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  1. Fulltext Effect of cathepsin D and prostate specific antigen on latent transforming growth factor-beta in breast cancer cell lines
    Wong SF, Seow HF, Lai LC
    Malays J Pathol, 2003 Dec;25(2):129-34.
    PMID: 16196369
    Transforming growth factor-beta (TGFbeta) is present, predominantly in latent forms, in normal and malignant breast tissue. The mechanisms by which latent TGFbeta is activated physiologically remain largely an enigma. The objective of this study was to assess whether the proteases, cathepsin D and prostate specific antigen (PSA) could activate latent TGFbeta1 and TGFbeta2 in conditioned media of the hormone-dependent MCF-7 and hormone-independent MDA-MB-231 human breast cancer cell lines, newly purchased from ATCC. Both of the cell lines were seeded in 6-well plates 2 days prior to treatment with varying concentrations of cathepsin D and PSA. Active TGFbeta1 and TGFbeta2 in the media were then measured by ELISA after 4, 8, 24 and 72 hours of treatment. TGFbeta1 and TGFbeta2 mRNA expression of both cell lines were measured by RT-PCR to determine whether any increase in level of active TGFbeta1 and TGFbeta2 was due to increased production. There was a significant increase in only active TGFbeta2 levels in the MDA-MB-231 cell line with both treatments. Cathepsin D and PSA did not have any effect on TGFbeta1 and TGFbeta2 mRNA expression. Cathepsin D and PSA were unable to activate latent TGFbeta1 and TGFbeta2 in these two breast cancer cell lines. A constant level of TGFbeta2 mRNA in the control and treated MDA-MB-231 cells suggests that the increase in level of active TGFbeta2 was not a result of increased production but was likely to be due to activation by a mechanism independent of cathepsin D and PSA.
    Matched MeSH terms: Culture Media, Conditioned/chemistry
  2. Fulltext Generation of dendritic cells from acute myeloid leukaemia cells and monocytes: our local experience
    Lim MN, Leong CF, Cheong SK, Seow HF
    Malays J Pathol, 2003 Dec;25(2):107-12.
    PMID: 16196366
    Dendritic cells (DC) are efficient and potent antigen-presenting cells. Pilot clinical trials indicated that DC loaded with tumour antigen could induce tumour-specific immune responses in various cancers including B-cell lymphoma, melanoma and prostate cancer. Owing to extensively low number of DC in the blood circulation, a variety of sources have been used to generate DC including monocytes, CD34+ stem cells and even with leukaemic blast cells. We demonstrate here a simple method to generate DC from acute myeloid leukaemia (AML) cells and monocytes from healthy donor or remission samples. AML cells or monocytes were cultured in RPMI 1640 media supplemented with foetal bovine serum or autologous serum where possible and different combinations of cytokines GM-CSF, IL-4 and TNF-alpha. The generated DC were evaluated for their morphology by phase contrast microscopy and May Grunwald Giemsa staining. Viability of cells was determined by trypan blue dye exclusion. Percentage of yields and immunophenotypes were carried out by flow cytometry. We found that cultured AML cells and monocytes developed morphological and immuno-phenotypic characteristics of DC. Monocytes are better than AML blast in generating DC and serve as a ready source for dendritic cell vaccine development.
    Matched MeSH terms: Cell Culture Techniques/methods*
  3. Cultural and dietary risk factors of oral cancer and precancer--a brief overview
    Zain RB
    Oral Oncol, 2001 Apr;37(3):205-10.
    PMID: 11287272
    This is an update on cultural and dietary risk factors for oral precancer and cancer. It is an overview on ethnic differences (where possible) and socio-cultural risk factors (tobacco/areca nut/betel quid, alcohol use and dietary factors) in relation to oral precancer and cancer. While studies were from Western countries, India and China, this update also attempts to include and highlight some studies conducted in the Asia-Pacific region.
