Plant growth promoting capability and genetic diversity of bacteria isolated from mud volcano and lime cave of Andaman and Nicobar Islands

Twenty four bacterial strains from four different regions of mud volcano and lime cave were isolated to estimate their diversity, plant growth promoting and biocontrol activities to use them as inoculant strains in the fields. An excellent antagonistic effect against four plant pathogens and plant growth promoting properties such as IAA production, HCN production, phosphate solubilization, siderophore production, starch hydrolysis and hydrolytic enzymes syntheses were identified in OM5 (Pantoea agglomerans) and EM9 (Exiguobacterium sp.) of 24 studied isolates. Seeds (Chili and tomato) inoculation with plant growth promoting strains resulted in increased percentage of seedling emergence, root length and plant weight. Results indicated that co-inoculation gave a more pronounced effects on seedling emergence, secondary root numbers, primary root length and stem length, while inoculation by alone isolate showed a lower effect. Our results suggest that the mixed inocula of OM5 and EM9 strains as biofertilizers could significantly increase the production of food crops in Andaman archipelago by means of sustainable and organic agricultural system.

Saccharomyces cerevisiae and non-Saccharomyces yeasts in grape varieties of the São Francisco Valley

The aims of this work was to characterise indigenous Saccharomyces cerevisiae strains in the naturally fermented juice of grape varieties Cabernet Sauvignon, Grenache, Tempranillo, Sauvignon Blanc and Verdejo used in the São Francisco River Valley, northeastern Brazil. In this study, 155 S. cerevisiae and 60 non-Saccharomyces yeasts were isolated and identified using physiological tests and sequencing of the D1/D2 domains of the large subunit of the rRNA gene. Among the non-Saccharomyces species, Rhodotorula mucilaginosa was the most common species, followed by Pichia kudriavzevii, Candida parapsilosis, Meyerozyma guilliermondii, Wickerhamomyces anomalus, Kloeckera apis, P. manshurica, C. orthopsilosis and C. zemplinina. The population counts of these yeasts ranged among 1.0 to 19 x 10(5) cfu/mL. A total of 155 isolates of S. cerevisiae were compared by mitochondrial DNA restriction analysis, and five molecular mitochondrial DNA restriction profiles were detected. Indigenous strains of S. cerevisiae isolated from grapes of the São Francisco Valley can be further tested as potential starters for wine production.

Identification of environmental mycobacteria isolated from Agra, north India by conventional & molecular approaches.

Background & objectives: Several environmental mycobacteria have been shown to be important human pathogens linked to immunomodulation especially in relation to effect on vaccination. Hence identification of mycobacteria to the species level is not only relevant to patient management but also to understand epidemiology of mycobacterial diseases and effect on vaccination. We undertook this study to assess the usefulness of various conventional and molecular methods in identification of environmental mycobacterial species from Agra, north India. Methods: One hundred nineteen isolates of environmental mycobacteria were grown from 291 (116 soil and 175 water) samples. These isolates were identified by standard biochemical tests, and a simple, rapid and cost-effective in-house developed gene amplification restriction analysis targeting 16S-23S rRNA spacer and flanking region and 16S rRNA sequencing. Results: Biochemical tests could clearly identify only 68.1 per cent (81/119) of isolates to species level. An in-house developed gene amplification - restriction analysis method could confirm the identity of 102 of 119 (85.7%) isolates and the remaining 17 isolates (14.3%) were confirmed by 16S rRNA sequencing also. These 119 environmental mycobacterial isolates, included several potentially pathogenic species such as M. fortuitum, M. chelonae, M. avium, M. marinum, M. manitobense, M. kansasii and others belonged to nonpathogenic species, M. terrae, M. smegmatis and M. flavescens. M. chelonae was isolated from water samples only whereas M. fortuitum was isolated from both water as well as soil samples. Interpretation & conclusion: The in-house developed gene amplification restriction analysis method though failed to accurately identify 14.3 per cent of isolates, facilitated rapid differentiation of most of environmental mycobacteria including potential pathogens from this area and thus would have diagnostic potential in cases with NTM infections. This combination strategy using PCR-RFLP and 16S rRNA sequencing may be useful for characterization of mycobacteria from similar environmental settings from other parts of world.

