Bacterial Histamine Production as a Function of Temperature and Time of Incubation

Optimal temperature, lower temperature limit, extent, and rate of histamine production in a tuna fish infusion broth (TFIB) varied for the strains of Proteus morganii, Klebsiella pneumoniae, Hafnia alvei, Citrobacter freundii, and Escherichia coli studied. P. morganii and K. pneumoniae produced large quantities of histamine in a relatively short incubation period (<24 hr) at 15°C, 30°C, and 37°C; production was fastest at 37°C. H. alvei, C. freundii, and E. coli produced toxicologically significant levels of histamine (>2500 nmoles/ml) only at 30°C and 37°C on prolonged incubation (≥48 hr). At 72 hr of incubation, optimal temperature for histamine production was 37°C for E. coli and C freundii; 30°C for P. morganii strain 110SC2, K. pneumoniae, and H. alvei; and 15°C for P. morganii strain JM. The lower temperature limits for production of toxicologically significant levels of histamine in TFIB were 7°C for K. pneumoniae; 15°C for both P. morganii strains; and 30°C for H. alvei, C. freundii, and E. coli.     

Biogenic amines produced by Enterobacteriaceae isolated from meat products

Biogenic amines in ground meat and processed meat products are one of the indicators to determine the poor quality raw material. Major histamine forming bacterium was Escherichia coli (strain EC04 with 65.88 mg/100 ml in Brain Heart Infusion medium), followed by the microorganisms Morganella morganii (strain MM4 with 8.40 mg/100 ml and strain MM7 with 8.28 mg/100 ml) and Proteus mirabilis (strain PM02 with 8.76 mg/100 ml), respectively. The highest putrescine production level was found in Citrobacter freundii and Enterobacter spp. strains, followed by Serratia grimesii, Proteus alcalifaciens, E. coli, Escherichia fergusonii, Morganella morganii, Proteus mirabilis, Proteus penneri and Hafnia alvei, respectively. The most important cadaverine producer was E. coli EC03 with a production level of 45.48 mg/100 ml, and of the strains Escherichia vulnaris EV01, Escherichia fergusonii EF06 produced 37.92 mg/100ml and 35.40 mg/100 ml, respectively. On the basis of our results, it can be concluded that the major biogenic amines produced by Enterobacteriaceae are putrescine, cadaverine, tyramine and histamine, both in culture medium and meat products.     

Antibacterial effect of food additives and detergents against histamine-producing bacteria on food contact material surfaces

We investigated the antibacterial activity of food additives and detergents against histamine-producing bacteria on food contact material surfaces. Based on minimum inhibitory concentration (MIC) testing with Morganella morganii NBRC3848, Raoultella planticola NBRC3317 and Enterobacter aerogenes NCTC10006, we screened nine food additives and four detergents with relatively high inhibitory potency. We prepared food contact material surfaces contaminated with histamine-producing bacteria, and dipped them into fourteen agents (100 μg/mL). Sodium hypochlorite, benzalkonium chloride, benzethonium chloride, n-hexadecyltrimethylammonium chloride and 1-n-hexadecylpyridinium chloride showed antibacterial activity against histamine-producing bacteria. We prepared low concentrations of the five agents (10 and 50 μg/mL) and tested them in the same way. Sodium hypochlorite showed high antibacterial activity at 10 μg/mL, and the other four showed activity at 50 μg/mL. So, washing the material surface with these reagents might be effective to prevent histamine food poisoning owing to bacterial contamination of food contact surfaces.     

Photobacterium phosphoreum caused a histamine fish poisoning incident

An incident of histamine fish poisoning (HFP) occurred due to the consumption of iwashi maruboshi (dried sardine) in Osaka, Japan in March 2002. A histamine-producing bacterial strain, YS4-7, was isolated from iwashi maruboshi that contained 1700 mg of histamine per kilogram. This strain was identified as Photobacterium phosphoreum by biochemical examinations and partial sequencing of 16S rDNA. P. phosphoreum YS4-7 showed greater capability as a histamine producer at 4 and 12 degrees C than Morganella morganii JCM 1672. Strain YS4-7 produced 546 mg of histamine per kilogram in a sardine homogenate stored for 12 h at 20 degrees C. M. morganii, Raoultella planticola and Hafnia alvei have been isolated from fish implicated in HFP incidents, whereas this is the first report of P. phosphoreum being the causative bacterium in a sporadic case of histamine food poisoning.     

