International Journal of Food Microbiology 128 (2008) 226–233

Contents lists available at ScienceDirect

International Journal of Food Microbiology

j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / i j f o o d m i c r o

Modelling the effect of temperature, carbon dioxide, water activity and pH on growth
and histamine formation by Morganella psychrotolerans
Jette Emborg ⁎, Paw Dalgaard
Department of Seafood Research, National Institute of Aquatic Resources, Technical University of Denmark, Søltofts Plads, Building 221, DK-2800, Kgs. Lyngby, Denmark

a r t i c l e i n f o a b s t r a c t

Article history: A mathematical model was developed to predict growth and histamine formation by Morganella psychrotolerans
Received 26 March 2008 depending on temperature (0–20 °C), atmosphere (0–100% CO2), NaCl (0.0–6.0%) and pH (5.4–6.5). Data from
Received in revised form 1 August 2008 experiments with both sterile tuna meat and Luria Bertani broth was used to develop the mathematical growth
Accepted 24 August 2008
and histamine formation model. The expanded Logistic model with a growth dampening parameter (m) of 0.7
was used as primary growth model. A primary model for histamine formation during storage was obtained by
Keywords:
Growth dampening
combining the expanded Logistic growth model with a yield factor (YHis/CFU). 120 maximum specific growth rate
Yield factor (µmax)-values were generated for M. psychrotolerans and used to model the combined effect of the studied
Tuna environmental parameters. A simple cardinal parameter type secondary model was used to model the effect of
Broth the four parameters on µmax. The maximum population density (log Nmax) was correlated with log (YHis/CFU) and a
Expanded Logistic model simple constrained polynomial (quadratic) secondary model was developed for the effect of the environmental
conditions on these model parameters. The developed model describes the effect of initial cell concentrations,
storage conditions and product characteristics on histamine formation. This is a significant progress compared to
previously available models for the effect of storage temperature only.
© 2008 Elsevier B.V. All rights reserved.

1. Introduction activity can be used to identify combinations of storage time, storage

Histamine fish poisoning (HFP) results from consumption of certain
marine finfish when these contain more than 500–1000 ppm histamine.
In seafood, histamine is formed by the bacterial enzyme histidine
decarboxylase (HDC, EC 4.1.1.22) and the substrate is so-called free
histidine which is part of the soluble extractives in finfish flesh.
Concentrations of free histidine can be as high as 10 000 to 20 000 ppm
for some finfish, particularly tuna species (Emborg et al., 2005; Fletcher et
al., 1995). Finfish becomes toxic when specific HDC-producing bacteria, at
some stage between catch and consumption, grow to high concentrations
and form significant amounts of histamine (Lehane and Olley, 2000;
Taylor, 1986). HFP is common and the intoxication occurs world-wide
(Dalgaard et al., 2008; DeWaal et al., 2006; McLauchlin et al., 2006). Since
the early 1990s, HFP has caused 32% of all the reported seafood-borne
incidents of human disease in England and Wales whereas the
corresponding value for the USA was 38%. HFP is a relatively mild
intoxication with allergy like symptoms, but due to its frequent occurrence
HFP contributes negatively to the consumer's image of seafood. In this
way, HFP is problematic due to economic losses and because it counteracts
potential health benefits from increased seafood consumption.
To reduce the frequency of HFP, information about the bacteria
actually responsible for histamine formation in the implicated products
is essential. Specifically, a quantitative description of their growth and
⁎ Corresponding author. Tel.: +45 45254918; fax: +45 45884774.
E-mail address: jem@aqua.dtu.dk (J. Emborg).
conditions and product characteristics that will reduce the risk of
histamine formation in toxic concentrations. However, compared to the
high number of HFP incidents, a surprisingly small number of studies
have identified histamine-producing bacteria directly from the impli-
cated finfish products (Dalgaard et al., 2008; Emborg and Dalgaard,
2006; Emborg et al., 2005; Havelka, 1967; Kanki et al., 2004; Kawabata
et al., 1956; Lerke et al., 1978; Sakabe, 1973; Taylor et al., 1979).
Interestingly, the early studies found mesophilic Enterobacteriaceae and
particularly Morganella morganii responsible for histamine formation
whereas, recent studies in Denmark and Japan found the psychroto-
lerant bacteria Morganella psychrotolerans and Photobacterium phos-
phoreum responsible for the production of histamine in toxic
concentrations. This new information is important because these
psychrotolerant bacteria can produce toxic concentrations of histamine
in seafood even when products are stored chilled as requested according
to the existing regulation for both EU and USA (EC, 2005; FDA/CFSAN,
2001). A recent study confirmed the importance of toxic histamine
formation during chilled storage of seafood. In fact, average concentra-
tions of more than 500 ppm histamine were observed in 18 out of 59
storage trials (31%) with naturally contaminated seafood when these
were stored between −1 °C and +5 °C (Dalgaard et al., 2008).
M. psychrotolerans has been responsible for the formation of toxic
concentrations of histamine in both fresh vacuum-packed (VP) tuna
and VP cold-smoked tuna that caused incidents of HFP (Emborg and
Dalgaard, 2006; Emborg et al., 2005). In addition, M. psychrotolerans
has been isolated as part of the dominating spoilage microflora in

0168-1605/$ – see front matter © 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.ijfoodmicro.2008.08.016
 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233 227

