Bioactive extracts of Carum copticum L. enhances efficacy of ciprofloxacin against MDR enteric bacteria.

The widespread occurrence of extended spectrum ß-lactamases (ESßLs) producing enteric bacteria and their co-resistance with flouroquinolones has impaired the current antimicrobial therapy. This has prompted the search for new alternatives through synergistic approaches with herbal extracts. In this study Carum copticum (seeds) was extracted first in methanol and then subsequently extracted in different organic solvents. MIC of plant extracts, ciprofloxacin and thymol was determined by broth micro-dilution method using TTC. Synergism between plant extracts and ciprofloxacin was assayed by the checkerboard method. Chemical constituents of active extracts were analyzed by GC-MS. Methanolic, hexane and ether extract of Carum copticum exhibited significant antibacterial activity with MIC values ranged from 0.25 mg/ml to 2.0 mg/ml. Synergy analysis between Carum copticum extracts and ciprofloxacin combinations revealed FIC index in the range of 0.093-0.25. About 81% ciprofloxacin resistant ESßL producing enteric bacteria were re-sensitized in the presence of 15.6-250 µg/ml of methanolic extract of Carum copticum. Moreover, ciprofloxacin showed 8 to 64 folds reduction in MIC in presence of 250 and 500 µg/ml of hexane extract. Whereas, 4-32 folds reduction in MIC of ciprofloxacin was achieved in the presence of 31.25 and 62.5 µg/ml of ether extract, indicating synergistic enhancement of drug activity. The chemical analysis of hexane and ether extracts by GC-MS revealed the common occurrence of one or more phenolic hydroxyl at different locations on benzene ring. This study demonstrated the potential use of herbal extract of Carum copticum in combination therapy against ESßL producing bacteria.

Bioactive extracts of Carum copticum and thymol inhibit biofilm development by multidrug-resistant extended spectrum ß-lactamase producing enteric bacteria.

The emergence and spread of multidrug-resistant (MDR) pathogenic bacteria is a clinical problem that requires novel anti-infective agents. Targeting pathogenic biofilms is considered a promising strategy to control bacterial infections. In this study, bioactive extracts of Carum copticum were investigated for their anti-biofilm efficacy against extended spectrum ß-lactamase (ESßL) producing MDR enteric bacteria. Thymol was also tested for its anti-biofilm properties, as gas chromatography-mass spectrometry revealed a high content (65.8%) of this phytochemical in the C. copticum methanolic extract. Biofilm inhibition was assessed in microtitre plates and further validated by light, electron and confocal laser microscopy. Sub-inhibitory concentrations of bioactive extracts of C. copticum and thymol significantly prevented biofilm development, ranging from 78.6 to 83.9% reductions. Microscopic analysis revealed that biofilms made by ESßL producing MDR enteric bacteria had a weakened structure, scattered microcolonies, and reduced cell density and thickness after exposure to the bioactive extracts and thymol.

Setting research priorities for maternal, newborn, child health and nutrition in India by engaging experts from 256 indigenous institutions contributing over 4000 research ideas: a CHNRI exercise by ICMR and INCLEN.

BACKGROUND: Health research in low- and middle- income countries (LMICs) is often driven by donor priorities rather than by the needs of the countries where the research takes place. This lack of alignment of donor's priorities with local research need may be one of the reasons why countries fail to achieve set goals for population health and nutrition. India has a high burden of morbidity and mortality in women, children and infants. In order to look forward toward the Sustainable Development Goals, the Indian Council of Medical Research (ICMR) and the INCLEN Trust International (INCLEN) employed the Child Health and Nutrition Research Initiative's (CHNRI) research priority setting method for maternal, neonatal, child health and nutrition with the timeline of 2016-2025. The exercise was the largest to-date use of the CHNRI methodology, both in terms of participants and ideas generated and also expanded on the methodology. METHODS: CHNRI is a crowdsourcing-based exercise that involves using the collective intelligence of a group of stakeholders, usually researchers, to generate and score research options against a set of criteria. This paper reports on a large umbrella CHNRI that was divided into four theme-specific CHNRIs (maternal, newborn, child health and nutrition). A National Steering Group oversaw the exercise and four theme-specific Research Sub-Committees technically supported finalizing the scoring criteria and refinement of research ideas for the respective thematic areas. The exercise engaged participants from 256 institutions across India - 4003 research ideas were generated from 498 experts which were consolidated into 373 research options (maternal health: 122; newborn health: 56; child health: 101; nutrition: 94); 893 experts scored these against five criteria (answerability, relevance, equity, innovation and out-of-box thinking, investment on research). Relative weights to the criteria were assigned by 79 members from the Larger Reference Group. Given India's diversity, priorities were identified at national and three regional levels: (i) the Empowered Action Group (EAG) and North-Eastern States; (ii) States and Union territories in Northern India (including West Bengal); and (iii) States and Union territories in Southern and Western parts of India. CONCLUSIONS: The exercise leveraged the inherent flexibility of the CHNRI method in multiple ways. It expanded on the CHNRI methodology enabling analyses for identification of research priorities at national and regional levels. However, prioritization of research options are only valuable if they are put to use, and we hope that donors will take advantage of this prioritized list of research options.