Formation of histamine and biogenic amines in cold-smoked tuna: an investigation of psychrotolerant bacteria from samples implicated in cases of histamine fish poisoning

Two outbreaks and a single case of histamine fish poisoning associated with cold-smoked tuna (CST) were reported in Denmark during 2004. The bacteria most likely responsible for histamine formation in CST implicated in histamine fish poisoning was identified for the first time in this study. Product characteristics and profiles of biogenic amines in the implicated products were also recorded. In the single poisoning case, psychrotolerant Morganella morganii-like bacteria most likely was responsible for the histamine production in CST with 2.2% +/- 0.6% NaCl in the water phase (WPS). In outbreak 1, Photobacterium phosphoreum most likely formed the histamine in CST with 1.3% +/- 0.1% WPS. In outbreak 2, which involved 10 persons, the bacteria responsible for histamine formation could not be determined. The measured concentrations of WPS were very low compared with those of randomly collected commercial samples of CST and cold-smoked blue marlin (4.1 to 12.7% WPS). Challenge tests at 5 degrees C with psychrotolerant M. morganii and P. phosphoreum in CST with 4.4% WPS revealed growth and toxic histamine formation by the psychrotolerant M. morganii-like bacteria but not by P. phosphoreum. In a storage trial with naturally contaminated CST containing 6.9% WPS, lactic acid bacteria dominated the microbiota, and no significant histamine formation was observed during the shelf life of about 40 days at 5 degrees C and of about 16 days at 10 degrees C. To prevent toxic histamine formation, CST should be produced with >5% WPS and distributed with a declared 5 degrees C shelf life of 3 to 4 weeks or less.     

Histamine content and histamine-forming bacteria in dried milkfish (Chanos chanos) products

Thirty-two dried milkfish products sold in five retail markets in southern Taiwan were purchased and tested to determine the occurrence of histamine and histamine-forming bacteria. Except for histamine and cadaverine, the average content of various biogenic amines in tested samples was less than 8.5 mg/100 g. Most of the tested dried milkfish products (78.1%) had histamine levels greater than the FDA guideline of 5 mg/100 g for scombroid fish and/or product, while fourteen of them (43.7%) contained > 50 mg/100 g hazard action level. Thirty histamine-producing bacterial strains, capable of producing 5.4 ppm to 562 ppm of histamine in trypticase soy broth supplemented with 1.0% L-histidine (TSBH), were identified as Enterobacter aerogenes (seven strains), Citrobacter sp. (one strain), Staphylococcusxylosus (ten strains), S. sciuri (one strain), Bacillus thuringiensis (two strains), Citrobacter freundii (five strains), Klebsiella pneumoniae (one strain) and E. cloacae (three strains), by 16S rDNA sequencing with PCR amplification.     

PCR detection and identification of histamine-forming bacteria in filleted tuna fish samples

Total of 14 filleted yellowfin tuna fish (Thunnus albacares) sold in wholesale fish market and supermarkets in Milan, Italy, were purchased and tested to determine microbial count, histamine level, histamine-forming bacteria, and their ability to produce histamine in culture broth. Although histamine level was less than 10 ppm, many samples showed high total viable bacterial and enterobacterial counts that reached dangerous levels after temperature abuse for short periods of time. A PCR assay targeting a 709-bp fragment of the histidine decarboxylase gene (hdc) revealed that 30.5% of the 141 enteric bacteria isolated from samples were positive and potentially able to produce histamine. The hdc positive strains were mainly isolated from fish bought at wholesale fish market, where we observed several possible risk factors, such as handling in poor and non-refrigerated conditions during fillet preparation. These positive strains were identified as Citrobacter koseri/Enterobacter spp. and Morganella morganii, by 16S/23S rRNA internal transcribed spacer amplification and 16S rRNA sequence analysis. The strains showed a variable ability of histamine production, with Morganella morganii being the most active histamine-producing species. A direct DNA extraction from fish and a PCR targeting the hdc gene showed a high degree of concordance with the results obtained through microbiological and chemical analyses, and could aid in the prompt detection of potentially contaminated fish products, before histamine accumulates. PRACTICAL APPLICATION: The use of methods for the early and rapid detection of bacteria producing biogenic amines is important for preventing accumulation of these toxic substances in food products. In this study, we used a molecular approach for the detection of histamine-forming bacteria in fish. PCR-based methods require expensive equipment and a high degree of training for the user, but are fast (< 24 h) and reliable. They now represent the best predictive methods to identify a potential risk factor in fish products during processing, storage, and marketing and can be used in the investigation of risk reduction strategies.     

