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Optimization of L-asparaginase production from endophytic Fusarium proliferatum using OFAT and RSM and its cytotoxic evaluation

Yap LS, Lee WL, Ting ASY

J Microbiol Methods, 2021 12;191:106358.
PMID: 34743930 DOI: 10.1016/j.mimet.2021.106358

L-asparaginase from endophytic Fusarium proliferatum (isolate CCH, GenBank accession no. MK685139) isolated from the medicinal plant Cymbopogon citratus (Lemon grass), was optimized for its L-asparaginase production and its subsequent cytotoxicity towards Jurkat E6 cell line. The following factors were optimized; carbon source and concentration, nitrogen source and concentration, incubation period, temperature, pH and agitation rate. Optimization of L-asparaginase production was performed using One-Factor-At-A-Time (OFAT) and Response surface methodology (RSM) model. The cytotoxicity of the crude enzyme from isolate CCH was tested on leukemic Jurkat E6 cell line. The optimization exercise revealed that glucose concentration, nitrogen source, L-asparagine concentration and temperature influenced the L-asparaginase production of CCH. The optimum condition suggested using OFAT and RSM results were consistent. As such, the recommended conditions were 0.20% of glucose, 0.99% of L-asparagine and 5.34 days incubation at 30.50 °C. The L-asparaginase production of CCH increased from 16.75 ± 0.76 IU/mL to 22.42 ± 0.20 IU/mL after optimization. The cytotoxicity of the crude enzyme on leukemic Jurkat cell line recorded IC50 value at 33.89 ± 2.63% v/v. To conclude, the enzyme extract produced from Fusarium proliferatum under optimized conditions is a potential alternative resource for L-asparaginase.

Matched MeSH terms: Microbiological Techniques/methods
Fulltext Back to basic: bio-burden on hands of health care personnel in tertiary teaching hospital in Malaysia

Wong JL, Siti Azrin AH, Narizan MI, Norliah Y, Noraida M, Amanina A, et al.

Trop Biomed, 2014 Sep;31(3):534-9.
PMID: 25382481 MyJurnal

Hands of Health Care Personnel (HCP) are one of the most common vehicles for the transmission of infection. Microorganisms can survive well on the hands of HCP for a certain duration. Therefore, the purpose of this study is to bring awareness to HCP that their hands can actually be contaminated with many microorganisms. These microbes on the hands of HCP can potentially infect their patients if they do not comply with the proper hand hygiene practice. This cross-sectional study was conducted at a randomly selected Intensive Care Unit (ICU) and general ward in a hospital. Twenty five HCP from each ward were randomly selected and their hands were imprinted on blood culture plates. Microorganism growth were quantified and identified. Data were analyzed and presented as descriptive analysis. One hundred blood agar plates were processed and analyzed. Majority (71%) of the samples had more than 50 colony-forming units (CFU) and only 17% of the samples had less than 25 CFU. Microorganisms identified include Staphylococcus spp., Acinetobacter spp., Enterobacteriaceae, Pseudomonas spp., Moraxella, Delftiaacidovorans and fungi. All isolated microorganisms were antibiotic sensitive strain. This study showed that the hands of HCP were contaminated with many microorganisms. Therefore, it is imperative that HCP must practice proper hand hygiene when taking care of their patients in the wards.

Matched MeSH terms: Microbiological Techniques
Occurrence and characterization of Candida nivariensis from a culture collection of Candida glabrata clinical isolates in Malaysia

Tay ST, Lotfalikhani A, Sabet NS, Ponnampalavanar S, Sulaiman S, Na SL, et al.

Mycopathologia, 2014 Oct;178(3-4):307-14.
PMID: 25022264 DOI: 10.1007/s11046-014-9778-9

