Resistant Genotypes Research Articles

Pooled prevalence of Escherichia coli phenotypic and genotypic antimicrobial resistance profiles in poultry: systematic review and meta-analysis

Pooled prevalence of Escherichia coli phenotypic and genotypic antimicrobial resistance profiles in poultry: systematic review and meta-analysis

Development of Faba Bean Genotypes for Broomrape Tolerant and Foliar Diseases Resistance in F\(_2\) Factorial Crosses

Two field experiments were conducted at Sakha Agricultural Research Station, Kafr El-Sheikh, Egypt. The first experiment was in a naturally Orobanche- free field and the second one was in a naturally Orobanche-infested field. In each experiment sixteen F2 crosses and their parents were sown. The sixteen F2 crosses were produced by a 4 x 4 factorial mating design in F1 and selfed to give F2-crosses. The data were then analyzed using a line x tester design. The main objective of this study was to develop new promising faba bean genotypes that are able to produce high yield, resistance to foliar diseases and broomrape tolerance. Significant differences among all genotypes mean square were observed for all studied traits under both growth conditions. The parents, crosses and their interaction mean squares were detected for all traits in the two environments. The mean squares of lines were significant and/or highly significant for flowering date and 100-seed weight under both conditions, while number of pods per plant was significant under normal condition, in addition to the number of seeds per plant and broomrape dry weight per plant under infested broomrape condition were significant and highly significant, respectively. Mean squares of testers were significant for chocolate spot at the two conditions; rust disease, plant height, number of pods and seeds, seed yield per plant and 100-seed weight under normal condition. Lines x testers mean squares were significant for all traits in both conditions, except for number of branches and 100-seed weight. L1 and L4 and the parental testers T4 were considered more resistance parents to foliar diseases under both environments. The crosses; L2 x T3, L2 xT4, L3 x T2, L3 x T4 and L4 x T4 were considered resistant to foliar disease and had an earlier flowering date. The testers, T3 and T4 showed significantly the highest values of number of seeds per plant, seed yield per plant and 100-seed weight under both conditions, while the crosses; L3 x T4 and L4 x T4 had the highest values for seed yield per plant and 100-seed weight under both conditions. The parental L2 and T4 were considered good combiners for chocolate spot and rust diseases under both environmental conditions, while L1 and T4 performed as good combiner parents for number of seeds and seed yield per plant. The cross; L2x T3 performed as good specific combiner cross for foliar diseases under both conditions. The crosses; L2x T3, L3xT4, L3 x T1and L4 x T4 had highly significant (sij) values for the No. of seeds and seed yield per plant under infested soil condition. The phenotypic coefficient of variation (PCV) values ranged from 5.52% to 84.01% and genotypic coefficient of variation (GCV) values ranged from 3.18% to 79.21%. Broad-sense heritability (H) ranged from 72.98% to 99.29%, and narrow sense heritability (h2) ranged from 2.3% to 21.38%. Expected genetic advance (Ga) ranged from 0.03 to 0.71, and predicted genetic advance (Ga%) ranged from 8.21% to 10.86%.

Exploring Metabolic Pathways and Phytohormonal Influence in Preharvest Sprouting Resistant and Susceptible Rice Genotypes

Exploring Metabolic Pathways and Phytohormonal Influence in Preharvest Sprouting Resistant and Susceptible Rice Genotypes

Serotypes, Antimicrobial Susceptibility, and Potential Mechanisms of Resistance Gene Transfer in Erysipelothrix rhusiopathiae Strains from Waterfowl in Poland

