In this contract stage, there have been aimed:
– setting a portfolio of laboratory analysis necessary to verify specific indicators of fish welfare in farms, during transport and slaughter, and conducting these analysis;
– establish a portfolio of specific rules for fish welfare during transport, its evaluation in an experiment involving two transport systems – by automobiles, in tanks and by water, in floating cages , systems quite different in terms of storage density and transport conditions (transportation management, water quality, physical factors – vibration, etc.)..
I) Laboratory tests useful in assessing fish welfare
Water quality (as fish environment) is most important in determining the level of fish welfare, so the main objective welfare indicator for these species is the physical – chemical and microbiological characteristics of water. Another indicator of fish welfare is the blood biochemical profile.
Therefore, in this phase of works we aimed to assess the welfare of fish in four farms (three for cyprinids rearing: unsystematic fishery Fundulea – Frunzăreni and systematic fisheries Ciocăneşti and Ulmu and one for salmonids rearing: applying the floating cage systems on Bicaz lake – Potoci) based on laboratory analysis of water (physical-chemical parameters and microbiological parameters) and blood biochemical parameters.
► Thus, in the Fundulea – Frunzăreni fishery there were collected water samples, of which were determined: pH, dissolved oxygen, ammonia, nitrites, nitrates, phosphates, residual chlorine , total iron, sulphates and detergents, but also was established fish serum profile: BUN, creatinine, phospate, calcium, albumin, alanine transaminase, cholesterol, triglycerides, glucose, lactate dehydrogenase.
Water sampling was performed automatically using GSP 4 sampler, and the analysis was done with Spectroquant Nova 60 photocolorimeter. To determine the constituents of blood serum biochemical profile was used the biochemical analyzer Vettest 8008 (all devices have been purchased from the research contract funds).
Blood collection was performed after anesthesia with Finquel: MS-222 . Fish were restraining by placing on a small table, then have been harvested blood either by caudal vein puncture with ventral approach, either through the dorsal aorta puncture. In fish where these methods have not worked (under 10 centimeters lenght), had been practiced the cardiac blood collection or caudal peduncle cut. Blood was collected directly in tubes without anticoagulant. Was performed a mixing of blood samples obtained from 10 specimens of common carp (Cyprinus carpio), achieving three blood samples (results are summarized in tab. 2). After syneresis, accelerated by heat and maintaining the samples in special coagulation tubes (with slope bottom and clotting ring ), serum was transported to our laboratory in FMV Bucharest, discipline of Animal Welfare, was preserved by freezing until analysis (1-3 days). Because up to present have not been established for fish a blood biochemical profile in our country or at international level, we used blood parameters reference levels present in works of foreign authors (Hemming TA, 1989, Jen-Lee Yang 2003; Luskova, V, 2002, Palmeiro BS, 2007, Thrall, MA, 2004). Researches of this project are first attempts at national level for fish welfare assessment based on fish blood profile. Our intention is to collect as much samples as possible to design a profile of blood biochemistry useful in assessing fish welfare (topic insufficiently addressed in Romania, but at European level has payed a significant attention-COST 867 European projects in progress, Benefish, Wealth, Fastfish, Eurofish, Welfish, Aquafirst).
Water quality was assessed on water harvested from three places, namely: the water admission, center of the pond and the draining point – for each place three samples were collected (surface, 2 m depth and 3 m depth). Of water samples collected were determined following physical – chemical parameters: the pH, dissolved oxygen, ammonia, phosphates, nitrates, nitrites, total iron, copper, chlorine, detergents (Table 1).
Tab. 1: Physical – chemical parameters of water quality in the Fundulea – Frunzareni fishery
|
Sampling place
|
Sampling point
|
Assessed parameters
|
|||||||||
|
pH
|
O2
|
NH4+
|
PO43 –
|
NO3–
|
NO2–
|
Fe
|
Cu
|
Cl2
|
Detergents
|
||
| Water admission |
surface
|
6.7
|
4.3
|
0
|
0.9
|
0
|
0.013
|
o.1
|
0.05
|
0.17
|
0.02
|
|
2 m
deep
|
10.25
|
0
|
1
|
0
|
|||||||
|
3 m
deep
|
8.15
|
nedec.
|
1.5
|
0
|
|||||||
|
Center of the pond
|
surface
|
7.5
|
5.6
|
0.2
|
0.8
|
0
|
|||||
|
2 m
deep
|
8.1
|
0.5
|
0.8
|
0
|
|||||||
|
3 m
deep
|
5.4
|
nedec.