BACKGROUND: Candida nivariensis and C. bracarensis have been recently identified as emerging yeast pathogens which are phenotypically indistinguishable from C. glabrata. However, there is little data on the prevalence and antifungal susceptibilities of these species.
OBJECTIVE: This study investigated the occurrence of C. nivariensis and C. bracarensis in a culture collection of 185 C. glabrata isolates at a Malaysian teaching hospital.
METHODS: C. nivariensis was discriminated from C. glabrata using a PCR assay as described by Enache-Angoulvant et al. (J Clin Microbiol 49:3375-9, 2011). The identity of the isolates was confirmed by sequence analysis of the D1D2 domain and internal transcribed spacer region of the yeasts. The isolates were cultured on Chromogenic CHROMagar Candida (®) agar (Difco, USA), and their biochemical and enzymic profiles were determined. Antifungal susceptibilities of the isolates against amphotericin B, fluconazole, voriconazole and caspofungin were determined using E tests. Clotrimazole MICs were determined using a microbroth dilution method.
RESULTS: There was a low prevalence (1.1 %) of C. nivariensis in our culture collection of C. glabrata. C. nivariensis was isolated from a blood culture and vaginal swab of two patients. C. nivariensis grew as white colonies on Chromogenic agar and demonstrated few positive reactions using biochemical tests. Enzymatic profiles of the C. nivariensis isolates were similar to that of C. glabrata. The isolates were susceptible to amphotericin B, fluconazole, voriconazole and caspofungin. Clotrimazole resistance is suspected in one isolate.
CONCLUSION: This study reports for the first time the emergence of C. nivariensis in our clinical setting.

Matched MeSH terms: Microbiological Techniques/methods
Fulltext First Two Cases of Fungal Infections Associated with Multi-drug Resistant Yeast, Fereydounia khargensis

Tap RM, Ramli NY, Sabaratnam P, Hashim R, Bakri AR, Bee LB, et al.

Mycopathologia, 2016 Aug;181(7-8):531-7.
PMID: 27010640 DOI: 10.1007/s11046-016-0002-y

The number of new fungal pathogens is increasing due to growing population of immunocompromised patients and advanced identification techniques. Fereydounia khargensis is a yeast and was first described in 2014 from environmental samples. As far as we know, this is the first report of human infections associated with F. khargensis. The yeasts were isolated from blood of a HIV-positive patient and pleural fluid of chronic renal failure patient. Amplification and sequencing of the internal transcribed spacer and the large subunit regions confirmed the identity of the isolates. Both isolates showed multi-drug resistance to antifungal agents tested.

Matched MeSH terms: Microbiological Techniques
Fulltext Synbiotics growth optimization of Bifidobacterium pseudocatenulatum G4 with prebiotics using a statistical methodology

Shuhaimi M, Kabeir BM, Yazid AM, Nazrul Somchit M

J Appl Microbiol, 2009 Jan;106(1):191-8.
PMID: 19054238 DOI: 10.1111/j.1365-2672.2008.03991.x

This study demonstrated the optimum growth of Bifidobacterium pseudocatenulatum G4 with prebiotics via statistical model.

Matched MeSH terms: Microbiological Techniques
An improved stool concentration procedure for the detection of Cryptosporidium oocysts in Orang Asli stool samples

Salleh FM, Moktar N, Yasin AM, Al-Mekhlafi HM, Anuar TS

J Microbiol Methods, 2014 Nov;106:143-145.
PMID: 25193442 DOI: 10.1016/j.mimet.2014.08.019

To improve the stool concentration procedure, we modified different steps of the standard formalin-ether concentration technique and evaluated these modifications by examining stool samples collected in the field. Seven samples were found positive by the modified formalin-ether concentration technique (M-FECT). Therefore, the M-FECT procedure provides enhanced detection of Cryptosporidium oocysts.

Matched MeSH terms: Microbiological Techniques/methods*
Update on laboratory diagnosis of amoebiasis

Saidin S, Othman N, Noordin R

Eur J Clin Microbiol Infect Dis, 2019 Jan;38(1):15-38.
PMID: 30255429 DOI: 10.1007/s10096-018-3379-3

Amoebiasis, an enteric protozoan disease caused by Entamoeba histolytica, is a public health problem in many developing countries, causing up to 100,000 fatal cases annually. Detection of the pathogenic E. histolytica and its differentiation from the non-pathogenic Entamoeba spp. play a crucial role in the clinical management of patients. Laboratory diagnosis of intestinal amoebiasis in developing countries still relies on labour-intensive and insensitive methods involving staining of stool sample and microscopy. Newer and more sensitive methods include a variety of antigen detection ELISAs and rapid tests; however, their diagnostic sensitivity and specificity seem to vary between studies, and some tests do not distinguish among the Entamoeba species. Molecular detection techniques are highly sensitive and specific and isothermal amplification approaches may be developed into field-applicable tests; however, cost is still a barrier for their use as a routine laboratory test method in most endemic areas. Laboratory diagnosis of extraintestinal amoebiasis faces challenges of lack of definitive detection of current infection and commercially available point-of-care tests. For both types of amoebiasis, there is still a need for highly sensitive and specific tests that are rapid and cost-effective for use in developing countries where the disease is prevalent. In recent years, new molecules of diagnostic value are being discovered and new tests developed. The advances in 'omics' technologies are enabling discoveries of new biomarkers that may help distinguish between different infection stages.

