MALAYSIAN JOURNAL OF MICROBIOLOGY
2021 - Volume 17
Aims: The development of an effective biocontrol formulation for inhibition of Ganoderma boninense, a well-known destructive pathogen in oil palm plantation is important to prolong the palm’s lifespan and reduce the losses due to this disease. In this paper, we present some new bioformulations with combination of different types of biocontrol agents in managing basal stem rot (BSR) disease.
Methodology: The effectiveness of the treatments designed as T1 (Trichoderma harzianum + Lecanicillium spp. + Streptomyces sundarbansensis + Pseudomonas aeruginosa), T2 (Penicillium simplicissimum + Lecanicillium sp. + S. sundarbansensis + P. aeruginosa), T3 (P. simplicissimum + P. aeruginosa) and T4 (LEStani®) was evaluated through treatment on the oil palm seedlings artificial infected by G. boninense and the results were expressed in disease severity index (DSI), bole severity index (BSI) and ergosterol content.
Conclusion, Significance and Impact of study: All tested treatments (T1-T4) managed to control the severity of the Ganoderma infection from continuously increasing when the treatments were applied either one month before or after artificial inoculation. The disease severity of infected seedlings without treatments had increased for almost 2-fold at the end of the trial. Moreover, T1 had the greatest inhibition of G. boninense with the lowest ergosterol content (a bioindicator of Ganoderma colonization) detected (676.67 g/mL), which is about 1.9-fold lower than control (1273.33 g/mL) without treatments and a BSI score of 1. Based on the effectiveness among the four tested biocontrol formulations, T1 is the most promising formulation to be further evaluated in the field for control of BSR disease. However, more research is needed in the future to estimate the effective amount for application in open environment.
Aims: The aim of this study was to screen lactic acid bacteria (LAB) isolates from fermented Sumbawa mare‘s milk that meet the requirements as starter cultures, and to evaluate the effect of the selected starter culture in improving the organoleptic quality of mare‘s milk fermentation.
Methodology and results: The LAB isolates (13 isolates) derived from naturally fermented Sumbawa mare‘s milk were firstly screened for acidification activity. Afterwards, the selected isolates were evaluated for the starter culture criteria such as technological properties (proteolytic test, lipolytic test, and exopolysaccharide production), food safety test (hemolytic test and antibiotic sensitivity test), antimicrobial activity test. The selected culture (SC) together with yogurt starter cultures (YC) and combination between the selected isolate and a mixture of both (MC) were used to ferment fresh mare’s milk. Six LAB isolates (DB7, BC10, DC4, BC9, DC10, and BC7) were obtained from the acidification screening. Isolate BC10 was the most potential isolate as starter culture due to its ability in terms of acidification and proteolytic activity, lack of lipolytic activity, no indication of pathogenic potency, as well as able to inhibit the growth of Escherichia coli ATCC 25922. However, this isolate was resistant to antibiotics kanamycin, trimethoprim, and cinoxacin. The isolate BC10 presented 99.99% sequence similarity with respect to Lactobacillus plantarum.
Conclusion, significance and impact of study: The selected starter culture (isolate BC10) was able to improve the organoleptic quality of fermented mare‘s milk especially aroma compared to the other starter cultures. Therefore, Lactobacillus plantarum BC10 is a potential isolate to be used as starter culture for mare’s milk fermentation.
Aims: Cholera epidemics have been occurred in Malaysia since 1991 till 2003 which can be proved from the records by the Infectious Diseases Division of the Ministry of Health. Moreover, there were also course of cholera epidemics from the year 1994 to 2003 which had been happened in Sarawak. Cholera outbreaks in Malaysia mostly caused by the El Tor O1 Vibrio cholerae serogroup. The aims of this study were to detect the presence of V. cholerae in clinical and environmental samples (n=28) from Limbang, Sarawak by collaboration with Sarawak Government Hospital and to detect the toxin genes from the isolates.
Methodology and results: All the isolates were sub-cultured in alkaline peptone water (APW). The boiled-cell method was used for DNA extraction. The total DNA extracted was amplified by polymerase chain reaction (PCR). Two types of PCR were used in this study which are 16S rRNA PCR and multiplex PCR. The results obtained from the study found out that 16 out of 28 (57.14%) samples were confirmed to be V. cholerae species. Four primers specific for V. cholerae were used in multiplex PCR (O1 type, O139 type, ctxA and ctxAB) to confirm the species type and the toxin genes. All samples shown positive for V. cholerae O1 serotype and 100% positive to all genes for the identification of ctxA and ctxAB genes.