|
4.2
|
0
|
|||||||
|
Draining point
|
surface
|
7.3
|
7.8
|
0.4
|
0.5
|
1
|
|||||
|
2 m
deep
|
10.4
|
0.5
|
0.7
|
7
|
|||||||
|
3 m
deep
|
5.6
|
0.2
|
4.1
|
5.1
|
|||||||
|
Limits for carp
|
5.5-8
|
> 4
|
<1
|
0.6 to 1
|
<50
|
<0.2
|
<2
|
0.3
|
0.01 to 0.03
|
N / A
|
|
As could be noticed, most physical chemical parameters of the water in Fundulea – Frunzareni fishery (pH, O2, NH4+, NO3–, NO2–, Fe, Cu) values were appropriate for rearing of cyprinids.
Phosphates in water samples harvested from 3 m depth exceeded acceptable limits for carp 1.5 times in water admission place and four times in the draining place. Residual chlorine in water samples exceeded the limits permitted 6 -13 times, most likely because of the recent water decontamination with chloramine.
Depending on temperature and concentration of active chlorine, this may cause damage to fish skin, gill and nervous disorders, resulting in poor welfare.
Tab. 2: Results of serum biochemical analysis in carp in the Fundulea – Frunzareni fishery
|
Specific parameters
|
|
Values obtained
|
Reference values
|
|
Blood urea nitrogen
BUN (mg / dl)
|
1
|
5.32
|
1.9 to 3.6
|
|
2
|
3.36
|
||
|
3
|
3.92
|
||
|
Creatinine
CREA (mg / dl)
|
1
|
0.20
|
0.07 to 0.09
|
|
2
|
0
|
||
|
3
|
0.20
|
||
|
Phosphate
PHOS (mg / dl)
|
1
|
16.07
|
6.5 to 7.3
|
|
2
|
16.06
|
||
|
3
|
16.00
|
||
|
Calcium
CA (mg / dl)
|
1
|
9.52
|
8-9
|
|
2
|
16
|
||
|
3
|
16
|
||
|
Albumin
ALB (g / dl)
|
1
|
11
|
0.76 – 0.85
|
|
2
|
12
|
||
|
3
|
11
|
||
|
Cholesterol
Chol (mg / dl)
|
1
|
63.07
|
127-184
|
|
2
|
98.46
|
||
|
3
|
94.23
|
||
|
Glucose
GLU (g / dl)
|
1
|
25.27
|
30-47 (96)
|
|
2
|
30.00
|
||
|
3
|
38.36
|
||
|
Alanine transaminase
ALT (U / l)
|
1
|
10
|
8.9 to 9.6
|
|
2
|
11
|
||
|
3
|
10
|
||
|
Lactate dehydrogenase
LDH (mg / dl)
|
1
|
> 2800
|
860 – 1200
|
|
2
|
2188
|
||
|
3
|
2247
|
||
|
Triglycerides
TRIGA (mg / dl)
|
1
|
91.81
|
N / A
|
|
2
|
144.5
|
||
|
3
|
158.18
|
Samples collected from the Fundulea – Frunzareni fishery – have recorded increasing for BUN, creatinine, phosphate, lactate dehydrogenase, albumin and slightly increasing of LDH and alanine transaminase. The causes of these changes are likely dehydration and intense muscular effort associated with the method of capturing fish (angling), and blood collecting (fish emersion and maintaining in high air temperature).[1]
BUN increasing could indicate kidney or gill disorders (excretory).
Finally, welfare is poor and negative aspects are stressed by the fish angling and blood sampling processing.
► The fish ponds in Ulmu and Ciocăneşti farms have the bottom from soil and the embankments from compacted clay. Ciocăneşti farm water supply is from the channels draining the Boianu-Sticleanu marsh and Ulmu farm from the upstream springs, precipitation and the Mostiştea lake.
Ulmu and Ciocăneşti farms growing common carp for production and fishing.
From the farm ponds have been collected water samples at different depths, as follows:
– Near the shore at 30 cm depth;
– In the center from 1.5 m depth;
– In the center, from the surface of the water.
From the samples collected have been determined following physical chemical parameters: pH, chlorine, detergents, dissolved oxygen, nitrates, nitrites, phosphates, total iron, copper and ammonia. The determined values were compared with the limits permitted under the Order 161/2006 regarding the surface water quality and the recommendations issued by Schlotfeldt H.J for carp rearing.
