Levels Of Phenotypic Resistance Research Articles

Phenotypic and Genotypic Characterization of Antimicrobial Resistance in Salmonella enterica Serovars from Colombian Pig Farms

Salmonella enterica is a globally important zoonotic microorganism that affects pigs and can enter the farm through various routes. This study aimed to determine the prevalence of S. enterica in water sources and pigs at Colombian pig farms, and to characterize the antimicrobial resistance of the isolates phenotypically and genotypically. Samples were collected from 103 farms including source water (n = 104), storage tank water (n = 103), drinking water (n = 103), and individual rectal swab samples (n = 1025). The presence of Salmonella was detected/identified using MDS-3M™ agar culture medium. Isolates were serotyped, and their antibiotic susceptibility was determined by minimum inhibitory concentration (MIC). Whole genome sequencing (WGS) was performed using Illumina NovaSeq, and bioinformatics analysis focused on serovar confirmation, MLST determination, and resistance gene detection. The overall between-farm prevalence of Salmonella enterica including all types of samples was 52.4% (54/103), with 6.4% of rectal swab samples and 21.3% of water samples found to be positive. Thirty serovars were identified using WGS, with the most common being S. Typhimurium var. monophasic (1,4,[5],12:i:-) (41.2%), S. Schwarzengrund (4.2%), and S. Saintpaul (4.2%). Salmonella Typhimurium and its monophasic variant were more commonly found in rectal swabs than the remaining serotypes (relative risk = 2.9, p < 0.0001), which were commonly found in the water samples (relative risk = 5.2, p < 0.0001). High levels of phenotypic resistance were observed, particularly to amikacin (99.2%), tetracycline (59.7%), chloramphenicol (55.5%), and ampicillin (42%). All isolates carried genes conferring resistance to aminoglycosides (aac(6′)-Iaa), quinolones (qnrB19), and tetracyclines (tetA). In conclusion, S. enterica is prevalent in Colombian pig farms including the water supply, with the S. Typhimurium monophasic variant being predominant, and antimicrobial resistance is widespread.

Phenotypic and genetic extended spectrum beta lactamase profiles of bacterial isolates from ICU in tertiary level hospital in Kenya

Background Bacterial infections in the Intensive Care Units are a threat to the lives of critically ill patients. Their vulnerable immunity predisposes them to developing bacteria-associated sepsis, deteriorating their already fragile health. In the face of increasing antibiotics resistance, the problem of bacterial infection in ICU is worsening. Surveillance of bacterial infections in ICUs and drug resistance will help to understand the magnitude of the problem it poses and inform response strategies. We assessed bacterial infections in ICU setting by identifying prevalent Gram-negative bacterial species and characterized their antibiotic susceptibility patterns. Methods Cross-sectional samples collected from Kenyatta National Hospital ICU between January and June 2021 were cultured and phenotypic identification of culture-positive samples performed using VITEK 2. Antibiotic susceptibility patterns were determined based on Antimicrobial Susceptibility Testing (AST) results. Cephalosporin-resistant Gram-negative bacteria were assessed by PCR to detect the presence of ESBL genes including ( bla CTX-M, bla SHV, bla TEM, bla OXA) Results and discussion Out of the 168 Gram-negative isolates, Acinetobacter baumanii was the most abundant (35%). Other isolates that were present at frequencies more than 15% are Klebsiella pneumoniae and Escherichia. coli. A. baumaniii is known to be a notorious bacterium in ICU due to its multidrug resistance nature. Indeed, A. baumanii isolates from Kenyatta National Hospital showed significantly high level of phenotypic resistance. Concordant with the high level of phenotypic resistance, we found high carriage of the ESBL genes among the isolates analysed in this study. Moreover, majority of isolates harboured all the four ESBL genes. Conclusion A high rate of phenotypic and genetic resistance was detected among the tested isolates. Resistance to cephalosporins was primarily driven by acquisition of the ESBL genes. The high prevalence rate of ESBL genes in ICU bacterial isolates shown in this study has a important implication for ICU patient management and general antibiotics use.

Occurrence and Antimicrobial Resistance of Staphylococcus aureus Isolated from Healthy Pet Rabbits.