Erysipelas is a significant problem in the waterfowl farming in Poland, and information on the characteristics of the Erysipelothrix rhusiopathiae strains causing this disease is limited. In this study, we determined the serotypes, antimicrobial susceptibility, and potential mechanisms of resistance gene transfer in E. rhusiopathiae isolates (n = 60) from domestic geese and ducks. We also developed a multiplex PCR for the detection of resistance genes. The antimicrobial susceptibility of the isolates was assessed using the broth microdilution method. Resistance genes, integrative conjugative element (ICE)-specific genes, phage-specific genes, and serotype determinants were detected by PCR. Multilocus sequence typing (MLST) was performed for selected resistant strains. The comparative analyses included 260 E. rhusiopathiae strains whose whole genome sequences (WGSs) are publicly available. E. rhusiopathiae isolates represented 7 serotypes, among which serotypes 5 (38.3%) and 1b (28.3%) were the most common. All strains were susceptible to β-lactams, and the vast majority of them were resistant to tetracycline (85%) and enrofloxacin (80%). The percentages of isolates resistant to other antimicrobials used ranged from 3.3% to 16.7%. Ten isolates (16.7%) were found to be multidrug resistant (MDR). The genotypic resistance profiles of the E. rhusiopathiae strains corresponded to their phenotypic resistance, and the amplification patterns obtained using the 10-plex PCR developed in this study were fully consistent with the results of single PCRs. The most prevalent resistance gene was tetM. In enrofloxacin-resistant strains, nonsynonymous mutations in the gyrA and parC genes were identified. The presence of ICE-specific genes was confirmed in resistant strains, and in MDR isolates of serotype 8 that represented sequence type (ST) 113, prophage DNA (Javan630-like) linked to the lsaE gene was additionally detected. The results indicate that β-lactam antibiotics should be the first choice for the treatment of waterfowl erysipelas in Poland. ICEs, including a transposon from the Tn916/Tn1545 family, and bacteriophages are most likely responsible for the transfer of resistance genes in E. rhusiopathiae.

Characterization of autochthone biological control agents for pear protection against the brown spot of pear disease caused by Stemphylium vesicarium

AbstractStemphylium vesicarium is an important phytopathogen for many plant species, that switches between necrotrophic and saprophytic lifestyles. In the last decade, it has severely affected pear production worldwide, causing the so-called brown spot of pears (BSP), which led to the replacement of several pear orchards in northern Italy with consequent important economic losses. In the field, the management of the disease still relies on fungicide application, though several resistant genotypes of the pathogen emerged through the years. In the present study, three different bacterial species have been isolated from an infected orchard and evaluated as potential biological control agents (BCAs) against a local isolate of Stemphylium vesicarium. Two bacterial isolates belonging to the Bacillus genus demonstrated a significant ability to inhibit S. vesicarium growth in vitro, both through direct antagonism and the emission of volatile organic compounds (VOCs). The cell-free supernatant from one of these isolates exhibited a strong biocontrol activity, particularly through compounds extracted in acidic conditions. The protective effectiveness of the two most promising BCAs was further validated on detached pear fruits infected with S. vesicarium. Overall, the findings suggest that these two BCAs have the potential to be developed into a bio-based alternative to fungicides to control BSP, thus contributing to a more sustainable pear production.

Downy mildew of millets - An overview

Amidst global food insecurity, malnutrition, agricultural challenges, and climate change, millet farming is emerging as a viable alternativedue to its high nutritional value, resilience to extreme weather conditions, and adaptability to marginal soils. Often referred to as “super-crops”, millets gained international recognition in March 2021 during the 75th session of the United Nations General Assembly. Millets rank among the most important crops globally, following rice, wheat, maize, and sorghum. However, they are susceptible to more than fifty diseases, with the most destructive being ‘Downy Mildew’. This disease is caused by oomycete pathogens such as Sclerospora graminicola, Peronosclerospora sorghi and Sclerophthora macrospora, posing a significant threat to millet production, with yield losses ranging from 50% to 100%. This review emphasizes the importance of millet and the devastating effects of downy mildew on its yields. It explores the physiological and histological changes induced by the disease, the characterization of effector protein, and the genetic variability in millet populations. Additionally, various management techniques for combating downy mildew are examined, including chemical treatments, induced resistance, organic-based approaches, cultural practices, resistant genotypes, and advancements in nanotechnology. By compiling current knowledge on millet disease and effective management strategies, this review aims to serve as a comprehensive resource for researchers and farmers, supporting sustainable millet farming and improving global food security.

