Multiple mechanisms of linezolid resistance in Staphylococcus haemolyticus detected by whole-genome sequencing.

Introduction. Linezolid is an effective therapeutic option for treating severe infections caused by multidrug-resistant Gram-positive organisms. Several mechanisms have been reported to be responsible for resistance to this antibiotic.Hypothesis or Gap Statement. Although several mechanisms of linezolid resistance have been reported in Staphylococcus haemolyticus, the prevalence and potential for horizontal transfer of resistance genes have not been fully characterized, particularly among S. haemolyticus isolates from India.Aim. To perform whole-genome sequencing (WGS) of linezolid-resistant S. haemolyticus isolates to characterize the resistance mechanisms.Methodology. WGS was performed for 16 linezolid-resistant S. haemolyticus isolates to check for the presence of cfr, optrA and poxtA genes and mutations in 23S rRNA and ribosomal proteins (L3, L4 and L22) that are possible mechanisms implicated in linezolid resistance. Sequence types were identified using MLST finder. The minimum inhibitory concentration (MIC) of linezolid was determined using the E-test method. Polymerase chain reaction (PCR) was carried out for the detection of the cfr gene.Results. The study documented three different mechanisms of linezolid resistance in S. haemolyticus. Thirteen of the 16 isolates were phenotypically resistant to linezolid, of which 12 were positive for the cfr gene. The G2603T mutation in 23S rRNA was found in the majority of the isolates (n=13). Ten isolates had the R138V mutation in L3 ribosomal protein. Twelve isolates with the cfr gene in combination with either G2603T or R138V mutations displayed extremely high MIC values. Surprisingly, three phenotypically sensitive isolates were found to be positive for the cfr gene but negative for other resistance mechanisms. Importantly, in almost half of the isolates the cfr gene was present on a plasmid. ST3 and ST1 were found to be the predominant sequence types.Conclusion. All phenotypically resistant isolates exhibited two or three linezolid resistance mechanisms. The cfr gene was found on plasmids in many isolates, demonstrating its potential for horizontal transfer to more pathogenic organisms.

Genome sequences of bla NDM-1 producing Acinetobacter lactucae isolated from immunocompromised patients in India.

Among the species within Acb-complex, Acinetobacter lactucae has not been frequently isolated from clinical settings, unlike Acinetobacter baumannii, which is an important nosocomial pathogen. We report the genomic sequences of A. lactucae strains (PKAL1732 and 1828C) harboring multiple-resistance determinants including metallo-ß-lactamase (bla NDM-1) isolated from immunocompromised patients admitted to a referral hospital in India.

Bacterial effluxome as a barrier against antimicrobial agents: structural biology aspects and drug targeting.

Antimicrobial resistance (AMR) is fast becoming a medical crisis affecting the entire global population. The bacterial membrane is the first layer of defense for the bacteria against antimicrobial agents (AMA), specifically transporters in the membrane efflux these AMA out of the bacteria and plays a significant role in the AMR development. Understanding the structure and the functions of these efflux transporters is essential to overcome AMR. This review discusses efflux transporters (primary, secondary, and tripartite), their domain architectures, substrate specificities, and efflux pump inhibitors (EPI). Special emphasis on nosocomial ESKAPEE (Enterococcus faecium., Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter spp. and Escherichia coli) pathogens, their multidrug efflux targets and inhibitors are discussed. Deep knowledge about the functioning of efflux pumps and their structural aspects will open up opportunities for developing new EPI, which could be used along with AMA as combination therapy to overcome the emerging AMR crisis.

Coexistence of blaNDM-1, blaOXA-51, blaOXA-23, and armA in conjunction with novel mutations detected in RND efflux pump regulators in tigecycline resistant clinical isolates of Acinetobacter baumannii.

This study has investigated a total of 51 Acinetobacter baumannii isolates for the prevalence of resistant determinants in tigecycline susceptible and non-susceptible clinical isolates of A. baumannii. Antimicrobial susceptibility testing revealed 74% of isolates were tigecycline resistant. Mutations in RND-efflux pump regulatory genes and the expression of efflux pump genes were measured in tigecycline resistant isolates. There was a strong co-relation between the blaNDM-1 and armA wherein majority of the isolates that are positive for blaNDM-1 have also harbored armA. Compared with TSAB (tigecycline susceptible A. baumannii), TNAB (tigecycline non-susceptible A. baumannii) isolates show increased distribution of blaNDM-1 (P = 0.048), blaIMP-1 (P< 0.0001) and blaOXA-51 (P = 0.0029) carbapenemase genes. The variants of RND-efflux pump regulatory genes due to amino-acid mutations in adeS (F12S, K84E, W61R, N268H and Q299R) and adeL (G21R and Q262R) were identified in tigecycline resistant isolates as well as ISAba1 mediated disruption of adeN were observed causing overexpression of adeIJK efflux pump. Additionally, mutations in adeRS were also associated with increased expression of adeABC efflux pump. Besides, TNAB isolates showed significantly (P< 0.0001) higher ability of biofilm formation as compared to TSAB isolates. The tigecycline resistance due to mutations in contemporary A. baumannii isolates having a higher ability to form biofilm may pose therapeutic difficulties.