DNA methylation of TPEF gene in head and neck squamous cell carcinoma cell lines

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common malignancy worldwide. The molecular mechanisms involved in the development and progression of these carcinomas are not well known. Abnormalities of genomic methylation patterns have been attributed a role in carcinogenesis and local de novo methylation at tumor suppressor loci was held to be involved in silencing of tumor suppressor genes. Using Ms APPCR, we previously isolated a hypermethylated fragment corresponded to the 5'end of TPEF gene from primary liver and lung cancer cells. To confirm the inactivation of TPEF gene by hypermethylation in HNSCC, we investigated correlation between methylation pattern and expression of TPEF in 10 HNSCC cell lines. In methylation analysis such as combined-bisulfite restriction analysis(COBRA) and bisulfite sequencing, only RPMI 2650 showed none methylated pattern and another 9 cell lines showed dense methylation. The TPEF gene expression level analysis using RT-PCR showed that these 9 cell lines had not or significantly low expression levels of TPEF as compared with RPMI 2650. In addition, the increase of TPEF reexpression by 5-AzaC as demethylating agent in 9 cell lines also indicated that TPEF expression was regulated by hypermethylation. These results of this study demonstrate that epigenetic silencing of TPEF gene by aberrant methylation could play an important role in HNSCC carcinogenesis.

Identification of Acinetobacter Genospecies by Analysis of rRNA Spacer Regions / 대한임상병리학회지

BACKGROUND: Recently, the genus Acinetobacter have become increasingly important as nosocomial pathogens. Colonization and infection caused by multiresistant epidemic strains are common manifestations in intensive care units. Up to now 21 genomic species have been described, of which 7 have been named. However, there is no approach about distribution of Acinetobacter species in Korea. The objective of this study was to evaluate the performance of PCR analysis for the accurate identification and distribution of Acinetobacter species. METHODS: tRNA intergenic spacer length polymorphism(tRNA-ILP), 16S-23S rRNA spacer length polymorphism(16S-23S ILP) and restriction analysis of the 16S-23S rRNA intergenic spacer sequences(RA 16S-23S) were performed to identify the 7 type strains and 83 clinical isolates of Acinetobacter. RESULTS: 1. In the 7 Acinetobacter type strains, tRNA-ILP data could be easily analyzed by visual comparision. Eighty one of 83(97.6%) clinical isolates of Acinetobacter were identified as A. baumannii and the others were identified as A. haemolyticus and A. calcoaceticus. 2. The 16S-23S ILP patterns were not distinguished among the species, except A. haemolyticus, A, johnonii and A. lwoffii. The three species could be discriminated by a number of specific DNA fragments. 3. In RA 16S-23S, restriction endonuclease Alu I was able to not only digest the amplification products but also gave characteristic restriction patterns for the 7 type strains. CONCLUSION: The tRNA-ILP analysis and RA 16S-23S can be used as a tool for the rapid identification of Acinetobacter species with a high degree of specificity.

Molecular Epidemiology of Methicillin-resistant Staphylococcus aureus Outbreak by Plasmid Restriction Analysis / 대한임상미생물학회지

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) continues to be a major cause of nosocomial infection and a molecular typing is necessary for proper epidemiologic investigations of sources and moles of spread in an outbreak. An nosocomial outbreak of MRSA in a neonatal intensive care unit at Ewha Womans University Mokdong Hospital was suspected. To investigate the clonality of isolates and control the spread of nosocomial outbreak, we performed plasmid restriction analysis of MRSA isolates from patients and medical staffs. METHODS: We studied 7 MRSA strains (umbilicus 4, blood 1, urine 1 and pus 1) from patients in a neonatal intensive care unit and the MRSA strains from nares and hands surveillance cultures of 26 medical staffs (4 medical doctors and 22 nurses). All MRSA strains were tested for antimicrobial susceptibility and plasmic analysis after EcoRI restriction. We analyzed the plasmid patterns of MRSA isolated from patients and compared with those from medical staffs. RESULTS: Ten MRSA strains (from 7 nares and 3 hands) were isolated from surveillance cultures of 26 medical staffs. Seven out of 10 MRSA strains from medical staffs revealed identical pattern of antibiogram which was the same pattern in all 7 MRSA strains from seven patients. Plasmid restriction patterns were classified 6 groups from A to F showing 2-10 bands. Six out of 7 MRSA strains from the patients showed group A(A1 5, A31) and 5 out of 10 MRSA strains from the medical staffs showed group A(A1 1, A21, A32, A41) and remainders showed different plasmid restriction analysis patterns. CONCLUSIONS: These results suggest that plasmid restriction analysis is a rapid, inexpensive, and good discriminating molecular typing of MRSA outbreak and is useful for the epidemiologic investigation of MRSA outbreaks in the clinical laboratory.

