Aquacultura Indonesiana (2015) 16 (1) : 29-32

ISSN 2477-6939

Detection of Antibiotic Residue from Shrimp Ponds and Their Environment in

East Java Province, Indonesia
Hari Suprapto, L. Sulmartiwi, and Sudarno

fisheries faculty, Airlangga University, Surabaya, Indonesia

Correspondence to: [email protected]

Abstract

Hari Suprapto, L. Sulmartiwi, and Sudarno. 2015. Detection of Antibiotic Residue from Shrimp
Ponds and Their Environment in East Java Province, Indonesia. Aquacultura Indonesiana, 16 (1): 29-32.
Indonesia shrimp products are often rejected by foreign countries due to high antibiotic content, whereas a few
years ago Japan and European countries are set 0.05 ppb permitted to consume by the people, but now the
restriction will become smaller, which is 0.1 ppb. Some farmer have topped using antibiotics but the residues were
still found in fish flesh. Possible origin of the antibiotic is from environment because residues are still exist in the
mud, water organism which lives in the pond. Samples originating from shrimp, mud, water and other biota were
examined. The purpose of this study was to determine the origin of antibiotic present in the cultivation or farming
system that does not use antibiotics in shrimp production.: a. Antibiotics obtained from ponds in the Tuban area is
1.8 mg/L (mud), 0.8 mg/L (water) and 2.1 (biota). b. Antibiotics obtained from ponds in Gresik were 0.6 mg/L
(mud), 0.3 mg/L (water) and 1.2 (biota). c. Antibiotics obtained from ponds in the area of Situbondo are 0.3 mg/L
(mud), 0.1 mg/L (water) and 0.6 (biota). d. a. Antibiotics obtained from ponds in the area are 0.3 mg/L
Probolinggo (mud), 0.2 mg/L (water) and 0.6 mg/L (biota).

Keywords : Antibiotic; Environment; Pond; Shrimp

Introduction tetracycline, sulfa drugs, trimethoprim, and

oxytetracycline Tribrissen (Paone, 2000).
Rejection of Indonesian shrimp products Spanggaard et al. (1993) reported that many bacteria
by some European countries and Japan occurred in fresh water resistant to antibiotics in Denmark.
few years ago due to high antibiotic content. Nygaard et al. (1992) also reported the use of
Previously the residue limit was 0.05 ppb but oxytetracycline and oxolinic acid causing bacteria
today 0.001 ppb. The use of vaccine to prevent the resistant to antibiotics. Furthermore Leano et al.
disease is recommended rather than the use of (1999) reported that Vibrio spp. and Aeromonas spp.
antibiotics. Antibiotics should be stopped because isolated from fish and shrimp were resistant to
it could cause some excesses of bad health and the streptomycin but sensitive to oxolinic acid. Now a
lot of bacteria are resistan to more than one
environment. Initially antibiotic is used to prevent
antibiotic. Vibrio harveyi and Vibrio are the most
diseases caused by bacteria, especially for the
frequent isolated (Peterson et al., 2002; Tendencia
luminous vibriosis in large scale of farming,
and de la Pena, 2001). The objective of this
oxytetracycline, oxolinic acid, chloramphenicol
experiment is to find the sources of antibiotics
and furazolidone are used through the feed. In the
contaminant in the farm, because according to
Philippines they used nitrofurans, erythromycin
farmers they had stopped the antibiotics
and sulfa drugs in shrimp culture (Baticados et al.,
application, but the residue is still found in fish
1990; Cobello, 2006). On salmon farming in
carcasses.
British Columbia, Canada in 2000, antibiotics
residues in salmon flesh are 0.1 mg/L (Paone, Materials and Methods
2000). Aso reported that crab consumed food
remains and salmon faeces contained residue of Sampling of water, mud and other biota
Further reported that crab eating food remains and
feces salmon containing residues Oxytetracycline of Water, mud and organism from suspected
0.8 and 3.8 mg/L. Bjorklund et al. (1990) reported pond contained antibiotic residues were collected.
that in Finland, although he did not perform A total of 100 mL of water was taken and then
measurements, antibiotic residues in wild fish and poured into a dark bottle, sealed and stored at
sediments were detected from the intestine of fish room temperature, 100 g of mud was taken and
and also a lot of isolated bacteria resistant to put into wide-mouthed dark bottles, while other
antibiotics. Aeromonas salmonicida causes organisms used as the samples were stored at are
furunculosis is also resistant to Romet-30,

