Multiple Antibiotic Resistance and Heavy Metal Resistance Profile of Bacteria Isolated from Giant Freshwater Prawn (Macrobrachium rosenbergii) Hatchery

In this article, antibiogram and heavy metal resistance profile of bacteria isolated from giant freshwater prawn （Macrobrachium rosenbergii） hatchery in Malaysia are described. Although giant freshwater prawn was introduced into Malaysia since the 1980s, there was no database information on antibiogram and heavy metal resistance profile of bacteria from giant freshwater prawn （34. rosenbergii） hatchery in Malaysia. Therefore, this study was carried out to determine the effectiveness of antibiotic and heavy metal resistance profile to control bacterial diseases in M. rosenbergii hatchery. The results can provide valuable information for local M. rosenbergii post-larval producer. Antibiotic sensitivity test was carried out by disk-diffusion method against 15 types of antibiotics as follows： oxolinic acid （2 μg）, ampicillin （10μg）, erythromycin （15μg）, furazolidone （15 μg）, lincomycin （15μg）, amoxicillin （25 μg）, colistin sulphate （25 μg）, doxycycline （30μg）, florfenicol （30 μg）, flumequine （30 lag）, nalidixic acid （30μg）, tetracycline （30μg）, oleandomycin （15μg）, fosfomycin （50μg）, and spiramycin （100 μg）, whereas heavy metal resistance profile of the present bacterial isolates was determined by 2-fold agar dilution technique. In this study, 5 types of bacteria were successfully isolated; they were Aeromonas spp. （n = 77）, Escherichia coli （n = 73）, Edwardsiella spp. （n = 62）, Salmonella spp. （n = 75）, and Vibrio spp. （n = 43）. The result showed that furazolidone was the most effective antibiotic to control the bacteria isolated in this study, approximately 89.7% of the bacterial isolates were sensitive to this antibiotic. Multiple antibiotic resistance （MAR） index indicated that the hatchery water source and M. rosenbergii post-larval and sediment tanks were at high-risk exposure to the tested antibiotic. Furthermore, all the tested heavy metals （Cd2＋, Cr6＋, Hg2＋, and Cu2＋） failed to inhibit the growth of the bacterial isolates. Therefore, it indicated that the water source of the hatchery is contaminated with both antibiotic residues and heavy metal.

A Simple and Efficient Protocol for the Mass Propagation of <i>Vanilla planifolia</i>

The present study describes a direct shoot regeneration-based micropropagation procedure for Vanilla planifolia. Two types of explant (i.e. shoot apex and stem nodal segment) were screened for their shoot induction potential following a three-month treatment with 6-benzylaminopurine (BAP) and &#945-Naphthaleneacetic acid (NAA). Results indicated that the shoot apices were poor candidates for shoot induction whereas the stem nodal segments showed potential for shoot initiation at a rate of up to 6 shoots/explant. Stem nodal segments were the most responsive as shoots formed (55 shoots) directly following treatments with 1 mg/L BAP at half strength MS medium after the third subculture. In addition, more shoots were produced on solid medium treatments compared to the liquid medium treatments in two strengths of mediums tested. Regenerated plantlets derived from the 1 mg/L of BAP treatment were induced to root following a one month culture in growth regulator-free MS medium. There was 90% survival rate of the rooted plantlets after acclimatization in the greenhouse. The findings in the present study would be helpful for large-scale mass propagation of Vanilla planifolia using this simple and efficient protocol.

Antibiotic Resistance and Heavy Metals Tolerance in Gram-Negative Bacteria from Diseased American Bullfrog (Rana catesbeiana) Cultured in Malaysia

A total of 140 bacterial isolates have been successfully isolated from various organs of diseased American bullfrog （Rana catesbeiana） cultured in Malaysia. The most frequently isolated bacteria was Edwardsiella spp. （46 isolates） followed by Aeromonas spp. （33 isolates）, Flavobacterium spp. （31 isolates）, and Vibrio spp. （30 isolates）. Majority of the bacterial isolates were found sensitive to furazolidone （85.0%）, chloramphenicol （85.0%）, oxolinic acid （90.0%）, florfenicol （95.0%）, and flumequine （97.5%）. On the other hand, most of the bacterial isolates were resistant to oleandomycin （77.5%） and lincomycin （87.5%）. Nitrofurantoin and flumequine can be inhibited the growth of all of Vibrio spp. whereas all isolates of Edwardsiella spp. were found sensitive to florfenicol and flumequine. Multiple antibiotic resistance （MAR） index were in range of 0.30-0.40, indicating that bacterial isolates from cultured bullfrogs may have received high risk exposure to the tested antibiotics. In addition, 90-100% of the isolates were resistant to copper, cadmium, and chromium. These results provided insight information on tolerance level of bacterial isolates from cultured bullfrogs to 21 antibiotics as well as heavy metals.

Antimicrobial Property of 2-Hydroxypropane-1,2,3-Tricarboxylic Acid Isolated from Citrus microcarpa Extract

This article described antimicrobial property and structure analysis of 2-hydroxypropane-1,2,3-tricarboxylic acid isolated from the crude extract of Citrus microcarpa. Presently, there was no report on compound from C. microcarpa that possessed antimicrobial property against fish pathogenic bacteria. Therefore, in this study, the bioactive principle in C. microcarpa extract was isolated using thin layer chromatography. It＇s structure was elucidated based on nuclear magnetic resonance （NMR） spectroscopic data, such as proton NMR （1HNMR）, correlation spectroscopy, carbon 13 NMR, and heteronuclear multiple bond correlation data. This study showed that the bioactive compound isolated from C. microcarpa was 2-hydroxypropane-1,2,3-tricarboxylic acid monohydrate. The minimum inhibitory concentration （MIC） values of crude C. microcarpa extract and its bioactive component, 2-hydroxypropane-1,2,3-tricarboxylic acid as well as commercially available synthetic 2-hydroxypropane-1,2,3-tricarboxylic acid, were determined against 18 isolates of Edwardsiella tarda and 7 bacterial reference strains, namely, Escherichia coli （ATCC 25922）, Citrobacter freundii （ATCC 8090）, Aeromonas hydrophila （ATCC 49140）, Pseudomonas aeruginosa （ATCC 35032）, Streptococcus agalatiae （ATCC 13813）, E. tarda （ATCC 15947）, and Yersinia enterocolitica （ATCC 23715）, using two-fold microdilution method. The MIC values for both the natural 2-hydroxypropane-1,2,3-tricarboxylic acid and the synthetic one were ranging from 15.6 to 62.5 mg mL-1, whereas that of the crude extract was ranging from 7.8 to 31.3 mg mL-1. These findings showed that both the crude extract and its bioactive component might have potential as antimicrobial agent for aquaculture use.