Bacteriocin Activity of Leuconostoc Mesenteroides Pbac1 Bacteria on Several Media

Bacteriocin is a proteinaceous compound that has bactericidal action against microorganisms. Bacteriocins from lactic acid bacteria are very potential as natural food biopreservatives. The aim of the research was to know the infl uence of the growth medium; MRS, CMG, LTB and CM on antimicrobial activity of Leuconostoc mesenteroides Pbac1. Growth inhibition zone determination has been carried out by antagonism assay, as well as diffusion method using Lactobacillus plantarum, Leuconostoc mesenteroides, Staphylococcus aureus, Lactobacillus pentosus and Lactobacillus acidilactici as indicator strains. The results showed that L. plantarum and L. acidilactici were the sensitive indicators. The best growth medium for antagonism assay of the two sensitive indicator bacteria was MRS, which showed inhibition zone diameter of 1.28 cm and 1.23 cm, respectively. The most active supernatant was produced by L. mesenteroides Pbac1 grown on MRS, which inhibited the growth of L. plantarum and L. acidilactici with respective zone diameter of 1.28 cm and 1.19 cm. Bacteriocin titre activity against sensitive indicator bacteria was 100 AU/ml. Based on the result, MRS was further utilized to study the effect of the different carbon sources i.e glucose, maltose and mannose on bacteriocin activity of L. mesenteroides Pbac1. The results showed that glucose was the best carbon source as indicated by the widest diameter of inhibition zone, i.e 1.23 cm, against both indicator strains. Bacteriocin titre activity of the latter study was 100 AU/ml

A Novel Integron in the Genome of Escherichia Coli Isolated From Indonesian Monitor Lizard (Varanus Spp).

The genotype of antibiotic resistance in natural isolates of Escherichia coli was determined through integron detection and characterization of the associated antibiotic resistance. E. coli SG2 isolated from Varanus salvator of Java demonstrated resistance to spectinomycin (50ng/ml) and streptomycin (SOng/ml). Integron detection indicated that eight isolates out of nine E. coli isolates possessed a conserved segment of the integron. Amplification of the inserted cassette of the integron in this SG2 isolate yielded a 1-kb DNA fragment. Sequence analyses indicated that this fragment was homologous with aad gene, which confirmed the resistance to spectinomycin/streptomycin. This is the first report on the presence of integron in the E. coli isolated from the environment

Heterologous Expression of a Chitinase Gene From Aeromonas Caviaein Pseudomonas Fluorescens

A transcriptional fusion for an Aeromonas caviae chitinase gene was constructed under the control of a constitutive promoter of the kanaraycin resistance gene (PKmR). The construct was inserted into a medium copy number broad host range plasmid vector to yield recombinant plasmid pAM340, which harbored transcriptional fusion PKmR- chi. Another transcriptional fusion, Ptac-chi, in a recombinant plasmid pAM630, was conducted as comparison. Triparental mating of E. coli carrying the recombinant plasmids with Pseudomotws fluorescens 5100, a phyllosphere bacterium, was performed. Pseudomonas fluorescens 5100 exconjugants were examined for constitutive expression of chitinase employing a spectrophotometric assay; they showed stronger chitin degradation activity than Escherichia coli transformants. Using a fungal antagonism plate assay, this chitinolytic P. fluorescens, however, could not inhibit selected phytopathogenic fungi

Direct PCR: Alternative Diagnostic Method for Diagnosis of Diphtheria Rapidly, Easily and Cost Effective

Some diseases require immediate and appropriate treatment to decrease the fatality risk patients incident, for example diphtheria. Time to help patients is very crucial since delay of therapy may increase the mortality cases up to 20 times. In other hands, conventional diagnostic methods (the gold standard) for diagnosis of diphtheria is time consuming and laborious. Therefore, an alternative diagnostic method which is rapid, easy and inexpensive is needed. In this case, direct PCR has been proved to reduce time and cost in laboratory examination. This study aimed to develop direct PCR as alternative diagnostic method for diagnosis of diphtheria rapidly, easily, and inexpensive. Fifteen samples include 10 isolates of Corynebacterium diphtheriae (toxigenic) and 3 isolates of Corynebacterium non- diphtheriae (nontoxigenic) and 2 clinical specimens (throat swab) was examined by performing direct PCR method and a standard PCR method was used for optimizing the protocols. Result showed that direct PCR can be used to amplify target genes correctly as well as standard PCR. All of C. diphtheriae samples showed bands at 168 bp (dtxR gene marker) and 551 bp (tox gene marker) while no band appeared in others. Direct PCR detected at least 71 CFU/uL of bacterial cells in samples. We concluded that direct PCR can be used for alternative diagnostic method for diagnosis of diphtheria which is rapid, easy and cost effective

