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Fulltext Postbiotic metabolites produced by Lactobacillus plantarum strains exert selective cytotoxicity effects on cancer cells

Chuah LO, Foo HL, Loh TC, Mohammed Alitheen NB, Yeap SK, Abdul Mutalib NE, et al.

BMC Complement Altern Med, 2019 Jun 03;19(1):114.
PMID: 31159791 DOI: 10.1186/s12906-019-2528-2

BACKGROUND: Lactobacillus plantarum, a major species of Lactic Acid Bacteria (LAB), are capable of producing postbiotic metabolites (PM) with prominent probiotic effects that have been documented extensively for rats, poultry and pigs. Despite the emerging evidence of anticancer properties of LAB, very limited information is available on cytotoxic and antiproliferative activity of PM produced by L. plantarum. Therefore, the cytotoxicity of PM produced by six strains of L. plantarum on various cancer and normal cells are yet to be evaluated.
METHODS: Postbiotic metabolites (PM) produced by six strains of L. plantarum were determined for their antiproliferative and cytotoxic effects on normal human primary cells, breast, colorectal, cervical, liver and leukemia cancer cell lines via MTT assay, trypan blue exclusion method and BrdU assay. The toxicity of PM was determined for human and various animal red blood cells via haemolytic assay. The cytotoxicity mode was subsequently determined for selected UL4 PM on MCF-7 cells due to its pronounced cytotoxic effect by fluorescent microscopic observation using AO/PI dye reagents and flow cytometric analyses.
RESULTS: UL4 PM exhibited the lowest IC50 value on MCF-7, RG14 PM on HT29 and RG11 and RI11 PM on HL60 cell lines, respectively from MTT assay. Moreover, all tested PM did not cause haemolysis of human, dog, rabbit and chicken red blood cells and demonstrated no cytotoxicity on normal breast MCF-10A cells and primary cultured cells including human peripheral blood mononuclear cells, mice splenocytes and thymocytes. Antiproliferation of MCF-7 and HT-29 cells was potently induced by UL4 and RG 14 PM respectively after 72 h of incubation at the concentration of 30% (v/v). Fluorescent microscopic observation and flow cytometric analyses showed that the pronounced cytotoxic effect of UL4 PM on MCF-7 cells was mediated through apoptosis.
CONCLUSION: In conclusion, PM produced by the six strains of L. plantarum exhibited selective cytotoxic via antiproliferative effect and induction of apoptosis against malignant cancer cells in a strain-specific and cancer cell type-specific manner whilst sparing the normal cells. This reveals the vast potentials of PM from L. plantarum as functional supplement and as an adjunctive treatment for cancer.
Fulltext Chemical Compositions of Brown and Green Seaweed, and Effects on Nutrient Digestibility in Broiler Chickens

Azizi MN, Loh TC, Foo HL, Akit H, Izuddin WI, Shazali N, et al.

Animals (Basel), 2021 Jul 20;11(7).
PMID: 34359273 DOI: 10.3390/ani11072147

This study aimed to analyse the nutritional properties, apparent ileal digestibility (AID) and apparent metabolisable energy (AME) of broiler chickens fed with brown seaweed (BS) and green seaweed (GS). Proximate analysis was performed to determine the nutrient composition of seaweed. The amino acids were determined using high-performance liquid chromatography (HPLC), and atomic absorption spectroscopy was used to determine the minerals content. The gross energy (GE) was determined using a fully automatic bomb calorimeter, and the AME value was calculated. Titanium dioxide (TiO2) was used as an indigestible marker to calculate the AID. A digestibility trial was conducted to investigate the effects of seaweeds on crude protein (CP), crude fibre (CF), ether extract (EE), dry matter (DM), organic matter (OM), amino acids (AA) and minerals digestibility, and AME on broiler chickens. Thirty-six broiler chickens were randomly distributed into two dietary treatment groups with six replicates and three birds per replicate. Results showed that brown and green seaweed was a source of macro and micronutrients. For the AME and AID of seaweed-based diets, the results showed that the AME value for BS and GS was 2894.13 and 2780.70 kcal/kg, respectively. The AID of BS and GS was 88.82% and 86.8% for EE, 82.03% and 80.6% for OM, 60.69% and 57.80% for CP, 48.56 and 44.02% for CF, and 17.97 and 19.40% for ash contents, respectively. Meanwhile, the AID of CP and CF was significantly higher for BS compared to the GS. Findings showed that the AID of various AA was 40.96 to 77.54%, and the AID of selected minerals (Ca, Na, K, Mg, Zn, Cu, Fe) for both BS and GS groups were above 90%.
Fulltext Dietary Supplementation of Postbiotics Mitigates Adverse Impacts of Heat Stress on Antioxidant Enzyme Activity, Total Antioxidant, Lipid Peroxidation, Physiological Stress Indicators, Lipid Profile and Meat Quality in Broilers