modified atmosphere packed (MAP) fresh tuna stored at 2 °C, frozen
and thawed MAP (26% CO2) garfish stored at 5 °C and lumpfish roe
stored at 5 °C (pH 5.4 and with 3.5–4.8% NaCl in the water phase)
(Basby et al., 1998; Dalgaard et al., 2006; Emborg and Dalgaard, 2006;
Emborg et al., 2006; Emborg et al., 2005). More precise quantitative
information, however, is not available concerning the effect of storage
conditions and product characteristics on growth and histamine
formation by M. psychrotolerans. Clearly, this makes it difficult to
determine if histamine formation by this bacterium represents an
important risk for a given chilled seafood product depending on
storage conditions and product characteristics.
The objective of the present study was to develop a mathematical
model that allows growth and histamine formation by M. psychroto-
lerans to be predicted in relevant seafoods. Growth and histamine
formation were determined for various combinations of storage
conditions (temperature, CO2) and product characteristics (NaCl/aw
and pH). Appropriate primary models for growth and histamine
formation during storage were selected and then secondary models
were developed for the effect of the environmental conditions on
relevant primary-model parameters including lag time (tlag), max-
imum specific growth rate (µmax), maximum population density
(Nmax) and the yield factor for histamine formation (YHis/CFU).
2. Materials and methods
2.1. Strains and pre-culturing
A mixture of four strains of M. psychrotolerans (Mix-Mp) was
studied. The strains (LMG 23374T = U2/3T, U2/5, JB-T11 and JB-T12)
were previously isolated from fresh and cold-smoked tuna (Emborg
and Dalgaard, 2006; Emborg et al., 2005). Prior to studies of growth
and histamine formation, the isolates were grown in brain heart
infusion broth (BHI CM225, Oxoid, Basingstoke, UK). The strains were
moved from −80 °C and initially grown for 24 h at 25 °C and
subsequently pre-cultured at 2–15 °C for 1–4 days depending on the
temperature to be used in the experiment. Pre-cultures were
harvested in the late exponential growth phase, corresponding to a
relative change in absorbance of 0.1 to 0.4 units at 540 nm (Novaspec
II, Pharmacia Biotech, Allerød, Denmark). The inoculum (Mix-Mp) was
prepared by mixing the four pre-cultures to obtain similar concentra-
tions of the isolates and then diluting as appropriate in 0.85% NaCl.
2.2. Growth and histamine formation
Experiments with tuna meat and broth containing relevant high
concentrations of the substrate histidine were used to quantify the
effect of temperature, CO2, NaCl/aw and pH on growth and histamine
formation by Mix-Mp. Table 1 shows the specific environmental
conditions studied using tuna meat or broth, respectively. Although it
is labour intensive to quantify histamine formation during storage of
tuna meat such experiments were carried out initially to evaluate
possible major differences in histamine production between tuna
meat and broth. Growth and histamine formation by M. psychrotoler-
ans were evaluated by inoculation of sterile tuna meat as no methods
for selective enumeration of this bacterium are available yet. Canned
tuna was used as a convenient source of sterile tuna meat with a high
concentration of free histidine.
2.2.1. Challenge studies with canned tuna
The effect of five different CO2 concentrations and of three
different storage temperatures on growth and histamine formation
was evaluated in experiments using canned tuna meat (Table 1, Code
A, B and C). Canned tuna in water (~ 1% NaCl), produced in Thailand
from yellowfin tuna (Thunnus albacares) meat, was obtained form a
local retailer (Amanda Seafood Ltd., Frederikshavn, Denmark). Tuna
meat from several cans (1350 g per can) was drained and mixed under
sterile conditions. 1% (v/w) of the diluted pre-culture Mix-Mp was
added to the tuna meat to obtain an initial concentration of 2–3 log
(CFU/g). The inoculum was added as three portions of 0.33% and after
each addition the tuna meat was mixed to distribute the inoculum.
Inoculated tuna meat and a batch of non-inoculated (control) tuna
meat were divided into 50 g portions and then packed in modified
atmospheres with different mixtures of N2 and CO2 (AGA, Copenha-
gen, Denmark) by using a Multivac C 500 packaging machine
(Multivac Ltd, Vejle, Denmark). A packaging film (NEN 40 HOB/
LLPDE 75, Amcore Flexibles, Horsens, Denmark) with low gas
permeability (0.45 ± 0.15 cm3 m− 2 d− 1 atm− 1 for O2 and 1.8 ± 0.6 cm3
m− 2 d− 1 atm− 1 for CO2) and a gas/tuna ratio of approximately 3:1 was
used. At regular intervals during storage samples were withdrawn in
duplicate for analysis of growth and histamine formation by M.
psychrotolerans as described in Section 2.2.3. The concentration of
NaCl and dry matter in the tuna meat was measured at the beginning
and at the end of the storage trial (See Section 2.2.3).

Table 1
Experiments used to determine the effect of four environmental parameters on growth and histamine formation by M. psychrotolerans

Type of experiments Experiments na Temp. (°C) % COb2 Water phase salt (%) pH
Growth and histamine formation
Canned tuna A 2 (10) 2 0, 17, 38, 66, 100 1.5 5.9
B 2 (6) 5, 10, 20 50 1.3 6.0
C 2 (2) 20 100 1.3 6.0
Brothc in Hungate tubes D 2 (10) 0, 5, 10, 15, 20 0 1.0 5.9
E 2 (4) 2 0 1.0 6.3, 6.5
F 2 (4) 2 0 5.0 6.2, 6.5
G 2 (4) 20 0 1.0 6.2, 6.5
H 2 (6) 20 0 5.0 5.9, 6.2, 6.5
I 2 (6) 5 0 2.0, 3.0, 4.0 5.9
J 2 (6) 10 0 3.0 5.6, 5.9, 6.3
K 2 (6) 10 0 2.0, 3.0, 4.0 5.9
L 2 (6) 5 0 3.0 5.5, 5.9, 6.3

Growth rates in brothc

Bioscreen C M 2 (22) 10 0 1.0 5.4, 5.5(2), 5.6, 5.8, 6.0,
6.3, 6.5, 5.9 (2), 6.0
N 2 (20) 10 0 0.0, 1.0, 2.0, 3.0, 4.0, 5.9
5.0, 5.3, 5.5, 5.8, 6.0
O 4 (8) 10, 17.5 0 1.0 5.9
a
Number of replicates for specific experiments. Total number of histamine and/or growth kinetics for each experiment is shown within brackets.
b
Equilibrium concentrations in headspace gas.
c
Amino acid-enriched LB Broth, Miller (244620 Difco™, Becton and Dickinson Company, Sparks, MD, USA).
228 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233

2.2.2. Experiments with broth instrument (Labsystems, Helsinki, Finland). Experiments were run in
Luria Bertani Broth, Miller (244620 Difco™, Becton and Dickinson duplicate (two microplates) and μmax-values for each treatment were
Company, Sparks, MD, USA) was used with added amino acids and lactic calculated from absorbance detection times of the serially diluted
acid to match the concentrations of these compounds in tuna meat (LB- cultures as previously described (Dalgaard and Koutsoumanis, 2001).
AA). LB-AA was prepared by mixing a previously sterilized (121 °C,
15 min) 10-fold stock solution of LB broth with a sterilized (121 °C, 2.3. Modelling and predicting the growth and histamine formation of M.
15 min) solution containing water with dissolved amino acids, lactic psychrotolerans
acid, buffer and NaCl. Final concentrations of the added amino acids,
buffer and lactic acid in LB-AA were: 50 mg/L L-arginine (A-3784 Sigma, Predictive food microbiology methods for development of math-
Steinheim, Germany), 250 mg/L L-lysine mono-hydrochloride (1.05700 ematical models to predict growth of microorganisms in foods under
Merck Darmstadt, Germany), 20 000 mg/L L-histidine mono-hydro- various environmental conditions are relatively well described
chloride (Sigma H-8125), 50 mg/L L-phenylalanine (78020 Fluka, Sigma- (McKellar and Lu, 2004; McMeekin et al., 1993). In comparison, models
Aldrich, Steinheim, Germany), 120 mg/L L-tyrosine hydrochloride for metabolite formation have been very little used within predictive
(Sigma T-2006), 7 g/L of H2KPO4 and 7 g/L of HK2PO4 and 20 700 mg/L food microbiology but extensively studied in relation to basic microbial
of a 60% w/w solution of sodium lactate (Sigma L-1375). pH was adjusted kinetics and fermentation technology (Bailey and Ollis, 1986; Pirt,
for sub-batches of LB-AA by using HCl/NaOH. 1975). The absolute rate of histamine formation (dHis/dt, mg/kg/h) can
Duplicate experiments were carried out for different combinations of be related to the absolute growth rate (dN/dt, CFU/g/h) by a constant
temperature, NaCl and pH (Table 1). Each sub-batch of LB-AA was yield factor for histamine formation (YHis/CFU, mg/CFU) (Eq. (1)). In food,
inoculated with Mix-Mp to a final concentration between 1 and 5 log bacteria can grow exponentially from very low initial concentrations
CFU/mL. Inoculated LB-AA were distributed into Hungate tubes (Bellco until they approach the maximum population density (Nmax, CFU/g). In
Hungate tubes, E. Pedersen og Søn Ltd., Oslo, Norway) which were filled this situation the dampening of growth close to Nmax can have an
with N2, sealed and stored in water baths at the different temperatures important effect on metabolites formation as shown in Fig. 1. To
studied. At regular intervals during storage samples were withdrawn for describe this growth dampening the expanded Logistic model (Eq. (2))
analysis of growth and histamine formation (See Section 2.2.3). includes the parameter m (Dalgaard, 2002; Turner et al., 1969). An
integrated version of this model was used in the present study (Eq. (3)).
2.2.3. Microbiological, physical and chemical analyses
15–25 g of canned tuna meat was diluted 10-fold in physiological saline dHis dN : 1000
¼ YHIs= : ð1Þ
(PS; 0.85% NaCl and 0.1% Bacto-peptone) and homogenised in a stomacher dt CFU dt
400 (Seward Medical, London, UK).10-fold dilutions of this homogenate or
of samples taken directly from inoculated broth were made in PS as

