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522100.9890Molecular cloning of the DNA gyrase genes from Methylovorus sp. strain SS1 and the mechanism of intrinsic quinolone resistance in methylotrophic bacteria. The genes encoding the DNA gyrase A (GyrA) and B subunits (GyrB) of Methylovorus sp. strain SS1 were cloned and sequenced. gyrA and gyrB coded for proteins of 846 and 799 amino acids with calculated molecular weights of 94,328 and 88,714, respectively, and complemented Escherichia coli gyrA and gyrB temperature sensitive (ts) mutants. To analyze the role of type II topoisomerases in the intrinsic quinolone resistance of methylotrophic bacteria, the sequences of the quinolone resistance-determining regions (QRDRs) in the A subunit of DNA gyrase and the C subunit (ParC) of topoisomerase IV (Topo IV) of Methylovorus sp. strain SS1, Methylobacterium extorquens AM1 NCIB 9133, Methylobacillus sp, strain SK1 DSM 8269, and Methylophilus methylotrophus NCIB 10515 were determined. The deduced amino acid sequences of the QRDRs of the ParCs in the four methylotrophic bacteria were identical to that of E. coli ParC. The sequences of the QRDR in GyrA were also identical to those in E. coli GyrA except for the amino acids at positions 83, 87, or 95. The Ser83 to Thr substitution in Methylovorus sp. strain SS1, and the Ser83 to Leu and Asp87 to Asn substitutions in the three other methylotrophs, agreed well with the minimal inhibitory concentrations of quinolones in the four bacteria, suggesting that these residues play a role in the intrinsic susceptibility of methylotrophic bacteria to quinolones.200516404155
124510.9890Mutation-based fluoroquinolone resistance in carbapenem-resistant Acinetobacter baumannii and Escherichia coli isolates causing catheter-related bloodstream infections. OBJECTIVE: We studied the presence of mutations in the chromosomal quinolone resistance-determining regions (QRDRs) of the fluoroquinolone targets gyrA and parC genes and detected the carbapenem resistance (CR) encoding genes among Acinetobacter baumannii and Escherichia coli isolates from catheter-related bloodstream infections (CRBSIs). METHODS: The study included 39 non-duplicate isolates of A. baumannii (14/39, 35.9%) and E. coli (25/39, 64.1%) isolated from 128 confirmed CRBSIs cases. Antimicrobial susceptibility testing was performed, followed by an evaluation of biofilm formation using the tissue culture plate method. The carbapenemase encoding genes were detected by multiplex polymerase chain reaction (PCR). The mutations in QRDRs of gyrA and parC genes were determined by singleplex PCR amplification followed by DNA sequencing and BlastN analysis in the GenBank database. DNA and the translated amino acid sequences were analyzed using the Mega7 bioinformatics tool. RESULTS: Multidrug-resistant (MDR) E. coli and A. baumannii isolates harbored CR encoding genes and combined gyrA and parC genes mutation. The specific substitutions observed in GyrA were Cys173Arg, Cys174Gly, Asp80Val, Tyr178ASP, Tyr84Gly, Glu85Lys, Ser172Leu, and Asp176Asn, while the specific substitutions observed in the ParC amino acid sequence were point mutation 62 Arg, Phe60Leu, Ils66Val, and Gln76Lys. Point mutation 62Arg was detected in two A. baumannii isolates, whereas Ser172Leu mutation was observed in two E. coli isolates. CONCLUSION: The presence of new single and multiple mutations in QRDR causes the emergence of MDR E. coli and A. baumannii infections in carbapenem-resistant Enterobacteriaceae in Egypt, requiring further investigation in Gram-negative bacteria.202337151743
124720.9889Antibiotic resistance determinants of multidrug-resistant Acinetobacter baumannii clinical isolates in Algeria. Antibiotic susceptibility testing was performed on 71 Acinetobacter baumannii clinical isolates, and presence of antibiotic resistance genes was screened for by PCR amplification and sequencing. Resistance rates were very high for aminoglycosides (22-80%), fluoroquinolones (>90%), and cephalosporins (>90%) but remained low for rifampin (2.8%) or null for colistin. Antibiotic resistance encoding genes detected were as follows: blaTEM-128 gene (74.6%), aph(3')-VI (50.7 %), aadA (63.4%), ant(2″)-I (14.1%), aac(3)-Ia (91.1%), aac(6')-Ib (4.2%), mutation Ser83Leu in gyrA (94.4%), double mutations Ser83Leu and Ser80Leu (or Ser84Leu) in gyrA and parC (69.0%), and mutation I581N in RRDR of the rpoB gene.201323688522
