High Cell Surface Hydrophobicity Research Articles

Identification and characterization of probiotic yeast isolated from digestive tract of ducks.

The objective of this study was to isolate and identify yeast strains from the digestive tract of ducks, and evaluate in vitro their potential as probiotics in poultry. The yeast strains were isolated using malt extract agar medium, and identified through morphological, physiological, and biochemical tests as well as sequence homology analyses of 26S rDNA D1/D2 region. A total of 35 yeast strains were isolated from the guts of Cherry Valley meat ducks, including seven strains of Saccharomyces cerevisiae (S. cerevisiae). These seven strains of S. cerevisiae were further screened for their use as alternative yeast probiotics strains for poultry feed. The yeast strains were characterized for their cell surface hydrophobicity, autoaggregation ability, and resistance to high temperature (30°C, 37°C, and 42°C), low pH (2.0, 3.0, and 4.0), bile salts (0.3% and 0.6%), and nutrition starvation (2, 4, 6, 8, 10, and 12 days). The isolates of WHY-2 and WHY-7 had a higher survival percentage at 37°C, pH 2.0, 0.60% poultry bile salts, and 10 days of nutrition starvation, with higher cell surface hydrophobicity and autoaggregation, when compared with the other isolates, suggesting that the isolates WHY-2 and WHY-7, could be used as probiotic candidates. The data obtained in this study could help in selecting probiotic yeast candidates for use in poultry industry.

Exploring the Probiotic and Compound Feed Fermentative Applications of Lactobacillus plantarum SK1305 Isolated from Korean Green Chili Pickled Pepper.

Herein, we explore the probiotic potentials and soybean meal (SBM) compound feed fermentative applications of Lactobacillus plantarum SK1305 strain isolated previously from Korean green chili pickled pepper (gochu-jangajji). The isolate exhibited higher acid (pH 2.5) and bile tolerance (0.3%, w/v) up to 2h and 4h, respectively. The cell-free culture supernatant (CFCS) displayed a broad spectrum antibacterial activities against various pathogens, which may be ascribed to high lactic acid production (L-form, 86.8 ± 0.8mM and D-form, 44.8 ± 0.2mM). Further, the strain displayed high cell-surface hydrophobicity (92.7 ± 1.0%), coupled with low auto-aggregation (23.6 ± 4.4%) but relatively higher co-aggregation properties with C. perfringens (49.6 ± 0.6%) as well as H2O2 (1.0mM) resistant property. Additionally, the isolate displayed significant DPPH free radical scavenging activity (55.2 ± 0.6%) and superoxide reducing ability in MAC-T cells. Considering safety, the isolate has no transmissible antibiotic resistant genes and harmful enzymes as well as non-hemolytic activities. Ushered by these appreciable probiotic properties, the isolate was used for solid state fermentation (SSF) of SBM compound feed. Notably, we observed a higher strain adaptability (> 1010CFU/g) following the production of L- (> 6.0 ± 0.0mM) and D-form (> 5.2 ± 0.3mM) lactic acid during fermentation for 8h. The methanolic extracts of fermented feed displayed high antibacterial and antioxidant activities, affirming the potential functional activities of fermented compound feeds. Therefore, L. plantarum SK1305 may act as a worthy inoculum toward fermentation of feed with enhanced nutritional properties.

Role and mechanism of cell-surface hydrophobicity in the adaptation of Sphingobium hydrophobicum to electronic-waste contaminated sediment.

Sphingomonads are isolated at exceptionally high frequency from organic polluted environments and assumed to be more hydrophobic than other Gram-negative bacteria. However, the potential roles of cell-surface hydrophobicity (CSH) in the cell survival in polluted environment, as well as the mechanisms underlying the CSH of sphingomonads, remain unclear. Sphingobium hydrophobicum C1T is a highly hydrophobic sphingomonad isolated from electronic-waste contaminated sediment. In this study, we found that exposure to the typical pollutants in electronic-waste contaminated sediment, such as the heavy metal ion Pb and the organic compound deca-brominated diphenyl ether (deca-BDE), resulted in the development of even higher CSH of the hydrophobic strain C1T; but no significant change was observed in the low CSH of its hydrophilic variant C2. The hydrophobic strain C1T achieved higher biomass yield in standing conditions and adsorbed more amounts of hydrophobic deca-BDE than its hydrophilic variant C2, suggesting that the high CSH potentially enhanced the adaptation of hydrophobic strain to colonize in sediment and adsorb hydrophobic nutrients. The identification of the bacterial cell-surface constituents showed that the high CSH of S. hydrophobicum was contributed greatly by outer-membrane proteins, particularly membrane transporters functioning as enhancers for nutrient uptake and stress sustainment. This study will enhance our understanding of the adaptive strategies of sphingomonads in contaminated environments. It will be of great importance to enhance the CSH of sphingomonads and utilize them in cleaning up the environment from organic pollution.