    Matched MeSH terms: Culture*
  4. Disease, disorder, or deception? Latah as habit in a Malay extended family
    Bartholomew RE
    J. Nerv. Ment. Dis., 1994 Jun;182(6):331-8; discussion 339-41.
    PMID: 8201305 DOI: 10.1097/00005053-199406000-00004
    Thirty-seven cases of latah are examined within the author's Malay extended family (N = 115). Based on ethnographic data collected and a literature review, cases are readily divisible into two broad categories: habitual (N = 33) and performance (N = 4). The first form represents an infrequent, culturally conditioned habit that is occasionally used as a learned coping strategy in the form of a cathartic stress response to sudden startle with limited secondary benefits (i.e., exhibiting brief verbal obscenity with impunity). In this sense, it is identical to Western swearing. Performers are engaged in conscious, ritualized social gain through the purported exploitation of a neurophysiological potential. The latter process is essentially irrelevant, akin to sneezing or yawning. It is concluded that latah is a social construction of Western-trained universalist scientists. The concept of malingering and fraud in anthropology is critically discussed.
    Matched MeSH terms: Culture*
  5. Comparison of culture-independent and dependent approaches for identification of native arsenic-resistant bacteria and their potential use for arsenic bioremediation
    Hamood Altowayti WA, Almoalemi H, Shahir S, Othman N
    Ecotoxicol Environ Saf, 2020 Dec 01;205:111267.
    PMID: 32992213 DOI: 10.1016/j.ecoenv.2020.111267
    Arsenic is a common contaminant in gold mine soil and tailings. Microbes present an opportunity for bio-treatment of arsenic, since it is a sustainable and cost-effective approach to remove arsenic from water. However, the development of existing bio-treatment approaches depends on isolation of arsenic-resistant microbes from arsenic contaminated samples. Microbial cultures are commonly used in bio-treatment; however, it is not established whether the structure of the cultured isolates resembles the native microbial community from arsenic-contaminated soil. In this milieu, a culture-independent approach using Illumina sequencing technology was used to profile the microbial community in situ. This was coupled with a culture-dependent technique, that is, isolation using two different growth media, to analyse the microbial population in arsenic laden tailing dam sludge based on the culture-independent sequencing approach, 4 phyla and 8 genera were identified in a sample from the arsenic-rich gold mine. Firmicutes (92.23%) was the dominant phylum, followed by Proteobacteria (3.21%), Actinobacteria (2.41%), and Bacteroidetes (1.49%). The identified genera included Staphylococcus (89.8%), Pseudomonas (1.25), Corynebacterium (0.82), Prevotella (0.54%), Megamonas (0.38%) and Sphingomonas (0.36%). The Shannon index value (3.05) and Simpson index value (0.1661) indicated low diversity in arsenic laden tailing. The culture dependent method exposed significant similarities with culture independent methods at the phylum level with Firmicutes, Proteobacteria and Actinobacteria, being common, and Firmicutes was the dominant phylum whereas, at the genus level, only Pseudomonas was presented by both methods. It showed high similarities between culture independent and dependent methods at the phylum level and large differences at the genus level, highlighting the complementarity between the two methods for identification of the native population bacteria in arsenic-rich mine. As a result, the present study can be a resource on microbes for bio-treatment of arsenic in mining waste.
    Matched MeSH terms: Culture Media/chemistry
  6. Characterization of a type I pullulanase from Anoxybacillus sp. SK3-4 reveals an unusual substrate hydrolysis
    Kahar UM, Ng CL, Chan KG, Goh KM
    Appl Microbiol Biotechnol, 2016 Jul;100(14):6291-307.
    PMID: 27000839 DOI: 10.1007/s00253-016-7451-6
    Type I pullulanases are enzymes that specifically hydrolyse α-1,6 linkages in polysaccharides. This study reports the analyses of a novel type I pullulanase (PulASK) from Anoxybacillus sp. SK3-4. Purified PulASK (molecular mass of 80 kDa) was stable at pH 5.0-6.0 and was most active at pH 6.0. The optimum temperature for PulASK was 60 °C, and the enzyme was reasonably stable at this temperature. Pullulan was the preferred substrate for PulASK, with 89.90 % adsorbance efficiency (various other starches, 56.26-72.93 % efficiency). Similar to other type I pullulanases, maltotriose was formed on digestion of pullulan by PulASK. PulASK also reacted with β-limit dextrin, a sugar rich in short branches, and formed maltotriose, maltotetraose and maltopentaose. Nevertheless, PulASK was found to preferably debranch long branches at α-1,6 glycosidic bonds of starch, producing amylose, linear or branched oligosaccharides, but was nonreactive against short branches; thus, no reducing sugars were detected. This is surprising as all currently known type I pullulanases produce reducing sugars (predominantly maltotriose) on digesting starch. The closest homologue of PulASK (95 % identity) is a type I pullulanase from Anoxybacillus sp. LM14-2 (Pul-LM14-2), which is capable of forming reducing sugars from starch. With rational design, amino acids 362-370 of PulASK were replaced with the corresponding sequence of Pul-LM14-2. The mutant enzyme formed reducing sugars on digesting starch. Thus, we identified a novel motif involved in substrate specificity in type I pullulanases. Our characterization may pave the way for the industrial application of this unique enzyme.
