Benha Veterinary Medical Journal 39 (2020) 105-110

Benha Veterinary Medical Journal

Official Journal Issued by Journal homepage: https://bvmj.journals.ekb.eg/

Faculty of
Veterinary Medicine Since 1990
Original Paper
Determination of histamine level in frozen fish in relation to bacterial contamination
Mohammed A. Hassan and Safa M. Hussam
Food Hygiene and control Department, Faculty of Veterinary Medicine, Benha University

ARTICLE INFO ABSTRACT

Keywords Histamine is a member of compounds called biogenic amines produced by decarboxylation

of free amino acids and present in a many types of foods. Production of amine in concert of
Coliform , the Biogenic Amines (BA) in fish represents a health risk to human food intoxication. In this
Enterobacteriaceae study ninety random frozen fish samples pictured by Scomber scombrus, Saurus and
Oreochromis niloticus (30 of each) were collected from markets in Kalyobia Governorate to
Histamine ,
work out the microorganism count of Coliform, Staphylococcus aureus and
Oreochromis niloticus Enterobacteriaceae and examined for the presence of amine by assay. The results of
S. aureus examination for frozen fish discovered that Enterobacteriaceae counts were recorded as 29
positive samples with percentage of (96.7%) from frozen Scomber scombrus samples, 26
Scomber scombrus
positive samples with percentage of (86.7%) from frozen Saurus samples and 24 positive
Received 06/12/2020 samples with percentage of (80%) from frozen Oreochromis niloticus . additionally, Coliform
Accepted 23/12/2020 counts were 27 positive samples with percentage of (90%) from frozen Scomber scombrus
Available On-Line samples,25 positive samples with percentage of (83.3%) from frozen Saurus samples and 23
20/01/2021 positive samples with percentage of (76.7) from frozen Oreochromis niloticus. Also, the study
discovered that S. aureus counts were 22 positive samples with percentage of (73.3%) from
examined Scomber scombrus , 17 positive samples with percentage of (56.7%) from Saurus
and 14 positive samples with percentage of (46.7%) from Oreochromis niloticus . Mistreatment
assay check, the results recorded that the amine mean values in examined fish samples were
twenty-nine.40 ± 0.31, 21.76 ±0.25 and 16.23±0.18 mg/kg for Scomber scombrus, Saurus and
Oreochromis niloticus severally. Once and for all, the appliance of early detection of biogenic
amines primarily amine manufacturing microorganism may scale back the health risk of amine
intoxication associated fish and fish product microbial deterioration. Improvement of a
selective medium to observe decarboxylating microorganism could also be a valuable tool.

1. INTRODUCTION quality. Sensory, chemical/biochemical and micro-

biological strategies conventionally wont to measure the
Fish quality is a complex concept involving many factors degree of fish freshness. Texture and color analysis
that embody for example: safety, nutritional quality, moreover as determination of post-mortem pH offer
handiness, convenience and integrity, freshness, intake indications of physical changes occurring in fish muscle
quality and therefore the obvious physical attributes of the throughout storage (Izumi, 2012).
species, size and product type (Abbas et al., 2008). The The Biogenic Amines (BA) are produced during the end of
quality of fish and fishery products represent a significant shelf life and thus their levels will be considered as a
concern in fish industry everywhere the globe (Huss et al., spoilage index instead of a high-quality index (Ozogul and
2003). Ozogul, 2006). Although several are found in fish, the most
Temperature and handling practices are necessary factors in common BA found in foods and causing histamine
determining the shelf life of all species of fish, if the fish poisoning and toxicity caused by histamine, cadaverine,
product is handled rigorously, the temperature at which is tyramine, agmatine, spermine, tryptamine, 2-
held, control its useful life. Temperature can manage the rate phenylethylamine, spermidine and ptomain (Ladero et al.,
of microorganism spoilage and enzyme breakdown, the 2010; Naila et al., 2010; Chong et al., 2011; Naila, et al.,
higher the temperature the quicker the fish spoils (Bagge- (2012).
Ravn et al., 2003) . Various species of family Enterobacteriaceae contain
Freezing preserves the taste , texture and nutritional value of histidine decarboxylase and have the ability to produce
foods better than the other strategies (George, 1997). histamine (Huang et al., 2010).
Freezing of fish cause many chemical changes as increasing Biogenic amines like ptomain, putrescine, spermidine and
in pH , total volatile base nitrogen, Thiobarbituric acid spermine don't have any adverse health effect, but
number , acid number, acid value and free fatty acids as a sometimes they react with nitrite to form carcinogenic
results of lipid oxidization in fish, whereas organoleptic nitrosamines (Onal et al., 2013).
attributes as odor, taste and texture were decreased Fresh fish don't contain free histamine however contain
throughout frozen storage period (Simeonidou et al., 1997). amino acid L-histidine. Histamine is made in fish by certain
Freshness assurance is one of the most necessary goals for microorganism capable of producing histidine
the fish industry as freshness is closely associated with decarboxylase enzyme which can converse the free