Conjugated linoleic acid production and probiotic assessment of Lactobacillus plantarum isolated from Pico cheese

Lactic acid bacteria isolated from a traditional Azorean cheese were screened for their ability to convert free linoleic acid to conjugated linoleic acid (CLA). Two strains of Lactobacillus plantarum were recognized as potential CLA producers. GC analysis identified cis-9, trans-11 C18:2 as the predominant isomer (10–14 μg/mL), followed by trans-9, trans-11 C18:2 (4–6 μg/mL). The CLA producing strains demonstrated strong biofilm capacity, high cell surface hydrophobicity and good auto-aggregation ability. These strains were capable of surviving in the presence of bile salts (0.3%) and pancreatin (0.1%), but only the highest CLA producer (L3C1E8) was able to resist low pH (2.5). Moreover, the CLA-producers showed good adhesion capacity to intestinal human cells (Caco-2 and HT-29) and were able to prevent colonization of Escherichia coli. Of the two strains, Lactobacillus plantarum L3C1E8 revealed superior probiotic properties and great potential for producing food products enriched in the two CLA isomers, cis-9, trans-11 C18:2 (60%) and trans-9, trans-11 C18:2 (25%).

Isolation and screening of Lactobacillus species from dogs for probiotic action

The present study aimed at the detection of suitable Lactobacillus species from dogs for usage as probiotic. A total of 67 rectal swabs from healthy pups were analyzed and 49 (73.1%) Lactobacillus isolates were identified based on morphological, biochemical characteristics and confirmed by genus specific PCR targeting 16S rRNA gene. A total of 20 isolates that showed strong aggregation, high cell surface hydrophobicity, acid and bile tolerance were screened for in-vitro antibacterial activity by agar well diffusion assay, where prominent inhibitory zones were observed against majority of the test pathogens. Reduction in antibacterial activity was noticed after neutralization, proteinase K and heat treatment of supernatants. Partial 16S rRNA nucleotide sequence analysis of seven Lactobacillus isolates that showed prominent in-vitro antibacterial activity revealed maximum sequence homology with Lactobacillus fermentum (for six isolates) and Lactobacillus agilis (for one isolate). The in-vitro antibacterial activity results highlighted the need for in-vivo studies in India to establish probiotic potential of canine faecal Lactobacillus species in the near future.

Variation of mucin adhesion, cell surface characteristics, and molecular mechanisms among Lactobacillus plantarum isolated from different habitats.

The adhesion ability to mucin varied greatly among 18 Lactobacillus plantarum isolates depending on their isolation habitats. Such ability remained at high level even though they were sequentially exposed to the gastrointestinal (GI) stresses. The majority of L. plantarum isolated from shrimp intestine and about half of food isolates exhibited adhesion ability (51.06-55.04%) about the same as the well-known adhesive L. plantarum 299v. Interestingly, five infant isolates of CIF17A2, CIF17A4, CIF17A5, CIF17AN2, and CIF17AN8 exhibited extremely high adhesion ranging from 62.69 to 72.06%. Such highly adhesive property correlating to distinctively high cell surface hydrophobicity was significantly weaken after pretreatment with LiCl and guanidine-HCl confirming the entailment of protein moiety. Regarding the draft genome information, all molecular structures of major cell wall-anchored proteins involved in the adhesion based on L. plantarum WCSF1, including lp_0964, lp_1643, lp_3114, lp_2486, lp_3127, and lp_3059 orthologues were detected in all isolates. Exceptionally, the gene-trait matching between yeast agglutination assay and the relevant mannose-specific adhesin (lp_1229) encoding gene confirmed the Msa absence in five infant isolates expressed distinctively high adhesion. Interestingly, the predicted flagellin encoding genes (fliC) firstly revealed in lp_1643, lp_2486, and lp_3114 orthologues may potentially contribute to such highly adhesive property of these isolates.