Humam AM, Loh TC, Foo HL, Izuddin WI, Awad EA, Idrus Z, et al.

Animals (Basel), 2020 Jun 05;10(6).
PMID: 32516896 DOI: 10.3390/ani10060982

The purpose of this work was to evaluate the impacts of feeding different postbiotics on oxidative stress markers, physiological stress indicators, lipid profile and meat quality in heat-stressed broilers. A total of 252 male Cobb 500 (22-day-old) were fed with 1 of 6 diets: A basal diet without any supplementation as negative control (NC); basal diet + 0.02% oxytetracycline served as positive control (PC); basal diet + 0.02% ascorbic acid (AA); or the basal diet diet + 0.3% of RI11, RS5 or UL4 postbiotics. Postbiotics supplementation, especially RI11 increased plasma activity of total-antioxidant capacity (T-AOC), catalase (CAT) and glutathione (GSH), and decreased alpha-1-acid-glycoprotein (α1-AGP) and ceruloplasmin (CPN) compared to NC and PC groups. Meat malondialdehyde (MDA) was lower in the postbiotic groups than the NC, PC and AA groups. Plasma corticosterone, heat shock protein70 (HSP70) and high density lipoprotein (HDL) were not affected by dietary treatments. Postbiotics decreased plasma cholesterol concentration compared to other groups, and plasma triglyceride and very low density lipoprotein (VLDL) compared to the NC group. Postbiotics increased breast meat pH, and decreased shear force and lightness (L*) compared to NC and PC groups. The drip loss, cooking loss and yellowness (b*) were lower in postbiotics groups compared to other groups. In conclusion, postbiotics particularly RI11 could be used as an alternative to antibiotics and natural sources of antioxidants for heat-stressed broilers.
Fulltext Inhibition of pathogenic and spoilage bacteria by a novel biofilm-forming Lactobacillus isolate: a potential host for the expression of heterologous proteins

Jalilsood T, Baradaran A, Song AA, Foo HL, Mustafa S, Saad WZ, et al.

Microb Cell Fact, 2015;14:96.
PMID: 26150120 DOI: 10.1186/s12934-015-0283-8

Bacterial biofilms are a preferred mode of growth for many types of microorganisms in their natural environments. The ability of pathogens to integrate within a biofilm is pivotal to their survival. The possibility of biofilm formation in Lactobacillus communities is also important in various industrial and medical settings. Lactobacilli can eliminate the colonization of different pathogenic microorganisms. Alternatively, new opportunities are now arising with the rapidly expanding potential of lactic acid bacteria biofilms as bio-control agents against food-borne pathogens.
Fulltext Effects of feeding different postbiotic metabolite combinations produced by Lactobacillus plantarum strains on egg quality and production performance, faecal parameters and plasma cholesterol in laying hens

Loh TC, Choe DW, Foo HL, Sazili AQ, Bejo MH

BMC Vet Res, 2014;10:149.
PMID: 24996258 DOI: 10.1186/1746-6148-10-149

Probiotics are beneficial bacteria that are able to colonize the host digestive system, increasing the natural flora and preventing colonization of pathogenic organisms and thus, securing optimal utility of the feed. However, commercial probiotic often do not meet the expected standards and the viability of the efficacy of these strains remains questionable. Another major issue has been highlighted in relation to the application of antibiotic resistant probiotics, the antibiotic resistant gene can be transferred between organisms. Recently, postbiotic metabolites produced from microbes have been extensively studied as feed additive in order to substitute in-feed antibiotics.
Fulltext Biodegradation of palm kernel cake by cellulolytic and hemicellulolytic bacterial cultures through solid state fermentation