Matched MeSH terms: Microbiological Techniques*
Comparison of three different methods for the presumptive detection of ESBL production in ceftazidime-resistant strains of Klebsiella pneumoniae

Palasubramaniam S, Parasakthi N

Malays J Pathol, 2001 Dec;23(2):73-8.
PMID: 12166595

Twenty-eight (28) strains of ceftazidime-resistant Klebsiella pneumoniae were isolated from blood cultures of in-patients from University Hospital, Kuala Lumpur between March 1995 and August 1996. Three methods were used to detect the production of ESBL enzymes by these strains. These three methods include the double-disc synergy test (DDST), inhibitor-potentiated disc-diffusion test (IPDD) and the E-test ESBL method. All strains could be identified as ESBL producers using the DDST method by a minimum of two beta-lactams and these included either a combination of ceftazidime and ceftriaxone with clavulanate respectively or cefotaxime and aztreonam with clavulanate respectively. Similarly using a combination of either cefotaxime and ceftriaxone with clavulanate or ceftriaxone and aztreonam with clavulanate respectively would have detected all strains as ESBL producers. The IPDD method could also detect for ESBL activity based on combinations of beta-lactam antibiotics with clavulanate respectively. All combinations of beta-lactam antibiotics could detect for ESBL activity in all the strains except a combination of either ceftazidime and aztreonam or cefotaxime and ceftriaxone with clavulanate respectively. The E-Test method using ceftazidime alone and in combination with clavulanate was found to be the most effective method in the presumptive identification of ESBL activity in all the strains.

Matched MeSH terms: Microbiological Techniques/standards*
Infrequent occurrence of vancomycin-resistant enterococci in poultry from Malaysian wet markets

Ong CH, Asaad M, Lim KC, Ngeow YF

Malays J Pathol, 2002 Dec;24(2):91-4.
PMID: 12887166

Fifty samples of chicken, duck and geese faeces were obtained from 13 wet markets in Kuala Lumpur to study the prevalence of vancomycin-resistant enterococci (VRE) among local market poultry. Biotyping of colonies grown on azide agar incubated at 45 degrees C yielded E. pseudoavium, E. faecalis, E. faecium and E. gallinarum from chicken faeces and E. malodoratus, E. faecalis, E. faecium, E. gallinarum, E. hirae/dispar, and E. durans from goose and duck faeces. On agar containing 6 mg/ l of vancomycin, one strain of E. flavescens was identified, giving a VRE detection rate of 2.0%. This isolate had a vancomycin M.I.C. of 8 mg/l as determined by the Etest, and the van C-3 gene that was identified by PCR followed by sequence analysis. The prevalence of VRE among poultry sold in local markets appears to be low, and may reflect the infrequent use of antimicrobials in our poultry farms. Nevertheless, the possibility of human acquisition of microbes via the food chain cautions against the use of antimicrobials in animal husbandry that may encourage the emergence and spread of multi-drug resistant organisms like the VRE among animal microbial flora.

Matched MeSH terms: Microbiological Techniques
Chaetomium globosum Cutaneous Fungal Infection Confirmed by Molecular Identification: A Case Report from Malaysia

Mohd Tap R, Sabaratnam P, Ahmad NA, Abd Razak MF, Hashim R, Ahmad N

Mycopathologia, 2015 Aug;180(1-2):137-41.
PMID: 25894509 DOI: 10.1007/s11046-015-9890-5

An 11-year-old girl presented with multiple blisters on her the right foot complicated with cellulitis. The conventional and molecular identification were performed on the culture. The internal transcribed spacer (ITS) region in rRNA gene of the isolate was amplified by PCR. The sequence of the amplified ITS region matched 99 % with that of Chaetomium globosum in the GenBank. This is the first report describing C. globosum causing cutaneous infection in Malaysia.