Background: Staphylococcus aureus is a ubiquitous microorganism and an opportunistic pathogen responsible for numerous diseases in humans and animals, characterized by different clinical pictures with acute or subacute course. S. aureus, due to its great adaptability and versatility in terms of infections and hosts, can be considered a relevant pathogen because of the harmful effects on animal health and its potential for transmission from animals to humans and vice versa. In recent years, a marked increase in multidrug-resistant S. aureus has been reported, posing a serious threat for disease management, food safety, and animal and human health as they limit available therapeutic options. In light of a growing interest of the scientific community for this micro- organism and considering the limited data availability on the prevalence of this pathogen in pet rabbits, the purpose of this research was to evaluate the presence of S. aureus in pet rabbits. Materials and Methods: From November 2021 to December 2022, nasal swabs were collected from 50 pet rabbits from private households in the Campania Region, southern Italy, and underwent analysis for S. aureus detection. Samples were enriched in broth, then inoculated onto nutrient and selective media, including Blood agar base supplemented with 7% sheep blood and Baird-Parker Agar Base, following standard laboratory protocols. Incubations in aerobic conditions at 37°C were performed for 24/48h for colony identification. Antimicrobial susceptibility testing for all S. aureus isolates was conducted using the disc diffusion method. Results: Our results reported the presence of S. aureus in 16/50 (32%) rabbits examined, showing high levels of phenotypic resistance to different antibiotics, in particular penicillin 10U (81.2%) and erythromycin 15 μg (62.5%). Conclusion: The study demonstrated that pet rabbits represent a significant reservoir of S. aureus and contributes to the knowledge on the phenotypic antimicrobial resistance of these bacteria in rabbits raised in a domestic environment.

Multidrug resistance in pathogenic Escherichia coli isolates from urinary tract infections in dogs, Spain.

Escherichia coli (E. coli) is a pathogen frequently isolated in cases of urinary tract infections (UTIs) in both humans and dogs and evidence exists that dogs are reservoirs for human infections. In addition, E. coli is associated to increasing antimicrobial resistance rates. This study focuses on the analysis of antimicrobial resistance and the presence of selected virulence genes in E. coli isolates from a Spanish dog population suffering from UTI. This collection of isolates showed an extremely high level of phenotypic resistance to 1st-3rd generation cephalosporins, followed by penicillins, fluoroquinolones and amphenicols. Apart from that, 13.46% of them were considered extended-spectrum beta-lactamase producers. An alarmingly high percentage (71.15%) of multidrug resistant isolates were also detected. There was a good correlation between the antimicrobial resistance genes found and the phenotypic resistance expressed. Most of the isolates were classified as extraintestinal pathogenic E. coli, and two others harbored virulence factors related to diarrheagenic pathotypes. A significant relationship between low antibiotic resistance and high virulence factor carriage was found, but the mechanisms behind it are still poorly understood. The detection of high antimicrobial resistance rates to first-choice treatments highlights the need of constant antimicrobial resistance surveillance, as well as continuous revision of therapeutic guidelines for canine UTI to adapt them to changes in antimicrobial resistance patterns.

Characterization of emamectin benzoate resistance in the diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae).

Plutella xylostella (L.) is a destructive pest of cruciferous crops worldwide that has evolved resistance to many insecticides. Here we examined the mode of inheritance, cross-resistance profile, and potential mechanisms of emamectin benzoate resistance in a field-derived strain of P. xylostella from Japan. A field-collected population of P. xylostella, was found to exhibit strong (> 150-fold) resistance to emamectin benzoate in insecticide bioassays when compared with a laboratory susceptible strain. Genetic analysis showed that resistance is inherited as an autosomal, recessive trait, and is conferred by a single or a few closely linked loci. The emamectin benzoate resistant strain also exhibited resistance to abamectin, lepimectin, chlorantraniliprole, lufenuron, spinetoram, indoxacarb, fipronil, dieldrin, endosulfan and lambda-cyhalothrin, demonstrating a remarkable multi-resistance profile. Insecticide bioassays employing inhibitors of detoxification enzymes revealed that piperonyl butoxide (PBO) increased the toxicity of emamectin benzoate in the resistant strain by ten-fold indicating the potential involvement of cytochrome P450 monooxygenases in avermectin resistance. Furthermore, cloning and sequencing of the primary receptor of avermectins, the GluCl channel, revealed the absence of target-site mutations in the resistant strain. Our data on the mode of inheritance and mechanisms of resistance to emamectin benzoate in a P. xylostella strain from Japan provide a foundation for the development of regional resistance management strategies. However, the high levels of phenotypic resistance in this strain to a diverse range of other insecticide classes available for control illustrate the challenges associated with the sustainable control of this important pest. © 2023 Society of Chemical Industry.

