Genotypic Resistance Profiles Research Articles

Microbiological Quality of Coconut Water Sold in the Grande Vitória Region, Brazil, and Phenogenotypic Antimicrobial Resistance of Associated Enterobacteria.

This study aimed to evaluate the microbiological quality of coconut water sold from street carts equipped with cooling coils or refrigerated at bakeries in the Grande Vitória Region, Brazil. Additionally, it assessed the phenotypic and genotypic antimicrobial resistance profiles of isolated enterobacteria. The results indicated that coconut water sold at street carts had lower microbiological quality compared to refrigerated samples, as evidenced by significantly higher counts of mesophilic microorganisms. Using MALDI-TOF, the following opportunistic pathogens were identified: Citrobacter freundii, Enterobacter bugandensis, E. kobei, E. roggenkampii, Klebsiella pneumoniae, and Kluyvera ascorbata. Three isolates-E. bugandensis, K. pneumoniae, and K. ascorbata-were classified as multidrug-resistant (MDR). Widespread resistance to β-lactams and cephalosporins was detected, and some isolates were resistant to quinolones, nitrofurans, and phosphonic acids. The gene blaCTX-M-2 was detected in C. freundii, E. bugandensis, E. kobei, and K. ascorbata. However, genes blaNDM, blaKPC, blaCMY-1, and blaCMY-2 were not detected in any isolate. The findings underscore the need to enhance good manufacturing practices in this sector to control the spread of antimicrobial resistance (AMR). To our knowledge, this is the first study documenting the presence of potentially pathogenic enterobacteria in coconut water samples and their associated phenotypic and genotypic AMR profiles.

Comparison of antimicrobial resistant Enterobacterales isolates from the dairy production environment in low and high zinc containing regions

Heavy metals occur naturally in the environment, and their concentration varies in soil across different regions. However, the presence of heavy metals may influence the antimicrobial resistance (AMR) in bacterial populations. Therefore, the objective of this study was to investigate and characterise the antimicrobial resistance profiles of Enterobacterales in soil and bovine milk filters from high and low zinc-containing regions in Ireland. In total, 50 soil samples and 29 milk filters were collected from two geographic locations with varying soil zinc concentrations. Samples were cultured for the enumeration and detection of Enterobacterales. Specifically, extended-spectrum beta-lactamase-producing Enterobacterales, carbapenem-resistant Enterobacterales and ciprofloxacin-resistant Enterobacterales were isolated using selective media. Species identification was performed using MALDI-TOF. The phenotypic resistance profiles of selected Enterobacterales were determined by disk diffusion testing, following EUCAST and CLSI criteria; while, the genotypic resistance profiles of the same isolates were determined by whole genome sequencing (WGS). Heavy metal concentrations were also measured for all soil samples. A total of 40 antimicrobial resistant Enterobacterales were identified in soil (n = 31) and milk filters (n = 9). The predominant species detected in the high zinc-containing region was Escherichia coli in both sample types (soil n = 10, milk filters n = 2), while in the low zinc-containing region Serratia fonticola was predominant in soil samples (n = 8) and E. coli in milk filters (n = 4). Ten E. coli isolates identified from soil samples in the high zinc-containing region were multidrug resistant, showing resistance to all the antimicrobials tested, except for carbapenems. The WGS findings confirmed the phenotypic resistance results. Moreover, zinc resistance-associated genes and genes encoding for efflux pumps were identified. The current study revealed distinct phenotypic resistance profiles of Enterobacterales in low and high zinc-containing regions, and highlighted the benefit of utilising milk filters for AMR surveillance in dairy production.

Widespread prevalence of plasmid-mediated blaCTX-M type extended-spectrum beta-lactamase Escherichia coli in backyard broiler production systems in the United States.

Extended-spectrum beta-lactamase (ESBL) Escherichia coli (E. coli) is an emerging pathogen of high concern given its resistance to extended-spectrum cephalosporins. Broiler chicken, which is the number one consumed meat in the United States and worldwide, can be a reservoir of ESBL E. coli. Backyard poultry ownership is on the rise in the United States, yet there is little research investigating prevalence of ESBL E. coli in this setting. This study aims to identify the prevalence and antimicrobial resistance profiles (phenotypically and genotypically) of ESBL E. coli in some backyard and commercial broiler farms in the U.S. For this study ten backyard and ten commercial farms were visited at three time-points across flock production. Fecal (n = 10), litter/compost (n = 5), soil (n = 5), and swabs of feeders and waterers (n = 6) were collected at each visit and processed for E. coli. Assessment of ESBL phenotype was determined through using disk diffusion with 3rd generation cephalosporins, cefotaxime and ceftazidime, and that with clavulanic acid. Broth microdilution and whole genome sequencing were used to investigate both phenotypic and genotypic resistance profiles, respectively. ESBL E. coli was more prevalent in backyard farms with 12.95% of samples testing positive whereas 0.77% of commercial farm samples were positive. All isolates contained a blaCTX-M gene, the dominant variant being blaCTX-M-1, and its presence was entirely due to plasmids. Our study confirms concerns of growing resistance to fourth generation cephalosporin, cefepime, as roughly half (51.4%) of all isolates were found to be susceptible dose-dependent and few were resistant. Resistance to non-beta lactams, gentamicin and ciprofloxacin, was also detected in our samples. Our study identifies prevalence of blaCTX-M type ESBL E. coli in U.S. backyard broiler farms, emphasizing the need for interventions for food and production safety.