Results interpretation (tables 3-4) revealed the following:
– PH of water in the two farms have values which framing within the admitted limits for carp rearing;
– Residual chlorine and dissolved oxygen were recorded values framing the water in first class of quality according to 161/2006 Order, favorable for carp rearing;
– Concentrations of nitrates and nitrites framing the water quality in Class II and are within the admitted limits set for carp rearing;
– Iron in water samples had values corresponding to quality class III for Ciocanesti farm and in class for Ulmu, the values framing within the admitted limits with regards to fish rearing – being lower than 2 mg / l;
– Phosphate concentrations had values corresponding to the Class V of water quality in the two farms according to 161/2006 Order, and in terms of standards set for carp exceeded the maximum permissible limits for 2 times for the water surface samples and 3 times for the samples harvested at 1.5 m depth, in both fisheries – Ulmu and Ciocăneşti.
Excess phosphates (similarly to nitrogen) can generate eutrophication, with negative consequences that decrease water transparency and dissolved oxygen concentration.
Tab. 3: Physical chemical parameters of water quality in Ulmu and Ciocanesti
fish farms:pH, chlorine, oxygen, nitrogen, copper
|
Farm
|
Sampling point
|
Assessed parameters
|
|||||
|
pH
|
Cl2
(mg / l)
|
O2
(mg / l)
|
NO3–
(mg / l)
|
Cu
(mg / l)
|
|||
|
Ciocanesti
|
Pond center;
1.5 m depth
|
6.8
|
0.15
|
6.0
|
3.0
|
–
|
|
|
Pond center,
surface
|
7.0
|
0.12
|
7.0
|
0
|
0.16
|
||
|
Shore
30 cm deep
|
6.7
|
0.09
|
6.9
|
undetectable
|
–
|
||
|
Ulmu
|
Pond center;
20 cm deep
|
7.3
|
0.18
|
9
|
undetectable
|
–
|
|
|
Pond center;
1.5 m depth
|
8.7
|
0
|
8.1
|
0.56
|
|||
|
Water supplying
30 cm deep
|
8.7
|
0.05
|
14.1
|
–
|
|||
|
Maximum admitted limits according to 161/2 006 Order: Water quality categories
|
I
|
6.5 – 9.5
|
N / A
max. 0.28 at
consumer
1342/91
|
7
|
1
|
background
|
|
|
II
|
6
|
3
|
2
|
||||
|
III
|
5
|
6
|
4
|
||||
|
IV
|
4
|
15
|
8
|
||||
|
V
|
<4
|
> 15
|
> 8
|
||||
|
Limitsfor carp rearing
|
5.5 – 8
|
0.01 – 0.03
|
min. 4 | max. 50 | max. 0.3 | ||
Tab. 4: Physical chemical water quality parameters in Ciocanesti and Ulmu
fish farms: detergents, P, Fe, nitrate, ammonium
|
Farm
|
Sampling point
|
Assessed parameters
|
|||||
|
Detergents
(mg / l)
|
P
(mg / l)
|
Fe
(mg /l)
|
NO2–
(mg/ l)
|
NH4+
(mg / l)
|
|||
|
Ciocanesti
|
Pondcenter;
1.5 m depth
|
–
|
1.8
|
0.2
|
0015
|
–
|
|
|
Pond center,
surface
|
0.08
|
1.1
|
0.8
|
–
|
undetected
|
||
|
Shore
30 cm deep
|
–
|
1.2
|
–
|
–
|
–
|
||
|
Ulmu
|
Pond center;
20 cm deep
|
–
|
7.7
|
0.9
|
0079
|
–
|
|
|
Pond center
1.5 m depth
|
0.06
|
3.2
|
1.4
|
–
|
undetected
|
||
|
Water supplying
30 cm deep
|
–
|
2.2
|
–
|
–
|
–
|
||
|
Maximum admitted limits
according to 161/2006 Order: water quality categories
|
I
|
1
|
0.05
|
background
|
0.01
|
0.2
|
|
|
II
|
1.5
|
0.1
|
0.1
|
0.06
|
0.3
|
||
|
III
|
2
|
0.2
|
0.3
|
0.12
|
0.6
|
||
|
IV
|
–
|
0.5
|
1
|
0.3
|
1.5
|
||
|
V
|
–
|
> 0.5
|
> 1
|
> 0.3
|
> 1.5
|
||
|
Limits for carp rearing
|
N / A
|
0.6 – 1
|
max. 2 | max 0.2 | max. 1 | ||
From the fishes in the Ciocăneşti and Ulmu ponds were taken blood samples too, following the same protocol above.
There were analyzed two blood samples from each farm, joining samples resulted from 10 carps.
