Microbial Quality and Safety of Anchovy Undergone Enzyme Maturation Treatment in Brine at Processing Plant in Albania
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Albanian j. agric. sci. 2020;19 (3): 45-49 Agricultural University of Tirana
RESEARCH ARTICLE (Open Access)
Microbial Quality and Safety of Anchovy Undergone Enzyme
Maturation Treatment in Brine at Processing Plant in Albania
FATMIRA SHEHU1*, RENIS MAÇI2, BIZENA BIJO1, ERMELINDA NEXHIPI2, HALIT MEMOÇI2
1
Department of Veterinary Public Health, Faculty of Veterinary Medicine (FVM), Kodër-Kamëz, Tirana, Albania
2
Department of Food Microbiology, Food Safety and Veterinary Institute (FSVI), "Aleksandër Moisiu" No. 10, Tirana,
Albania *Corresponding author; E-mail: mshehu@ubt.edu.al
Abstract
The fish processing industry is increasingly becoming an important component in fishery sector and currently
has as significant contribution in labor market in Albania. We aimed to assess the microbiological status of
processed anchovies (Engraulis spp.) in brine from different Albania manufacturers and to identify the
contamination factor associated with these products. A total of 150 anchovies samples were collected during the
year 2018. The total aerobic mesophilic bacteria (AMB) concentration varied from 2.1 to 2.6 Log10 CFU/g in
40% of the samples analysed. Escherichia coli were present in low level in 4 out of 27 (15%) and density of
coagulase positive staphylococci varied from 2.0 to 2.2 Log10 CFU/g in 5% (4 out of 83 samples), while neiher
Salmonella spp. nor Listeria monocytogenes was found respectively in 15 and 25 anchovy samples.
Microbiological hazard analysis of practices allowed to identify the sensitive steps where hygiene measures
need to be emphasized for an improved quality control. The fluctuations in the microbial loads of AMB,
coagulase positive staphylococci, E. coli, were mainly ouline to inadequate handling or processing procedures,
thus suggesting the need for an enhancement of staff training activities and for a reorganization of tasks as part
of implementing HACCP system and complying with EU requirements.
Keywords: anchovy; E.coli; Salmonella spp; L. monocytogenes; coagulase positive staphylococci
1. Introduction
Anchovy Engraulis encrasicolus is a coastal pelagic product is marketed regionally and (ii) industrial scale
and euryhaline species that represents the only production, EU approved plants
European species of the family Engraulidae, with a that it is partly based on imports and catches from
widespread distribution. Anchovies are small salt Albanian Adriatic area [14] (Figure 1). Physico
water fish having silvery with blue-green backs. In chemical requirements for dried salted anchovies are
Albania they usually never grow larger than 13.21 ± as follows: Sodium chloride, percent by weight, max
1.17 cm. [13] Anchovies prefer the warmer waters (d.b.) 15; Water activity (aw), max 0.75; Acid
around the world, where they swim in massive insoluble ash, percent by weight, max. (d.b.) 1.5.[7]
schools. They are related to the herring belonging to The spoilage in salted and dried fish during
the family Clupeidae. Anchovy production can vary conservation is mainly due to bacterial actions
widely from year to year. The fish can appear in large especially the salt tolerant halophilic bacteria of
schools for decades and disappear the next year, and Serratia sp. Acording to a study [15], the
then reappear the year after that [8]. In Albania it is microbiological analysis of anchovy product indicate
marketed fresh, frozen, salted or marinated and the activity of the bacteria during the storage period,
mainly exported to Italy [9]. There are two scales of inspite of their reduced water activity . According to a
production of processed anchovy in Albania; (i) study the salted and pressed anchovies showed an
small-scale production of Albania anchovy where initial count of 2 × 102 log cfu/ g, which went up to
*Corresponding author: Fatmira Shehu; E-mail: mshehu@ubt.edu.al
Special Issue of the Conference:Agriculture a Life Science with Roots in Applied Biology; 3 -4 Dec.2020. (Accepted for public. 17.01.2020)
ISSN: 2218-2020, © Agricultural University of TiranaShehu et al., 2019
6.4 × 103 log cfu/ g during the 5 weeks of storage. increased to 4.1 × 102 log cfu/ g. The increase in total
This product did not show any halophilic count upto plate count during storage is due to the high water
3rd week of storage, thereafter recorded a halophilic activity.