Comparative investigation on a hexane-degrading strain with different cell surface hydrophobicities mediated by starch and chitosan.

Bioremediation usually exhibits low removal efficiency toward hexane because of poor water solubility, which limits the mass transfer rate between the substrate and microorganism. This work aimed to enhance the hexane degradation rate by increasing cell surface hydrophobicity (CSH) of the degrader, Pseudomonas mendocina NX-1. The CSH of P. mendocina NX-1 was manipulated by treatment with starch and chitosan solution of varied concentrations, reaching a maximum hydrophobicity of 52%. The biodegradation of hexane conformed to the Haldane inhibition model, and the maximum degradation rate (ν max) of the cells with 52% CSH was 0.72mg (mg cell)-1·h-1 in comparison with 0.47mg (mg cell)-1·h-1 for cells with 15% CSH. The production of CO2 by high CSH cells was threefold higher than that by cells at 15% CSH within 30h, and the cumulative rates of O2 consumption were 0.16 and 0.05mL/h, respectively. High CSH was related to low negative charge carried by the cell surface and probably reduced the repulsive electrostatic interactions between hexane and microorganisms. The FT-IR spectra of cell envelopes demonstrated that the methyl chain was inversely proportional to increasing CSH values, but proteins exhibited a positive effect to CSH enhancement. The ratio of extracellular proteins and polysaccharides increased from 0.87 to 3.78 when the cells were treated with starch and chitosan, indicating their possible roles in increased CSH.

Effect of low-concentration rhamnolipid biosurfactant on Pseudomonas aeruginosa transport in natural porous media.

The effect of low-concentrations of monorhamnolipid biosurfactant on transport of Pseudomonas aeruginosa ATCC 9027 in natural porous media (silica sand and a sandy soil) was studied with miscible-displacement experiments using artificial groundwater as the background solution. Transport of two types of cells was investigated, glucose- and hexadecane-grown cells with lower and higher cell surface hydrophobicity (CSH), respectively. The effect of hexadecane presence as a residual non-aqueous phase liquid (NAPLs) on transport was also examined. A clean-bed colloid deposition model was used to calculate deposition rate coefficients (k) for quantitative assessment. Significant cell retention was observed in the sand (81% and 82% for glucose- and hexadecane-grown cells, respectively). Addition of a low-concentration rhamnolipid solution enhanced cell transport, with 40 mg/L of rhamnolipid reducing retention to 50% and 60% for glucose- and hexadecane-grown cells, respectively. The k values for both glucose- and hexadecane-grown cells correlate linearly with rhamnolipid-dependent CSH represented as bacterial-adhesion-to-hydrocarbon rate of cells. Retention of cells by the soil was nearly complete (>99%). Addition of 40 mg/L rhamnolipid solution reduced retention to 95%. The presence of NAPLs in the sand increased the retention of hexadecane-grown cells with higher CSH. Transport of cells in the presence of the NAPL was enhanced by rhamnolipid at all concentrations tested, and the relative enhancement was greater than in was in the absence of NAPL. This study shows the importance of hydrophobic interaction on bacterial transport in natural porous media and the potential of using low-concentration rhamnolipid for facilitating the transport in subsurface for bioaugmentation efforts.