 
dN N m
appropriate. M. psychrotolerans was determined by spread plating (25 °C, ¼ N : μ max : 1− ð2Þ
dt Nmax
2 d) on tryptone soya agar (TSA, CM131, Oxoid, Basingstoke, UK).
Concentrations of histamine in tuna and broth were quantified by
HPLC. Post-column derivatisation with ο-phthalaldehyde (OPA, Sigma
Log Nt ¼ Log N0 tbt lag
P1378, Steinheim, Germany) was followed by excitation at 354 nm and
fluorescence detection at 430 nm as previously described (Jørgensen
et al., 2000a). Analyses were performed on filter sterilized (0.20 μm)
broth or after extraction of canned tuna meat samples (15 g) with 7.5%


   !
trichloroacetic acid (1.00807 Merck, Darmstadt, Germany).
Storage temperature in all experiments was continuously recorded by
Log Nt ¼ Log Nmax = 1þ
Nmax m
N0
 
−1 : exp −μ max :m: t−tlag
 1=m
t≥tlag

data loggers (TinytagPlus, Gemini Data Loggers Ltd., Chichester, UK). For ð3Þ
packed (previously canned) tuna meat the equilibrium concentration of 2.3.1. Primary modelling of growth and histamine formation
CO2 in the headspace gas during storage was measured by a gas analyser Eq. (3) was used to estimate lag time (tlag, h), maximum
(Combi Check 9800-1; PBI Dansensor, Ringsted, Denmark). pH in broth specific growth rate (µmax, h− 1) and maximum population density
was measured directly while pH in tuna meat was measured after dilution
of 10 g in 10 mL of demineralised water using a PHM 250 Analyser
(MetroLab™, Radiometer, Copenhagen, Denmark). The percentage of dry
matter was determined for 2 g of tuna meat after heating (102 °C, 24 h)
(AOAC International, 2007a). The concentration of NaCl in tuna meat was
determined by automated potentiometric titration (AOAC International,
2007b). The concentrations of lactic acid and free amino acids in tuna
meat were determined by different HPLC methods as previously
described (Barkholt and Jensen, 1989; Dalgaard and Jørgensen, 2000).

2.2.4. Growth rates determined in broth by absorbance measurements

In a total of 50 growth experiments the effect of 19 different
combinations of temperature (10.0 and 17.5 °C), NaCl (0.0–6.0%) and
pH (5.4–6.0) (Table 1) on the maximum specific growth rate (µmax) of
Mix-Mp was determined using LB-AA (See Section 2.2.2) and
automated absorbance measurements. For each experiment Mix-Mp
was 10-fold serially diluted to final concentrations of 105, 104, 103, 102,
101 CFU/mL in LB-AA with a specific NaCl concentration and pH. From
each of these dilutions, 250 μL inoculated LB-AA was transferred into Fig. 1. Growth (bold lines, log (CFU/g)) and histamine formation (fine lines, ppm).
Growth was simulated by the classical Logistic model (solid lines, Eq. (2), m = 1) and by
honeycomp 2 microplates (with 100 wells), added 50 μL sterile paraffin the expanded Logistic model (Eq. (2), dashed lines m = 0.4, dotted lines m = 0.25). For
oil and incubated. Changes in absorbance at 540 nm were measured each of the growth curves histamine formation was predicted using Eq. (4) with a
automatically at regular time intervals by using a Bioscreen C constant yield factor (YHis/CFU) of 10− 7.2 mg histamine/CFU.
 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233 229

(Nmax, CFU/g or CFU/mL) from individual viable count growth
curves determined for tuna meat (n = 18) or LB-AA (n = 52)
(Table 1). Eq. (3) was fitted to log-transformed concentrations of
M. psychrotolerans and for all viable count growth curves the
parameter m, that characterizes the dampening of growth when
N t approaches Nmax, was fixed to 0.4, 0.7, 1.0 and 1.3. It was not
attempted to determine the value of m by curve fitting as stable
estimates of this parameter cannot always be obtained (Dalgaard
and Koutsoumanis, 2001).
A primary model for histamine formation during storage was ob-
tained by combining the expanded Logistic growth model (Eq. (3)) with
a yield factor (YHis/CFU, mg histamine/CFU) (Eq. (4)).
Hist ¼ His0 þ YHis= : ðNt −N0 Þ : 1000 ð4Þ
CFU
In Eq. (4) Hist and His0 (ppm) are the concentrations of histamine
(ppm) at time t and 0, respectively whereas Nt and No (CFU/g or CFU/
mL) are the corresponding concentrations of M. psychrotolerans. Eq.
(4) was fitted to non-transformed data for histamine concentrations
(Hist, ppm) to estimate values of the yield factor (YHis/CFU) at specific
environmental conditions. When fitting a specific histamine forma-
tion curve, the values for Nt in Eq. (4) were obtained from Eq. (3)
(Nt = 10logNt) with the values of No, tlag, µmax and Nmax obtained from
the corresponding growth curve. Thus, the only parameter estimated
from each histamine formation curve was YHis/CFU. Each histamine