Evaluation of yield and stability of sugar beet (beta vulgaris L.) genotypes using GGE biplot and AMMI analysis

Rhizomania is the most destructive sugar beet disease in the world, and in recent years, it has widespread in most of the sugar beet growing areas in Iran. Since the control of this soil-borne disease is a difficult task, the use of resistant genotypes is known as the best measure against the disease. The ultimate goal of sugar beet breeders is to produce genotypes that can be used in both infected and non-infected fields without any reduction in terms of yield and quality. Twenty-one sugar beet genotypes along with four controls were evaluated in randomized complete block design with four replications in fields with natural infection to rhizomania in five research stations. Important sugar beet traits including root yield, sugar yield, sugar content, and white sugar yield were evaluated for two years (2021 and 2022). For all traits, location was the main source of variation that spanned 33 to 55% of the total sum of the square followed by the location×year×genotype accounted for 3–40% of the variation. Based on the results of analysis of variance, multivariate stability parameters were computed to evaluate the genotypes’ stability. The first two principal components (IPCA1 and IPCA2) generated by GGE biplot contributed for 31.3 and 17.5% difference in genotype×environment interaction for root yield, respectively. According to the GGE biplot, genotypes RM-11 and RM-12 were identified as the winning genotypes across environments for both root yield and white sugar yield traits whereas AMMI model identified RM-14 and RM-9 (for root yield) and RM-1 (for white sugar yield) as best genotypes. Based on the ideal genotype ranking, RM-11 and RM-10 were the best performer with a high mean yield as well as stability in the studied environments. The biplot rendered using the weighted average of absolute scores (WAASB) and root yield and white sugar yield identified RM-11 and RM-9 as superior genotypes in terms of yield and stability. The selected genotypes can be used in breeding programs to transfer the disease resistance and cultivar development.

Rising orthotospovirus incidence in India: Challenges and advances in management

Orthotospoviruses (OV) have emerged as a significant agricultural threat in India, posing a severe risk to critical crops, including tomato, chilli, watermelon and groundnut. This review explores the rising incidence of OV, including Groundnut Bud Necrosis Orthotospovirus (GBNV), Watermelon Bud Necrosis Orthotospovirus (WBNV), Capsicum Chlorosis Orthotospovirus (CaCV), Peanut Yellow Spot Orthotospovirus (PYSV), Iris yellow spot Orthotospovirus (IYSV) and Tomato Spotted Wilt Orthotospovirus (TSWV), along with the challenges in their management. These viruses have led to severe yield losses, sometimes causing complete crop failure due to their wide host range and the polyphagous nature of thrips. This further complicates control efforts by facilitating rapid spread across diverse crops and regions. The review highlights the multifaceted challenges in managing OV and thrips, including the lack of durable host resistance, limited diagnostic capabilities, and difficulties in controlling thrips populations. Current management strategies, including cultural practices, chemical control, biological control and resistant genotype development, have shown limited efficacy in providing long-term solutions. Recent advancements in biotechnological approaches, such as RNA interference, are discussed as promising pathways for improved virus management. The review underscores the need for genome editing techniques, such as CRISPR/Cas9, which offer the capacity to develop virus-resistant plants by targeting essential viral or vector genes to disrupt transmission cycles. Additionally, using nanoparticles for targeted delivery of antiviral compounds and novel detection tools presents another innovative solution to effectively mitigate the impact of OV on Indian agriculture.

Characterizing Escherichia coli carrying plasmid-mediated AmpC β-lactamases to optimize detection in a diagnostic laboratory setting.