EVALUATION OF PROBITOIC POTENTIAL OF LACTOBACILLUS STRAINS FROM FERMENTED FOODS AND FEACES OF INFANTS

Lactobacillus strains are a major part of the probiotics, microflora of the intestine and of fermented foods. The aim of this study was to evaluate the potential probiotics of six Lactobacillus strains (L. fermentum 39-183; L. plantarum subsp.plantarum P-8; L. casei ATCC 334; L. rhamnosus ATCC 8530, L. brevis KB 290 and L. fermentum JMC 7776). Probiotic properties such as acid tolerance, bile resistance, bacteriocin-like activity, cell surface hydrophobicity and antibiotic resistance were assessed. In vitro results obtained showed that all Lactobacillus strains tested were able to meet the basic requirements for probiotic functions as they demonstrated probiotic characteristics such as tolerance to pH 2.0 and 2% bile salt. All Lactobacillus strains inhibited the growth of E. coli, Staphylococcus aureus and Salmonella Typhi. Among strains tested, L. plantarum subsp.plantarum P-8 showing inhibitory is very promising with inhibition zone ranging between 6.5 to 12.7 mm. The results for cell surface hydrophobicity and susceptibility against antibiotics also showed that L. fermentum JMC 7776 and L. plantarum subsp.plantarum P-8 had higher cell surface hydrophobicity than the rests. All Lactobacillus tested were resistant to vancomycin and susceptible to streptomycin. The results obtained in this investigation will be used to select potentially probiotic strains for in vivo study

Characterization of fructophilic lactic microbiota of Apis mellifera from the Caucasus Mountains

Microbial symbionts of honeybee colony are considered as promising tools to support the honeybee population welfare. The majority of existing honeybee microbiota studies is focused on genetic description of the honeybee-associated microbiome fingerprints. The lack of a deeper knowledge on the bacterial community colonizing the honeybee niche, which may be helpful in encouraging industrial applications of this microbiota, led us to undertake this study. The biodiversity of the cultivable fructophilic lactic acid bacteria (FLAB) isolated from adult honeybee intestine and beebread samples was studied. Phenotypic properties of probiotic interest, such as the adhesive potential using in vitro models and adhesion determinants, were also investigated. Antibiotic resistance profiles as reliable markers to evaluate the impact of long-term and current exposure of honeybees to antibiotics were phenotypically determined on the isolated lactic acid bacteria (LAB). The mannose-specific adhesion and high cell surface hydrophobicity found in the studied FLAB isolates sheds light on the effective adaptation of microbiota to specific ecologic niche. It is the first report of phenotypically detected antibiotic resistance profiles of honeybee endogenous bacteria and the first account of minimum inhibitory concentration (MIC) values for four antibiotics used in beekeeping practice.

Sphingobium hydrophobicum sp. nov., a hydrophobic bacterium isolated from electronic-waste-contaminated sediment.

Four hydrophobic bacteria were isolated from sediment at Guiyu, an electronic-waste recycling site in southeastern China. The isolates had high cell surface hydrophobicity with microbial-adhesion-to-hydrocarbon score of 71.4 %. 16S rRNA gene sequences of the strains all showed highest similarity to the hydrophilic Sphingobium xenophagum DSM 6383T (99.9 % 16S rRNA gene sequence similarity), followed by Sphingobiumczechense DSM 25410T (97.1 %). However, DNA-DNA hybridization revealed that the isolates and S. xenophagum DSM 6383T exhibited low DNA-DNA relatedness with a hybridization value of 54.5±0.5 %. The genomic DNA G+C content was 64.2 mol% and the predominant quinone was ubiquinone Q-10. Spermidine was the major polyamine component. The major fatty acids were C18 : 1ω7c, C16 : 1ω7c, C16 : 0, C14 : 0 2-OH and C14 : 0. In contrast to its closest relative S. xenophagum DSM 6383T, the isolates had a much higher proportion of C16 : 0 and C14 : 0 and a much lower proportion of C18 : 1ω9t. Sphingoglycolipid was present and diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylmonomethylethanolamine were detected in the polar lipid pattern. Phosphatidyldimethylethanolamine and phosphatidylcholine, which are present in S. xenophagum DSM 6383T, were not detected in the isolates. Results of DNA-DNA relatedness, cell surface hydrophobicity, fatty acids, polar lipids, and biochemical and physiological properties reveal that the isolates represent a novel species of the genus Sphingobium, for which the name Sphingobium hydrophobicum sp. nov. is proposed. The type strain is C1T (=CCTCC AB 2015198T=KCTC 42740T).

The role of Proteus mirabilis cell wall features in biofilm formation.