Pharmaco-virological outcomes and genotypic resistance profiles among children and adolescents receiving a DTG-based regimen in Togo.

Few data are available on the real-world efficacy of receiving tenofovir-lamivudine-dolutegravir (DTG) as HIV treatment, particularly among young people in West Africa. Here, we evaluated pharmaco-virological outcomes and resistance profiles among Togolese children and adolescents. A cross-sectional study was conducted in Lomé, Togo, enrolling antiretroviral-treated people with HIV aged from 18 months to 24 years. Plasma HIV-1 viral load and antiretroviral concentrations were measured. Next-Generation Sequencing (NGS) of protease, Reverse Transcriptase (RT) and integrase was performed on all samples with viral load >200 c/mL. Drug resistance mutations (DRMs) were identified and interpreted using the ANRS-MIE algorithm. 264 participants were enrolled (median age=17 years), 226 received a DTG-based regimen for a median of 20.5 months. Among them, virological suppression at the 200 c/mL threshold in 80.0% of the participants. Plasma DTG concentrations were adequate (i.e., >640 ng/mL), suboptimal and below the limit of quantification in 74.1%, 6.7% and 19.2% of participants receiving DTG, respectively. Overall, viruses resistant to any of Nucleoside RT Inhibitors, Non-NRTIs, and protease inhibitors were found in 52%, 66% and 1.6% of participants, respectively. A major integrase inhibitor DRM was observed in 9.4% (n=3/32, R263K, E138A-G140A-Q148R, and N155H) of participants with a viral load >200 c/mL. These first findings in such a large series of adolescents in a low-income country, showed a good virological response of 80% and the presence of an integrase DRM in 9.4% of the virological failures, supporting the need to monitor DTG drug resistance to reduce the risk of resistance acquisition.

Molecular genetic characterization analysis of a novel HIV-1 circulating recombinant form (CRF156_0755) in Guangdong, China.

The characteristic of human immunodeficiency virus type 1 (HIV-1) is its susceptibility to erroneous replication and recombination, which plays a crucial role in the diverse and dynamic variation of HIV-1. The spread of different subtypes in the same population often leads to the emergence of circulating recombination forms (CRFs). At present, the main recombinant subtypes of HIV-1 in China are CRF07_BC, CRF01_AE, CRF08_BC and B' subtypes, while CRF55_01B has become the fifth major epidemic strain in China after rapid growth in recent years since it was first reported in 2013. In this study, we obtained five nearly full-length genomes (NFLGs) and one half-length genome from five different cities in Guangdong. Here, we focused on analyzing their characteristics, parental origin and drug resistance. Plasma samples were collected from six HIV-1 infected patients in Guangdong Province who had no epidemiological association with each other. The NFLGs of HIV-1 were amplified in two overlapping segments by the near-terminal dilution method. The positive products were sequenced directly to obtain genomic sequences. The recombinant patterns and breakpoints of the NFLGs were determined using the Simplot software and confirmed by the maximum likelihood trees for segments using the IQ-TREE and BEAST software. The genotypic resistance profiles of the protease reverse transcriptase and integrase were resolved by the Stanford HIV drug resistance database. The six genomes shared highly similar recombinant pattern, with the CRF55_01B backbone substituted by CRF07_BC segments, therefore assigned as CRF156_0755. The evolutionary analysis of the segments showed that CRF07_BC segments were not clustered with the Chinese MSM variants in the CRF07_BC lineage. All the five NFLGs were identified with the non-nucleoside reverse-transcription inhibitors (NNRTIs) resistance mutation V179E. With the accumulation and evolution of recombination between CRF55_01B and CRF 07_BC, the prevalence of more recombinant strains of CRF55_01B and CRF 07_BC may occur. Therefore, it is necessary to strengthen the identification and monitoring of the recombination of CRF55_01B and CRF 07_BC.

Co-production of metallo-β-lactamase and OXA-type β-lactamases in carbapenem-resistant Acinetobacter baumannii clinical isolates in North East India.

