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Microencapsulation of Lactobacillus plantarum 299v and its storage in kuini juice

Jian Ting Lai ; Ka Wai Lai ; Li Yu Zhu ; Kar Lin Nyam ; Liew Phing Pui

Malaysian Journal of Microbiology.2020;16(4):235-244. doi:10.21161/mjm.190398

Aims: Probiotics are living microorganism, when administrated in sufficient quantity can exert beneficial effect to the host. This study focused on the microencapsulation by co-extrusion to increase the viability of Lactobacillus plantarum 299v (Lp299v) in gastrointestinal conditions, and its storage stability in kuini juice at refrigerated (4 °C) and ambient temperature (25 °C). Methodology and results: Lp99v was encapsulated with 1.5% w/v sodium alginate and chitosan coating (0.1% w/v) and yielded a microencapsulation efficiency of 97.71%. The Lp299v microbeads produced were spherical in shape and exhibited a mean microbeads size of 618.75 ± 25.85 µm. Acid and bile tolerance of both free and encapsulated Lp299v were tested in simulated gastric juice (SGJ) for 2 h and in simulated intestinal juice (SIJ) for 4 h, respectively. The encapsulated Lp299v maintained above 108 CFU/mL after exposure to artificial gastrointestinal juice, whereas a significant loss of viability was observed in the free cells. The storage stability of encapsulated Lp299v in kuini juice was determined during 4 weeks of storage at 4 °C and 25 °C. Results showed that encapsulated Lp299v was capable to remain viable (107 CFU/mL) for at least 4 weeks in a refrigerated condition. However, free Lp299v did not survived under both refrigerated and ambient temperature as the storage period extended. Conclusion, significance and impact of study Lp299v entrapped in chitosan-coated alginate microbeads produced by co-extrusion method is able to enhance the viability of Lp299v above the minimum recommended level in harsh environment (gastrointestinal conditions and low pH of kuini juice).
Cell Encapsulation ; Lactobacillus plantarum

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Identification and characterization of antifungal compounds from a Burkholderia strain against plant pathogenic fungi

Theodorus Eko Pramudito ; Delia Agustina ; Widyah Budinarta ; Thi Kim Ngan Nguyen ; Cahya Prihatna ; Antonius Suwanto

Malaysian Journal of Microbiology.2020;16(4):245-252. doi:10.21161/mjm.190524

Aims: Biocontrol of fungal plant pathogens using beneficial microorganisms is a safer alternative over synthetic fungicides. PHP12 is a bacterial strain isolated from healthy oil palm rhizosphere and is closely related to the recently described Burkholderia stagnalis, a member of the Burkholderia cepacia complex. This study aimed to characterize the antifungal activity spectrum of PHP12 and identify the antifungal compounds produced by the strain. Methodology and results: The antifungal activity of PHP12 was characterized by growing fungal strains in the presence and absence of PHP12 and measuring the radius of the antifungal zone. PHP12 inhibited the growth of fungal pathogens including Ganoderma boninense, Curvularia oryzae, Phellinus noxius and Colletotrichum capsici. However, PHP12 did not inhibit the growth of Trichoderma asperellum, a known fungal biocontrol agent. The antifungal compounds of PHP12 were precipitated using ammonium sulfate and further purified with HPLC followed by identification using Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC/ESI-MS). The LC/ESI-MS analysis showed the presence of an oligopeptide with a molecular weight of 1210.63 Da. The peptide consists of heavily modified amino acids that are linked by a hexose residue. Conclusion, significance and impact of study Although characteristics of the antifungal compounds are similar to other antifungal peptides from Burkholderia such as occidiofungin, there have been no reports of antifungal peptides from B. stagnalis with the corresponding molecular weight or fragmentation profile. The novelty of the compound, as well as its antifungal spectrum, makes PHP12 an interesting strain to be investigated further as a biocontrol agent.
Fungicides, Industrial ; Burkholderia cepacia complex--pathogenicity

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Isolation and identification of phosphate solubilizing bacteria from indigenous microorganisms (IMO) of cow rumen in East Java, Indonesia as eco-friendly biofertilizer

