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Table 2: Concentrations of Lead, Zinc, Chrome and Cadmium in Muscles of Fish (ug / kg)

Species of Fish





































95 ±2

















Heavy metals in water:

The results showed that Zn was the highest concentrated heavy metal in sampled water; the mean concentration of Zn in the water samples was significantly higher than that of Pb,Cd and Cr levels (p< 0.05). Pb contents in water sampled from all station were 122±8 ppb and 82±4

ppb. The concentration of Zn was highest in Station 1 and the lowest was in Station 5. Also, the highest concentrations of Cr ranged from 82±4to 28±2 ppb (Table 3). The order of heavy metals concentration in water samples in all Station was Zn >Pb> Cr > Cd.

Table 3: Concentrations of Heavy Metals in waters of Gorgan Bay and Caspian Sea (ppb)

No. of


























































































Heavy metals in sediments:

The results showed that Zn was the highest concentrated heavy metalin sediments; the mean concentration of Zn in the sediments was significantly higher than that of Pb, Cd and Cr levels (p < 0.05). Zn contents in sediments of station 1and station 8 were 1213±8ppb and 598±4ppb. The highest concentrations of Pb ranged from 1021±9 to324±3 ppb (Table 4).The order of the heavy metals in sediments in all regions was Zn >Pb> Cr > Cd.

Table 4: Concentrations of heavy metals in sediments (|rg/kg Dry Weight)

No. of Station






Max Min

Max Min

Max Min

Max Min


687±7 528±4

126±2 106±3

282±4 127±5

1213±8 824±6


721±7 489±5

293±4 131±4

232±6 98±3

926±7 721±6


721±8 234±3

159±2 102±2

281±3 175±1

986±7 721±7


1021±9 324±3

302±5 98±2

315±7 141±5

1012±12 481±6


927±8 412±4

201±7 102±3

302±6 89±1

875±8 702±7


726±6 624±7

181±3 121±3

179±5 99±3

1015±11 609±5


915±8 521±6

181±6 69±2

241±6 197±6

902±7 517±3


598±4 408±3

100±1 63±1

104±2 63±1

598±4 402±3


The Cr and Cd concentrations in water and sediment recorded in this study are consistent with the findings of (Obire, et al.2003). Suspension of sediments into the water body may increase the metal concentration in the water. Sediment is the major depository of metals in some cases, holding more than 99% of total amount of a metal present in the aquatic system (Odiete 1999). In addition, heavy rain fall leads to farm draining. Large amounts of pesticides containing metal compounds are brought via surface run off from the farms to the river, contributing highly to the agricultural pollution.Chemical fertilizers containing Ni and Pb are used in agricultural industries of the regions around Gorgan Bay. Fungicides are used in fish farming in order to prevent epidemic diseases,such as fin erosion. Fungicides and algaecides are used in the paper industry for pulp making, in order to protect papers from mould development; these preparations contain copper compounds (Diagomanolin et al. 2004; Rauret et al. 1988). Gorgan Bay is receiving organic matter in amount exceeding its natural purification capacity due to high population and industrial growth. Pb, Cu, Cr and Ni compounds are used as pigments in the painting industry and as anti-fouling agents in marine paints (Diagomanolin et al., 2004; Ramelow et al. 1992). Otherwise, Local industries in Gorgan and Aq-Qala (the cities of Golestan province) use

Cr, Cd and Zn compounds in electroplating. In the past, natural purification and dilution were usually sufficient (Saad et al. 1994).Also; the wastewater of these industries is discharged to Gorgan coast and its related rivers directly, without any remediation; only a simple physical screening is being performed. Several sources for the increased metal concentrations appear likely (Diagomanolin et al. 2004).The results implies that of all the water, sedimentand fish species examined, the likelihood of obtaininghigh Pb dosage from eating fish from the study area ismore apparent than that of Cd and Cr. Pb is known to cause serious concern to health, but the contamination of the sea and sea products does not appear to be a matter for serious concern (Clark, 1997). The safe limits for heavy metals in seafood vary from region to region (Ashraf, 2006). The concentrations of Cr, Cd and Pb evaluated in the fish selected from Gorgan Bay indicated that consumption of fishes sampled from Caspian Sea may not cause hazardous effects on human health. However, determined values of Pb, Zn and Cd in the water samples were below the recommended standard limits (standard limit for Pb and Zn is about 0.1_0.2 mg/L and for Cd is 0.5 mg/L), but Cr values were higher than tolerable standards (0.05mg/mL; Joint FAO/WHO food standards programme, 1990). On the other hand, Pb and Cd intake from daily food is considered to be20 - 400 ppb and 400 - 500 ppb. So, the measured values of Pb and Cd in the fish samples were lower than standard detected metal concentrations in food(Emami-Khansari et al. 2005; Joine FAO/WHO food standards programme, 1990).