Histamine formation by histamine-forming bacteria and yeast in mustard pickle products in Taiwan

The occurrence of histamine, histamine-forming bacteria and yeast were tested in 37 mustard pickle products sold in both retail markets and supermarkets in southern Taiwan. Aerobic plate count (APC), total coliform, and Escherichia coli were also tested for microbiological quality. Salt content, pH value, titratable acidity and sulphite content were determined for quality of mustard pickle products. Only one retail market sample and one supermarket sample had 8.9 and 7.4 mg histamine per 100 g products, although the average content for each of the nine biogenic amines was less than 2 mg/100 g. Ten histamine-forming bacterial strains and 6 histamine-producing yeast strains capable of producing 8.7 to 1260 ppm of histamine in trypticase soy broth (TSB) supplemented with 1% l-histidine (TSBH) were identified as Staphylococcus capitis (four strains),Staphylococcus pasteuri (two strains), Enterobacter cloacae (four strains), Candida glabrata (two strains) and Candida rugosa (four strains). S. capitis, which was previously reported to be halotolerant, was a potent histamine-former, capable of producing more than 1000 ppm of histamine in TSBH in the presence of 0.5–10% NaCl. The numbers of the aerobic plate count (APC) in all samples were below the Taiwanese regulatory level of 5 log CFU/g. None of the samples contained total coliform or E. coli. The values of pH, salt content, titratable acidity and sulphite content in all samples ranged from 3.8% to 5.0%, 2.0% to 10.0%, 0.21% to 1.18% and <2.0–1876 ppm, respectively.     

Histamine contents of salted seafood products in Taiwan and isolation of halotolerant histamine-forming bacteria

Fifty-seven salted seafood products sold in the fishing village stores in Taiwan, including salted fish product, salted mollusc product and salted shrimp product, were tested to determine the occurrence of histamine and histamine-forming bacteria. Although the average content of each of nine different biogenic amines in all samples was less than 5.0 mg/100 g, 10.5% (6/57) of tested samples had the histamine content greater than the 5.0 mg/100 g allowable limit suggested by the US Food and Drug Administration. One histamine-producing bacterial strain, capable of producing 78.5 ppm of histamine in trypticase soy broth supplemented with 1.0% l-histidine (TSBH), was identified as Bacillus megaterium. The B. megaterium isolate was a halotolerant bacterium which grew well to an elevated NaCl concentration of 15% in TSBH medium. Besides, it had a consistent ability to produce >300 ppm of histamine at 10% NaCl concentration in TSBH medium after 72 h.     

Acid phosphatase/phosphotransferases from enteric bacteria

We have investigated the enzymatic phosphorylation of nucleosides and found that Morganella morganii phoC acid phosphatase exhibits regioselective pyrophosphate (PP(i))-nucleoside phosphotransferase activity. In this study, we isolated genes encoding an acid phosphatase with regioselective phosphotransferase activity (AP/PTase) from Providencia stuartii, Enterobacter aerogenes, Escherichia blattae and Klebsiella planticola, and compared the primary structures and enzymatic characteristics of these enzymes with those of AP/PTase (PhoC acid phosphatase) from M. morganii. The enzymes were highly homologous in primary structure with M. morganii AP/PTase, and are classified as class A1 acid phosphatases. The synthesis of inosine-5'-monophosphate (5'-IMP) by E. coli overproducing each acid phosphatase was investigated. The P. stuartii enzyme, which is most closely related to the M. morganii enzyme, exhibited high 5'-IMP productivity, similar to the M. morganii enzyme. The 5'-IMP productivities of the E. aerogenes, E. blattae and K. planticola enzymes were inferior to those of the former two enzymes. This result underlines the importance of lower K(m) values for efficient nucleotide production. As these enzymes exhibited a very high degree of homology at the amino acid sequence level, it is likely that local sequence differences in the binding pocket are responsible for the differences in the nucleoside-PP(i) phosphotransferase reaction.     