521430.9888Comparative genomic analysis of a new tellurite-resistant Psychrobacter strain isolated from the Antarctic Peninsula. The Psychrobacter genus is a cosmopolitan and diverse group of aerobic, cold-adapted, Gram-negative bacteria exhibiting biotechnological potential for low-temperature applications including bioremediation. Here, we present the draft genome sequence of a bacterium from the Psychrobacter genus isolated from a sediment sample from King George Island, Antarctica (3,490,622 bp; 18 scaffolds; G + C = 42.76%). Using phylogenetic analysis, biochemical properties and scanning electron microscopy the bacterium was identified as Psychrobacter glacincola BNF20, making it the first genome sequence reported for this species. P. glacincola BNF20 showed high tellurite (MIC 2.3 mM) and chromate (MIC 6.0 mM) resistance, respectively. Genome-wide nucleotide identity comparisons revealed that P. glacincola BNF20 is highly similar (>90%) to other uncharacterized Psychrobacter spp. such as JCM18903, JCM18902, and P11F6. Bayesian multi-locus phylogenetic analysis showed that P. glacincola BNF20 belongs to a polyphyletic clade with other bacteria isolated from polar regions. A high number of genes related to metal(loid) resistance were found, including tellurite resistance genetic determinants located in two contigs: Contig LIQB01000002.1 exhibited five ter genes, each showing putative promoter sequences (terACDEZ), whereas contig LIQB1000003.2 showed a variant of the terZ gene. Finally, investigating the presence and taxonomic distribution of ter genes in the NCBI's RefSeq bacterial database (5,398 genomes, as January 2017), revealed that 2,623 (48.59%) genomes showed at least one ter gene. At the family level, most (68.7%) genomes harbored one ter gene and 15.6% exhibited five (including P. glacincola BNF20). Overall, our results highlight the diverse nature (genetic and geographic diversity) of the Psychrobacter genus, provide insights into potential mechanisms of metal resistance, and exemplify the benefits of sampling remote locations for prospecting new molecular determinants.201829479501
121240.9886Virulence Factors and Antimicrobial Resistance of Uropathogenic Escherichia coli EQ101 UPEC Isolated from UTI Patient in Quetta, Balochistan, Pakistan. Infectious diseases have been tremendously increasing as the organisms of even normal flora become opportunistic and cause an infection, and Escherichia coli (E. coli EQ101) is one of them. Urinary tract infections are caused by various microorganisms, but Escherichia coli is the primary cause of almost 70%-90% of all UTIs. It has multiple strains, possessing diverse virulence factors, contributing to its pathogenicity. Furthermore, these virulent strains also can cause overlapping pathogenesis by sharing resistance and virulence factors among each other. The current study is aimed at analyzing the genetic variants associated with multi-drug-resistant (MDR) E. coli using the whole genome sequencing platform. The study includes 100 uropathogenic Escherichia coli (UPEC) microorganisms obtained from urine samples out of which 44% were multi-drug-resistant (MDR) E. coli. Bacteria have been isolated and antimicrobial susceptibility test (AST) was determined by disk diffusion method on the Mueller-Hinton agar plate as recommended by the Clinical and Laboratory Standards Institute (CLSI) 2020, and one isolate has been selected which shows resistance to most of the antibiotics, and that isolate has been analyzed by whole genome sequencing (WGS), accompanied by data and phylogenetic analysis, respectively. Organisms were showing resistance against ampicillin (10 μg), cefixime (5 μg), ceftriaxone (30 μg), nalidixic acid (30 μg), ciprofloxacin (5 μg), and ofloxacin (5 μg) on antimicrobial susceptibility test. WGS were done on selected isolate which identified 25 virulence genes (air, astA, chuA, fyuA, gad, hra, iha, irp2, iss, iucC, iutA, kpsE, kpsMII_K1, lpfA, mchF, ompT, papA_F43, sat, senB, sitA, terC, traT, usp, vat, and yfcV) and seven housekeeping genes (adk, fumC, gyrB, icd, mdh, purA, and recA). Among resistance genes, seven genes (TolC, emrR, evgA, qacEdelta1, H-NS, cpxA, and mdtM) were identified to be involved in antibiotic efflux, three AMR genes (aadA5, mphA, and CTX-M-15) were involved in antibiotic inactivation, and two genes (sul1 and dfrA14) were found to be involved in antibiotic drug replacement. Our data identified antibiotic resistance and virulence genes of the isolate. We suggest further research work to establish region-based resistance profile in comparison with the global resistance pattern.202337727279