    Matched MeSH terms: Culture Media/chemistry*
  7. In vitro generation of functional insulin-producing cells from lipoaspirated human adipose tissue-derived stem cells
    Mohamad Buang ML, Seng HK, Chung LH, Saim AB, Idrus RB
    Arch Med Res, 2012 Jan;43(1):83-8.
    PMID: 22374243 DOI: 10.1016/j.arcmed.2012.01.012
    BACKGROUND AND AIMS: Tissue engineering strategy has been considered as an alternative treatment for diabetes mellitus due to lack of permanent pharmaceutical treatment and islet donors for transplantation. Various cell lines have been used to generate functional insulin-producing cells (IPCs) including progenitor pancreatic cell lines, embryonic stem cells (ESCs), umbilical cord blood stem cells (UCB-SCs), adult bone marrow stem cells (BMSCs), and adipose tissue-derived stem cells (ADSCs).

    METHODS: Human ADSCs from lipoaspirated abdominal fat tissue was differentiated into IPCs following a two-step induction protocol based on a combination of alternating high and low glucose, nicotinamide, activin A and glucagon-like peptide 1 (GLP-1) for a duration of 3 weeks. During differentiation, histomorphological changes of the stem cells towards pancreatic β-islet characteristics were observed via light microscope and transmission electron microscope (TEM). Dithizone (DTZ) staining, which is selective towards IPCs, was used to stain the new islet-like cells. Production of insulin hormone by the cells was analyzed via enzyme-linked immunosorbent assay (ELISA), whereas its hormonal regulation was tested via a glucose challenge test.

    RESULTS: Histomorphological changes of the differentiated cells were noted to resemble pancreatic β-cells, whereas DTZ staining positively stained the cells. The differentiated cells significantly produced human insulin as compared to the undifferentiated ADSCs, and its production was increased with an increase of glucose concentration in the culture medium.

    CONCLUSIONS: These initial data indicate that human lipoaspirated ADSCs have the potential to differentiate into functional IPCs, and could be used as a therapy to treat diabetes mellitus in the future.

    Matched MeSH terms: Culture Media; Cell Culture Techniques
  8. Evaluation of Tualang honey as a supplement to fetal bovine serum in cell culture
    Kannan TP, Ali AQ, Abdullah SF, Ahmad A
    Food Chem Toxicol, 2009 Jul;47(7):1696-702.
    PMID: 19394390 DOI: 10.1016/j.fct.2009.04.020
    The aim of this study was to evaluate Tualang honey as a supplement to fetal bovine serum in cell cultures using MTT assay, chromosome aberration test and gene expression analyses. The MTT assay showed the highest percentage of cell proliferation (105.3% increment than control) of human osteoblast cell line (CRL 1543) in 0.0195% honey in Dulbecco's modified eagle medium supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. There was enhanced cell proliferation corresponding to the decrease in concentrations of honey as indicated by the mitotic index values when the osteoblast cell line was incubated at 37 degrees C for 48 hours. There were no chromosome aberrations both in the honey treated as well as distilled water treated (negative control) cell lines. In the case of gene expression analyses, fibroblast cell lines (CCL 171) were treated with honey (0.0195%) for 24 and 48 hours separately. Though there was over expression for the bcl-xl gene at both 24 and 48 hours, under expression for bcl-xs gene at 24 hours and over expression at 48 hours and under expression for both c-myc and p53 genes at both 24 and 48 hours, none of them were statistically significant in altering the expression of mRNA.
    Matched MeSH terms: Culture Media/chemistry*
  9. Fulltext Detection of secreted antimicrobial peptides isolated from cell-free culture supernatant of Paenibacillus alvei AN5
    Alkotaini B, Anuar N, Kadhum AA, Sani AA
    J Ind Microbiol Biotechnol, 2013 Jun;40(6):571-9.
