Biomedical subjects

F M Rombouts

Publications and source records attributed to F M Rombouts.

At least 19 recordsLinked to original sources

PubMed1996

Model for the combined effects of temperature, pH, and sodium lactate on growth rates of Listeria innocua in broth and Bologna-type sausages.

A modified Monod equation was successfully applied to describe the maximum specific growth rate of Listeria innocua in a broth model in the presence of various concentrations of sodium lactate or NaCl. The combined effects of temperature and pH were assessed by translating the parameters of the modified Monod equation mu(m), alpha, and p') as functions of pH and/or temperature. As a result, the area in which the growth rate could be predicted was extended to include as a variable not only the salt concentration but also pH and temperature. The number of parameters needed to describe the experimental data was thereby reduced from 48 to 4 (NaCl) and from 42 to 5 (sodium lactate). The decline in the goodness of fit that accompanied the reduction in the number of parameters was within statistically acceptable ranges. The resulting model was compared with a polynomial fit, and it was proposed that the former was more suitable for the purpose of this study. The broth model for sodium lactate was evaluated with Bologna-type sausages. Because of the "worst-case" design of the broth model, it was necessary to reestimate one or all parameters to obtain a good description of the growth rate of L. innocua in the meat product. However, the simplicity of the model and the practical usefulness of its parameters offer considerable prospects for its use in predictive microbiology.

Food Microbiology↗

PubMed1995

Antibiotic susceptibility of campylobacter isolates from sewage and poultry abattoir drain water.

In this study, the in vitro susceptibility of 209 campylobacter strains to the quinolones nalidixic acid, flumequine, ciprofloxacin, enrofloxacin, and to ampicillin, tetracycline and erythromycin was tested by the disk diffusion method. The strains were isolated from poultry abattoir effluent (DWA) and two sewage purification plants (SPA and SPB). Sewage purification plant SPA received mixed sewage, including that from a poultry abattoir, whereas SPB did not receive sewage from any meat-processing industry. The quinolone resistance of the DWA isolates ranged from 28% for enrofloxacin to 50% for nalidixic acid. The strains isolated from the sewage purification plants were more susceptible to the quinolones with a range of 11-18% quinolone resistance for SPB isolates to 17-33% quinolone resistance for SPA isolates. The susceptibility criteria as recommended by National Committee Clinical Laboratory Standards (USA) cannot readily be employed for campylobacter isolates. This investigation shows that the resistance of campylobacter bacteria is highest in the plant receiving sewage from a poultry slaughterhouse. Monitoring of antibiotic resistance of aquatic Campylobacter spp. is important, as surface waters are recognized as possible sources of infection.

Abattoirs↗

PubMed1995

The speciation and subtyping of campylobacter isolates from sewage plants and waste water from a connected poultry abattoir using molecular techniques.

In this study the distribution of phenotypes of campylobacter strains in sewage and surface waters was investigated by subtyping and by speciation of isolates from various aquatic environments. These environments included two municipal sewage plants (SPA and SPB) and waste water from a poultry abattoir (WWA). Both the sewage plants SPA and SPB collected domestic and industrial waste, and SPA received drain water from WWA. SPB received no waste water from any meat-processing plant. The isolates were speciated by PCR and subtyped by PCR/RFLP based on the flagellin PCR products. From all three reservoirs, no Campylobacter lari was isolated, and approximately 80% of the isolates could be identified as C. jejuni and the rest belonged to the C. coli species. The PCR/RFLP typing technique has a high discrimination level and was reproducible between two separate laboratories. The 182 isolates tested yielded 22 distinct Dde I profiles. The results indicate that strains with profiles found in poultry are also detectable in waste water presumed to be solely from domestic and human sources. In addition some strains were unique to the known poultry-related sources, suggesting that avian-specific strains, non-pathogenic to man, may exist in the environment. In contrast some strains were unique to human waste indicating the potential importance of non-poultry sources of infection. No seasonality was observed in the profile distribution. So, at least in the Netherlands, it is unlikely that infections caused by contaminated surface waters contribute to the seasonality of human campylobacteriosis.

Abattoirs↗

PubMed1995

Degradation and fermentation of fructo-oligosaccharides by oral streptococci.

Fructo-oligosaccharides (FOS) are claimed to have a positive effect on the intestinal flora. They are being used in functional foods in Japan and Europe. This group have tested the degradation of two commercial FOS preparations by oral streptococci in order to predict the cariogenicity of these products. Both preparations could be fermented to some extent by the species of oral streptococci tested. The enzymes necessary for the degradation of FOS were inducible. Each strain showed a specific degradation pattern. All strains, particularly Streptococcus mutans rapidly produced acid, mainly lactic acid. Streptococcus mitis also produced high concentrations of acetic acid. Plaque formation by Strep. mutans was similar to the sucrose control. It is concluded that FOS are cariogenic to a similar extent as sucrose.