Multidrug resistance and transferability of blaCTX-M among extended-spectrum ß-lactamase-producing enteric bacteria in biofilm.

This study aimed to investigate the occurrence of biofilm-forming extended-spectrum ß-lactamase (ESBL)-producing enteric bacteria in hospital wastewater and to evaluate their antibiotic resistance behaviour and transferability of the plasmid-encoded blaCTX-M gene in biofilm. ESBL production was confirmed using the combined disc test and Etest. Amplification of blaCTX-M was performed by PCR. Antibiotic susceptibility was evaluated using the disc diffusion assay and broth dilution method. Transfer of blaCTX-M in planktonic and biofilm state was performed by broth mating and filter mating experiments, respectively. Among 110 enteric bacteria, 24 (21.8%) isolates belonging to Escherichia coli, Klebsiella pneumoniae and Enterobacter cloacae were found to produce ESBL and formed varying levels of biofilm in vitro. Presence of blaCTX-M was detected in 18 (75%) ESBL-producing isolates. A many fold increase in resistance to antibiotics was observed in biofilm. Among ESBL-producers, seven isolates could transfer the blaCTX-M gene by conjugation, with transfer frequencies ranging from 2.22×10(-4) to 7.14×10(-2) transconjugants/recipient cell in the planktonic state and from 3.04×10(-3) to 9.15×10(-1) in biofilm. The transfer frequency of blaCTX-M was significantly higher in biofilm compared with the planktonic state, and co-transfer of ciprofloxacin resistance was also detected in five isolates. This study demonstrates that biofilm-forming ESBL-producing enteric bacteria with a greater transfer frequency of resistance genes will lead to frequent dissemination of ß-lactam and fluoroquinolone resistance genes in environmental settings. The emergence and spread of such multidrug resistance is a serious threat to animal and public health.

Emergence of ciprofloxacin-resistant extended-spectrum ß-lactamase-producing enteric bacteria in hospital wastewater and clinical sources.

This study aimed to evaluate the incidence of ciprofloxacin-resistant extended-spectrum ß-lactamase (ESBL)-producing enteric bacteria in hospital wastewater and clinical sources. Enteric bacteria, mainly Escherichia coli, were isolated from clinical sources (urinary tract and gastrointestinal tract infections; 80 isolates) and hospital wastewater (103 isolates). The antibiotic resistance profile and ESBL production of the isolates were investigated by disc diffusion assay and combined disc diffusion test, respectively. Plasmid profiling was performed by agarose gel electrophoresis, and elimination of resistance markers was performed by a plasmid curing experiment. Antibiotic susceptibility testing revealed a high incidence of ß-lactam resistance, being highest to ampicillin (88.0%) followed by amoxicillin, ceftriaxone, cefpodoxime, cefotaxime, aztreonam, cefepime and ceftazidime. Among the non-ß-lactam antibiotics, the highest resistance was recorded to nalidixic acid (85.7%). Moreover, 50.8% of enteric bacteria showed resistance to ciprofloxacin. Among 183 total enteric bacteria, 150 (82.0%) exhibited multidrug resistance. ESBL production was detected in 78 isolates (42.6%). A significantly higher incidence of ciprofloxacin resistance was observed among ESBL-producing enteric bacteria both in clinical (P=0.0015) and environmental isolates (P=0.012), clearly demonstrating a close association between ESBL production and ciprofloxacin resistance. Plasmid profiling of selected ESBL-positive strains indicated the presence of one or more plasmids of varying sizes. Plasmid curing resulted in loss of ciprofloxacin and cefotaxime resistance markers simultaneously from selected ESBL-positive isolates, indicating the close relationship of these markers. This study revealed a common occurrence of ciprofloxacin-resistant ESBL-producing enteric bacteria both in hospital wastewater and clinical sources, indicating a potential public health threat.

Alpha 1 antitrypsin deficiency in children with chronic liver disease in North India.

OBJECTIVE: We attempted to determine the role of alpha-1-antitrypsin (AAT) deficient variants as an etiologic factor for chronic liver disease in North Indian children. DESIGN: This study investigated 1700 children (682 retrospectively and 1018 prospectively) (840 CLD, 410 neonatal cholestasis and 450 without liver disease) for AAT deficiency. SETTING: Tertiary referral center, All India Institute of Medical Sciences, New Delhi. PATIENTS: Of 1250 liver disease patients, 98 (7.8%) were suspected to be AAT deficient on the basis of screening tests (low serum AAT levels and/or absent/faint alpha-1-globulin band on serum agarose electrophoresis and/or diastase resistant PAS positive granules on liver biopsy). MAIN OUTCOME MEASURES: AAT deficient Z or S allele in suspected patients. RESULTS: Z or S allele was not observed on phenotyping (1700 subjects), or with PCR-RFLP, SSCP and sequencing done in 50 of 98 suspected AAT deficient patients. A novel mutation G-to-A at position 333 in exon V was found in two siblings having positive immunohistochemistry for AAT on liver biopsy, both of whom had significant liver disease with portal hypertension. CONCLUSION: In conclusion, AAT deficiency as an etiologic factor for chronic liver disease in childhood appeared to be uncommon in North India.