Fig. 2. Growth (open symbols) and histamine formation (solid symbols) by Morganella psychrotolerans. Data from experiment A, D and I (Table 1). Fig. 2A) Experiment D with broth at
0 °C (squares) and at 10 °C (triangles). Fig. 2C) Experiment A with modified atmosphere packed tuna meat at 2 °C with equilibrium CO2 concentration in the headspace gas of 38%
(squares) and 99.6% (circles). Fig. 2E) Experiment I with broth at 5 °C with 2% NaCl (squares), 3% NaCl (circles) and 4% NaCl (triangles). Maximum specific growth rates (µmax) of M.
psychrotolerans as affected by (Fig. 2B) temperature, (Fig. 2D) equilibrium CO2 concentrations at 2 °C and (Fig. 2F) water activity at 5 °C.
230 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233

formation curve was fitted with four different but fixed degrees of described when Eq. (4) was expanded with a maintenance coefficient
growth dampening (m = 0.4, 0.7, 1.0 and 1.3). The corresponding (M) (Eq. (6)). Eq. (6) was fitted to histamine formation curves by using
residual sum of squares (RSS) was recorded to determine the most Microsoft Excel 2003 (Microsoft Corp., Redmond, WA, USA) and
appropriate m-value. manual adjustment of YHis/CFU and M to minimize RSS.

2.3.2. Secondary modelling of growth and histamine formation 3. Results and discussion
To model the effect of the environmental parameters temperature
(T, °C), NaCl (water phase salt or aw), pH and CO2 (%) on the maximum 3.1. Growth and histamine formation
specific growth rate (µmax) a combination of a square root and a
cardinal parameter type model was used (Eq. (5), Ross and Dalgaard, The studied environmental parameters (Table 1) markedly influ-
2004). Concentrations of water phase salt (WPS) were determined enced both growth and histamine formation by M. psychrotolerans.
from concentrations of NaCl and water in fish meat. %WPS were Importantly M. psychrotolerans was able to grow to high concentra-
transformed into the corresponding water activity (aw) by using the tions and form toxic concentrations of histamine (above 500–
following equation: aw = 1 − 0.0052471 ⁎ %WPS − 0.00012206 ⁎ %WPS2 1000 ppm) at environmental conditions corresponding to those of
(Resnik and Chirife, 1988). Appropriate transformations of terms for chilled fresh and lightly preserved seafood (Fig. 2). These conditions
the different environmental parameters in Eq. (5) were determined by included 0 °C (Fig. 2A), 100% CO2 at 2 °C (Fig. 2C) and 4% NaCl at 5 °C
plotting square root-transformed or non-transformed µmax-values (Fig. 2E). Specific experiments were not designed as part of the
against values of the relevant individual environmental parameters present study to test if growth and histamine formation by M.
(Fig. 2). psychrotolerans were similar in tuna meat and broth (Table 1).
However, the average difference between observed and fitted square




μ max ¼ μ max−ref :
T−Tmin 2
Tref −Tmin
:
aw −aw min
aw opt −aw min ð: 1−10pHmin −pH Þ
:
CO2 max −CO2 2
CO2 opt −CO2 max
:n
root-transformed µmax-values were 0.03 ± 0.04 for tuna meat (n = 18)
and −0.005 ± 0.03 for broth (n = 102). For log(YHis/CFU) the residuals
ð5Þ were −0.07 ± 0.14 for tuna meat (n = 18) and 0.05 ± 0.11 for broth
(n = 50). These similar residuals suggest there was no important
In Eq. (5), µmax-ref corresponds to µmax at the selected reference difference for growth and histamine formation in tuna meat and in the
temperature (Tref = 20 °C) whereas Tmin, aw opt, aw min, pHmin, CO2 opt LB-AA broth used in the present study.
and CO2 max are model parameters (cardinal parameters) describing A simple strait line relation was observed between storage tem-
the effect of temperature, aw, pH and CO2 on µmax (Ross and Dalgaard, perature and square root-transformed µmax-values for M. psychroto-
2004). For M. psychrotolerans CO2 opt and aw opt were assumed to be 0 lerans (Fig. 2B). This suggests that sub-optimal temperatures (0–20 °C)
and 1, respectively. Terms for each of the environmental parameters in influence growth rates of M. psychrotolerans in the same way as
Eq. (5) have a value between 0 and 1. In this way Eq. (5) corresponds to observed for numerous other bacteria (Ratkowsky et al., 1982; Ross
the gamma-concept (Zwietering et al., 1992) but with the effect of and Dalgaard, 2004). The estimated theoretical minimum growth
interaction between environmental parameters (ξ) being taken into temperature (Tmin) of −6.2 °C for M. psychrotolerans (Fig. 2B, Table 2)
account as previously studied e.g. for growth of Listeria (Le Marc et al., was about 7 °C lower than the reported Tmin-values for M. morganii
2002; Mejlholm and Dalgaard, 2007). (Emborg and Dalgaard, 2008-this issue; Ratkowsky et al., 1983), but
A total of 120 square root-transformed µmax data (Table 1) were similar to Tmin-values determined for psychrotolerant pseudomonads
used to estimate values of the parameters in Eq. (5). Firstly, the value (Neumeyer et al., 1997; Ratkowsky et al., 1982) and slightly higher than
of CO2 max was estimated from data obtained in experiment A the Tmin-value for spoilage bacteria in iced fresh fish including H2S-
(Table 1). Subsequently, Eq. (5), with a fixed CO2 max-value, was fitted producing Shewanella and P. phosphoreum (Dalgaard, 2002).
(n = 120) to estimate µmax-ref, Tmin, aw min and pHmin by non-linear Increasing concentrations of CO2 reduced the growth rate and
regression. Data from experiment A, D, M and N (n = 62) could have delayed the histamine formation by M. psychrotolerans (Fig. 2C). The
been used to estimate values of µmax-ref, Tmin, aw min, pHmin and CO2 CO2 max-value of 266% CO2, corresponding to a partial pressure of
max by modelling each environmental parameter separately (Table 1, 2.7 atm, (Table 2, Fig. 2D) indicates M. psychrotolerans was more CO2
Fig. 2). The applied two step approach allowed data from experiment resistant than psychrotolerant pseudomonads and H2S-producing
B, C and D–L (n = 48) to contribute to the secondary growth model
although these experiments were primarily carried out to quantify
and model histamine formation (Table 1).
Table 2
The effect of the environmental parameters on tlag, Nmax and YHis/ Parameter values in secondary growth and histamine formation models for M.
CFU was described by simple secondary models. tlag from experiment psychrotolerans
A–L (n = 70) was related to µmax by calculation of relative lag times
Models and parameters Estimated values Standard errors Other information
(RLT = tlag/generation time = tlag ⁎ µmax/Ln(2)) (Ross and Dalgaard,
Maximum specific growth rate (µmax) model: Eq. (5)
2004). Data from experiment A–L (n = 68) and a simple constrained µmax-ref, h− 1 0.535 0.018 n = 120
linear (quadratic) model (Eq. (7)) was used to describe the effect of Tmin, °C −6.22 0.474 RSS = 0.127
important environmental parameters on log(Nmax). Finally, a correla- aw min 0.963 0.0005 r2 = 94.5%
tion between log(Nmax) and log(YHis/CFU) was established (n = 68). pHmin 5.12 0.051 RMSE = 0.033
CO2 max , %CO2 266 29 r2 = 90.0%
2.3.3. Curve fitting and statistical analysis
Maximum cell density model: Eq. (7) (Log Nmax)
SigmaStat for Windows Version 3.10 (Systat Software Inc. San Jose, b0 −2321 653 n = 68
CA, USA) was used for curve fitting and statistical analysis. Data were b1 4659 1327 RSS = 6.07
reported as mean ± standard deviation. Relation between the values of b2 −0.00914 0.00140 r2 = 79.8%
m with lowest RSS for histamine formation (Eqs. (3) and (4)) and the b3 −2330 674 RMSE = 0.31