The Detection of Mutations and Genotyping of Drug-Resistant Mycobacterium tuberculosis Strains Isolated from Patients in the Rural Eastern Cape Province.

Drug-resistant tuberculosis (DR-TB) is still a major public health concern in South Africa. Mutations in M. tuberculosis can cause varying levels of phenotypic resistance to anti-TB medications. There have been no prior studies on gene mutations and the genotyping of DR-TB in the rural Eastern Cape Province; hence, we aimed to identify DR-TB mutations, genetic diversity, and allocated lineages among patients in this area. Using Xpert® MTB/RIF, we assessed the rifampin resistance of sputum samples collected from 1157 patients suspected of having tuberculosis. GenoType MTBDR plus VER 2.0 was used for the detection of mutations causing resistance to anti-TB medications. The next step was to spoligotype 441 isolates. The most prevalent rifampin resistance-conferring mutations were in rpoB codon S531L in INH-resistant strains; the katG gene at codon S315TB and the inhA gene at codon C-15TB had the most mutations; 54.5% and 24.7%, respectively. In addition, 24.6% of strains showed mutations in both the rpoB and inhA genes, while 69.9% of strains showed mutations in both the katG and rpoB genes. Heteroresistance was seen in 17.9% of all cases in the study. According to spoligotyping analysis, Beijing families predominated. Investigation of the evolutionary lineages of M. tuberculosis isolates can be carried out using the information provided by the study's diversity of mutations. In locations wherein these mutations have been discovered, decision-making regarding the standardization of treatment regimens or individualized treatment may be aided by the detection frequency of rpoB, katG, and inhA mutations in various study areas.

Antimicrobial Susceptibility and Resistance Mechanisms in Mannheimia haemolytica Isolates from Sheep at Slaughter.

Mannheimia haemolytica is the main pathogen contributing to pneumonic pasteurellosis in sheep. The aim of this study was to investigate the antimicrobial resistance levels in M. haemolytica isolates from the lungs of slaughtered sheep and to examine the genetic resistance mechanisms involved. A total of 256 M. haemolytica isolates, 169 from lungs with pneumonic lesions and 87 from lungs without lesions, were analyzed by the disk diffusion method for 12 antimicrobials, and the whole genome of 14 isolates was sequenced to identify antimicrobial resistance determinants. Levels of phenotypic resistance ranged from <2% for 10 antimicrobials (amoxicillin, amoxicillin-clavulanic, ceftiofur, cefquinome, lincomycin/spectinomycin, gentamicin, erythromycin, florfenicol, enrofloxacin, and doxycycline) to 4.3% for tetracycline and 89.1% for tylosin. Six isolates carried tetH genes and four isolates carried, in addition, the strA and sul2 genes in putative plasmid sequences. No mutations associated with macrolide resistance were identified in 23 rDNA sequences, suggesting that the M. haemolytica phenotypic results for tylosin should be interpreted with care in the absence of well-established epidemiological and clinical breakpoints. The identification of strains phenotypically resistant to tetracycline and of several resistance genes, some of which were present in plasmids, highlights the need for continuous monitoring of susceptibility patterns in Pasteurellaceae isolates from livestock.

Phenotypic and genetic extended spectrum beta lactamase profiles of bacterial isolates from ICU in tertiary level hospital in Kenya