    PMID: 23508455 DOI: 10.1007/s10295-013-1259-5
    An antimicrobial substance produced by the Paenibacillus alvei strain AN5 was detected in fermentation broth. Subsequently, cell-free culture supernatant (CFCS) was obtained by medium centrifugation and filtration, and its antimicrobial activity was tested. This showed a broad inhibitory spectrum against both Gram-positive and -negative bacterial strains. The CFCS was then purified and subjected to SDS-PAGE and infrared spectroscopy, which indicated the proteinaceous nature of the antimicrobial compound. Some de novo sequencing using an automatic Q-TOF premier system determined the amino acid sequence of the purified antimicrobial peptide as Y-S-K-S-L-P-L-S-V-L-N-P (1,316 Da). The novel peptide was designated as peptide AN5-1. Its mode of action was bactericidal, inducing cell lysis in E. coli ATCC 29522 and S. aureus, and non-cell lysis in both S. marcescens and B. cereus ATCC 14579. Peptide AN5-1 displayed stability at a wide range of pH values (2-12) and remained active after exposure to high temperatures (100 °C). It also maintained its antimicrobial activity after incubation with chemicals such as SDS, urea and EDTA.
    Matched MeSH terms: Culture Media, Conditioned/chemistry*
  10. Fulltext Collagen type I promotes osteogenic differentiation of amniotic membrane-derived mesenchymal stromal cells in basal and induction media
    Akhir HM, Teoh PL
    Biosci Rep, 2020 12 23;40(12).
    PMID: 33245097 DOI: 10.1042/BSR20201325
    Collagen has been widely shown to promote osteogenesis of bone marrow mesenchymal stromal cells (BM-MSCs). Due to the invasive procedure of obtaining BM-MSCs, MSCs from other tissues have emerged as a promising alternative for regenerative therapy. MSCs originated from different sources, exhibiting different differentiation potentials. Therefore, the applicability of collagen type I (COL), combining with amniotic membrane (AM)-MSCs was examined through proliferation and differentiation assays together with the expression of surface markers and genes associated with stemness and differentiation under basal or induction conditions. No increase in cell growth was observed because AM-MSCs might be directed toward spontaneous osteogenesis. This was evidenced by the calcium deposition and elevated expression of osteogenic genes when AM-MSCs were cultured in collagen plate with basal media. Under the osteogenic condition, reciprocal expression of OCN and CEBPA suggested a shift toward adipogenesis. Surprisingly, adipogenic genes were not elevated upon adipogenic induction, although oil droplets deposition was observed. In conclusion, our findings demonstrated that collagen causes spontaneous osteogenesis in AM-MSCs. However, the presence of exogenous inductors could shift the direction of adipo-osteogenic gene regulatory network modulated by collagen.
    Matched MeSH terms: Culture Media/metabolism*
  11. Fulltext Biosafety evaluation of culture-expanded human chondrocytes with growth factor cocktail: a preclinical study
    Al-Masawa ME, Wan Kamarul Zaman WS, Chua KH
    Sci Rep, 2020 12 09;10(1):21583.
    PMID: 33299022 DOI: 10.1038/s41598-020-78395-y
    The scarcity of chondrocytes is a major challenge for cartilage tissue engineering. Monolayer expansion is necessary to amplify the limited number of chondrocytes needed for clinical application. Growth factors are often added to improve monolayer culture conditions, promoting proliferation, and enhancing chondrogenesis. Limited knowledge on the biosafety of the cell products manipulated with growth factors in culture has driven this study to evaluate the impact of growth factor cocktail supplements in chondrocyte culture medium on chondrocyte genetic stability and tumorigenicity. The growth factors were basic fibroblast growth factor (b-FGF), transforming growth factor β2 (TGF β2), insulin-like growth factor 1 (IGF-1), insulin-transferrin-selenium (ITS), and platelet-derived growth factor (PD-GF). Nasal septal chondrocytes cultured in growth factor cocktail exhibited a significantly high proliferative capacity. Comet assay revealed no significant DNA damage. Flow cytometry showed chondrocytes were mostly at G0-G1 phase, exhibiting normal cell cycle profile with no aneuploidy. We observed a decreased tumour suppressor genes' expression (p53, p21, pRB) and no TP53 mutations or tumour formation after 6 months of implantation in nude mice. Our data suggest growth factor cocktail has a low risk of inducing genotoxic and tumorigenic effects on chondrocytes up to passage 6 with 16.6 population doublings. This preclinical tumorigenicity and genetic instability evaluation is crucial for further clinical works.