Carbapenem-resistant Acinetobacter baumannii poses a significant threat to public health globally, especially due to its ability to produce multiple carbapenemases, leading to treatment challenges. This study aimed to investigate the antibiotic resistance pattern of carbapenem-resistant A. baumannii isolates collected from different clinical settings in North East India, focusing on their genotypic and phenotypic resistance profiles. A total of 172 multidrug-resistant A. baumannii isolates were collected and subjected to antibiotic susceptibility test using the Kirby-Bauer disk diffusion method. Various phenotypic tests were performed to detect extended-spectrum β-lactamase (ESBL), metallo-β-lactamase (MBL), class C AmpC β-lactamase (AmpC), and carbapenem hydrolyzing class D β-lactamase (CHDL) production among the isolates. Overexpression of carbapenemase and cephalosporinase genes was detected among the isolates through both phenotypic and genotypic investigation. The antibiotic resistance profile of the isolates revealed that all were multidrug-resistant; 25% were extensively drug-resistant, 9.30% were pan-drug-resistant, whereas 91.27% were resistant to carbapenems. In the genotypic investigation, 80.81% of isolates were reported harbouring at least one metallo-β-lactamase encoding gene, with blaNDM being the most prevalent at 70.34%, followed by blaIMP at 51.16% of isolates. Regarding class D carbapenemases, blaOXA-51 and blaOXA-23 genes were detected in all the tested isolates, while blaOXA-24, blaOXA-48, and blaOXA-58 were found in 15.11%, 6.97%, and 1.74% isolates respectively. Further analysis showed that 31.97% of isolates co-harboured ESBL, MBL, AmpC, and CHDL genes, while 31.39% of isolates co-harboured ESBL, MBL, and CHDL genes with or without ISAba1 leading to extensively drug-resistant or pan drug-resistant phenotypes. This study highlights the complex genetic profile and antimicrobial-resistant pattern of the isolates circulating in North East India, emphasizing the urgent need for effective infection control measures and the development of alternative treatment strategies to combat these challenging pathogens.

Comparison of genotypic and phenotypic antimicrobial resistance profiles of Salmonella enterica isolates from poultry diagnostic specimens.

The spread of antimicrobial-resistant bacteria is a significant concern, as it can lead to increased morbidity and mortality in both humans and animals. Whole-genome sequencing (WGS) is a powerful tool that can be used to conduct a comprehensive analysis of the genetic basis of antimicrobial resistance (AMR). We compared the phenotypic and genotypic AMR profiles of 97 Salmonella isolates derived from chicken and turkey diagnostic samples. We focused AMR analysis on 5 antimicrobial classes: aminoglycoside, beta-lactam, phenicol, tetracycline, and trimethoprim. The overall sensitivity and specificity of WGS in predicting phenotypic antimicrobial resistance in the Salmonella isolates were 93.4% and 99.8%, respectively. There were 16 disagreement instances, including 15 that were phenotypically resistant but genotypically susceptible; the other instance involved phenotypic susceptibility but genotypic resistance. Of the isolates examined, 67 of 97 (69%) carried at least 1 resistance gene, with 1 isolate carrying as many as 12 resistance genes. Of the 31 AMR genes analyzed, 16 were identified as aminoglycoside-resistance genes, followed by 4 beta-lactam-resistance, 3 tetracycline-resistance, 2 sulfonamide-resistance, and 1 each of fosfomycin-, quinolone-, phenicol-, trimethoprim-, bleomycin-, and colistin-resistance genes. Most of the resistance genes found were located on plasmids.

Multidrug resistant Vibrio spp. identified from mussels farmed for human consumption in Central Italy.

This study investigated phenotypic and genotypic antimicrobial resistance profiles of Vibrio strains identified from Mytilus galloprovincialis farmed for human consumption in the Adriatic Sea Central Italy. A total of 475 mussels (M. galloprovincialis) were involved in the present study, and culture-dependent microbiological methods permitted to identify a total of 50 Vibrio strains that were tested for antibiotic susceptibility followed by the genetic determinant detections. Antibiograms showed resistance against ampicillin (36.0%), amoxicillin-clavulanic acid (30.0%), gentamycin (14.0%), and imipenem (18.0%). Biomolecular assays amplified a total of 264 antibiotic resistance genes harbored by both susceptible and resistant Vibrio species. Among resistance genes, aacC2 (62.0%) and aadA (58.0%) for aminoglycosides, blaTEM (54.0%) for beta-lactams, qnrS (24.0%) for quinolones, tetD (66.0%) for tetracyclines, and vanB (60.0%) for glycopeptides were mainly amplified by PCR assays. Vibrio genus is involved in the antibiotic resistance phenomenon diffusion in the aquatic environments, as demonstrated by the harboring of many genetic determinants representing a kind of genetic "dark world".

