Histamine, Cadaverine and Putrescine Forming Bacteria from Ripened Spanish Semipreserved Anchovies

Histidine decarboxylase activity and production of putrescine and cadaverine were assessed in 200 bacterial isolates from ripened semipreserved Spanish anchovies. Highest levels of histidine decarboxylase activity were observed in Morganella morganii, with 2336.74 ± 356.32 ppm of histamine, produced in laboratory media, after 24 hr incubation at 37°C. Microorganisms producing histamine also produced detectable putrescine and cadaverine. The Niven medium was used to evaluate the 200 isolates for histidine decarboxylase activity, after incubation 24 hr at 37°C. An enzymic technique was used to distinguish false positives and to quantify bacterial histamine levels. The number of microorganisms was the most important factor in the accumulation of histamine.     

Histamine Production by Morganella morganii in Mackerel, Albacore, Mahi-mahi, and Salmon at Various Storage Temperatures

Morganella morganii was studied for its growth and histamine formation in mackerel, albacore, mahi‐mahi, and salmon stored at various temperatures from ‐30 °C to 37 °C. The optimal temperature for histamine formation was 25 °C. Mackerel, albacore, and mahi‐mahi were shown as good substrates for histidine decarboxylation by M. morganii at elevated temperatures (> 15 °C). M. morganii inoculated in all fish species including salmon formed histamine above the FDA guideline. Their growth was controlled by cold storage of the fish at 4 °C or below, but histamine formation was controlled only by frozen storage. Although histamine was not detected in any frozen samples, it accumulated rapidly in the previously frozen fish stored at 25 °C.     

Histamine Production in Soy Extended Tuna Salads

Histamine production in tuna salads extended with textured soy protein (TSP) was evaluated. Salads were inoculated with five known histamine-producing bacteria and held at 8°C, 24°C, and 37°C for up to 48 hr. Addition of 30% TSP to tuna salads resulted in higher initial pH and favorable growth conditions for microorganisms and histidine decarboxylase activity. Addition of 15% TSP provided an initial pH for maximal enzyme and histamine production but somewhat slower microbial growth. Tuna salad extended with either 15% or 30% TSP developed toxic levels of histamine (>50 mg/l00 g) when held at either 24° or 37°C for 6 hr. Nonextended tuna salads did not develop toxic levels of histamine even when inoculated with known histamine-producing bacteria and held at 24° or 37°C for 48 hr.     

Bacterial Growth and Histamine Production on Vacuum Packaged Tuna

Vacuum and nonvacuum packaged samples of tuna or tuna spiked with Klebsiella oxytoca T₂, Morganella morganii JM or Hafnia alvei T₈ were stored at 2°C and 10°C and examined for bacterial growth and histamine production on day 3, 6, 10 and 15. Unlike those stored at 2°C, tuna samples stored at 100°C showed significant bacterial growth over time and had high levels of histamine (>200 mg/100g tuna on day 15). Vacuum packaging did not show any beneficial effect in controlling bacterial growth and histamine production on tuna samples at these two temperatures. Low temperature storage was more effective than vacuum packaging in controlling histamine production by the bacteria tested.     

Production of biogenic amines by<Emphasis Type="Italic"> Morganella morganii</Emphasis>,<Emphasis Type="Italic"> Klebsiella pneumoniae</Emphasis> and<Emphasis Type="Italic"> Hafnia alvei</Emphasis> using a rapid HPLC method

The histidine decarboxylating activity and production of biogenic amines by Morganella morganii (NCIMB, 10466), Klebsiella pneumoniae (NCIMB, 673) and Hafnia alvei (NCIMB, 11999) were investigated using a rapid HPLC method. Derivatisation of the bacterial samples was carried out using benzoyl chloride. A gradient elution system was used for analysis with a mixture of acetonitrile and HPLC grade water. Bacterial strains not only produce histamine in histidine-enriched broth but also the other biogenic amines. The chromatographic results show that bacterial strains are also capable of producing spermine and spermidine in histidine-enriched broth. Bacterial ammonia production by all three strains was clearly detected since ammonia is generated during the degradation of histidine. The study demonstrates that the highest histamine production was obtained by Morganella morganii, followed by Klebsiella pneumoniae, and the lowest with the Hafnia alvei. Therefore, Morganella morganii and Klebsiella pneumoniae have strong histidine decarboxylase activity since they are prolific histamine-forming bacteria     