Restriction enzyme analysis of the chloroplast DNA of Phaseolus vulgaris L. vr. Rio Negro / Análise de restrição do DNA cloroplástico de Phaseolus vulgaris vr. Rio Negro

The chloroplast DNA of Phaseolus vulgaris L. vr. Rio Negro was isola ted from chloroplasts obtained by descontiuous sucrose gradient centrifugation. The restriction analysis with the enzymes HindIII, EcoRI and BamHI and their combination, allowed to identified more than 20 fragments of 18 to 0.65kb. The size of Phaseolus vulgaris L. cp DNA was estimated in 140kb with the presence of a repeat sequence of about 22kb.

O DNA cloroplástico do cultivar Rio Negro (Phaseolus vulgaris L.) foi isolado a partir de cloroplastos obtidos por gradiente descontínuo de sacarose. A análise de restrição com as enzimas HindIII, EcoRI e BamHI e a combinação destas, permitiu a identificação de mais de 20 fragmentos na faixa de 18 a 0.65kb. O tamanho do cp DNA de Phaseolus vulgaris L. foi estimado em 140kb com a existência de sequências repetidas de aproximadamente 22kb.

Detection and Typing of Herpes Simplex Virus by Polymerase Chain Reaction / 대한피부과학회지

BACKGROUND: Herpes simplex virus(HSV) infections are very common viral illnesses in dermatology and they have shown a tendency to increase in prevalence and incidence, and they would seem to have become more prevalent recently. This has resulted in an increased need for the rapid diagnosis of these infections. It has also become important to recognize the types of HSV in the clinical setting, because the two types differ in their natural histories and prognoses. OBJECTIVE: The purpose of this study is to detect and type HSV DNA by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). We investigated the relationship between the clinical manifestations and type using PCR. METHODS: The study population consisted of 40 cases of HSV infections and 10 cases of varicella-zoster virus infections as negative controls. We used a pair of primers designated by Sakaoka et. al and performed restriction analysis after PCR. RESULTS: No specific amplification was observed using the varicella-zoster virus. A total of 40 patients were examined by PCR and 28 were positive. Of 17 patients with lesions which developed above the umbilicus, l0 were positive. Out of 8 patients with lesions below the umbilicus, 5 were positive. Virus genomes were detected in the 13 out of 15 patients with eczema herpeticum. CONCLUSION: The method used in this study is useful in differentiating HSV-1 from HSV-2 in fections. These data suggest that the PCR results were nearly consistent with the clinical manifestations.

Detection and Typing of Herpes Simplex Virus by Polymerase Chain Reaction / 대한피부과학회지

BACKGROUND: Herpes simplex virus(HSV) infections are very common viral illnesses in dermatology and they have shown a tendency to increase in prevalence and incidence, and they would seem to have become more prevalent recently. This has resulted in an increased need for the rapid diagnosis of these infections. It has also become important to recognize the types of HSV in the clinical setting, because the two types differ in their natural histories and prognoses. OBJECTIVE: The purpose of this study is to detect and type HSV DNA by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). We investigated the relationship between the clinical manifestations and type using PCR. METHODS: The study population consisted of 40 cases of HSV infections and 10 cases of varicella-zoster virus infections as negative controls. We used a pair of primers designated by Sakaoka et. al and performed restriction analysis after PCR. RESULTS: No specific amplification was observed using the varicella-zoster virus. A total of 40 patients were examined by PCR and 28 were positive. Of 17 patients with lesions which developed above the umbilicus, l0 were positive. Out of 8 patients with lesions below the umbilicus, 5 were positive. Virus genomes were detected in the 13 out of 15 patients with eczema herpeticum. CONCLUSION: The method used in this study is useful in differentiating HSV-1 from HSV-2 in fections. These data suggest that the PCR results were nearly consistent with the clinical manifestations.