* Corresponding author: [email protected]

105
 Hassan and Hussam (2020) BVMJ 39 (2): 105-110

histidine, naturally present within the muscle of some fish, were picked up and sublimate on culture medium slopes for
to histamine (FAO/WHO, 2013). more identification.
High amounts of BA are produced considerably throughout
processing and storage of food as a results of microbial 2.2. Determination of histamine by ELISA (Leszczynskai et
contamination and inadequate storage conditions through al., 2004):
decarboxylation of specific free amino acids by exogenous - Intended use and principle of the test:
decarboxylase enzymes released by microorganisms related This enzyme immunoassay is for the quantitative
to seafood. Biogenic amines like histamine, ptomain and determination of histamine in plasma and urine as well as
ptomain are considered as indicators of fish spoilage (Lee different tissues of the body. In combination with
et al., 2015; Biji et al., 2016). supplementary kit (available for purchase separately, cat. no.
Therefore, the goal of this study is to judge the effect of BA E-1100), the assay is performed for the determination of
microorganism producing histamine on quality of frozen histamine release in heparinized whole blood and tissues of
fish the body. First, histamine is quantitively acylated. The
subsequent competitive ELISA kit uses the microtiter plate
2. MATERIAL AND METHODS format. The antigen is bound to the solid phase of the
microtiter plate. The acylated standards controls, samples,
A total of ninety random frozen fish samples represented by and the solid phase bound analyte compete for a fixed
Scomber scombrus , Saurus and Oreochromis niloticus (30 number of antiserum binding sites. After the system is in
of each) were collected from completely different fish equilibrium, free antigen and free antigen-antiserum
markets settled in Kalyobia Governorate. Every sample was complexes are removed by washing. The antibody bound to
kept in a separate sterile bag and preserved in an ice box then the solid phase is detected by an anti-rabbit IgG- peroxidase
transferred to the laboratory below complete sterile conjugate using TMB as a substrate. The reaction is
conditions while not undue delay and examined as quickly monitored at 450 nm. Quantification of unknown samples is
as doable. The collected samples were subjected to achieved by comparing their absorbance with a reference
bacteriological examination to see the connection between curve prepared with known standard concentrations.
the presence of bound microorganism and therefore the
quality of such examined frozen fish. - Test procedure:
All reagents and samples are allowed to reach room
2.1. Bacteriological examination: temperature prior to use. Measurement in duplicates is
2.1.1. Preparation of samples (ISO 4833-1, 2013): Twenty- recommended.
five grams of the sample, 225 ml of sterile organic 2.2.1. Preparation of reagents:
compound water were side and totally mixed mistreatment  Wash Buffer
sterile liquidizer for one.5 minutes, from that 10-fold serial Dilute the 20 ml wash buffer concentrate with distilled water
dilutions was ready. The ready samples were subjected to the to a final volume of 1,000 ml. Storage up to 6 months at 4 –
subsequent examinations. 8°C.
2.1.2. Enterobacteriaceae Count (ISO 4833-1, 2013)  Acylation Diluent
One ml from every of the antecedently ready dilution was The Acylation Diluent has a freezing point of 18.5°C. To
transferred into 2 separate sterile Petri-dishes to that around ensure that the Acylation Diluent is liquid when being used,
fifteen ml of sterile thawed and tempered Violet Red it must be ensured that the Acylation Diluent has reached
digestive fluid aldo hexose agar (45C) were side. once room temperature and forms a homogeneous, crystal-free
thorough commixture, the inoculated plates were allowed to solution before being used. Alternative the Acylation
solidify before being incubated at 37C for twenty-four Diluent can be stored at room temperature (20–25°C)
hours All purple colonies were then counted and therefore separate from the other kit components.
the family Enterobacteriaceae count per gram was  Acylation Reagent
calculated on plates containing 30-300 colonies and every Reconstitute each vial with 1.25 mL Acylation Diluent. The
count was recorded severally. Acylation Reagent has to be newly prepared prior to the
2.1.3. Total coliform count: assay (not longer than 1 hour in advance). If more than 1.25
Violet Red digestive fluid agar medium , constant technique mL is needed, pool the contents of 2 or 3 vials and mix
of the previous pour plate methodology was applied thoroughly.
mistreatment Violet Red digestive fluid agar medium. The
plates were incubated at 37C for twenty-four hours. All red 2.2.2. Sample preparation and acylation:
colonies measurement zero.5 metric linear unit in diameter  Pipette 25 µl of standards, 25 µl of controls, 25 µl of plasma
on the plates were then counted and therefore the average samples, 10 µl of urine samples, or 50 µl of supernatant from
range of colonies decided. the release test* into the respective wells of the reaction
2.1.4. Determination of Staph. aureus count (FDA, 2001): plate.
One ml from every of antecedently ready serial dilutions was  Add 25 µl of acylation buffer to all wells.
cover Baired Parker agar plate employing a sterile bent glass  Add 25 µl of acylation reagent to all wells.
spreader. The plates were preserved in upright position till  Incubate for 45 min at rt (20-25°c) on a shaker (approx. 600
the matter is absorbed by agar for regarding ten min, or rpm).
placed in up right within the brooder for regarding one hour.  Add 200 µl of distilled water to all wells.
The inoculated and management plates were inverted and  Incubate for 15 min. At rt (20-25°c) on a shaker (approx. 600
incubated at 37°C for forty-eight hours. once that they were rpm).
examined for colony character. The developed colonies  Take 25 µl of the prepared standards, controls, and samples
(shiny black colonies) were enumerated as presumptive for the Histamine ELISA.
Staph. aureus count/g was calculated. Also, the colonies * For the release test the Histamine Release supplementary
kit (available for purchase separately, cat. no. BA E-1100)
has to be used.

