Disulfiram enhances meropenem activity against NDM- and IMP-producing carbapenem-resistant Acinetobacter baumannii infections.

OBJECTIVES: To evaluate the in vitro and in vivo efficacy of the FDA-approved drug disulfiram in combination with meropenem against MBL-expressing carbapenem-resistant Acinetobacter baumannii. METHODS: Chequerboard and antibiotic resistance reversal analysis were performed using 25 clinical isolates producing different MBLs. Three representative strains harbouring NDM, IMP or non-MBL genes were subjected to a time-kill assay to further evaluate this synergistic interaction. Dose-dependent inhibition by disulfiram was assessed to determine IC50 for NDM-1, IMP-7, VIM-2 and KPC-2. Further, to test the efficacy of meropenem monotherapy and meropenem in combination with disulfiram against NDM- and IMP-harbouring A. baumannii, an experimental model of systemic infection and pneumonia was developed using BALB/c female mice. RESULTS: Chequerboard and antibiotic reversal assay displayed a synergistic interaction against MBL-expressing A. baumannii strains with 4- to 32-fold reduction in MICs of meropenem. In time-kill analysis, meropenem and disulfiram exhibited synergy against NDM- and IMP-producing carbapenem-resistant A. baumannii (CRAb) isolates. In vitro dose-dependent inhibition analysis showed that disulfiram inhibits NDM-1 and IMP-7 with IC50 values of 1.5 ± 0.6 and 16.25 ± 1.6 µM, respectively, with slight or no inhibition of VIM-2 (<20%) and KPC-2. The combination performed better in the clearance of bacterial load from the liver and spleen of mice infected with IMP-expressing CRAb. In the pneumonia model, the combination significantly decreased the bacterial burden of NDM producers compared with monotherapy. CONCLUSIONS: These results strongly suggest that the combination of disulfiram and meropenem represents an effective treatment option for NDM- and IMP-associated CRAb infections.

The unusual glycine-rich C terminus of the Acinetobacter baumannii RNA chaperone Hfq plays an important role in bacterial physiology.

Acinetobacter baumannii is a Gram-negative nosocomial pathogen that causes soft tissue infections in patients who spend a long time in intensive care units. This recalcitrant bacterium is very well known for developing rapid drug resistance, which is a combined outcome of its natural competence and mobile genetic elements. Successful efforts to treat these infections would be aided by additional information on the physiology of A. baumannii Toward that end, we recently reported on a small RNA (sRNA), AbsR25, in this bacterium that regulates the genes of several efflux pumps. Because sRNAs often require the RNA chaperone Hfq for assistance in binding to their cognate mRNA targets, we identified and characterized this protein in A. baumannii The homolog in A. baumannii is a large protein with an extended C terminus unlike Hfqs in other Gram-negative pathogens. The extension has a compositional bias toward glycine and, to a lower extent, phenylalanine and glutamine, suggestive of an intrinsically disordered region. We studied the importance of this glycine-rich tail using truncated versions of Hfq in biophysical assays and complementation of an hfq deletion mutant, finding that the tail was necessary for high-affinity RNA binding. Further tests implicate Hfq in important cellular processes of A. baumannii like metabolism, drug resistance, stress tolerance, and virulence. Our findings underline the importance of the glycine-rich C terminus in RNA binding, ribo-regulation, and auto-regulation of Hfq, demonstrating this hitherto overlooked protein motif to be an indispensable part of the A. baumannii Hfq.

High-sensitivity detection of human malaria parasites by the use of rapid diagnostic tests and nested polymerase chain reaction in burdened communities of North East India.

BACKGROUND AND OBJECTIVES: The study aimed to evaluate the diagnostic performance of malaria through microscopy and rapid diagnostic test (RDT) analysis performed locally and the accuracy evaluated by nested polymerase chain reaction (PCR) for diagnosis of Plasmodium falciparum from hotspot regions of North East (NE) India. MATERIALS AND METHODS: One thousand one hundred and seventy-three blood samples were collected for identification of P. falciparum infection using microscopy and RDT analysis. DNA was extracted from whole blood using QIAamp DNA blood mini kit, and nested PCR was performed to confirm P. falciparum for evaluating sensitivity and specificity from various epidemiological surveys and geographical areas of NE India. RESULTS: Of 1173 symptomatic malaria suspected patients, 15.6% (183/1173) patients were diagnosed as malaria positive by RDT and 67.94% cases (53/78) with microscopy. Of 183 malaria-positive patients, 42.62% (78/183) were diagnosed with P. falciparum and 84.61% (66/78) further confirmed to be P. falciparum positive by nested PCR. High sensitivity (97.9%) and low specificity (2.03%) of the RDT and high sensitivity (99.1%) and low specificity (0.9%) in microscopy against nested PCR results was statistically significant (P < 0.05). Epidemiological comparisons expressed highest incidences in Manipur (51.11%) followed by Meghalaya (48.93%) and Assam (35.16%). Overall incidence rate among the genders was observed to be higher in males than in females. CONCLUSIONS: Our findings suggest that PCR, RDT and microscopy can potentially determine hotspots at moderate transmission intensities, but PCR testing has a diagnostic advantage as transmission intensity falls. Therefore, malaria control programs should consider PCR testing when the prevalence of infection is low.