Journal of FisheriesSciences.com

Keywords

Oreochromis niloticus; Biochemical and histopathological changes; Nutritional values

Introduction

Fish and other aquatic animals are critical to providing a high-value protein source and important micronutrients for much of the world. As production from capture fisheries remains stable, aquaculture’s critical role in meeting increased demand for aquatic food products is driving researchers to assess the sustainability of the industry. In addition, consumers are becoming increasingly concerned with the environmental and ethical impacts of their food choices [1]. In Egypt, aquaculture has become an increasingly important activity as an immediate source of animal protein required for the country's growing population [2].

Fish parasites constitute the main cause of economic losses in aquaculture, including various types of pathogens (protozoa, trematodes, cestodes, nematodes, acanthocephalan and crustacean parasites), those causing, deformity, weight loss mortality etc [3]. Parasitic diseases of fish reduce the amount and quality of food available to peoples around the globe. It is imperative, therefore, to investigate the relationship between the environmental factors as it affects the parasites that affect production and quality of fishes [4]. Hematological parameters constitute an important tool that reveals the health state of fish. Parasitism may induce lowered growth and haematological alteration [5].

Fish diseases and histopathology, with a broad range of causes can be used as indicators of environmental stress, since they provide a definite biological end point of histological exposure, it is a mechanism which can provide an indication of fish health by determining early injury to cells and can therefore be considered an important tool to determine the effect of parasites on fish tissue [6]. Morevere, histopathological biomarkers can be sensitive indicators of subcellular stress in organisms exposed over short and long periods to a range of pollutants [7].

Materials and Methods

Abbasa Fish Farm, one of the most productive of the Egyption fish farms, is located in EL-Sharkia Governorate between latitudes 30° 32 ' North and latitudes 31° 44' East. The fish farm is about 2200 feddans. A total of 846 cultivated fishes Oreochromis niloticus were collected seasonally during the period from October, 2015 to September, 2017 formed the material for the present study Fishes were anaesthetized and blood was collected from the caudal blood vessels by a sterile syringe and kept forphysiological analysis. After dissection, a known weight of muscles was kept under freezing condition at 4°C until the latter examinations. Moreover, the clinical, postmortem, parasitological, physiological, histopathological and nutritional values were conducted.

Parasitological studies

Macroscopic examination

Macroscopic examination was done according to Syme [8] for the detection of any parasites in different parts of the fish body by nacked eyes and hand lens.

Identification of isolated parasites

Protozoans were identified according to Ahmed et al. [9]. Monogenatic trematodes were identified according to Eissa [3]. Cestodes and nematodes were identified according to Paperna [10]. Acanthocephala were identified according to Ramadan [11]. While crustacean parasites were identified according to Eissa [3].

Physiological studies

Blood samples were divided into three parts, the first part kept in clean and dry tubes and centrifuged for 15 min for separation of serum (collected serum was stored at -20°C) for aspartate aminotransferase (ASAT), alanine aminotransferase (ALAT) and total protein, The second part was taken on a dry and clean Eppendorf tube with sodium fluoride as anticoagulant factor and centrifuged for 15 min for separation of plasma for glucose analysis. The third part was taken on a dry and clean Eppendorf tube with EDTA solution for measuring hemoglobin (Hb), Red Blood Cell (RBCS), White Blood Cells (WBCs), Packed Cell Volume (PCV) [12].

Nutritional value of the fish muscles

Muscle samples were analyzed for moisture, protein; lipid, carbohydrates and ash were determined by using the standard process of [13].

Histopathological studies

Autopsy samples were taken from the liver, gills, muscles, skin, and eye of fish of different sized groups and fixed in 10% formol saline for twenty four hour. Washing was done in tap water then serial dilutions of alcohol (methyl, ethyl and absolute ethyl) were used for dehydration. Specimens were cleared in xylene and embedded in paraffin at 56 degree in hot air oven for twenty four hours. Paraffin bees wax tissue blocks were prepared for sectioning at 4 microns by slidge microtome. The obtained tissue sections were collected on glass slides, deparaffinized and stained with hematoxylin and eosin stain and examined under the electric light microscope [14].

