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ABD-AL-AAL, Z.et al.

DIGENETIC TREMATODES OF THE LITTLE EGRET, EGRETTA GARZETTA, AND POSSIBILITY OF TRANSMISSION TO OREOCHROMIS NILOTICUS

AT EL-ABBASSA FISH FARMS, EGYPT

ABD-AL-AAL, Z.1, O. H.AMER2, A. I. I.BADAWY2 AND A.M.M. EL-ASHRAM3

  1. Dept. of Zoology, Fac. of Science.
  2. Dept. of Parasitology, Fac. Vet. Med., ZagazigUniversity
  3. Fish diseases Dept., Central Lab. for Aquaculture Research (El-Abbassa), Agricultural Research Center, Egypt.

Abstract

Migratory aquatic birds consider as a potential reservoir hosts for many parasitic diseases that

may affect seriously both farmed fish and the public health. In this study, the digenetic trematodes of the migratory fish-eating bird, Egretta garzetta, and the possibility of transmission to Oreochromis niloticus at El-Abbassa fish farms, Sharkia province were investigated. Out of examined 75 E. garzetta, 45 (60%) were infected with digenetic trematodes. Four mature species were identified, in which Euclinostomum heterostomum was the most prevalent (44%), followed by Clinostomum complanatum (12%), Apharyngostrigea cornu (8%) and Strigea falconis (4%). In one bird (1.3%), an unidentified encysted metacercaria species was found on the subcutaneous and underlying muscular fascia for the first time in this bird species. Investigation of 350 O. niloticus for digenetic trematodes encystation revealed a total infection rate of 31.14%, represented by five types of encysted metacercariae including Centrocestus species (11.1%), Prohemistomum species (10.9%), Posthodiplostomum species (7.7%), C. complanatum (0.9%) and E. heterostomum (0.6%). It appears that the migratory E. garzetta could be a source of infection to O. niloticus at El-Abbassa fish farms with the zoonotically important parasite, C. complanatum, as well as E. heterostomum. The study recommends investigation of other fish-eating birds for determination of reservoirs of parasitic diseases at this region.

INTRODUCTION

Migratory fish-eating birds should be considered as a potential reservoir hosts for many parasitic diseases that affect seriously the birds, fish and the public health (Torres et al., 1991; Kijewska et al., 2002; Barson and Marshall, 2004 and Mattiucci et al., 2008). On the other hand, fish-borne zoonotic trematodes represent an important group of human parasites, which transmitted to man through eating of raw or insufficiently cooked fish and consider a serious risk factor on the public health (Thu et al., 2007 and Trung Dung et al., 2007).According to the reports of the world health organization (WHO), the number of people currently infected with fish-borne trematodes exceed 18 million and the number of people at risk worldwide estimated at more than half a billion (Thu et al., 2007). Regarding this point of view, this study was conducted to investigate the digenetic trematodes of the fish-eating bird, E. garzetta, and possibility of transmission to O. niloticus at fish farms of El-Abbassa, Sharkia province through investigation of this fish species for digenetic trematodes encystation.

MATERIALS AND METHODS

Birds

Seventy five little Egret, E. garzetta, were captured at El-Abbassa fish farms, El-Abbassa, Sharkia province during the period extended from October 2007 to April 2008. Birds were submitted to the laboratory at the department of parasitology, faculty of veterinary Medicine, ZagazigUniversity, identified according to Brown et al. (1982) and examined for the digenetic trematodes.

Examination of birds for digenetic trematodes and preparation of permanent specimens

Birds were opened longitudinally, the skin was reflected and the underlying subcutaneous tissues, the body cavity and the viscera were examined by the naked eyes. The digestive system, beginning from the peak to the cloaca, was opened and any found parasites were collected. Intestinal mucosal scrap as well as the contents were washed several times with physiological saline, and after sedimentation, the sediments were examined under dissecting microscope for parasites. The recovered adult digenetic trematodes were relaxed in the refrigerator, fixed in formalin 10% and stained with acetic acid alum carmine stain according to the technique of Kruse and Pritchard (1982). The encysted metacercaria, recovered from the subcutaneous tissues of birds, as well as the excysted ones (by gentle pressure between two glass slides) were directly photographed without staining.