The frequency of kdr and ace-1 alleles in Anopheles gambiae s.l. before and during indoor residual spraying (IRS) implementation and four years after IRS withdrawal in three districts in Atacora, Benin

BackgroundIndoor residual spraying (IRS) was first implemented in the Atacora department, Benin from 2011 to 2012 using bendiocarb (carbamate) followed by annual spraying with pirimiphos-methyl (organophosphate) from 2013 to 2018. Before and after IRS implementation in Atacora, standard pyrethroid insecticide-treated bed nets were the main method of vector control in the area. This study investigated the knockdown resistance (kdr) gene (L1014F) and the acetylcholinesterase (ace-1) gene (G119S), before and during IRS implementation, and 4-years after IRS withdrawal from Atacora. This was done to assess how changes in insecticide pressure from indoor residual spraying may have altered the genotypic resistance profile of Anopheles gambiae s.l.MethodIdentification of sibling species of An. gambiae s.l. and detection of the L1014F mutation in the kdr gene and G119S mutation in ace-1 genes was done using molecular analysis. Allelic and genotypic frequencies were calculated and compared with each other before and during IRS implementation and 4 years after IRS withdrawal. The Hardy–Weinberg equilibrium and genetic differentiation within and between populations were assessed.ResultsPrevalence of the L1014F mutation in all geographic An. gambiae s.l. (An. gambiae s.s., Anopheles. coluzzii, Anopheles. arabiensis, and hybrids of “An. gambiae s.s. and An. coluzzii”) populations increased from 69% before IRS to 87% and 90% during and after IRS. The G119S allele frequency during IRS (20%) was significantly higher than before IRS implementation (2%). Four years after IRS withdrawal, allele frequencies returned to similar levels as before IRS (3%). Four years after IRS withdrawal, the populations showed excess heterozygosity at the ace-1 gene and deficit heterozygosity at the kdr gene, whereas both genes had excess heterozygosity before and during IRS (FIS < 0). No genetic differentiation was observed within the populations.ConclusionsThis study shows that the withdrawal of IRS with bendiocarb and pirimiphos-methyl may have slowed down the selection of individual mosquitoes with ace-1 resistance alleles in contrast to populations of An. gambiae s.l. with the L1014F resistance allele of the kdr gene. This may suggest that withdrawing the use of carbamates or organophosphates from IRS or rotating alternative insecticides with different modes of action may slow the development of ace-1 insecticide-resistance mutations. The increase in the prevalence of the L1014F mutation of the kdr gene in the population, despite the cessation of IRS, could be explained by the growing use of pyrethroids and DDT in agriculture and for other domestic use. More observational studies in countries where carbamates or organophosphates are still being used as public health insecticides may provide additional insights into these associations.Graphical

Multi-drug Resistant Acinetobacter baumannii: Phenotypic and Genotypic Resistance Profiles and the Associated Risk Factors in Teaching Hospital in Jordan

BackgroundThis study aimed to determine the prevalence of Antimicrobial Resistance Genes (ARGs), with a focus on colistin resistance in clinical A. baumannii, as well as the risk factors associated with A. baumannii infection in Jordanian patients. MethodsIn total, 150 A. baumannii isolates were obtained from patients at a teaching hospital. The isolates were tested for antimicrobial susceptibility using disc diffusion and microdilution methods. PCR amplification was used to detect ARGs, and statistical analysis was conducted to evaluate the influence of identified risk factors on the ARGs acquisition. ResultsMore than 90% of A. baumannii isolates were resistant to monobactam, carbapenem, cephalosporins, Fluoroquinolones, penicillin, and β-lactam agents. Moreover, 20.6% of the isolates (n = 31) were colistin-resistant. Several ARGs were also detected in A. baumannii isolates. Univariate analysis indicated that risk factors and the carriage of ARGs were significantly associated P ≤ (0.05) with gender, invasive devices, immunodeficiency, systemic diseases, tumors, and covid-19. Logistic regression analysis indicated seven risk factors, and three protective factors were associated with the ARGs (armA, strA, and strB) P ≤ (0.05). In contrast, tetB and TEM were associated with 2 risk factors each P ≤ (0.05). ConclusionOur study indicates a high prevalence of MDR A. baumannii infections in ICU patients, as well as describing the case of colistin-resistant A. baumannii for the first time in Jordan. Additionally, the risk factors associated with ARGs-producing A. baumannii infections among ICU patients suggest a rapid emergence and spread of MDR A. baumannii without adequate surveillance and control measures.