Samples taken from Ciocăneşti showed elevated levels of creatinine, aspartate aminotransferase, phosphate, lactate dehydrogenase, alkaline phosphatase, and slightly elevated levels of calcium and albumin due to dehydration and intense muscular effort combined with the methods of catching and fish restraining. In samples taken from Ulmu farm ponds was observed, in addition to issues from Ciocanesti, a significant increase in cholesterol, transaminases and glucose, which might suggest hepatopancreatice lesions (Table 5a and b).
Following research it was found that the fish welfare in Ciocăneşti is acceptable, while the actual welfare of fish in the Ulmu farm is poor.
Tab. 5 – Results of serum biochemical analysis in
carps from Ciocăneşti (A) and Ulmu (B) farms
A
|
Serum parameters analyzed
|
Blood sample
|
Results
|
Reference Values
|
|
Urea nitrogen
BUN (mg / dl)
|
Ciocanesti 1
|
3
|
1.9 to 3.6
|
|
Ciocanesti 2
|
2
|
||
|
Creatinine
CREA (mg / dl)
|
Ciocanesti 1
|
0.2
|
0.07 to 0.09
|
|
Ciocanesti 2
|
0.4
|
||
|
Phosphate
PHOS (mg / dl)
|
Ciocanesti 1
|
> 16.1
|
6.5 to 7.3
|
|
Ciocanesti 2
|
> 16.1
|
||
|
Calcium
CA (mg / dl)
|
Ciocanesti 1
|
10.8
|
8-9
|
|
Ciocanesti 2
|
10.7
|
||
|
Albumin
ALB (g / dl)
|
Ciocanesti 1
|
1.2
|
0.76 – 0.85
|
|
Ciocanesti 2
|
1.2
|
||
|
Cholesterol
CHOL (mg / dl)
|
Ciocanesti 1
|
157
|
127-184
|
|
Ciocanesti 2
|
163
|
||
|
Glucose
GLU (g / dl)
|
Ciocanesti 1
|
36
|
30-47 (96)
|
|
Ciocanesti 2
|
26
|
||
|
Total protein
TP (g / dl)
|
Ciocanesti 1
|
3
|
2.5 to 3.5 g / dl
|
|
Ciocanesti 2
|
–
|
||
|
N blood
NH3 (mg / dl)
|
Ciocanesti 1
|
890
|
1070 – 1250
|
|
Ciocanesti 2
|
–
|
||
|
Alanine transaminase
ALT (U / l)
|
Ciocanesti 1
|
<10
|
8.9 to 9.6
|
|
Ciocanesti 2
|
<10
|
||
|
Aspartate aminotransferase
AST (U / l)
|
Ciocanesti 1
|
331
|
121-124
|
|
Ciocanesti 2
|
–
|
||
|
Lactate dehydrogenase
LDH (mg / dl)
|
Ciocanesti 1
|
> 2800
|
860 – 1200
|
|
Ciocanesti 2
|
> 2800
|
||
|
Alkaline phosphatase
ALP (U / l)
|
Ciocanesti 1
|
58
|
14-24
|
|
Ciocanesti 2
|
–
|
||
|
Total bilirubin
TBIL (mg / dl)
|
Ciocanesti 1
|
<0.2
|
0.2 – 2
|
|
Ciocanesti 2
|
–
|
||
|
Creatine kinase
CK (U / l)
|
Ciocanesti 1
|
> 2036
|
1150 – 9700
|
|
Ciocanesti 2
|
> 2036
|
||
|
Triglycerides
TRIGA (mg / dl)
|
Ciocanesti 1
|
172
|
N / A
|
|
Ciocanesti 2
|
173
|
||
|
Amylase
Amylase (U / l)
|
Ciocanesti 1
|
83
|
N / A
|
|
Ciocanesti 2
|
–
|
B
|
Serum parameters analyzed
|
Blood sample
|
Results
|
Reference Values
|
|
Urea nitrogen
BUN (mg / dl)
|
Ulmu 1
|
3
|
1.9 to 3.6
|
|
Ulmu 2
|
4
|
||
|
Creatinine
CREA (mg / dl)
|
Ulmu 1
|
0.3
|
0.07 to 0.09
|
|
Ulmu 2
|
0.6
|
||
|
Phosphate
PHOS (mg / dl)
|
Ulmu 1
|
> 16.1
|
6.5 to 7.3
|
|
Ulmu 2
|
> 16.1
|
||
|
Calcium
CA (mg / dl)
|
Ulmu 1
|
13.3
|
8-9
|
|
Ulmu 2
|
13.8
|
||
|
Albumin
ALB (g / dl)
|
Ulmu 1
|
