Advanced Journal of Microbiology Research

Advanced Journal of Microbiology Research Vol. 2016

Available online at http://internationalscholarsjournals.org/journal/ajmr

© 2015 International Scholars Journals

Full Length Research Paper

Fingerprinting of repetitive DNA sequences in the genus Anabaena using PCR- based techniques

Anbalagan Ezhilarasi* and Narayanaswamy Anand

Centre for Advanced Studies in Botany, University of Madras, Guindy Campus, Chennai–600025, India.

Accepted 23 March, 2015

Abstract

In this study, ten species of Anabaena were used to test the congruence between the traditional morphological classification system and the present molecular classification system. The electrophoretic patterns for 10 cyanobacterial strains belonging to the genera (Anabaena) were used for molecular analysis using the repeated PCR technique. Genetic diversity was assessed using the banding patterns of repetitive DNA sequences including the HIP1 sequences, enterobacterial repetitive intergenic consensus [ERIC] sequences and REP sequences that are present in the cyanobacterial genome. PCR with different sets of repetitive oligonucleotides as primers were used to evaluate the various repetitive DNA sequences. The dendrograms based on the HIP1, ERIC and REP sequences did not show a clear correlation with morphological dendrogram based on traditional classification.

Key words: Anabaena, cyanobacteria, highly iterated palindromic sequences, enterobacterial repetitive intergenic consensus sequences, repetitive extragenic palindromic sequences, polymerase chain reaction.

Advanced Journal of Microbiology Research Vol. 2016

Available online at http://internationalscholarsjournals.org/journal/ajmr

© 2015 International Scholars Journals

Full Length Research Paper

The effect of cultivation conditions on the mycelial growth of a dark-septate endophytic isolate

Ya-li Lv, Li-hua Sun, Fu-sheng Zhang, Yue Zhao and Shun-xing Guo*

Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, P. R. China.

Accepted 9 March, 2016

Abstract

The EF-37 isolate, one of DSE fungi, is beneficial to the growth and development of its host plant, Saussurea involucrata Kar. et Kir. The cultivation requirements including basic culture medium, temperature, light, pH, carbon source and nitrogen compounds were studied for their effects on mycelial growth of a dark-septate endophytic (DSE) fungus EF-37 by using one-factor-at-a- time method. Potato dextrose agar (PDA) was the best medium for the growth of endophyte EF-37. Our studies showed that 20°C, 24 h dark cultivation and pH 7 significantly influenced the growth of endophyte EF-37 on PDA medium. Moreover, glucose and calcium nitrate were found to be the best nutrients for EF- 37 growth. Under the optimal cultivation conditions, DSE fungus EF-37 isolate could grow actively. This is the first study about the effect of cultivation conditions on the growth of this strain, which provides the preparatory knowledge for the biological characteristics of DSE fungus EF-37.

Key words: Cultural conditions, dark-septate endophytic (DSE) fungus EF-37, mycelial growth, optimization, Saussurea involucrata Kar. et Kir.

Advanced Journal of Microbiology Research Vol. 2016

Available online at http://internationalscholarsjournals.org/journal/ajmr

© 2016 International Scholars Journals

Full Length Research Paper

The identification of the first isolate of influenza B virus using a duplex RT- PCR DNA sequencing in Saudi Arabia (B/Riyadh/01/2007)

Fahad N. Almajhdi

Department of Botany and Microbiology, Center of Excellence in Biotechnology Research, College of Sciences, King Saudi University, Saudi Arabia. E-mail: [email protected]. Tel: +966-4677778. Fax: +966-4675833.

Accepted 24 February, 2016

Abstract

Although Influenza virus infections threaten thousands of lives each year worldwide and bear the risk of been an epidemic, little is known about the circulating strains in Saudi Arabia. The availability of a rapid and accurate diagnostic approach is essential for effective treatment and disease control. In the current study, we optimized a duplex RT-PCR assay for the simultaneous detection of influenza A and B viruses in clinical samples. The developed assay was utilized for testing 100 nasopharyngeal aspirates collected from young children hospitalized with acute respiratory tract infection in Riyadh. Influenza B not A viruses, were identified as possible causes of the disease syndrome. Recovery of influenza B virus from clinical samples was achieved by serial passage in chicken embryos and the virus isolate was designated as Influenza B/Riyadh/01/2007. The identity of B/Riyadh/01/2007 was confirmed by sequencing the RT-PCR product. The sequenced data was submitted to the Gene Bank under the accession number GU135839. An almost complete homology was recognized with all Influenza B virus strains available on Gene Bank. Although several studies suggested the presence of influenza B virus in respiratory tract infection of Saudi children, this is the first report that describes the isolation and partial identification of the circulating virus. Further studies that fully characterize the isolated strain are ongoing.

Key words: Influenza B virus, isolation, RT-PCR, DNA sequencing, Saudi Arabia.

Advanced Journal of Microbiology Research Vol. 2016

Available online at http://internationalscholarsjournals.org/journal/ajmr

© 2016 International Scholars Journals

Full Length Research Paper

Development and evaluation of colloidal gold immunochromatographic strip for detection of Escherichia coli O157

Xihong Zhao¹, Xiaowei He¹*, Wenmei Li ^1,3, Yao Liu², Liansheng Yang¹ and Jihua Wang³

¹College of Light Industry and Food Sciences, South China University of Technology, Guangzhou 510640, China.

²Zhongshan Supervision Testing Institute of Quality and Metrology, Zhongshan 528403, China.

³Guangzhou Wondfo Biotech CO., LTD, 510530, Guangzhou, China.

