NR3C1 gene identified a novel missense variant located on exon 5 (NM_000176.3, c.1706 G&amp;gt;A) resulting in substitution of arginine to glutamine at amino acid 569 (GR R569Q) in the Ligand-Binding Domain (LBD) of GR. We characterized transcriptional activity of GR R569Q in HEK 293T cells transiently co-transfected with either GR WT or GR R569Q, together with a luciferase reporter gene construct placed under control of a promoter containing two glucocorticoid responsive elements. We observed a decreased transcriptional activity of GR R569Q (EC50 4.55 nM [3.8;5.3]) compared to GR WT (EC50 0.19 nM [0.15; 0.24]), in response to dexamethasone. Interestingly, GR R569Q exerted a dominant negative effect on GR WT, with a decrease in transcriptional activity of 80% when stimulated with 1 nM dexamethasone (2-way ANOVA test, P&amp;lt;0.0001), which was blunted when stimulated with a 100-fold higher concentration of dexamethasone. As the variant is located within the second nuclear localization signal, we studied nuclear translocation in response to 1 nM cortisol for 1 h. As expected, nuclear-cytoplasmic ratio was lower in patients’ fibroblasts (2.3 ± 0.14 vs 3.0 ± 0.98, ANOVA 2-Way, P &amp;lt; 0.0001) compared with control fibroblasts, as quantified by automated high-throughput microscopy. Three-dimensional modeling revealed that arginine substitution leads to loss of a hydrogen bond stabilizing the LBD structure. This may result in a lower ligand affinity that can be bypassed by high concentrations of agonist. We are currently testing receptor affinity and reversibility of phenotype after hydrogen bond restoration by directed mutagenesis (c.1705 C&amp;gt;A, p.Q569K). Conclusion: Overall, we identified a novel variant of the NR3C1 gene associated with fluctuating GR sensitivity depending on ligand concentration. This finding may provide new insights into mechanisms involved in GR sensitivity. Presentation: Friday, June 16, 2023

Plant Stomata: An Unrealized Possibility in Plant Defense against Invading Pathogens and Stress Tolerance.

Stomata are crucial structures in plants that play a primary role in the infection process during a pathogen's attack, as they act as points of access for invading pathogens to enter host tissues. Recent evidence has revealed that stomata are integral to the plant defense system and can actively impede invading pathogens by triggering plant defense responses. Stomata interact with diverse pathogen virulence factors, granting them the capacity to influence plant susceptibility and resistance. Moreover, recent studies focusing on the environmental and microbial regulation of stomatal closure and opening have shed light on the epidemiology of bacterial diseases in plants. Bacteria and fungi can induce stomatal closure using pathogen-associated molecular patterns (PAMPs), effectively preventing entry through these openings and positioning stomata as a critical component of the plant's innate immune system; however, despite this defense mechanism, some microorganisms have evolved strategies to overcome stomatal protection. Interestingly, recent research supports the hypothesis that stomatal closure caused by PAMPs may function as a more robust barrier against pathogen infection than previously believed. On the other hand, plant stomatal closure is also regulated by factors such as abscisic acid and Ca2+-permeable channels, which will also be discussed in this review. Therefore, this review aims to discuss various roles of stomata during biotic and abiotic stress, such as insects and water stress, and with specific context to pathogens and their strategies for evading stomatal defense, subverting plant resistance, and overcoming challenges faced by infectious propagules. These pathogens must navigate specific plant tissues and counteract various constitutive and inducible resistance mechanisms, making the role of stomata in plant defense an essential area of study.

Transient eco-evolutionary dynamics early in a phage epidemic have strong and lasting impact on the long-term evolution of bacterial defences.

Organisms have evolved a range of constitutive (always active) and inducible (elicited by parasites) defence mechanisms, but we have limited understanding of what drives the evolution of these orthogonal defence strategies. Bacteria and their phages offer a tractable system to study this: Bacteria can acquire constitutive resistance by mutation of the phage receptor (surface mutation, sm) or induced resistance through their CRISPR-Cas adaptive immune system. Using a combination of theory and experiments, we demonstrate that the mechanism that establishes first has a strong advantage because it weakens selection for the alternative resistance mechanism. As a consequence, ecological factors that alter the relative frequencies at which the different resistances are acquired have a strong and lasting impact: High growth conditions promote the evolution of sm resistance by increasing the influx of receptor mutation events during the early stages of the epidemic, whereas a high infection risk during this stage of the epidemic promotes the evolution of CRISPR immunity, since it fuels the (infection-dependent) acquisition of CRISPR immunity. This work highlights the strong and lasting impact of the transient evolutionary dynamics during the early stages of an epidemic on the long-term evolution of constitutive and induced defences, which may be leveraged to manipulate phage resistance evolution in clinical and applied settings.

Distribution of Macrolide, Lincosamide, and Streptogramin B and Detection of erm Genes in Staphylococcus aureus from Wounds in Uyo, Nigeria

The increasing resistance to macrolide, lincosamide, and streptogramin B (MLSB) among methicillin-resistant Staphylococcus aureus (MRSA) is a challenge. The study determined the constitutive and inducible MLSB resistance and erm genes in S. aureus from wounds using the erythromycin-clindamycin D-zone test and a multiplex polymerase chain reaction assay, respectively. Of the 260 patients recruited in the study, S. aureus was isolated from the wounds of 102 patients, giving a prevalence rate of 39.2%. Of the 102 S. aureus isolates, 32.4% were methicillin-sensitive S. aureus (MSSA), and 67.6% were MRSA. The prevalence of S. aureus in wounds was higher in females (39.8%) than in males (38.4%). The highest prevalence of S. aureus was found in divorce patients (59.1%) and those residing in urban areas (39.7%). There was no statistical difference between the occurrence of MRSA and MSSA in wounds based on the sex (p = 0.97), age (p = 0.08), and marital status (p = 0.41) of the patients. All (100%) MRSA were resistant to Cefoxitin, and chloramphenicol, while ˂ 50% of MRSA were resistant to Tetracycline, Ciprofloxacin, and Gentamycin. Of the 69 MRSA isolates from the wounds, 21.7% and 26.1% were iMLSB and cMLSB phenotypes, respectively. Nine (9) MSSA were iMLSB phenotypes, and eight MSSA were cMLSB phenotypes. Among the 12 representative isolates, three (3) MRSA and one (1) MSSA isolate possessed the ermC gene. This study has revealed that screening tests for iMLSB-resistant S. aureus strains are critical for the therapeutic management of wound infections caused by S. aureus.