Prevailing parasitic diseases affecting Oreochromis aureus in Lake Burullus

Abstract

Lake Burullus is an important source of fish production in Egypt; it produces 20.5% of the Egyptian fisheries' production. There is intense controversy about the heavy metal pollution in Burullus water and its effects on fish health and safety for human consumption. Heavy metals represent a major concern for aquatic life and could negatively affect fish health. Agricultural and industrial water drainage represents a considerable part of the lake water supply. The present work was conducted to determine heavy metal concentrations in lake water and blue tilapia Oreochromis aureus musculature. Water samples were collected from six locations to determine cadmium, copper, lead, zinc and iron. Sixty O. aureus fish samples were also collected from the same sampling points to assess the prevalent parasites infesting fish and to determine the heavy metal (Cd, Cu, Pb, Zn and Fe) concentrations in fish musculature then study the relationship between heavy metals concentration and prevailing fish parasites. Results indicated that 53.34% of the examined fish were infested with encysted metacercaria. Centrocestus formosanus, Prohemistomum vivax, and Euclinostomum heterostomum were retrieved from gills, musculature, hepatopancreas and the posterior kidney. The parasitic intensity in fish tissues was between 1 and 9 cyst g⁻¹. Centrocestus formosanus was identified using the polymerase chain reaction in the gill tissues of 16 fish. The gill parasitic copepod Lamproglena monodi was identified in one fish. Degenerative changes such as thickening, corrugation, and destruction of gill filament are the most dominant pathological changes in infested fish gills. Heavy metal concentrations in water samples were at normal levels, except for copper and iron in the southern part of the lake. All heavy metals in fish musculature were below the permissible limits. The parasitic infestation was more dominant in the northern part of the lake than in the southern region; this could be due to elevated copper concentration in the southern part of the lake that could negatively affect the survival of the first intermediate host and parasite cercaria. In conclusion, captured fish from Lake Burullus were safe for human consumption, and heavy metal pollution in lake water does not represent a severe risk.

Introduction

Lake Burullus is the second natural Egyptian Lake in size after Manzala; it extends over 462 km², of which 360 km² is open water. Burullus is now the first natural source of fisheries production in Egypt. Lake productivity reached 81,146 tons in 2020 (Mehanna et al. 2023). Lake Burullus is situated between the Rosetta and Damietta branches of the River Nile along the Mediterranean coast and lies between longitude 30° 30′ and 31° 10′ E and latitude 31° 21′ and 31° 35′ N. Burullus is connecting to the Mediterranean Sea through El-Burg inlet that allows water exchange between Lake and the Mediterranean (Abd el-sadek et al. 2022).

Lake Burullus produces 3.98% of the total fish production in Egypt, representing 20.5% of fisheries production. Tilapia production from Burullus was 46,775 tons in 2019, which equals 33.32% of the total captured tilapia in Egypt and 57.6% of the lake fisheries productivity (GFARD 2021). Nile tilapia is the most important fish species; Egypt is ranked as the third tilapia producer worldwide in 2021 (El-Sayed and Fitzsimmons 2023). Captured tilapia has higher economic value and marketability than cultured tilapia; captured tilapia is organic, more delicious, and completely natural (Komolka et al. 2020).

O. aureus is primarily a fresh and brackish water fish; it is distributed in a wide range of habitats such as streams, rivers, lakes, and ponds. It has a high tolerance to salinity and temperatures (Al-Wan and Mohamed 2019).

Radwan et al. (2022) isolated Centrocestus formosanus, Diplostomum tilapiae, Opisthorchis sp., Cyanodiplostomum sp., and Prohemistomum sp. from Nile tilapia collected from Lake Burullus. They also reported that iron, zinc, lead, copper, arsenic and cadmium levels were 0.865, 0.7, 0.088, 0.07, 0.05 and 0.004, respectively.