environmental parameters was evaluated by multiple linear regres- Yield factor model: Eq. (8) (Log YHis/CFU)
sion and by using an F-test to identify significant factors. The same α −0.148 0.185 n = 68
methods were used to identify the studied environmental parameters β −0.881 0.023 RSS = 1.05
that significantly influenced log(Nmax) (Eq. (7)). An F-test was also r2 = 95.7%
RMSE = 0.13
used to test if histamine formation curves were significantly better
 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233 231

with 3–5% NaCl. It is expected, even though not documented in the

present study, that increasing concentrations of CO2 and NaCl results
in filaments of normal sized cells of M. psychrotolerans. For these
filaments it is likely that one colony observed in an agar plate originate
from several attached cells of normal size and that CO2 and NaCl,
therefore reduced the cell concentration determined as viable counts
more than the corresponding histamine formation.

3.2. Modelling and predicting the growth and histamine formation for M.
psychrotolerans

3.2.1. Primary modelling of growth and histamine formation

For the 70 histamine formation curves generated in the present
study a value of 0.7 for the growth dampening parameter m in Eq. (3)
resulted in lowest RSS-values on average. For different specific
experiments m-values of 0.4, 0.7, 1.0 and 1.3 all resulted in the lowest
RSS-value but no significant effect of the studied environmental
parameters on m-values with lowest RSS was identified by multiple
linear regression (P N 0.05). The m-value of 0.7 was therefore selected
as the most appropriate to use.
Previously, Bermejo et al. (2004) evaluated growth and histamine
formation for a mixed microflora from jack mackerel. In that study
both the appropriate growth dampening parameter and the yield
factor for histamine formation differed markedly between experi-
ments with broth and fish muscle. The m-value of 0.7 from the present
study, however, cannot be directly compared to data from Bermejo
et al. (2004) as they studied a mixed microflora. Jørgensen et al.
(2000b) used the classical Logistic model with m of 1.0, and a constant
Fig. 3. Growth (circles) and histamine formation (triangles) by Morganella psychrotoler- yield factor to describe growth and histamine formation by P.
ans in broth at 2 °C. Data show replicate 1 (Fig. 3A) and replicate 2 (Fig. 3B) from phosphoreum in cold-smoked salmon. Other more flexible models,
experiment E (Table 1) with pH 6.3. Growth curves were fitted with Eq. (3) and
however, were not evaluated in that study.
histamine formation curves were described using Eq. (3) together with Eq. (1) (solid
lines) and Eq. (3) together with Eq. (6) (dashed lines). Fitted parameter values for As shown in Fig. 3 histamine formation curves could be appro-
growth and histamine formation are shown in Table 3. priately described by combining a growth model (Eq. (3)) and a simple
yield factor (Eq. (1)). However, not all histamine formation curves
seemed appropriately fitted by this simple approach as shown in
Shewanella, approximately as CO2 sensitive as lactic acid bacteria and Fig. 3B. Expanding the differential form of the simple histamine
less CO2 resistant than P. phosphoreum (Dalgaard, 2002). The aw min- formation model (Eq. (1)) by adding a maintenance coefficient (M, mg
value of 0.963 (Fig. 2F, Table 2) suggests M. psychrotolerans was histamine/CFU/h) as shown in Eq. (6) improved the ability of the
slightly less NaCl-resistant than other psychrotolerant and Gram model to describe the histamine formation in some experiments
negative bacteria from fresh and lightly preserved seafood including (Fig. 3B and Table 3). An F-test showed that 42 of the 70 histamine
pseudomonads, H2S-producing Shewanella and P. phosphoreum (Dal- formation curves were better fitted (P b 0.05) by Eq. (6) compared to
gaard, 2002). This, however, is not in agreement with data from Eq. (1). It is important to note, however, that introducing M into Eq. (6)
challenge tests where M. psychrotolerans grew faster than P. had little effect on estimated values of the yield factor (YHis/CFU)
phosphoreum in vacuum-packed cold-smoked tuna with 4.4 ± 0.8% (Table 3) as well as on the predicted time to formation of histamine
WPS (Emborg and Dalgaard, 2006). One explanation for this could be concentrations below 2000 ppm (Fig. 3). Consequently, to predict
that the smoke components in that product inhibited the growth of P.
phosphoreum more than the growth of M. psychrotolerans. The effect of
smoke components on growth of these bacteria, however, is not Table 3
known and further research is relevant since both organisms have Evaluation of primary histamine formation model for M. psychrotolerans
been involved in outbreaks of HFP due to smoked seafood (Emborg Parameter values and statisticsa
and Dalgaard, 2006).
Ab Bb
Increasing concentrations of CO2 and NaCl reduced both µmax and
Log (N0, CFU/mL) 1.4 1.4
Nmax for M. psychrotolerans (Fig. 2C, E). Interestingly, a close to ten-fold Log (Nmax, CFU/mL) 8.3 8.3
(88%) reduction of Nmax from 108.3 CFU/g to 107.4 CFU/g, as observed Lag time, hours 32 10
with 2% NaCl compared to 4% NaCl did not result in a proportional µmax, hours− 1 0.0390 0.0385
decrease in histamine formation. In fact, the corresponding histamine Eq. (1)
Log (YHis/CFU, mg/CFU) −7.530 −7.481
concentrations decreased from 10 200 ppm to 7400 ppm (27%). That Residual sum of squares (RSS) 1.01 ⁎ 106 3.95 ⁎ 106
each cell of M. psychrotolerans seems to produce more histamine with Eq. (6)
4% NaCl compared to 2% NaCl might be explained by an apparent Log (YHis/CFU, mg/CFU) −7.525 −7.550
elongation of cells when exposed to salt stress. Previously elongated Maintenance coefficient, Log(mg/CFU/h)) −12.30 −10.55
Residual sum of squares (RSS) 9.80 ⁎ 105 1.88 ⁎ 106
cells have been observed for Salmonella and Listeria when exposed to
Difference between Eqs. (1) and (6)
high NaCl concentrations (Geng et al., 2003; Hazeleger et al., 2006; Yield factor (mg/CFU) 1% 15%
Mattick et al., 2000; Mukhopadhyay et al., 2006). In those studies it Residual sum of squares (RSS) 3% 52%
was documented that filaments formed by Salmonella and Listeria a
Corresponding growth and histamine formation curves are shown in Fig. 3.
were composed of several normal sized cells. For M. psychrotolerans b
Replicates from a growth and histamine formation experiment with M. psychrotolerans
Emborg (2007) observed a marked elongation of cells when grown in amino acid-enriched LB Broth, Miller at 2 °C (pH 6.3, 1% water phase salt and 0% CO2).
232 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233
histamine formation of importance in seafood a simple model with a
yield factor but without a maintenance coefficient (i.e. Eq. (1)) seemed
sufficient from a practical point of view and this approach was used
within the present study (Eqs. (1) and (4)).


