| In 2003, the genus Gallibacterium was established within the family of Pasteurellaceae by Christensen et al. based on 16S rRNA gene sequences. In 2009, Bisgaard resported that the genus Gallibacterium currently includes four species: Gallibacterium anatis, Actinobacillus salpingitidis, Gallibacterium melopsittaci and Gallibacterium trehalosifermentans. And the Gallibacterium anatis is the representative species. Two biovars are described within Gallibacterium anatis, a haemolytic biovar haemolytica and a non-haemolytic, biovar anatis. Now a little research on the Gallibacterium anatis was resported in domestic. To further understand the infection of Gallibacterium anatis in parts of farms in China, thirty flocks from Henan, Shanxi, Shandong provinces were investigated for the occurrence of Gallibacterium anatis, in which fifteen layer flocks with sapingitis, two SPF flocks, six healthy layer flocks, five rooster flocks and two breeder roosters were included. Meantime, in order to carry out the molecular epidemiology, molecular genetics and pathogenesis of Gallibacterium anatis, the partial characteristic of Gallibacterium anatis isolates were studied, including cultural characteristics, biochemical properties, drug resistance and pathogenicity. The results were as follows:1. One hundred and seven layers from fifteen sapingitis flocks in Henan and Shanxi provinces were submitted for necropsy and the following organs were examined for bacteria: choana, trachea, lung, heart, liver, spleen, ovary, oviduct, duodenum and cloaca. Seventy-six Gallibacterium anatis strains and one hundred and eighty E.coli strains were isolated by morphological observation, differential cultivation, PCR and sequence analysis. And the infection rate of Gallibacterium anatis in the sapingitis flocks was up to 42.9%, indicating that Gallibacterium anatis was prevalence in the sapingitis flocks of Henan and Shanxi provinces. Among the one hundred and seven layers with sapingitis, eleven layers were infected Gallibacterium anatis singly, which illustrated that Gallibacterium anatis participated in pathological processes of salpingitis. Among the forty-five layers infected Gallibacterium anatis, thirty-five layers were infected E.coli simultaneously. Thus, the exact role of Gallibacterium anatis played in salpingitis, primarily, secondary or opportunist needed to be proved by subsequent researches.2. By morphological observation, differential cultivation, PCR, ten Gallibacterium anatis strains were isolated from ten tissues of thirty-three clinically healthy layers from six different farms in Henan and Shanxi provinces. No Gallibacterium anatis was isolated from thirty-eight SPF chickens in Shandong province. And the infection rate of Gallibacterium anatis in clinically healthy layers was up to 27.3%, indicating the existence of clinically healthy layer flocks. But the rate reduced by 15.6% compared with salpingitis fowls, which indicated that chicken salpingitis was an important clinical feature of Gallibacterium anatis.3. Twenty roosters from five different farms and nine breeders from another two farms in Henan province were submitted for necropsy and the following organs were examined for bacteria: choana, trachea, lung, heart, liver, spleen, duodenum and cloaca. By morphological observation, differential cultivation, PCR, no Gallibacterium anatis was isolated from tewnty roosters. However, eight Gallibacterium anatis strains were isolated from nine breeders, showing that the breeders chould infect Gallibacterium anatis. One of the eight Gallibacterium anatis isloations wasγ-hemolytic on sheep blood agar plates, which may be useful to the pathogenic study of Gallibacterium anatis.4. Sixty-four Gallibacterium anatis isolates from different flocks were selected to study the cultural characteristics, biochemical properties, drug resistance. The results showed Gallibacterium anatis on sheep/ chicken/duck blood agar plates growed well. However, no growth on SS agar plates. After the culture of Gallibacterium anatis in different PH value of LB broth, the optimal PH value range for growth was determined at 7.0～8.0. The culture of Gallibacterium anatis on sheep blood agar plates in different temperatures indicated that Gallibacterium anatis growed well at 31～40℃, and in this range, the lower the temperature the more obvious hemolytic. In addition, each strain differences in sensitivity to temperature and PH. Known the cultural characteristics was useful to improve the rate of isolation and separation efficiency. Biochemical test results showed that Gallibacterium anatis isolates were different in some biochemical characteristics. The Gallibacterium anatis isolates had different levels of drug resistance on gentamicin, streptomycin, ampicillin, penicillin, doxycycline, norfloxacin, and sulfa drugs, and the resistance rate ranged from 44% to 96.9%, which had brought difficulty to the control of Gallibacterium anatis.5. It is the first successful reproduction of Gallibacterium anatis infection in SPF layer. SPF 112-day-old, 164-day-old and 184-day-old hens were experimentally infected with Gallibacterium anatis and /or Escherichia coli which were isolated from the hens with salpingitis. Results showed that, compared to uninfected control group, mean age at first egg of 112-day-old hens in Gallibacterium anatis and Gallibacterium anatis/ Escherichia coli infection group were delayed 7 and 13 days; egg production in 50 days after infection were decreased by 69.7% and 87.8%; and the rate of poor-quality eggs were increased by 70% and 75% respectively. However, there were no significant differences in egg production and clinical situation between hens infected with E.coli only and uninfected control. SPF 164-day-old hens infected with E.coli and Gallibacterium anatis/E.coli were dead, while the hens infected with Gallibacterium anatis only were healthy clinically, except for the egg production was dropped by 33%, the rate of the poor-quality eggs was increased by 23% compared to the control group. The Gallibacterium anatis could be detected and shed from the 112-day-old hens until 50 days after infection and from the 164-day-old and 184-day-old hens until 16 days after infection respectively. Salpingitis appeared in SPF hens infected with Gallibacterium anatis during 16 to 50 days experimental period. However, we didn't observe typical oviduct cysts. The results confirmed the pathogenicity of Gallibacterium anatis. The infection of Gallibacterium anatis had serious impacts on the age at first egg, egg production and egg quality. While the pathology damage was not obviously in short time. But E.coli infection could enhance the pathogenicity of Gallibacterium anatis. The cohabitation infection experiment of 184-day-old hens confirmed that Gallibacterium anatis could be transmitted laterally, and the whole flock got infected in two days. But no Gallibacterium anatis were isolated from the eggs layed from the SPF hens infected with Gallibacterium anatis, which indicated that Gallibacterium anatis was spread from flock to flock mainly by horizontal transmission through infected birds instead of vertical transmission by eggs.
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