The aim of this study was determination of the heavy metals concentration in four species of most consumed fish, water and sediment sampled from Southern coast of Caspian Sea. Then, the heavy metal values were compared with standard limits. The results showed that level of the heavy metal in the evaluated samples are in standard range. However, there should be continuous environmental pollution monitoring to check heavy metals hazard. In order to control heavy metals level in water, sediment and fish (aqua culture)of Caspian Sea, quality control of water of farmlands,quality control of input and output water, and assessment and control of the metals content in water of the sea which

was exposed to water contamination,resulted from farmlands and industries have wide importance. Also, heavy metals content of input soils to Caspian Sea, which enter with rain and rivers form local farmlands and industries, should be assayed.Guidance for people and farmers about the instructions for use of pesticides or fertilizers, determination of sedimentation process in Caspian Sea, control of house wastewaters spreading in rivers and crops,establishment of reference laboratories equipped with analytical apparatuses etc are necessary.


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Inhibition ability of probiotic, Saccharomyces cerevisia

and Bacillus spore blend, against Ichthyophthirius multifiliis and study of its effect on growth of Guppy (Poecilia reticulata) ornamental fish larvae

Javad Sahandi1*, Christina Tadiri2, Hojatollah Jafaryan1, Javad Bagheri Dorbadam1

1 Fishery department, Faculty of natural resource, Gonbad Kavous University, Golestan Iran

2 Department of biology, McGill University, Montreal, Canada * Corresponding author:sahandijavad@gmail.com


Fish have managed to colonize most bodies of water from the deepest ocean to the highest mountain pools. Because of this ubiquity, they have been widely exploited by man for food. Additionally some species with attractive colors are kept as pets. Guppies (Poecilia reticulata) are one such ornamental species which are bred and exported to other countries by ornamental fish companies. This species was discovered in the middle of the 19th century. The two main aspects that determine the trade and prosperity of the ornamental fish industry are health and nutrition of the ornamental fishes. Success of larval rearing depends mainly on the availability of suitable food that is readily consumed, efficientlydigested and provides the required nutrients to support good growth and health (Tovar-Ramirez et al. 2002; Wache et al. 2006).Therefore, the use of probiotics in aquaculture has generated much interest and would be efficient on growth and fish survival rate.

Material and methods

Guppy larvae were obtained from a local ornamental fish store with the initial body weight 28.42 ± 13.84 mg. They were divided in 3 experimental treatments and a control with two replicates. Each 15 liter tank was stocked with 10guppies. Water temperature was maintained at 27 C, and water quality was monitored and recorded daily. 30% of

rearing water was siphoned and replaced every two days.Fish were hand-fed to apparent satiation, three times a day (7:00 am; 3:00 pm; 11:00 pm), with an Iso-nitrogenous (41% protein) diet, supplemented with dried yeast (Saccharomyces cerevisia) at levels, of 0%, 0.5%, 1%, 1.5%. Yeast concentrations were mixed with 50 mL distilled water and then sprayed into the food and mixed part by part. Guppy growth parameters wereassessed in terms of Condition factor (CF), Specific growth rate (SGR), Food conversion ratio (FCR), Relative food intake (RFI) and Growth coefficient efficiency (GCE) at the end of experiment. After weighing, 5 remaining fish in each expremental tank were challenged with Ichthyophthirius multifiliisto measure level of resistance to infection in each group. Fish were fed on the control diet and similar water temperature and quality parameters were maintained as observed in growth trial for 10 days. Upon termination of experiment, each fish was individually examined for the numberoflchthyophthirius multifiliis present. Total number of Ichthyophthirius multifiliis'specimens present on each fish were counted and recorded to estimate the intensity of infection.The differences in growth rates and parameters among the different experimental treatments were calculated using a one- way ANOVA followed by Duncan's multiple range tests to examine which of them varied significantly (SPSS ver. 19 software).