Meli Astriani ; Siti Zubaidah ; Abdul Latief Abadi ; Endang Suarsini

Malaysian Journal of Microbiology.2020;16(4):253-262. doi:10.21161/mjm.190536

Aims: Phosphate is an essential nutrient required for plant growth, but its solubility in the soil is relatively low (0.1%). Microbes can dissolve phosphate to meet crop requirements. This study aimed to isolate phosphate solubilizing bacteria from indigenous microorganisms (IMO) of cow rumen. Methodology and results: The selection of isolates on a Pikovskaya medium was using a clear zone index and a spectrophotometer for phosphate solubilization measurements. Hypersensitivity was tested on tobacco leaves and tested antagonists within isolates. The results found that four selected isolates had the highest phosphate dissolving potential, namely, MTA1, SMAD1, SMAD2, and SMAD3. The culture of selected isolates on plate media showed that the morphological characters of the four colonies are the same. They had round form (circular), the edge of the colony were smooth, flat elevation, white and cream color. Isolate MTA1 had the highest phosphate solubilizing activity compared to the others. The isolate that showed the highest phosphate solubilizing activity were identified based on 16S rRNA gene. The result of molecular identification showed that strain MTA1 was closely related to Lactobacillus plantarum with a similarity level of 99%. L. plantarum performed the highest ability to form a clear zone (7.66 mm). The highest concentration of soluble phosphate was observed on day 5 (278.42 mg/L). Conclusion, significance, and impact of the study Lactobacillus plantarum which was isolated from the IMO of cow rumen in East Java, Indonesia was identified as one of the phosphate solubilizing bacteria that are useful for the development of eco-friendly biofertilizer. The application of phosphate solubilizing microbes can be used to increase the soil fertility.
Fertilizers--microbiology ; Indonesia (East Java)

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Distribution and prevalence of antibiotic resistant bacteria in fish farms in East Malaysia

Samuel Lihan ; Nurul Asyiqin Jamil ; Mohd Azizul Hafiz Jamian ; Toh Seng Chiew ; Olaide Olawunmi Ajibola ; Sabella Justin ; Flonia Benet ; Lee Nung Kion

Malaysian Journal of Microbiology.2020;16(4):263-274. doi:10.21161/mjm.190522

Aims: Aquaculture has grown tremendously in Malaysia over the past decades. However, guaranteeing aquaculture sustainability is a big challenge in terms of maintaining continuous output with a safe environment. Furthermore, the cultured species should be free from antibiotic resistance bacterial and antibiotic residue. This study aimed to monitor the existence and prevalence of antibiotic resistant bacteria associated with aquaculture farms in Sarawak. Methodology and results: Samples of water, sediment and fish were collected from five aquaculture farms within Sarawak. The samples were plated on trypticase soy agar and incubated at 28 °C for 24 h. A total of 204 bacterial isolates were isolated and analysed by (GTG)5-fingerprinting to determine genetic similarity among the bacterial isolates, so that representatives could be selected from similar clonal isolates. Based on the (GTG)5 profiles, 50 representative isolates were chosen for species identification using 16S rRNA sequencing. The identified bacteria were tested against 25 antibiotics using standard disk diffusion method. The 16S rRNA analysis revealed that the isolates constitute of 14 genera of bacteria including Bacillus (38%), Exiguobacterium (16%), Enterobacter (14%), Aeromonas (6%), Acinetobacter (4%), Citrobacter (4%), Staphylococcus (4%), Achromobacter (2%), Chitinophaga (2%), Fictibacillus (2%), Plesiomonas (2%), Pseudomonas (2%), Pseudoxanthomonas (2%) and Stenotrophomonas (2%). The antibiotic resistance analysis revealed that the highest percentage of resistance was recorded against streptomycin (75.0%), followed by ampicillin (66.0%), ceftriaxone (50.0%), rifampin (43.3%), aztreonam (36.8%) and ceftazidime (31.6%). Resistance to more than two antibiotics was observed in 40.0% of isolates with an overall multiple antibiotic resistant (MAR) index ranging from 0 to 0.79. Conclusion, significant and impact of study The variability of antibiotic resistance patterns exhibited by different bacterial species suggests a dependence on selective pressures exhibited in different geographical locations. Our results show that the occurrence of MAR bacteria in an aquaculture environment with unknown history of antibiotics usage in the aquaculture system is possible, indicating a need to continuously monitor the presence of antibiotic resistant bacteria in the aquaculture system.
Drug Resistance, Microbial ; Aquaculture ; Malaysia