1.5
|
0.76 – 0.85
|
|
Ulmu 2
|
1.7
|
||
|
Cholesterol
CHOL (mg / dl)
|
Ulmu 1
|
228
|
127-184
|
|
Ulmu 2
|
223
|
||
|
Glucose
GLU (g / dl)
|
Ulmu 1
|
121
|
30-47 (96)
|
|
Ulmu 2
|
133
|
||
|
Total protein
TP (g / dl)
|
Ulmu 1
|
4.0
|
2.5 to 3.5 g / dl
|
|
Ulmu 2
|
4.5
|
||
|
N blood
NH3 (mg / dl)
|
Ulmu 1
|
> 950
|
1070 – 1250
|
|
Ulmu 2
|
> 950
|
||
|
Alanine transaminase
ALT (U / l)
|
Ulmu 1
|
18
|
8.9 to 9.6
|
|
Ulmu 2
|
30
|
||
|
Aspartate aminotransferase
AST (U / l)
|
Ulmu 1
|
263
|
121-124
|
|
Ulmu 2
|
758
|
||
|
Lactate dehydrogenase
LDH (mg / dl)
|
Ulmu 1
|
> 2800
|
860 – 1200
|
|
Ulmu 2
|
> 2800
|
||
|
Alkaline phosphatase
ALP (U / l)
|
Ulmu 1
|
28
|
14-24
|
|
Ulmu 2
|
32
|
||
|
Total bilirubin
TBIL (mg / dl)
|
Ulmu 1
|
0.1
|
0.2 – 2
|
|
Ulmu 2
|
0.5
|
||
|
Creatine kinase
CK (U / l)
|
Ulmu 1
|
> 2036
|
1150 – 9700
|
|
Ulmu 2
|
> 2036
|
||
|
Triglycerides
TRIGA (mg / dl)
|
Ulmu 1
|
158
|
N / A
|
|
Ulmu 2
|
164
|
||
|
Amylase
Amylase (U / l)
|
Ulmu 1
|
109
|
N / A
|
|
Ulmu 2
|
121
|
► There have been harvested water samples from Bicaz Lake, inside the trout farm, from depths of 10 m, 20 m and 35 m, from adults and juvenile floating cages. From the samples have been determined following physical chemical parameters: dissolved oxygen, chlorine, pH, phosphates, nitrates, nitrites, ammonia, phenols, copper, sulfates and detergents (Table 6).
Tab 6 – Mean physical chemical parameters values of the water in Potoci – Bicaz farm
|
Sampling point
|
Assessed element
|
||||||||||
|
O2
mg / l
|
Cl2
mg / l
|
pH
|
P
mg / l
|
NO2–
mg / l
|
NH4+
mg / l
|
NO3–mg / l
|
Phenols
ng / l
|
Cu
mg / l
|
SO42 –
mg / l
|
Deterg.
mg / l
|
|
|
Bicaz Lake – 10 m depth
|
9.5
|
0.03
|
7.7
|
0.1
|
0.014
|
Ned.
|
2.0
|
0.04
|
0.01
|
23
|
0.01
|
|
Bicaz Lake – 20 m depth
|
13.5
|
0.07
|
7.1
|
0.2
|
0.006
|
Ned.
|
1.0
|
0.14
|
0.02
|
31
|
0
|
|
Bicaz Lake – 35 m depth
|
11.5
|
0.03
|
7.3
|
0.2
|
0.004
|
Ned.
|
1
|
0.04
|
0.01
|
23
|
0
|
|
Adult trout floating cage
|
9.3
|
0.02
|
8.0
|
0.3
|
0.015
|
Ned.
|
0
|
0.01
|
0.02
|
22
|
0.02
|
|
Juvenile trout floating cage
|
9.1
|
0.02
|
7.9
|
0.2
|
0.016
|
Ned.
|
Ned.
|
0.04
|
0.015
|
24
|
0.02
|
| Admitted limits for trout |
6 mg
(Min)
|
0.01 to 0.03
|
6 to 7.2
|
0.2
|
<0.2
|
<1
|
<20
|
6-7
|
0.1
|
2-7
|
–
|
Analyzing the data from the table could be noticed the following:
– dissolved oxygen concentrations is appropriate, the minimum allowed for trout being 6 mg/l;
– chlorine, for most samples, recorded values close to the maximum permissible values and even higher than the limits for samples harvested from 20 m depth (for 2.33 times);
– water pH framing in within the acceptable limits for trout samples harvested from 10, 20, 35 m depths and reveals alkaline values to those harvested directly from the adults and juveniles cages.