Examination of O. niloticus for encysted metacercariae

Three hundred and fiftyO. niloticus were collected from El-Abbassa fish farms between the period extended from October, 2007 to April, 2008 and examined for digenetic trematodes encystation. Fish were macroscopically examined, both externally and internally, for presence of macroscopic encysted metacercariae. Direct compression between two glass slides as well as using the standard pepsin digestion solution (Meyer and Olsen, 1971), as well as, haematoxylin and eosin stained histopathological sections from the gills (Bancroft et al., 1996) were used for recovery and isolation of encysted metacercariae. The recovered metacercariae were identified according to Paperna (1996).

RESULTS

Incidence of digenetic trematodes in the little Egret, E. garzetta

Investigation of 75 E. garzetta collected from El-Abbassa fish farms, Sharkia during the period extended from October, 2007 – April, 2008 for infection with digenetic trematodes revealed that 45 birds (60%) were infected (Table, 1). Four mature digenetic trematodes species were recovered from the digestive tracts of examined birds. Two species were recovered from the pharynx and oesophagus of infected birds, in which Euclinostomum heterostomum was the most prevalent (44%), followed by Clinostomum complanatum (12%). Apharyngostrigea cornu (8%) and Strigea falconis (4%) were recovered from the small intestine of infected birds. The intensity of infection of E. garzetta with mature digenetic trematodes worms was particularly low, where the recovered maximum number of worms per infected bird was 6, 6, 8, and 2 for E. heterostomum, C. complanatum, A. cornu, and S.falconis, respectively. In only one case (1.3%), an unidentified macroscopic encysted metacercaria species was detected in the subcutaneous and underlying muscular fascia of the thigh region of onebird (Table, 2).

Table 1. Incidence of digenetic trematodes in little Egret, E. garzetta, captured at El-Abbassa fish farms

Bird / No. examined / Infected / %
E. garzetta / 75 / 45 / 60

Table 2. Incidence of digenetic trematode species in little Egret, E. garzetta, hunted at El-Abbassa fish farms

Birds / E. garzetta
No. examined / 75
Digenetic trematodes / Site / Infected / % / Intensity of infection
(Max. No/Bird)
E. heterostomum / Pharynx and oesophagus / 33 / 44 / 6
C. complanatum / 9 / 12 / 6
A. cornu / Small intestine / 6 / 8 / 8
S. falconis / 3 / 4 / 2
unidentified encysted metacercaria / Subcutaneous and underlying muscular fascia / 1 / 1.3 / 18

Prevalence of encysted metacercariae of digenetic trematodes in O. niloticus

Among 350 examined O. niloticus collected from El-Abbassa fish farms between the period extended from October, 2007 to April, 2008, 109 (31.14%) proved to be infected with digenean encysted metacercariae. Five species of metacercariae could be identified. High infection rate with Centrocestus species (11.1%) was recorded, followed by Prohemistomum species (10.9%), Posthodiplostomum species (7.7%), C. complanatum (0.9%), and E. heterostomum (0.6%). The highest infection rate of O. niloticus with the metacercariae (52%) was observed in October, 2007, while the lowest infection rate (10%) was observed in December, 2007 and January, 2008 (Table 3).

Table 3. Prevalence and monthly fluctuation of encysted metacercariae of digenetic trematodes in O. niloticus at El-Abbassa fish farms

