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Objective The inhibitory effects of essential oils including fennel, juniper and kalonji from Foeniculum Vulgare , Juniperus Osteosperma and Nigella Sativa on multi drug resistant clinical isolates were investigated. All the oils have been evaluated for phytochemical constituents, antibacterial activity and TLC bioautography assay. Methods Preliminary phytochemical analysis was performed. The antibacterial potential of essential oils from fennel, juniper and kalonji fennel, juniper and kalonji was evaluated by agar well diffusion method against multi drug resistant clinical isolates. The antibacterial effect was investigated using the TLC-bioautographic method. Results Preliminary phytochemical analysis demonstrated the presence of most of the phytochemicals including saponins, cardiac glycosides, steroids, terpenoids, flavonoids and tannins. Antibacterial activity of essential oils was assessed on eight multi-drug resistant (MDR) clinical isolates from both Gram-positive and Gram-negative bacteria and two standard strains. All the oils tested showed significant to moderate antibacterial activity toward all tested strains except Acinetobacter sp and Staphylococcus aureus MRSA. The maximum zone of inhibition was found to be 25±0.12 mm for juniper oil followed by 21±0.085 mm for kalonji oil against Staphylococcus aureus 2. Thin layer chromatography and bioautography assay demonstrated well-defined growth inhibition zones against Staphylococcus aureus 2 and E. coli for juniper essential oil in correspondence with tannins observed at Rf values of 0.07 and 0.57. Conclusions Based on the present study, the essential oils from juniper and kalonji possess antibacterial activity against several multi drug resistant pathogenic bacteria and thus can be used as a base for the development of new potent drugs and phytomedicine.

Seventy isolates of Macrophomina phaseolina recovered from different host plants were assessed for DNA polymorphism using two molecular techniques: microsatellite primed polymerase chain reaction (MSP-PCR) under both touchdown (T) and non-touchdown (NT) PCR conditions and primers corresponding to disperse repetitive sequence-based polymerase chain reaction (rep-PCR). Fingerprints obtained by rep-PCR were compared with those of MSP-PCR. Even though these methods yielded intraspecific polymorphisms, yet different levels of discrimination could be obtained. A partial correlation was apparent between the molecular techniques used. Some of the genetic groups ⁄ genotypes were supported by both the molecular markers employed in the study, thus confirming their relationship. Thirty nine MSP (T), 55 MSP (NT) and 53 rep-PCR genotypes were identified with discrimination indices of 0.962, 0.993 and 0.99, respectively. Our results have shown that rep-PCR is a rapid, inexpensive technique that is highly reproducible and almost as discriminatory as MSP-PCR for genotyping M. phaseolina isolates and is highly suitable for understanding disease epidemiology at molecular level. Suggesting, thereby, that it is a robust technique employed for genotypical and phylogenetic studies for determining taxonomical diversity and phylogenetic structure of the economically important fungal pathogen of cluster bean. The data presented here will help researchers to design effective strategies for deployment of resistant germplasm in cluster bean (Cymopsis tetragonoloba) growing regions in the country and worldwide.

Phenotypic and genetic diversity of 59 Macrophomina phaseolina isolates collected from various host species growing in or near cluster bean (Cyamopsis tetragonoloba) fields in four states of north and north-west India were characterized using RAPD and PCR–RFLPs of the ITS region. These isolates, and 11 from various hosts from culture collections, were classified into three mycelial phenotypes: dense, feathery and restricted, based on variable growth patterns on nutrient agar containing 120 mm chlorate. Pathogenicity of isolates was evaluated by measuring the length of stem lesions 21 days post-inoculation on the susceptible cluster bean genotype FS 277. Isolates showed considerable variation in aggressiveness, with the isolates from cluster bean with dense chlorate phenotype producing relatively higher lesion lengths on cluster bean plants. The results of the RAPD assay clearly distinguished the isolates on the basis of chlorate phenotype and host origin. Isolates from a single host were generally similar to each other, but differed distinctly from those from other hosts. Chlorate-sensitive isolates were distinct from chlorate-resistant isolates within a given host. A high degree of polymorphism in restriction patterns of the ITS region, including part of 25S rDNA, has been reported for the first time in the charcoal rot fungus.