Plasmid-mediated AmpC β-lactamases are a cause of acquired cephalosporin resistance in Gram-negative bacteria. However, consensus regarding the optimal detection method is yet to be achieved and varies depending on local epidemiology and laboratory capacity. We determined the acquired genotypic resistance mechanisms of 250 Escherichia coli isolates with a positive AmpC screen, defined as cefoxitin MIC ≥ 8 mg/L and a positive AmpC double- disc diffusion test, using in-house designed high-resolution melt PCR, detecting plasmid-acquired genes from the CIT and DHA families. A proportion of these isolates (n = 170, 68%) underwent further genotypic characterization using whole- genome sequencing (WGS). Of 250 isolates with a positive screening test, 72 (28.8%) were determined to carry an acquired AmpC gene. There was 100% concordance between PCR and WGS in the identification of acquired AmpC genes. The phenotypic criteria were then assessed to determine their utility in predicting acquired AmpC gene carriage. Criteria 1 (cefoxitin MIC > 8 mg/L plus ceftriaxone MIC > 1 mg/L and/or ceftazidime MIC > 1 mg/L) yielded a sensitivity of 93.1% and a specificity of 47.8%. Criteria 2 (cefoxitin MIC > 16 mg/L plus ceftriaxone MIC > 4 mg/L) had a sensitivity of 33.3% and a specificity of 98.9%. DHA genes, whose expression may be induced following antibiotic exposure, were present in 19% of isolates testing susceptible to ceftriaxone (MIC ≤ 1 mg/L) and were significantly more likely than CIT genes to be detected in susceptible isolates (P < 0.0001). These findings highlight the importance of using genotypic methods to detect acquired AmpC resistance in E. coli isolates that meet phenotypic screening criteria.IMPORTANCEDetection of transmissible AmpC resistance remains a challenging problem for diagnostic laboratories, especially in Escherichia coli where the expression of its intrinsic AmpC gene can result in phenotypic resistance patterns indistinguishable from plasmid-mediated resistance. In conjunction with whole- genome sequencing (WGS), we describe the development and performance of a novel melt-curve PCR to identify the two most prevalent plasmid-mediated AmpC gene families: CIT and DHA. We then describe phenotypic testing algorithms that incorporate this PCR and can differentiate these from non-acquired resistance in E. coli. It is important to distinguish these, not only to spare patients from unnecessarily being treated with infection control precautions, but also to identify plasmid-mediated genes, especially of the DHA family, that have been associated with inducible drug resistance to third- generation cephalosporins.

Machine Learning for Prediction of Resistance Scores in Wheat (Triticum aestivum L.)

ABSTRACTMachine learning methods were shown to improve the prediction accuracies of genomic prediction of resistance scores compared to methods like RR‐BLUP, which were originally designed for metric rather than ordinal response values. We conducted a cross‐validation study with 361 wheat genotypes evaluated for five fungal diseases. Our objective was to compare the prediction accuracy and the ability to identify the most resistant genotypes of 19 genomic prediction approaches. Each approach consisted of a different combination of prediction method (RR‐BLUP, an alternative method with heterogeneous marker variances, Bayesian generalized linear regression with an ordinal response, support vector machine, gradient boosting machine and random forest), predictor (single SNP markers, LD‐based haplotype blocks, 250 variables generated with an autoencoder and SNPs identified with incremental feature selection) and response value (untransformed and logit‐transformed resistance scores). In our dataset, RR‐BLUP was consistently among the methods with the largest prediction accuracies and the best abilities to identify resistant genotypes in four of five investigated traits. However, in P. triticina, using gradient boosting machine and random forest instead of RR‐BLUP increased the prediction accuracy from 0.64 to 0.71, indicating that machine learning methods may have an advantage over linear models in genomic prediction. We also found that even though there was a positive correlation between the prediction accuracy and Cohen's , a measure to judge how well the most resistant genotypes can be identified, the correlation is not perfect and a large value for the prediction accuracy does not necessarily translate into an equally large value.

Genome‐Wide Association Mapping and Genomic Prediction of Septoria nodorum Blotch Resistance in Central European Winter Wheat (Triticum aestivum L.)

ABSTRACTSeptoria nodorum blotch (SNB) is a fungal disease of wheat caused by the necrotrophic fungus Parastagonospora nodorum (Berk.), considered as one of the most devastating fungal diseases affecting winter wheat (Triticum aestivum L.). The complex inheritance of resistance to SNB poses significant challenges to breeding programmes. Improving selection precision and identifying novel resistance QTLs are crucial for enhancing SNB resistance. This study investigated strategies for crop genetic improvement, including genome‐wide association mapping (GWAS) and genomic prediction (GP), within a practical breeding programme. A population of 1500 winter wheat breeding lines was phenotyped for SNB resistance over 5 years across 19 geographical locations under natural infection conditions. Despite highly unbalanced breeder's data, medium to high heritabilities for SNB resistance were achieved. GWAS identified 11 significant marker‐trait associations for SNB resistance across Chromosomes 2A, 2B, 4B, 4D, 5D and 7B. GP fivefold cross‐validation analysis revealed a predictive ability of 0.52 for SNB resistance. The resistant wheat genotypes and SNP markers identified in this study will be valuable assets for future breeding efforts to enhance SNB resistance in wheat.