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Genotypic profiles of virulent genes detected among the Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa isolated from swiftlets in Borneo

Sui Sien Leong ; Samuel Lihan ; Hwa Chuan Chia

Malaysian Journal of Microbiology.2020;16(4):275-284. doi:10.21161/mjm.190554

Aims: The occurrence of multiple pathogenic Enterococcus faecalis, Escherichia coli, and Pseudomonas aeruginosa are important nosocomial and hazardous infection clinically challenge worldwide. Thus, the aim of this study was to screen for the virulent genes profiles to ascertain their prevalence in swiftlets in Borneo. Methodology and results: The Enterococci, E. coli and P. aeruginosa bacteria were isolated from the swiftlets’ faeces and air inside swiftlet houses, which located in the Southern, Central and Northern regions of Borneo. The isolates were identified to the species level by 16S rRNA sequencing assay. Specific primers were designed for detection of the potential virulence genes in E. faecalis (ace, AS, efaA and gelE), E. coli (stx) and P. aeruginosa (oprL) by PCR assay. A total of 38 Enterococci, 26 of E. coli and 2 of P. aeruginosa fecal and airborne bacteria were identified. Sixty-seven percent of E. faecalis isolates were detected positive for four virulence genes, 27% possessed three (AS, efaA, gelE) genes and 6% possessed two (ace, AS) genes. There were no stx genes detected among all the E. coli isolates. The oprL gene was detected in all the P. aeruginosa isolates. Conclusion, significance and impact of study Virulence genes are important in the pathogenesis of both clinical and avian infections which considered to be a serious public health threat. The high incidence of virulence genes detection in E. faecalis and P. aeruginosa indicates these genes were widely disseminated among the bacteria found in swiftlet houses, suggesting the important issues in the pathogenesis of infections and diseases which may cause potential health risks to humans.
Virulence--genetics

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Orthosiphon stamineus water extracts inhibit human herpes virus 1 KOS-1 and acyclovir-resistant strains by virucidal activity and suppressing virus early infection

Maysam Dahham Habboo ; Norefrina Shafinaz Md. Nor ; Nazlina Ibrahim

Malaysian Journal of Microbiology.2020;16(4):285-293. doi:10.21161/mjm.190568

Aim: To determine the efficacy and mode of action of hot and cold water extracts of Orthosiphon stamineus leaves against two strains of human herpes virus 1 (HHV-1) i.e. KOS-1 and acyclovir (ACV)-resistant UKM-1 (UKM-1) strains. Methodology and results: Hot and cold water extracts of O. stamineus were not cytotoxic to vero cells as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) (MTT) assay with 50% cytotoxicity concentration (CC50) values of 3.4 and 3.3 mg/mL respectively. Antiviral activity was determined by plaque reduction assay in post-treatment, pre-treatment and virucidal assays followed by time-addition and time removal assay to relate with the stages during the viral infection cycle. Both extracts displayed antiviral activity against HHV-1 KOS-1 and HHV-1 UKM-1 strains with 50% effective concentration (EC50) values between 0.12-0.15 mg/mL in reducing plaque formation. The calculated selectivity indices (SI) were 23 and 28 for hot and cold water extract respectively, indicating that they have good potential as antiviral agent. The extracts were virucidal towards both HHV-1 KOS-1 and HHV-1 UKM-1 strains which may directly affects the virus structure. This is supported with the fact that exposure of the extracts inhibit viral attachment and penetration to the vero cells. In time-of addition assay, both extracts were effective during the early stage of virus infection cycle for HHV-1 KOS-1 strain which is in parallel with the results from the attachment and penetration studies. For HHV-1 UKM-1 strain, contact to the extracts at any time during post-infection inhibits virus replication and also progeny release. Conclusion, significance and impact of study Cold and hot water extracts of O. stamineus have good potential as antiviral agent against HHV-1 strain KOS-1 and more importantly against UKM-1 strain which is ACV-resistant. The extracts displayed virucidal effect and inhibition of early virus replication cycle involving viral attachment and penetration to cells.
Orthosiphon--adverse effects