Results

Seasonal dynamics of parasitic infection in different groups of O. niloticus

Results (Table 1) showed that, the highest value of protozoans in O. niloticus, was recorded during summer (12.77%) and the autumn (12.18%) and the lowest value (7.81%) was obtained during winter. However, prevalence of trematodes was declined gradually from 38.63% during summer to 27.10% during spring. While, the maximum prevalence of cestoda (5.92%) were detected during summer and the minimum value (0.88%) was observed during winter. On the other hand, the highest prevalence of acanthocephalan parasites was observed during spring (8.88%) and the lowest (4.39%) during winter. The highest prevalence of crustacean was recorded during summer and the lowest during autumn; being 14.33 & 4.06%, respectively.

Table 1: Seasonal dynamics of parasitic infection in O. niloticus, collected from Abbasa Fish Farm

Physiological studies

Data (Table 2) revealed that, Red Blood Cell count (RBCs), hemoglobin content and Packed Cell Volume (PCV) decreased in the infected Oreochromis niloticus, compared with the uninfected ones. Also, they were higher in female than in male groups. However, white blood cell counts were increased with the increasing infection. Also, WBCs values in females were higher than in males. Concerning O. niloticus, data showed increased activity of aspartate aminotransferase activity (ASAT) and alanine aminotransferase (ALAT) in the infected fish than the uninfected one of both sized groups. On the other hand, glucose increased with the increasing infection. Regarding to O. niloticus the highest values of total protein in the serum were recorded in uninfected small males and large females; being 4.13 ± 0.12 g/dl in the former and 4.31 ± 0.09 g/dl in the latter. While, the lowest values were recorded in the infected samples of the same sized groups; being 3.15 ± 0.14 g/dl in male groups and 3.00 ± 0.14 g/dl in female ones.

Table 2: Effect of parasitic infection on haematological and biochemical parameters in Oreochromis niloticus, collected from Abbasa Fish Farm, according to sex and sized groups.

Results revealed that, protein contents in O. niloticus, decreased with infection and increasing sized group. It was relatively higher in females than in males of this fish. Concerning O. niloticus, lipid contents in the fish males was fluctuated between 4.91 ± 0.28% in the uninfected group of small fishes and 5.65 ± 0.012% in the infected large sized groups. In females, however, it was varied from 4.76 ± 0.61% in uninfected small fishes to 5.46 ± 0.35% in the infected ones. Moreover, carbohydrate contents in the muscles decreased with infection and increasing sized groups. In general it was higher in females than in males, except uninfected large group. Results showed that, moisture in O. niloticus was slightly increased with infection and increasing sized groups. Regards to ash contents in male fishes was peaked in the uninfected large fishes (2.31 ± 0.41%) and declined in the infected small one (1.75 ± 0.21%). In females, however, it was varied from 1.76 ± 0.21% in uninfected small fishes to 2.68 ± 0.35% in the infected ones (Table 3).

Table 3: Effect of parasitic infection on the nutritional value (protein, lipid, carbohydrates, moisture and Ash) in the muscles of Oreochromis niloticus, collected from Abbasa Fish Farm, according to sex and sized groups.

Histopathological alterations on the different organs of the infected fishes

Microscopic examination of the gills of infected O. niloticus, with trematods revealed desquamation of primary and secondary gill filament epithelium and subepithelium infiltration of lymphocytes (Figure 1). Degenerated parasitic cyst at the tip of the primary gill filament; surrounded with moderately thick fibrous capsule with a prominent round infiltrated cells in the inter filamentous tissue was also observed. Epithelium of primary and secondary gill filament, revealed increased number of primary and secondary mucous secreting cells and subepithelial lymphocytic infiltration (Figures 2 and 3).

Figure 1: Photomicrograph of gills of O. niloticus, showing desquamation gill filament epithelium (yellow arrow) and sub epithelium infiltration of lymphocytes (black arrow) (H. and E. stain- X 40).