209350.9885Are Enterobacteriaceae and Enterococcus Isolated from Powdered Infant Formula a Hazard for Infants? A Genomic Analysis. Powdered infant formulas (PIF) are the most used dietary substitutes that are used in order to supplement breastfeeding. However, PIF are not sterile and can be contaminated with different microorganisms. The objective of this study was to genomically characterize Enterobacteriaceae (ENT) and Enterococcus strains that were isolated from PIF. Strains were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and whole-genome sequencing (WGS). Genomic typing, detection of virulence, and resistance profiles and genes were performed with the Ridom SeqSphere+ software; the comprehensive antibiotic resistance database (CARD) platform; ResFinder and PlasmidFinder tools; and by the disk diffusion method. Nineteen isolates from PIF were analyzed, including ENT such as Kosakonia cowanii, Enterobacter hormaechei, Franconibacter helveticus, Mixta calida, and lactic acid bacteria such as Enterococcus faecium. The strains exhibited resistance to beta-lactams, cephalosporins, and macrolides. Resistance genes such as AcrAB-TolC, marA, msbA, knpEF, oqxAB, fosA, bla(ACT-)(7), bla(ACT-)(14,)qacJ, oqxAB(,)aac(6')-Ii, and msr(C); and virulence genes such as astA, cheB, cheR, ompA ompX, terC, ironA, acm, and efaAfm, adem were also detected. All the analyzed strains possessed genes that produced heat-shock proteins, such as IbpA and ClpL. In PIF, the presence of ENT and Enterococcus that are multiresistant to antibiotics-together with resistance and virulence genes-pose a health risk for infants consuming these food products.202236429148
541660.9885Limited predictive power of known resistance genes for phenotypic drug resistance in clinical Mycobacterium abscessus complex from Beijing in China. Mycobacterium abscessus complex (MABC) is an emerging pathogen with intrinsic multidrug resistance. Genomic sequencing technology has been widely applied to predict bacterial resistance in other bacteria, but the catalog of known resistance-determining genes to explain phenotypic resistance in the MABC is incomplete for many antibiotics. Eighty-one MABC strains were isolated from sputum samples of patients with pulmonary disease in the Beijing Chest Hospital. All isolates were tested for minimum inhibitory concentrations (MICs) to eight antibiotics and underwent whole-genome sequencing (WGS). Of the total 81 MABC isolates, six strains exhibited clarithromycin (CLM) resistance by day 3 in culture, but only one (16.7%, 1/6) contained a mutation in the rrl gene. All M. abscessus strains contained the erm (41)28T (100.0%, 49/49) polymorphism and exhibited CLM-induced resistance after 14 days in culture. Of the 61 imipenem-resistant strains, 12 (19.7%, 12/61) had mutations in the bla gene. Although there were four (4.9%) amikacin-resistant, nine (11.1%) linezolid-resistant, eight (9.9%) clofazimine-resistant, 23 (28.4%) bedaquiline-resistant, and 27 (33.3%) cefoxitin-resistant strains, no known mutations associated with resistance to these antibiotics were found. These results suggest that the explanatory power of known resistance genes for clinical MABC resistance is limited and that other unidentified genes or novel resistance mechanisms may be involved.202540422286
614970.9885Characterization and whole-genome sequencing of an extreme arsenic-tolerant Citrobacter freundii SRS1 strain isolated from Savar area in Bangladesh. Citrobacter freundii SRS1, gram-negative bacteria, were isolated from Savar, Bangladesh. The strain could tolerate up to 80 mmol L(-1) sodium arsenite, 400 mmol L(-1) sodium arsenate, 5 mmol L(-1) manganese sulfate, 3 mmol L(-1) lead nitrate, 2.5 mmol L(-1) cobalt chloride, 2.5  mmol L(-1) cadmium acetate, and 2.5 mmol L(-1) chromium chloride. The whole-genome sequencing revealed that the genome size of C. freundii SRS1 is estimated to be 5.4 Mb long, and the G + C content is 51.7%. The genome of C. freundii SRS1 contains arsA, arsB, arsC, arsD, arsH, arsR, and acr3 genes for arsenic resistance; czcA, czcD, cbiN, and cbiM genes for cobalt resistance; chrA and chrB genes for chromium resistance; mntH, sitA, sitB, sitC, and sitD genes for manganese resistance; and zntA gene for lead and cadmium resistance. This novel acr3 gene has never previously been reported in any C. freundii strain except SRS1. A set of 130 completely sequenced strains of C. freundii was selected for phylogenomic analysis. The phylogenetic tree showed that the SRS1 strain is closely related to the C. freundii 62 strain. Further analyses of the genes involved in metal and metalloid resistance might facilitate identifying the mechanisms and pathways involved in high metal resistance in the C. freundii SRS1 strain.202336332226