Factors Affecting Amine Production by a Selected Strain of Lactobacillus bulgaricus

Lactobacillus bulgaricus-52 showed maximum growth and amine production in MRS broth in 24 hr at 37°C. The optimum pH for the production of histamine, tyramine, and tryptamine by L. bulgaricus-52 was 5.0. Highest yields of the different amines were obtained in the absence of NaCI in the growth medium. A concentration of even 0.5% NaCI had a slight inhibitory affect on the synthesis of amines. A similar trend was observed in terms of amine production by L bulgaricus-52 in milk samples held at 37°C for different time intervals.     

Growth and histamine production by Morganella morganii under various temperature conditions

The generation times (G) for Morganella morganii were determined using absorbance measurements (at 0–10°C) or conductance measurements (at 15–25°C). Growth was rapid at 15–25°C (G = 2.6 at 15°C and G = 1.08 h at 25°C) but at the lower temperatures growth was greatly reduced. Growth and histamine production by M. morganii in histidine-containing broth (HDB) and in mackerel (packed in air-permeable plastic bags) was studied. It was found that large amounts of histamine are formed at low temperatures (0–5°C), where no growth takes place, following storage at higher temperatures (10–25°C). Measurements of conductance can be used to separate between samples with no or only insignificant content of histamine (< 50 ppm) which had detection times (DT) above 10 h and those with elevated content of histamine (> 50 ppm) which had shorter DT.     

Escherichia coli O157: H7 Growth in Laboratory Media as Affected by Phenolic Antioxidants

Five strains of Escherichia coli O157:H7 with ATCC 11775 E. coli were grown in brain heart infusion (BHI) broth (pH 5.8, adjusted with citric acid) and treated with butylated hydroxyanisole (BHA), butylated hy-droxytoluene (BHT), tertiary butylhydroquinone (TBHQ), and propyl gallate (PG) individually or combined. Additives ranged from 100–400 ppm with inocula levels between 5 and 104 CFU/mL in tissue culture plates or in flasks; samples were incubated at 4°C or 37°C for 24 hr. Additive antimicrobial efficacy varied with inoculum level and incubation temperature. BHA at <200 ppm was bactericidal on all strains. Poly-hydroxyl additives (TBHQ, PG) were less effective at 4°C. BHA-BHT combinations were synergistic at 4°C.     

Increased Aflatoxin Production by Aspergillus parasiticus Under Conditions of Cycling Temperatures

The effects of cycling temperatures (5°C for 12 hr and 25°C for 12 hr) on aflatoxin production by Aspergillus parasiticus NRRL 2999 in yeast extract sucrose (YES) medium were studied. Cycling temperatures, after preincubation at 25°C for various times, resulted in more aflatoxin B₁, G₁, and total aflatoxin production than did constant incubation at either 25°C, which is generally considered to be the optimum for aflatoxin production, or 15°C, which is the same total thermal input as the 5-25°C temperature cycling. With increased preincubation time at 25°C, toxin production increased and the lag phase of growth was shortened or not evident. Cultures that were preincubated at 25°C for 1, 2, and 3 days prior to onset of temperature cycling showed the greatest increase in maximum aflatoxin production over the 25°C and 15°C constant temperatures. Cultures that were not preincubated at 25°C but subjected to constantly fluctuating temperatures produced maximum amounts of aflatoxin equivalent to cultures incubated at a constant 25°C. The maximum aflatoxin production at all temperatures studied occurred during the late log phase of growth and at pH minimums. Aflatoxins were found in higher concentrations in the broth than the mycelia under temperature cycling conditions, at 15°C, and at 25°C during the first 21 days of incubation, whereas greater amounts of toxin were retained in mycelium at 25°C in the later incubation period (28-42 days).     