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Prevalence, associated risk factors and antibiotic resistance pattern of bacterial uropathogens among pregnant women in a tertiary care hospital of Bangladesh

Sohana Al Sanjee ; Masudur Rahman ; Md. Ekramul Karim ; Umme Salma Sigma

Malaysian Journal of Microbiology.2020;16(4):294-301. doi:10.21161/mjm.190595

Aims: The study was designed to determine the prevalence of urinary tract infection (UTI) in pregnant women depending on their various clinical and socio-demographic factors, and to assess the antibiotic susceptibility pattern of the responsible uropathogens in a tertiary care hospital of Dhaka, Bangladesh. Methodology and results: A total of 100 midstream urine samples were collected from pregnant women and different clinical and socio-demographic variables viz. age, gestational weeks, living conditions, and level of education associated with UTI were determined. Bacterial isolation was carried out using blood and MacConkey agar and identified according to their phenotypic characteristics. Antibiogram profiling of the isolates was done by disc diffusion method. From 48% of positive UTI samples, the highest bacteriuria was recorded within the age group of 26-30 years (n=19; 59.38%) and in both, 1st and 3rd trimester period (50%). There was no significant association between the studied risk factors and bacteriuria, except for the age of the pregnant women. Most predominantly isolated bacteria was Escherichia coli (n=39; 81.25%), followed by Klebsiella pneumoniae (n=9; 18.75%). In E. coli, the highest resistance was recorded against ceftriaxone (87.18%), followed by cephalexin (84.61%) and ceftazidime (79.49%); whereas K. pneumoniae showed 100% resistance to ceftriaxone and cephalexin. Netilmicin was found as the only effective antibiotic against E. coli showing 100% sensitivity. For K. pneumoniae, azithromycin, imipenem, chloramphenicol, gentamicin, ciprofloxacin, amikacin and nitrofurantoin were found as the most efficacious drugs. Conclusion, significance and impact of study As the emergence of drug resistance is ever increasing, the study necessitates the continuous surveillance of antibiotic susceptibility of uropathogens to ensure safety and better treatment to the mother and fetus.
Urinary Tract Infections--microbiology ; Drug Resistance, Microbial ; Pregnant Women ; Bangladesh (Daka)

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Prevalence of antibiotic resistant bacteria isolated from raw chicken meat

Sarannia Thanganathan ; Astha Latchumi Sanglidevan ; Kamariah Hasan

Malaysian Journal of Microbiology.2020;16(4):302-311. doi:10.21161/mjm.190633

Aims: Antibiotics are widely used in poultry industry for treatment, control and in preventing the spread of infectious diseases among chicken flocks. The uncontrolled use of antibiotic causes the emergence of antibiotic resistant bacteria which is a major concern worldwide. The aim of this study is to isolate and molecularly identify antibiotic resistant bacteria using raw chicken meat samples from farm, supermarket, wet market as well as free-range chicken. Methodology and results: A total of 34 isolates were obtained through primary screening based on their ability to grow on streptomycin, kanamycin, ampicillin and cefazolin antibiotic plates. Kirby-Bauer disc diffusion test performed on the 34 isolates showed that they were highly resistant to oxacillin (97%) and penicillin (94%) followed by ampicillin (64%), cefazolin (50%), tetracycline (32%), erythromycin (24%), ciprofloxacin (21%) and least resistance towards gentamycin (6%). Eight isolates with the highest antibiotic resistance, were selected for molecular identification using 16S rDNA sequencing. Analysis of the 16S rDNA sequence using BLASTN and phylogenetic tree constructed on the selected isolates revealed that five different species of antibiotic resistant bacteria namely Escherichia coli, Klebsiella sp., Chryseobacterium gleum, Comamonas testosteroni and Bacillus cereus were successfully identified from the different types of chicken sample. Conclusion, significance and impact of study The excessive use of antibiotic in the poultry farm industries had caused the emergence of antibiotic resistant bacteria which can harm the health of people consuming chicken meat. To overcome this crisis, antibiotic usage in the poultry farm industries should be regulated.
Drug Resistance, Bacterial ; Poultry