Background: Bacterial infections in the ICUs are a threat to the lives of critically ill patients. Moreover, their vulnerable immunity predisposes them to developing bacteria-associated sepsis, further deteriorating their already fragile health. In the face of increasing drug resistance menace, the problem of bacterial infection in ICU is likely to worsen. This study sought to assess bacterial infections in ICU setting by identifying prevalent gram-negative bacterial species and characterizing their antibiotic susceptibility patterns. Phenotypic and genetic resistance were determined among gram-negative isolates from ICU patients. Methods: Cross-sectional samples collected from Kenyatta National Hospital ICU between January and June 2021 were cultured for subsequent analysis. Phenotypic identification of culture-positive samples was performed using VITEK 2 while Antibiotic susceptibility patterns were determined based on Antimicrobial Susceptibility Testing (AST) results. Cephalosporin-resistant gram-negative bacteria were assessed by PCR to detect the presence of ESBL genes. Results and discussion: Out of the 168 gram-negative isolates, Acinetobacter baumanii was the most abundant (35%). Other isolates that were present at frequencies more than 15% are Klebsiella pneumoniae and Escherichia. coli. A. baumaniii is known to be a notorious bacterium in ICU due to its multidrug resistance nature. Indeed, A. baumanii isolates from Kenyatta National Hospital showed significantly high level of phenotypic resistance. Concordant with the high level of phenotypic resistance, we found high carriage of the ESBL genes among the isolates analysed in this study. Moreover, majority of isolates harboured all the four ESBL genes. Conclusion: A high rate of phenotypic and genetic resistance was detected among the tested isolates. Resistance to cephalosporins was primarily driven by acquisition of the ESBL genes. The high-rate multidrug resistance in ICU bacterial isolates in this study has a crucial implication for ICU patient management and general antibiotics use.

Antibiotic resistance of microorganisms isolated from cows with subclinical mastitis

Cow mastitis is a polyetiological disease involving pathogenic microorganisms. Antibiotics are used to treat sick animals, which contributes to the development of resistant clones of bacteria, and consumption of the corresponding milk leads to the spread of antibiotic-resistant microorganisms. The purpose of the work is to determine strains of antibiotic-resistant bacteria in the milk of cows suffering from subclinical mastitis. Bacteriological studies of mammary gland secretion samples were performed by inoculating the sediment obtained after centrifugation of the examined material on an agarised nutrient medium (with 5% ram erythrocytes). The study of antimicrobial sensitivity was performed by the disk diffusion method. It was established that pathogens with multiple resistance to antibiotics circulate among cows suffering from mastitis on the farm, which indicates their indiscriminate use in animal therapy schemes, in particular mastitis. It was found that the overall level of phenotypic resistance to antimicrobial agents of the examined isolates was within 19-33% (in Staphylococcus spp. – 19.9%, in E. coli – 33.2%, in Enterococcus spp. – 32.5%). The analysis of the results allowed assigning the culture study to the category of multiresistant: 31.6% of cultures of Staphylococcus spp. demonstrated resistance to five groups of antibiotics; 54.5% of Escherichia cultures – up to five groups of antibiotics; 37.5% of cultures of Enterococcus spp. resistant to three groups of antibiotics. It was established that 100% of the studied cultures of Staphylococcus spp. are sensitive to netilmicin, sparfloxacin, gatifloxacin and clarithromycin. Therewith, 100% of the examined cultures of Enterococcus spp. sensitive to gentamicin, norfloxacin, and nitrofurantoin, and 100% of the examined E. coli cultures were sensitive to cephalothin, meropenem, gentamicin, tobramycin, amikacin, nitroxoline, and gatifloxacin. Thus, dairy cows suffering from subclinical mastitis are a source of bacteria with multiple resistance to antibiotics and carry risks for consumers due to the consumption of dairy products contaminated with pathogens of zoonotic diseases, which requires proper control by veterinary service specialists

Antibiotic resistance and biofilm formation in Staphylococcus aureus isolated from dairy cows at the stage of subclinical mastitis in northern Kazakhstan.

Staphylococcus aureus is an important causative agent of subclinical bovine mastitis worldwide. The aim of this research was to study the ability of S. aureus to form biofilms. Additionally, we examined the genes involved in cell resistance and sensitivity to antibiotics. Samples were collected from December 2020 to May 2021 from Simmental and black-and-white cows. The study was carried out on a total number of 643cows, of which 278 (23 %) were in the subclinical mastitis stage. Finally, 64 S. aureus isolates were isolated and identified. The highest level of phenotypic resistance was observed to antibiotics of the tetracycline (tetracycline - 48.4 %, doxycycline - 32.8 %) and -lactam (ampicillin - 45.3 %, penicillin - 45.3 %) groups. The genes encoding antibiotic resistance were characterized with the polymerase chain reaction method: blaZ in 30isolates, mecA in 1isolate, ermC in 15isolates, aph (3) in 2isolates, tetK in 19isolates, tetM in 9isolates. The tested S. aureus isolates had the ability to form biofilms in 76.6 % ( ) of cases. Of these, 69.4 % were resistant to at least one antibiotic. The obtained results have shown that S. aureus, identified in cows with subclinical mastitis, was resistant mainly to tetracycline and -lactam antibiotics. In addition, S. aureus isolates expressed resistance genes to the above drugs and had the ability to form biofilm. This study will help to identify the extent of antibiotic resistance and monitor S. aureus contamination of raw milk.