Biofilms formed by Proteus mirabilis strains are a serious medical problem, especially in the case of urinary tract infections. Early stages of biofilm formation, such as reversible and irreversible adhesion, are essential for bacteria to form biofilm and avoid eradication by antibiotic therapy. Adhesion to solid surfaces is a complex process where numerous factors play a role, where hydrophobic and electrostatic interactions with solid surface seem to be substantial. Cell surface hydrophobicity and electrokinetic potential of bacterial cells depend on their surface composition and structure, where lipopolysaccharide, in Gram-negative bacteria, is prevailing. Our studies focused on clinical and laboratory P. mirabilis strains, where laboratory strains have determined LPS structures. Adherence and biofilm formation tests revealed significant differences between strains adhered in early stages of biofilm formation. Amounts of formed biofilm were expressed by the absorption of crystal violet. Higher biofilm amounts were formed by the strains with more negative values of zeta potential. In contrast, high cell surface hydrophobicity correlated with low biofilm amount.

Aerobic degradation of BDE-209 by Enterococcus casseliflavus: Isolation, identification and cell changes during degradation process

Decabromodiphenyl ether (BDE-209) is one of the most commonly used brominated flame retardants that have contaminated the environment worldwide. Microbial bioremediation has been considered as an effective technique to remove these sorts of persistent organic pollutants. Enterococcus casseliflavus, a gram-positive bacterium capable of aerobically transforming BDE-209, was isolated by our team from sediments in Guiyu, an e-waste dismantling area in Guangdong Province, China. To promote microbial bioremediation of BDE-209 and elucidate the mechanism behind its aerobic degradation, the effects of BDE-209 on the cell changes of E. casseliflavus were examined in this study. The experimental results demonstrated that the high cell surface hydrophobicity (CSH) of E. casseliflavus made the bacteria absorb hydrophobic BDE-209 more easily. E. casseliflavus responded to BDE-209 stress, resulting in an increase in cell membrane permeability and accumulation of BDE-209 inside the cell. The differential expression of intracellular protein was analyzed through two-dimensional gel electrophoresis (2-DE). More than 50 differentially expressed protein spots were reproducibly detected, including 25 up, and 25 down regulated after a 4 days exposure. Moreover, the apoptotic-like cell changes were observed during E. casseliflavus mediated degradation of BDE-209 by means of flow cytometry.

Diverse effect of surfactants on pyrene biodegradation by a Pseudomonas strain utilizing pyrene by cell surface hydrophobicity induction

Pseudomonas aeruginosa is an efficient pyrene degrader that can utilize pyrene through induction of high cell surface hydrophobicity. The effect of three surfactants, Triton X-100, Tween 80 and rhamnolipid JBR 515 on rate and extent of pyrene (100 mg l−1) degradation was evaluated through batch studies. Among the three surfactants Triton X-100 demonstrated highest pyrene solubilization. All surfactants were used at 10 times the critical micelle concentration (CMC) so as to ensure notable enhancement in solubilization. However, growth of P. aeruginosa on pyrene and pyrene degradation was inhibited in the presence of Triton X-100 possibly due to accumulation of toxic metabolites formed due to partial utilization of Triton X-100. Tween 80 supported higher rate of pyrene degradation due to higher biomass generation during simultaneous utilization of pyrene and Tween 80 as carbon source. In presence of rhamnolipid JBR 515 preferential degradation of surfactant over pyrene resulted in decrease in specific growth rate and pyrene utilization rate. Maximum biodegradation rates (MBR) for pyrene were 0.458, 1.111 and 0.154 mg l−1 h−1 in absence of any surfactant, in presence of Tween 80 and rhamnolipid JBR 515, respectively. The results suggest that Tween 80 is the preferred surfactant for remediation of pyrene using P. aeruginosa RS1 strain.