dHis dN
¼ YHis=CFU : þ M : N : 1000 ð6Þ
dt dt
3.2.2. Secondary modelling of growth and histamine formation
The secondary µmax-model for M. psychrotolerans (Eq. (5)) explained
94.5% of the variation in the experimental data (Table 2) corresponding
to a residual mean square error (RMSE) of 0.033. The inclusion of an
interaction term (ξ) in Eq. (5) did not affect the parameter values or
residual sum of squares (RSS). However, this may change when more
data becomes available in the growth boundary region.
Lag times for M. psychrotolerans depended on the environmental
parameters and RLT showed some variability being 2.55±2.82 (n=70).
These values are similar to data previously reported for E. coli and Listeria
monocytogenes (Augustin and Carlier, 2000; Ross, 1999). RLT depends on
the initial physiological condition of cells (Ross and Dalgaard, 2004) and
the observed variability for M. psychrotolerans probably reflects the pre-
culturing at different temperatures between 2 °C and 15 °C.
The maximum population density for M. psychrotolerans decreased
with (i) increasing concentrations of CO2 and (ii) increasing concen-
tration of NaCl, corresponding to decreasing aw-values (Fig. 2C and E).
In fact, it was shown by multiple linear regression that aw and CO2 had
a statistically significant effect on log(Nmax) (P b 0.05). Eq. (7) described
79.8% of the variation in the log(Nmax) data (Table 2) corresponding to
a RMSE of 0.308. To avoid modelling of errors, Eq. (7) was constrained
by A logðNmax Þ
ACO2 b0 corresponding to b2 b 0 and by A logAaðNwmax Þ N0 correspond-
ing to b1 + 2b3 N 0. The fitted parameter values with these constrains
are reported in Table 2. It was previously observed that CO2 reduced
Nmax for pseudomonas (Willocx et al., 1993) and that the final optical
density for cultures of Escherichia coli decreased with decreasing aw
(Krist et al., 1998). However, in many predictive modelling studies a
similar effect of NaCl and CO2 has not been significant and this effect is
for example not included in either the ComBase Predictor (www.
combase.cc) or the Pathogen Modeling Program (http://www.arserrc.
gov/mfs/PMP7_Start.htm).
LogðNmax Þ ¼ b0 þ b1 : aw þ b2 : kCO2 þ b3 : ðaw Þ2 ð7Þ
YHis/CFU was influenced by the environmental parameters but a
simple secondary model was obtained by relating log(YHis/CFU) and log
(Nmax) (Eq. (8), Table 2). The RMSE was 0.13.
 