Month / No. of examined fishes / Total infected / Metacercariae
Macroscopic / Microscopic
E. heterostomum / C. complanatum / Posthodiplostomum sp. / Centrocestus sp. / Prohemistomum sp.
No. / % / No. / % / No. / % / No. / % / No. / % / No. / %
October / 50 / 26 / 52 / 0 / 0 / 0 / 0 / 15 / 30 / 4 / 8 / 7 / 14
November / 50 / 22 / 44 / 0 / 0 / 0 / 0 / 12 / 24 / 5 / 10 / 5 / 10
December / 50 / 5 / 10 / 0 / 0 / 0 / 0 / 0 / 0 / 3 / 6 / 2 / 4
January / 50 / 5 / 10 / 0 / 0 / 0 / 0 / 0 / 0 / 3 / 6 / 2 / 4
February / 50 / 9 / 18 / 0 / 0 / 0 / 0 / 0 / 0 / 6 / 12 / 3 / 6
March / 50 / 20 / 40 / 0 / 0 / 0 / 0 / 0 / 0 / 10 / 20 / 10 / 20
April / 50 / 22 / 44 / 2 / 4 / 3 / 6 / 0 / 0 / 8 / 16 / 9 / 18
Total / 350 / 109 / 31.14 / 2 / 0.6 / 3 / 0.9 / 27 / 7.7 / 39 / 11.1 / 38 / 10.9

Morphological features of the detected digenetic trematodes

Mature worms recovered from E. garzetta

C. complanatum (Plate 1, A)

The worm was pear shape, with a narrow anterior end which widens greatly at the level of the ventral sucker. It measured 5.5 mm in length and 3 mmin breadth. The oral sucker was terminal and measured 234 µm x 450 µm, while the ventral one measured 1.1 mm x 1.0 mm and apart from the oral sucker by 1.1 mm. The intestinal caeca were simple and reached the posterior end of the body. The testes were tandem in position, and lied near the middle of the body and measured 0.8 mm x 1.2 mm and 0.6 mm x 1.4 mm. The ovary was ovoid, between the testis and measured 0.7 mm x 1.0 mm. The ootype located beside the ovary and measured 0.4 mm x 0.3 mm. The vitellaria were present laterally and extended from the level of the ventral sucker to the end of the body. The eggs were oval, operculated and measured 144 µm x 72 µm.

E. heterostomum (Plate 1, B)

The body was truncate anteriorly, then widens greatly near beginning of the lateral branching of the intestinal caeca and broadly rounded posteriorly. It measured 1.9 cm x 3.5 mm. The oral sucker was subterminal, lied about 0.5 mm from the anterior end and measured 0.4 mm x 0.48 mm, while the ventral one measured 1.7 mm x 1.6 mm. The oesophagus leads to an intestinal caecum, which bifurcates anterior to the ventral sucker into two main intestinal branches, which in turn give rise to 10 – 13 simple secondary blind diverticula begins behind the ventral sucker to the end of the body. The testes were lobulated, tandem in position and lied caudally in the last third of the body. The anterior testis measured 1.8 mm x 2.1 mm, while the posterior one measured 1.8 mm x 1.6 mm. The ovary was ovoid, between the testis and measured 1.1 mm x 1.5 mm. The ootype was beside the ovary and measured 0.8 mm x 0.6 mm. The vitellaria were present laterally and extended from the level of the ventral sucker to the end of the body. The eggs were oval, operculated and measured 130 µm x 76 µm.

S. falconis (Plate 1, C)

The body was divided into fore body, measuring 1.0 x mm 0.98 mm, and an elongated cylindrical hind body, measuring 4.5 mm x 0.95 mm. The oral sucker was rounded and measured 144 µm in diameter, while the ventral sucker was larger and measured 252 µm in diameter and lied behind the oral one by about 180 µm. A bilobed tribocytic organ was located caudally in the fore body. The testes were tandem, in the second half of the hind body and measured 594 µm x 576 µm and 630 x 576 µm for the anterior and posterior testis, respectively. The ovary was oval, measured 234 µm x 360 µm and lied anterior to the testes. The uterus had a thin loop, laterally and opens in a genital cone caudally. The eggs were oval, operculated and measured 108 µm x 90 µm. The vitellarine glands were in form of small follicles, along the whole lateral side of the hind body and present also caudally in the fore body.