Alshelmani MI, Loh TC, Foo HL, Lau WH, Sazili AQ

ScientificWorldJournal, 2014;2014:729852.
PMID: 25019097 DOI: 10.1155/2014/729852

Four cellulolytic and hemicellulolytic bacterial cultures were purchased from the Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Culture (DSMZ) and the American Type Culture Collection (ATCC). Two experiments were conducted; the objective of the first experiment was to determine the optimum time period required for solid state fermentation (SSF) of palm kernel cake (PKC), whereas the objective of the second experiment was to investigate the effect of combinations of these cellulolytic and hemicellulolytic bacteria on the nutritive quality of the PKC. In the first experiment, the SSF was lasted for 12 days with inoculum size of 10% (v/w) on different PKC to moisture ratios. In the second experiment, fifteen combinations were created among the four microbes with one untreated PKC as a control. The SSF lasted for 9 days, and the samples were autoclaved, dried, and analyzed for proximate analysis. Results showed that bacterial cultures produced high enzymes activities at the 4th day of SSF, whereas their abilities to produce enzymes tended to be decreased to reach zero at the 8th day of SSF. Findings in the second experiment showed that hemicellulose and cellulose was significantly (P < 0.05) decreased, whereas the amount of reducing sugars were significantly (P < 0.05) increased in the fermented PKC (FPKC) compared with untreated PKC.
Fulltext Characterization of cellulolytic bacterial cultures grown in different substrates

Alshelmani MI, Loh TC, Foo HL, Lau WH, Sazili AQ

ScientificWorldJournal, 2013;2013:689235.
PMID: 24319380 DOI: 10.1155/2013/689235

Nine aerobic cellulolytic bacterial cultures were obtained from the Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Culture (DSMZ) and the American Type Culture Collection (ATCC). The objectives of this study were to characterize the cellulolytic bacteria and to determine the optimum moisture ratio required for solid state fermentation (SSF) of palm kernel cake (PKC). The bacteria cultures were grown on reconstituted nutrient broth, incubated at 30°C and agitated at 200 rpm. Carboxymethyl cellulase, xylanase, and mannanase activities were determined using different substrates and after SSF of PKC. The SSF was conducted for 4 and 7 days with inoculum size of 10% (v/w) on different PKC concentration-to-moisture ratios: 1 : 0.2, 1 : 0.3, 1 : 0.4, and 1 : 0.5. Results showed that Bacillus amyloliquefaciens 1067 DSMZ, Bacillus megaterium 9885 ATCC, Paenibacillus curdlanolyticus 10248 DSMZ, and Paenibacillus polymyxa 842 ATCC produced higher enzyme activities as compared to other bacterial cultures grown on different substrates. The cultures mentioned above also produced higher enzyme activities when they were incubated under SSF using PKC as a substrate in different PKC-to-moisture ratios after 4 days of incubation, indicating that these cellulolytic bacteria can be used to degrade and improve the nutrient quality of PKC.
Cloning and in silico characterization of two signal peptides from Pediococcus pentosaceus and their function for the secretion of heterologous protein in Lactococcus lactis

Baradaran A, Sieo CC, Foo HL, Illias RM, Yusoff K, Rahim RA

Biotechnol Lett, 2013 Feb;35(2):233-8.
PMID: 23076361 DOI: 10.1007/s10529-012-1059-4

Fifty signal peptides of Pediococcus pentosaceus were characterized by in silico analysis and, based on the physicochemical analysis, (two potential signal peptides Spk1 and Spk3 were identified). The coding sequences of SP were amplified and fused to the gene coding for green fluorescent protein (GFP) and cloned into Lactococcus lactis pNZ8048 and pMG36e vectors, respectively. Western blot analysis indicated that the GFP proteins were secreted using both heterologous SPs. ELISA showed that the secretion efficiency of GFP using Spk1 (0.64 μg/ml) was similar to using Usp45 (0.62 μg/ml) and Spk3 (0.58 μg/ml).
Fulltext Live recombinant Lactococcus lactis vaccine expressing aerolysin genes D1 and D4 for protection against Aeromonas hydrophila in tilapia (Oreochromis niloticus)

Anuradha K, Foo HL, Mariana NS, Loh TC, Yusoff K, Hassan MD, et al.