Conclusion, significance and impact of study: From this study, it showed that multiplex PCR can be used for research purposes in molecular genetics field involving cholera outbreak.
Aims: The attention for new and effective anticancer drugs but less toxic is increasing over time. Streptomyces is the most important and well-known source of their bioactive compound production with useful bioactivities. This work aimed for evaluation of the anticancer potential of methanolic extract of Streptomyces sp. strain KSF 83 against non-cancerous cell lines (CCD-841-CoN), breast (MCF-7, MDA-MB-231) and colon cancer cell lines (HT-29, HCT-116).
Methodology and results: The characteristic of the strain KSF 83 was identified by morphology and 16S rRNA sequencing and results confirmed that the strain belonged to the genus of Streptomyces. The crude substance was produced via submerged fermentation from the strain and methanol solvent was used to extract the culture filtrate. Methanolic extract possessed low toxicity against CCD-841-CoN with only 18% of inhibition activity at the 400 µg/mL. Among all tested cancer cells, the methanolic extract was able to inhibit the growth of all cancer cells tested with MCF-7 was the highest anticancer activity recorded. The methanolic extract also exhibited cytotoxicity in a range of EC50 of 65.79 μg/mL to 262.40 μg/mL. This study revealed the anticancer potential of *Streptomyces *sp. strain KSF 83, which could be sources of prospective anticancer drugs against breast and colon cancer.
Conclusion, significance and impact of study: The extract of KSF 83 was non-toxic toward normal cell lines and able to inhibit the growth of breast and cancer cell lines, thus it can be a potential source of the anticancer drug against breast and colon cancer.
Aims: To evaluate the antibacterial efficacy of ethyl acetate extract of Aspergillus flavus IBRL-C8 against Gram-positive and Gram-negative bacteria.
Methodology and results: In this experiment, an endophytic fungus which identified as A. flavus IBRL-C8 was extracted using ethyl acetate and methanol, from Senna siamea, prior to in vitro antibacterial test on eight Gram-bacteria. The results were significantly more enunciated to the ethyl acetate extract since the Gram-bacteria signified 9.0 to 20.0 mm of inhibition zones on Muller Hinton Agar (MHA) during disc diffusion assay. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of the extract were ranged from 125–1000 µg/mL and 125–2000 µg/mL, respectively. Time-kill assay depicted the ethyl acetate extract of A. flavus IBRL-C8 exceptionally retarded methicillin-resistant Staphylococcus aureus (MRSA) and also manifested extended antibacterial activity. The maximum reduction in cell numbers occurred at 2MIC concentration (250 µg/mL) during the interval time of 16 h. The malformations noticed from microscopic observations where the transformation of structural annihilation from regular spherical morphology to non-spherical shape with an irregular surface and also disruption around the cell membrane when the MRSA treated with ethyl acetate extract of A. flavus IBRL-C8.
Conclusion, significance and impact of study: This study proposed the ethyl acetate extract of A. flavus IBRL-C8 as a potential antibacterial agent against MRSA infection, which can be useful in pharmaceutical application.
Aims: This study aimed to isolate and identify fungi involved in causing diseases to Vanilla planifolia as well as to study their pathogenicity level in causing disease.
Methodology and results: The diseased parts of vanilla plants were collected from vanilla farms located in Pahang and Sabah, Malaysia from May 2015 to May 2016. Diseases tissue transplantation was adopted to isolate the fungi for morphology identification prior to the polymerase chain reaction (PCR) amplification of internal transcribed spacer (ITS) regions using universal primers for fungi, ITS1 and ITS4. After being isolated, the fungi pathogenicity was tested on detached fresh and mature vanilla leaves. A total of 22 fungal isolates were identified, Fusarium fujikuroi and F. oxysporum were the two most recovered species, followed by Colletotrichum gloeosporioides, Fusarium sp., F. proliferatum and F. solani. Pathogenicity test revealed a significantly high pathogenicity of F. oxysporum and C. gloeosporiodes (p<0.01) on detached vanilla leaf, with high level of damage.