Taxonomic position, antibiotic resistance and virulence factor production by Stenotrophomonas isolates from patients with cystic fibrosis and other chronic respiratory infections

BackgroundThe potential pathogenic role of Stenotrophomonas maltophilia in lung disease and in particular in cystic fibrosis is unclear. To develop further understanding of the biology of this taxa, the taxonomic position, antibiotic resistance and virulence factors of S. maltophilia isolates from patients with chronic lung disease were studied.ResultsA total of 111 isolates recovered between 2003 and 2016 from respiratory samples from patients in five different countries were included. Based on a cut-off of 95%, analysis of average nucleotide identity by BLAST (ANIb) showed that the 111 isolates identified as S. maltophilia by Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/MS) belonged to S. maltophilia (n = 65), S. pavanii (n = 6) and 13 putative novel species (n = 40), which each included 1–5 isolates; these groupings coincided with the results of the 16S rDNA analysis, and the L1 and L2 ß-lactamase Neighbor-Joining phylogeny. Chromosomally encoded aminoglycoside resistance was identified in all S. maltophilia and S. pavani isolates, while acquired antibiotic resistance genes were present in only a few isolates. Nevertheless, phenotypic resistance levels against commonly used antibiotics, determined by standard broth microbroth dilution, were high. Although putative virulence genes were present in all isolates, the percentage of positive isolates varied. The Xps II secretion system responsible for the secretion of the StmPr1–3 proteases was mainly limited to isolates identified as S. maltophilia based on ANIb, but no correlation with phenotypic expression of protease activity was found. The RPF two-component quorum sensing system involved in virulence and antibiotic resistance expression has two main variants with one variant lacking 190 amino acids in the sensing region.ConclusionsThe putative novel Stenotrophomonas species recovered from patient samples and identified by MALDI-TOF/MS as S. maltophilia, differed from S. maltophilia in resistance and virulence genes, and therefore possibly in pathogenicity. Revision of the Stenotrophomonas taxonomy is needed in order to reliably identify strains within the genus and elucidate the role of the different species in disease.

Levels of phenotypic resistance and the pattern of gene mutations of M. tuberculosis

The adjustment of the minimum inhibitory concentrations for anti-tuberculosis drugs used in classical cultivation methods and the obtaining of more accurate data on the level of drug resistance, makes it possible to evaluate cases with different results to phenotypic and genotypic methods of anti-tuberculosis resistance testing. The evaluation of different types of genotypic resistance, with the description of mutations that confer low or high resistance, is consistent with the level of phenotypic resistance, and also makes it possible to adjust treatment regimens, which ultimately will positively influence the duration and results of treatment. Whenever testing of M. tuberculosis resistance by molecular methods allows, the results should be reported with the specific mutations detected and the description of the clinical implications of this mutation.

Genome-wide association mapping for resistance to bacterial blight and bacterial leaf streak in rice.