SSEL,a selective enrichment broth for simultaneous growth of Salmonella enterica,Staphylococcus aureus,Escherichia coli O157: H7, and Listeria monocytogenes

Salmonella enterica, Staphylococcus aureus, Escherichia coli O157: H7, and Listeria monocytogenes may contaminate similar types of food and cause foodborne disease. The objective of this study was to develop a selective enrichment broth for simultaneous enrichment of Salmonella enterica, Staphylococcus aureus, Escherichia coli O157: H7, and Listeria monocytogenes (SSEL) using nalidixic acid, acriflavine, lithium chloride, and sodium cholate as selective agents. Developed SSEL broth not only enriched the target pathogens to 5 log10 CFU/ml after 18 hr incubation at 37°C with 10–100 CFU/mL of inoculation concentration, but also could successfully support the simultaneous enrichment of target pathogens with similar growth rates and inhibit the growth of most nontarget bacteria effectively. The enrichment effect of SSEL was confirmed by artificial contamination test coupled with multiplex PCR. In summary, SSEL has been shown to be a promising multiplex selective enrichment broth for the detection of the four pathogens on a single-assay platform.     

Temperature and NaCl Affect Growth and Survival of Escherichia coli 0157:H7 in Poultry-Based and Laboratory Media

In tryptic soy broth (TSB) and a poultry extract broth (PB) with 0 to 10% (w/v) NaCl incubated at 37°C, growth of E. coli 0157:H7 was inhibited at 28% NaCl whereas at 10°C, growth was inhibited at 24% NaCl in TSB and at 26% NaCl in PB. The bacterium did not grow at 4°C. Increased NaCl-sensitivity observed at 10°C was a bacteriostatic effect that was ineffective with increasing incubation temperature. At 10°C, E. coli 0157:H7 was more salt-tolerant in PB than in TSB, although PB growth rates were lower. Findings suggest that PB may be a more suitable medium for testing E. coli 0157:H7 in poultry products. Cells of E. coli 0157:H7 that were exposed to refrigeration (4°C) and/or NaCl for 24 days did not grow on MacConkey agar with 1% sorbitol.     

Biogenic amine production by<Emphasis Type="Italic"> Morganella morganii</Emphasis> and<Emphasis Type="Italic"> Klebsiella oxytoca</Emphasis> in tuna

Biogenic amine formation in tuna inoculated with two micro-organisms reported as powerful in vitro histamine producers, Morganella morganii and Klebsiella oxytoca, was investigated. Inoculated and non-inoculated tuna samples were stored in ice, under refrigeration at 8–10 °C and at 20±1 °C. Ten biogenic amines were studied and changes were found only in cadaverine and histamine in inoculated samples. Therefore, the in vitro ability of K. oxytoca to form cadaverine and that of both microorganisms to form histamine was confirmed in inoculated samples. Storage temperature had a crucial effect on the amount and intensity of amine production. On the other hand, the potential ability of M. morganii to form putrescine was not shown in the inoculated samples, regardless of the storage temperature.     

Evaluation of 54° and 60°C Holding Temperatures on Aerobic Plate Counts and Coliform Determinations of Control and Post Cooking Contaminated Beef Roast

Fabricated beef roasts were cooked to an internal temperature of 60°C and the surface inoculated with a suspension of Escherichia coli at a level of lo⁶ cells/cm². Roasts were then stored in a warming oven at 54° and 60°C for 1, 2, 6, and 12 hr. Under these conditions the destruction of this strain of E. coli is quite rapid (less than 6 hr) at the 60°C holding temperature. The holding temperatures do allow the growth of thermoduric organisms naturally present in the product to initiate exponential growth after 6 hr at 54°C. The higher holding temperature reduced the rate of multiplication of the thermoduric organisms.     