The growth and nutrient utilization data obtained from this study are presented in Table 3. At the end of experiment, it was found that the weight gain (%) had no significant difference (P>0.05) in among all yeast supplemented groups.

Table 3. The performance of growth parameters in reared Guppy fish


Treatment D1




WG (mg)

10.73 ± 7.63a

11.05 ± 8.08a

10.78 ± 6.95a

9.31 ± 4.53a

CF %

1.97 ± 1.12a

2.23 ± 1.35a

2.90 ± 2.80a

2.22 ± 1.47a


1.17 ± 0.15a

1.14 ± 0.11a

1.15 ± 0.13a

1.14 ± 0.10a


6.39 ± 2.18a

6.58 ± 1.86a

6.53 ± 1.90a

6.38 ± 1.41a


34.91 ± 30.60a

28.87 ± 22.34a

31.01 ± 26.20a

28.82 ± 20.26a


40.99 ± 36.45a

42.37 ± 41.63a

40.65 ± 33.85a

32.26 ± 20.75a

Likewies, no significant difference (p<0.05) was observed among yeast supplementation groups for food conversion Ratio (FCR) and specific growth rate (SGR) (P>0.05). No significant difference was observed in condition factor between groups (P>0.05). The same result was observed in relative food intake (RFI) and growth coefficient efficiency (GCE). Weight means of groups, in Figure 1(P>0.05).







gi ie 345 340 335 330 325 320 315 310

305 328.57 ±

213.80 337.5 ±

226.38 330 ± 194.65




320.58 ±




Figure1. Weight Mean Among Treatments

Group D3, which was fed with 1.5% yeast showed no mortality, while high mortality rate (30%) was observed in group D1, which was fed with the 0.5% yeast supplemented diet. Group D2 (20) and the control (10) the next highest mortality, respectively, after D1.

Table 4. Response of Guppy fish reared on artificial feed with different concentration of

Saccharomyces cerevisia when exposed to Ichthyophthirius multifiliisinfection

Diet with Saccharomyces cerevisia

Parameter Control -

_D1 (0.5 %)      D2 (1 %) D3 (1.5 %)

No. of fish exposed to

Ichthyophthirius multifiliis        5 5 5 5


Average number of

Ichthyophthirius multifiliis ± 020b 15 ± bc        14 ± 134bc 00 ± 00a

observed at each fish

Values in the same row with different superscripts are significantly different (P < 0.05).

The lowest infestation after challenge infection was observed in group D3, which was fed with 1.5 % yeast supplementation and 1 x 109 CFU/100g (P>0.05). There was a significant difference between D3 and all other groups. No significant difference was observed between D1 and D2 or between D1, D2 and the control (P>0.05).


This specie is the best known as a good choice for beginner aquarists, since they are hardy and reproduce rapidly. Our probiotic food fed fish exhibited no significant difference in growth performance when compared to the control food fed fish. This result implies that ornamental guppies utilize dietary nutrients with the same efficiency, regardless of whether or not the food is supplemented with yeast. In contrast, Kumar et al. (2006) and Ghosh et al. (2004)founda significant improvement in weight gain and survival in carps fed food containingBacillus. The present study found no significant impact of yeast supplementation onFCR and SGR,in contrast to Noh et al. (1994) and Lara-Flores et al. (2003), who reported that the use of Saccharomyces cerevisia in Carp diet and Nile Tilapia improved nutrient utilization. In our study the lowest FCR, RFI and GCE rates were observed in D2 group. Jafaryan et al. (2008) also reported that the probiotic Bacillus supplemented diet

significantly increased the weight, length and SGR of fish when compared with those on a control diet without probiotic supplementation, but in our study Bacillus spores at different concentrations did not show significant difference in growth rate among treatments. We did, however find a significant difference in survival rate. Group D3 (fed feed with 1.5 % yeast) showed 100% survival rate. The highest survival rate was related to group D3, followed by D2 then the control, then D1. In contrast to our result, Wache et al. (2006) reported that the use of Saccharomyces cerevisiae in diets for Oncorhynchus mykiss didn't show any significant difference in survival rate. Sahandi et al. (2010) reported that the use of probiotic increases survival rate Hypophthalmichthys molitrix larvae. The same result was also observed by Jafaryan et al. (2010) about Sasan (Cyprinus carpio). Yeast are able to provide P-glucans and nucleotides that stimulate the immune system of fish (Sahoo and Mukherjee 2001; Li et al. 2004). Significant difference was observed among groups after 10 days. Group D3 showed the highest response against Ichthyophthirius multifiliis, and no "ICH" parasites were found on fish skin. There was no significant difference among other groups (P>0.05), but significant difference observed between D3 and the other treatments (P<0.05). Similar results were reported by Harikrishnan et al. (2010) about gold-fish, by Abutbul et al. (2004) about tilapia and by Rao et al. (2006) about Indian carp. Similarly, the use of Saccharomyces cerevisia was found increase disease resistance in rosy barb and black tetra fish challenged with Aeromonashydrophila and Psudomonasfluorecens (Turenau et al. 2000).