Fish parasites are the most prevalent diseases affecting wild fishes in their natural habitat; fish parasites are responsible for considerable mortality, particularly in smaller fishes (fries and fingerlings); parasites also can induce decreased fecundity and productivity (Ali and Aboyadak 2018). Parasites attract increasing interest from ecologists as potential indicators of environmental pollution (Uckun 2017). Pollution can increase parasitism by affecting the host defense mechanisms, thereby increasing host susceptibility or increasing the population of the intermediate or final hosts (Lafferty and Kuris 1999). Blanar et al. (2009) reported that pollutants negatively affect parasitism in aquatic animals. Sures and Nachev (2022) recorded that the pollutants directly influence the composition and diversity of parasite communities, particularly the free-living (larval) parasite stages, as the larval stages are in immediate contact with the environment. Pollutants induce lethal responses and lower the transmission efficiency of parasites which directly affect the dynamics of aquatic parasite populations.

Yearly, about 4.1 billion square meters of agriculture drainage enters the lake from several drains: El-Burullus drain (East), Drains number 7, 8, 9, and 11 located at the southern side of the lake, Brimbal canal on the western side receives freshwater from Rosetta branch, El-Burullus drain in the Western North of the lake, Terra and El-Gharbia drains. The lake also receives about one hundred million m³ per year of precipitation, increasing the water-residence time to about 2.5 months (Shakweer and Radwan 2004). Agriculture and industrial drainage may contain variable amounts of heavy metals that could affect the nature of lake water and aquatic life (Mao et al. 2023).

Heavy metals can find their way to fish bodies either by direct uptake from the water or indirectly through vegetation on contaminated algae, invertebrates, or smaller fishes. Most aquatic organisms can concentrate metals in their tissues (Bawuro et al. 2018). Heavy metals have a direct toxic effect on fish and could affect normal organ physiological functions if they exceed the acceptable level (Shahjahan et al. 2022).

Therefore, the present work aimed to determine the prevalent parasitic diseases affecting O. aureus. Estimating heavy metal concentrations in Lake water and fish musculature, then studying the possible correlation between parasitic infestation and heavy metal pollution.

Materials and methods

Study area

This study was conducted in April 2022; six points were selected for fish and water sampling:

Point (I): Boughaz El-Burullus (Lake–Mediterranean Sea connection).

Point (II): Bar-Baharry fish farms discharge.

Point (III): Baltim (Tera drain, El-Gharbia drain, and El-Burullus drain).

Point (IV): Drain No. 7.

Point (V): El-Shakhloba (drain No. 8 and 9).

Point (VI): Al-Hoks (drain No.11).

The specific positions of the different sampling points sites were recorded by the global positioning system (GPS) as presented in Table 1 and Fig. 1.

Table 1.

Fish and water sampling points

Sampling points	Location	Latitude	Longitude
I	El-Boughaz	31 33.326	31 4.513
II	Bar-Baharry	31 34.22	30 59.172
III	Baltim	31 32.464	30 57.865
IV	Drain No. 7	31 31.518	30 57.264
V	El-Shakhloba	31 29.818	30 56.598
VI	Al-Hoks	31 28.288	30 56.300

Fig. 1.

Lake Burullus map showing sampling points

Fish sampling

Ten blue tilapia (O. aureus) samples were collected from each sampling point using a gill net. Fish ranged between 39 and 42 g in body weight and 11–13.4 cm in total length. Fish samples were maintained separately in a polyethylene bag and placed in an ice box. Collected samples were transported to the fish diseases laboratory, inspected and dissected for parasitological examination. Muscular tissue samples were taken from three fishes at each sampling point; fish skin was removed using a plastic knife to avoid metal contamination, and then the dorsal musculature was dissected, labelled and maintained at − 20 °C.

Clinical and postmortem examination

The clinical examination was performed to observe abnormal signs or lesions. The internal examination was conducted in the laboratory according to the method indicated by El-Bahar et al. (2019).

Parasitological examination

Fish samples were observed for any external parasite on the skin, opercular chamber, and mouth cavity and then were dissected to detect internal parasites. Fish internal organs and gills were removed, placed in a Petri dish, and examined separately. The gastrointestinal tract was opened and inspected according to the method described by Eissa et al. (2012, 2013). Small pieces from the liver, kidney, gills, and musculature were compressed between 2 slides and microscopically examined for encysted metacercaria. Encysted metacercariae was counted in 100 mg of infested tissues to determine parasite intensity.