J Appl Microbiol, 2010 Nov;109(5):1632-42.
PMID: 20602654 DOI: 10.1111/j.1365-2672.2010.04789.x

To evaluate a live recombinant Lactococcus lactis vaccine expressing aerolysin genes D1 (Lac-D1ae) and/or D4 (Lac-D4ae) in protection against Aeromonas hydrophila in tilapia (Oreochromis niloticus).
Fulltext Molecular characterisation of new organisation of plnEF and plw loci of bacteriocin genes harbour concomitantly in Lactobacillus plantarum I-UL4

Tai HF, Foo HL, Abdul Rahim R, Loh TC, Abdullah MP, Yoshinobu K

Microb Cell Fact, 2015;14:89.
PMID: 26077560 DOI: 10.1186/s12934-015-0280-y

Bacteriocin-producing Lactic acid bacteria (LAB) have vast applications in human and animal health, as well as in food industry. The structural, immunity, regulatory, export and modification genes are required for effective bacteriocin biosynthesis. Variations in gene sequence, composition and organisation will affect the antimicrobial spectrum of bacteriocin greatly. Lactobacillus plantarum I-UL4 is a novel multiple bacteriocin producer that harbours both plw and plnEF structural genes simultaneous which has not been reported elsewhere. Therefore, molecular characterisation of bacteriocin genes that harboured in L. plantarum I-UL4 was conducted in this study.
Egg production, faecal pH and microbial population, small intestine morphology, and plasma and yolk cholesterol in laying hens given liquid metabolites produced by Lactobacillus plantarum strains

Choe DW, Loh TC, Foo HL, Hair-Bejo M, Awis QS

Br Poult Sci, 2012;53(1):106-15.
PMID: 22404811 DOI: 10.1080/00071668.2012.659653

1. Various dosages of metabolite combinations of the Lactobacillus plantarum RI11, RG14 and RG11 strains (COM456) were used to study the egg production, faecal microflora population, faecal pH, small intestine morphology, and plasma and egg yolk cholesterol in laying hens. 2. A total of 500 Lohmann Brown hens were raised from 19 weeks to 31 weeks of age. The birds were randomly divided into 5 groups and fed on various treatment diets: (i) basal diet without supplementation of metabolites (control); (ii) basal diet supplemented with 0·3% COM456 metabolites; (iii) basal diet supplemented with 0·6% COM456 metabolites; (iv) basal diet supplemented with 0·9% COM456 metabolites; and (v) basal diet supplemented with 1·2% COM456 metabolites. 3. The inclusion of 0·6% liquid metabolite combinations, produced from three L. plantarum strains, demonstrated the best effect in improving the hens' egg production, faecal lactic acid bacteria population, and small intestine villus height, and reducing faecal pH and Enterobacteriaceae population, and plasma and yolk cholesterol concentrations. 4. The metabolites from locally isolated L. plantarum are a possible alternative feed additive in poultry production.
Effects of feeding metabolite combinations produced by Lactobacillus plantarum on growth performance, faecal microbial population, small intestine villus height and faecal volatile fatty acids in broilers

Thanh NT, Loh TC, Foo HL, Hair-Bejo M, Azhar BK

Br Poult Sci, 2009 May;50(3):298-306.
PMID: 19637029 DOI: 10.1080/00071660902873947