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 Aquacultura Indonesiana Vol. 16 No.1, June 2015 : 29-32

used as the sample will be stored in the -80C to added to the total homogenate to obtain the
avoid spoilage. Prior to the analysis, the biota is number of Chloramphenicol entirely. Amount is
tawed at room temperature. then divided by the total weight of shrimp meat to
get g Chloramphenicol in shrimp meat.
Antibiotics (Chloramphenicol)
Results and Discussion
Chloramphenicol was analyzed according
to Thomson (1966) and Horwitz (1980). Briefly, Organic matter and excessive shrimp feed
cuticle and the head taken from 3 pieces of shrimp will dissolve in water and will accumulate in pond
of each sample then weighted, crushed and mixed bottom. Excessive organic material will cause
with 4 volumes of 0.1 M KH2PO4 buffer, pH 4.5 problems in the ecosystem of brackishwater pond
sterile. Half of the buffer is poured to the shrimp ecosystem because the excessive of organic
in the mixer, blended for 10 sec in a low speed matter is the habitat of several pathogens bacteria
and 10 sec in high speed. The other half is and will spend the oxygen. Excessive feeding may
blended for 10 sec of low speed to avoid foaming. cause the elevation of NH3-N and NO2 in the
The mixture was then centrifuged in 50 mL tubes pond. Some plants and animals can absorb
for 20 min at 10,000 rpm at 4ºC, removed the ammonia in the water; they can serve as a
debris material. Supernatant was transferred into a biological filter. Most farmers used biofilter such
clean, sterile tube and its volume is measured. as mussels, milkfish, tilapia or mullet with density
Supernatant will be devided in two parts, not of 3 pieces of fish/m2 and. Vibrio bacteria will
diluted and diluted with ratio of 1: 3 with 0.1 M thrive in water containing a high organic matter
KH2PO4 buffer, pH 4.5. Blended and then washed (Austin and Austin, 1988) as well as Aeromonas.
with a half volume of 0.1 M KH 2PO4 buffer, pH Organic material causing plankton blooming and
4.5 for 10 sec in low-speed, stored separately. interfere growth of fish and shrimp. Water
Standard curve prepared by using shrimp containing high organic matter needs to be
homogenate without using Chloramphenicol, but reduced by draining the ponds. This pond
using 0.1 N to final concentration of functions as a reduction of organic matter from
oxytetracycline (OTC) by 0.1, 0.2, 0.4, 0.8 and water conservancy shrimp because of abundant
1.6 mg/mL OTC. A total of 0.4 mg/mL OTC is organic material to be used by plankton to grow
used as a control. Each of a total 18 wells for each and as the main meal of tilapia. Ammonia and
standard curve and the 18 wells for reference. some other gases can partially evaporate in the
Zone size was measured and then compared with aerated pond. In addition, to the suspended matter
reference (control). A reference obtained from an will be deposited in these ponds. Biological filter
average of 90 wells. Rectification zone size is not able to remove the solids present in the
obtained from the increase or decrease the water, but the point is to change harmful
difference between the average sizes for the substances such as ammonia into harmless
reference zone which same concentration and substances such as nitrate. For this purpose few
average size of the grand reference zone. bacteria such as Nitrosomonas sp. can be added to
Suspension of spores of Bacillus cereus (ATCC help the nitrification process in the water. The
No. 11 778) prepared in antibiotic medium A main biological filter is an autotrophic bacterium,
(Nifco, Detroit Michigan) on agar plate. Aliquots although algae, yeast and protozoa and other
(200 mL) was used to test the cylinder is placed in small animals sometimes help turn waste from the
stainless steel, 6 cylinder for each plate, and 3 body of water. Soil has an important role in
plates for each test. Nine wells of each shrimp shrimp aquaculture, and almost all type of land is
homogenates prepared, as well as references there suitable for shrimp farming. Soil contains high
are 9 wells (0.4 g Chloramphenicol/mL). Zone of organic matter is not suitable for aquaculture due
inhibition resulting from dilution of shrimp to the decomposition of organic matter requires a
homogenates were then compared with the lot of oxygen. The results of the analysis use
references above, the results were then calculated chromatography presented in the Table 1.
by dilution of the homogenate of shrimp that Table 1. showed that in Gresik ponds,
references a result close to the concentration. mud, water and crabs as ponds organisme
Total Chloramphenicol in shrimp (g) is calculated contains of antibiotics chloramphenicol. The crabs
by multiplying the concentration of the tested contains antibiotic much higher than in the mud
suspension with total volume (mL) of supernatant. and water. Initially antibiotic was used to prevent
Chloramphenicol in the washing water (g) is diseases caused by bacteria, such as luminous

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 Detection of Antibiotic Residue from Shrimp Ponds and Their Environment in East Java Province, Indonesia (H. Suprapto et al.)