    Matched MeSH terms: Culture Media/pharmacology*
  12. Fulltext Long term treatment by mesenchymal stem cells conditioned medium modulates cellular, molecular and behavioral aspects of adjuvant-induced arthritis
    Nazemian V, Manaheji H, Sharifi AM, Zaringhalam J
    Cell Mol Biol (Noisy-le-grand), 2018 Jan 31;64(1):19-26.
    PMID: 29412789 DOI: 10.14715/cmb/2018.64.2.5
    Neuroinflammation plays a crucial role in expression of symptoms of numerous autoimmune and neurodegenerative diseases such as pain during rheumatoid arthritis. Overproduction of pro-inflammatory cytokines and activation of intracellular signaling pathways have been strongly implicated in the generation of pathological pain states, particularly at central nervous system sites and induction of spinal neuroinflammatory symptoms. The wide ranges of research to define new therapeutic approaches, including neuroimmune-modulators like stem cells are in progress. Mesenchymal stem cells conditioned medium (MSC-CM) has anti-inflammatory factors which can regulate the immune responses. The aim of this study was to investigate the effect of administration of MSC-CM on behavioral, cellular and molecular aspects of adjuvant-induced arthritis in male Wistar rats. Complete Freund's adjuvant (CFA)-induced arthritis (AA) was caused by single subcutaneous injection of CFA into the rat's hind paw on day 0. MSC-CM was administered daily (i.p.) and during the 21 days of the study after injection. Hyperalgesia, Edema, Serum TNF-α levels and p38MAPK and NF-κB activities were assessed on days 0,7,14 and 21 of the study. The results of this study indicated the role of MSC-CM in reducing inflammatory symptoms, serum TNF-α levels and activity of intracellular signaling pathway factors during different phases of inflammation caused by CFA. It seems that MSC-CM treatment due to its direct effects on inhibition of intracellular signaling pathways and pro-inflammatory cytokines can alleviate inflammatory symptoms and pain during CFA-induced arthritis.
    Matched MeSH terms: Culture Media, Conditioned/pharmacology
  13. Polymicrobial biofilm formation by Candida albicans, Actinomyces naeslundii, and Streptococcus mutans is Candida albicans strain and medium dependent
    Arzmi MH, Alnuaimi AD, Dashper S, Cirillo N, Reynolds EC, McCullough M
    Med Mycol, 2016 Nov 01;54(8):856-64.
    PMID: 27354487 DOI: 10.1093/mmy/myw042
    Oral biofilms comprise of extracellular polysaccharides and polymicrobial microorganisms. The objective of this study was to determine the effect of polymicrobial interactions of Candida albicans, Actinomyces naeslundii, and Streptococcus mutans on biofilm formation with the hypotheses that biofilm biomass and metabolic activity are both C. albicans strain and growth medium dependent. To study monospecific biofilms, C. albicans, A. naeslundii, and S. mutans were inoculated into artificial saliva medium (ASM) and RPMI-1640 in separate vials, whereas to study polymicrobial biofilm formation, the inoculum containing microorganisms was prepared in the same vial prior inoculation into a 96-well plate followed by 72 hours incubation. Finally, biofilm biomass and metabolic activity were measured using crystal violet and XTT assays, respectively. Our results showed variability of monospecies and polymicrobial biofilm biomass between C. albicans strains and growth medium. Based on cut-offs, out of 32, seven RPMI-grown biofilms had high biofilm biomass (HBB), whereas, in ASM-grown biofilms, 14 out of 32 were HBB. Of the 32 biofilms grown in RPMI-1640, 21 were high metabolic activity (HMA), whereas in ASM, there was no biofilm had HMA. Significant differences were observed between ASM and RPMI-grown biofilms with respect to metabolic activity (P <01). In conclusion, biofilm biomass and metabolic activity were both C. albicans strain and growth medium dependent.