247880.9885Study on the resistance mechanism via outer membrane protein OprD2 and metal β-lactamase expression in the cell wall of Pseudomonas aeruginosa. The aim of the present study was to evaluate the imipenem-resistant mechanism via the outer membrane protein (OMP) OprD2 and metal β-lactamase expression in the cell wall of Pseudomonas aeruginosa. The Pseudomonas aeruginosa was clinically separated and validated by VITEK-2 full-automatic bacteria analyzer. Drug resistance, sensitive antibiotics and minimum inhibitory concentration (MIC) were tested using the drug sensitivity analysis system. The phenotype positive strains of MBL genes were screened using the Kirby-Bauer diffusion method by adding metal ion-chelating agent EDTA on the imipenem susceptibility paper. IMP-1, VIM-1 and SPM metaloenzyme genes were tested by polymerase chain reaction (PCR)-telomeric repeat amplification protocol (TRAP). The OMP OprD2 genes were tested by PCR-TRAP, and the protein expression was tested using western blot analysis. The location of OMP OprD2 was confirmed using the sodium salicylate inhibition test. The results showed that 80 portions (40%) of MBL-positive strains were screened out of 200 specimens. Imipenem-resistant Pseudomonas aeruginosa (IRPA) and MIC values were significantly higher than quality control bacteria and control bacteria (P<0.05). A total of 35 cases with IMP-1 positive, 20 with VIM-1 positive, 16 with SPM positive, 5 with 2 positive genes and 4 with 3 positive genes were screened among MBL positive strains. A total of 150 portions (75%) of OprD2 deficiencies were screened from 200 specimens. The standard strains and sensitive strains showed OprD2 protein bands at 45 kDa while no OprD2 protein bands appeared in OprD2 deficiency strains. It was in accordance with gene detection. In conclusion, OMP OprD2 deficiency and MBL phenotype positivity may be important mechanisms of IRPA.201627882088
245490.9885Colistin resistance in Gram-negative bacteria analysed by five phenotypic assays and inference of the underlying genomic mechanisms. BACKGROUND: Colistin is used against multi-drug resistant pathogens, yet resistance emerges through dissemination of plasmid-mediated genes (mcr) or chromosomal mutation of genes involved in lipopolysaccharide synthesis (i.e. mgrB, phoPQ, pmrCAB). Phenotypic susceptibility testing is challenging due to poor diffusion of colistin in agar media, leading to an underestimation of resistance. Performance of five phenotypic approaches was compared in the context of different molecular mechanisms of resistance. We evaluated Vitek 2® (bioMérieux, AST N242), Colistin MIC Test Strip (Liofilchem Diagnostici), UMIC (Biocentric), and Rapid Polymyxin™ NP test (ELITechGroup) against the standard broth microdilution (BMD) method. We used whole genome sequencing (WGS) to infer molecular resistance mechanisms. We analysed 97 Enterobacterales and non-fermenting bacterial isolates, largely clinical isolates collected up to 2018. Data was analysed by comparing susceptibility categories (susceptible or resistant) and minimal inhibitory concentrations (MIC). Susceptibility category concordance is the percentage of test results sharing the same category to BMD. MIC concordance was calculated similarly but considering ±1 MIC titre error range. We determined genomic diversity by core genome multi locus sequencing typing (cgMLST) and identified putative antimicrobial resistance genes using NCBI and CARD databases, and manual annotation. RESULTS: Of 97 isolates, 54 (56%) were resistant with standard BMD. Highest susceptibility category concordance was achieved by Rapid Polymyxin™ NP (98.8%) followed by UMIC (97.9%), Colistin E-test MIC strip (96.9%) and Vitek 2® (95.6%). Highest MIC concordance was achieved by UMIC (80.4%), followed by Vitek 2® (72.5%) and Colistin E-test MIC strip (62.9%). Among resistant isolates, 23/54 (43%) were intrinsically resistant to colistin, whereas 31/54 (57%) isolates had acquired colistin resistance. Of these, mcr-1 was detected in four isolates and mcr-2 in one isolate. Non-synonymous mutations in mgrB, phoQ, pmrA, pmrB, and pmrC genes were encountered in Klebsiella pneumoniae, Escherichia coli, and Acinetobacter bereziniae resistant isolates. Mutations found in mgrB and pmrB were only identified in isolates exhibiting MICs of ≥16 mg/L. CONCLUSIONS: The Rapid Polymyxin™ NP test showed highest categorical concordance and the UMIC test provided MIC values with high concordance to BMD. We found colistin resistance in diverse species occurred predominantly through spontaneous chromosomal mutation rather than plasmid-mediated resistance.202134798825