Determination of antibiotic resistance patterns and genotypes of Escherichia coli isolated from wild birds

BackgroundCurbing the potential negative impact of antibiotic resistance, one of our era's growing global public health crises, requires regular monitoring of the resistance situations, including the reservoir of resistance genes. Wild birds, a possible bioindicator of antibiotic resistance, have been suggested to play a role in the dissemination of antibiotic-resistant bacteria. Therefore, this study was conducted with the objective of determining the phenotypic and genotypic antibiotic resistance profiles of 100 Escherichia coli isolates of gull and pigeon origin by using the Kirby-Bauer disk diffusion method and PCR. Furthermore, the genetic relationships of the isolates were determined by RAPD-PCR.ResultsPhenotypic antibiotic susceptibility testing revealed that 63% (63/100) and 29% (29/100) of E. coli isolates were resistant to at least one antibiotic and multidrug-resistant (MDR), respectively. With the exception of cephalothin, to which the E. coli isolates were 100% susceptible, tetracycline (52%), kanamycin (38%), streptomycin (37%), ampicillin (28%), chloramphenicol (21%), trimethoprim/sulfamethoxazole (19%), gentamicin (13%), enrofloxacin (12%) and ciprofloxacin (12%) resistances were detected at varying degrees. Among the investigated resistance genes, tet(B) (66%), tet(A) (63%), aphA1 (48%), sul3 (34%), sul2 (26%), strA/strB (24%) and sul1 (16%) were detected. Regarding the genetic diversity of the isolates, the RAPD-PCR-based dendrograms divided both pigeon and gull isolates into five different clusters based on a 70% similarity threshold. Dendrogram analysis revealed 47–100% similarities among pigeon-origin strains and 40–100% similarities among gull-origin E.coli strains.ConclusionsThis study revealed that gulls and pigeons carry MDR E. coli isolates, which may pose a risk to animal and human health by contaminating the environment with their feces. However, a large-scale epidemiological study investigating the genetic relationship of the strains from a "one health" point of view is warranted to determine the possible transmission patterns of antibiotic-resistant bacteria between wild birds, the environment, humans, and other hosts.3bRmtrrdv9vbDYjBBmhJ2hVideo

Analysis of Phenotypic and Genotypic Susceptibility to Clarithromycin and Amikacin of Mycobacterium abscessus Complex Strains Isolated from Cystic Fibrosis Patients.

Mycobacterium abscessus complex infections are ever on the rise. To curb their increasing evolution, we performed an in-depth study of 43 clinical isolates of cystic fibrosis patients obtained from 2009 to 2020. We identified their subspecies, uncovered their genotypic resistance profiles, characterised their antibiotic-resistant genes, and assessed their phenotypic antibiotic susceptibilities. The phenotypic and genotypic methods showed total agreement in terms of resistance to clarithromycin and amikacin. Of the 43 clinical strains, 28 belonged to M. abscessus subsp. abscessus (65.1%), 13 to M. abscessus subsp. massiliense (30.2%), and 2 to M. abscessus subsp. bolletii (4.6%). The resistant rates for clarithromycin and amikacin, the two main drugs against M. abscessus complex pulmonary infections, were 64.2% and 14.2%, respectively. We found three strains of M. abscessus subsp. abscessus that showed heteroresistance in the rrl and rrs genes, and these strains also presented double-resistance since they were macrolide- and aminoglycoside-resistant. M. abscessus subsp. abscessus showed a high minimum inhibitory concentration (MIC) and a resistant percentage larger than or equal to 88% to cefoxitin, ciprofloxacin, moxifloxacin, doxycycline, imipenem, and trimethoprim-sulfamethoxazole. These results show a panorama of the high resistance of Mycobacterium abscessus complex to current drugs for cystic fibrosis patients. Thus, other treatment methods are urgently needed.

Phenotypic and Genotypic Antimicrobial Resistance Profile of Salmonella spp. Isolated from Kampung Chicken Carcasses

Kampung chicken meats have been widely consumed in Indonesia as well as broiler chicken. However, the extensive rearing allowed multidrug-resistant (MDR) bacteria exposure to kampung chicken, including through horizontal gene transfer. This study aimed to observe the correlation between the phenotypic and genotypic antimicrobial resistance profile of Salmonella spp. isolated from kampung chicken carcasses. Phenotypic antimicrobial resistance was evaluated by Kirby-Bauer's disk diffusion method, while the detection of drug resistance genes in seventeen isolates of Salmonella was carried out by PCR. All (17/17) isolates were susceptible to ampicillin and chloramphenicol. Most isolates of Salmonella were resistant to erythromycin (82%; 14/17), while the decreased susceptibility (intermediate category) most occurred in oxytetracycline (82%; 14/17). Salmonella Typhimurium showed a resistance pattern to more antimicrobial groups than S. Newport and S. Weltevreden. Several antimicrobial resistance genes (blaTEM, tetG, cmlA, gyrA) were present in all (17/17) isolates of Salmonella spp. Resistance to antimicrobial agents and the presence of resistance genes were not always related. This study could provide beneficial information regarding the transmission of antimicrobial resistance among Salmonella spp. from kampung chickens.