106
 Hassan and Hussam (2020) BVMJ 39 (2): 105-110

bacteriological profile of the examined frozen fish samples

2.2.3. Histamine ELISA: (Feldman et al., 2003). All data were presented as mean ±
 Pipette 25 µl of the acylated standards, controls, and samples Standard error of mean (SEM). Differences in mean of
into the appropriate wells of the histamine microtiter strips. analyzed data were considered significant at P≤0.05.
 Pipette 100 µl of the histamine antiserum into all wells and
cover plate with adhesive foil. 3. RESULTS
 Incubate for 3 hours at rt (20-25°c) on a shaker (approx. 600
rpm). Table (1) revealed that Enterobacteriaceae was isolated
 Alternatively, shake the histamine microtiter strips briefly from examined frozen fish samples represented as 29
by hand and incubate for 15 – 20 hours at 2 – 8°C. (96.7%) from Scomber scombrus , 26(86.7%) from Sảurus
 Remove the foil. Discard or aspirate the contents of the wells and 24(80%) from Oreochromus niloticus . Meanwhile, the
and wash each well 4 times thoroughly with 300 µl wash mean values of Enterobacteriaceae count (cfu/g) in the
buffer. Blot dry by tapping the inverted plate on absorbent examined samples were 2.09x104± 0.41x104, 1.27x 104 ±
material. 0.19x104 and 5.45x103 ± 0.74x103 from Mackerel, Sảurus
 Incubate for 30 min at rt (20-25°C) on a shaker (approx. 600 and Oreochromus niloticus respectively. Furthermore
rpm). ;Table (2) revealed that coliform was isolated from
 Discard or aspirate the contents of the wells and wash each examined frozen fish samples represented as 27(90%) from
well 4 times thoroughly with 300 µl wash buffer. Blot dry Scomber scombrus , 25 (83.3%) from Sảurus and 23 (76.7%)
by tapping the inverted plate on absorbent material. from Oreochromus niloticus respectively.
 Pipette 100 µl of the substrate into all wells and incubate for Tables (3) revealed that Staph. aureus was isolated from
20-30 min at rt (20-25°C) on a shaker (600 rpm). Avoid examined frozen fish samples represented as 22(73.3%)
exposure to direct sunlight. from Scomber scombrus , 17(56.7%) from Sảurus and
 Add 100 µl of the stop solution to each well and shake the 14(46.7%) from Oreochromus niloticus . Meanwhile, the
microtiter plate to ensure a homogeneous distribution of the mean values of Staph. aureus count (cfu/g) in the examined
solution. samples were 4.35x103± 0.62x103, 1.81x 103 ± 0.29x103 and
 Read the absorbance of the solution in the wells within 10 9.1x102 ± 2.06x102 from Mackerel, Sảurus and
minutes, using a microplate reader set to 450 nm with a Oreochromus niloticus, respectively.
reference wavelength between 620 nm and 650 nm. Table (4 ) indicated analysis of histamine levels mg% in the
examined samples of frozen fish with mean value of 29.40 ±
2.2.4. Calculation of results: 0.31, 21.76 ±0.25 and 16.23±0.18 for Scomber scombrus ,
Concentration of the standards
Saurus, Oreochromus niloticus, respectively .
Standard A B C D E F
Table (5) indicate correlation coefficient (r) between
Histamine (ng/mL = µg/L) 0 0.5 1.5 5 15 50 histamine levels Vs. bacteriological quality of the examined
Histamine (nmol/L) 0 4.5 13.5 45 135 450 frozen fish samples.
Conversion: Histamine (ng/mL) x 9 = Histamine (nmol/L) Table 1 Statistical analysis of Enterobacteriaceae counts (cfu/g) in the
examined samples of frozen fish (n=30).
 The calibration curve is obtained by plotting the absorbance
+ve samples
readings (calculate the mean absorbance) of the standards Frozen fish species Min Max Mean ± S.E*
No. %
(linear, y-axis) against the corresponding standard
2.09×104 ±
concentrations (logarithmic, x- axis). Use a non-linear Scomber scombrus 29 96.7 1.9×103 6.2×104
0.41×104
regression for curve fitting (e.g., spline, 4- parameter, 1.27×104 ±
Saurus 26 86.7 1.1×103 3.5×104
0.19×104
akima). The concentrations of the plasma samples and the Oreochromis 5.45×103 ±
24 80 7.0×102 8.9×103
controls can be read directly from the standard curve. niloticus 0.74×103

Table 2 Statistical analysis of Coliform counts (cfu/g) in the examined

2.2.5. Quality control: samples of frozen fish (n=30).
It is recommended to use control samples according to state +ve samples
and federal regulations. Use controls at both normal and Frozen fish species No. % Min Max Mean ± S.E*
pathological levels. The kit controls, or other commercially 1.15×104 ±
Scomber scombrus 27 90 1.3×103 2.8×104
available controls, should fall within established confidence 0.17×104
7.92×103 ±
limits. The confidence limits of the kit controls are printed Saurus 25 83.3 1.0×102 1.2×104
1.24×103
on the QC- Report. Oreochromis
23 76.7 1.0×102 5.1×103
3.07×103 ±
niloticus 0.51×103