1. Four combinations of metabolites produced from strains of Lactobacillus plantarum were used to study the performance of broiler chickens. 2. A total of 432 male Ross broilers were raised from one-day-old to 42 d of age in deep litter pens (12 birds/pen). These birds were divided into 6 groups and fed on different diets: (i) standard maize-soybean-based diet (negative control); (ii) standard maize-soybean-based diet + Neomycin and Oxytetracycline (positive control); (iii) standard maize-soybean-based diet + 0.3% metabolite combination of Lactobacillus plantarum RS5, RI11, RG14 and RG11 strains (com3456); (iv) standard maize-soybean-based diet + 0.3% metabolite combination of L. plantarum TL1, RI11 and RG11 (Com246); (v) standard maize-soybean-based diet + 0.3% metabolite combination of L. plantarum TL1, RG14 and RG11 (Com256) and (vi) standard maize-soybean-based diet + 0.3% metabolite combination of L. plantarum TL1, RS5, RG14 and RG11 (Com2356). 3. Higher final body weight, weight gain, average daily gain and lower feed conversion ratio were found in all 4 treated groups. 4. The addition of a metabolite combination supplementation also increased faecal lactic acid bacteria population, small intestine villus height and faecal volatile fatty acids and faecal Enterobacteriaceae population.
Fulltext Effects of postbiotic supplementation on growth performance, ruminal fermentation and microbial profile, blood metabolite and GHR, IGF-1 and MCT-1 gene expression in post-weaning lambs

Izuddin WI, Loh TC, Samsudin AA, Foo HL, Humam AM, Shazali N

BMC Vet Res, 2019 Sep 02;15(1):315.
PMID: 31477098 DOI: 10.1186/s12917-019-2064-9

BACKGROUND: Postbiotics have been established as potential feed additive to be used in monogastric such as poultry and swine to enhance health and growth performance. However, information on the postbiotics as feed additive in ruminants is very limited. The aim of this study was to elucidate the effects of supplementation of postbiotics in newly-weaned lambs on growth performance, digestibility, rumen fermentation characteristics and microbial population, blood metabolite and expression of genes related to growth and volatile fatty acid transport across the rumen epithelium.
RESULTS: Postbiotic supplementation increased weight gain, feed intake, nutrient intake and nutrient digestibility of the lambs. No effect on ruminal pH and total VFA, whereas butyrate and ruminal ammonia-N concentration were improved. The lambs fed with postbiotics had higher blood total protein, urea nitrogen and glucose. However, no difference was observed in blood triglycerides and cholesterol levels. Postbiotics increased the population of fibre degrading bacteria but decreased total protozoa and methanogens in rumen. Postbiotics increased the mRNA expression of hepatic IGF-1 and ruminal MCT-1.
CONCLUSIONS: The inclusion of postbiotics from L. plantarum RG14 in newly-weaned lambs improved growth performance, nutrient intake and nutrient digestibility reflected from better rumen fermentation and microbial parameters, blood metabolites and upregulation of growth and nutrient intake genes in the post-weaning lambs.
Fulltext Enhancement of Versatile Extracellular Cellulolytic and Hemicellulolytic Enzyme Productions by Lactobacillus plantarum RI 11 Isolated from Malaysian Food Using Renewable Natural Polymers

Mohamad Zabidi NA, Foo HL, Loh TC, Mohamad R, Abdul Rahim R

Molecules, 2020 Jun 03;25(11).
PMID: 32503356 DOI: 10.3390/molecules25112607

Lactobacillus plantarum RI 11 was reported recently to be a potential lignocellulosic biomass degrader since it has the capability of producing versatile extracellular cellulolytic and hemicellulolytic enzymes. Thus, this study was conducted to evaluate further the effects of various renewable natural polymers on the growth and production of extracellular cellulolytic and hemicellulolytic enzymes by this novel isolate. Basal medium supplemented with molasses and yeast extract produced the highest cell biomass (log 10.51 CFU/mL) and extracellular endoglucanase (11.70 µg/min/mg), exoglucanase (9.99 µg/min/mg), β-glucosidase (10.43 nmol/min/mg), and mannanase (8.03 µg/min/mg), respectively. Subsequently, a statistical optimization approach was employed for the enhancement of cell biomass, and cellulolytic and hemicellulolytic enzyme productions. Basal medium that supplemented with glucose, molasses and soybean pulp (F5 medium) or with rice straw, yeast extract and soybean pulp (F6 medium) produced the highest cell population of log 11.76 CFU/mL, respectively. However, formulated F12 medium supplemented with glucose, molasses and palm kernel cake enhanced extracellular endoglucanase (4 folds), exoglucanase (2.6 folds) and mannanase (2.6 folds) specific activities significantly, indicating that the F12 medium could induce the highest production of extracellular cellulolytic and hemicellulolytic enzymes concomitantly. In conclusion, L. plantarum RI 11 is a promising and versatile bio-transformation agent for lignocellulolytic biomass.
Fulltext Growth Performance, Cytokine Expression, and Immune Responses of Broiler Chickens Fed a Dietary Palm Oil and Sunflower Oil Blend Supplemented With L-Arginine and Varying Concentrations of Vitamin E