Log YHis=CFU ¼ α þ β : LogðNmax Þ ð8Þ
Clearly, the maximum concentration of histamine formed depends
on the available concentration of the substrate i.e. the free amino acid
histidine. The canned tuna meat used in the present study contained
from 7000 to 8600 ppm (n =6) of free histidine resulting in a maximum
theoretical histamine formation within the range of 5000 to 6150 ppm.
This corresponded well with the concentrations of histamine actually
produced by M. psychrotolerans (Fig. 2C). However, when the developed
model is used to predict histamine formation in fish flesh with low
concentrations of free histidine the theoretical maximum histamine
formation must be taken into account by introducing an absolute rate of
histamine formation (dHis/dt) equal to zero from the time when the
substrate is consumed and no longer is available for histamine formation.
To evaluate histamine formation, simple empirical models have been
suggested previously for the effect of low storage temperatures (−1.1 to
15.6 °C) and higher storage temperatures (21.1 to 37.8 °C) on skipjack
tuna (Frank and Yoshinaga, 1987; Frank et al., 1983). These models
however, have not been successfully validated for other fish species
(Dalgaard et al., 2008; Frank and Yoshinaga, 1987; Frank et al., 1983).
Models relating growth and histamine formation during storage have
been suggested for P. phosphoreum in cold-smoked salmon at 5 °C
(Jørgensen et al., 2000b), for M. morganii (Torres et al., 2002) and for a
mixed microflora (Bermejo et al., 2004) in mackerel. However, secondary
mathematical models to predict the effect of different combinations of
storage conditions and product characteristics on histamine formation in
seafood have not previously been developed. Thus, the model developed
in the present study for M. psychrotolerans represent considerable
progress with respect to prediction of histamine formation depending on
both storage conditions and product characteristics.
Evaluation and product validation of the developed M. psychrotolerans
growth and histamine formation model is important and this aspect
has been included as part of separate study (Emborg and Dalgaard, 2008-
this issue). In brief, they showed that the new model developed in the
present study successfully predicted growth of M. psychrotolerans in
seafood at constant and variable storage temperature. The only
unacceptable prediction was obtained for cold-smoked tuna and further
evaluation of the growth model is needed for naturally contaminated
products and seafood with high concentration of NaCl (Emborg and
Dalgaard, 2008-this issue). The new M. psychrotolerans-histamine-forma-
tion model on average provided slightly conservative (fail-safe) prediction
for time to toxic concentrations of histamine in seafood (500 and
2000 ppm). Possible improvements of the model are discussed by Emborg
and Dalgaard (2008-this issue).
The M. psychrotolerans-histamine-formation model developed in the
present study can support decisions concerning histamine formation in
chilled finfish products. Predictions are not highly accurate but
information on potential histamine formation depending on storage
conditions, product characteristics and initial concentration of M.
psychrotolerans can be obtained rapidly and seems to correspond well
with much more costly and time consuming challenge studies (Emborg
and Dalgaard, 2008-this issue). The developed model has been included
the Seafood Spoilage and Safety Predictor software version 3 from 2008
and in this way the model becomes accessible and easy to use (www.
difres.dk/micro/sssp/). To evaluate histamine formation for various
seafoods and storage conditions it is relevant to develop similar model
for other histamine-producing bacteria, particularly, M. morganii and P.
phosphoreum. Such new histamine formation models may be developed
by using the same kinetic approach as applied in the present study
where a classical growth model with variable growth dampening (Eq.
(3)) has been related to histamine formation by a constant yield factor
(Eq. (4)). Within predictive food microbiology models for growth and
metabolite formation have been little studied. Nevertheless, the
importance of such models to predict formation of microbial toxins or
production of metabolites responsible for food spoilage seems con-
siderable. Predictive microbiology growth models typically focus on lag
time and growth rate to predict the time required to reach critical cell
concentrations of importance to food safety or quality. To model growth
and metabolite formation it becomes important to have more precise
information on how the maximum cell density (Nmax) and yield factors
depend on storage conditions and product characteristics.
Acknowledgements
The authors thank Tina Dahl Dewitt, Lina Pedersen and Nadereh
Samieian for excellent technical assistance. Most of the research was
carried out within the EU Integrated Project SEAFOODplus, contract
no. FOOD-CT-2004-506359. Partial financing of the work by the
European Union is gratefully acknowledged.
References
AOAC International, 2007a. AOAC official methods of analysis, Method 950.46, Moisture
in meat. 18th ed. AOAC International, Gaithersburg, Maryland, USA.
AOAC International 2007b. AOAC official methods of analysis. 18th ed. Method 976.18,
Salt (chlorine and sodium chloride) in seafood. Potentiometric method, in
 J. Emborg, P. Dalgaard / International Journal of Food Microbiology 128 (2008) 226–233
combination with 937.07, Fish and marine products. Treatment and preparation of
sample procedure, and 971.27, Sodium chloride in canned vegetables. Potentio-
metric method. AOAC International, Gaithersburg, Maryland, USA.
Augustin, J.-C., Carlier, V., 2000. Mathematical modelling of the growth rate and lag time
for Listeria monocytogenes. International Journal of Food Microbiology 56, 29–51.
Bailey, J.E., Ollis, D.F., 1986. Biochemical Engineering Fundamentals, 2 edn. McGraw-Hill
Book Company, New York, USA.
Barkholt, V., Jensen, A.L., 1989. Amino-acid analysis — determination of cysteine plus
half-cystine in proteins after hydrochloric-acid hydrolysis with a disulfide
compound as additive. Analytical Biochemistry 177, 318–322.
Basby, M., Jeppesen, V.F., Huss, H.H., 1998. Characterization of the microflora of lightly
salted lumpfish (Cyclopterus lumpus) roe stored at 5 °C. Journal of Aquatic Food
Product Technology 7, 35–51.
Bermejo, A., Mondaca, M.A., Roeckel, M., Marti, M.C., 2004. Bacterial formation of
histamine in jack mackerel (Trachurus symmetricus). Journal of Food Processing and
Preservation 28, 201–222.
Dalgaard, P., 2002. Modelling and predicting the shelf-life of seafood. In: Bremner, H.A.
(Ed.), Safety and Quality Issues in Fish Processing. Woodhead Publishing Ltd.,
Cambridge, England, pp. 191–219.
Dalgaard, P., Jørgensen, L.V., 2000. Cooked and brined shrimps packed in a modified
atmosphere have a shelf life of N 7 months at 0 °C, but spoil in 4–6 days at 25 °C.
International Journal of Food Science and Technology 35, 431–442.
Dalgaard, P., Koutsoumanis, K., 2001. Comparison of maximum specific growth rates
and lag times estimated from absorbance and viable count data by different
mathematical models. Journal of Microbiological Methods 43, 183–196.
Dalgaard, P., Madsen, H.L., Samieian, N., Emborg, J., 2006. Biogenic amines formation
and microbial spoilage in chilled garfish (Belone belone belone) — effect of modified
atmosphere packaging and previous frozen storage. Journal of Applied Microbiol-
ogy 101, 80–95.
Dalgaard, P., Emborg, J., Kjølby, A., Sørensen, N.D., Ballin, N.Z., 2008. Histamine and
biogenic amines — formation and importance in seafood. In: Børresen, T. (Ed.),
Improving seafood products for the consumer. Woodhead Publishing Ltd.,
Cambridge, England, pp. 292–324.
DeWaal, C.S., Hicks, G., Barlow, K., Alderton, L., Vegosen, L., 2006. Foods associated with
food-borne illness outbreaks from 1990 through 2003. Journal of Food Protection
26, 466–473.
EC, 2005. Commission regulation (EC) No 2073/2005 of 15 November 2005 on
microbiological criteria for foodstuffs. Official Journal of the European Union 338,1–25.