Matched MeSH terms: Microbiological Techniques
Meningitis Caused by Rhodotorula mucilaginosa in HIV-Infected Patient: A Case Report and Review of the Literature

Mohd Nor F, Tan LH, Na SL, Ng KP

Mycopathologia, 2015 Aug;180(1-2):95-8.
PMID: 25739670 DOI: 10.1007/s11046-015-9879-0

Rhodotorula species are increasingly being identified as a cause of fungal infection in the central nervous system, especially in patients with compromised immunity. The diagnosis could easily be missed due to low index of suspicion, as cryptococcus meningitis and cerebral toxoplasmosis are more common amongst immunocompromised hosts. To date, there are six cases of Rhodotorula-related meningitis reported, and three are associated with human immunodeficiency virus infection. In this report, a case of a Malaysian male with underlying human immunodeficiency virus infection who developed Rhodotorula mucilaginosa meningitis is presented. High-grade fever and severe headaches were the complaints presented in three previous case reports. India ink and nigrosin stainings were performed in the two previous reports and both revealed positive results. R. mucilaginosa were isolated from the culture of the patient's cerebrospinal fluid in all three previous reports. Predominant lymphocyte infiltration in the cerebrospinal fluid examination was documented in two reports. CD4 counts were above 100/µl in two previously published reports, while another report documented CD4 count as 56/µl. Amphotericin B and itraconazole are identified to be the first line of antifungal used and as the maintenance therapy, respectively. The possibility of relapse cannot be excluded as it was reported in the first report. It was also revealed that the current case has almost similar clinical presentation and therapeutic outcome as compared to the published reports, but some differences in diagnostic details were to be highlighted.

Matched MeSH terms: Microbiological Techniques
Fulltext Identification of local clinical Candida isolates using CHROMagar Candida™ as a primary identification method for various Candida species

Madhavan P, Jamal F, Chong PP, Ng KP

Trop Biomed, 2011 Aug;28(2):269-74.
PMID: 22041745

The objective of our study was to study the effectiveness of CHROMagar Candida™ as the primary identification method for various clinical Candida isolates, other than the three suggested species by the manufacturer. We studied 34 clinical isolates which were isolated from patients in a local teaching hospital and 7 ATCC strains. These strains were first cultured in Sabouraud dextrose broth (SDB) for 36 hours at 35ºC, then on CHROMagar plates at 30ºC, 35ºC and 37ºC. The sensitivity of this agar to identify Candida albicans, Candida dubliniensis, Candida tropicalis, Candida glabrata, Candida rugosa, Candida krusei and Candida parapsilosis ranged between 25 and 100% at 30ºC, 14% and 100% at 35ºC, 56% and 100% at 37ºC. The specificity of this agar was 100% at 30ºC, between 97% and 100% at 35ºC, 92% and 100% at 37ºC. The efficiency of this agar ranged between 88 and 100% at 30ºC, 83% and 100% at 35ºC, 88% and 100% at 37ºC. Each species also gave rise to a variety of colony colours ranging from pink to green to blue of different colony characteristics. Therefore, the chromogenic agar was found to be useful in our study for identifying clinical Candida isolates.

Matched MeSH terms: Microbiological Techniques/methods*
Fulltext DNA barcoding survey of Trichoderma diversity in soil and litter of the Colombian lowland Amazonian rainforest reveals Trichoderma strigosellum sp. nov. and other species

López-Quintero CA, Atanasova L, Franco-Molano AE, Gams W, Komon-Zelazowska M, Theelen B, et al.

Antonie Van Leeuwenhoek, 2013 Nov;104(5):657-74.
PMID: 23884864 DOI: 10.1007/s10482-013-9975-4

The diversity of Trichoderma (Hypocreales, Ascomycota) colonizing leaf litter as well as the rhizosphere of Garcinia macrophylla (Clusiaceae) was investigated in primary and secondary rain forests in Colombian Amazonia. DNA barcoding of 107 strains based on the internal transcribed spacers 1 and 2 (ITS1 and 2) of the ribosomal RNA gene cluster and the partial sequence of the translation elongation factor 1 alpha (tef1) gene revealed that the diversity of Trichoderma was dominated (71 %) by three common cosmopolitan species, namely Trichoderma harzianum sensu lato (41 %), Trichoderma spirale (17 %) and Trichoderma koningiopsis (13 %). Four ITS 1 and 2 phylotypes (13 strains) could not be identified with certainty. Multigene phylogenetic analysis and phenotype profiling of four strains with an ITS1 and 2 phylotype similar to Trichoderma strigosum revealed a new sister species of the latter that is described here as Trichoderma strigosellum sp. nov. Sequence similarity searches revealed that this species also occurs in soils of Malaysia and Cameroon, suggesting a pantropical distribution.