Longitudinal Assessment of Virological Failure and Antiretroviral Drug Resistance among Treatment-naive Subjects Living with HIV.

Human immunodeficiency virus (HIV) infection, the etiological agent of acquired immunodeficiency syndrome (AIDS), is a serious public health issue. Therapeutic measures have been successful in increasing the survival and improving the quality of life. However, some treatment-naive subjects living with HIV present resistance-associated mutations as a result of late diagnosis and/or mutant strain infections. The objective of this study was to identify the virus genotype and assess the antiretroviral resistance profile based on the results of HIV genotyping in treatment-naive subjects living with HIV, after six months of taking antiretroviral therapy. This was a prospective cohort study on treatment-naive adults living with HIV attending a specialized outpatient clinic in southern Santa Catarina State, Brazil. The participants were interviewed and had blood samples drawn. The genotypic antiretroviral drug resistance profile was examined in patients with detectable viral loads. 65 treatment-naive subjects living with HIV were recruited for this study. After six months of taking antiretroviral therapy, resistance-associated mutations were observed in 3 (4.6%) subjects living with HIV. Subtype C was identified as the circulating subtype in southern Santa Catarina State, and L10V, K103N, A98G, and Y179D were the most common mutations found in treatment-naive subjects.

Pangenomic analyses of antibiotic-resistant Campylobacter jejuni reveal unique lineage distributions and epidemiological associations.

Application of whole-genome sequencing (WGS) to characterize foodborne pathogens has advanced our understanding of circulating genotypes and evolutionary relationships. Herein, we used WGS to investigate the genomic epidemiology of Campylobacter jejuni, a leading cause of foodborne disease. Among the 214 strains recovered from patients with gastroenteritis in Michigan, USA, 85 multilocus sequence types (STs) were represented and 135 (63.1 %) were phenotypically resistant to at least one antibiotic. Horizontally acquired antibiotic resistance genes were detected in 128 (59.8 %) strains and the genotypic resistance profiles were mostly consistent with the phenotypes. Core-gene phylogenetic reconstruction identified three sequence clusters that varied in frequency, while a neighbour-net tree detected significant recombination among the genotypes (pairwise homoplasy index P<0.01). Epidemiological analyses revealed that travel was a significant contributor to pangenomic and ST diversity of C. jejuni, while some lineages were unique to rural counties and more commonly possessed clinically important resistance determinants. Variation was also observed in the frequency of lineages over the 4 year period with chicken and cattle specialists predominating. Altogether, these findings highlight the importance of geographically specific factors, recombination and horizontal gene transfer in shaping the population structure of C. jejuni. They also illustrate the usefulness of WGS data for predicting antibiotic susceptibilities and surveillance, which are important for guiding treatment and prevention strategies.

Whole genome analysis and antimicrobial resistance of Neisseria gonorrhoeae isolates from Ghana.

Gonorrhoea is a major public health concern. With the global emergence and spread of resistance to last-line antibiotic treatment options, gonorrhoea threatens to be untreatable in the future. Therefore, this study performed whole genome characterization of Neisseria gonorrhoeae collected in Ghana to identify lineages of circulating strains as well as their phenotypic and genotypic antimicrobial resistance (AMR) profiles. Whole genome sequencing (WGS) was performed on 56 isolates using both the Oxford Nanopore MinION and Illumina MiSeq sequencing platforms. The Comprehensive Antimicrobial Resistance Database (CARD) and PUBMLST.org/neisseria databases were used to catalogue chromosomal and plasmid genes implicated in AMR. The core genome multi-locus sequence typing (cgMLST) approach was used for comparative genomics analysis. In vitro resistance measured by the E-test method revealed 100%, 91.0% and 85.7% resistance to tetracycline, penicillin and ciprofloxacin, respectively. A total of 22 sequence types (STs) were identified by multilocus sequence typing (MLST), with ST-14422 (n = 10), ST-1927 (n = 8) and ST-11210 (n = 7) being the most prevalent. Six novel STs were also identified (ST-15634, 15636-15639 and 15641). All isolates harboured chromosomal AMR determinants that confer resistance to beta-lactam antimicrobials and tetracycline. A single cefixime-resistant strain, that belongs to N. gonorrhoeae multiantigen sequence type (NG-MAST) ST1407, a type associated with widespread cephalosporin resistance was identified. Neisseria gonorrhoeae Sequence Typing for Antimicrobial Resistance (NG-STAR), identified 29 unique sequence types, with ST-464 (n = 8) and the novel ST-3366 (n = 8) being the most prevalent. Notably, 20 of the 29 STs were novel, indicative of the unique nature of molecular AMR determinants in the Ghanaian strains. Plasmids were highly prevalent: pTetM and pblaTEM were found in 96% and 92% of isolates, respectively. The TEM-135 allele, which is an amino acid change away from producing a stable extended-spectrum β-lactamase that could result in complete cephalosporin resistance, was identified in 28.5% of the isolates. Using WGS, we characterized N. gonorrhoeae strains from Ghana, giving a snapshot of the current state of gonococcal AMR in the country and highlighting the need for constant genomic surveillance.