Acetates↗

PubMed1995

A di- and tripeptide transport system can supply Listeria monocytogenes Scott A with amino acids essential for growth.

Listeria monocytogenes takes up di- and tripeptides via a proton motive force-dependent carrier protein. This peptide transport system resembles the recently cloned and sequenced secondary di- and tripeptide transport system of Lactococcus lactis (A. Hagting, E. R. S. Kunji, K. J. Leenhouts, B. Poolman, and W. N. Konings, J. Biol. Chem. 269:11391-11399, 1994). The peptide permease of L. monocytogenes has a broad substrate specificity and allows transport of the nonpeptide substrate 5-aminolevulinic acid, the toxic di- and tripeptide analogs, alanyl-beta-chloroalanine and alanyl-alanyl-beta-chloroalanine, and various di- and tripeptides. No extracellular peptide hydrolysis was detected, indicating that peptides are hydrolyzed after being transported into the cell. Indeed, peptidase activities in response to various synthetic substrates were detected in cell extracts obtained from L. monocytogenes cells grown in brain heart infusion broth or defined medium. The di- and tripeptide permease can supply L. monocytogenes with essential amino acids for growth and might contribute to growth of this pathogen in various foods where peptides are supplied by proteolytic activity of other microorganisms present in these foods. Possible roles of this di- and tripeptide transport system in the osmoregulation and virulence of L. monocytogenes are discussed.

Amino Acids↗

PubMed1995

Characterization of uptake and hydrolysis of fluorescein diacetate and carboxyfluorescein diacetate by intracellular esterases in Saccharomyces cerevisiae, which result in accumulation of fluorescent product.

Flow cytometry is a rapid and sensitive method which may be used for the detection of microorganisms in foods and drinks. A key requirement for this method is a sufficient fluorescence staining of the target cells. The mechanism of staining of the yeast Saccharomyces cerevisiae by fluorescein diacetate (FDA) and 5- (and 6-)carboxyfluorescein diacetate (cFDA) was studied in detail. The uptake rate of the prefluorochromes increased in direct proportion to the concentration and was not saturable, which suggests that transport occurs via a passive diffusion process. The permeability coefficient for cFDA was 1.3 x 10(-8) m s-1. Once inside the cell, the esters were hydrolyzed by intracellular esterases and their fluorescent products accumulated. FDA hydrolysis (at 40 degrees C) in cell extracts could be described by first-order reaction kinetics, and a rate constant (K) of 0.33 s-1 was calculated. Hydrolysis of cFDA (at 40 degrees C) in cell extracts was described by Michaelis-Menten kinetics with an apparent Vmax and Km of 12.3 nmol.min-1.mg of protein-1 and 0.29 mM, respectively. Accumulation of fluorescein was most likely limited by the esterase activity, since transport of FDA was faster than the hydrolysis rate. In contrast, accumulation of carboxyfluorescein was limited by the much slower transport of cFDA through the cell envelope. A simple mathematical model was developed to describe the fluorescence staining. The implications for optimal staining of yeast cells with FDA and cFDA are discussed.

Biological Transport, Active↗

PubMed1995

Temperature-dependent membrane fatty acid and cell physiology changes in coccoid forms of Campylobacter jejuni.

The effect of temperature and the availability of nutrients on the transition of spiral Campylobacter jejuni cells to coccoid forms was investigated. Ageing of spiral C. jejuni cells in either nutrient-poor or nutrient-rich environments resulted in the formation of nonculturable coccoid cells at 4, 12, and 25 degrees C after different periods, with the cells incubated at 4 degrees C in nutrient-deficient media remaining culturable the longest. To study the phenomenon, ATP levels, protein profiles, and fatty acid compositions were monitored under conditions where the transition from spiral to coccoid cells occurred. During storage, the levels of intracellular ATP were highest in cells incubated at low temperatures (4 and 12 degrees C) and remained constant after a small initial decrease. During the transformation from spiral to coccoid forms, no alteration in protein profiles could be detected; indeed, inhibition of protein synthesis by chloramphenicol did not influence the transition. Furthermore, DNA damage by gamma irradiation had no effect on the process. Membrane fatty acid composition of cocci formed at low temperatures was found to be almost identical to that of spiral cells, whereas that of cocci formed at 25 degrees C was clearly different. Combining these results, it is concluded that the formation of cocci is not an active process. However, distinctions between cocci formed at different temperatures were observed. Cocci formed at 4 degrees C show characteristics comparable to those of spirals, and these cocci may well play a role in the contamination cycle of C. jejuni.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗

PubMed1995

An ATP-dependent L-carnitine transporter in Listeria monocytogenes Scott A is involved in osmoprotection.