Metode Cepat Ekstraksi Dna Corynebacterium Diphtheriae Untuk Pemeriksaan Pcr

Diagnosis of diphtheria caused byCorynebacterium diphtheriaeshould be done immediately since delay of therapy may cause 20-fold increase rate of death. One method of rapid diagnostic to identify diphtheria is by using polymerase chain reaction (PCR). The fundamental issue of this method depends on the DNA, either its quality or quantity. The simple DNA extraction method, which is using mechanical/physical principles with a little of chemical reagents (such as boiling method and the use of sodium hydroxide (NAOH)), will have some benefits, such as easy to be performed, low cost, fast, and environmentally friendly. This study aimed to evaluate effectivity and efficiency of boiling method with NaOH to extract DNA of C. diphtheriae compared to the use of a commercial diagnostic kit for PCR assay. We used C. diphtheriae toxygenic(NCTC 10648) isolates, which are grown in blood agar plates. We then prepared the suspensions of cell/colony in aquadest with several dilutions. Each dilution was extracted using boiling method, NaOH and controlled with the use of a commercial diagnostic kit (QiAmp DNA Minikit). The results were evaluated quantitatively with spectrophotometer and qualitatively with gel electrophoresis. The results showed that the extracted DNA from boiling method with NaOH has an adequate quality and quantity for PCR assay (up to 9 CFU/uL cell/reaction). Therefore, it can be summarized that boiling method with NaOH is effective and efficient to be applied in PCR assay forC. diphtheriae.Key words: boiling extraction method, NaOH, C.diphtheriae, PCRAbstrakKematian kasus difteri yang disebabkan oleh Corynebacterium diphtheriaedapat meningkat 20 kali lipat karena keterlambatan pengobatan sehingga penegakan diagnosis harus dilakukan sesegera mungkin. Salah satu metode diagnostik yang cukup cepat untuk mendeteksi penyakit difteri adalah pemeriksaan polymerase chain reaction(PCR). Keberhasilan pemeriksaan PCR dipengaruhi oleh kualitas dan kuantitas DNA. Metode ekstraksi DNA sederhana menggunakan prinsip mekanik/fisika dengan reagen kimia minimalis, seperti metode pemanasan (boiling)dan penggunaan (NAOH) memiliki beberapa kelebihan, diantaranya mudah dilakukan, biaya murah, waktu yang dibutuhkan singkat dan lebih ramah lingkungan. Penelitian ini bertujuan untuk menilai efektifitas dan efisiensi metode boilingdan NAOH untuk ekstraksi DNA C.diphtheriae dibandingkan dengan penggunaan kit komersial dalam pemeriksaan PCR. Sampel berupa isolat C.diphtheriae toksigenik (NCTC 10648), ditumbuhkan dalam blood agar, kemudian dibuat suspensi sel/koloni dalam aquadest dengan beberapa kali pengenceran. Masing-masing pengenceran diekstraksi dengan metode boilingdan NaOH yang terkontrol dengan kit komersial (QiAmp DNA Minikit). Hasil ekstraksi dinilai secara kuantitatif dengan spektrofotometer dan secara kualitatif dengan gel elektroforesis. Hasil penelitian menunjukkan bahwa DNA hasil ekstraksi dengan metode boilingdan NaOH mempunyai kualitas dan kuantitas yang cukup untuk pemeriksaan PCR hingga 9 CFU/uL sel bakteri/reaksi sehingga dapat disimpulkan bahwa metode boiling dan NaOH cukup efektif dan efisien untuk diaplikasikan pada metode PCR untuk C.diphtheriae.Kata Kunci: boiling, NaOH, C.diphtheriae, PC

Seleksi Galur-Galur Leuconostoc Yang Mempunyai Aktivitas Bakteriostatin Terhadap Berbagai Bakteri Indikator

Lactid Acid Bacteria (LAB) are known to produce bacteriocins which have antimicrobi- al activity, and possessed to be developed as antibiotic complement. This study aimed to characterize bacteriocins activity from Leuconostoc strains isolated previously from local sources, and to optimize pH and incubation temperature as well. A well diffusion agar assay for zone inhibition method and bacteriocin potency assay performing minimum in- hibition concentration (MIC) have been done. Bacterial indicators used in this study are Leu. mesenteroides TISTR 120, and JCM 6124, Staphylococcus aureus FNCC 0047, Lis- teria monocytogenes FNCC 0156, Escherichia coli FNCC 0183, Pseudomonas aeruginosa FNCC 0063, Salmonella typhi FNCC 0165 and Bacillus subtilis FNCC 0061. Catalase, Trypsin and Protease K were also used for confirmation test. Results revealed that both Leu. mesenteroides MBF2-5 and MBF7-17 possessed bacteriocin activity although against Leu. mesenteroides TISTR 120 and JCM 6124 indicators strains. The optimum pH for bacteriocin potency assay for both Leuconostoc strains MBF2-5 and MBF7-17 was pH 6, whereas the optimum incubation temperature was 32 oc with MIC value of 90% and 80%, respectively