YgiV promoter mutations cause resistance to cystobactamids and reduced virulence factor expression in Escherichia coli

Antimicrobial resistance is one of the major health threats of the modern world. Thus, new structural classes of antimicrobial compounds are needed in order to overcome existing resistance. Cystobactamids represent one such new compound class that inhibit the well-established target bacterial type II topoisomerases while exhibiting superior antibacterial and resistance-breaking properties. Understanding potential mechanisms of emerging resistances is crucial in the development of novel antibiotics as they directly impact the future therapeutic application and market success. Therefore, the frequency and molecular basis of cystobactamid resistance in Escherichia coli was analyzed. High-level resistant E. coli mutants were selected and found to harbor single nucleotide polymorphisms in the promotor region of the ygiV gene, causing an upregulation of the respective protein. These stable mutations are contrary to what was observed as a resistance genotype in the structurally related albicidins, where ygiV gene amplifications were identified as causing resistance. Overexpression of YgiV in the mutants was additionally amplified upon cystobactamid exposition, showing further adaptation to this compound class under treatment. YgiV binds cystobactamids with high binding affinity, thereby preventing their interaction with the antimicrobial targets topoisomerase IV and DNA gyrase. In addition, we observed a substantial impact of YgiV on in vitro gyrase activity by leading to increased DNA cleavage and concurrent reduction in the efficacy of cystobactamids in inhibiting gyrase supercoiling activity. Furthermore, we identified co-upregulation of membrane-modifying proteins, such as EptC, and the transcriptional regulator QseB. This presumably contributes to the observed reduced motility and fimbrial protein expression in resistant mutants, resulting in a reduced expression of virulence factors and potentially pathogenicity, associated with ygiV promotor mutations.

Evaluation of genetic diversity and genome-wide association studies of resistance to bacterial wilt disease in potato.

The development of novel improved varieties adapted to unstable environmental conditions is possible through the genetic diversity of breeding materials. Potato is among the most important food crops worldwide, however, there are still significant hindrances to breeding gains attributed to its autotetraploid and highly heterozygous genome. Bacterial wilt caused by the Ralstonia solanacearum species complex (RSSC) is an important disease affecting potato among many economically important crops worldwide. No cultivated potato genotypes have shown a satisfactory level of resistance to bacterial wilt. Nevertheless, resistance can play a crucial role in effective integrated disease management. To understand the genetic landscape of bacterial wilt resistance in cultivated potato, we evaluated the diversity of 194 accessions from the International Potato Centre (CIP) using 9,250 single nucleotide polymorphisms (SNPs) and their associations to the response to bacterial wilt disease evaluated over two independent trials. Twenty-four accessions showed high resistance throughout both trials. Genetic diversity analysis revealed three major clusters whose subgroups were mostly represented by CIP clones derived from common parents. Genome-wide association analyses have shown six major hits: two on chromosome 8, and one on each chromosome 2, 4, 5, and 9. These results facilitate genetic dissection of bacterial wilt resistance and marker-enabled breeding in elite genotypes for potato breeding initiatives.

Molecular Confirmation for Gall Midge (Biotype 3) Resistance in Phenotypically Resistant Rice Genotypes Using Functional Markers

This study performed molecular screening of rice genotypes, which showed no gall infestation in field conditions at the Regional Agricultural Research Station (RARS), Jagtial, during rabi, 2021-22. using functional markers such as “gm3del3” for the gm3 gene, “LRR del” for the Gm4 gene, and “PRP” for the Gm8 gene, six genotypes were analyzed. WGL-1145 contained all three resistance genes (gm3, Gm4, and Gm8). Additionally, three genotypes namely WGL-1147, WGL-1127, and Kakai had the Gm4 and Gm8 genes, while RP-5332-54-11-8-2-13 had only Gm8 and IR72476-B-P-9-3-1-1 possessed only gm3. These findings contribute to developing new rice varieties with resistance to gall midge, which can improve crop yields and food security in affected regions. Some promising rice varieties may be used as donors in breeding programs aimed at creating pyramided lines with durable gall midge resistance.

Response to oxalic acid: an important supplement screening against stem rot resistance in groundnut (Arachis hypogaea L.)