Neutral-pH waters show a low buffering power and tendency to acidification.
– Phosphates have registered values close to permissible limit values (0.2 mg / l) in all samples except those taken from adult trout floating cage, where the limit was exceeded by 1.5 times;
– Nitrites and nitrates in all samples are within acceptable limits for trout of 0.2 mg / l and 20 mg / l water;
– Copper had values below 0.1 mg /L: the limit for trout;
– sulphates from all samples showed levels exceeding the limits (2-7 mg / l) by 3.2 – 4.4 times;
– Detergents recorded values within acceptable limits, and phenols exceeded permissible limits in all samples.
There were also conducted microbiological analysis: APC (aerobic plate count), FPC (fungal plate count) ant total coliforms (Table 7).
Analyzing the results, there is noticed a big microbial loadup, the number of germs, fungi and total coliforms being high. In water, microorganisms find favorable conditions for life. They can have as origin indigenous biocenosis, being included in specific biocenosis (planktonic, neustoni, benthic, epibiotic) and have alohtonic origin (those who accidentally reach the water from the soil, wind disperssed, from the rain water, those derived from human, animal or industrial activities) and ubiquitous (microorganisms that can normally live in water, soil and air).
Among indigenous microorganisms, can be mentioned: bacteria, fungi, protozoa and microalgae.
In the water is present all physiological types of bacteria (heterotrophic, autotrophic, chimioautotrofe etc.).. Microfungi and yeasts from the water may come from the soil or from the ill animals (the pathogen ones).
Between fish and microorganisms have been established rcomensalism relationships. From salmonids intestine, have been isolated over 200 species of bacteria. Maximum density of microorganisms in water is 1012 cells / ml.
Survival time of bacteria in water is inversely proportional with the organic and microbiological load of water.
Tab. 7 – Results of microbiological examination of water: Potoci – Bicaz trout farm
|
Sampling site
|
Assessed element
|
||
|
APC/ ml
|
FPC/ ml
|
Total coliform
|
|
|
Bicaz Lake – 10 m depth
|
660
|
17
|
128
|
|
Bicaz Lake – 20 m depth
|
594
|
45
|
136
|
|
Bicaz Lake – 35 m depth
|
550
|
18
|
260
|
|
Juvenile floating cage 85
|
900
|
1
|
687
|
|
Juvenile floating cage 104
|
260
|
1
|
714
|
|
Adults floating cage 214
|
374
|
16
|
520
|
|
Adults floating cage 221
|
300
|
4
|
360
|
II) establishing a portfolio of specific rules for fish welfare during transport, the welfare assessment in an experiment involving two transport systems – by automobiles, in transport tanks and by water, in transport floating cages
► live fish transportation and particulary native trout (present research followed the transport of this species) is made for the following reasons: restocking, juveniles saving when water levels drop due to drought or in winter, for human consumption of fish.
Regarding transportation, following issues interest: water temperature, fish density, transport vehicle speed, chemical and microbiological parameters of water. Water temperature should be maintained during the entire transport period at 10 ° C (range in 8-12 ° C interval). For this, in the transport containers are being added chunks of ice and there is ensured a permanent control by thermometry.
Concerning density, the rules in tab. 8 must be applied.
Tab.8 – Transport capacities of different containers (Bud, I, 2004)
|
Transportation
|
° C
|
Transport time
h
|
Juveniles
4-5 wk.
No.
|
Juveniles
5 – 6 months
No.
|
Adult trout
Kg
|
|
10 l recipients
|
10-12
|
1
|
1500 – 2000
|
150-300
|
0.7
|
|
100 l recipients
|
10-12
|
1
|
2000-16000
|
2000 – 3000
|
7.0
|
|
200 l tanks
|
10-12
|
1
|
50000-100000
|
1500-2500
|
6.0
|
|
50 l plastic bags
|
10-12
|
5-10
|
2000 – 3000
|
400-600
|
1.5
|
Immediately after the arrival at the destination point, must be provide a resting period of 30-40 minutes for calming the fish. In order to avoid thermal shock, the are gradually mixed water from the transport container with water from the launching site.
If fishnets are used, they must be integer, clean, decontaminated and iron frames need to be covered with cloth.
Admitted losses (DOA – Death on Arrivals) during transport shall not exceed the values in Table 9.