Ten different isolates of Macrophomina phaseolina from cluster bean (Cyamopsis tetragonoloba) selected on the basis of wide geographical distance and potassium chlorate sensitivity, showed remarkable variation in virulence on the susceptible cluster bean genotype, FS 277. Four different host inoculation techniques were employed for resistance screening of six elite cluster bean genotypes under artificial infection conditions using the most pathogenic isolate, MP-6D. The symptoms induced in the plant tissues from seedling to maturity following artificial inoculation clearly simulated lesions produced under natural conditions. Both greenhouse and field experiments demonstrated significant differences in all parameters-plant weight, plant death, root lesion length and stem lesion length-in all the genotypes tested from seedling stage to pod set. The host inoculation methods described in this study might be useful for screening and analysis of crop varieties' resistance to various other disease infes...

Xanthomonas axonopodis pv. cyamopsidis causes bacterial blight of cluster bean, an industrially important legume. A total of 76 isolates of X. axonopodis pv. cyamopsidis obtained from cluster bean fields of north and north-west India were identified at the pathovar level using virulence trials on the susceptible cluster bean genotype, PNB which confirmed their phytopathogenic specialization on cluster bean. Different polymerase chain reaction (PCR)-based techniques, such as, random amplified polymorphic DNA (RAPD)–PCR, repetitive (rep)-PCR and IS1112-PCR were successfully employed to elucidate the pathogen population structure and intrapathovar relationship among a set of 25 X. axonopodis pv. cyamopsidis isolates. There was a significant variation in their pathogenicity and genetic structure. The pathogenicity data were found to be in line with the molecular data to some extent. RAPD markers revealed a high level of genetic diversity across the isolates employed in comparison with the other molecular markers employed. A weak correlation was observed among the various molecular markers employed as each technique explores genetic variation differently. There was no correlation between DNA amplification product patterns and geographic sites of isolation, suggesting that this bacterium has spread largely through exchange of infected plant germplasm. This is the first report on the molecular characterization of X. axonopodis pv. cyamopsidis isolates from India.

The in vitro antibacterial activity of various solvents and water extracts of aloe vera, neem, bryophyllum, lemongrass, tulsi, oregano, rosemary and thyme was assessed on 10 multi-drug resistant clinical isolates from both Gram-positive and Gram-negative bacteria and two standard strains including Staphylococcus aureus ATCC 25923 and Escherichia coli ATCC 25922. The zone of inhibition as determined by agar well diffusion method varied with the plant extract, the solvent used for extraction, and the organism tested. Klebsiella pneumoniae 2, Escherichia coli 3 and Staphylococcus aureus 3 were resistant to the plant extracts tested. Moreover, water extracts did not restrain the growth of any tested bacteria. Ethanol and methanol extracts were found to be more potent being capable of exerting significant inhibitory activities against majority of the bacteria investigated. Staphylococcus aureus 1 was the most inhibited bacterial isolate with 24 extracts (60%) inhibiting its growth whereas Escherichia coli 2 exhibited strong resistance being inhibited by only 11 extracts (28%). The results obtained in the agar diffusion plates were in fair correlation with that obtained in the minimum inhibitory concentration tests. The minimum inhibitory concentration of tulsi, oregano, rosemary and aloe vera extracts was found in the range of 1.56-6.25 mg/ml for the multi-drug resistant Staphylococcus aureus isolates tested whereas higher values (6.25-25 mg/ml) were obtained against the multi-drug resistant isolates Klebsiella pneumoniae 1 and Escherichia coli 1 and 2. Qualitative phytochemical analysis demonstrated the presence of tannins and saponins in all plants tested. Thin layer chromatography and bioautography agar overlay assay of ethanol extracts of neem, tulsi and aloe vera indicated flavonoids and tannins as major active compounds against methicillin-resistant Staphylococcus aureus.