Accepted 24 February, 2016

Abstract

Escherichia coli O157:H7 is a serious and common human pathogen that can cause diarrhoea, haemorrhagic colitis, and haemolytic uraemic syndrome (HUS). In this study, the synthesis and identification of colloidal gold particles and antibody-colloidal gold conjugates probe specific to E. coli O157:H7 were performed, and the preparation of colloidal gold immunochromatographic strip based the biotin-streptavidin system was developed for detection of E. coli O157:H7.Monodispersional nanogold colloid was synthesized and preparation of nanogold-labeled polyclonal antibody probe to E. coli O157:H7 by citrate method. Combination of antibody with nanogold particles was also characterized by UV-visible (UV-vis) light absorption spectra and transmission electron microscopy (TEM). Furthermore, nanogold-labeled probe was used to develop an immunochromatographic (IC) strip for E. coli O157:H7 analysis. With this method, analysis could be completed in less than 10 min. Examination of the 65 known strains (36 E. coli O157 strains and 29 serotypes other than E. coli) showed 98.5% specifity and 100% sensitivity, only yield a false-positive reaction with Salmonella choleraesuis . The sensitivity of the IC strip was tested using 10-fold dilution E. coli O157 in foods, could be detected at a minimum of 2.3 × 10³ CFU/ml without enrichment and 2.3 CFU/ml after enrichment. Application of IC strip test were performed on 265 water samples, 340 beef samples, 208 milk samples and 120 cake samples after enrichment, the specificity of the strip was 99.2, 97.9, 94.6 and 94.9%, respectively. The sensitivity of the strip was 100% agreement with tradition culture method. The established method is very useful for monitoring E. coli O157 containment in food samples.

Key words: Escherichia coli O157, colloidal gold immunochromatographic assay, rapid test.

Advanced Journal of Microbiology Research Vol. 2016

Available online at http://internationalscholarsjournals.org/journal/ajmr

© 2015 International Scholars Journals

Full Length Research Paper

Investigation of different infectious pathways used by Streptococcus suis type 2 and other bacteria in mice

Xingjian Cao, Zhaoming cao, Guohua Tao, Wei Xie, Xiang Chen and Shuo Wang

Center of Clinical Laboratory Medicine, the Second Affiliated Hospital of Nantong University, Nantong 226001, China.

Accepted 19 March, 2015

Abstract

The purpose of this study was to investigate the infectious pathways used by Streptococcus suis type 2 and enterohemorrhagic Escherchia coli O26 (EHEC O26) in mice. Six-week mice were intraperitoneally injected with mitomycin (0.2 mg/mouse) to decrease the protective immunity. The mice were then challenged with S. suis type 2 and EHEC O26 strains through wounded skin, stomach lavaging and intraperitoneal injection. Enterococcus faecalis ATCC 29212 and E. coli ATCC25922 were used as negative controls. Clinical and microbiological examinations were performed for all the infected mice routinely. The mortality rate of the mice infected with S. suis type 2 strain through wounded skin was 60% within 2 - 5 days. The mortality rate of the mice infected with S. suis type 2 strain through stomach lavaging was 60% within 7 - 10 days. S. suis type 2 can be isolated from the puncturing fluid of heart in the dead mice. Mortality was not observed in mice infected by S. suis type 2 and E. faecalis ATCC 29212 strains through intraperitoneal injection. However, intraperitoneal injection with EHEC O26 and E. coli ATCC25922 caused death of mice within 10 h. We demonstrated an important pathway used by S. suis type 2 strain in mouse infection model. Intraperitoneal injection with gram negative bacteria (E. coli ATCC25922 and EHEC O26) caused non-specific death in mice. These results provided fundamental bases for investigating the pathogenesis of S. suis type 2, designing vaccine and evaluating the efficiency of antimicrobial drugs.

Key words: Streptococcus suis type 2, enterohemorrhagic Escherchia coil O26, model, mouse.

Advanced Journal of Microbiology Research Vol. 2016

Available online at http://internationalscholarsjournals.org/journal/ajmr

© 2015 International Scholars Journals

Full Length Research Paper

ISSR for comparison of cross-inoculation potential of Colletotrichum capsici causing chilli anthracnose

Kanchalika Ratanacherdchai^1*, Hong-Kai Wang², Fu-Cheng Lin² and Kasem Soytong³

¹International College, King Mongkut’s Institute of Technology Ladkrabang, Ladkrabang, Bangkok 10520, Thailand.

²Biotechnology Institute, School of Agriculture and Biotechnology, Zhejiang University, Hangzhou 310029, P. R. China.

³Department of Plant Pest Management Technology, Faculty of Agricultural Technology, King Mongkut’s Institute of Technology Ladkrabang, Ladkrabang, Bangkok 10520, Thailand.

Accepted 9 November, 2015

Abstract

Thirty-four isolates of Colletotrichum spp. including 2 species, C. gloeosporioides and C. capsici, from anthracnose on Bell pepper, Long cayenne pepper and Bird’s eye chilli were isolated and their pathogenicity was proven via fruit inoculation. Pathogenicity tests divided pathogenic potential into low, medium and high virulence groups. It is clearly revealed that C. capsici from the three tested hosts expressed the highest virulent isolates. Cross-inoculation of three high virulent isolates of C. capsici in accordance with three chilli varieties showed that all isolates could produce anthracnose symptom in the same lesions. All tested isolates developed lesions after co-inoculation of all hosts. Inter simple sequence repeat (ISSR) analysis indicated that there are two distinct groups of C. gloeosporioides and C. capsici. Furthermore, genetic diversity was correlated with geographic distribution, while there was no clear relationship between genetic diversity and pathogenic variability. But it is clearly demonstrated that whereas C. gloeosporioides appears in the same geographic area as C. capsici, it causes lower disease incidence.

Key words: Anthracnose, chilli, Colletotrichum capsici, cross-inoculation.