2.2.6. Calibration: Table 3 Statistical analysis of Staph. aureus counts (cfu/g) in the examined
The binding of the antisera and the enzyme conjugates and samples of frozen fish (n=30).
the activity of the enzyme used are temperature dependent, +ve samples
and the extinction values may vary if a thermostat is not Frozen fish species No. % Min Max Mean ± S.E*
used. The higher the temperature, the higher the extinction Scomber scombrus 22 73.3 5.0×102 9.0×103
4.35×103 ±
0.62×103
values will be. The extinction values also depend on the 1.81×103 ±
Saurus 17 56.7 2.0×102 4.0×103
incubation times. The optimal temperature during the 0.29×103
Oreochromis 9.10×102 ±
Enzyme Immunoassay is between 20-25°C. In cases of niloticus
14 46.7 1.0×102
2.0×103
2.06×102
overflow, read the absorbance of the solution in the wells
within 10 minutes, using a microplate reader set to 405 nm. Table 4 Statistical analysis of histamine levels "mg%" in the examined
samples of frozen fish (n=30).
Frozen fish species Min Max Mean ± S.E
2.3. Statistical Analysis:
The obtained results were statistically evaluated by Scomber scombrus 4.65 51.78 29.40 ± 0.31

application of Analysis of Variance (ANOVA) test as well Saurus 4.02 38.93 21.76 ± 0.25

as correlation coefficient between the histamine limits Vs Oreochromis niloticus 3.51 27.47 16.23 ± 0.18