    Matched MeSH terms: Culture Media/chemistry*
  14. High level expression of Glomerella cingulata cutinase in dense cultures of Pichia pastoris grown under fed-batch conditions
    Seman WM, Bakar SA, Bukhari NA, Gaspar SM, Othman R, Nathan S, et al.
    J Biotechnol, 2014 Aug 20;184:219-28.
    PMID: 24910973 DOI: 10.1016/j.jbiotec.2014.05.034
    A Pichia pastoris transformant carrying the cutinase cDNA of Glomerella cingulata was over-expressed in a 5L bioreactor (2.0L working volume) under fed-batch conditions. Bioreactor experiments rely on varying selected parameters in repeated rounds of optimisation: here these included duration of induction, pH and temperature. Highest cell densities (320gL(-1) wet cell weight) with a cutinase production of 3800mgL(-1) and an activity of 434UmL(-1) were achieved 24h after induction with methanol in basal salt medium (at pH 5 and 28°C). Characterisation of the cutinase showed that it was stable between pH 6 and pH 11, had an optimum pH of 8.0 and retained activity for 30min at 50°C (optimum temperature 25°C).The preferred substrates of G. cingulata cutinase were the medium- to long-chain ρ-nitrophenyl esters of ρ-nitrophenylcaprylate (C8), ρ-nitrophenyllaurate (C12) and ρ-nitrophenylmyristate (C14), with the highest catalytic efficiency, kcat/Km of 7.7±0.7mM(-1)s(-1) for ρ-nitrophenylcaprylate. Microscopic analyses showed that the G. cingulata cutinase was also capable of depolymerising the high molecular weight synthetic polyester, polyethylene terephthalate.
    Matched MeSH terms: Batch Cell Culture Techniques/methods*
  15. Bioprospecting of indigenous marine microalgae with ammonium tolerance from aquaculture ponds for microalgae cultivation with ammonium-rich wastewaters
    Katayama T, Nagao N, Kasan NA, Khatoon H, Rahman NA, Takahashi K, et al.
    J Biotechnol, 2020 Nov 10;323:113-120.
    PMID: 32768414 DOI: 10.1016/j.jbiotec.2020.08.001
    We isolated fifty-two strains from the marine aquaculture ponds in Malaysia that were evaluated for their lipid production and ammonium tolerance and four isolates were selected as new ammonium tolerant microalgae with high-lipid production: TRG10-p102 Oocystis heteromucosa (Chlorophyceae); TRG10-p103 and TRG10-p105 Thalassiosira weissflogii (Bacillariophyceae); and TRG10-p201 Amphora coffeiformis (Bacillariophyceae). Eicosapentenoic acid (EPA) in three diatom strain was between 2.6 and 18.6 % of total fatty acids, which were higher than in O. heteromucosa. Only A. coffeiformi possessed arachidonic acid. Oocystis heteromucosa naturally grew at high ammonium concentrations (1.4-10 mM), whereas the growth of the other strains, T. weissflogii and A. coffeiformi, were visibly inhibited at high ammonium concentrations (>1.4 mM-NH4). However, two strains of T. weissflogii were able to grow at up to 10 mM-NH4 by gradually acclimating to higher ammonium concentrations. The ammonium tolerant strains, especially T. weissflogii which have high EPA contents, were identified as a valuable candidate for biomass production utilizing NH4-N media, such as ammonium-rich wastewater.
    Matched MeSH terms: Cell Culture Techniques/methods
  16. Selective media and real-time PCR improves diagnosis of melioidosis in community-acquired pneumonia in a low-incidence setting in Kuala Lumpur, Malaysia
    Subakir H, Chong YM, Chan YF, Hasan MS, Jamaluddin MFH, Pang YK, et al.
    J Med Microbiol, 2020 Jan;69(1):49-51.