Khatun J, Loh TC, Foo HL, Akit H, Khan KI

Front Vet Sci, 2020;7:619.
PMID: 33195499 DOI: 10.3389/fvets.2020.00619

This study set out to examine the combined effects of the supplementation of a dietary palm oil (PO) and sunflower oil (SO) blend, 0. 25% L-Arginine (L-Arg), and different levels of vitamin E (Vit E) on growth performance, fat deposition, cytokine expression, and immune response in broilers. A total of 216 1-day-old male broiler chicks (Cobb500) were randomly distributed into six dietary groups as follows: Diet 1: 6% palm oil (negative control); Diet 2: PO and SO blend (4% palm oil and 2% sunflower oil) + 0.25% L-Arg (positive control); Diet 3: (PO and SO blend + 0.25% L-Arg) + 20 mg/kg Vit E; Diet 4: (PO and SO blend + 0.25% L-Arg) + 50 mg/kg Vit E; Diet 5: (PO and SO blend + 0.25% L-Arg) + 100 mg/kg Vit E; and Diet 6: (PO and SO blend + 0.25% L-Arg) + 150 mg/kg Vit E. Weight gain and serum IgG and IgM increased while feed conversion ratio, fat deposition, and plasma cholesterol decreased in broilers fed Vit E with the oil blend and L-Arg, compared to those fed the negative control (Diet 1). Expression of IFN and TNF-α were reduced, whereas TGF-ß1 was up-regulated as the level of Vit E increased in the broiler diets. In summary, the combination of oil blend, L-Arg, and Vit E at a level of 50 mg/kg increased the performance and altered the expression of cytokines that may positively influence immune function in broiler chickens.
Fulltext Supplementation of postbiotic RI11 improves antioxidant enzyme activity, upregulated gut barrier genes, and reduced cytokine, acute phase protein, and heat shock protein 70 gene expression levels in heat-stressed broilers

Humam AM, Loh TC, Foo HL, Izuddin WI, Zulkifli I, Samsudin AA, et al.

Poult Sci, 2021 Mar;100(3):100908.
PMID: 33518339 DOI: 10.1016/j.psj.2020.12.011

The aim of this work was to evaluate the impacts of feeding different levels of postbiotic RI11 on antioxidant enzyme activity, physiological stress indicators, and cytokine and gut barrier gene expression in broilers under heat stress. A total of 252 male broilers Cobb 500 were allocated in cages in environmentally controlled chambers. All the broilers received the same basal diet from 1 to 21 d. On day 22, the broilers were weighed and grouped into 7 treatment groups and exhibited to cyclic high temperature at 36 ± 1°C for 3 h per day until the end of the experiment. From day 22 to 42, broilers were fed with one of the 7 following diets: negative control, basal diet (0.0% RI11) (NC group); positive control, NC diet + 0.02% (w/w) oxytetracycline (OTC group); antioxidant control, NC diet + 0.02% (w/w) ascorbic acid. The other 4 other groups were as follows: NC diet + 0.2% cell-free supernatant (postbiotic RI11) (v/w), NC diet + 0.4% cell-free supernatant (postbiotic RI11) (v/w), NC diet + 0.6% cell-free supernatant (postbiotic RI11) (v/w), and NC diet + 0.8% cell-free supernatant (postbiotic RI11) (v/w). Supplementation of different levels (0.4, 0.6, and 0.8%) of postbiotic RI11 increased plasma glutathione peroxidase, catalase, and glutathione enzyme activity. Postbiotic RI11 groups particularly at levels of 0.4 and 0.6% upregulated the mRNA expression of IL-10 and downregulated the IL-8, tumor necrosis factor alpha, heat shock protein 70, and alpha-1-acid glycoprotein levels compared with the NC and OTC groups. Feeding postbiotic RI11, particularly at the level of 0.6%, upregulated ileum zonula occludens-1 and mucin 2 mRNA expressions. However, no difference was observed in ileum claudin 1, ceruloplasmin, IL-6, IL-2, and interferon expression, but downregulation of occludin expression was observed as compared with the NC group. Supplementation of postbiotic RI11 at different levels quadratically increased plasma glutathione peroxidase, catalase and glutathione, IL-10, mucin 2, and zonula occludens-1 mRNA expression and reduced plasma IL-8, tumor necrosis factor alpha, alpha-1-acid glycoprotein, and heat shock protein 70 mRNA expression. The results suggested that postbiotics produced from Lactiplantibacillus plantarum RI11 especially at the level of 0.6% (v/w) could be used as an alternative to antibiotics and natural sources of antioxidants in poultry feeding.
Feeding of different levels of metabolite combinations produced by Lactobacillus plantarum on growth performance, fecal microflora, volatile fatty acids and villi height in broilers