107
 Hassan and Hussam (2020) BVMJ 39 (2): 105-110

Table 5 Correlation coefficient (r) between histamine levels Vs. handlers, inadequate clean instruments, impure water, such
bacteriological quality of the examined frozen fish samples.
contamination represent a public health hazard, this agrees
Frozen fish species Scomber scombrus Saurus O.niloticus with the statement reported by (Thatcher and Clark, 1975).
Bacterial counts: Staphylococcus food poisoning is caused by ingestion of
Enterobacteriacae count +0.72** +0.66** +0.54*
Coliform count +0.63** +0.58** +0.49* food containing performed toxins secreted by
Staph. aureus count +0.59** +0.50* +0.47* Staphylococcus aureus and characterized by nausea,
* Significant correlation. ** High significant correlation
vomiting, abdominal pain and prostration usually with
symptom while not fever , through 1-6 hours after ingestion
4. DISUCSSION of contaminated food (Eley, 1996).
Dealing with results presented in Table (4), it is evident that
Fish and their products are important source of protein, and of histamine levels mg% in the examined samples of frozen
other elements necessary for the healthy body maintenance. fish with mean value of 29.40 ± 0.31, 21.76 ±0.25 and
fish constitute an important food component for a large 16.23±0.18 for Scomber scombrus, Saurus, Oreochromus
section of the world population, the fish quality is of major niloticus, respectively. Results are nearly similar to those
concern to the food processors, consumers, and public health obtained by Brink et al., (1990), Rodtong et al., (2005),
authorities. Provision of safe, wholesome and acceptable Gonzaga et al. (2009), Tao et al. (2010), Sigh et al. (2012)
fish and fish products, though control of contamination is and Humaid and Mamdoh (2014), whereas, it was higher
essential from food safety point of view. the quality of fish than those obtained by Auerswald et al. (2006) and
degrades due to a complex process in which physical, Rajapaksha and Jayakody, (2008), in contrast results were
chemical and microbiological forms of deterioration are lower than those obtained by Kim et al. (2009). Biogenic
implicated (Adebayo-Tayo et al., 2012) . amines are biologically active compounds present in variety
The detection of potential contaminants in harvested ocean of food e.g. fish the presence of biogenic amines in these
food is terribly crucial to confirm safety of consumer. foods is an indicator of spoilage (Vaciana-Nogues et al.,
Further analysis on the microbial quality assessment of fish 1997). The consumption of histamine may result in
and fishery products ought to be undertaken. Cooking toxicological effects to consumers like high blood pressure,
properly immediately before consumption is additionally an headache, diarrhea, rash and localized inflammation
effective method of reducing or eliminating risk of fresh (Rawles et al., 1996). Poisoning with amine will cause
fish-borne diseases pseudo-allergic reaction, in other words, it can produce
The results in table (1) showed that the mean values of symptoms like urticarial, symptom or spasm of bronchi, the
Enterobacteriaceae count /gm of Scomber scombrus, Saurus content of histamine is thought to be a criterion of the quality
and Oreochromus niloticus are 2.09x104 ± 0.41x104 , 1.27x of food (Fletcher et al., 1998). Histamine is produced in fish
104 ± 0.19x104 and 5.45x103 ± 0.74x 103 respectively. The tissue by the action of bacterial enzymes with the optimum
number of positive samples of Mackerel are 29 with percent temperature of amine production being 25 °C (Kim et al.,