    PMID: 31750812 DOI: 10.1099/jmm.0.001108
    Introduction.Burkholderia pseudomallei (melioidosis) is an important cause of community-acquired pneumonia (CAP) in the tropics. Selective medium is recommended for laboratory diagnosis with non-sterile respiratory samples, while PCR is not routinely used due to variable reported performance. The effectiveness of these diagnostic modalities varies by site.Aim. To compare selective media and real-time PCR (qPCR) with routine media in detecting B. pseudomallei in CAP respiratory samples in a low-incidence setting in Kuala Lumpur, Malaysia.Methodology. Respiratory samples were routinely cultured on blood, chocolate and MacConkey agar (RESP-ROUTINE), and compared to culture on selective Ashdown medium (RESP-SELECTIVE) and qPCR. The gold standard was routine culture of B. pseudomallei from any site (ALL-ROUTINE).Results.B. pseudomallei was detected in 8/204 (3.9 %) samples. Overall sensitivity rates differed (P=0.03) for qPCR (100%), RESP-SELECTIVE (87.5%) and RESP-ROUTINE (50%). There was a trend towards lower median days to positive culture for RESP-SELECTIVE (1 day) compared to RESP-ROUTINE (2 days, P=0.08) and ALL-ROUTINE (2 days, P=0.06). Reagent costs for each additional detection were USD59 for RESP-SELECTIVE and USD354 for PCR.Conclusions. In a low-incidence setting, selective culture of respiratory samples on Ashdown was more sensitive and allowed quicker identification than routine media, at reasonable cost. Blood cultures are critical, confirming four cases missed by routine respiratory culture. Selective medium is useful in early pneumonia (pre-sepsis) and resource-limited settings where blood cultures are infrequently done. Real-time PCR is costly, but highly sensitive and useful for high-risk patients with diabetes, cancer or immunosuppressants, or requiring ventilation or intensive care.
    Matched MeSH terms: Culture Media/chemistry*
  17. Fulltext Lactobacillus plantarum strain HT-W104-B1: potential bacterium isolated from Malaysian fermented foods for control of the dermatophyte Trichophyton rubrum
    Danial AM, Medina A, Magan N
    World J Microbiol Biotechnol, 2021 Feb 24;37(4):57.
    PMID: 33625606 DOI: 10.1007/s11274-021-03020-7
    The objective was to screen and evaluate the anti-fungal activity of lactic acid bacteria (LABs) isolated from Malaysian fermented foods against two Trichophyton species. A total of 66 LAB strains were screened using dual culture assays. This showed that four LAB strains were very effective in inhibiting growth of T. rubrum but not T. interdigitale. More detailed studies with Lactobacillus plantarum strain HT-W104-B1 showed that the supernatant was mainly responsible for inhibiting the growth of T. rubrum. The minimum inhibitory concentration (MIC), inhibitory concentration, the 50% growth inhibition (IC50) and minimum fungicide concentration (MFC) were 20 mg/mL, 14 mg/mL and 30 mg/mL, respectively. A total of six metabolites were found in the supernatant, with the two major metabolites being L-lactic acid (19.1 mg/g cell dry weight (CDW)) and acetic acid (2.2 mg/g CDW). A comparative study on keratin agar media showed that the natural mixture in the supernatants predominantly contained L-lactic and acetic acid, and this significantly controlled the growth of T. rubrum. The pure two individual compounds were less effective. Potential exists for application of the natural mixture of compounds for the treatment of skin infection by T. rubrum.
    Matched MeSH terms: Culture Media/metabolism
  18. Fulltext Characterization and antimicrobial activities of two Streptomyces isolates from soil in the periphery of Universiti Putra Malaysia
    Zin NZ, Tasrip NA, Desa MN, Kqueen CY, Zakaria ZA, Hamat RA, et al.
    Trop Biomed, 2011 Dec;28(3):651-60.
    PMID: 22433896 MyJurnal
    This study was to assess the identification and antimicrobial activities of two actinomycete isolates. The two isolates designated as B8 and C2, were isolated from a patch of soil in the peripheral area of Universiti Putra Malaysia by streaking on starch casein agar after standard serial dilution procedures. Their antimicrobial activities were first evaluated against eight clinical laboratory strains namely Bacillus sp., Enterococcus sp., Escherichia coli, Klebsiella sp., Pseudomonas sp., Salmonella sp., Staphylococcus aureus, and Staphylococcus epidermidis by perpendicular streak method on Mueller Hinton and Tryptic Soy agar. In both media, a broad-spectrum antibacterial activity was observed for both isolates, with B8 against all the test bacteria and C2 against five of them (Bacillus sp., E. coli, Pseudomonas sp., S. aureus and S. epidermidis). Re-assessment against E. coli ATCC 25922 and S. aureus ATCC 25923 strains by similar method showed antibacterial activities by isolate B8 against both ATTC strains while C2 only against S. aureus ATCC 25923. Streptomyces griseus ATCC 10137 was included in the later experiment and showed antibacterial activity against both ATCC strains. Subsequently, the two isolates were identified by PCR/sequencing techniques and phylogenetic analysis to be Streptomyces species (>93% homology based on 16S rRNA and rpoB genes). Characterization on cultural characteristic and viable count at different temperatures (37ºC and 28ºC), on different microbiological media (AIA, ISP-2, MHA, NA, PDA and TSA), were performed. More morphological features were observed on ISP-2 for both isolates. A higher growth yield was also observed at 28ºC in all media but in comparing that between the two isolates, isolate B8 outnumbered C2 at all experimental conditions. The observed variation in cultural traits and growth yield indicate unique properties between the two antibiotic-producing isolates.