Concordance between Genotypic and Phenotypic Drug-Resistant Profiles of Shigella Isolates from Taiyuan City, Shanxi Province, China, 2005 to 2016.

Antimicrobial resistance in Shigella spp. is a global public health concern. In this study, the AMR phenotypic profiles of 10 kinds of antibiotics were compared with the genotypic profiles using genomic analysis of 218 Shigella isolates from Taiyuan City, Shanxi Province, China, 2005 to 2016. Core genome Multilocus Sequence Typing (cgMLST) based on the EnteroBase Escherichia/Shigella scheme was used to obtain the genetic relatedness of Shigella isolates. Multiple-drug resistance was observed in 96.79% Shigella spp., and the resistance to antimicrobial agents varied between S. flexneri and S. sonnei. The genotypic results correlated well with the phenotypic profiles with concordance rates of 96.42% and 94.50% in S. flexneri and S. sonnei isolates, respectively, from Taiyuan City, Shanxi Province. The sensitivity and specificity of the genotypic antimicrobial susceptibility testing (AST) were 97.56% and 95.34% for S. flexneri, and 95.65% and 93.31% for S. sonnei isolates, respectively. A discrepancy of genotypic and phenotypic AST results existed in some cephalosporin- and azithromycin-resistant Shigella isolates; there were no clear resistance patterns to predict ciprofloxacin resistance. There were major discrepancies between genotypic and phenotypic AST in the genotypically resistant but phenotypically susceptible isolates. The drug-resistance patterns and essential drug-resistance genes to predict the phenotypic drug-resistant profiles were the discrepancies between S. flexneri and S. sonnei isolates. Phylogenetic analysis showed that isolates of the same cluster but with different antibiotic-resistance gene patterns occurred because of the loss or gain of antibiotic-resistance genes located in the plasmids and multidrug-resistance islands. IMPORTANCE Antimicrobial resistance in Shigella spp. has become a global public health concern. In this study, we identified the antimicrobial susceptibility testing (AST) characteristics based on genomic sequences of 218 Shigella isolates and analyzed the correlation between genotypic and phenotypic antibiotic resistance profiles of Shigella spp., especially for fluoroquinolone, macrolides, and third-generation cephalosporins. Our results show that the genotypic results correlated with the phenotypic profiles with concordance rates of 96.42% and 94.50% in S. flexneri and S. sonnei isolates, respectively. The drug-resistance patterns and essential drug-resistance genes to predict the phenotypic drug-resistant profiles of S. flexneri and S. sonnei isolates in Taiyuan city were distinct. The discrepancy between genotypic and phenotypic AST was considerable in the genotypically resistant but phenotypically susceptible isolates. The information on drug resistance and resistance genes in this study can offer more details on the prevalence of drug resistance of Shigella spp.

HIV-1 genotypic profiling ensures effective response to third-line antiretroviral therapy in Cameroon.

In order to limit the emergence of human immunodeficiency virus (HIV) drug resistance in a context of limited antiretroviral options, we sought to evaluate the efficacy of third-line (3L) regimens considering HIV genotypic resistance profile at initiation of 3L in Cameroon. A cohort-study was conducted from January-September 2020 among patients initiating a 3L antiretroviral therapy regimen at the Yaoundé Central Hospital. HIV-1 protease-reverse transcriptase was sequenced at the Chantal Biya international reference center for research on HIV/AIDS prevention and management and results were interpreted using Stanford HIVdbv8.3. Good virological response (viral load < 390 copies/mL) was assessed after 12 months using OPP-ERA platform. Statistical analyses were performed using Epi Info v7.2.2.6, with P < .05 considered statistically significant. Of the 38 patients initiating 3L with an available genotyping (42% female; median age, 49 [39-57] years), median cluster of differentiation type 4 count and viral load were 173 [34-374] cells/μL and 169,322 [30,382-551,826] copies/mL, respectively. At enrollment, all patients harbored resistance to reverse transcriptase inhibitors and 66% (25/38) to protease-inhibitors, although 63% (24/38) were still susceptible to darunavir/ritonavir. Preferred 3L regimen was dolutegravir + darunavir/r + tenofovir + lamivudine (51%) and median duration on 3L was 21 [17-32] months. Interestingly, 82% (31/38) of the participants achieved good virological response on 3L, regardless of genotypic profile at recruitment, variations in 3L regimens (P = .9) and baseline cluster of differentiation type 4 count (P = .3). Despite the high burden of reverse transcriptase inhibitor - and protease inhibitor boosted by ritonavir drug resistance, genotyping-guided 3L regimens is accompanied by virological success in most patients. This high efficacy, most likely due to use of high genetic barrier antiretrovirals, requires continuous adherence support alongside close monitoring for long-term effectiveness in similar programmatic settings.

