Evaluation of lysine-iron-cystine-neutral red for the detection of salmonellas in the Bactometer

Ninety-five salmonellas and 40 non-salmonellas were screened in the Bactometer using lysine-iron-cystine-neutral red (LICNR) in order to evaluate the selectivity of the medium for the detection of salmonellas. Results for blackening of the medium in the well (indicating hydrogen sulphide, H₂S, production) and step size are presented. Five out of 95 salmonellas tested failed to produce blackening in the well, four of these five are known to be non-H₂S producers. Although salmonellas generally gave a larger step value than non-salmonellas, this criterion could not be used to distinguish reliably the two groups of organisms.     

Rapid impedance detection of salmonellas in confectionery using modified LICNR broth

A commercially available broth, with the addition of inhibitors, was used for the rapid impediometric detection of salmonellas in confectionery. Pre-enrichment in skimmed milk was followed by lysine-iron-cystine-neutral red broth in a Bactometer 123 system. Results were obtained 3d earlier than is possible with conventional microbiological tests. Some false positives were obtained predominantly with Citrobacter freundii but this problem is also frequently encountered with traditional methods. Organisms responsible for false positives may be isolated and identified more rapidly than is possible by conventional methods.     

Rapid impedance detection of salmonellas in confectionery using modified LICNR broth

A commercially available broth, with the addition of inhibitors, was used for the rapid impediometric detection of salmonellas in confectionery. Pre-enrichment in skimmed milk was followed by lysine-iron-cystine-neutral red broth in a Bactometer 123 system. Results were obtained 3 d earlier than is possible with conventional microbiological tests. Some false positives were obtained predominantly with Citrobacter freundii, but this problem is also frequently encountered with traditional methods. Organisms responsible for false positives may be isolated and identified more rapidly than is possible by conventional methods.     

A note on the isolation of salmonellas from environmental samples using three formulations of Rappaport's broth

The efficiency of isolation of salmonellas from 412 seagull intestines and 247 polluted water samples was compared using three formulations of Rappaport's medium (RV, RV-soya and R25). A modification of RV medium (RV-soya) was shown to be the most efficient of the three media and it was shown that the duration of incubation of this medium could be restricted to 24 h. Inoculation ratios were compared for RV and RV-soya and 1:100 was shown to be significantly better than 1:20. It is concluded that RV-soya is at least as efficient as the standard RV medium and its use can therefore be recommended.     

Voltammetric detection of trimethylamine using immobilized trimethylamine dehydrogenase on an electrodeposited goldnanoparticle electrode

A voltammetric enzyme electrode was developed based on nicotinamide-independent trimethylamine dehydrogenase (TMADH, EC 1.5.99.7), which catalyses the oxidation of trimethylamine (TMA) to dimethylamine and formaldehyde. A quaternized osmium hydrogel polymer, poly(vinylimidazole-[Os(4,4′-dimethyl-2,2′-bipyridine)₂Cl]^+/2+) with ethylamine (PVI-Os-EA), was prepared as a potential redox mediator in an electrochemical biosensor. TMA was detected using TMADH that was co-immobilized with an osmium hydrogel polymer on electrodeposited gold nanoparticles (Au-NPs) on screen-printed carbon electrodes (SPCEs). The Au-NPs deposited onto SPCEs provided about a three times higher electrochemical response compared to that of a planar gold electrode. As TMA was catalyzed by wired TMADH, the electrical signal was monitored at 0.3 V versus Ag/AgCl by cyclic voltammetry and chronoamperometry. The anode currents increased linearly in proportion to the TMA concentration over the 0 ∼ 2.5 mM range with a detection limit of 1 μM (R = 0.9972).     

Effect of carbohydrate source on alfalfa somatic embryogenesis

Cultures of alfalfa (Medicago sativa L. clone RA-3) were screened for their response to a wide variety of carbohydrate sources in the presence or absence of sucrose. Maltose, maltotriose, and soluble starch all act to improve the morphology and eventual conversion to plantlets of somatic embryos. Glucose, sucrose, and other carbohydrates do not have a similar effect. Additional studies were carried out with maltose since this carbohydrate also gives the highest embryo yield. The concentration optimum for maltose is about 4% (w/v). Maltose acts independently of sucrose and in a way which suggests that it serves as a nutritional, rather than as an osmotic, effector of embryogenesis. The effect of maltose on embryogenesis is dependent on the presence of NH₄⁺ whose optimum is approximately 15 mM. Embryogenesis on maltose will not occur in the absence of NH₄⁺. A highly effective regeneration medium can be achieved by including NH₄⁺ and amino acids, especially proline, in a maltose-containing regeneration medium. Maltose is also useful in increasing embryo formation in genotypes which show low regeneration.     