Molecular study

C. formosanus in infested O. aureus gills was detected by PCR following the method described by Jaruboonyakorn et al. (2022). Briefly, infected primary gill filaments were dissected under SFX-51 binocular stereomicroscope, Optika, Italy, and the infested filaments were subjected to pepsin digestion protocol following Duflot et al. (2021) for isolating EMC. EMC was collected by adding phosphate buffer to the digestion solution followed by centrifugation at 6000 rpm for 2 min at ambient temperature, and then the sedimented metacercaria was collected. Collected EMCs were subjected to DNA extraction as described by Ali et al. (2022). PCR was performed to detect C. formosanus by amplifying a partial region of the 28S rRNA gene after a few modifications. CFCytb primer designed by Jaruboonyakorn et al. (2022), F: 5′ GCT ATA TGT AGC GGA TGT TGA C 3′ and R: 5′ TCT TTC CAC GTC TAA CAC CC 3′, was used in the molecular study. The reaction mixture consisted of 1.5 μl from each primer (Intron, Korea), five μl from genomic DNA, 12 μl from 2× master mix (i-StarMAXTM II, Intron), and five μl nuclease-free water. Target gene amplification was carried out in SensoQuest Labcycler, Germany. The amplification cycle started with initial pre-denaturation at 94 °C for 5 min, followed by 35 cycles of denaturation at 94 °C for 1 min, annealing at 55 °C for 1 min, and elongation at 72 °C for 1 min. The run ended with a final extension step at 72 °C for 7 min. The PCR products were analyzed by 1.5% agarose gel electrophoresis as described by Soliman et al. (2022).

Histopathological study

After the parasitological examination, the infested fish tissues (gill filament, small cubes of infested hepatopancreas, posterior kidney, and muscular tissue) were fixed in a 10% neutral buffered formalin solution. After seven days, the fixed tissues were dehydrated in ascending-grade ethyl alcohol and cleared in xylene. Cleared tissues were placed in soft then hard paraffin wax and sectioned into 5-micron thickness sections. Tissue sections were mounted over a glass slide and stained with hematoxylin and eosin (H & E). Stained sections were examined and captured with OPTIKA® microscope with a digital camera, OPTIKA, Italy.

Water analysis

Physicochemical water parameters

Water temperature, dissolved oxygen, and salinity were determined at each sampling site using a multi-parameter electrochemical analyzer model, C6030 Consort, Belgium.

Heavy metals determination in water samples

Water samples were collected at 25–50 cm depth from the surface in a polyethylene bottle and kept in the ice tank. Three water samples were collected from each sampling point in between 1 km. One liter of each water sample was filtered using Wittman filter paper no (10) and re-filtered with a Polyvinylidene Difluoride membrane filter (0.45 μm pore diameter). The filtrated sample was concentrated to a final volume of 100 mL, acidified to pH 4.5 with 6 N analytical grade hydrochloric acid, and then preserved at 8 °C till analysis.

Glassware was soaked in diluted nitic acid (10%), washed with hydrochloric acid (10%) to remove any possible heavy metal residues, and finally washed several times with deionized distilled water (Abdel-Kader and Mourad 2023).

Cadmium, copper, lead, zinc, and iron (Cd, Cu, Pb, Zn, and Fe) concentrations in filtered water samples were assayed according to the method described by Elaarabi et al. (2022) in the atomic absorption spectrophotometer model Shimadzu AA-6800, Shimadzu, Japan using analar grade analytical standards Sigma, Germany.

Heavy metals determination in fish musculature samples:

Cd, Cu, Pb, Zn, and Fe were assayed in muscle samples as described by Dar et al. (2021). One gram of the dorsal musculature of each fish sample was taken, and the muscle samples from three fish in each sampling point were pooled together and then added to 6 ml nitric acid + 3 ml perchloric acid in a glass jar. The jar was tightly closed and left overnight at room temperature for predigest. The glass jar was placed on the hot plate at 100 °C until all muscle tissue dissolved. The samples were cooled to room temperature, diluted with deionized H₂O, filtered, completed using deionized H₂O to 10 ml, and filtrated with a 0.45 μm filter as indicated by Ali et al. (2021). Heavy metals were measured by atomic absorption spectrophotometer, Shimadzu AA-6800, and the results were expressed as µg g⁻¹.