Conclusion, significance and impact of study: This study provides valuable information on fungi-associated diseases on vanilla plants grown in Malaysia and can be used for future development in disease management.
Aims: This study aimed to screen the plant growth-promoting fluorescent bacteria (FLB) which isolated from the healthy rice rhizophere and to evaluate its biocontrol and growth promotion properties against Pyricularia oryzae on aerobic rice seedling of MARDI Aerob 1.
Methodology and results: King’s B agar with glycerol was used as the selective medium to isolate FLB from the healthy rice rhizosphere soil. All FLB obtained were in vitro screened for antagonistic activities against P. oryzae using dual culture, volatile substances and hydrogen cyanide productions. The potential FLB isolates were further evaluated on rice seedling early growth promotion before identified using 16S rRNA gene sequencing. A total of 24 FLB were isolated from the healthy rice rhizosphere soil in Setiu, Terengganu, Malaysia. Isolates: FLB4, FLB5, FLB7 and FLB10 scored the total of percentage inhibition radial growth (PIRG) values ranged 99.5-105.0%. Further seedling growth promotion screening revealed that FLB4, FLB7 and FLB10 were significantly improved seedling growth with vigor index of 378.32%, 461.53% and 335.60% over control (133.31%). 16S rRNA sequencing identified that FLB7 as Bacillus subtilis and the FLB4 and FLB10 as Pseudomonas putida.
Conclusion, significance and impact of study: The selected FLB isolates (FLB4, FLB7 and FLB10) are potential to be developed as biological control agents against P. oryzae with growth promoting property on aerobic rice seedling.
Aims: Lytic polysaccharide monooxygenase (LPMO) is an enzyme capable of cleaving glycoside bonds of recalcitrant polysaccharides through an oxidative mechanism. LPMO activity, in synergy with hydrolytic enzymes, increases the production of monomer sugars from the biodegradation of lignocellulose. This study was aimed at evaluating actinomycete S2 strain LPMO activity based on the release of xylose as one of reducing sugar and hydrogen peroxide (H2O2) in the course of lignocellulosic biodegradation.
Methodology and results: The oxidation activity of LPMO from actinomycete S2 strain was measured by using the substrate of Avicel supplemented with ascorbic acid and copper ions (Cu2+) to identify its effect on the release of xylose as one of reducing sugar. The optimum incubation time for the LPMO production was also conducted. Further, H2O2 quantitative analysis was performed as by-product of LPMO activity and 16S rRNA gene sequence of actinomycete S2 strain were subsequently determined. We found that supplementation of 1 mM ascorbic acid and 0.2 mM Cu2+ increased xylose as one of reducing sugar production by up to 5-fold from 255.03 to 1290 μg/mL after an optimal incubation period of 6 days. Based on H2O2 production, the LPMO activity of actinomycete S2 strain was 0.019 ± 0.001 U/mL. There is likelihood that LPMO activity derived from actinomycete S2 strain has a synergistic effect with the activity of other lignocellulose-degrading enzymes. This actinomycete showed 99% similarity to the 16S rRNA gene sequence of Streptomyces avermitilis strain EAAG80.
Conclusion, significance, and impact of study: LPMO enzyme activity from actinomycete S2 strain as determined by the production of reducing sugar and H2O2 was greatly increased by supplementation with ascorbic acid as an electron donor and Cu2+ ions. To the best of our knowledge, this is the first elucidation of LPMO activity from an indigenous Indonesian actinomycete.
Aims: Coral diseases have emerged over the last several decades, causing a loss of live coral cover in the Caribbean and Indo-Pacific reefs. Hence, microbiological and disease cultural techniques are commonly used to investigate their causative microbial agents. This is the first study to identify the potential of pathogenic Vibrio spp. isolated from apparently white syndrome (WS) coral disease in Tioman Island Marine Park using biochemical and molecular techniques.