    Matched MeSH terms: Culture Media/chemistry
  19. Fulltext Cloning and expression of Toxoplasma gondii dense granular protein 4 (GRA4) in Pichia pastoris
    Lau YL, Hasan MT, Thiruvengadam G, Idris MM, Init I
    Trop Biomed, 2010 Dec;27(3):525-33.
    PMID: 21399595
    GRA4 of Toxoplasma gondii has been shown to prompt IgG, IgM and IgA responses in previous studies and is thus considered one of the major immunogenic proteins from T. gondii that can be used for both diagnostics purposes and vaccine development. This study seeks to clone and express the GRA4 in Pichia pastoris, which has numerous advantages over other systems for expression of eukaryotic proteins. In order to achieve this, the gene was cloned into the pPICZα A expression vector, which was then incorporated into the P. pastoris genome via insertional integration for expression of the recombinant protein, under the AOX1 promoter. The antigen was expressed along with the prepro sequence of the α-factor of yeast so that it could be excreted out of the P. pastoris cells and obtained from the medium. Upon SDS-PAGE analysis it was found that the recombinant protein was expressed optimally as a 40 kDa protein after 96 hours of induction with 0.75% of methanol. The expressed GRA4 protein showed discrepancy in size with the calculated molecular mass. This may be attributed to the various posttranslational modifications including glycosylation and phosphorylation. Despite the difference in molecular weight, the recombinant protein was able to detect toxoplasmosis in Western blot format. The recombinant GRA4 was expressed with an intact polyhistidine-tag, which could be used for future purification of the antigen.
    Matched MeSH terms: Culture Media/chemistry
  20. Fulltext Paracrine Effects of Adipose-Derived Stem Cells on Matrix Stiffness-Induced Cardiac Myofibroblast Differentiation via Angiotensin II Type 1 Receptor and Smad7
    Yong KW, Li Y, Liu F, Bin Gao, Lu TJ, Wan Abas WA, et al.
    Sci Rep, 2016 10 05;6:33067.
    PMID: 27703175 DOI: 10.1038/srep33067
    Human mesenchymal stem cells (hMSCs) hold great promise in cardiac fibrosis therapy, due to their potential ability of inhibiting cardiac myofibroblast differentiation (a hallmark of cardiac fibrosis). However, the mechanism involved in their effects remains elusive. To explore this, it is necessary to develop an in vitro cardiac fibrosis model that incorporates pore size and native tissue-mimicking matrix stiffness, which may regulate cardiac myofibroblast differentiation. In the present study, collagen coated polyacrylamide hydrogel substrates were fabricated, in which the pore size was adjusted without altering the matrix stiffness. Stiffness is shown to regulate cardiac myofibroblast differentiation independently of pore size. Substrate at a stiffness of 30 kPa, which mimics the stiffness of native fibrotic cardiac tissue, was found to induce cardiac myofibroblast differentiation to create in vitro cardiac fibrosis model. Conditioned medium of hMSCs was applied to the model to determine its role and inhibitory mechanism on cardiac myofibroblast differentiation. It was found that hMSCs secrete hepatocyte growth factor (HGF) to inhibit cardiac myofibroblast differentiation via downregulation of angiotensin II type 1 receptor (AT1R) and upregulation of Smad7. These findings would aid in establishment of the therapeutic use of hMSCs in cardiac fibrosis therapy in future.
    Matched MeSH terms: Culture Media, Conditioned/pharmacology
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