Results

Clinical and postmortem examination

All of the examined fishes were normal and healthy. Minute yellowish foci of encysted metacercaria (EMC) were observed on the liver surface, and two whitish nodules (3 mm in diameter) were present on the posterior kidney of one fish.

Parasitological examination and molecular study

Thirty-three fish were infested (55%), in which 32 fish were infested with encysted metacercaria of digenetic trematodes, parasitic copepod Lamproglena monodi was isolated from the gill tissue of one fish. The parasitic infestation ranged between 10 and 40% in the southern part of the lake and increased markedly (70–100) % in the northern part of the lake (Table 2).

Table 2.

Infestation percent at each sampling point

Point	I	II	III	IV	V	VI
Infestation %	70	90	100	20	40	10

Thirty-two fish were infested with the encysted metacercaria of digenetic trematodes, 20 fish were infested with encysted metacercaria in gills, seven fishes had an infestation in the liver, 3 in muscular tissue, and only two in the posterior kidney (Table 3).

Table 3.

Distribution of encysted metacercaria in fish tissues

Encysted Metacercaria	Number of infested fish
	Gills	liver	Muscle	Kidney
Centrocestus formosanus	16	0	0	0
Prohemistomum vivax	4	7	3	0
Euclinostomum heterostomum	0	0	0	2

Centrocestus formosanus cysts were present in the gill tissue of 16 fish (Fig. 2a), and Fig. 2b showed the characteristic amplicon bands that appeared at 609 bp. The microscopical examination revealed that 14 fish were infested with Prohemistomum vivax metacercaria, four in the gills, seven in the liver, and three in the muscles (Fig. 2c & d). Euclinostomum heterostomum metacercaria was detected in the posterior kidney of two fishes (Fig. 2e and f).

Fig. 2.

a Compression smear showed Centrocestus formosanus encysted metacercaria in two adjacent primary gill filaments (blue arrows), X = 100. b Agarose gel electrophoresis showed the characteristic bands for C. formosanus at 609 bp, lane L is 100 bp ladder, lane N is the negative control, and lanes (1–16) are the positive samples. c and d Prohemistomum vivax encysted metacercaria in gill and musculature tissue of O. aureus, X = 200. e Posterior kidney of O. aureus with two Euclinostomum heterostomum metacercarial cysts (yellow arrow). F E. heterostomum trematode

The intensity of C. formosanus in fish gills ranged between 1 to 7 cysts per primary gill filament; the cyst was mainly present at the tip of gill lamellae, while the intensity of P. vivax was 0.33 to 1 cyst per gill filament. Both parasite cysts were oval to rounded. C. formosanus cyst size was ranged between 100 and 160 µm in diameter.

The intensity of P. vivax in the fish liver was 1 to 9 cysts per gram, but it was 1 to 2 cysts per gram in the dorsal musculature. Parasite cyst was rounded in shape and 315 to 335 µm in diameter.

Infestation with E. heterostomum was reported in two fish; 3 mm diameter parasitic cysts were present in the posterior kidney and were 3 mm in diameter. E. heterostomum trematode was 5.4 mm in length and 2.65 mm in width (Fig. 2f).

The examined fish samples were free from gastrointestinal parasites (either trematode, cestode or nematode).

The parasitic copepod (Lamproglena monodi) was 2.85 mm in length; its body consisted of the cephalothorax (0.69 mm in length & 0.43 mm in width), thorax (1.425 mm in length & 0.51 mm in width), genital segment (0.365 mm in length & 0.392 mm in width). The body ended with the three-segment abdomen (Fig. 3); the abdomen length was 0.96 mm and 0.29 mm in the first abdominal segment width, while the width of the third abdominal segment was 0.23 mm. The thorax has three pairs of legs 0.06 mm in length.