Histamine formation by enterococci isolated from home-made goat cheeses

A survey was made of the histamine-producing capability of enterococci isolated from goat cheeses. The strains, 130 Streptococcus faecium and 106 S. faecalis, were grown in Trypticase Soy Broth Histidine medium (TSBH) at 35 degrees C for 24 h and the histamine formed was determined by fluorometry. Forty-one (31.5%) of the S. faecium strains and 2 (1.9%) of the S. faecalis strains produced histamine. The largest amount detected was 4.0 micrograms histamine/ml TSBH. Compared with the amounts of histamine produced by some Gram-negative bacteria, the histamine production by enterococci seems to be low. Under the present conditions the enterococci seem to have no relevance from a histamine intoxication point of view.     

Detection of Morganella morganii,a prolific histamine former,by the polymerase chain reaction assay with 16S rDNA-targeted primers

A polymerase chain reaction (PCR) assay for the rapid and sensitive detection of the most prolific histamine former, Morganella morganii, was developed. 16S rDNA targeted PCR primers were designed, and the primer specificity and sensitivity of the PCR assay were evaluated. The 16S rDNA sequence (1,503 bp) for M. morganii showed 95% identity to those for enteric bacteria, i.e., Enterobacter spp., Klebsiella spp., Citrobacter spp., Hafnia alvei, Proteus spp., and Providencia spp. The unique primers for M. morganii were designed on the basis of the variable regions in the 16S rDNA sequence. The primers showed positive reactions with all M. morganii strains tested. However, PCR amplification was not detected when the primers were tested with other enteric or marine bacteria. When the sensitivity of the assay was evaluated, M. morganii was detected at levels ranging from 10(6) to 10(8) CFU/ml in albacore homogenate after the PCR amplification. The sensitivity of the assay was greatly improved with the enrichment of samples, and 9 CFU of M. morganii per ml of albacore homogenate was detected after 6 h of enrichment at 37 degrees C.     

Control of Histamine‐Producing Bacteria and Histamine Formation in Fish Muscle by Trisodium Phosphate

Scombrotoxin fish poisoning remains the primary cause of seafood poisoning outbreaks despite preventive guidelines. The purpose of this study was to investigate the use of pH for the control of growth and histamine formation by histamine‐producing bacteria in fish muscle. We examined pH effects on growth and histamine formation in tuna fish infusion broth and in inoculated tuna and mahi‐mahi fish muscle. Histamine production was significantly less for all bacterial strains at pH 8.5 compared to pH 5.5 in tuna fish infusion broth with no significant difference in growth. Elevated pH due to phosphate treatment of fish muscle tissues significantly reduced histamine formation with no effect on the growth of histamine‐producing bacteria. This study revealed that phosphate treatment of mahi‐mahi and tuna fish muscle resulted in significantly lower histamine production over 4 d of storage at 10 °C. Phosphate treatment of fish muscle may serve as a secondary barrier in addition to FDA recommended time and temperature controls for reducing public health concerns of scombrotoxin fish poisoning.     

Significant histamine formation in tuna (Thunnus albacares) at 2 degrees C--effect of vacuum- and modified atmosphere-packaging on psychrotolerant bacteria

Occurrence and importance of psychrotolerant histamine producing bacteria in chilled fresh tuna were demonstrated in the present study. The objective was to evaluate microbial formation of histamine and biogenic amines in chilled fresh tuna from the Indian Ocean and stored either vacuum-packed (VP) or modified atmosphere-packed (MAP). Firstly, biogenic amines and the dominating microbiota were determined in VP tuna involved in an outbreak of histamine fish poisoning in Denmark. Secondly, the microbiota of fresh MAP tuna was evaluated at the time of processing in Sri Lanka and chemical, microbial and sensory changes were evaluated during storage at 1-3 degrees C. To explain the results obtained with naturally contaminated tuna the effect of VP and MAP on biogenic amine formation by psychrotolerant bacteria was evaluated in challenge tests at 2 degrees C and 10 degrees C. The VP tuna that caused histamine fish poisoning had a histamine concentration of >7000 mg/kg and this high concentration was most likely produced by psychrotolerant Morganella morganii-like bacteria or by Photobacterium phosphoreum. Similar psychrotolerant M. morganii-like bacteria dominated the spoilage microbiota of fresh MAP tuna with 60% CO2/40% N2 and formed >5000 mg/kg of histamine after 24 days at 1.7 degrees C. These psychrotolerant bacteria were biochemically similar to M. morganii subsp. morganii and their 16S rDNA (1495 bp) showed >98% sequence similarity to the type strain of this species. Toxic concentrations of histamine were produced at 2.1 degrees C in inoculated VP tuna by both the psychrotolerant M. morganii-like bacteria (7400+/-1050 mg/kg) and P. phosphoreum (4250+/-2050 mg/kg). Interestingly, MAP with 40% CO2/60% O2, in challenge tests, had a strong inhibitory effect on growth and histamine formation by both the psychrotolerant M. morganii-like bacteria and P. phosphoreum. In agreement with this, no formation of histamine was found in naturally contaminated fresh MAP tuna with 40% CO2/60% O2 during 28 days of storage at 1.0 degrees C. To reduce current problems with histamine fish poisoning due to VP tuna it is suggested, for lean tuna loins, to replace vacuum packaging with MAP containing approximately 40% CO2 and approximately 60% O2.     

