Mismatch amplification mutation assay-polymerase chain reaction: A method of detecting fluoroquinolone resistance mechanism in bacterial pathogens

The mismatch amplification assay is a modified version of polymerase chain reaction (PCR) that permits specific amplification of gene sequences with single base pair change. The basis of the technique relies on primer designing. The single nucleotide mismatch at the 3' proximity of the reverse oligonucleotide primer makes Taq DNA polymerase unable to carry out extension process. Thus, the primers produce a PCR fragment in the wild type, whereas it is not possible to yield a product with a mutation at the site covered by the mismatch positions on the mismatch amplification mutation assay (MAMA) primer from any gene. The technique offers several advantages over other molecular methods, such as PCR-restriction fragment length polymorphism (RFLP) and oligonucleotide hybridization, which is routinely used in the detection of known point mutations. Since multiple point mutations in the quinolone resistance determining region play a major role in high-level fluoroquinolone resistance in Gram-negative bacteria, the MAMA-PCR technique is preferred for detecting these mutations over PCR-RFLP and sequencing technology.

CTX-M type extended-spectrum ?-lactamase in Escherichia coli isolated from extra-intestinal infections in a tertiary care hospital in south India

Background & objectives: Infections caused by extended-spectrum ?-lactamase (ESBL)-producing Escherichia coli carrying blaCTX-M genes have been spreading globally, but there are geographical variations in the type of blaCTX-Mgenes prevalent and there are scanty data from India. This study was conducted to determine the CTX-M type ESBLs in E. coli isolates obtained from clinical specimens from patients with extra-intestinal infections attending a tertiary care hospital in south India. Methods: ESBL-producing E. coli isolated from patients with extra-intestinal infections were subjected to PCR using CTX-M group-specific primers. From a representative isolate, full-length CTX-M-15 gene was amplified and sequenced. An internal fragment of this gene was sequenced in 10 representative isolates. Results: Of the 300 isolates of E. coli tested, 88 per cent carried CTX-M genes and blaCTX-M-15was the most dominant gene present in 90 per cent of the positive isolates. Most (91%) of the isolates positive for blaCTX-M were sensitive to meropenem. Interpretation & conclusions: Our findings showed blaCTX-M-15 to be the dominant gene. Based on the data on antimicrobial susceptibility, cefoperazone-sulbactum could be an antimicrobial of choice.

Plasmid-mediated fluoroquinolone resistance associated with extra-intestinal Escherichia coli isolates from hospital samples

Background & objectives: Infection from fluoroquinolone-resistant extra-intestinal Escherichia coli is a global concern. In this study, isolation and characterization of fluoroquinolone-resistant extra-intestinal E. coli isolates obtained from hospital samples were undertaken to detect plasmid-mediated quinolone resistance (PMQR) genes. Methods: Forty three isolates of E. coli obtained from patients with extra-intestinal infections were subjected to antibiogram to detect fluoroquinolone resistance. The mechanism of fluoroquinolone resistance was determined by the detection of PMQR genes and mutations in quinolone resistance determining region (QRDR). Results: Of the 43 isolates, 36 were resistant to nalidixic acid (83.72%) and 28 to ciprofloxacin (65.11%). Eight E. coli isolates showed total resistance to both the antimicrobials without any minimum inhibitory concentration. The detection of PMQR genes with qnr primers showed the presence of qnrA in two, qnrB in six and qnrS in 21 isolates. The gene coding for quinolone efflux pump (qepA) was not detected in any of the isolates tested. The presence of some unexpressed PMQR genes in fluoroquinolone sensitive isolates was also observed. Interpretation & conclusions: The detection of silent PMQR genes as observed in the present study presents a risk of the transfer of the silent resistance genes to other microorganisms if present in conjugative plasmids, thus posing a therapeutic challenge to the physicians. Hence, frequent monitoring is to be done for all resistance determinants.

An unusual case of unresolving tunnel infection in a patient on continuous ambulatory peritoneal dialysis

Atypical mycobacteria remain a rare cause of peritoneal dialysis catheter-related tunnel infection (TI) and poses serious risk because of the resistant nature to most antibiotic therapy. Non-tubercular mycobacterial infections lead to chronicity requiring peritoneal dialysis catheter removal. We report an 82-year-old male, with diabetic nephropathy who had a coinfection with Staphylococcus hominis and Mycobacterium abscessus who presented with pus discharge at exit site and TI. He was treated with relocation of the extraperitoneal part of the catheter with a new exit site without catheter removal and multidrug mycobacterial therapy.

Twenty-one episodes of peritonitis in a continuous ambulatory peritoneal dialysis patient: What is the root cause?

A 51-year-old female, with non-alcoholic liver cirrhosis, portal hypertension, type 2 diabetes mellitus, autosomal dominant polycystic kidney disease with a clipped cerebral aneurysm and chronic kidney disease stage 5 was on continuous ambulatory peritoneal dialysis (CAPD) for 6.5 years elsewhere. She came for opinion on continuation of CAPD as she had 21 episodes of peritonitis in 76 months. Her blood pressure was 80/50 mmHg. She was on haemodialysis with a temporary central access for 2 weeks. She had no abdominal tenderness, and exit site looked normal. Fluid was negative for Mycobacterium tuberculosis. Laparoscopically, we replaced the catheter with a new swan-neck Tenckhoff double-cuff catheter through a different exit site in the same sitting. Catheter-tip biofilm culture isolated Enterococcus casseliflavus. Peritoneal sampling biopsy showed evidence of fibrosis. She has adequate ultrafiltration and is currently on automated peritoneal dialysis for 5 months.

The effect of indocyanine green‑mediated photodynamic therapy as an adjunct to scaling and root planing in the treatment of chronic periodontitis: A comparative split‑mouth randomized clinical trial.

Objective: The aim of this study was to assess the effect of adjunctive photodynamic therapy (PDT) (using 810 nm diode laser and Indocyanine green as photosensitizer) in chronic periodontitis. Materials and Methods: Patients with untreated chronic periodontitis were included. Treatment was done according to a split mouth design. All sites received periodontal treatment comprising scaling and root-planing (SRP). Test group were additionally treated with PDT. Plaque Index (PI), Gingival Index (GI), Probing Pocket Depth (PPD) and Relative Attachment Level (RAL) were evaluated at baseline, 1 month and 3 months. Results: Mean baseline values for PI, GI, PPD and RAL were not different in the test group and control group. Statistical significant difference in PPD and RAL, 3 months after treatment was seen in test group as compared to the control group. Conclusions: In patients with chronic periodontitis, clinical outcomes of conventional SRP can be improved by adjunctive PDT.