Listeria monocytogenes is a gram-positive, psychotrophic, food-borne pathogen which is able to grow in osmotically stressful environments. Carnitine (beta-hydroxy-L-tau-N-trimethyl aminobutyrate) can contribute significantly to growth of L. monocytogenes at high osmolarity (R. R. Beumer, M. C. te Giffel, L. J. Cox, F. M. Rombouts, and T. Abee, Appl. Environ. Microbiol. 60:1359-1363, 1994). Transport of L-[N-methyl-14C]carnitine in L. monocytogenes was shown to be energy dependent. Analysis of cell extracts revealed that L-carnitine was not further metabolized, which supplies evidence for its role as an osmoprotectant in L. monocytogenes. Uptake of L-carnitine proceeds in the absence of a proton motive force and is strongly inhibited in the presence of the phosphate analogs vanadate and arsenate. The L-carnitine permease is therefore most likely driven by ATP. Kinetic analysis of L-carnitine transport in glucose-energized cells revealed the presence of a high-affinity uptake system with a Km of 10 microM and a maximum rate of transport (Vmax) of 48 nmol min-1 mg of protein-1. L-[14C]carnitine transport in L. monocytogenes is significantly inhibited by a 10-fold excess of unlabelled L-carnitine, acetylcarnitine, and tau-butyrobetaine, whereas L-proline and betaine display, even at a 100-fold excess, only a weak inhibitory effect. In conclusion, an ATP-dependent L-carnitine transport system in L. monocytogenes is described, and its possible roles in cold adaptation and intracellular growth in mammalian cells are discussed.

Adenosine Triphosphate↗

PubMed1995

The effects of processing on the availability of lysine in kenkey, a Ghanaian fermented maize food.

The effects of processing steps such as soaking, fermentation, cooking and drying on the availability of lysine in kenkey were investigated. Soaking increased lysine availability by 21% and 22% for maize and maize-cowpea mixtures, respectively. Cooking of soaked samples further improved lysine availability by 68% and 31% for maize and maize-cowpea mixtures, respectively. Further significant improvements in lysine availability were effected by fermentation and cooking and values of 3.42 and 4.43 g/16 g N were recorded, respectively for maize and maize-cowpea doughs fermented for 4 days and cooked for 3 h. Cabinet drying had no significant effect on lysine availability, but drum drying of fermented maize and maize-cowpea doughs significantly lowered lysine availability in the resulting kenkey. A 1:1 mixture of cabinet and drum dried flours gave a product with higher available lysine content than the drum dried flour.

Biological Availability↗

PubMed1994

2-O-methyl-D-mannose residues are immunodominant in extracellular polysaccharides of Mucor racemosus and related molds.

In this study, the structure of the immunodominant carbohydrate epitope of the extracellular polysaccharides from mold species belonging to the order Mucorales reactive with rabbit IgG antibodies was elucidated. An exo-alpha-D-mannanase which was able to abolish the antigenicity of these polysaccharides completely was purified and characterized, and the activity was compared with that of an alpha-D-mannosidase. Analysis of the monomeric reaction products after enzymatic treatment revealed the presence of 2-O-methyl-D-mannose residues. This compound is a constituent of the polysaccharides from the mold genera Mucor, Rhizopus, Rhizomucor, Absidia, Syncephalastrum, and Thamnidium, and its occurrence in fungi has not been reported until now. Two mannan fractions which are highly reactive with rabbit IgG were isolated from the extracellular polysaccharides of Mucor racemosus and characterized with ethylation analysis. The role of the newly found 2-O-methyl-D-mannose residues in the immunoreactivity was assessed by specific degradation of these mannans with the exo-alpha-D-mannanase and subsequent ethylation analysis. It was concluded that the immunodominant carbohydrates reactive with rabbit IgG are chains composed of a single terminal non-reducing 2-O-methyl-D-mannose residue, alpha (1-2)-linked to a short sequence of alpha(1-2)-linked D-mannose residues.

Antibodies, Fungal↗

PubMed1994

Modelling growth rates of Listeria innocua as a function of lactate concentration.