Fig. 3.

The crustacean gill parasitic copepod Lamproglena monodi, parasite body consists of cephalothorax, thorax carries three pairs of legs, genital segment and abdomen consists of three segments, X = 50

Histopathological study

Infested fish gills showed many degenerative changes, such as thickening, corrugation, and destruction of the supporting cartilaginous core. Pathological changes were more prominent when multiple cysts were present in the same gill filament (Fig. 4a and b). Destruction and sloughing of secondary gill lamellae with epithelial lifting were also observed. The posterior kidney of E. heterostomum-infested fish showed shrunk glomerular thought and widened Bowman’s space with interstitial hemorrhages around the cyst wall (Fig. 4c and d). P. vivax cysts in hepatopancreas and muscular tissue were surrounded with fibrous capsules (Fig. 4e and f).

Fig. 4.

a and b O. aureus gills infested with Centrocestus formosanus cyst (black arrows), cysts induced degeneration of the primary gill filaments (F), with destruction of cartilaginous core (blue arrows), epithelial lifting (green arrows) and thickened secondary gill filament (T), H & E, X = 40 (a), X = 100 (b). c and d Euclinostomum heterostomum cercarial cysts (M) in the posterior kidney, the cyst is surrounded with fibrous cyst wall (C), hemorrhages around cyst (H) and degenerated glomeruli (Yellow arrow), H & E, X = 40 (c), X = 100 (d). e Prohemistomum vivax encysted metacercaria (M) in the hepatopancreas, cyst surrounded by fibrous cyst wall (C), X = 400. F encysted metacercaria of Prohemistomum vivax (M) in the musculature of O. aureus, X = 100

Physicochemical water parameters at different sampling points

There was a minoructuation shift in the water temperature that ranged between (16 and 18.1) °C, but there was a considerable variation between sampling points in the dissolved oxygen (5.76–10.84) ppm and salinity (3–13.9) ppt. Generally, all values tend to be higher in the northern than the southern portion of the lake. The physiochemical water characteristics at different sampling sites are represented in Table 4.

Table 4.

Physicochemical water parameters at different sampling sites

Sampling point	Temperature (°C)	Salinity PPT	Dissolved oxygen (ppm)
I	18	13.9	10.84
II	18.1	7.6	7.04
III	16.5	6.1	8.15
IV	16.3	4	6.4
V	17.8	5.2	5.76
VI	16	3	6.72

I El Boughaz, II Bar-Baharry, III Baltim, IV Drain No.7, V El-Shakhloba, VI Al-Hoks

Heavy metals concentration in lake water in µg L⁻¹

Cadmium, lead, and zinc concentration in lake water was within the permissible limit and ranged and ranged between (0.94 to 2.88), (3.15 to 9.42) and (3.75 to 32.2) µg L⁻¹, respectively. On the other hand, copper and iron levels showed the greatest fluctuation among the tested elements. Copper and iron levels exceeded the acceptable value in three sampling points (Drain No. 7, El-Shakhloba, and Al-Hoks). Copper and iron levels reached 63.6 and 458.5 µg L⁻¹, respectively. Trace element concentration in lake water is shown in Table 5.

Table 5.

Heavy metals concentration in lake water at different sampling sites in µg L⁻¹

Element	Point
	I	II	III	IV	V	VI	PL
Cd	0.94 ± 0.075	1.23 ± 0.11	2.09 ± 0.35	1.47 ± 0.12	1.36 ± 0.1	2.88 ± 0.16	3
Cu	0.84 ± 0.11	15.72 ± 2.34	13.21 ± 2.03	58.1 ± 4.1	71.5 ± 7.3	63.6 ± 5.67	50
Pb	3.15 ± 0.38	8.21 ± 1.05	9.42 ± 1.4	6.2 ± 0.55	2.36 ± 0.22	5.1 ± 0.5	10
Zn	3.75 ± 0.25	25 ± 1.84	3.87 ± 0.11	32.2 ± 4.3	25.8 ± 4.07	21.73 ± 4.7	2000
Fe	51.87 ± 8.34	75.31 ± 4.38	147.2 ± 9.7	326.1 ± 42.95	297.6 ± 4.8	458.5 ± 32.5	300