BackgroundStem rot, caused by the soil-borne pathogen Sclerotium rolfsii, pose a serious challenge in the groundnut (Arachis hypogaea L) cultivation. Although this disease is widespread globally but had most adverse impact in groundnut growing regions of United States, India, and Australia. The pathogen primarily targets the crown region of the plant, resulting in systemic collapse and potentially leading to yield losses up to 80%. Effective genetic control measures are essential to mitigate the impact of this disease on groundnut production. Realizing the time and resource-consuming complex field-based phenotyping, the availability of easy and repeatable phenotyping methods may fasten the process of donor and gene discovery efforts.ResultsMulti-season phenotyping was performed for stem rot on 184 minicore germplasm accessions, including checks, under two conditions: sick field screening and response to oxalic acid assay. This study demonstrated medium to high heritability (52–63% broad-sense heritability) and significant environmental influence (36%). The response to the oxalic acid assay showed a high proportion of similarity (approximately 80%) with the percent mortality observed in the sick field indicating an easy way of performing precise phenotyping. Notably, seven genotypes—ICG163, ICG721, ICG10479, ICG875, ICG11457, ICG111, and ICG2857—exhibited stable resistance, with less than 30% mortality against stem rot disease. Among these, ICG163, ICG875, and ICG111 displayed low mortality and consistent stability across multiple seasons in both the sick field and controlled conditions of the oxalic acid assay.ConclusionsThe oxalic acid assay developed in this study effectively complements field phenotyping, as a reliable method for assessing stem rot resistance. Seven resistant genotypes identified through this assay can be utilized for the introgression of stem rot resistance into elite genotypes. Given the significant influence of the environment on stem rot resistance, it is essential to implement multi-season phenotyping to obtain precise results. Furthermore, the response to oxalic acid serves as a valuable supplement to traditional field phenotyping, since maintaining uniform disease pressure during field screenings is often challenging.

Metabolite Profiling of Annual Ryegrass Cultivars to Assess Disease Resistance and Susceptibility.

Direct immersion solid-phase microextraction coupled with gas chromatography-mass spectroscopy was used to create chemical fingerprints of annual ryegrass cultivars (Lolium rigidum). Extracts made of the inflorescences of four cultivars and one accession of annual ryegrass were assessed to identify differential metabolites between those resistant to and susceptible to bacterial galls associated with annual ryegrass toxicity (ARGT). Numerous compounds were identified. Principal component analysis showed distinct clustering of metabolites from disease-resistant and disease-susceptible cultivars. Partial least-squares-discriminant analysis identified sterols, esters, aldehydes, and terpenes that correlated with resistance to galls formation. Esters, sterols, phenols, heterocyclics, fatty acids, organofluorides, and siloxanes were predominant in resistant genotypes, whereas alcohols, aldehydes, terpenes, and hydrocarbons were predominant in susceptible genotypes. The identification of differentially expressed metabolites provides potential chemical markers to guide breeding strategies for ARGT resistance in ryegrass.

Population structure and markers analysis associated with powdery mildew resistance in pumpkin, Cucurbita pepo L

Pumpkin (Cucurbita pepo L.) is an economically important vegetable and summer crop. The seeds have high nutritional value due to their edible oil and protein content with a high medicinal value. Pumpkin is attacked by several fungal diseases; one of which is powdery mildew (PM) caused by Podosphaera xanthii race-0. PM causes severe disease on pumpkin leading to major yield losses globally. In this study, we analyzed the genetic diversity and population structure of fourteen local pumpkin cultivars, all under Cucurbita pepo L. based on dominance in the region using 15 Inter-Simple-Sequence-Repeats-(ISSR) primer-pairs in association with resistance against PM as well as regulating the expression of three resistant marker genes i.e. SGT1, bHLH and PSK via qRT-PCR at three stages with 10 days' interval post-inoculation-stages-(SPI) with the powdery mildew agent, P. xanthii race-0. The results showed that ISSR markers demonstrated 13 primers out of 15 primers generated 169 distinct amplified bands, out of which 143 bands showed polymorphism. The mean polymorphism information content (PIC), marker index (MI), effective-number of alleles (I), expected heterozygosity (H), and Nei's gene diversity parameters were 0.354, 3.344, 0.334, 0.205 and 0.200, respectively. According to PIC, MI, I, H, and Nei indices evaluation, ISSR10 primer had the best performance in genetic diversity of the related pumpkin population. NTSYS pc 2.02e software and several similarity coefficients (r = 0.964), placed the related genotypes into five distinct groups. Of which, ‘Maragheh1′ and ‘Shahreza’ as resistant ones with 9.4 % and 6.1 % disease severity (DS), and ‘Gorgan’, ‘Urmia1′ and 3 and ‘Maragheh3’ with 79.6–84.8 % DS, as susceptible ones into two distinct groups, respectively. Further, the marker genes SGT1, bHLH and PSK expression level changes showed that the effect of variable factors including different genotypes (susceptible and resistant), disease-causing fungus, SPI, and the mutual effect of these factors is significant with the highest increase in resistant genotypes. Considering the relative compatibility in the classification of pumpkin genotypes in the dendrograms of DS and ISSR markers, it can be expected that increasing the number of markers in evaluating the genetic diversity of these genotypes makes it possible to differentiate pumpkin genotypes resistant and susceptible to PM disease based on the degree of disease resistance. The markers associated with resistance are recommended for use in marker assisted breeding.