A. cornu (Plate 1, D)

The body of the worm consisted of a pear shaped fore body, which measured 1.6 mm x 1.0 mm, and an elongated cylindrical hind body, measured 3.3 mm x 0.8 mm. The oral sucker was terminal, rounded in shape and measured 180 µm in diameter. The ventral sucker measured 306 µm in diameter and lied behind the oral one by about 360 µm. A triangular shaped Tribocytic organ was located caudally in the fore body. The testes were tandem, in the second half of the hind body and measured 630 µm x 540 µm and 720 x 486 µm for the anterior and posterior testis, respectively. The ovary measured 180 µm x 360 µm and lied anterior to the testes. The uterus had a small ascending limb and contained oval, operculated eggs which measured 126 µm x 68 µm. The vitellaria were in form of small follicles, present laterally in the whole hind body and extended in the fore body reaching the tribocytic organ.

Unidentified encysted metacecaria from E. garzetta (Plate 2)

The encysted metacercaria appeared as macroscopic globoid whitish cysts, measured about 0.96 mm in diameter. Excysted metacercaria was fragile and ruptured spontaneously under the glass cover after a short period of examination with the microscope. The metacercaria showed a somewhat pointed anterior end and a much rounded posterior one.

Encysted metacercaria from O. niloticus

C. complanatum (Plate 3, A)

The body was convex dorsally and concave ventrally and measured 5.1 mm x 2 mm. The oral sucker was small and subterminal, while the ventral one was larger and situated in the anterior third of the body. The intestinal caeca were simple, reaching the posterior end of the body. The testes were lobulated, tandem in position and located in the second half of the body, and the ovary between them. Vetillaria extended from level of the ventral sucker to the end of the body.

E. heterostomum (Plate 3, B)

The body was truncate anteriorly and more rounded posteriorly. It measured 6.6 mm x 1.1 mm. The oral sucker was small, rounded and subterminal, while the ventral sucker was larger and located in the first third of the body. Both intestinal caeca showed about 10 – 13 simple lateral blind diverticula. The testes were lobulated and located in the second half of the body, and the ovary between them.

Prohemistomum species (Plate 3, C & D)

The cysts were spherical and surrounded with an easily ruptured fragile outer cyst wall. It measured 0.26 – 0.3 mm (aver. 0.28 mm) in diameter. The excysted metacercaria appeared oval in shape, with a somewhat pointed anterior end and a round posterior one.

Posthodiplostomum species (Plate 3, E)

The cysts were transparent and ovoid in shape. It measured 0.98 – 1.0 mm (aver. 0.99 mm) in length and 0.7 – 0.9 mm (aver. 0.8 mm) in breadth. The metacercaria was enclosed in a double layered cyst wall, in which the outer layer was thick and dark in colour, while the inner one was thin.

Centrocestus species(Plate 3, F & G)

A double layered oval or elliptical and hyaline transparent cysts. It measured 1.9 - 2.0 mm (aver. 1.95 mm) in length and 0.13 – 0.14 mm (aver. 0.13 mm) in breadth. The outer cyst wall was thin and fragile, while the inner one was thick and difficult to be ruptured. The body of the metacercaria was pigmented.

Plate 1. Mature digenetic trematodes recovered from E. garzetta at El-Abbassa fish farms (Alum carmine stain). A: C. complanatum (Bar= 1.6 mm); B: E. heterostomum (Bar= 5 mm); C: S. falconis (Bar= 1.2 mm); D: A. cornu (Bar= 0.8 mm).

Plate 2. Unidentified encysted metacercaria recovered from the subcutaneous and underlying muscular fascia of E. garzetta. A: Photograph of the encysted metacercaria (Arrows) on the subcutaneous and underlying muscular fascia (Bar= 10 mm); B: Micrograph of the encysted metacercaria (Bar= 0.5 mm); C: Metacercaria out from the cyst wall (Bar= 0.6 mm); D: Excysted metacercaria (Bar= 0.4 mm).

Plate 3. Encysted metacercariae recovered from O. niloticus at El-Abbassa fish farms. A: C. complanatum (Alum carmine stain) (Bar= 0.8 mm); B: E. heterostomum (Alum carmine stain) (Bar= 1.1 mm); C & D: Prohemistomum species (Arrows) (Unstained) (Bar= 0.1 mm); E: Posthodiplostomum species (Unstained) (Bar= 0.6 mm); F: Centrocestus species encysted metacercaria in gills (Arrow) (Unstained) (Bar= 1.3 mm); Histopathological section in gills showing Centrocestus species metacercaria (Arrow) (Haematoxylin and Eosin stain) (Bar= 2.7 mm).

