A.M. Yazid^1*, M. Shuhaimi¹, A.M. Ali², M.H. Ghazali², J. Normah¹, A.B. Fatimah³,
N.A. Nur Atiqah4 and A. Reezal3

¹Department of Food Technology, ²Department of Biotechnology, ³Department of food Science, Faculty of Food Science and
Biotechnology, Universiti Putra Malaysia, 43400 Serdang, Selangor,
⁴Department of Paediatrics, Kuala Lumpur Hospital, Jalan Pahang, 50586 Kuala Lumpur, Malaysia

(Received 10 April 1999 / Accepted 29 July 1999)

Abstract.
The ability of eighteen strains of genus Bifidobacterium spp. to survive in a simulated gastric pH and tolerance to bile acids exposure was investigated. Four strains namely, B. bifidum (ATCC 35914), B. adolescentis (ATCC 11146), B. breve (ATCC 15698) and B. infantis (ATCC 27920) showed excellent growth following 90 min exposure to extreme acidic condition (pH 2.5). Strains B. breve (ATCC 15701) and B. adolescentis (ATCC 15706) showed good acid tolerance and another 2 strains (B. adolescentis ATCC 15705 and B. longum ATCC 15707) possessed moderate ability to tolerate exposure to low pH. The remaining 10 strains did not survive the extreme acid condition. The ability to grow in the presence of bile acids was variable among strains. Four strains (B. longum ATCC 15707 and ATCC 15708, B. adolescentis ATCC 11146 and B. asteroides ATCC 25910) showed good tolerance and another 5 strains (B. breve ATCC 15700 and 15701, B. asteroides ATCC 25909, B. bifidum ATCC 35914 and B. infantis ATCC 27920) showed moderate tolerance and the rest were sensitive to bile acids. No correlation was observed between the ability to survive extreme acidic condition and the ability to grow in the presence of bile. The strains that possess excellent acid tolerance were not necessarily tolerant to bile, and vice versa.

Keywords: Bifidobacteria, tolerance, pH, bile acids

A. R. Omar^1* and K. A. Schat

^*1Department of Veterinary Pathology and Microbiology, Faculty of Veterinary Medicine and Animal Science, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia, Department of Microbiology and Immunology,
College of Veterinary Medicine, Cornell University, Ithaca, NY 14850, USA.

(Received 8 December 1997 / Accepted 10 January 1998)

Abstract.
Marek's disease virus (MDV) genes with homology to herpes simplex virus genes encoding for immediate early, early and late proteins and genes unique to MDV have been cloned and sequenced. Similarly, MDV genes that have significant homology to known oncogenes have also been characterized. Chickens vaccinated against this disease induce neutralizing antibody response and probably cytotoxic T lymphocyte activity that confer protection against subsequent challenge.

Keywords: Molecular Biology, Immunology, Marek's Disease Virus

Aris Tri Wahyudi, Antonius Suwanto*, Tedja Imas and Aris Tjahjoleksono

Department of Biology, Faculty of Mathematics and Sciences, Bogor Agricultural University, Jl. Raya Pajajaran, Bogor 16144, Indonesia.

(Received 9 August 1997 / Accepted 18 November 1997)

Abstract.
The genes, inaZ from Pseudomonas syringae, xylE from P. putida and the kanamycin resistance gene (Km^R) of mini-Tn5Km1, were studied for their capabilities and reliabilities as molecular markers in three acid tolerant Bradyrliizobium japonicum strains. Conjugation was used to transfer each of these genes to B. japonicum strains. Mating was conducted on membrane filter (0.45 gm, Millipore) using modified Luria Agar. The results showed that all of the genes were able to be transferred to acid tolerant B. japonicum strains by conjugation. All of these bacteria were able to express the inaZ and Km^R genes. However, none of the exconjugants was able to express the xylE gene. Acid tolerant B. japonicum strains that carried each of these genes were able to form root nodules in either siratro or soybean plant. The KmR gene had high stability as tested after nodulation. experiments. This gene was subsequently used as a genetic marker in a competition study of acid tolerant B. japonicum strains for nodule occupancy in soybean plant. This study was conducted using acid tolerant soybean cultivar (B 09) grown in Leonard jars using nitrogen free nutrient solution (pH 4.5 + Al 50 µM). Mixtures of acid tolerant B. japonicum strains and USDA 110 were inoculated in 1:1 ratio and nodules were harvested 30 days after planting. The results indicated that one exconjugant strain (11.71Km) dominated root nodules. Thus this strain has the potential to be developed as soybean inoculant in acid soils.

Keywords: Acid Aluminium Tolerance, Bradyrhizobium japonicum Marker Genes, Competition

Nooraini Abdulrashid* and Brian S. Hartley

*Faculty of Science, Universiti Teknologi Malaysia, Locked Bag 791, 80990 Johor Bahru, Malaysia, Centre for Biotechnology, Imperial College of Science, Technology and Medicine, London, SW7 2AZ, England

(Received 12 December 1997 / Accepted 16 February 1998)

Abstract.
The screening for the Aspergilus niger glucoamylase cDNA has led to the discovery that a full length cDNA in a pUC vector is lethal to Escherichia coli. Initial screening has indicated that full length glucoamylase cDNAs were initially present in the cDNA libraries but deletion mutants arose during the purification of the clones. The deletion mutants contained identical incomplete cDNAs lacking 35 nucleotides downstream of the initiation codon. The missing portion encodes part of an 18 amino acid signal peptide responsible for the translocation of the A. niger glucoamylase extracellularly. Therefore, to reconstruct a full length cDNA, synthetic oligonucleotides which represent the missing portion of the gene were ligated with the incomplete cDNA. However, several attempts to reconstruct a full length cDNA failed when the cDNA was in pUC18, a high copy number expression vector which utilizes the lac promoter. Fusion of a double stranded oligonucleotide representing the missing portion of the cDNA with a truncated cDNA led to the isolation of one deletion mutant which completely lost the entire glucoamylase cDNA except for the signal sequence. Incomplete cDNA lacking part of the signal sequence was then subcloned into a promoterless vector pSP72 and ligated with synthetic oligonucleotides representing the missing portion. This finally yielded full length 2.0 kb glucoamylase cDNA. In addition, it was found that subcloning of the full length glucoamylase cDNA into pT7T3 18U, another high copy number vector equipped with the lac promoter, also resulted in deletion of part of the cDNA to 1.5 kb.

Keywords: DNA Instability, Toxicity