Rapid detection of salmonellas in raw meats using a fluorescent antibody-microcolony technique

A fluorescent antibody-microcolony technique was developed for the rapid detection of salmonellas in pure cultures. Examination of microcolonies made the detection of salmonellas by epifluorescence microscopy easier and more reliable than using fluorescent antibody and single cells. After a study of the most effective selective enrichment media for increasing the number of salmonellas, the technique was examined with various samples of raw meats. It was able to detect salmonellas in 24 h and appeared to be as sensitive as conventional cultural techniques. Of the 101 samples studied, complete agreement was obtained with conventional methods for 94 but six apparently false positive results and one false negative result occurred. and accepted 22 June 1989     

Rapid detection of salmonellas in raw meats using a fluorescent antibody-microcolony technique

A fluorescent antibody-microcolony technique was developed for the rapid detection of salmonellas in pure cultures. Examination of microcolonies made the detection of salmonellas by epifluorescence microscopy easier and more reliable than using fluorescent antibody and single cells. After a study of the most effective selective enrichment media for increasing the number of salmonellas, the technique was examined with various samples of raw meats. It was able to detect salmonellas in 24 h and appeared to be as sensitive as conventional cultural techniques. Of the 101 samples studied, complete agreement was obtained with conventional methods for 94 but six apparently false positive results and one false negative result occurred.     

A comparison of strontium selenite broth and mannitol-selenite-cystine broth in the isolation of salmonellae from egg products.

Of 103 isolations of Salmonella sp from egg pulp and albumen, 92 were made using strontium selenite enrichment broth and 54 were made using mannitol-selenite-cystine enrichment broth. The results suggest that the former medium may be used successfully for enrichment and subsequent detection of salmonellae in egg products.     

A complete protocol using conductance for rapid detection of salmonellas in confectionery materials

Increased confidence in conductimetric detection of salmonellas was achieved by combining a bacteriophage-based test with use of a selenite cystine trimethylamine oxide dulcitol medium and a modified lysine decarboxylase broth. All 81 Salmonella isolates tested were detected and few of the 39 non-salmonellas gave false positives. Results from the screening of 43 inoculated product samples further support the use of this simple, rapid method for routine salmonella testing in the food industry.     

Enzymes metabolizing dimethylamine, trimethylamine and trimethylamine N-oxide in the yeast Sporopachydermia cereana grown on amines as sole nitrogen source

Abstract Sporopachydermia cereana , an ascosporogenous yeast, grew on dimethylamine, trimethylamine or trimethylamine N -oxide as sole nitrogen sources and produced mono-oxygenases for dimethylamine and trimethylamine that were significantly more stable than the corresponding enzymes found in Candida utilis . No trimethylamine mono-oxygenase activity was found in S. cereana grown on dimethylamine. In cells grown on trimethylamine N -oxide (but not on the other nitrogen sources), evidence for an enzyme metabolizing the N -oxide, possibly an aldolase, but more probably a reductase was obtained. All these activities showed a similar requirement for the presence of FAD or FMN in the extract buffer during isolation to retain activity. Amine mono-oxygenase activities showed a similar sensitivity to inhibitors, including proadifen hydrochloride and carbon monoxide as the corresponding enzymes in C. utilis . The trimethylamine N -oxide-dependent oxidation of NADH was more sensitive to inhibition by EDTA, N -ethylmaleimide and β-phenylethylamine than the mono-oxygenases, and less sensitive to KCN, and activity was significantly higher with NADPH than was observed with the 2 mono-oxygenases.     

Experimental enzyme-linked amperometric immunosensors for the detection of salmonellas in foods

Two enzyme-linked amperometric immunosensors specific for salmonellas were developed as rapid methods for quantifying and detecting these organisms in pure cultures and foods. Both used alkaline phosphatase as the enzyme reporter molecule but one system used phenyl phosphate as the substrate followed by the electrochemical detection of phenol at a polarized platinum electrode. The other system incorporated an enzyme amplification step and relied on the electrochemical detection of a reduced mediator, ferrocyanide. Both assays were rapid (4 h) and specific and generated salmonella-dependent signals above 10⁴ cfu/ml (phenyl phosphate system) or 10⁵ cfu/ml (enzyme amplified system) in pure cultures and samples of several foods, although the results with beef samples showed considerable variation. Both systems were able to detect low (1–5 cfu/g or /ml) numbers of salmonellas in foods after non-selective (18 h) and selective (22 h) enrichment steps but four samples, out of 147, gave false positive results. False positive results were eliminated by reducing the enrichment steps to 6 h and 18 h respectively (90 samples).