2453100.9884Prevalence and molecular determinants of colistin resistance among commensal Enterobacteriaceae isolated from poultry in northwest of Iran. BACKGROUND: The emergence of colistin-resistant Enterobacteriaceae from human and animal sources is a public health concern as this antibiotic is considered to be the last line therapeutic option for infections caused by multidrug-resistant Gram-negative bacteria. Here we aimed to determine the prevalence of colistin resistance, among enterobacteria isolated from poultry and the possible underlying colistin resistance mechanisms. METHODS: A collection of 944 cloacal samples were obtained from poultry and screened for colistin resistance. To uncover the molecular mechanism behind colistin resistance, the presence of plasmid encoded colistin resistance genes mcr-1, mcr-2, mcr-3 and mcr-4 was examined by PCR. The nucleotide sequences of the mgrB, pmrA, pmrB, phoP, phoQ, crrA and crrB genes were determined. The genetic relatedness of the colistin resistant (ColR) isolates was evaluated by Multilocus sequence typing. Three ColR mutants were generated in vitro by repetitive drug exposure. RESULTS: Overall from 931 enteric bacteria isolated from poultry samples obtained from 131 farms, nine ColR bacteria (0.96%) with high level colistin resistance (MICs ≥ 64 mg/L) were detected all being identified as K. pneumoniae. The 9 ColR bacteria originated from different farms and belonged to 7 distinct Sequence types including ST11 (22.2%) and ST726 (22.2%) being the most prevalent STs followed by ST37, ST74, ST485, ST525 and novel sequence type 3380 (11.1% each). mcr-type genes were not detected in any isolate. In 88.8% of the isolates (n = 8), MgrB was inactivated by Insertion of IS elements (IS1-like, IS3-like, IS5-like families, positions + 75, + 113, + 117, + 135) and nonsense mutations at codons 8, 16, 30. All ColR isolates harboured wild type PmrA, PhoP, PhoQ or polymorphic variants of PmrB. Sequence analysis of the CrrB revealed a familiar S195N and 4 novel I27V, T150R, F303S and K325R substitutions. PmrB T93N substitution and mgrB locus deletion were identified in two laboratory induced ColR mutants and one mutant lacked alteration in the studied loci. In one ColR isolate with wild type MgrB an A83V substitution was detected in CrrA. CONCLUSION: It is concluded from our results that colistin resistance in the studied avian K. pneumoniae isolates was mostly linked to alterations identified within the mgrB gene.201930728861
1263110.9884Antimicrobial Resistance and Antimicrobial Activity of Staphylococcus lugdunensis Obtained from Two Spanish Hospitals. Staphylococcus lugdunensis is a coagulase-negative-staphylococci (CoNS) that lately has gained special attention in public health as a human pathogen and also as a bacteriocin-producer bacteria. In this study, we characterized 56 S. lugdunensis isolates recovered from human samples in two Spanish hospitals. Antimicrobial susceptibility testing was performed and antimicrobial resistance and virulence genotypes were determined. Antimicrobial activity (AA) production was evaluated by the spot-on-lawn method against 37 indicator bacteria, including multidrug-resistant (MDR) isolates, and the presence of the lugD gene coding for lugdunin bacteriocin was analyzed by PCR. The antibiotic resistance detected was as follows (% resistance/genes detected): penicillin (44.6%/blaZ), oxacillin (1.8%/mecA on SCCmec-V), erythromycin-clindamycin inducible (7.1%/erm(C), msrA), tetracycline (5.3%/tetK), gentamicin and/or tobramycin (3.6%/ant(4')-Ia, acc(6')-aph(2″)), and fosfomycin (21.4%). A MDR phenotype was detected in 5% of isolates. Twenty-one of the S. lugdunensis isolates showed susceptibility to all 20 antibiotics tested (37.5%). The screening for AA revealed 23 antimicrobial producer (AP) isolates with relevant inhibition against coagulase-positive-staphylococci (CoPS), including both methicillin-susceptible and -resistant S. aureus. The lugD gene was detected in 84% of the 56 S. lugdunensis isolates. All of the AP S. lugdunensis isolates (n = 23) carried the lugD gene and it was also detected in 24 of the non-AP isolates, suggesting different gene expression levels. One of the AP isolates stood out due to its high antimicrobial activity against more than 70% of the indicator bacteria tested, so it will be further characterized at genomic and proteomic level.202235893538