Matched MeSH terms: Microbiological Techniques
A Hospital-based Study on the Local Epidemiology of Pneumonia Including the Contribution of Legionella Pneumonia

Lourdesamy Anthony AI, Zam Z, Hussin N

Malays J Med Sci, 2020 Dec;27(6):79-88.
PMID: 33447136 DOI: 10.21315/mjms2020.27.6.8

Background: In real-life practice, only 20% of hospitalised pneumonia cases have an identified etiology. The usage of Legionella urine antigen test (LUAT) in developed nations revolutionised case detection rates. Accordingly, our objectives were to study the microbiological etiology for hospitalised pneumonia patients and the diagnosis of Legionella pneumonia.
Methods: A prospective, observational single-centre study was conducted where all 504 cases that were consecutively admitted for pneumonia were enrolled. Blood and sputum samples obtained were used to identify pathogens using standard microbiological culture methods. The urine samples collected were tested using the ImmunocatchTMLegionella immunochromatographic (ICT) urine antigen test.
Results: A microbiological diagnosis was only achieved in 104 cases (20.6%) and a Gram-negative infection predominance was observed. Culture-positive cases required longer hospitalisation (8.46 days versus 5.53 days; P < 0.001) and the higher usage of antipseudomonal antibiotics (23.1% versus 8.3%; P < 0.001). Only 3 cases (0.6%) were diagnosed with Legionella pneumonia.
Conclusion: The local pathogen distribution is diverse compared to other regions. Culture-negative pneumonia is common and significantly differs from culture-positive pneumonia. Legionella pneumophila serotype 1 is not a common cause of pneumonia and LUAT did not help demystify the cause of culture-negative pneumonia.

Matched MeSH terms: Microbiological Techniques
Isolation of indigenous larvicidal microbial control agents of mosquitos: the Malaysian experience

Lee HL, Seleena P

Southeast Asian J. Trop. Med. Public Health, 1990 Jun;21(2):281-7.
PMID: 2237596

A nationwide screening program searching for microbial control agents of mosquitos was initiated in Malaysia in 1986. A total of 725 samples were collected and 2,394 bacterial colonies were isolated and screened for larvicidal activity. From such screening, 20 Bacillus thuringiensis, 6 B. sphaericus, 1 Clostridium bifermentans and 2 Pseudomonas pseudomallei larvicidal isolates were obtained. Of these, a new B. thuringiensis named as subspecies malaysianensis was found, while the C. bifermentans was also a new anaerobe individualized as serovar malaysia. It was concluded that this screening program was highly successful.

Matched MeSH terms: Microbiological Techniques*
Comparison of microplate- and bottle-based methods to age yeast for chronological life span assays

Kwong MMY, Lee JW, Samian MR, Watanabe N, Osada H, Ong EBB

J Microbiol Methods, 2019 12;167:105743.
PMID: 31629019 DOI: 10.1016/j.mimet.2019.105743

This study compared the chronological life span and survival of Saccharomyces cerevisiae aged in a microplate or bottle, under different aeration and calorie restriction conditions. Our data shows that limited aeration in the microplate-aged culture contributed to slower outgrowth but extended yeast CLS compared to the bottle-aged culture.

Matched MeSH terms: Microbiological Techniques/instrumentation; Microbiological Techniques/methods*
Quantitative assessment of antimalarial activities from Malaysian Plasmodium falciparum isolates by modified in vitro microtechnique

Hoon AH, Lam CK, Wah MJ

Antimicrob Agents Chemother, 1995 Mar;39(3):626-8.
PMID: 7793863

Malaysian, TGR (Thailand), and Gambian (West African) Plasmodium falciparum isolates were cultured in vitro by the candle jar method and were characterized for their susceptibilities to present antimalarial drugs by the modified in vitro microtechnique. Results showed that 93 and 47% of the Malaysian isolates were resistant at 50% inhibitory concentrations of 0.1415 to 0.7737 and 0.1025 to 0.1975 microM, respectively, while the rest were susceptible to choloroquine and cycloguanil at 0.0376 and 0.0306 to 0.0954 microM, respectively. All isolates were susceptible to mefloquine, quinine, and pyrimethamine at 0.0026 to 0.0172, 0.0062 to 0.0854, and 0.0149 to 0.0663 microM, respectively. In contrast, the Gambian isolate was susceptible to multiple drugs at 0.0024 to 0.0282 microM; TGR was resistant to chloroquine at 0.8147 microM but was susceptible to mefloquine, quinine, cycloguanil, and pyrimethamine at 0.0024, 0.0096, 0.0143, and 0.0495 microM, respectively.