Emborg, J. , 2007. Morganella psychrotolerans — identification, histamine formation and
importance for histamine poisoning. Ph.D. thesis. Technical University of Denmark,
Danish Institute for Fisheries Research. Lyngby, Denmark, 97 pp.
Emborg, J., Dalgaard, P., 2006. Formation of histamine and biogenic amines in cold-
smoked tuna: an investigation of psychrotolerant bacteria from samples implicated
in cases of histamine fish poisoning. Journal of Food Protection 69, 897–906.
Emborg, J., Dalgaard, P., 2008. Growth, inactivation and histamine formation of Mor-
ganella psychrotolerans and Morganella morganii — development and evaluation of
predictive models. International Journal of Food Microbiology 128, 234–243 (this
issue).
Emborg, J., Laursen, B.G., Dalgaard, P., 2005. Significant histamine formation in tuna
(Thunnus albacares) at 2 °C — effect of vacuum- and modified atmosphere-
packaging on psychrotolerant bacteria. International Journal of Food Microbiology
101, 263–279.
Emborg, J., Dalgaard, P., Ahrens, P., 2006. Morganella psychrotolerans sp. nov., a
histamine-producing bacterium isolated from various seafoods. International
Journal of Systematic and Evolutionary Microbiology 56, 2473–2479.
FDA/CFSAN 2001. Scombrotoxin (histamine) formation, Fish and Fishery Products Hazards
and Controls Guidance. 3 edn, Food and Drug Administration, Center for Food Safety
and Applied Nutrition, Office of Seafood, Washington, DC, USA, pp. 83–102.
Fletcher, G.C., Summers, G., Winchester, R.V., Wong, R.J., 1995. Histamine and histidine
in New Zealand marine fish and shellfish species, particularly kahawai (Arripis
trutta). Journal of Aquatic Food Product Technology 4, 53–74.
Frank, H.A., Yoshinaga, D.H., 1987. Table for estimating histamine formation in skipjack
tuna, Katsuwonus pelamis, at low nonfreezing temperature. Marine Fisheries
Review 49, 67–70.
Frank, H.A., Yoshinaga, D.H., Wu, I.P., 1983. Nomograph for estimating histamine formation
in skipjack tuna at elevated temperatures. Marine Fisheries Review 45, 40–44.
Geng, T., Kim, K.P., Gomez, R., Sherman, D.M., Bashir, R., Ladisch, M.R., Bhunia, A.K., 2003.
Expression of cellular antigens of Listeria monocytogenes that react with mono-
clonal antibodies C11E9 and EM-7G1 under acid-, salt- or temperature-induced
stress environments. Journal of Applied Microbiology 95, 762–772.
Havelka, B., 1967. The role of bacteria from Hafnia genus in the origination of histamine
in tunny meat. Ceskoslovenska hygiena 12, 343–352.
Hazeleger, W.C., Dalvoorde, M., Beumer, R.R., 2006. Fluorescence microscopy of NaCl-
stressed, elongated Salmonella and Listeria cells reveals the presence of septa in
filaments. International Journal of Food Microbiology 112, 288–290.
Jørgensen, L.V., Dalgaard, P., Huss, H.H., 2000a. Multiple compound quality index for
cold-smoked salmon (Salmo salar) developed by multivariate regression of biogenic
amines and pH. Journal of Agricultural and Food Chemistry 48, 2448–2453.
233
Jørgensen, L.V., Huss, H.H., Dalgaard, P., 2000b. The effect of biogenic amine production
by single bacterial cultures and metabiosis in cold-smoked salmon. Journal of
Applied Microbiology 89, 920–934.
Kanki, M., Yoda, T., Ishibashi, M., Tsukamoto, T., 2004. Photobacterium phosphoreum
caused a histamine fish poisoning incident. International Journal of Food
Microbiology 92, 79–87.
Kawabata, T., Ishizaka, K., Miura, T., Sasaki, T., 1956. Studies on the food poisoning
associated with putrefaction of marine products. VII. An outbreak of allergy-like
food poisoning caused by ‘sashimi’ of Parathunnus mebachi and the isolation of
causative bacteria. Bulletin of the Japanese Society of Scientific Fisheries 22, 41–47.
Krist, K.A., Ross, T., McMeekin, T.A., 1998. Final optical density and growth rate; effects of
temperature and NaCl differ from acidity. International Journal of Food Micro-
biology 43, 195–203.
Le Marc, Y., Huchet, V., Bourgeois, C.M., Guyonnet, J.P., Mafart, P., Thuault, D., 2002.
Modelling the growth kinetics of Listeria as a function of temperature, pH and
organic acid concentration. International Journal of Food Microbiology 73, 219–237.
Lehane, L., Olley, J., 2000. Histamine fish poisoning revisited. International Journal of
Food Microbiology 58, 1–37.
Lerke, P.A., Werner, S.B., Taylor, S.L., Guthertz, L.S., 1978. Scombroid poisoning — report
of an outbreak. Western Journal of Medicine 129, 381–386.
Mattick, K.L., Jørgensen, F., Legan, J.D., Cole, M.B., Porter, J., Lappin-Scott, H.M.,
Humphrey, T.J., 2000. Survival and filamentation of Salmonella enterica serovar
Enteritidis PT4 and Salmonella enterica serovar Typhimurium DT104 at low water
activity. Applied and Environmental Microbiology 66, 1274–1279.
McKellar, R.C., Lu, X., 2004. Modeling Microbial Responses in Food. CRC Press, Boca
Raton, USA. 343 pp.
McLauchlin, J., Little, C.L., Grant, K.A., Mithani, V., 2006. Scombrotoxic fish poisoning.
Journal of Public Health 28, 61–62.
McMeekin, T.A., Olley, J., Ross, T., Ratkowsky, D.A., 1993. Predictive Microbiology: Theory
and Application. Research Studies Press Ltd., Taunton, UK. 340 pp.
Mejlholm, O., Dalgaard, P., 2007. Modeling and predicting the growth boundary of
Listeria monocytogenes in lightly preserved seafood. Journal of Food Protection 70,
70–84.
Mukhopadhyay, A., He, Z.L., Alm, E.J., Arkin, A.P., Baidoo, E.E., Borglin, S.C., Chen, W.Q.,
Hazen, T.C., He, Q., Holman, H.Y., Huang, K., Huang, R., Joyner, D.C., Katz, N., Keller,
M., Oeller, P., Redding, A., Sun, J., Wall, J., Wei, J., Yang, Z.M., Yen, H.C., Zhou, J.Z.,
Keasling, J.D., 2006. Salt stress in Desulfovibrio vulgaris Hildenborough: an
integrated genomics approach. Journal of Bacteriology 188, 4068–4078.
Neumeyer, K., Ross, T., McMeekin, T.A., 1997. Development of a predictive model to
describe the effect of temperature and water activity on the growth of spoilage
pseudomonads. International Journal of Food Microbiology 38, 45–54.
Pirt, S.J., 1975. Principles of Microbial and Cell Cultivation. Blackwell Scientific
Publication, Oxford, England. 274 pp.
Ratkowsky, D.A., Olley, J., McMeekin, T.A., Ball, A., 1982. Relationship between
temperature and growth-rate of bacterial cultures. Journal of Bacteriology 149, 1–5.
Ratkowsky, D.A., Lowry, R.K., McMeekin, T.A., Stokes, A.N., Chandler, R.E., 1983. Model
for bacterial culture growth rate throughout the entire biokinetic temperature
range. Journal of Bacteriology 154, 1222–1226.
Resnik, S.L., Chirife, J., 1988. Proposed theoretical water activity values at various
temperatures for selected solutions to be used as reference sources in the range of
microbial-growth. Journal of Food Protection 51, 419–423.
Ross, T., 1999. Predictive Microbiology for the Meat Industry. Meat and Livestock
Australia. 196 pp.
Ross, T., Dalgaard, P., 2004. Secondary models. In: McKellar, R.C., Lu, X. (Eds.), Modeling
Microbial Responses in Food. CRC Press, Boca Raton, pp. 63–150.
Sakabe, Y., 1973. Studies on allergylike food poisoning 2. Studies on the relation
between storage temperature and histamine production. Journal of the Nara
Medical Association 24, 257–264.
Taylor, S.L., 1986. Histamine food poisoning: toxicology and clinical aspects. CRC Critical
Reviews in Toxicology 17, 91–128.
Taylor, S.L., Guthertz, L.S., Leatherwood, M., Lieber, E.R., 1979. Histamine production by
Klebsiella pneumoniae and an incident of scombroid fish poisoning. Applied and
Environmental Microbiology 37, 274–278.
Torres, S., Roeckel, M., Marti, M.C., 2002. Histamine formation by Morganella morganii
isolated from Trachurus murphyii (Chilean mackerel). Latin American Applied
Research 32, 209–214.
Turner, M.E., Blumenst, B.A., Sebaugh, J.L., 1969. A generalization of the Logistic law of
growth. Biometrics 25, 577–580.
Willocx, F., Mercier, M., Hendrickx, M., Tobback, P., 1993. Modeling the influence of
temperature and carbon-dioxide upon the growth of Pseudomonas fluorescens.
Food Microbiology 10, 159–173.
Zwietering, M.H., Wijtzes, T., De Wit, J.C., Van't Riet, K., 1992. A decision support system
for prediction of the microbial spoilage in foods. Journal of Food Protection 55,
973–979.