Occurrence of Histamine-Forming Bacteria in Albacore and Histamine Accumulation in Muscle at Ambient Temperature

Distribution of histamine‐producing bacteria in fresh albacore and their proliferation in muscle during storage at 25 °C were monitored. Histamine formers constituted a minor portion of the bacterial flora in albacore. Only weak histamine formers were detected in the gill and skin of fresh fish after enrichment. Histamine formers were isolated from the muscles when APC reached >10⁷ CFU/g during storage. Hafnia alvei was most prevalent in both fresh and temperature‐abused fish. The most prolific histamine former, Morganella morganii, was rarely isolated even from spoiled muscles. The prevalence of weak histamine formers resulted in low levels of histamine accumulation in muscle, 67.1 mg/100 g, for 6 d of storage.     

Application of chitosan–alginate microspheres for the sustained release of bacteriophage in simulated gastrointestinal conditions

This study was designed to evaluate the acid stability, release property and antimicrobial efficacy of Escherichia coli O157:H7 bacteriophages encapsulated in chitosan–alginate microspheres under the simulated gastrointestinal conditions. The bacteriophages belonging to Myoviridae family were stable at the pH above 4 in trypticase soy broth. The chitosan–alginate microspheres exhibited protective effect on the viability of bacteriophages in the simulated gastric conditions at pH 2.0 and pH 2.5, showing 4.8 and 5.6 log PFU mL^‐1, respectively, after 1 h of incubation at 37 °C. The release per cent of bacteriophages from microspheres gradually increased up to 65% in the simulated intestinal condition (pH 7.5) at 37 °C for 6 h. The lytic efficacy of chitosan‐ and alginate‐encapsulated bacteriophages against E. coli O157:H7 was significantly maintained in the simulated intestinal conditions to 10 h of incubation (1.3 log reduction). The results suggest that the chitosan–alginate microspheres can be used as a reliable delivery system for bacteriophages.     

Heat resistance in Escherichia coli and its implications on ground beef cooking recommendations in Canada

This study assessed the adequacy of the current cooking recommendations in relation to heat resistant Escherichia coli by evaluating eight potentially heat resistant E. coli strains (four generic and four E. coli O157:H7) along with AW1.7. The D_60°C-values for these strains varied from 1.3 to 9.0 min, with J3 and AW1.7 being the least and most heat resistant strains, respectively. The D_60°C-values for E. coli 62 and 68 were similar and were not affected by growth medium, while the heat resistance of C37, J3, and AW1.7 varied with the growth medium. When heated in extra lean ground beef (100 g) in vacuum pouches, the mean D_54°C, D_57°C, and D_60°C-values were 44.8, 18.6, and 2.9 min for C37, 13.8, 6.9, and 0.9 min for J3, and 40.5, 9.1, and 6.1 min for AW1.7. Burger temperatures continued to rise after being removed from heat when the target temperature was reached, by 3–5°C, and resting of 1 min would result in a destruction of 133, 374 and 14 log C37, J3 and AW1.7. These findings along with the very low occurrence of heat resistant E. coli expected in ground beef show that cooking ground beef to 71°C should be adequate.     

Effects of Frozen Storage on Survival of Staphylococcus aureus and Enterotoxin Production in Precooked Tuna Meat