Matched MeSH terms: Microbiological Techniques
Fulltext Mobile Phones: A Possible Vehicle of Bacterial Transmission in a Higher Learning Institution in Malaysia

Hikmah N', Anuar TS

Malays J Med Sci, 2020 Mar;27(2):151-158.
PMID: 32788850 MyJurnal DOI: 10.21315/mjms2020.27.2.15

Background: Mobile phones (MPs) have become one of the most indispensable accessories in social and professional life. Though they offer plenty of benefits, MPs are prolific breeding grounds for infectious pathogens in communities. Thus, the aim of this study was to identify the prevalence of bacterial contamination and determine antimicrobial susceptibility pattern of Staphylococcus aureus (S. aureus) from MPs.
Methods: A cross-sectional study was conducted from March to July 2019 on 126 students and 37 laboratory staff/clinical instructors' MPs from the Faculty of Health Sciences, Universiti Teknologi MARA, Malaysia by a simple random sampling technique. Along with the questionnaire, a swab sample from each participant's MPs was collected and transported to the microbiology laboratory for bacterial culture as per standard microbiological procedures and antimicrobial susceptibility test by the disc diffusion technique. Data were analysed by the Statistical Package for Social Sciences Programme version 24.
Results: All of the tested MPs were contaminated with either single or mix bacterial agents. Bacillus spp. (74.8%), coagulase-negative staphylococci (CoNS; 47.9%) and S. aureus (20.9%) were the most predominant bacterial isolates, whilst the least isolate was Proteus vulgaris (P. vulgaris) (2.5%). Oxacillin resistance was seen in 5.9% of S. aureus isolate. A comparison of bacteria type and frequency among gender showed a significant difference with P. vulgaris (P = 0.003) and among profession showed a significant difference with S. aureus (P = 0.004).
Conclusion: The present study indicates that MPs can serve as a vector for both pathogenic and non-pathogenic organisms. Therefore, full guidelines about restricting the use of MPs in laboratory environments, hand hygiene and frequent decontamination of MPs are recommended to limit the risk of cross-contamination and healthcare-associated infections caused by MPs.

Matched MeSH terms: Microbiological Techniques
Three stage cultivation process of facultative strain of Chlorella sorokiniana for treating dairy farm effluent and lipid enhancement

Hena S, Fatihah N, Tabassum S, Ismail N

Water Res, 2015 Sep 1;80:346-56.
PMID: 26043271 DOI: 10.1016/j.watres.2015.05.001

Reserve lipids of microalgae are promising for biodiesel production. However, economically feasible and sustainable energy production from microalgae requires optimization of cultivation conditions for both biomass yield and lipid production of microalgae. Biomass yield and lipid production in microalgae are a contradictory problem because required conditions for both targets are different. Simultaneously, the mass cultivation of microalgae for biofuel production also depends extremely on the performance of the microalgae strains used. In this study a green unicellular microalgae Chlorella sorokiniana (DS6) isolated from the holding tanks of farm wastewater treatment plant using multi-step screening and acclimation procedures was found high-lipid producing facultative heterotrophic microalgae strain capable of growing on dairy farm effluent (DFE) for biodiesel feedstock and wastewater treatment. Morphological features and the phylogenetic analysis for the 18S rRNA identified the isolated strains. A novel three stage cultivation process of facultative strain of C. sorokiniana was examined for lipid production.

Matched MeSH terms: Microbiological Techniques/methods
Optimization of Salmonella Typhi biofilm assay on polypropylene microtiter plates using response surface methodology

Ganjali Dashti M, Abdeshahian P, Sudesh K, Phua KK

Biofouling, 2016;32(4):477-87.
PMID: 26963754 DOI: 10.1080/08927014.2015.1135328

The objective of this study was to develop an optimized assay for Salmonella Typhi biofilm that mimics the environment of the gallbladder as an experimental model for chronic typhoid fever. Multi-factorial assays are difficult to optimize using traditional one-factor-at-a-time optimization methods. Response surface methodology (RSM) was used to optimize six key variables involved in S. Typhi biofilm formation on cholesterol-coated polypropylene 96-well microtiter plates. The results showed that bile (1.22%), glucose (2%), cholesterol (0.05%) and potassium chloride (0.25%) were critical factors affecting the amount of biofilm produced, but agitation (275 rpm) and sodium chloride (0.5%) had antagonistic effects on each other. Under these optimum conditions the maximum OD reading for biofilm formation was 3.4 (λ600 nm), and the coefficients of variation for intra-plate and inter-plate assays were 3% (n = 20) and 5% (n = 8), respectively. These results showed that RSM is an effective approach for biofilm assay optimization.

Matched MeSH terms: Microbiological Techniques/methods; Microbiological Techniques/standards

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