The effect of sodium lactate (NaL) concentration on growth of Listeria innocua in a yeast-extract/peptone broth at pH 5.5, 6.0, 6.5 and 7.0 at 4, 10, 20 and 30 degrees C was modelled with the modified Gompertz model. NaCl was used as a reference to distinguish between the water activity effect and the specific inhibitory effect of NaL. Minimum inhibitory concentrations (MIC) of NaCl appeared to be significantly higher than MIC values of NaL, indicating that NaL had a specific inhibitory effect on growth of L. innocua. The MIC values of NaL and NaCl were not much influenced by the temperature. The pH of the growth medium was shown to have influence on the MIC values of NaL but not on the MIC values of NaCl. Total growth inhibition of L. innocua at low pH (5.5) took place at lower NaL concentrations (217 mM) than at neutral pH (1071-1339 mM), indicating that the undissociated lactic acid plays a role in the mechanism of inhibition. However, MIC values for undissociated acid increased with decreasing pH from 0.8 mM at pH 7 to 5 mM at pH 5.5. It is therefore likely that besides acidification of the cytoplasm due to diffusion of undissociated acid into the cell, other mechanisms are involved. Growth rates at NaL concentrations between 0 and the MIC value decreased progressively with increasing concentrations down to 0 at the MIC value, and were strongly influenced by both temperature and pH. Growth rates in the presence of NaCl were influenced by the temperature only. It was shown that a modified Monod equation with three parameters was effective for description of growth rates of L. innocua at NaL and NaCl concentrations over the whole experimental range.

Cell Division↗

PubMed1994

Effect of exogenous proline, betaine, and carnitine on growth of Listeria monocytogenes in a minimal medium.

Three Listeria monocytogenes strains isolated from food or food-processing environments were used to assess the response of this species to salinity in a chemically defined minimal medium. Growth in a minimal medium containing five essential amino acids and glucose as a carbon and energy source was comparable to growth in a rich medium (brain heart infusion broth). In the absence and presence of 3% NaCl the final cell numbers reached in minimal medium were 10(9) and 10(7) CFU/ml, respectively. Growth under the latter conditions could not be detected by spectrophotometry by measuring A660. Apparently, this technique was not suitable for these experiments since the detection level was > 10(7) CFU/ml. Exogenously added proline (10 mM), trimethylglycine (betaine) (1 mM), and beta-hydroxy-gamma-N-trimethyl aminobutyrate (carnitine) (1 mM) significantly stimulated growth under osmotic stress conditions in minimal medium at both 37 and 10 degrees C. Betaine and carnitine are present in foods derived from plants and animals, respectively. Therefore, these compounds can contribute significantly to growth of L. monocytogenes in various foods at high osmolarities.

Amino Acids↗

PubMed1994

Energy-dependent, carrier-mediated extrusion of carboxyfluorescein from Saccharomyces cerevisiae allows rapid assessment of cell viability by flow cytometry.

Carboxyfluorescein diacetate is a nonfluorescent compound which can be used in combination with flow cytometry for vital staining of yeasts and bacteria. The basis of this method is the assumption that, once inside the cell, carboxyfluorescein diacetate is hydrolyzed by nonspecific esterases to produce the fluorescent carboxyfluorescein (cF). cF is retained by cells with intact membranes (viable cells) and lost by cells with damaged membranes. In this report, we show that Saccharomyces cerevisiae extrudes cF in an energy-dependent manner. This efflux was studied in detail, and several indications that a transport system is involved were found. Efflux of cF was stimulated by the addition of glucose and displayed Michaelis-Menten kinetics. A Km for cF transport of 0.25 mM could be determined. The transport of cF was inhibited by the plasma membrane H(+)-ATPase inhibitors N,N'-dicyclohexylcarbodiimide and diethylstilbestrol and by high concentrations of tetraphenylphosphonium ions. These treatments resulted in a dissipation of the proton motive force, whereas the intracellular ATP concentration remained high. Transport of cF is therefore most probably driven by the membrane potential and/or the pH gradient. The viability of S. cerevisiae was determined by a two-step procedure consisting of loading the cells with cF followed by incubation at 40 degrees C in the presence of glucose. Subsequently, the fluorescence intensity of the cells was analyzed by flow cytometry. The efflux experiments showed an excellent correlation between the viability of S. cerevisiae cells and the ability to translocate cF. This method should prove of general utility for the rapid assessment of yeast vitality and viability.

Adenosine Triphosphatases↗

PubMed1993

Characterization and frequency distribution of species of lactic acid bacteria involved in the processing of mawè, a fermented maize dough from Benin.