PL: WHO permissible limit (Sankhla et al. 2021), N = 3, Values are Mean ± SE

Heavy metals in O. aureus musculature in µg g⁻¹ wet weight

The tested heavy metal concentrations were under the acceptable permissible limit recommended by WHO in muscles. Cadmium and copper concentrations ranged between (0.048–0.24) and (0.31–1.13) µg g⁻¹, respectively. The lead level was also under the accepted value in all sampling sites and was between 0.05 and 0.45 µg g⁻¹. Zinc and iron were also in the normal range between (3.84–7.24) and (8.73–27.3) µg L⁻¹, respectively. The heavy metal concentration in fish musculature is presented in Table 6.

Table 6.

Heavy metal concentrations in fish musculature in µg g⁻¹

Sampling point	I	II	III	IV	V	VI	PL
Cd	0.15	0.048	0.14	0.082	0.065	0.24	0.5
Cu	0.31	0.45	0.61	0.73	0.88	1.13	30
Pb	0.18	0.23	0.45	0.15	0.05	0.38	0.5
Zn	7.24	3.84	5.86	4.21	5.79	5.31	40
Fe	27.3	10.35	20.8	8.73	13.10	24.8	50

PL: WHO permissible limit (WHO 2006)

Discussion

Burullus is the most important northern lake in Egypt; it has a wide range of biological diversity, and so it was declared as a nature reserve under the International Ramsar Convention. The present work was conducted to determine heavy metal content in lake water and fish musculature and study the relation to prevailing parasitic diseases affecting O. aureus.

Regarding the physiochemical water parameters, water temperature, dissolved oxygen, and salinity tend to be higher in the northern than the southern part of the lake. Generally, all the recorded values are ideal for fish growth, reproduction, and aquatic life (Null et al. 2017). The agriculture drains downstream at the southern part represent the main water supply source of the lake. The water current moves toward the northern part; agriculture drains are narrow and deep, so the water temperature is lower than the surrounding, while the lake body is shallow (0.4 m in depth), allowing more sunlight penetration and raising temperature toward the northern part. Dissolved oxygen is extremely important for aquatic life (Abo-Taleb et al. 2020); the dissolved oxygen content was higher in the northern part than in the southern region. This change may be attributed to the aeration process throughout water passage under the effect of wind and the huge surface area of the lake with the role of photosynthesis. On the other side, the agriculture drains are reached in organic matter content which consumes oxygen for oxidation (chemical oxygen demand), which lowers the dissolved oxygen level in water (Sorour et al. 2003). Water salinity follows the same manner as temperature, dissolved oxygen, and water salinity increased toward the northern part and reached the maximum value (13.9 ppt) at El-Boughaz which connects the lake to the Mediterranean Sea and allows the entrance of salt water to the lake. Water parameter values were nearly like the previous values recorded by (Dewidar and Khedr 2005; ElTohamy et al. 2014; Alprol et al. 2021).

Heavy metals are the most dangerous pollutants in aquatic ecosystems due to their high toxicity and tendency to accumulate in living organisms. There is a serious debate about the heavy metal contamination of Lake Burullus; some researchers indicated the presence of high levels of heavy metals in the water (Melegy et al. 2019). They reported that contamination originated from the growing human population associated with expansion in agriculture, industrial activities, and urban and rural development. Heavy metals are serious pollutants that persist in the environment and are not subject to biological degradation like other synthetic organic chemicals (Venkatesan et al. 2022).