This study investigated the survival of Staphylococcus aureus in precooked tuna meat for producing canned products during frozen storage (?20 ± 2 °C) as well as its growth and enterotoxin production at 35 to 37 °C after the storage. Samples (50 ± 5 g) of precooked albacore (loin, chunk, and flake) and skipjack (chunk and flake) tuna were inoculated with 5 enterotoxin‐producing strains of S. aureus at a level of approximately 3.5 log CFU/g and individually packed in a vacuum bag after 3 h incubation at 35 to 37 °C. Vacuum‐packed samples were stored in a freezer (?20 ± 2 °C) for 4 wk. The frozen samples were then thawed in 37 °C circulating water for 2 h and incubated at 35 to 37 °C for 22 h. Populations of S. aureus in all precooked tuna samples decreased slightly (<0.7 log CFU/g) after 4 wk of storage at ?20 ± 2 °C, but increased rapidly once the samples were thawed and held at 35 to 37 °C. Total S. aureus counts in albacore and skipjack samples increased by greater than 3 log CFU/g after 6 and 8 h of exposure to 35 to 37 °C, respectively. All samples became spoiled after 10 h of exposure to 35 to 37 °C, while no enterotoxin was detected in any samples. However, enterotoxins were detected in albacore loin and other samples after 12 and 24 h of incubation at 35 to 37 °C, respectively. Frozen precooked tuna meat should be used for producing canned tuna within 6 to 8 h of thawing to avoid product spoilage and potential enterotoxin production by S. aureus in contaminated precooked tuna meat.     

Microbiological quality of retail fresh fish fillets in The Netherlands

A total of 242 samples of ready-for-sale fish fillets of validated good sensory quality was examined for colony counts at 20, 30 and 37°C, Enterobacteriaceae at 37°C, Escherichia coli, Salmonella and Vibrio parahaemolyticus in 10 g aliquots. Staphylococcus aureus and yeast and mould propagules. Gram negative pathogens were not detected in any sample. The following reference values were found attainable: colony counts at 30°C, 10⁶ g^?1; E. coli 10 g^?1; S. aereus 10² g^?1; yeast and mould propagules 10⁴ g^?1. These reference values include, as customary, a tolerance of about 20% of samples exceeding the stated levels without, however, reaching the next log₁₀ level.     

The ability of biogenic amines and ammonia production by single bacterial cultures

The amino acid decarboxylating activity and production of biogenic amines, trimethylamine and ammonia by Morganella morganii (two strains), Klebsiella pneumoniae (three strains), Hafnia alvei (two strains), Enterococcus faecalis, Photobacterium phosphoreum, Micrococcus sp., Psychrobacter immobilis, Corynebacterium sp., Vibrio fischeri, Vibrio harveyi and Pseudomonas putida were investigated using a rapid HPLC method. In a laboratory medium containing amino acid (histidine, ornithine, lysine, tyrosine and arginine), not all bacterial strains produced the biogenic amines but most of them produced histamine, putrescine, cadaverine and ammonia. Cadaverine production by Klebsiella pneumoniae (8152), Klebsiella pneumoniae (673), Klebsiella pneumoniae (2122), Hafnia alvei (6578), Hafnia alvei (11999), Vibrio fischeri (25) Vibrio harveyi (42) and Pseudomonas putida (10936) was 531, 422, 532, 485, 472, 343, 547 and 343 mg/l, respectively in lysine decarboxylase broth. Tyramine was produced in highest concentration (526 mg/l) by Enterococcus faecalis (775). Agmatine was not produced apart from Psychrobacter immobilis (100) in an arginine decarboxylase broth.     

Improved Tenderness of Restructured Beef Steaks by a Microbial Collagenase Derived from Vibrio B-30

To reduce connective tissue toughness Vibrio B-30 collagenase was evaluated for its ability to degrade muscle collagen in a restructured beef (RSB) product. Dissected muscles pooled from clods of 4 USDA choice chucks were made into three RSB products: untrimmed control (C), hand trimmed of epimysium (T) and untrimmed and treated with 0.01% collagenase (E). RSB products were held for 24 hr at either 2°C or 11°C and were analyzed cither raw or after heating (40°C for 1 hr). Aerobic microbial load remained low (1–2X10² cfu/g) during the 24 hr 11°C incubation. Heated samples of enzyme treated RSB had lower Lee-Kramer shear values than controls. In both raw and heated samples enzyme treatment increased hydroxyproline solubility. An enzyme treatment of 24 hr at 11°C followed by heating produced maximum tenderization. Although optimum treatment conditions need to be defined by further research, Vibrio B-30 collagenase appeared to be efficacious for the tenderization of RSB products.