Using genome-wide SNP association mapping, a total of 77 and 7 loci were identified for rice bacterial blight and bacterial leaf streak resistance, respectively, which may facilitate rice resistanceimprovement. Bacterial blight (BB) and bacterial leaf streak (BLS) caused by Gram-negative bacteria Xanthomonas oryzae pv. oryzae (Xoo) and X. oryzae pv. oryzicola (Xoc), respectively, are two economically important diseases negatively affecting rice production. To mine new sources of resistance, a set of rice germplasm collection consisting of 895 re-sequenced accessions from the 3000 Rice Genomes Project (3K RGP) were screened for BB and BLS resistance under field conditions. Higher levels of BB resistance were observed in aus/boro subgroup, whereas the japonica, temperate japonica and tropical japonica subgroups possessed comparatively high levels of resistance to BLS. A genome-wide association study (GWAS) mined 77 genomic loci significantly associated with BB and 7 with BLS resistance. The phenotypic variance (R2) explained by these loci ranged from 0.4 to 30.2%. Among the loci, 7 for BB resistance were co-localized with known BB resistance genes and one for BLS resistance overlapped witha previously reportedBLS resistance QTL. A search for the candidates in other novel loci revealed several defense-related genes that may be involved in resistance to BB and BLS. High levels of phenotypic resistance to BB or BLS could be attributed to the accumulation of the resistance (R) alleles at the associated loci, indicating their potential value in rice resistance breeding via gene pyramiding. The GWAS analysis validated the known genes underlying BB and BLS resistance and identified novel loci that could enrich the current resistance gene pool. The resources with strong resistance and significant SNPs identified in this study are potentially useful in breeding for BB and BLS resistance.

Phenotypic and molecular characterization of quinolone resistance of enteropathogens isolated from diarrhea in young children in Koula-Moutou/Gabon

ABSTRACT In the face of the growing phenomenon of multidrug-resistant bacteria, studies on the sensitivity profiles of enteric pathogens are required to better manage the remaining antibiotic capital. The objective of this study was to investigate the sensitivity and to determine the genetic basis of quinolone-resistant enteric diarrheal pathogens isolated from children in the city of Koula-Moutou, Gabon. Thirty strains of the Salmonella genera, five strains of Shigella and two strains of Yersinia (2) were analyzed. The Kirby–Bauer method was used to determine the sensitivity of these isolates to five molecules in the quinolone family. The presence of chromosomal (gyrA) and plasmid (PMQR) quinolone resistance genes, including the QNR complex (qnrA, qnrB, and qnrS) were investigated by PCR. These enteric pathogens exhibited high levels of resistance to quinolones, particularly ofloxacin (100%) and ciprofloxacin (97.3%). Most isolates of Salmonella were found to be resistant to nalidixic acid (86.7%), however all isolates of the Shigella and Yersinia genera were susceptible to this antibiotic. Among the 37 strains, 21.6% had the gyrA gene, 13.5% had the qnrB and qnrS genes but the qnrA gene was not detected. Enteric pathogens isolated from the diarrheal feces of the children in Koula-Moutou exhibited high levels of phenotypic resistance to quinolones. This resistance is especially due to mutations occurring on the chromosomal determinant gyrA which also played a role in the reduced sensitivity of these strains to fluoroquinolones. The resistance is also linked to a lesser extent to the acquisition of the PMQR genes, notably qnrB and qnrS. In conclusion, this study highlights the high prevalence rate of enteric pathogens isolated from diarrheal feces of children in Gabon and shows the presence of the QNR complex genes (qnrB and qnrS) in the strains.

Genotypic and Phenotypic Characterization, Antibiotic Resistance and Virulence Patterns of Staphylococcus aureus Isolated form Goat Mastitis

Objective: To determine the phenotypic and genotypic characterization of the virulence and antimicrobial susceptibility of S. aureus. Design: Descriptive study Samples:175 raw goat milk samples Procedures: A total of 87 S. aureus strains was isolated from goat with mastitis in north-west cost of Egypt. PCR technique was used to detect genes coding some virulence and antimicrobial properties while phenotypic antimicrobial susceptibility was determined by the disc diffusion method. Results: Most S. aureus isolates were resistant to amoxycillin (89.7%), penicillin G (88.5%), and ampicillin (86%) while were less resistant to other members of β-lactam group as cefoxitin (17.2%) and ceftriaxone (10.3%).The lowest resistant rates were toward fluroquinolone group members; ciprofloxacin, levofloxacin and ofloxacin in rates 1.2%, 2.3% and 4.5%, respectively. Tetracycline showed the highest level of phenotypic resistance (34.6%) and prevalence of a tetK gene (39.1%). The virulence factors, spa and coa exhibited prevalence rate of 6.9% and 8%, respectively with significant relationship between them. Conclusion and Clinical relevance: The problem of antimicrobial resistance needs strict measures to limit the indiscriminate use of antibiotics and concerted research efforts to limit the spread of resistant bacteria, which will negatively affect human health and control of the infectious diseases in both humans and animals.