Loh TC, Thanh NT, Foo HL, Hair-Bejo M, Azhar BK

Anim Sci J, 2010 Apr;81(2):205-14.
PMID: 20438502 DOI: 10.1111/j.1740-0929.2009.00701.x

The effects of feeding different dosages of metabolite combination of L. plantarum RS5, RI11, RG14 and RG11 strains (Com3456) on the performance of broiler chickens was studied. A total of 504 male Ross broilers were grouped into 7 treatments and offered different diets: (i) standard corn-soybean based diet (negative control); (ii) standard corn-soybean based diet +100 ppm neomycin and oxytetracycline (positive control); (iii) standard corn-soybean based diet + 0.1% metabolite combination of L. plantarum RS5, RI11, RG14 and RG11 strains (Com3456); (iv) standard corn-soybean based diet + 0.2% of Com3456; (v) standard corn-soybean based diet + 0.3% of Com3456 (vi) standard corn-soybean based diet + 0.4% of Com3456 and (vii) standard corn-soybean based diet + 0.5% of Com3456. Supplementation of Com3456 with different dosages improved growth performance, reduced Enterobacteriaceae and increased lactic acid bacteria count, and increased villi height of small intestine and fecal volatile fatty acid concentration. Treatment with 0.4% and 0.2% Com3456 had the best results, especially in terms of growth performance, feed conversion ratio and villi height among other dosages. However, the dosage of 0.2% was recommended due to its lower concentration yielding a similar effect as 0.4% supplementation. These results indicate that 0.2% is an optimum level to be included in the diets of broiler in order to replace antibiotic growth promoters.
Effects of feeding fermented fish on egg cholesterol content in hens

Loh TC, Law FL, Goh YM, Foo HL, Zulkifli I

Anim Sci J, 2009 Feb;80(1):27-33.
PMID: 20163464 DOI: 10.1111/j.1740-0929.2008.00591.x

This study was conducted to investigate the effects of feeding fermented fish (FF) to layers on laying performance, and polyunsaturated fatty acid and cholesterol levels in eggs and plasma. A total of 96, 13-week-old Babcock B380 pullets were used in this study. They were randomly assigned to four numerically equal groups with eight replicates per treatment, three birds per replicate. All the birds were housed in individual cages. The dietary treatments were: Control diet, without FF; FF3 diet containing 3% (w/w) FF, FF6 diet containing 6% (w/w) FF and FF9 diet containing 9% (w/w) FF. The study was carried out for 16 weeks inclusive of two weeks of adjustment. Weekly feed intake and egg production were recorded. Blood plasma cholesterol and fatty acid profiles were assayed at the end of the experiment. FF did not enhance (P > 0.05) egg mass but (P < 0.05) decreased egg weight slightly. However, egg yolk cholesterol and plasma cholesterol concentrations were reduced (P < 0.05) by FF. The n-6:n-3 fatty acids ratio in the egg yolk (Control = 7.9, FF9 = 6.2) and plasma (Control = 10.6, FF9 = 6.2) were decreased by feeding FF. Moreover, FF was able to increase (P < 0.05) the docosahexaenoic acid (DHA) concentrations in egg yolk and plasma. In conclusion, this study demonstrated that FF increased DHA and reduced egg yolk cholesterol in poultry eggs.
Fulltext Promising discovery of beneficial Escherichia coli in the human gut