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Antimicrobial, cytotoxicity and preliminary phytochemical determination of commonly used medicinal plants to treat oral cavity, urinary tract and gut infections by inhabitants of Borabu sub-county, Nyamira County, Kenya

Eric Omori Omwenga ; Francisco Maria Goycoolea ; Andreas Hensel ; Anakalo Shitandi

Malaysian Journal of Microbiology.2020;16(4):312-322. doi:10.21161/mjm.200697

Aims: The study aimed at determining the antimicrobial activities and cytotoxicity properties of medicinal plants collected from southwestern Kenya. Methods and results: A total of 23 ethanol extracts of selected medicinal plants were bio-assayed against Gramnegative bacterial strains (Escherichia coli NU14, Helicobacter pylori ATCC 700824, and Porphyromonas gingivalis ATCC 33277). Cytotoxicity tests were also carried out on mammalian cell lines (AGS, KB, and TR146). Preliminary type of phytochemical compounds present in the extracts was determined by thin-layer chromatography. Cassia didymobotrya plant extract (1 mg/mL) had strong antimicrobial activity against P. gingivalis (average zone of inhibition of 21.70 ± 0.88 mm, MIC 0.13 ± 0.00 mg/mL and MBC 0.50 ± 0.00 mg/mL). Escherichia coli was resistant to all the extracts bioassayed. Leonotis nepetifolia (15.80 ± 0.20 mm) and Clerodendrum myriacoides (14.20 ± 0.44 mm) showed only moderate activity against H. pylori. Cell cytotoxicity results indicated a dose-dependent response against KB, TR146 and AGS cell lines with C. didymobotrya having IC50 values of 47.64 and 704.00 µg/mL on KB and TR146 cell lines, respectively. L. nepetifolia and C. myriacoides did produce IC50 of 0.1883 mg/mL and 0.1061 mg/mL against the AGS cell line respectively. Conclusion, significance and impact of the study Most of the extracts had no or weak activity against test isolates, but C. didymobotrya leaves extracts showed strong activity against P. gingivalis. C. didymobotrya can offer alternative medicare to P. gingivalis conditions.
Plants, Medicinal--adverse effects ; Kenya (Southwestern)

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Phytochemicals with antifungal properties: Cure from nature

Amal A. M. Elgharbawy ; Nurhusna Samsudin ; Farah Fadwa Benbelgacem ; Yumi Zuhanis Has-Yun Hashim ; Hamzah Mohd. Salleh ; Jacinta Santhanam

Malaysian Journal of Microbiology.2020;16(4):323-345. doi:10.21161/mjm. 190551

Aims: The exploration of natural products with innovative uses is dynamic and expanding rapidly. Medicinal plants have fascinated many researchers that subsequently lead to research publications highlighting plant extracts with wide range of secondary metabolites such as flavonoids, alkaloids, glycosides, quinones, terpenoids, tannins and saponins that exhibit antimicrobial activities and disease control. The concentration of these bioactive compounds in each plant species varies based on the pathosystem and environmental conditions. This study aims to uncover the various types of phytochemicals with antifungal properties. Methodology and results: Seven categories of plant-based antifungal compounds were reviewed, which are terpenoids, saponins, phenolic compounds, coumarins, alkaloids, essential oils and peptides, with examples and structures of some available compounds. The mechanism of action of each category of phytochemical was discussed. Also, the impact of some compounds was explained and elaborated. Conclusion, significance and impact of study It is of a great importance to explore natural plant fighters against fungal infection. Those active plant components do not only have antifungal properties, but they also help in the healing process and some even exhibit anticancer activities. The development and knowledge of antifungal activities from plant extracts have the potential for applications in antifungal therapy. Since the exact description of how antifungal compounds function in the human body is still unclear more studies are required to unveil phytochemicals’ properties and to elucidate their effects on living cells.
Phytochemicals--therapeutic use ; Antifungal Agents

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