of 96.7, Saurus are 26 with percent of 86.7 and Oreochromus 2009).
niloticus are 24 with a percent of 80. Food concerned as a Fish is implicated in HFP (histamine fish producing) include
vehicle in food borne illness, sometimes contain bacteria of both scombroid fish (mackerel, tuna and saury) and non-
uncertain significance to human health, among these scombroid fish (sardine, anchovies, blue fish) , as they
organisms Citrobacter, Klebsiella, Proteus and Enterobacter. contain large amount of free histidine (Lehane and Olley,
Some of these bacteria belong to the normal intestinal flora 2000). The hazardous level of histamine for human health
of animals and man, and many of them seem as a food has been suggested as fifty mg%, although, low levels as five
contamination. Certain members of Citrobacter are mg% are reported in histamine poisoning (Huss et al., 2003).
suspected to cause enteric infection (Cruickshank et al., Biogenic amines in fish as precursors of nitrosamines that is
1975) carcinogens (Yurchenko and decompose, 2006). The
The entire cluster of family Enterobacteriaceae is of limited presence of biogenic amines in food, is considered a health
use within the examination of food that has probably been problem because of its physiological and toxic effects (Onal,
concerned in disease outbreak. On the opposite hand, 2007).
observation of processed for safety, would like family Histamine or scombroid poisoning (scombrotoxism)(HFP)
Enterobacteriaceae because the indicator of choice (Mosel et is hypersensitivity associated with the consumption of
al., 1995) decomposed fish with toxic histamine levels (4500 ppm)
Concerning the data presented in Table (2), the mean value (Hungerford, 2010). He additionally cleared the reason for
of coliform count/gm in examined Scomber scombrus , observation Bas in sea foods is twofold: as indices of
Saurus and Oreochromus niloticus is 1.15x104 ± 0.17x104, decomposition and to prevent the potential toxicity on
7.92x103± 1.24x103 and 3.07x 103 ± 0.51x 103, respectively. human health. Symptoms of HFP vary from mild urticaria
The number of positive samples of Mackerel are 27 with and oral allergic reactions like syndrome to life threating
percent of 90 , Saurus are 25 with percent of 83.3 and cardiovascular reactions that can be mistaken for sea food
Oreochromus niloticus are 23 with percent of 76.7 allergy (Lionte, 2010).
Dealing with results presented in Table (3), it is evident that Temperature is the most important factor contributing to
the mean value of Staph. aureus counts in examined biogenic amines formation (Chong et al., 2011). High levels
Scomber scombrus, Saurus and Oreochromus niloticus are of Bas may be prevented by application of good hygiene
4.35x103 ± 0.62x 103 ,1.81x 103± 0.29x103 and 9.10x102 ± practices and proper temperature throughout handling,
2.06x102, respectively. delivery and storage (Visciano et al., 2012).
Staphylococcus species are major contaminants in fish and The content of BAs differ according to species for example
fishery products. The presence of Staph. aureus in food is Scombridae family, like tuna and bonito and Clupeidae
usually associated with improper manipulation by food family, like sardines characterized by the presence of high
handlers, who are frequently contaminated with this levels of free amine in their muscle, additionally according
microorganism (Hatakka et al., 2000). Presence of to the season of the year , genetics , environment , food ,
Staphylococcus in fish indicates its contamination from