Biosurfactant from Pseudomonas species with waxes as carbon source – Their production, modeling and properties

This study reports the production of biosurfactant from Pseudomonas aeruginosa and Pseudomonas fluorescens with waxes namely, n-hexadecane and n-eicosane as carbon sources. The latter organism produced 9.8 and 8.2gL−1 of rhamnolipid containing C8 and C10 fatty acid with n-eicosane and n-hexadecane as substrates, respectively, whereas, the former produced 5.7 and 11.5gL−1 of C10 and C12 fatty acids. Both the microorganisms were observed to degrade 85–90% of the waxes in one day and achieved very high cell surface hydrophobicity in a short period and indicates their ability of quick adherence to hydrophobic surfaces. The produced biosurfactant was stable upto 100°C temperature, 8MPa pressure, 20% (w/v) salinity and alkaline pH. Reaction kinetics was mathematically modeled using the Monod and Logistic growth model. The Logistic growth model was observed to fit the experimental values of biomass formed, biosurfactant produced, and the paraffins (waxes) degraded satisfactorily. The enhanced physico-chemical properties of biosurfactant with paraffin substrates, such as high bacterial adherence, wax degrading ability, emulsification activities, and surface tension reducing ability indicates the potential of these organisms in addressing oil spill, flow assurance and enhanced oil recovery process.

Survival of vancomycin-intermediate Staphylococcus aureus on hospital surfaces

Contaminated surfaces play an important role in the transmission of certain pathogens that are responsible for healthcare-associated infections. Although previous studies have shown that meticillin-resistant Staphylococcus aureus (MRSA) can survive on dry surfaces at room temperature, no published data regarding vancomycin-intermediate S. aureus (VISA) are available to date. To compare the survival time on different types of surfaces, cell-surface hydrophobicity, adherence to abiotic surfaces and biofilm formation of meticillin-susceptible S. aureus (MSSA), MRSA and VISA. Survival of the S. aureus strains was tested on latex, cotton fabric, vinyl flooring and formica. Cell-surface hydrophobicity was determined using the hydrocarbon interaction affinity method. Adhesion to abiotic surfaces was tested on granite, latex (gloves), glass, vinyl flooring and formica. Biofilm formation was evaluated at 6, 12, 24 and 48 h. All of the samples survived on the vinyl flooring and formica for at least 40 days. VISA survived on both surfaces for more than 45 days. All of the strains were highly hydrophobic. VISA adhered to latex, vinyl flooring and formica. Biofilm formation increased for all of the tested strains within 6-24 h. VISA present high survival, adherence and cell-surface hydrophobicity. Therefore, as the treatment of patients with VISA is a significant challenge for clinicians, greater care with cleaning and disinfection of different types of surfaces in healthcare facilities is recommended because these may become important reservoirs of multi-resistant pathogens.

Distribution of Candida albicans and non-albicans Candida species in oral candidiasis patients: Correlation between cell surface hydrophobicity and biofilm forming activities

ObjectivesThe purposes of this investigation were to study the prevalence of Candida albicans and non-albicans Candida (NAC) species from oral candidiasis patients and evaluate the cell surface hydrophobicity (CSH) and biofilm forming capacity of the clinical isolates Candida species from oral cavity. DesignThis study identified a total of 250 Candida strains isolated from 207 oral candidiasis patients with PCR-RFLP technique. CSH value, total biomass of biofilm and biofilm forming ability of 117 oral Candida isolates were evaluated. ResultsC. albicans (61.6%) was still the predominant species in oral candidiasis patients with and without denture wearer, respectively, followed by C. glabrata (15.2%), C. tropicalis (10.4%), C. parapsilosis (3.2%), C. kefyr (3.6%), C. dubliniensis (2%), C. lusitaniae (2%), C. krusei (1.6%), and C. guilliermondii (0.4%). The proportion of mixed colonization with more than one Candida species was 18% from total cases. The relative CSH value and biofilm biomass of NAC species were greater than C. albicans (p<0.001). Ninety-two percent of oral isolates NAC species had biofilm forming ability, whereas 78% of C. albicans were biofilm formers. Furthermore, the significant difference of relative CSH values between biofilm formers and non-biofilm formers was observed in the NAC species (p<0.005), whereas the difference was not statistically significant in C. albicans. ConclusionThe frequency of the NAC species colonization in oral cavity was gradually increasing. The possible contributing factors might be high cell surface hydrophobicity and biofilm forming ability. The relative CSH value could be a putative factor for determining biofilm formation ability of the non-albicans Candida species.

