Epidemiological and Bacteriological Studies on Dead-in-Shell Embryos

Several types of bacteria have been associated with infection of yolk sac and death of chicken embryos. The most common of these are Staphylococcus, Streptococcus, Klebsiella, Escherichia coli, Enterobacter, Citrobacter, Proteus, Salmonella, Pseudomonas spp. and Mycoplasma [1-4]. Fungi have been also reported with dead-in-shell embryos [5,4]. Bacteria that cause dead-in-shell embryos are usually those of the normal bacterial flora of the intestinal tract, skin or feather. The bacteria can migrate to yolk and cause death of the embryos and hatched chicks. Bacteria may also gain entrance to the egg as a result of the ovary and ovarian follicles (transovarian transmission) [1].


Introduction
Several types of bacteria have been associated with infection of yolk sac and death of chicken embryos. The most common of these are Staphylococcus, Streptococcus, Klebsiella, Escherichia coli, Enterobacter, Citrobacter, Proteus, Salmonella, Pseudomonas spp. and Mycoplasma [1][2][3][4]. Fungi have been also reported with dead-in-shell embryos [5,4]. Bacteria that cause dead-in-shell embryos are usually those of the normal bacterial flora of the intestinal tract, skin or feather. The bacteria can migrate to yolk and cause death of the embryos and hatched chicks. Bacteria may also gain entrance to the egg as a result of the ovary and ovarian follicles (transovarian transmission) [1].
The purposes of the present study were: 1) to determine the bacterial causes of dead-in-shell chicken embryos; 2) to exhibiting the epidemiologic state of these isolated bacteria; 3) to report certain recommendations to minimize the deployment of death of embryos in the hatcheries.

Materials and Methods
Ten unhacthed solid eggs were collected at the end of the incubation period from each of three different local hatcheries in Erbil province. Egg collection was done twice monthly over a period of 18 months (2011-2012). During each visit, 15 unhacthed eggs were collected randomly to form one sample.
In the laboratory, a sample of five eggs was chosen as a representative sample from each hatchery. The eggs were washed thoroughly with a disinfectant (2% tincture iodine) and after dryness they were mopped with alcohol. Opening of the eggs was done aseptically and yolk contents were collected into sterile containers. In case of eggs with fully developed embryos, the unabsorbed yolk was used for pooling. Culture media that were used for bacterial isolation were Blood agar, MacConkey agar and Mannital salt agar. Identification of bacterial isolates were done on the basis of their colonial, morphological, cultural and biochemical properties [6,7].

Results
The types, numbers, and percentages of bacterial isolates done deadin-shell embryos from the three hatcheries are presented in Table 1. A total of 37 bacterial isolates were encountered in the three hatcheries.
The total isolated bacteria from the three hatcheries arranged in order of decreasing frequency were Escherichia coli (18 isolates), Pseudomonas spp. The number of dead-in-shell embryos with yolk sac infection in association with the various bacteria isolated from the three hatcheries and the severity of lesions in various organs of the dead-in-shell embryos are presented in Table 2.
From data presented in this Table 2 it becomes apparent that the most severe lesions were associated with Escherichia coli. All of the examined internal organs exhibited vascular changes and they were red congested in color and hemorrhagic.

Discussion
In this study, the prevalence rates of dead-in-shell chicken embryos in the three local hatcheries were 37%, 21.6% and 40.5% respectively. From these figures it would seem that the condition is quite common in hatcheries in Erbil Province. However a similar study has been in the Nenevha Province and according to that study it was possible to evaluate these figures. Bacteria that were isolated in this study are similar to those isolated by other workers in Nenevha (1) and other parts of the world [2,4,8]. Escherichia coli constituted 48.6% of the bacteria isolated from the three hatcheries. This finding is in agreement with that reported by others working on the same topic and in the same geographic location [1,9,10]. Similarly, Escherichia coli are responsible for the largest numbers of dead embryos in the three hatcheries (7 dead embryos). These lesions were consistent with Escherichia coli infection and they were similar to those reported in the literature [1].
In view of the high prevalence of dead-in-shell chicken embryos in local hatcheries, it could be recommended that: • All trays used for hatching should be disinfected before being used for the next hatch. Disinfection could be accomplished though dipping of trays in a tank of suitable disinfectant with formaldehyde in the hatcher.
• Egg used in the hatch must be cleaned and disinfectant in a manner similar to that used in cleaning and disinfection of the trays used for hatching. In fact, the two processes could be done together.
• Eggs to be used for hatching must be obtained from a wellknown and good breeder "code" (source of hatching eggs).
• All hatcheries must be under veterinary supervision and they must be visited periodically by the veterinarians to assure clean and healthy hatching.