Figure 2: Photomicrograph of gills of O. niloticus, showing parasitic cyst embedded between lamellar epithelium with surrounding fibrous connective tissue capsule (H. and E. stain- X 100).

Figure 3: Photomicrographs of gills of O. niloticus, showing parasitic cyst embedded between lamellar epithelium with surrounding fibrous connective tissue capsule at the area of parasitic attachment is completely destroyed(A). Gills infested with Lamproglena parasite (B) (H. and E. stain- X 40, 100).

Histopathological examination of skin and muscles of the infected O. niloticus, showed encysted metacercaria among muscle fibers; it was represented by central degenerated parasitic element, surrounded by thin fibrous capsule, followed by aggregation of eosinophilic granular cells and lymphocytes. Morever, the affected muscle fibers showed distortion, degeneration and interstitial edema (Figure 4). Morever, muscle fibers showed interstitial edema, mild lymphocytic infiltration prominent field of cohenheim and longitudinal striations. Hyaline degeneration in some fibers "swelling and eosinophilia" and pyknotic nuclei were also detected (Figure 5).

Figure 4: Photomicrograph of skin and muscles of O. niloticus, showing encysted metacercaria (yellow arrow), fibrous capsule (red arrow) and aggregation of eosinophilic granular cells (green arrow) (H. and E. stain- X 100).

Figure 5: Photomicrograph of muscle of infected O. niloticus, showing interstitial edema (blue arrow) mild lymphocytic infiltration (yellow arrow) and pyknotic nuclei (green and red arrow) (H. and E. stain- X 40, 100).

Results (Figure 6) showed that, the liver of infected O. niloticus, have focal replacement of the hepatic parenchyma by cystic caseated material surrounded by aggregation of lymphocytes and melanomacrophages and degenerated parasitic cysts. The hepatic parenchyma showed small degenerated parasitic cyst surrounded by aggregation of lymphocytes. The hepatic cells showed hydropic degeneration. The hepatoportal pancrease revealed interstitial edema and infiltration of lymphocytes and melanomacrophages (yellow arrow). The bile duct is cystically fibroses. Concerning to infected O. niloticus, eyes, scleral congestion and edema were the main changes detected in the examined eyes (Figure 7).

Figure 6: Photomicrographs of the liver of infected O. niloticus, showing cystic caseated material (red arrow) aggregation of lymphocytes (blue arrow) degenerated parasitic cyst (brown arrow) hydropic degeneration (green arrow) edema (yellow arrow) bile duct cystically fibrosed (black arrow) (H. and E. stain- X 40,100).

Figure 7: Photomicrograph of the eye of infected O. niloticus, showing scleral congestion (red arrow) and edema (yellow arrow) (H. and E. stain- X 40,100).

Discussion

Parasites are attracting increasing interest from parasite ecologists as bio indicators of environmental pollution originating from human activities, due to the variety of ways in which they respond to such anthropogenic pollution [15].

The highest prevalence of protozoan of O.niloticus, was observed during summer and the lowest during winter. These findings was agree with that obtained by Abu El-Wafa [16] and Noor El-Deen [17] whom recorded that, the highest prevalence was recorded during summer and autumn and disagree with Soror [18] who revealed that, the highest prevalence was recorded during winter and spring. However, the highest values of prevalence encysted metacercariae in O. niloticus were recorded during summer and the lowest values were observed during winter. Similar observations were previously reported by many authors including [19-21]. The warm water during summer may increases the activity of cercarial emergence and penetration. Similar observation was cited previously by Harrod and Griffiths [22]. Results revealed that, the maximum prevalence of cestoda parasitic in O. niloticus, was recorded during summer and the minimum during winter; this could berelated to the availability of intermediate hosts of these parasites at these seasons and increase the feeding activity in warm temperature. These results nearly agreement with the findings recorded by Negm El-Din et al. [23,24]. The presence of fewer internal cestode parasites in O. niloticus could be attributed to its resistance to parasitic infections. This may be explained by the fact that O. niloticus species rarely succumb to disease epidemics and have a remarkable power of recovery from infections [25].