108
 Hassan and Hussam (2020) BVMJ 39 (2): 105-110

sex, physiological stage, storage period and sampled tissue Public Health Risks of Histamine and other Biogenic
(Lee et al., 2012). Amines from Fish and Fishery Products. Meeting report.
Biogenic amines may be used as quality index , once 11. Food and Drug Administration (FDA). (2001). Fish and
formed by bacterial activity and are resistant to thermal fisheries products hazards and controls guidance, 3rd ed. US
treatment ,thus reflecting the quality of the raw material and FDA Center for Food Safety and Applied Nutrition,
hygienic conditions of food processing (Sagratini et al., Maryland.
2012). 12. Food and Drug Administration (FDA). (2011). Fish and
Preventive measures of freezing and cooling directly after Fishery Products Hazards and Controls Guidance, 4th ed.
death can prevent rapid development of enzyme histidine Department of Health and Human Services, Food and Drug
decarboxylase, because the hazard control is no possible Administration, Center for Food Safety and Applied
after formation of enzyme (Anon, 2001). Histamine Nutrition, Washington DC.
formation in fish doesn't stop at four c or below; it stops at 13. George , R.M (1997): Freezing ystems. un: nuality in
frozen storage only (Rossano et al., 2006). Toxicological , Frozen Food. Fdited dy Friczson M.C., and Hung , Yen-
levels of biogenic amines is very difficult to determine Con. Chapman and Hall, USA. P.484.
because it depends on individual characteristics and the 14. Huss, H.H .; Ababouch L. and Gram L. (2003): Assessment
presence of other amine, however , a maximum total Bas and management of seafood safety and quality , FAO
levels of 750-900 ppm has been proposed (Ladero et al., Fisheries Technical Paper 444, Rome, 230 pp.
2010) 15. u O “unternational tandards Organization” (1995):
Microbiology of food and animal feeding stuffs. ISO 10272:
5. CONCULOSIONS 1995 (E) International Standards Organization, Geneva,
Switzerland.
The current study proved that high bacterial counts of 16. u O “unternational tandards Organization” (4833-1: 2013):
Enterobacteriaceae, Coliform and Staph. ảureus is of high Microbiology of food chain- Horizontal method for the
vital correlation with histamine level enumeration of microorganisms. Part I; Colony count at
30C by the pour plate technique. International Standards
Organization, Geneva, Switzerland.
6. REFERENCES
17. u O “unternational tandards Organization” (4832: 2006):
Microbiology of food and animal feeding stuffs. Horizontal
1. Abbas,K.A.; Mohamed , M.; Jamilah, B. and Ebrahimian, method for the enumeration of coliforms: Colony count
M.(2008): A review on correlation between fish freshness technique. International Standards Organization, Geneva,
and pH during cold storage , American Journal of Switzerland.
Biochemistry and Biotechnology , 4:416-421. 18. u O “unternational tandards Organization” (16649: 2001):
2. Bagge-Ravan, D. Y.; Hjelm, M.; Christiansen, N.J.; Microbiology of food and animal feeding stuffs. Horizontal
Johansen, C. and Gram, L. (2003): The microbial ecology of method for the enumeration of glucuronidase- positive
processing equipment in different fish industries- analysis of
Escherichia coli- Part 2; Colony count technique at 44C
the microflora during processing and following cleaning and
using 5-bromo-4-chloro-3-indoly-D-glucuronide.
disinfection . International Journal of food Microbiology.
International Standards Organization, Geneva, Switzerland.
87, 239-250
19. Joshi, P.A. and Bhoir, V.S. (2011). Study of Histamine