Histamine production by Enterobacter aerogenes in sailfish and milkfish at various storage temperatures

Enterobacter aerogenes was studied for its growth and ability to promote the formation of total volatile base nitrogen (TVBN) and histamine in sailfish (Istiophorus platypterus) and milkfish (Chanos chanos) stored at various temperatures from -20 to 37 degrees C. The optimal temperature for bacterial growth in both fish species was 25 degrees C, whereas the optimal temperature for histamine formation was 37 degrees C. The two fish species inoculated with E. aerogenes, when not properly stored at low temperatures such as 15 degrees C for 36 h, formed histamine at above the U.S. Food and Drug Administration hazardous guideline level of 50 mg/100 g. Milkfish was a better substrate than sailfish for histamine formation by bacterial histidine decarboxylation at elevated temperatures (> 15 degrees C). Although higher contents of TVBN were detected in the spiked sailfish than milkfish during the same storage time at temperatures above 15 degrees C, the use of the 30-mg/100 g level of TVBN as a determination index for fish quality and decomposition was not a good criterion for assessing potential histamine hazard for both fish species. Bacterial growth was controlled by cold storage of the fish at 4 degrees C or below, but histamine formation was stopped only by frozen storage. Once the frozen fish samples were thawed and stored at 25 degrees C, histamine started to accumulate rapidly and reached levels greater than the hazardous action level in 36 h.     

Determination of histamine and histamine-forming bacteria in tuna dumpling implicated in a food-borne poisoning

An incident of food-borne poisoning causing illness in seven victims, due to ingestion of tuna dumpling, occurred in March 2006, in Chiayi Prefecture, southern Taiwan. The leftovers of the victims’ tuna dumpling and the five other tuna dumpling samples from five other retail stores were collected and tested to determine the occurrence of histamine and histamine-forming bacteria. The levels of pH, salt content, aerobic plate count (APC), total volatile basic nitrogen (TVBN), total coliform (TC) and Escherichia coli in all samples ranged from 6.08 to 6.43, 0.46% to 0.81%, 5.90 to 8.95 log CFU/g, 6.38 to 21.29 mg/100 g, 750 to 8000 most probable number (MPN)/g, and <3 to 1000 MPN/g, respectively. The suspected tuna dumpling contained 160.8 mg/100 g of histamine greater than the hazard action level of 50 mg/100 g set by the US Food and Drug Administration (FDA) for tuna fish. Given the allergy-like symptoms of the victims and the high histamine content in the suspected tuna dumpling, this food-borne poisoning was strongly suspected to be due to histamine intoxication. In addition, although thirteen histamine-producing bacteria strains capable of producing 8.1–19.7 ppm of histamine in trypticase soy broth (TSB) supplemented with 1.0% l-histidine (TSBH), were identified as Enterobacter sp. (three strains), Pantoea agglomerans (two strains), Klebsiella variicola (four strains) and Serratia marcescens (four strains), by 16S rDNA sequencing with PCR amplification, they were not determined to be the main contributors to histamine accumulation in suspected tuna dumpling.     