Tab.9 – The maximum losses during transportation (Bud, I, 2004)
|
Age
|
Transported species
|
||
|
Native trout
%
|
Rainbow trout
%
|
Other salmonid species
%
|
|
|
Eggs
|
2.5
|
2.0
|
3.0
|
|
Juveniles 1 year
|
5.5
|
5.0
|
6.0 to 7.0
|
|
Trout 2 years
|
3.5-4
|
2.5 to 3.0
|
4.0 to 5.0
|
|
Trout 3 years
|
2.0 to 3.0
|
1.5 to 2.0
|
3.0 to 4.0
|
|
Breeding 3 – 6 years
|
1.0 to 1.5
|
1.0
|
1.0 to 2.0
|
► There has been conducted a comparative study concerning two modes of transport – automobile transport in tanks and transport in floating cage (on water, between two salmonids farm on Bicaz Lake) – see Fig.
The first study follows the particular situation of a trout transport for repopulation (90 g average weight of individuals), from Brasov to Potoci – Bicaz farm in containers with a total volume of 6 m3 (2 tanks of 3 m3 each, the applied density being 250 Kg / tank). Each tank has a hole in which enters the aeration tube in connection with oxygen cylinder and manometer. Transportation period was of 18 hours.
On destination, unloading is done in fish nets, in which fish are transferred to a pool fixed on a mobile console (on rails), with this are brought from the point of unloading to the lake, in a fish transport floating cage (hight difference level of 10 to 12 m).
As a positive aspect of management at the destination place, are to be mentioned the use of clean and decontaminated fishnets, in which the rigid iron frame was covered with cloth to avoid injuries.
As disavantages, are to be mentioned a too shor resting period after transport (10-15 minutes) and the lack of water homogenization of destination place (Bicaz lake) with the water from the transport containers in order to habituate fish to local temperature and chemistry.
Also, the water draining from the transport tanks and subsequently transfer of the fish generated a critical situation: fish left in the bottom of transport tanks after water discharge came into contact with air for a long period: 5-10 minutes.
Another negative aspect was the high transport density: 250 kg fish / tanks of 3 m3 (the allowed limit being 90 kg / m3, respectively 6 kg/200 l – tab. 8). Overcrowding generated behavioral changes, obvious before fish unloading and during the unloading (fast swimming, restlessness, frequent jumps outside the water).
To determine the physical chemical and microbiological parameters of water during transport, there have been collected water samples from each tanks. In the samples there have been determined following parameters: dissolved oxygen, residual chlorine, pH, phosphates, nitrites, nitrates, ammonia, phenols, copper, sulphates and detergents (Table 10).
Tab. 10 – Water quality during road transport in tanks: chemical parameters
|
Sampling point
|
Assessed element
|
||||||||||
|
O2
mg / l
|
Cl2
mg / l
|
pH
|
P
mg / l
|
NO2–
mg / l
|
NH4+
mg / l
|
NO3–mg / l
|
Phenols
ng / l
|
Cu
mg / l
|
SO42 –
mg / l
|
Deterg.
mg / l
|
|
|
Tank 1
|
14.9
|
0.37
|
7.0
|
4.4
|
0.027
|
13.2
|
0
|
0.55
|
0.23
|
35
|
0.02
|
|
Tank 2
|
15.3
|
0.42
|
7.1
|
4.55
|
0.030
|
11.5
|
2
|
1.11
|
0.20
|
35
|
0.02
|
| Admitted limits for trout |
6
(Min)
|
0.01 to 0.03
|
6 to 7.2
|
0.2
|
<0.2
|
<1
|
<20
|
6-7
|
0.1
|
2-7
|
–
|
Analyzing the data in Table 10 could be noticed that: dissolved oxygen recorded relatively high values, respectively from 14.9 to 15.3 mg / l (minumum admitted limit for trout being 6 mg/l of water). Excess of dissolved oxygen in water is often adverse, increasing the metabolic rate, leading to paralysis in extreme cases. Residual chlorine exceeded the permissible limit for trout for 12 times in T1 (Tank 1) and 14 times in Tank 2 (T2). Excessive chlorine in water may cause fish agitation, postural changes, spasmodic movements of the mouth and caudal fins, bronchial congestion, paleness of fish body surface and suffocation. For phosphates too have been exceeded permissible limit for trout by 22 times. Nitrogen is found in ponds as salts: nitrates and ammonium salts, caming from the decomposition of dead organic substance in water due to nitrogen fixation by the nitrifying bacteria. Nitrites and nitrates have values much below the permissible limit in all water samples. Ammonium ion values exceeded 12 times the admitted limit. Through residual chlorine, nitrite ions penetrate the body of fish at gills level. In blood, nitrites are bounded to hemoglobin, forming methemoglobin, with the consequence of the oxygen transport capacity reduction, gills and blood turning into brown.