3. Cicero, A. ; Cammilleri , G. ; Galluzzo, F.G. ; Calabrese, I.
Forming Bacteria in Commercial fish samples of Kalyan
;Pulvirenti, A. ; Giangrosso ,G. ; Cicero , N. ; Cumbo , V.;
city. Int. J. Cur. Sci. Res., 1(2): 39 – 42.
Vella ,A. ; Macaluso , A. and Ferrantelli , V. (2020).
20. Kok, T.; Worswich, D. and Gowans, E. (1996): Some
Histamine in Fish Products Randomly Collected in Southern
serological techniques for microbial and viral infections. In
Italy: A 6-Year Study. J Food Prot., 16: 241 - 248.
Practical Medical Microbiology (Collee, J.; Fraser, A.;
4. Egyptian Organization for Standardization (ES), (2006):
Marmion, B. and Simmons, A., eds.), 14th ed., Edinburgh,
Methods of analysis and testing for meat. Part 9:
Churchill Livingstone, UK.
determination of total volatile nitrogen (TVN).ES:63-
21. Krieg and Holt, J.G. (1984): Bergey's Manual of systematic
9/2006.
Bacteriology. Vol. 1, Baltimore, MD : Williams & Wilkins
5. Egyptian Organization for Standardization (ES), (2006):
22. Ladero, V.; Calles-Enríquez, M.; Fernández, M. and Alvarez
Methods of analysis and testing for meat. Part 10:
M.A. (2010).Toxicological effects of dietary biogenic
determination of thiobarbituric acid (TBA). ES: 63-10/2006.
amines. Curr. Nutr. Food Sci., 6: 145 – 156.
6. Egyption Organization for Standardization and Quality
23. Lehane, L. and Olley, J. (2000). Histamine fish poisoning
Control (EOS),(2005): Reports related to No. 1-889-2005
revisited. Int. J. Food Microbiol., 58: 1 – 37.
for frozen fish. Egyptian Standards, Ministry of Industry,
24. Leszczynska, J.; Wiedlocha, M. and Pytasz, U. (2004): The
Egypt.
histamine content in some samples of food products. Czech
7. Ekici, K. and Alisarli, M. (2008). Histamine formation and
J. Food Sci., 22: 81–86.
microbiological changes in endemic Chalcalburnus tarichi
25. Macfaddin, J. F. (2000): Biochemical tests for identification
Pallas 1811 (Inci Kefali) stored at 4oC. Arch Med Vet., 40:
medical bacteria. Williams and Wilkins Press, INC.
95 - 98.
Baltimore, Md. 21202 USA.
8. Feldman, D.; Ganon, J.; Haffman, R. and Simpson, J.
26. Pearson, D. (2006): Chemical Analysis of Foods. 11th Ed,
(2003): The solution for data analysis and presentation
Publishing Co., Churchill Livingstone, Edinburgh, London,
graphics. 2nd Ed., Abacus Lancripts, Inc., Berkeley, USA.
United Kingdom.
9. Food and Drug Administration "FDA” (2001):
27. Onal, A.; Tekkeli, S.E.K. and Onal, C. (2013). A review of
Staphylococcus aureus. Bacteriological analytical manual
th the liquid chromatographic methods for the determination
.8 Ed. Chapter12. Academic Press, Gaithersburg, UK.
biogenic amines in foods. Food Chem., 138: 509 –515.
10. Food and Agriculture Organization of the United
28. Rahimi, E.F.; Nayebpour, and Alian, F. (2012).
Nations/World Health Organization (FAO/WHO). (2013).
Determination of histamine in canned tuna fish using ELISA
method. Am. Eurasian. J. Toxicol. Sci., 4: 64 - 66.

109
 Hassan and Hussam (2020) BVMJ 39 (2): 105-110

29. Sabry, M.A.; Hayam, A.A.M.; Ashour, R.M. and Hamza, E. Laboratory Screening of Biogenic Amines Producing
(2019). Histamine- Producing Bacteria and Histamine Bacteria Potentially Threatens Human Health in Some
Induction in Retail Sardine and Mackerel from Fish Markets Egyptian Fish and Fish. Journal of Fisheries and Aquatic
in Egypt. Food borne Pathog. Dis., 16: 597 - 603. Science, 12 (3) 134-140
30. Soliman, W.S; Shaapan, R.M.; Mohamed, L.A.; Younes, A.
M.; Elgendy, M.Y. and Salah El din, D. A. (2017).

110