Formation and characterisation of aerobic sludge aggregates in a lab-scale activated sludge system

Sludge aggregates were developed under ambient conditions in the aeration tank of a lab-scale activated sludge system. Different strategies were employed to accelerate the aggregation process viz.: i) initiating aggregation with large-dense flocs; ii) supplementing sewage with food-waste leachate containing readily biodegradable substrates; iii) periodic aerobic starvation; iv) addition of support materials to nucleate aggregate formation. Four identical reactors were set-up, each with a different support material viz.: R1: control; R2: unburnt carbon from bagasse flyash; R3: commercial activated carbon; R4: zeolite 4A. Optical and scanning electron microscopy, supported by particle size measurement by sieving indicated the development of aggregates with mean diameter of 0.5 mm in all the four reactors after 90 days of continuous operation. The aggregates showed improved physical properties (high cell-surface hydrophobicity, high integrity coefficient, low sludge volume index, high settling velocity) and chemical properties (high extracellular polymeric substances or EPS content). The process resulted in an average of 72% to 76% COD reduction with a biomass concentration of 8 to 9 g/L in the reactor. Interestingly, introduction of the support materials into the aeration tank did not enhance the aggregation process.

Substrate Uptake and Subcellular Compartmentation of Anoxic Cholesterol Catabolism in Sterolibacterium denitrificans

Cholesterol catabolism by actinobacteria has been extensively studied. In contrast, the uptake and catabolism of cholesterol by Gram-negative species are poorly understood. Here, we investigated microbial cholesterol catabolism at the subcellular level. (13)C metabolomic analysis revealed that anaerobically grown Sterolibacterium denitrificans, a β-proteobacterium, adopts an oxygenase-independent pathway to degrade cholesterol. S. denitrificans cells did not produce biosurfactants upon growth on cholesterol and exhibited high cell surface hydrophobicity. Moreover, S. denitrificans did not produce extracellular catabolic enzymes to transform cholesterol. Accordingly, S. denitrificans accessed cholesterol by direction adhesion. Cholesterol is imported through the outer membrane via a putative FadL-like transport system, which is induced by neutral sterols. The outer membrane steroid transporter is able to selectively import various C27 sterols into the periplasm. S. denitrificans spheroplasts exhibited a significantly higher efficiency in cholest-4-en-3-one-26-oic acid uptake than in cholesterol uptake. We separated S. denitrificans proteins into four fractions, namely the outer membrane, periplasm, inner membrane, and cytoplasm, and we observed the individual catabolic reactions within them. Our data indicated that, in the periplasm, various periplasmic and peripheral membrane enzymes transform cholesterol into cholest-4-en-3-one-26-oic acid. The C27 acidic steroid is then transported into the cytoplasm, in which side-chain degradation and the subsequent sterane cleavage occur. This study sheds light into microbial cholesterol metabolism under anoxic conditions.

Characteristics of Lactic Acid Bacteria Isolated from Gastrointestinal Tract of Cemani Chicken and Their Potential Use as Probiotics

The aims of this study were to screen and characterize lactic acid bacteria (LAB) isolated from gastrointestinal (GI) tract of Cemani chicken, one of Indonesian local chicken and to investigate their potential use as probiotics. LAB were isolated from GI tract using MRSA and GYPA media and incubated anaerobically. Selected LAB were determined their probiotic properties with several assays. Identification of selected LAB was based on 16S rDNA sequences, morphological and biochemical characteristics. Ninety five bacteria were isolated and characterized as lactic acid bacteria (Gram positive, catalase negative, non sporeforming and acid producing). Twenty four isolates of LAB demonstrated antimicrobial activity against Escherichia coli JCM 1649 and Salmonella enteritidis B2586, and three selected isolates, i.e. CCM011, CSP004, and CVM002 showed the highest inhibition activity. The isolates had characters of high cell surface hydrophobicity and inter-isolate coaggregation ability of LAB, high survival at low pH, high phytase and protease activity (but no amylase and lipase activity), weak coaggregation with pathogen and no resistance to the examined antibiotics. The isolates were identified based on sequence analysis of 16S rRNA gene as Lactobacillus salivarius , however, each isolate had different profiles of sugar fermentation. Therefore the three LAB isolates had potential application as probiotics for chicken. Key words: Cemani chicken, gastrointestinal tract, lactic acid bacteria, probiotic