Abstract
The live attenuated vaccine strain, SG9R, has been used against fowl typhoid worldwide, but it can revert to the pathogenic smooth strain owing to single nucleotide changes such as nonsense mutations in the rfaJ gene. As SG9R possesses an intact Salmonella plasmid with virulence genes, it exhibits dormant pathogenicity and can cause fowl typhoid in young chicks and stressed or immunocompromised brown egg-laying hens. To tackle these issues, we knocked out the rfaJ gene of SG9R (named Safe-9R) to eliminate the reversion risk and generated detoxified strains of Safe-9R by knocking out lpxL, lpxM, pagP, and phoP/phoQ genes to attenuate the virulence. Among the knockout strains, live ΔlpxL- (Dtx-9RL) and ΔlpxM-9R (Dtx-9RM) strains induced remarkably less expression of inflammatory cytokines in chicken macrophage cells, and oil emulsion (OE) Dtx-9RL did not cause body weight loss in chicks. Live Dtx-9RM exhibited efficacy against field strain challenge in one week without any bacterial re-isolation, while the un-detoxified strains showed the development of severe liver lesions and re-isolation of challenged strains. Thus, SG9R was optimally detoxified by knockout of lpxL and lpxM, and Dtx-9RL and Dtx-9RM might be applicable as OE and live vaccines, respectively, to prevent fowl typhoid irrespective of the age of chickens.
Highlights
SG9R is a live attenuated vaccine strain that has long been inoculated subcutaneously to provide protection against fowl typhoid (FT) caused by Salmonella enterica serovar Gallinarum biovar Gallinarum (SG) and the food poisoning risk posed by S. enterica serovar Enteritidis (SE) infection in egg-laying hens in several countries [1,2]
To test the pathogenicity of the detoxified strains, five 1 week-old chicks were assigned to the same scheme of groups and inoculated with the same dose as above
The residual pathogenicity of live detoxifified and un-detoxifified strains was compared in the PPEEMMmmooddeel.l.NNoolelseisoinosnosromr imldilldeslieosniosnins itnhethlievelirvwererweeorbesoerbvsedrvwedithwtihthe athdemaindmistirnaitsitornatoiofnDotxf -D9RtxL-9aRnLdaDntdx-D9tRxM-9,RrMes,preecstpiveectlyiv,ealny,datnhdeythweyerwe enreegnaetigvaetifvoer fboarcbtearcitaelrriaelriseo-ilsaotiloanti.oHn.owHeovwere,vSear,feS-a9fRe-a9nRdaSnGd9SRGc9aRusceadumseoddmeroadteertoatseetvoerseevleesrieonlessaionndswanerde wpoesrieptiovseitfiovrebfaocrtberaicatlerreia-ilsroel-aitsioolnat(i4o/n5(a4n/d5 3a/n5d), 3r/es5p),ercetisvpeelcyti(vTealbyl(eT2a)b.le 2)
Summary
SG9R is a live attenuated vaccine strain that has long been inoculated subcutaneously to provide protection against fowl typhoid (FT) caused by Salmonella enterica serovar Gallinarum biovar Gallinarum (SG) and the food poisoning risk posed by S. enterica serovar Enteritidis (SE) infection in egg-laying hens in several countries [1,2]. SG9R was transformed into a rough strain by a nonsense mutation in the rfaJ gene [3]. SG9R has the potential to restore pathogenicity, and has residual pathogenicity of its own. It can be transmitted vertically, and mortality and re-isolation can occur if there is immune suppression by insufficient nutrition or infection by immunosuppressive pathogens [3,6]. As SG9R inoculation decreases the growth rate in young chicks, SG9R vaccination has usually been recommended after 6 weeks of age, resulting in the lack of implementation of proper measures to protect against FT during the most susceptible period [7,8]
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