Genotypic Antimicrobial Resistance Profiles of Diarrheagenic Escherichia coli and Nontyphoidal Salmonella Strains Isolated from Children with Diarrhea and Their Exposure Environments in Ethiopia.

Antimicrobial resistance (AMR) poses a significant global threat, particularly in low- and middle-income countries, such as Ethiopia, where surveillance is limited. This study aimed to predict and characterize the AMR profiles of diarrheagenic Escherichia coli (DEC) and nontyphoidal Salmonella (NTS) strains isolated from human, animal, food, and environmental samples using whole genome sequencing. A total of 57 NTS and 50 DEC isolates were sequenced on an Illumina NextSeq 550. The ResFinder and PointFinder tools were employed to identify antimicrobial resistance genes (ARGs) and point mutations. Salmonella serotypes were determined using SeqSero. The analysis identified at least one ARG in every NTS sample and 78% of the DEC isolates, with 22 distinct ARGs in the NTS samples and 40 in the DEC samples. The most prevalent ARGs were aac(6')-Iaa and aph(3')-Ib, which predict aminoglycoside resistance in 100% of NTS and 54% of DEC isolates, respectively. Other commonly identified ARGs include sul2, aph(6)-Id, blaTEM-1B , and tet(A), which confer resistance to folate inhibitors, aminoglycosides, β-lactams, and tetracycline. Some ARGs predicted phenotypic multidrug resistance in both DEC and NTS isolates. All identified β-lactam ARGs, except for blaTEM -1D, conferred resistance to more than three antibiotics. Interestingly, blaCTX- M-15 was found to confer resistance to nine antibiotics, including third-generation cephalosporins, in 18% of DEC and 3.5% of NTS isolates. DEC isolates from children exhibited the highest ARG diversity. Notably, genes such as aph(3″)-Ib, aph(6)-Id, sul2, and tet(A) were detected across all sample types, including water sources, although some ARGs were exclusive to specific sample types. Point mutations mediating AMR were detected in several genes, with mutations associated with nucleotide substitution being the most frequent. This genotypic AMR profiling revealed the presence of widespread drug-resistant NTS and DEC strains in Ethiopia. Robust and sustained AMR surveillance is essential for monitoring the emergence and spread of these resistant pathogens.

Development of rapid, efficient and cost effective screening technique for testing arecanut against Phytophthora meadii incitant of fruit rot disease

To accelerate identification of disease resistant arecanut germplasm or hybrids against Phytophthora, it is very much imperative to develop bioassays which could differentiate resistant and susceptible cultivars efficiently. Here, developed a cost effective and rapid technique called “Detached Leaf Assay” to identify resistant germplasm at the seedling stage itself. Standardized zoospore production in highly virulent Phytophthora meadii (P19) by incubating under 12 hours light and dark regime. Zoospore suspension adjusted to 105spores ml-1 in petri plates. Subsequently, surface sterilized arecanut leaves were floated in zoospore suspension and incubated at temperature of 24±1°C. Disease symptoms, including water-soaked lesions, were recorded three days after inoculation. Infection lesion increased from 1 to 7.3 cm2. The pathogen was re-isolated and confirmed with the original culture. The assay was successfully validated to screen arecanut accessions, wild types and hybrids against P. meadii. This technique is the first to be developed, and it is simple, cost-effective, and faster. It also provides consistent infection and could be effectively utilized to screen arecanut germplasm or hybrids against P. meadii in the seedling stage itself.• Developed a cost effective, efficient and rapid screening technique• The technique was validated to identify resistant genotypes against Phytophthora meadii at the seedling stage.