Abutbul S, Golan-Goldhirsh A, Barazani O, Zilberg D. Use of Rosmarinusoffi- and immunostimulants on innate immunity of goldfish infected with AeromonasBogut, I., Milakovic,

Ghosh, K., Sen, S.K. & Ray, A.K. (2004) Growth and survival of rohu, Labeorohita (Hamilton) spawn fed diets fermented with intestinal bacterium, Bacillus circulans. ActaIchthyol.Piscat., 34, 155-165.

Harikrishnan R, Balasundaram C, Heo MS 2004 Effect of chemotherapy, vaccines cinalis as a treatment against Streptococcus iniae in tilapia (Oreochromis sp.). Aquaculture238:97-105.

Jafaryan H, Soltani M, Adineh H, Barzali H, Mazandarani R, (2010) Supplementation of experiment diet of Sasan (Cyprinus carpio carpio L.) larvae by useful microorganism for elevation of health and production , 2nd international congress on aquatic animal health management and diseases, Tehran, Iran, October 26-27 (2010).

Jafaryan,H., Asadi, R. & Bagheri, A. (2008) The promotion of growth parameters and feeding effciency of Acipenser nudiventris larvae by using of probiotic Bacillus via bioencapsulation of Artemia urmiana. In: Resource management, natural, human and material resources for the sustainable development of aquaculture. Aquacult. Europe 08.

Kumar, R., Mukherjee, S.C., Prasad, K.P. & Pal, A.K. (2006) Evaluation of Bacillus

subtilis as a probiotic to Indian major carp Labeorohita (Ham.). Aquac. Res., 37,


Lara-Flores M, Olvera-Novoa MA, Guzman-Mendez BE, Lopez-Madrid W. 2003. Use of the bacteria Streptococcus faecium and Lactobacillus acidophilus, and the yeast Saccharomyces cerevisiae as growth promoters in Nile tilapia (Oreochromisniloticus). Aquaculture 216: 193-201.

Li P, Lemis DH, Gatlin DM, 2004 Dietary oligonucleotide influence immune response and resistance of hybrid striped bass (Moronic chrysops x M. saxatilis) to Streptococcus iniae infection, fish and shell fish immunol 16:561-569.

Noh SH, Han K, Won TH, Choi YJ. 1994. Effect of antibiotics, enzyme, yeast culture and probiotics on the growth performance of Israeli carp. Korean J Anim Sci 36:


Rao YV, Das BK, Jyotyrmayee P, Chakrabarti R. 2006 Effect of Achyranthesaspera on the immunity and survival of Labeorohita infected with Aeromonashydrophila. Fish Shellfish Immunol 20:263-73.

Sahandi J, Soltani M, Jafaryan H, Rozbehfar R, Babaei S, Dehestani M, (2010) The using bioencapsulated rotifer (Brachionus plicatilis) with spores of Bacillus latrospores and Bacillus licheniformis for enhancement growth and decreasing of mortality in silver carp larvae, 2nd international congress on aquatic animal health management and diseases, Tehran, Iran, October 26-27 (2010).

Sahoo PK, Mukherjee SC, 2001 Effect of dietry P-1,2glucan on immune response and disease resistance of healthy and aflatoxin P1 induced immunocompromisedrohu (Labeorohita Hamilton). Fish and shellfish Immunol 11:683-695

Tovar-Ramirez D, Zambonino J, Cahu C, Gatesoupe FJ, Vazquez-Juarez R, Lesel R. 2002. Effect of live yeast incorporation in compound diet on digestive enzyme activity in sea bass (Dicentrarchuslabrax) larvae. Aquaculture 204: 113-123.

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