Quantitative analysis of the growth of Salmonella stanley during alfalfa sprouting and evaluation of Enterobacter aerogenes as its surrogate

Raw seed sprouts have been implicated in several food poisoning outbreaks in the last 10 years. Few studies have included investigations of factors influencing the effectiveness of testing spent irrigation water, and in no studies to date has a nonpathogenic surrogate been identified as suitable for large-scale irrigation water testing trials. Alfalfa seeds were inoculated with Salmonella Stanley or its presumptive surrogate (nalidixic acid-resistant Enterobacter aerogenes) at three concentrations (-3, -30, and -300 CFU/g) and were then transferred into either flasks or a bench top-scale sprouting chamber. Microbial concentrations were determined in seeds, sprouts, and irrigation water at various times during a 4-day sprouting process. Data were fit to logistic regression models, and growth rates and maximum concentrations were compared using the generalized linear model procedure of SAS. No significant differences in growth rates were observed among samples taken from flasks or the chamber. Microbial concentrations in irrigation water were not significantly different from concentrations in sprout samples obtaihed at the same time. E. aerogenes concentrations were similar to those of Salmonella Stanley at corresponding time points for all three inoculum concentrations. Growth rates were also constant regardless of inoculum concentration or strain, except that lower inoculum concentrations resulted in lower final concentrations proportional to their initial concentrations. This research demonstrated that a nonpathogenic easy-to-isolate surrogate (nalidixic acid-resistant E. aerogenes) provides results similar to those obtained with Salmonella Stanley, supporting the use of this surrogate in future large-scale experiments.     

A risk-based sampling plan for monitoring of histamine in fish products

In 2008, the French Institute for Public Health Surveillance reported an increase in the number of histamine food poisoning outbreaks and cases in France. The aim of this study was to propose a new monitoring plan for characterizing consumers' exposure to histamine through fishery products. As fish products of concern are numerous, we proposed that the number of samples allocated for a fish category be chosen based on the risk associated with the category. Point risk estimates of histamine poisoning were assessed with the Risk Ranger tool. Fresh fish with high histidine content was found to contribute most to the number of cases. The (estimated) risks associated with the consumption of canned and deep-frozen fish appear marginal as compared with the risk associated with fresh fish with high histidine concentrations. Accordingly, we recommend excluding canned and deep-frozen fish from the monitoring plan, although these risk estimates can be biased. Within a category, samples were proportional to the relative food consumption of the different fishes. The spatial and seasonal consumption patterns were also taken into account for the design of the new monitoring plan. By testing appropriate numbers of samples from categories of fish products of concern, this plan will permit investigation of trends or comparison of product categories presenting risks of histamine poisoning.     

Source and identification of histamine-producing bacteria from fresh and temperature-abused albacore.

Histamine-producing bacteria were isolated from fresh and temperature-abused albacore using two different isolation procedures. Typically, the bacterial isolates on Niven's or modified Niven's medium produced negligible or low levels of histamine (<300 ppm) in histamine enumeration broth. The most frequently found species using this approach was Hafnia alvei. By prescreening on selective media (eosin methylene blue [EMB] agar for enteric bacteria; deMan Rogosa Sharpe agar for lactic acid bacteria: KF streptococcus agar for streptococci; pseudomonas isolation [PI] agar for pseudomonads; and staphylococcus medium 110 agar for staphylococci) prior to plating on histidine decarboxylase differential media, detection rate of true histamine formers increased. Prolific histamine producers capable of forming >1,000 ppm histamine in culture broth were isolated when PI and EMB agars were used for prescreening. Among the selective media tested, EMB agar was most effective in selecting high histamine producers, as demonstrated by the highest rate of true positives based on histamine analysis. Histamine-producing isolates were mostly enteric bacteria, including Morganella morganii, H. alvei, Klebsiella spp., Citrobacter freundii, Enterobacter spp., and Serratia spp. M. morganii isolated on PI agar from temperature-abused albacore muscle was found to be the highest histamine former. This species was not isolated from fresh albacore. while other enteric bacteria were frequently detected on the gills. However, only a few species isolated from both fresh and temperature-abused muscles were identified as high histamine formers.     

液体培养条件下产气肠杆菌与阴沟肠杆菌产生物胺交互作用研究

产气肠杆菌和阴沟肠杆菌是发酵香肠中产尸胺和腐胺的主要肠杆菌,常同时被检测分离出来。本研究将产气肠杆菌和阴沟肠杆菌按7:1、5:3、4:4、3:5、1:7的比例混合接种培养,及分别纯菌培养,发现在48h培养过程中,产气肠杆菌有较强的产尸胺能力,而阴沟肠杆菌的产腐胺能力则相对较强;两者混合接种培养时,在产尸胺方面存在明显的协同作用,几乎所用混合体系的尸胺产量都高于两个纯菌体系(P＜0.01),在产腐胺方面亦存在协同交互作用,但相对较弱,混合体系的腐胺产量一直介于两个纯菌体系之间。