The present research indicated that lead, cadmium, and zinc in water were within the acceptable level and did not represent any hazard to aquatic life, other studies including (Masoud et al. 2011; Melegy et al. 2019; Alprol et al. 2021) confirmed the presence of metals within the normal range in lake water. It was evident that the lead level was higher in the northern part of the lake than in the southern region, which could be attributed to the intensive fisheries activity in the lake-northern using a huge number of fishing boats. The bottom of the fishing boat is coated with lead-containing paints (antifouling paint) to prevent the attachment of aquatic creatures like the barnacle and other crustacea that make boats slower and consume more fuel. Lead leakage into water from the paints increases the metal concentration in water. Gottesfeld (2015) also reported the same result. Cadmium and zinc levels tend to be higher in the southern part of the lake; this may be attributed to extensive fertilizer usage in agriculture processes; agriculture fertilizers contain a considerable level of both elements (Kubier et al. 2019), while cadmium and zinc levels were lower in the northern part due to dilution factor.

Copper and iron are essential minerals for humans, animals, and plants; copper and iron catalyze many metabolic and biological processes (Radakovich and Olver 2022). Copper and iron levels varied among the sampling sites; levels were higher and exceeding the (WHO 2004) guideline limits (50 & 300 μg/l respectively) at drain outlets (Drain No. 7, El-Shakhloba and Al-Hoks), then levels decreased toward the sea to be within the normal range. Copper is an essential component in agriculture practices; copper is used as a fertilizer; Copper sulfate is a potent pesticide for molluscs and worms (Husak 2015). Heavy copper use in agriculture explains the high copper levels in some water samples. Iron levels follow the same manner as copper; iron fertilizer chelated acts as a foliar fertilizer for plants and enters in chlorophyll synthesis (Zhang et al. 2022); this illustrates high iron levels in drain water.

The present work results are indeed different from (Melegy et al. 2019; Farouk et al. 2020) findings; they indicated higher levels of the studied elements; this difference could be attributed to different sampling time (2010–2015) while the present research is the most recent as samples were collected at 2022 after dredging of El-Boughaz that connects the lake with Mediterranean that allow water exchange and the entrance of seawater to the lake that decreasing pollutant and heavy metals concentration in lake water.

Tilapia is the dominant fish species in Burulus Lake, so it was selected for studying heavy metal residues. Results proved that heavy metals in fish musculature are below the WHO (2004) permissible limits, are acceptable by the international legislation limits of many other organizations and are considered safe for human consumption. Despite copper and iron being present at a high level in the water from some locations, it was within the normal range in fish samples collected from the same area. This finding may be referred to fish movement from the highly polluted region to another area lower in heavy metal concentrations. This result comes in complete harmony with El-Batrawy et al. (2018) findings; they found heavy metals below the hazardous range in tilapia musculature collected from Burulus Lake.

Collected fish samples were clinically normal and devoid of any external abnormalities. Internally minute faint foci were present in the liver and musculature, and large cysts were present in the posterior kidney for encysted metacercaria. Meanwhile, there was an inverse relationship between parasitic infection and heavy metal content in lake water; iron and copper levels exceeded the acceptable value in the southern part of the lake. In this region, the parasitic infestation was a minimum (10–40)% when compared with the northern part (70–100)% this may be attributed to the toxic effect of copper on the snails (the first intermediate host of digenetic trematodes). Copper has a molluscicidal effect at a dose of 40–64.3 mg L⁻¹ and is also toxic to miracidia and cercariae (Ibrahim et al. 2022). Accordingly, Cross et al. (2001) found that increased iron or copper levels decreased the swimming speed and viability of trematode cercariae which subsequently depress the cercarial infectivity for the second intermediate host (fish). The present research was in complete harmony with Radwan et al. (2022), who found a negative correlation between heavy metal pollution and parasitic infestation in cultured and wild Oreochromis niloticus. Accordance with the present research results (Blanar et al. 2009; Sures and Nachev 2022), indicated a negative correlation between parasitic infestation and pollution.

Encysted metacercariae infestation represents the most common parasitic affection of O. aureus; this could be due to the spreading of the intermediate host (snails) and the final host (birds) in the wild natural habitat and ecosystem of Burullus Lake. In accordance, Eissa et al. (2013) mentioned that encysted metacercariae infestation is the most common parasite affecting wild fish.