Blood is a good bio-indicator of the health of any organism; it also acts as a pathological reflector of the whole body. Hence, haematological parameters are important in diagnosing the functional status of the fish (host) infested by parasites and also to evaluate the physiological condition and nutritional state of fish [26].

The present study exhibited that, the reduction in RBCs count, Haemoglobin (Hb) value and packed cell volume (PCV) in the infected fishes may be as a result of the parasitic infestation that often leads to anemia. These observations agree with Tavares-Dias et al. [27] whom recorded decreasing in the erythrocyte numbers and hematocrit percentage in O. niloticus infected by parasites. Similar observation were recorded by Martins et al. [5] whom found that, the reduction in haemoglobin content in the infected catfish occurred as a result of the parasitic infection that often leads to anemia.

Moreover, the increase in WBCs count occurred as a pathological response since WBCs play a major role during infestation by stimulating the haemopoietic tissues and immune system to produce antibodies and chemical substances which work as defense agent against infection [28]. The obtained result was in agreement with those reported by many authors including Mlay et al. [29-31] whom concluded that, the positive correlation between WBCs and prevalence of parasites infection i.e. WBCs increased according to intensity of infection. Changes in WBCs and differential leucocyte counts have been reported to play an important role in assessing the health status of the fish. On other hand, the activities of ASAT and ALAT were significantly higher in the infected fishes when compared to the uninfected ones. Similar finding was observed by Nnabuchi et al., [26] whom reported that AST, ALT and Urea showed a significant increase in O. niloticus infected with external protozoa and monogenetic trematodes. Also, the increases in blood sugar level in the infected fishes may be due to increase in the breakdown of liver glycogen or due to decreased synthesis of glycogen from glucose. Hyperglycemic condition in naturally as wellas experimentally stressed fishes may be due to impairment in the hormone level in blood involved in the carbohydrate metabolism. These findings were in agreement with Ali and Ansari [31]. On the other hand, many fish diseases were found to alter the concentration of total protein, albumin and globulin. It has been shown that total serum protein content in the fish infected with parasites decreased significantly [32]. The depletion in protein and carbohydrate contents in the muscles of infected fishes may be due to the nutritional needs for parasites make them compete with the host's needs. The decrease in protein content may contribute to the effect of gut parasites which feed on gut contents of host fish. Result were in agreement with those recorded by Habiballah et al., [33,34].The increase of lipid in the muscles observed in the present investigation owing to the attack by various parasites might be the result of host's defensive mechanism to with stand the effect of infestation. In order to compensate the loss, due to intensive attack of parasites, the host tries to produce more of the nutrients. Results declared that, rise in moisture as a result of influence of parasites there was also a concomitant relation of moisture owing to multiple infestation, while the protein content showed an inverse relationship. Result was also similar to that observed by Mozhdeganloo and Heidarpour [35].

Histopathological examinations of fish collected from fish farm revealed, much higher incidence of pathological alterations. The effect of parasites in the present study can be detected using an analysis of pathologies, where the pathological alterations in fish are net result of adverse biochemical and physiological changes within the present finding were in agreement with [15]. The histopathological examination of the gill, liver, skin and muscles of the infected fish indicated that, the organs most affected by infection of parasites. This is similar to the observation of Rahman et al. [36,37].

Conclusion and Recommendation

From this study, we demonstrated that pathological outcomes and adverse health effects on fish could be related to habitat use, preferences and environmental variables associated with these. Fish farmers and sellers should be progressive on the potential risk of parasitic infestation in fishes in order to avoid economic loss and more studies on parasites to be conducted, selection of hosts and parasites as indicators of pollution and other stressors. Since some of these parasites are zoonotic to fish eating birds and mammals, the fish consumer should avoid eating half cooked fish. That is fish should be very well cooked before eating, this request for raising awareness in fish health management and the application of appropriate control measures.

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