count of 1.0 × 102 log cfu/ g, which marginally
Figure 1. Map of the Adriatic Sea showing the sampling localities. (Ruggeri et al., 2016)
Food-poisoning bacteria in most of the cases are not well as sensory and nutritional quality of fish,
considered to be found on fish in their natural habitat application of various preservation methods is
in the water. It is when they come into contact with necessary. Various hurdles are available, most of
uncleaned surfaces and are poorly handled so they can which act by inhibiting microbial growth and/or
transmit the potentially dangerous, food-poisoning inactivate microbial cells (e.g., chilling, curing and
bacteria. Thus, contamination of fish with pathogenic acidifying) [11]
microorganisms may raise safety concerns with regard
to fish used for human consumption and thus
2. Material and Methods
microbiological hazards are of great importance for
public health. Several potential foodborne hazards 2.1.1 Samples collection
reside naturally in the marine or freshwater environ-
ment. It is generally accepted that microorganisms can A total of 150 processed anchovies in brine were
be found on fish skin and gills because of exposure to collected from 7 fishery plant located in four districts
contaminated water (e.g., Escherichia coli, Vibrio in Abania.
spp., etc)[12]. In addition, pathogens such as L.
monocytogenes, Salmonella spp., and S. aureus can
contaminate fish and fish products from utensils,
personnel, and food processing environment during
handling and various processes for example, gutting
and filleting (Huss et al., 2000). to inhibit and/or
inactivate spoilage microorganisms and foodborne
pathogens, to ensure microbial safety and stability, asShehu et al., 2019
Table 1. Geografical distribution of anchovy processing plant in Albania and sample collection.
Locations of processsing Plant N. of samples %
Durrës (n=1) 11 7.3%
Elbasan (n=2) 72 48.0%
Lezhë (n=3) 36 24.0%
Shkodër (n=1) 31 20.7%
Total (n=7) 150 100%
N, number of samples analysed; n number of processing plants; %, percentage.
subcultured and confirmed by appropriate
2.1. Sample preparation and growth media
morphological and biochemical assays (β-hemolysis
Detection of pathogens of interest Salmonella and and rhamnose, xylose, and mannitol fermentation) and
Listeria monocytogenes. identified using the API® Listeria. The limit of
To start the analysis 25 gram of salted anchovies were sensitivity for the detection of L. monocytogenes was
weighted and homogenized in a stomacher for 1 min one cell in 25 g samples.
and then were inoculated into 225± 5 ml BPW and Enumeration of aerobic mesophilic bacteria. One
subject to primary enrichment. milliliter of the selected dilutions was transferred into
Another 25 gram of salted anchovies were weighted the Petri dishes, then 15 mL of Plate Count Agar
and homogenized in a stomacher for 1 min and then (PCA) medium at 45 °C was poured into each 90 mm
were inoculated into 225± 5 ml HFR and subject to plate. Petri dishes were inverted and incubated at 30
primary enrichment. °C for 72 h. Colonies in plates were counted and
For the detection of Salmonella spp., the salted viable counts in the test sample per gram were
anchovies homogenate was incubated at 37 °C for 24 calculated.
h. After this pre-enrichment step, 1 mL and 0.1 mL of Enumeration of E.coli. One milliliter of the selected
the pre-enrichment BPW were transferred, dilutions was transferred into the Petri dishes, then 15
respectively, in 10mL of Muller Kauffmann mL of TBX medium at 45 °C was poured into each 90
Tetrathionate Broth and Rappaport Vassiliadis broth mm plate. Petri dishes were inverted and incubated at
and incubated, respectively, at 37 °C and 41.5 °C for 44 °C for 24 hour. Colonies displaying the typical
24 h. After that, a loop of the two selective broths morphological characteristics (blue to blue-green) in
were spread on Xylose Lysine Deoxycholate (XLD) plates.
agar and Hectoen Enteric Agar plates and incubated at Enumeration of coagulae positive staphylococci.