1333120.9884Virulence-encoding genes related to extraintestinal pathogenic E. coli and multidrug resistant pattern of strains isolated from neonatal calves with different severity scores of umbilical infections. Umbilical infections in calves comprise a major cause of neonatal mortality and have been related to a variety of microorganisms. E. coli is an opportunistic enteropathogen characterized by a diversity of virulence factors (VF). Nonetheless, the gene profiles that encode VF associated with umbilical infections in calves and their effect on the clinical severity remains unclear. In this scenario, microbial identification (with an emphasis on E. coli), was carried out among 150 neonatal calves (≤30 days of age) with umbilical infections, where the omphalopathies were clinically scored as mild, moderate, or severe. Also, a panel of 16 virulence-encoding genes related to extraintestinal pathogenic E. coli (ExPEC) were investigated, i.e., fimbriae/adhesins (sfa/focDEa, papA, papC, afaBC), toxins (hlyA, sat, cnf1, cdt), siderophores (iroN, irp2, iucD, ireA), invasins (ibeA), and serum resistance (ompT, traT, kpsMT II). Bacteria and yeasts isolates were identified using mass spectrometry. Bacteria, yeasts, and fungi were isolated in 94.7% (142/150) of neonatal calves sampled. E. coli was the agent most frequently isolated (59/150 = 39.3%), in pure culture (27/59 = 45.8%) and combined infections (32/59 = 54.2%), although a great variety (n = 83) of other species of microorganisms were identified. Clinical severity scores of 1, 2, and 3 were observed in 32.2% (19/59), 23.7% (14/59), and 44.1% (26/59) of E. coli infections, respectively. The ExPEC genes detected were related to serum resistance (traT, 42/59 = 72.2%; ompT, 35/59 = 59.3%, kpsMTII, 10/59 = 17%), invasins (ibeA, 11/59 = 18.6%), siderophores (iucD, 9/59 = 15.3%; iroN, 8/59 = 13.6%), and adhesins/fimbriae (papA, 8/59 = 13.6%; papC, 15/59 = 9.6%). The presence of each virulence gene was not associated with the case's clinical score. Among all isolates, 89.8% (53/59) showed in vitro resistance to sulfamethoxazole/trimethoprim and 59.3% to ampicillin (35/59), while 94.1% (55/59) revealed a multidrug resistant profile. Great complexity of bacteria, yeast, and fungi species was identified, reinforcing the umbilical infections of neonatal calves as a polymicrobial disorder. The high occurrence of E. coli (39.3%) highlights the role of this pathogen in the etiology of umbilical infections in calves. Furthermore, a panel of ExPEC genes was investigated for the first time among calves that were clinically scored for case severity. The high prevalence of traT and ompT indicates that these serum resistance-related genes could be used as biomarkers for further investigations of ExPEC isolates from umbilical infections. Our results contribute to the etiological investigation, clinical severity scoring, antimicrobial resistance pattern, and virulence-related to ExPEC genes involved in umbilical infections of neonatal calves.202336427660
5216130.9883Unraveling the draft genome and phylogenomic analysis of a multidrug-resistant Planococcus sp. NCCP-2050(T): a promising novel bacteria from Pakistan. Planococcus is a genus of Gram-positive bacteria known for potential industrial and agricultural applications. Here, we report the first draft genome sequence and phylogenomic analysis of a CRISPR-carrying, multidrug-resistant, novel candidate Planococcus sp. NCCP-2050(T) isolated from agricultural soil in Pakistan. The strain NCCP-2050(T) exhibited significant resistance to various classes of antibiotics, including fluoroquinolones (i.e., ciprofloxacin, levofloxacin, ofloxacin, moxifloxacin, and bacitracin), cephalosporins (cefotaxime, ceftazidime, cefoperazone), rifamycins (rifampicin), macrolides (erythromycin), and glycopeptides (vancomycin). Planococcus sp. NCCP-2050(T) consists of genome size of 3,463,905 bp, comprised of 3639 annotated genes, including 82 carbohydrate-active enzyme genes and 39 secondary metabolite genes. The genome also contained 80 antibiotic resistance, 162 virulence, and 305 pathogen-host interaction genes along with two CRISPR arrays. Based on phylogenomic analysis, digital DNA-DNA hybridization, and average nucleotide identity values (i.e., 35.4 and 88.5%, respectively) it was suggested that strain NCCP-2050(T) might represent a potential new species within the genus Planococcus. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-023-03748-z.202337663752
