Abstarct: In present study we evaluated the effect of formaldehyde on decreasing the contamination rate of egg shell, yolk and yolk sac. 240 hatching eggs from broiler breeder farm and hatchery as well as 60 newly-hatched chicks (5 stages) were taken for bacteriological examinations. Firstly egg shells were rinsed with peptone water solution and then these solutions were transferred to cysteine selenite media and were incubated, also these solutions were incubated separately. Then we sterilized egg shells and broke shells, yolks were homogenized and transferred to peptone water and cysteine selenite media and were incubated. Furthermore we sterilized abdominal skin of each newly-hatched chicks, thereafter necropsied,yolk sacs were homogenized and transferred to peptone water and cysteine selenite media and then were incubated. After that we cultivated on McConky agar, blood agar and XLD from colonies and incubated them. Finally we used differential diagnostic media. At first stage from 18.33 % of egg shells, at second stage from 11.66 % of egg shells, at third stage from 8.33 % of egg shells and at fourth stage from 10 % of egg shells was separated Alcaligenes faecalis. Also at fifth stage from 11.66 % of yolk sacs was separated Alcaligenes faecalis. In any stages did not separate non-coliform gram-negative bacteria from yolk. Based on this research the existence of non- coliform gram-negative bacteria on the hatching egg shells is ordinary and natural. Egg collection as soon as possible after laying and properly disinfection with formaldehyde must be lead to a significant contamination reduction of non- coliform gram-negative bacteria with which the risk of bacteria penetration in to the yolk will decrease dramatically, but formaldehyde could not effect significantly (p = 0.323) on these contaminations. Only contamination reduction between stages 1,3 was significant (p = 0.049) so we conclude it is related to secondary fumigation by formaldehyde.

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