Nakkarach A, Foo HL, Song AA, Nitisinprasert S, Withayagiat U

3 Biotech, 2020 Jul;10(7):296.
PMID: 32550113 DOI: 10.1007/s13205-020-02289-z

Ingested dietary fibres are hydrolysed by colon microbiota to produce energy-providing short-chain fatty acids (SCFA) that stimulate anti-inflammatory effects. SCFA-producing bacteria were screened from bacteria isolated from human faeces using bromothymol blue as an acid indicator and gas chromatography for SCFA profiling. The beneficial functions (antagonistic activity, haemolytic activities, antibiotic susceptibility, mucus adherent percentage and toxin gene detection) were evaluated for the top five SCFA-producing bacteria isolated from three healthy volunteers that identified as Escherichia coli strains. They produced acetic, propionic, isobutyric, butyric, isovaleric, valeric and caproic acids at average concentrations of 15.9, 1.8, 1.1, 1.9, 1.8, 2.7 and 3.4 mM, respectively. The SCFA production by E. coli strains was rapidly increased during the first 8 h of incubation and gradually decreased after 16 h of incubation. All E. coli strains showed acid and bile tolerance, resulting in a survival rate greater than 70% with no haemolytic activity, mucus adherence greater than 40% and susceptibility to conventional antibiotics. Hence, the selected E. coli strains exhibited promising probiotic properties with neither enterotoxin nor LPS producibility was detected. The present results confirm the existence of friendly and harmless E. coli strains in human microbiota as potential probiotics.
Fulltext Anti-cancer and anti-inflammatory effects elicited by short chain fatty acids produced by Escherichia coli isolated from healthy human gut microbiota

Nakkarach A, Foo HL, Song AA, Mutalib NEA, Nitisinprasert S, Withayagiat U

Microb Cell Fact, 2021 Feb 05;20(1):36.
PMID: 33546705 DOI: 10.1186/s12934-020-01477-z

BACKGROUND: Extracellular metabolites of short chain fatty acids (SCFA) excreted by gut microbiota have been reported to play an important role in the regulation of intestinal homeostasis. Apart from supplying energy, SCFA also elicit immune stimulation in animal and human cells. Therefore, an attempt was conducted to isolate SCFA producing bacteria from healthy human microbiota. The anti-cancer and anti-inflammatory effects of extracellular metabolites and individual SFCA were further investigated by using breast, colon cancer and macrophage cells. Toxin, inflammatory and anti-inflammatory cytokine gene expressions were investigated by RT-qPCR analyses in this study.
RESULTS: Escherichia coli KUB-36 was selected in this study since it has the capability to produce seven SCFA extracellularly. It produced acetic acid as the main SCFA. It is a non-exotoxin producer and hence, it is a safe gut microbiota. The IC50 values indicated that the E. coli KUB-36 metabolites treatment elicited more potent cytotoxicity effect on MCF7 breast cancer cell as compared to colon cancer and leukemia cancer cells but exhibited little cytotoxic effects on normal breast cell. Furthermore, E. coli KUB-36 metabolites and individual SCFA could affect inflammatory responses in lipopolysaccharide-induced THP-1 macrophage cells since they suppressed inflammatory cytokines IL-1β, IL-6, IL-8 and TNF-α well as compared to the control, whilst inducing anti-inflammatory cytokine IL-10 expression.
CONCLUSION: SCFA producing E. coli KUB-36 possessed vast potential as a beneficial gut microbe since it is a non-exotoxin producer that exhibited beneficial cytotoxic effects on cancer cells and elicited anti-inflammatory activity simultaneously. However, the probiotic characteristic of E. coli KUB-36 should be further elucidated using in vivo animal models.

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