Lactic acid bacteria involved in the natural fermentation of both home-produced and commercial mawè were investigated during a 72 h fermentation period. Lactobacillus spp. constitute the majority (94%) of the strains of the lactic acid bacteria isolated, among which 89% represent the Betabacterium group. They include L. fermentum (biotype cellobiosus) (41%), L. fermentum or L. reuteri (19%), L. brevis (26%), L. confusus (less than 2%), L. curvatus (less than 1%) and L. buchneri (less than 1%). Other isolated lactic acid bacteria were L. salivarius, Lactococcus lactis, Pediococcus pentosaceus, Pediococcus acidilactici and Leuconostoc mesenteroides. Several species were detected at the early stage of fermentation, but the final stage was dominated by L. fermentum (biotype cellobiosus) and L. fermentum or L. reuteri totalling 90% of the isolated strains. The trend was the same for both home-produced and commercial mawè. No strains of L. plantarum, generally reported as dominating lactic acid bacteria at the final stage of fermentation of most plant foods, were isolated.

Benin↗

PubMed1993

Minimum inhibitory concentration (MIC) of sodium lactate for pathogens and spoilage organisms occurring in meat products.

Pathogens and spoilage organisms occurring in meat products were screened in laboratory media, according to a method of Eklund (1983), in order to determine the specific inhibitory effect of lactate on growth of these microorganisms under optimum growth conditions (pH 6.5, 20 degrees C). In general, Gram-positive bacteria were more sensitive towards lactate than Gram-negative bacteria. It was shown especially, that strains that were able to grow at water activities of 0.95 and below in the presence of NaCl (Staphylococcus aureus, Listeria monocytogenes, Brochothrix thermosphacta) were inhibited by sodium lactate (NaL). It appeared, that yeasts were resistant to large amounts of NaL (> 10% w/v). However, NaL had a specific inhibitory effect on growth of these organisms when compared to the effect of NaCl. These results indicate that lactate addition to food products with a pH near neutrality offers good prospects for shelf life prolongation.

Animals↗

PubMed1993

Inhibition of the growth of yeasts in fermented salads.

Salads composed of vegetables and/or meat in an oil-in-water emulsion were prepared by fermentation for 7 h at 42 degrees C or 45 degrees C with strains of Lactobacillus spp. Their stability towards spoilage yeasts was studied using Saccharomyces cerevisiae, Saccharomyces exiguus and Torulaspora delbrueckii, isolated from salads, as well as Pichia membranaefaciens and Zygosaccharomyces bailii. Salads fermented with good lactic starters usually had pH values of < or = 4.2 and lactic acid concentrations of 0.28 to 0.43% (w/w). High numbers of spoilage yeasts (and production of large volumes of CO2) were not attained in these salads, provided the initial concentration of spoilage yeasts was sufficiently low (< or = 100 CFU/g). Inhibition of spoilage yeasts in lactic fermented salads is probably due to lactic acid, the low storage temperature and the low residual oxygen concentration.

Animals↗

PubMed1993

The production and partial characterization of a monoclonal IgG antibody specific for moulds belonging to the order Mucorales.

A monoclonal antibody (mAb) was raised against extracellular polysaccharides from Mucor racemosus after intrasplenic immunization of mice. An indirect ELISA and a dot-blot assay were developed with this mAb. The IgG antibody was found to be very specific for all mould species tested belonging to the order of Mucorales, except species belonging to the genus Mortierella sensu stricto. No cross-reactions were observed with other moulds or yeasts. The immunoreactivity of the polysaccharides of these moulds with this mAb is based on carbohydrate epitopes, in which fucose residues probably play an important role. The mAb may be suited for specific detection of species of the genera Mucor, Rhizopus, Rhizomucor, Thamnidium, Absidia, Syncephalastrum and species belonging to the Mortierella isabellina group in food, and possibly for diagnosis of mucormycosis in humans.

Animals↗

PubMed1992

Detection of fungal carbohydrate antigens by high-performance immunoaffinity chromatography using a protein A column with covalently linked immunoglobulin G.

Fungal carbohydrate antigens were analysed by high-performance immunoaffinity chromatography (HPIAC) with immunoglobulin G (IgG) antibodies raised against extracellular polysaccharides of Mucor racemosus. The protein A-IgG complex was covalently bound with dimethyl pimelimidate, which enabled the use of strong acidic buffers to release the tightly bound antigens from the column. Prior to pulsed-amperometric detection, an anion-micromembrane suppressor was used to raise the pH of the effluent to above 12 without dilution. The HPIAC system provides a sophisticated method for the rapid and sensitive detection of antigenic oligomeric carbohydrates in biological samples and is proposed as an alternative to quantitative enzyme-linked immunosorbent assay techniques.

Antigens, Fungal↗