Vibriosis shrimp culture. Antibiotics were October.

administered via the feed (medicated feed). In other The antibiotic analytical results from
countries such as the Philippines, antibiotics used in samples from the area Tuban (Table 2.) showed
shrimp farming are nitrofurans, erythromycin, and that same as Gresik, mud, water and crabs may
sulfa drugs (Baticados et al., 1990; Primavera et al., contains the chloramphenicol. Furthermore Leano
1993) and several other drugs. Uncontrolled use of et al. (1999) reported that Vibrio spp. and
antibiotics will left the residue in fish and the Aeromonas spp. isolated from fish and shrimp
environment. For example, in fish farming in British were resistant to streptomycin but sensitive to
Columbia, Canada in 2000, residues of antibiotics in acid oxolinic. Now a lot of bacteria were resistant
salmon flesh is 0.1 mg/L (Paone, 2000). This to more than one antibiotic. Vibrio harveyi and
residue also found in several places in East Java Vibrio spp. and other species are the most
with range from 1-2 mg/L. In dry season antibiotic frequent isolated (Tendencia and de la Pena,
concentration in some place is high. 2001). The use of drug for cultivation in the USA
Crab eats feed remaining and fish faeces should get permission from the FDA's Center for
may contains residues of oxytetracycline 0.8 and Veterinary Medicine. Drug generally used mixed
3.8 mg/L. Bjorklund et al. (1990) reported that in in feed, and it supposed 75% the fish can use it
Finland, although he did not perform and not mostly discharged into the environment.
measurements of antibiotic residues in wild fish, So the case like in ornamental fish not happened.
from sediments and intestine of fish but a lot of Ornamental fish are closer to humans will
resistant antibiotics bacteria can be isolated. transmit the bacteria that were resistant to more
Aeromonas salmonicida causes furunculosis is human. Indiscriminate use of antibiotics in
also resistant to Romet-30, tetracycline, sulfa Equador, which will cause outbreak of Vibrio
drugs, trimethoprim, and oxytetracycline cholerae in the USA. Which are resistant bacteria
Tribrissen (Paone, 2000). Spanggaard et al. can be transmitted from fish to human. Table 3.
(1993) reported that many bacteria in fresh water Antibiotic analytical results from samples from
resistant to antibiotics in Denmark. Nygaard et al. the area Pasuruan June-October. Results of
(1992) also reported the use of oxytetracycline antibiotic analysis from Probolinggo (June-
and oxolinic acid causes the bacteria resistant to October) see Table 4.
other antibiotics. Table 2. Antibiotic analytical
results from samples from the area Tuban June-

Table 1. Results of antibiotic analysis from Gresik (June-October, 2011)

Sample Sample quantity Antibiotic concentration Antibiotic in feed Antibiotic
Mud 100 g 0.6 mg/L 1000 mg/kg Chloramphenicol
Water 100 mL 0.3 mg/L 1000 mg/ kg Chloramphenicol
Pond organism/crabs 2 1.2 mg/L 1000 mg/kg Chloramphenicol

Table 2. Results of antibiotic analysis from Tuban (June-October, 2011)

Sample Sample quantity Antibiotic concentration Antibiotic in feed Antibiotic
Mud 100 g 1.8 mg/L 1000 mg/kg Chloramphenicol
Water 100 mL 0.8 mg/L 1000 mg/ kg Chloramphenicol
Pond 2 2.1 mg/L 1000 mg/kg Chloramphenicol
organism/crabs

Table 3. Results of antibiotic analysis from Pasuruan (June-October, 2011)

Sample Sample quantity Antibiotic concentration Antibiotic in feed Antibiotic
Mud 100 g 0.3 ppb 1000 mg/kg Chloramphenicol
Water 100 mL 0.1 ppb 1000 mg/ kg Chloramphenicol
Pond organism/crabs 2 0.6 ppb 1000 mg/kg Chloramphenicol

Table 4. Results of antibiotic analysis from Probolinggo (June-October, 2011)

Sampel Sample quantity Antibiotic concentration Antibiotic in feed Antibiotic
Mud 100 g 0.3 ppb 1000 mg/kg Chloramphenicol
Water 100 mL 0.2 ppb 1000 mg/kg Chloramphenicol
Pond 2 0.6 ppb 1000 mg/kg Chloramphenicol
organism/crabs

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 Aquacultura Indonesiana Vol. 16 No.1, June 2015 : 29-32

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