The effect of next-generation, dual-active-ingredient, long-lasting insecticidal net deployment on insecticide resistance in malaria vectors in Benin: results of a 3-year, three-arm, cluster-randomised, controlled trial

Insecticide resistance among malaria vector species now occurs in 84 malaria-endemic countries and territories worldwide. Novel vector-control interventions, including long-lasting insecticidal nets (LLINs) that incorporate new active ingredients with distinct modes of action, are urgently needed to delay the evolution and spread of resistance and to alleviate reversals in malaria-control gains. We aimed to assess the longitudinal effect of two dual-active-ingredient LLINs on insecticide resistance during a cluster-randomised, controlled trial in Benin. This 3-year, three-arm, cluster-randomised, controlled trial was conducted between Oct 17, 2019, and Oct 24, 2022, in three districts in southern Benin, to compare the effects of LLINs containing chlorfenapyr-pyrethroid or pyriproxyfen-pyrethroid with LLINs containing pyrethroid only. In 19 292 mosquitoes (Anopheles gambiae sensu lato) collected over 36 months-3 months of baseline followed by 3 years post-intervention-we measured longitudinal phenotypic insecticide resistance profiles using bioassays and genotypic resistance profiles using quantitative, real-time, reverse transcriptase PCR of metabolic resistance genes in two clusters per trial group. The trial was registered with ClinicalTrials.gov, NCT03931473. In all three trial groups, a significant effect of LLINs on insecticide resistance selection was evident, with the median lethal dose (LD50) of α-cypermethrin approximately halving between baseline and 12 months post-LLIN distribution (pyrethroid-only LLIN cluster 21: LD50 78·78 μg/ml [95% CI 65·75-94·48] vs 35·93 [29·41-43.86] and cluster 31: 79·26 [65·40-96·44] vs 38·71 [30·88-48·53]; chlorfenapyr-pyrethroid LLIN cluster 43: 104·30 [82·97-133·58] vs 43·99 [35·30-54·86]; and pyriproxyfen-pyrethroid LLIN cluster 36: 63·76 [52·14-77·75] vs 37·96 [30·88-46·69] and cluster 53: 77·67 [57·63-104·56] vs 39·72 [29·26-53·97]). Over the subsequent 2 years, the LD50 of α-cypermethrin increased past baseline values in all three trial groups (year 3 pyrethroid-only LLIN cluster 21: 141·01 [111·70-181·90] and cluster 31: 115·15 [93·90-143·09]; chlorfenapyr-pyrethroid LLIN cluster 43: 97·00 [77·24-123·54] and cluster 55: 126·99 [102·34-161·26]; and pyriproxyfen-pyrethroid LLIN cluster 36: 142·29 [112·32-184·84] and cluster 53: 109·88 [79·31-157·70]). We observed minimal reductions in chlorfenapyr susceptibility and variable but significant reductions in fertility after pyriproxyfen exposure, with an overall trend of increasing susceptibility across trial years. Several metabolic genes were implicated in resistance selection, including CYP6P4 in the pyriproxyfen-pyrethroid LLIN group, which encodes an enzyme known to metabolise pyriproxyfen in vitro, and CYP6P3 and CYP9K1 in the chlorfenapyr-pyrethroid LLIN group, both of which encode enzymes that are involved in pro-insecticide activation. After 24 months of use, chlorfenapyr-pyrethroid LLINs no longer mitigated pyrethroid resistance selection in this area of southern Benin, which has high malaria transmission dominated by highly resistant A gambiae sensu lato. This finding raises issues for current net-procurement schedules, which are based on an operational net lifespan of 3 years. Knowledge of the effects of next-generation LLINs on insecticide-resistance selection is crucial for the pragmatic design of prospective resistance-management strategies. UNITAID and The Global Fund to Fight AIDS, Tuberculosis and Malaria.