1250140.9883Distribution of 16S rRNA methylases among different species of Gram-negative bacilli with high-level resistance to aminoglycosides. 16S rRNA methylases confer high-level resistance to most aminoglycosides in Gram-negative bacteria. Seven 16S rRNA methylase genes, armA, rmtA, rmtB, rmtC, rmtD, rmtE and npmA, have been identified since 2003. We studied the distribution of methylase genes in more than 200 aminoglycoside-resistant Gram-negative clinical isolates collected in 2007 at our hospital in Shanghai, China. 16S rRNA methylase genes were amplified by polymerase chain reaction (PCR) among 217 consecutive clinical isolates of Gram-negative bacilli resistant to gentamicin and amikacin by a disk diffusion method. 16S rRNA methylase genes were present in 97.5% (193/198) of clinical isolates highly resistant to amikacin (≥512 μg/ml), with armA and rmtB detected in 67.2 and 30.3% of strains, respectively, while no 16S rRNA methylase genes were detected in 19 strains with amikacin minimum inhibitory concentration (MIC) ≤256 μg/ml. armA or rmtB genes were detected in 100% of 104 strains of Enterobacteriaceae, and these two genes were equally represented (49 vs. 55 strains). Genes for armA or rmtB were detected in 94.7% (89/94) of Acinetobacter baumannii and Pseudomonas aeruginosa strains, and armA was predominant (84 vs. 5 strains with rmtB). No rmtA, rmtC, rmtD or npmA genes were found. Enterobacterial repetitive intergenic consensus sequence (ERIC-PCR) indicated that armA and rmtB genes were spread by both horizontal transfer and clonal dissemination.201020614151
1246150.9883Ciprofloxacin-resistant Gram-negative isolates from a tertiary care hospital in Eastern India with novel gyrA and parC gene mutations. BACKGROUND: Expanded-spectrum quinolones (ciprofloxacin) are highly effective against gram-negative bacteria, but significant resistance to quinolones has been increasingly reported. We sought to evaluate the prevalence of gram-negative ciprofloxacin-resistant isolates (CRIs) from our hospital and their mechanism of action. METHODS: Gram-negative CRIs were identified as per standard procedures and confirmed using the Ezy MICTM Strip (HiMedia). DNA from 67 CRIs was amplified for the quinolone resistance-determining region (QRDR) and plasmid-mediated quinolone resistance genes. Thirty isolates positive for QRDR DNA were sequenced by Sanger's method to detect mutation. RESULTS: Of the isolates, 42.5% were found to be CRIs, the majority (74.42%) from inpatient departments, and E scherichia coli (64.19%) was the predominant isolate. Among the CRIs, 24.55% were ESBL producers and 35.29% were multidrug resistant. The polymerase chain reaction results showed the majority were amplified by QRDR target regions of gyrA (35.4%) while 4.61% were amplified for the plasmid-mediated fluoroquinolone resistance region of the qnrB gene. Further sequencing of QRDR-positive genes showed point mutations with amino acid changes at codons Ser83 and Asp87 in the gyrA gene and Ser80, Glu84, and Leu88 positions in the parC gene. CONCLUSION: Ciprofloxacin resistance observed in our study was mostly due to point mutations. Hence, strategies for rational use of ciprofloxacin and adherence to the dose and duration of treatment could be helpful to prevent selection and spread of mutant CRIs/strains.202235035040
2293160.9883Mechanisms of Resistance in Clinical Isolates of Enterobacter cloacae that Are Less Susceptible to Cefepime than to Ceftazidime. Thirty-two Enterobacter cloacae strains that are less susceptible to cefepime than to ceftazidime were collected. This unique phenotype of 8 strains was confirmed using the agar dilution method. OXA1, OXA10, OXA31 and OXA35 were detected in 3, 2, 3, and 2 strains, respectively, whereas all strains were negative for PSE-1 genes. OXA genes were also identified in the plasmid DNA of 5 strains, but only 2 strains were positive in a conjugation experiment. The acrA, acrB and tolC genes were identified in 4, 4 and 6 strains, respectively. Decreased expression of the acrA mRNA and overexpression of the acrB and tolC mRNAs were observed using real-time RT-PCR. Most of the bacteria (n=7) stably expressed the marA gene, which is a regulatory gene in the AcrAB-TolC multidrug efflux system, whereas all strains were negative for ramA. The acrA, acrB, tolC, acrR and marA genes were similar to the genes in reference strains in GenBank, with nucleotide homologies of 96%, 98%, 98%, 98% and 100%, respectively. In conclusion, the mechanism of resistance of Enterobacter cloacae with less susceptibility to cefepime than to ceftazidime is associated with the overexpression of AcrAB-TolC and the production of OXA1, XA10, OXA31 and OXA35.201829970440
