Histamine production by Klebsiella pneumoniae and an incident of scombroid fish poisoning.

A histamine-producing strain of Klebsiella pneumoniae was isolated from a sample of tuna sashimi implicated in an outbreak of scombroid fish poisoning. None of the other nine gram-negative bacterial strains isolated from the tuna sashimi was capable of equivalent histamine production. Bacterial histamine production was monitored in a tuna fish infusion broth (TFIB), and the implicated K. pneumoniae was capable of producing 442 mg of histamine per 100 g of tuna in TFIB in 7 h under controlled incubation conditions. Only 12 of 50 other K. pneumoniae strains, representing 5 distinct biochemical types, which had been originally isolated from foods, were able to produce such levels of histamine in TFIB. No correlation was found between histamine production and other biochemical characteristics or antibiotic resistance. Of the 12 histamine-producing strains, 11 belonged to type 2, which is characterized as indole negative with positive reactions in the urea and Voges-Proskauer tests. However, only 50% of the type 2 strains examined produced high levels of histamine in TFIB. Additionally, the implicated K. pneumoniae strain and one other strain belonged to type 1, which is characterized by positive reactions in the indole, urea, and Voges-Proskauer tests.

Comparison of thin-layer chromatographic detection methods for histamine from food extracts.

The specificity and sensitivity of four visualization reagents, ninhydrin, o-phthalaldehyde (OPT), fluorescamine and o-diacetylbenzene (DAB), for the detection of histamine on thin-layer chromatograms developed in several optimal solvent systems were compared. While the ninhydrin reagent is the most sensitive with a detection limit of 0.4 nmoles of histamine, the DAB and fluorescamine reagents offer the most potential as histamine-specific visualization reagents. The OPT reagent was unsatisfactory because the fluorescent sports developed with this reagent were unstable. Of 19 amines tested in addition to histamine, ninhydrine reacted with 17, fluorescamine with 8 and DAB with 15. Since only histidyl-leucine, octopamine, serotonin and tryptophan interfere in the resolution of histamine in any of the solvent systems used, DAB provides some advantages by not reacting with histidyl-leucine. In addition, with the acetone-ammonia (95:5) solvent system, which adequately separates histamine from all other tested amines, no background interference problems were noted when using DAB as the visualization reagent. Background interference problems with the acetone-ammonia system were evident with the other visualization reagents. The sensitivity of the DAB and fluorescamine methods (4 nmoles of histamine) allows both reagents to act as semi-quantitative screening agents for potentially toxic levels of histamine in food extracts. With a variety of food samples, DAB visualization following development in acetone-ammonia (95:5) was the optimal thin-layer chromatographic method.

Thin-layer chromatographic screening methods for histamine in tuna fish.

Twelve solvent systems were tested for their ability to separate histamine and histidine on a variety of thin-layer coatings. The best solvent-adsorbent systems were: chloroform-methanol-ammonia (2:2:1), methanol-ammonia (20:1), acetone-ammonia (95:5), and double development with (a) n-butanol-acetone-water (2:2:1) and (b) chloroform-methanol-ammonia (12:7:1), all on silica-gel layers. Ninhydrin was used as the visualization reagent. These four systems were then evaluated for their potential use as rapid screening procedures in the detection of possibly deleterious levels of histamine in tuna fish. Successful separation of histamine from the other ninhydrin-positive components of methanolic tuna fish extracts was achieved with all four systems. A sample from a lot of tuna implicated in human illness was found to have a histamine level considerably higher than tuna purchased from a local retail outlet or an extract spiked to a histamine level considered to be a threshold value for toxicity symptoms. The methanol-ammonia (20:1) and chloroform-methanol-ammonia (2:2:1) systems, used with silica-gel plates, are the most promising for rapid preliminary screening of tuna fish extracts for histamine.

Time Course of Steroid Biosynthesis and Metabolism in Haplopappus heterophyllus.

A single dose of mevalonic acid-2-(14)C was administered simultaneously to 6 Haplopappus heterophyllus plants. They were harvested at intervals ranging from 3 days to 6 months. Four groups of biosynthetically related sterols were found to be radioactive in each plant, and the changes in radioactivity with time were studied. The most striking finding was a radioactive phenolic material present only in the 6-month plant, which had flowered.