37 °C for 24 h. Five colonies (or all if less than 5 Decimal dilutions in sterile BPW were prepared. One
CFU) showing the typical Salmonella morphology milliliter of the selected dilutions was transferred into
were cultured in TSB broth overnight at 37 °C and the the Bair Parker Medium. Petri dishes were inverted
confirmation was performed using API 20E and and incubated at 37°C for 24 hour.
polyvalent salmonella antisera Poly O Poly H and Vi. Coagulase positive black colonies on the selective
For the detection of Listeria monocytogenes, the media typical of Staphylococcus aureus were
method consisted of a double enrichment in half enumerated.
Fraser and Fraser selective broths. Salted anchovies
2.2. Sample testing
homogenate in half Fraser broth was incubated at 30
°C for 24 h. The second step of the enrichment was Samples were analyzed by an accredited laboratory
carried out in Fraser broth for 48 h at a temperature of [4] in Albania. Microbiological analysis on food
37 °C. A loop from cultures obtained in half Fraser samples was carried out according to standard ISO
and Fraser broths were plated on ALOA agar and methods as follow.
Oxford Agar. After incubation at 37 °C for 24–48 h, Total aerobic mesophilic plate count was tested
the colonies of presumptive L. monocytogenes were according to ISO 4833-1:2013 (Anonymous, 2013).Shehu et al., 2019
Escherichia coli glucuronidase positive at 44°C were 3. Results and Discussion
performed according to ISO 16649-2:2001 [2]
The sampling, carried out in different facilities was
Coagulase positive staphylococci count was
focused on the testing of only one food category
performed in line with ISO 6888-1:1999 [1]. Presence
related to anchovy undergone enzyme maturation
of Salmonella spp. was tested according to ISO 6579
treatment in brine. The microbiological testing on
part one 2017 (Anonymous, 2017a) and Listeria
salted anchovies is presented in Table 2.
monocytogenes to ISO 11290-1:2017 [4].
Table 2. Results of Salmonella spp., L. monocytogenes, enumeration of aerobic mesophilic colony count
(AMB), coagulase positive staphylococci, in processed anchovies with brine.
Microbiological criteria No of sample % Evaluation
Salmonella spp. 15 10.0% Not detected in 25 g
L. monocytogenes 25 16.7% Not detected in 25 g
Coagulase positive staphylococci 83 55.3% 4 out of 83 sample values between 2.0 to 2.2
Log10 CFU/g
Eshctericia coli count 27 18.0% 4 out of 27 values (less than 2 Log)
Total Plate Count 5 3.3% 3 out of 5 sample values between (2.1 to 2.6
Log10 CFU/g)
N, number of samples analysed; CFU, colony forming unit.
other species)—Part 1: Technique using Baird-
4. Conclusions
Parker agar medium. Geneva.
The results obtained from this study indicate the need 2. Anonymous. (2001). ISO 16649-2:2001
to improve the hygienic conditions of the environment Microbiology of food and animal feeding
and personal hygiene of operators. To reduce the risk stuffs—Horizontal method for the
of microbial contamination of environmental origin, enumeration of beta-glucuronidase-positive
the most immediate strategy is to take maximum care Escherichia coli—Part 2: Colony-count
of the cleaning and disinfection of work surfaces . technique at 44 degrees C using 5-bromo-4-
From this point of view, special care should be taken chloro-3-indolyl beta-D-glucuronide. Genva.