The R Package PROSPER: An Environment for Modeling Weed Population Dynamics and the Evolution of Herbicide Resistance

Weed management is a challenge for farmers worldwide, and the problem is exacerbated by the spread of weed herbicide resistance. Simulation models that combine population dynamics and genetics are valuable tools for predicting the impact of competing management options on weed density, allele frequency, and phenotypic resistance levels. The new R package PROSPER provides functions for the forward simulation of weed population dynamics on a field scale, the selection of individuals according to their sensitivity to herbicides, and the recombination of alleles during reproduction. Objects are provided to enter and save model parameters in a clear structure, and to save output data for further processing in R. The basic functions are extensible with R code. PROSPER combines individual-based population dynamics with monogenic or polygenic diploid inheritance and flexible selection pressure. Stochasticity can be included at all model steps. Two examples of the population dynamics of two annual weed species with herbicide resistance are presented. All parameters and the models are available in PROSPER. In addition to simulation, PROSPER is intended for sharing and publishing population dynamic parameters and models, which is easily done thanks to R.

Resting behaviour of malaria vectors in highland and lowland sites of western Kenya: Implication on malaria vector control measures.

Understanding the interactions between increased insecticide resistance and resting behaviour patterns of malaria mosquitoes is important for planning of adequate vector control. This study was designed to investigate the resting behavior, host preference and rates of Plasmodium falciparum infection in relation to insecticide resistance of malaria vectors in different ecologies of western Kenya. Anopheles mosquito collections were carried out during the dry and rainy seasons in Kisian (lowland site) and Bungoma (highland site), both in western Kenya using pyrethrum spray catches (PSC), mechanical aspiration (Prokopack) for indoor collections, clay pots, pit shelter and Prokopack for outdoor collections. WHO tube bioassay was used to determine levels of phenotypic resistance of indoor and outdoor collected mosquitoes to deltamethrin. PCR-based molecular diagnostics were used for mosquito speciation, genotype for knockdown resistance mutations (1014S and 1014F) and to determine specific host blood meal origins. Enzyme-linked Immunosorbent Assay (ELISA) was used to determine mosquito sporozoite infections. Anopheles gambiae s.l. was the most predominant species (75%, n = 2706) followed by An. funestus s.l. (25%, n = 860). An. gambiae s.s hereafter (An. gambiae) accounted for 91% (95% CI: 89-93) and An. arabiensis 8% (95% CI: 6-9) in Bungoma, while in Kisian, An. arabiensis composition was 60% (95% CI: 55-66) and An. gambiae 39% (95% CI: 34-44). The resting densities of An. gambiae s.l and An. funestus were higher indoors than outdoor in both sites (An. gambiae s.l; F1, 655 = 41.928, p < 0.0001, An. funestus; F1, 655 = 36.555, p < 0.0001). The mortality rate for indoor and outdoor resting An. gambiae s.l F1 progeny was 37% (95% CI: 34-39) vs 67% (95% CI: 62-69) respectively in Bungoma. In Kisian, the mortality rate was 67% (95% CI: 61-73) vs 76% (95% CI: 71-80) respectively. The mortality rate for F1 progeny of An. funestus resting indoors in Bungoma was 32% (95% CI: 28-35). The 1014S mutation was only detected in indoor resitng An. arabiensis. Similarly, the 1014F mutation was present only in indoor resting An. gambiae. The sporozoite rates were highest in An. funestus followed by An. gambiae, and An. arabiensis resting indoors at 11% (34/311), 8% (47/618) and 4% (1/27) respectively in Bungoma. Overall, in Bungoma, the sporozoite rate for indoor resting mosquitoes was 9% (82/956) and 4% (8/190) for outdoors. In Kisian, the sporozoite rate was 1% (1/112) for indoor resting An. gambiae. None of the outdoor collected mosquitoes in Kisian tested positive for sporozoite infections (n = 73). The study reports high indoor resting densities of An. gambiae and An. funestus, insecticide resistance, and persistence of malaria transmission indoors regardless of the use of long-lasting insecticidal nets (LLINs). These findings underline the difficulties of controlling malaria vectors resting and biting indoors using the current interventions. Supplemental vector control tools and implementation of sustainable insecticide resistance management strategies are needed in western Kenya.