Copper exceeded the permissible limit (0.1 mg / l) in all samples by 2.3 times. The role of this metal is related with the formation of hemoglobine and some enzymes. As salts (copper sulphate) is used to stop the eutrophication phenomenon. Copper increasing in harvested samples can be explained by its existence in the transport water (in Brasov fishery being corrected water parameters for algal control).
Detergents from water are within the acceptable limits.
Microbiological water tests results are presented in Table. 11.
Tab. 11 – Water quality during road transport in tanks: microbiological parameters
|
Sampling site
|
Assessed element
|
||
|
APC / ml
|
FPC / ml
|
Total coliforms
|
|
|
Tank 1
|
0
|
7
|
0
|
|
Tank 2
|
0
|
2
|
0
|
| Admitted limits for trout rearing |
*
|
–
|
–
|
Analyzing the data from the table, it appears that no germs or total coliforms were developed at 37 ° C. Regarding the FPC, there was a higher number of fungi in tank no. 1. Finally, we analyzed mortality in the two trout tanks and we found a 18-20% rate in T1 and 25% in T2.
The causes of this high mortality were the transportation period (18 hours), overcrowding and high residual chlorines concentration (which is explained by insufficient rinse of tanks after decontamination with chloramine).
The second mode of transportation was clearly superior in terms of ensuring the needs of fish (ecological-etological-and physiological ones). The transfer was done in floating cage on water with very low speed, air contact being minimized. Density was significantly better, fits to the standards (500 kg fish from 90-100 m3 of water within the nets marking the limits of floating cages).
Regarding the behavior of fish, this was was normal (swimming in exploratory purpose, the lack of abnormal patterns: restlessness, frequent swimming in circles or jumping outside the water, crowded on the water surface etc.)..
Mortality recorded for transport was up to 4%.
The physical – chemical and microbiological water quality was similar in transport tanks and in floating cages from the Bicaz lake (see tab. 6-7).
The presented issues suggest that the welfare level in transport tanks is very poor and in floating cages are good. This was proved by blood tests conducted on serum samples in 1:10 dilution, values showing a pronounced alteration of fish blood chemistry panel in transport tanks in comparison with those sampled in floating cages.
Tab. 12 – Biochemical parameters from blood samples collected from fish in tanks and floating cages (transportation)
|
Parameters determined
|
Vehicle transport tank 1
|
Vehicle transport tank 2
|
Floating cages transport between farms
|
Reference values from literature *
|
|
BUN (mg / dl)
|
23.0
|
19.0
|
7.0
|
30
|
|
PHOS (mg / dl)
|
27.8
|
32.9
|
16.1
|
14
|
|
Ca (mg / dl)
|
3.5
|
7.2
|
10.6
|
16-29
|
|
Mg (mg / dl)
|
4.08
|
4.89
|
3.23
|
N / A
|
|
TP (g / dL)
|
0.01
|
0.01
|
0.6
|
0.3 – 3.08
|
|
Alb (g / dl)
|
0.0
|
0.0
|
–
|
N / A
|
|
ALT (U / l)
|
261
|
100
|
54
|
23 – 43.20
|
|
AST (U / l)
|
1698
|
514
|
136.4
|
29 – 52.45
|
|
Chol (mg / dl)
|
–
|
194
|
126
|
125-225
|
|
NH3 (μmol / l)
|
3500
|
3206
|
4497
|
N / A
|
|
Trig (mg / dl)
|
–
|
225
|
44
|
121-206
|
|
Glu (mg / dl)
|
161
|
164
|
23
|
20-70
|
|
LDH (U / l)
|
12 390
|
13 295
|
280
|
26-60
|
|
CREA (mg / dl)
|
0.2
|
0.9
|
0.4
|
N / A
|
|
Uric (mg / dl)
|
0.288
|
0.420
|
0.220
|
N / A
|
|
ALKP (U / l)
|
137
|
140
|
110
|
46.1 – 60.5
|
|
TBIL (mg / dl)
|
0.290
|
0.530
|
1
|
N / A
|
|
Amylase (U / l)
|
779
|
1032
|
529
|
N / A
|
|
LIPA (U / l)
|
201
|
–
|
151
|
N / A
|
|
CK (U / l)
|
11 669
|
617
|
93
|
48.8 – 50.6
|
[1] There are scientific studies that recommended blood sampling in fish while they are immersed, using a intraaortic cannula (biochemical parameters being much closer to real ones in this situation – TA Heming, 1989). Sampling methods will also be considered in further research, although rises special problems.