DISCUSSION

This study was conducted to investigate the digenetic trematodes of the migratory fish-eating bird, little Egret (E. garzetta), and the possibility of their transmission to O. niloticus fish farms at El-Abbassa region, Sharkia province. Sixty percent of birds harboured mature worms of four digenetic trematode species. A similar high prevalence rate of adult digenea in Egrets was also recorded previously (Ryang et al., 1991 and Aohagi et al., 1992 and Navarro et al., 2005).

Results of this study revealed that the prevalence rates of E. heterostomum and C. complanatum in E. garzetta were 44% and 12%, respectively. As early stated(Aohagi et al., 1992), clinostomatids were prevalent in Egrets and other fish eating birds. The wide geographical distribution and high infection rates of clinostomatids might be related to the wide range of birds acting as final hosts as well as fish acting as intermediate hosts for these worms as previously reported (Aohagi et al., 1992 and Chung et al., 1995).

In the present study,A. cornu was recovered from the small intestine of 8% of the examined E. garzetta. Similar findings were documented in Ardea cinerea and E. garzetta in Spain (Navarro et al., 2005).

As described by Ryang et al. (1991), birds of the family ardiedae were proven to be final hosts for the strigeid digenean, S. falconis, where the authors isolated S. falconis from 83% of Ardea alba in Korea. In comparison to clinostomatids, the infection rate of E. garzetta with strigeids was low during this study. This is may be attributed to the host's diet as well as abundance of the intermediate hosts of the worms.

In one E. garzetta, an unidentified species of subcutaneous metacercaria was detected. Although encysted metacercariae of digenetic trematodes were not previously detected from little egret, E. garzetta, but this finding could be supported by the findings of Kramer et al. (1996), who recorded a 38-year-old man suffering from bronchospasms and isolated later a mesocercaria, most likely that of Alaria spp. or Strigea spp, from the subcutaneous nodule. The authors stated that eating undercooked wild goose meat during a hunting trip was the most likely source of infection; and also, with the findings ofKrone and Streich (2000), who recovered the metacercaria of Strigea falconispalumbi from the connective tissue of the neck of 10% - 58% of the Eurasian buzzards in different localities in Germany.

Except minor differences in the measurements, morphological features of the detected mature digenetic trematodes in this study were in close agreement with the preceding descriptions (Yamaguti, 1958; El-Naffar et al., 1980; Amer and Gattas, 1993 and Chung et al., 1995). The difference in measurements of the worms might be attributed to the birds, from which the parasites were collected and the methods of preparation of the parasites for examination.

Examination of O. niloticus revealed five types of digenean encysted metacercariae, with a total infection rate of 31.14%. Similar records of high infection rate of O. niloticus with digenean metacercariae were also described (El-Ashram, 2003 and Abd El Rahman, 2005).

In the present study, the prevalence rates of C. complanatum and E. heterostomum encysted metacercariae in O. niloticus were 0.9% and 0.6%. These low infection rates were unexpected; especially the prevalence rates of adult parasites of both species in the E. garzetta were high. As well as results of the previous related studies (El-Ashram, 2003), who found that 34% and 29% of this fish species were infected with these types of metacercariae, respectively; and that of Abd El Rahman (2005), who found that 45% and 11.8% of O. niloticus were infected with Clinostomum species and Euclinostomum species metacercariae, respectively. This is can be returned to the difference in sampling periods and the ponds from which the examined fishes were obtained.

Centrocestus species metacercariae were detected in 11.1% of examined O. niloticus. Clinostomum species, Euclinostomum species, Posthodiplostomum species and Centrocestus species proved to be commonly affected tilapias and other fresh water fishes (El-Nobi, 1998 and Dzikowski et al., 2003). Prohemistomum species and Posthodiplostomum species metacercariae affected 10.9% and 7.7% of the examined tilapia species.