Rapid phenotypic evolution in multidrug-resistant Klebsiella pneumoniae hospital outbreak strains.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) increasingly cause high-mortality outbreaks in hospital settings globally. Following a patient fatality at a hospital in Beijing due to a blaKPC-2-positive CRKP infection, close monitoring was put in place over the course of 14 months to characterize all blaKPC-2-positive CRKP in circulation in the hospital. Whole genome sequences were generated for 100 isolates from blaKPC-2-positive isolates from infected patients, carriers and the hospital environment. Phylogenetic analyses identified a closely related cluster of 82 sequence type 11 (ST11) isolates circulating in the hospital for at least a year prior to admission of the index patient. The majority of inferred transmissions for these isolates involved patients in intensive care units. Whilst the 82 ST11 isolates collected during the surveillance effort all had closely related chromosomes, we observed extensive diversity in their antimicrobial resistance (AMR) phenotypes. We were able to reconstruct the major genomic changes underpinning this variation in AMR profiles, including multiple gains and losses of entire plasmids and recombination events between plasmids, including transposition of blaKPC-2. We also identified specific cases where variation in plasmid copy number correlated with the level of phenotypic resistance to drugs, suggesting that the number of resistance elements carried by a strain may play a role in determining the level of AMR. Our findings highlight the epidemiological value of whole genome sequencing for investigating multi-drug-resistant hospital infections and illustrate that standard typing schemes cannot capture the extraordinarily fast genome evolution of CRKP isolates.

Delay in effective therapy in anidulafungin-resistant Candida tropicalis fungaemia: Potential for rapid prediction of antifungal resistance with whole-genome-sequencing.

This study investigated the feasibility of using whole-genome sequencing (WGS) for the prediction of antifungal resistance in anidulafungin-resistant Candida tropicalis candidaemia isolates. Next-generation sequencing was performed for three anidulafungin-resistant C. tropicalis isolates on an Illumina MiSeq system with in-house bioinformatics analysis. Mutations in Fks1p associated with anidulafungin resistance were identified. Other mutations associated with varying levels of phenotypic resistance to fluconazole were also identified. These data demonstrate the potential to predict antifungal resistance using WGS. With improving technology, real-time WGS may be used for tailoring effective antifungal therapy in patients with candidaemia.

Status of insecticide resistance in malaria vectors in Kwale County, Coastal Kenya

BackgroundThe strategy for malaria vector control in the context of reducing malaria morbidity and mortality has been the scale-up of long-lasting insecticidal nets to universal coverage and indoor residual spraying. This has led to significant decline in malaria transmission. However, these vector control strategies rely on insecticides which are threatened by insecticide resistance. In this study the status of pyrethroid resistance in malaria vectors and it’s implication in malaria transmission at the Kenyan Coast was investigated.ResultsUsing World Health Organization diagnostic bioassay, levels of phenotypic resistance to permethrin and deltamethrin was determined. Anopheles arabiensis showed high resistance to pyrethroids while Anopheles gambiae sensu stricto (s.s.) and Anopheles funestus showed low resistance and susceptibility, respectively. Anopheles gambiae sensu lato (s.l.) mosquitoes were further genotyped for L1014S and L1014F kdr mutation by real time PCR. An allele frequency of 1.33% for L1014S with no L1014F was detected. To evaluate the implication of pyrethroid resistance on malaria transmission, Plasmodium falciparum infection rates in field collected adult mosquitoes was determined using enzyme linked immunosorbent assay and further, the behaviour of the vectors was assessed by comparing indoor and outdoor proportions of mosquitoes collected. Sporozoite infection rate was observed at 4.94 and 2.60% in An. funestus s.l. and An. gambiae s.l., respectively. A higher density of malaria vectors was collected outdoor and this also corresponded with high Plasmodium infection rates outdoor.ConclusionsThis study showed phenotypic resistance to pyrethroids and low frequency of L1014S kdr mutation in An. gambiae s.l. The occurrence of phenotypic resistance with low levels of kdr frequencies highlights the need to investigate other mechanisms of resistance. Despite being susceptible to pyrethroids An. funestus s.l. could be driving malaria infections in the area.