Dolutegravir-associated resistance mutations after first-line treatment failure in Brazil

BackgroundSince January 2017, the recommended first-line antiretroviral regimen in Brazil is the fixed-dose combination of tenofovir plus lamivudine with dolutegravir (TL + D). According to the literature, integrase resistance-associated mutations (INRAMs) are rarely found upon virologic failure to first-line dolutegravir plus two nucleoside reverse transcriptase inhibitors. We evaluated the HIV antiretroviral genotypic resistance profile of patients referred for genotyping in the public health system who failed first-line TL + D after at least six months of therapy on or before December 31, 2018.MethodsHIV Sanger sequences of the pol gene were generated from plasma of patients with confirmed virologic failure to first-line TL + D in the Brazilian public health system before December 31, 2018.ResultsOne hundred thirteen individuals were included in the analysis. Major INRAMs were detected in seven patients (6.19%), four with R263K, one with G118R, one with E138A, and one with G140R. Four patients with major INRAMs also had the K70E and M184V mutations in the RT gene. Sixteen (14.2%) additional individuals presented minor INRAMs, and five (4,42%) patients had both major and minor INRAMS. Thirteen (11.5%) patients also presented mutations in the RT gene selected by tenofovir and lamivudine, including four with both the K70E and M184V mutations and four with only M184V. The integrase mutations L101I and T124A, which are in the in vitro pathway for integrase inhibitor resistance, were found in 48 and 19 patients, respectively. Mutations not related to TL + D, thus probable transmitted resistance mutations (TDR), were present in 28 patients (24.8%): 25 (22.1%) to nucleoside reverse transcriptase inhibitors, 19 (16.8%) to non-nucleoside reverse transcriptase inhibitors, and 6 (5.31%) to protease inhibitors.ConclusionsIn marked contrast to previous reports, we report a relatively high frequency of INRAMs among selected patients failing first-line TL + D in the public health system in Brazil. Possible reasons for this discrepancy include delays in detecting virologic failure, patients inadvertently on dolutegravir monotherapy, TDR, and/or infecting subtype.

Antimicrobial resistance and genomic characterization of Salmonella enterica isolates from chicken meat.

This study investigated genotypic and phenotypic antimicrobial resistance profiles, phylogenic relatedness, plasmid and virulence composition of 39 Salmonella enterica strains isolated from chicken meat samples using whole genome sequencing (WGS) technology. Four distinct serotypes were identified; Salmonella Minnesota (16/39, 41%), Salmonella Infantis (13/39, 33.3%), Salmonella Enteritidis (9/39, 23.1%), and one isolate was detected for Salmonella Kentucky (1/39, 2.6%), with sequence types (STs) as followed: ST548, ST32, ST11, and ST198, respectively. Phenotypic resistance to tetracycline (91.2%), ampicillin (82.4%), sulfisoxazole (64.7%), and nalidixic acid (61.6%) was the most observed. Resistome analysis revealed the presence of resistance genes to aminoglycosides, β-lactamase, sulfonamides, trimethoprim, phenicol, lincosamide, macrolides, and tetracyclines. Plasmidome showed the presence of eight incompatibility groups, including IncA/C2, IncFIB(K)_1_Kpn3, Col440I_1, IncR, IncX1, IncI1_1_Alpha, IncFIB(S)/IncFII(S), IncHI2/IncHI2A, IncX2 and ColpVC plasmids across the 39 genomes. Three resistance genes, sul2, tetA and blaCMY-2, were predicted to be located on IncA/C2 plasmid in S. Minnesota isolates, whereas all S. Infantis isolates were positive to IncFIB(K)_1_Kpn3 plasmid that carries bla CTX-M-65 gene. Eleven Salmonella pathogenicity islands and up to 131 stress and/or virulence genes were identified in the evaluated genomes. Phylogenetic analysis showed four phylogroups that were consistent with the identified ST profiles with a high level of inter-diversity between isolates. This is the first genomic characterization of Salmonella isolates from retail chicken meat in Saudi Arabia using WGS technology. The availability of Salmonella genomes from multiple geographic locations, including Saudi Arabia, would be highly beneficial in future source-tracking, especially during epidemiological surveillance and outbreak investigations.