for the work surfaces which enable the development
3. Anonymous. (2013). ISO 4833-1:2013
of microbial biofilms, aggregates of bacteria
Microbiology of the food chain—Horizontal
immersed in a mucopolysaccharide matrix that
method for the enumeration of
protects them from the action of disinfectants.
microorganisms—Part 1: Colony count at 30
Although checking the necessary certificates for
°C by the pour plate technique. Geneva.
incoming materials in terms of food safety is
sufficient for preventive measures, it is also advisable 4. Anonymous. (2017a). ISO 6579-1:2017
to send occasional samples to an accredited laboratory Microbiology of the food chain—Horizontal
for verification procedures which is the part of method for the detection, enumeration and
HACCP plan. serotyping of Salmonella—Part 1: Detection of
Salmonella spp. Geneva.
5. Acknowledgements
5. Anonymous. (2017b). ISO 11290-1:2017
This work was supported by Department of Food Microbiology of the food chain—Horizontal
Microbiology, Food Safety and Veterinary Institute method for the detection and enumeration of
(FSVI). Listeria monocytogenes and of Listeria spp.
6. References —Part 1: Detection method.
6. Anonymous. (2017c). ISO/IEC 17025 General
1. Anonymous. (1999). ISO 6888-1:1999 Requirements for the Competency of Testing
Microbiology of food and animal feeding and Calibration Laboratories. Geneva.
stuffs—Horizontal method for the
enumeration of coagulase-positive 7. Codex Alimentarius Commission. (1998).
staphylococci (Staphylococcus aureus and Report of the Twenty-Third Session of the
Codex Committee on Fish and FisheryShehu et al., 2019
Products. ALINORM 99/18 Draft Standard for and fish products: A review. Annals of
Dried Salted Anchovies. CX 5/15 CL 1998/23- Microbiology, 66 (1), 1–15.
FFP.
13. Kristo R, Harizaj F, Maci A, & Kolitari J.
8. Donald, N. J., & Beamish, R. J. Synchrony of Growth Indicators of Engraulis encrasicolus,
marine fish catches and climate and ocean (Linnaeus, 1758) Populations in Albanian
regime shifts in the North Pacific Ocean. Waters for Two Seasons of their Lifecycle.
Marine and Coastal Fisheries: Dynamics. Albanian Journal of Agricultural Sciences,
Management, and Ecosystem Science, 2009. 1, 2015. v. 14 /n.1.
155–168.
14. Ruggeri, P., Splendiani, A., Occhipinti, G.,
9. EUMOFA. (2018). European market Fiorvanti, T., Santojanni, A., Leonori, I., De
observatory for fisheries and aquaculture Felice, A., Arneri, E., Procaccini, G., Catanese,
products – Case study, processed anchovy in G., Tičina, V., Bonanno, A., Nisi Cerioni, P.,
italy. Maritime Affairs and Fisheries. Giovannotti, M., Grant, W. S., & Caputo
Barucchi, V.. Biocomplexity in Populations of
10. Huss, H. H., Reilly, A., & Embarek, P. K. B.
European Anchovy in the Adriatic Sea. 2016,
Prevention and control of hazards in seafood.
PLOS ONE, 11(4), e0153061. https:// doi.org/
Food Control, 2000. 11(2), 149–156.
10.1371/ journal. pone.0153061
11. Leistner, L. Basic aspects of food preservation
15. Siriskar, D. A., Khedkar, G. D., & Lior, D.
by hurdle technology. International Journal of
Production of salted and pressed anchovies
Food Microbiology, 2000. 55 (1–3), 181–186.
(stolephorus sp.) and it’s quality evaluation
12. Novoslavskij, A., Terentjeva, M., Eizenberga, during storage. Journal of Food Science and
I., Valciņa, O., Bartkevičs, V., & Bērziņš, A. Technology, 2013. 50 (6), 1172–1178. PubMed.
(2016). Major foodborne pathogens in fish https:// doi.org/ 10.1007/s13197-011-0450-9You can also read