Molecular study assured the microscopical identification of Centrocestus formosanus in the studied O. aureus gills; in the present work, C. formosanus is the most prevailing metacercarial infestation affecting fish that was augmented by (Chai 2019; Jaruboonyakorn et al. 2022; Radwan et al. 2022). Fish gills are the target organ for C. formosanus; this could be due to hypervascularization, high activity, mobility of gills, and contact with water. Metacercaria attached to fish gills during the respiration process. Melanoides tuberculate is the intermediate host, and many species of piscivorous birds representing the final host of C. formosanus are normal inhabitants in Burullus Lake, explaining the endemicity of encysted metacercariae among lake fishes.

O. aureus gills infested with C. formosanus showed thickening, corrugation, and destruction of the cartilaginous core; degenerative changes were more prominent when multiple adjacent cysts were present in the same gill filament. These degenerative changes were nearly like (Rezaie et al. 2017; Sumuduni et al. 2018) observations. The metacercarial cysts induce a pressure atrophy and inflammatory reaction that destroys the infected gill filament.

Prohemistomum vivax encysted metacercaria was the second prevailing tissue parasite affecting O. aureus, cercarial cysts isolated from the liver, gills, and muscular tissue (Elaswad et al. 2021; Abd-ELrahman et al. 2023) also identified P. vivax metacercaria from O. niloticus and Clarias gariepinus collected from Nile turbinate. Pathologically, P. vivax did not induce any inflammatory or tissue reaction in the liver and muscular tissue of infested fish; this could be due to the thick fibrous tissue capsule surrounding the cyst and isolating it from the tissue and small cyst size, Abd-ELrahman et al. (2023) have also reported a similar finding in Nile tilapia infested with P. vivax.

E. heterostomum is one of the most common metacercarial affection of tilapia species (Abd-ELrahman et al. 2023; Shinn et al. 2023). Degenerated shrined glomerular thought and widened Bowman’s with the presence of hemorrhages around the cyst wall was observed in infested tilapia kidney; similarly, Kaur et al. (2016) reported detachment of renal tubules epithelial cells, tubular occlusion and necrosis, Shareef and Abidi (2015) also found hyperplasia and hypertrophy of tubular epithelium with fibrosis and hemorrhage in renal tubules of E. heterostomum infested fish. The pathological lesions and tissue destruction in the posterior kidney are associated with the pressure induced by the parasite cyst on the surrounding tissue.

Lamproglena monodi is an ectoparasitic copepod classified under Lamproglena genus and family Lernaeidae. In the present study, the L. monodi female was isolated from the gills of O. aureus, representing 3% of the recovered parasitic infestations. L. monodi prevalence in O. niloticus collected from the river Nile and Abassa fish farm was 44.8 and 6.5% (Abdel-Gaber et al. 2017; Al Malki et al. 2021), which was much higher than the present study results; this difference may be due to the presence of some agricultural pesticide residues in Lake water that could affect water insects including copepods.

Conclusion

Encysted metacercaria was the most prevalent parasitic disease affecting O. aureus; there was an inverse relationship between parasitic infection and heavy metal content in lake water. The toxic effect of copper on the first intermediate host of digenetic trematodes could be the most probable cause. Cadmium, lead, and zinc concentrations were within the permissible limit in Lake Burullus water, while copper and iron exceeded the acceptable value in the southern part. All heavy metal concentrations were in the normal range in fish musculature and did not represent any hazard to the human consumer.

Author contributions

IMA: heavy metal analysis, help in writing the original draft, reviewing & editing. MAH: water and fish samples collection. NGA: Study design, clinical and post-mortem examinations, parasitological examination and parasites identification, molecular study, writing, editing and reviewing the original draft.

Funding

The present work is not funded by any organization and funded only by the authors.

Data availability

The data that support the findings of this study are available in the sublimity file and any other data will be available from the corresponding author upon request.

Declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

Oreochromis aureus samples were handled, transported, examined, and euthanized following the National Advisory Committee for Laboratory Animals Research (NACLAR, 2004) and guidelines for the care and use of fish in teaching and research (CCAC, 2005). This work was approved by the Institutional Care of Aquatic Organisms and Experimental Animals (NIOF-IACUC) Committee, NIOF.

Consent for publication

All the authors approved to submission to Journal of Parasitic Diseases.