1334170.9882Intimin (eae) and virulence membrane protein pagC genes are associated with biofilm formation and multidrug resistance in Escherichia coli and Salmonella enterica isolates from calves with diarrhea. OBJECTIVES: This study aimed to evaluate the association of the intimin (eae) and pagC genes with biofilm formation and multidrug resistance (MDR) phenotype in Escherichia coli and Salmonella enterica collected from calves with diarrhea. RESULTS: Fecal samples (n: 150) were collected from calves with diarrhea. Of 150 fecal samples, 122 (81.3%) were culture positive and 115/122 (94.2%) were Gram-negative bacteria. Among them, E. coli (n = 64/115, 55.6%) was the most common isolate followed by S. enterica (n = 41/115, 35.6%). Also, 10 (8.6%) isolates were other Enterobacteriaceae bacteria including Klebsiella and Proteus species. Eighty-nine isolates (77.4%) from calf diarrhea, including 52 (81.3%) E. coli and 37 (90.2%) S. enterica were MDR. The eae and pagC genes were detected in 33 (51.5%) E. coli and 28 (68.3%) S. enterica isolates, respectively. There was a strong association between these genes and biofilm formation and MDR phenotype (P-value = 0.000). All E. coli isolates carrying the eae gene were biofilm producers and MDR. Also, all pagC-positive S. enterica isolates were MDR and 25 (89.3%) isolates of them produced biofilm.202236221149
2289180.9882Comprehensive Molecular Profiling of AcrAB-TolC Efflux Pump Genes in Salmonella typhi Isolates from Typhoid Infected Patients. Salmonella typhi is a facultative anaerobic, rod-shaped, Gram-negative bacterium that causes typhoid fever, a potentially fatal systemic infection. This study aimed to characterize antibiotic susceptibility patterns, mutations at the molecular level, and efflux pump genes in clinical isolates. In this study, blood samples (n = 2950) were collected from suspected typhoid-infected patients, and 380 (12.88%) bacterial isolates were found, comprising 144 (37.89%) Gram-positive and 236 (62.10%) Gram-negative bacteria. S. typhi was identified in 95 isolates (25%), corresponding to an overall prevalence of 3.22%. Biochemical identification was performed by Analytical Profile Index (API) 20-E strips, and molecular identification was done by partial 16S rRNA gene using PCR. The S. typhi isolates were categorized into multidrug-resistant (MDR), 13 (13.68%), and extensively drug-resistant (XDR), 82 (86.31%), and their resistance patterns were recorded. Ampicillin (98.94%) and chloramphenicol (93.68%) showed the highest antibiotic resistance profiles, while azithromycin and meropenem exhibited no resistance. Numerous mutations were found in acrA, acrB, and tolC genes after sequencing; TolC (MDR) showed the highest score (16 points), and AcrB (MDR) displayed the lowest score (9 points). I-Mutant 2.0 was used to assess mutations and calculate the reliability index (RI), whereas trRosetta and Discovery Studio were used to predict and refine 3D protein models. Consensus sequences of the selected genes were analyzed to construct phylogenetic trees illustrating evolutionary relationships with other Salmonella enterica serovars. The study emphasizes the concerning multidrug resistance of S. typhi isolates as well as notable mutations (genetic changes) that may affect efflux pump activity and contribute to resistance.202540844743
2131190.9882Alistipes Bacteremia in Older Patients with Digestive and Cancer Comorbidities, Japan, 2016-2023. The clinical characteristics of Alistipes bacteremia remain insufficiently understood. We retrospectively analyzed 13 cases of Alistipes bacteremia at a tertiary care center in Japan. Of the 13 patients, 7 were male and 6 female; 10 were >65 years of age. Of 9 patients with comorbidities, 7 had solid tumors or hematological malignancies and 11 had gastrointestinal symptoms. Isolates identified were Alistipes finegoldii in 4 cases, A. onderdonkii in 4, A. putredinis in 3, A. indistinctus in 2, and A. ihumii in 1. Ten strains exhibited low MICs against β-lactam/β-lactamase inhibitors and metronidazole. We observed high MICs against penicillin, ceftriaxone, and minocycline. Several strains harbored antimicrobial resistance genes, including adeF, tet(Q), cfxA3, cfxA4, and ermG. Twelve patients received β-lactam/β-lactamase inhibitors; 2 patients with solid tumors experienced septic shock and died. Alistipes bacteria can translocate from the gastrointestinal tract into the bloodstream, particularly in cases of inflammation, obstruction, or perforation, leading to severe infections.202540133031