Methodology and results: The Vibrio colonies were isolated from 108 samples of WS infected corals (Acropora cytherea and Montipora aequituberculata) including seawater, sediment and algae found adjacent to infected coral colonies. A total of one hundred representative Vibrio isolates were characterized and most of them (n=50) were identified as V. vulnificus, V. alginolyticus and Photobacterium damselae following biochemical analysis. The molecular analysis revealed six Vibrio spp. (V. coralliilyticus, V. hepatarius, V. brasiliensis, V. tubiashi, V. campbellii, V. ishigakensis) and one Photobacterium rosenbergii. V. coralliilyticus isolated from all infected coral samples may be highly responsible for the sign of WS disease.
Conclusion, significance and impact of study: The findings of this study provide baseline data and information on potential coral pathogens identified in the coastal waters of Tioman Island. Etiological disease study is suggested to validate their severity and virulence factors in the future.
Aims: Knowledge of the Trichoderma taxa is important for both control efficiency and environmental conservation. Therefore, the objective of this study is to isolate and identify Trichoderma species from various rhizosphere soil samples using phenotypic and molecular characterization.
Methodology and results: Native Trichoderma spp. were isolated from agricultural fields in 17 sites from seven states of Malaysia. These isolates were characterized via morphological observation and molecular phylogenetic analysis based on the translation elongation factor-1α (tef1-α) gene. About 42 isolates were classified into eight Trichoderma species, which are Trichoderma asperellum, T. hamatum, T. harzianum, T. koningiopsis, T. rodmanii, T. spirale, T. viride and T. virens. Comparison of DNA sequences of tef1-α showed that the isolates were 98–100% similar to respective Trichoderma species from GeneBank, thus confirming the fungal identity. Phylogenetic trees of maximum likelihood (ML) dataset of tef1-α inferred that the isolates were clustered according to species.
Conclusion, significance and impact of study: Findings in the present study will be beneficial for the purposes of biodiversity conservation and plant disease management using biocontrol agents.
Aims: Acne is a common skin disease among teenagers and also affects other ages. It occurs when the oil and dead skin cells plug into the hair follicles and causing pimples or whitehead. Although antibiotics have been used for many years in treating acne, the widespread use of it has led to the development of bacterial resistant, which resulted in unsuccessful treatment. Thus, in this study, Andrographis paniculata (AP) herbal formulation gel is proposed in order to determine its effectiveness in treating acne. Three different methodologies were used to compare the antimicrobial effect of A. paniculata herbal gel against acne-associated pathogens.
Methodology and results: Well diffusion, disc diffusion and broth dilution methods were applied to evaluate the antimicrobial effect of AP herbal gel at concentrations of 1.5% (w/w), 2.5% (w/w) and 5.0% (w/w) onto selected pathogens associated with acne which consisted of Staphylococcus epidermidis, Staphylococcus aureus, Propionibacterium acnes and Candida albicans. Among the three methods, broth dilution showed the best antimicrobial effect towards all microorganisms used. AP herbal gel at concentration 2.5% (w/w) showed the optimum antimicrobial effect of S. aureus and C. albicans, while 5.0% (w/w) exhibited the best antimicrobial activities for P. acnes and S. epidermidis.
Conclusion, significance and impact of study: Broth dilution method appears to be a reliable method for the determination of antimicrobial effects for the pathogens tested. In addition, AP herbal formulation gel has great potential to treat acne effectively.
Aims: Metarhizium anisopliae is an entomopathogenic fungus (EPF) that exists naturally in the environment and potentially be used as a biological control agent against many insect pests. This study aims to evaluate the effect of nutrient additives on the yield and viability of M. anisopliae spore and to determine the optimum incubation period for maximum spore production.
Methodology and results: In this study, M. anisopliae was cultivated by solid-state fermentation using rice as a growth medium. Three different nutrient additives were examined which aimed to maximize the production of M. anisopliae spores. Among the three nutrient additives evaluated, yeast (1.84 ± 0.04 g) supported better growth and spore production than molasses (0.58 ± 0.04 g) and palm oil (0.47 ± 0.09 g). The incubation period between 2–6 weeks produced higher spore yield (0.97 ± 0.02 g spores) at week 4 with a better spore viability (86.30 ± 0.45%) at week 2.
Conclusion, significance and impact of study: Hence, it is suggested that the optimum incubation period is between 2 and 6 weeks after inoculation, and M. anisopliae could be mass produced in large quantities on rice substrate with the addition of yeast as the nutrient additives.