Eimeria tenella 5802 Hemorrhagic cecal coccidiosis Protective immunity against Eimeria infections strongly depends on the cellular immune responses involving T cells, natural killer cells (NK) and macrophages (Wang et al., 2009). Chickens the only host of Eimeria tenella (Wiki: Eimeria tenella). Eimeria tenella is a species of Eimeria that causes hemorrhagic cecal coccidiosis in young poultry. Monoxeous life cycle with definitive (only) host as chickens. Extremely host-specific. Acquired via fecal contamination of food and water (oral fecal route). Goes thru endogenous merogony in crypts of lieberkuhr (intestinal ceca of chicken) and gametogony in epithelial cells of small intestines. Fusion of microgamete and macrogamete forms unsporulated zygote which is released with feces of chicken. Zygote sporulates after 1-5 days and becomes infective. Diagnosis is based on finding oocysts in feces. No effective treatment but can reduce rate of infection via prophylates, anti-coccidial drugs and vaccination of baby chicks (Wiki: Eimeria tenella). Baboon Papio cynocephalus 9556 Bank vole Clethrionomys glareolus 447135 Bear Ursus americanus 9643 Birds Passeroidea 175121 Brown Trout Salmo trutta 8032 Buffalo Bison bison 9901 Carnivores Vulpes 9625 Cat Felis catus 9685 Catfishes Siluriformes 7995 Cattle Bos taurus 9913 Chicken Gallus gallus 9031 Chimpanzee Pan troglodytes 9598 chinchillas Chinchillidae 10150 Copper Pheasant Syrmaticus soemmerringii 9067 Deer Cervus elaphus 9860 Deer mouse Peromyscus maniculatus 10042 Dog Canis familiaris 9615 Ducks Anas 8835 Ferret Mustela putorius furo 9669 Fish Hyperotreti 117565 Gerbil Gerbillina 10045 Goat Capra hircus 9925 Gray wolf Canis lupus 9612 Guinea pig Cavia porcellus 10141 Hamster Mesocricetus auratus 10036 Horse Equus caballus 9796 Human Homo sapiens 9606 Macaque Macaca fascicularis 9541 Mongolian Gerbil Meriones unguiculatus 10047 Monkey Platyrrhini 9479 Mouse Mus musculus 10090 None None Parrot Psittacidae 9224 Pig Sus scrofa 9823 Rabbit Oryctolagus cuniculus 9986 Rainbow trout Oncorhynchus mykiss 8022 Rat Rattus 10114 Raven Corvus corax 56781 sei whale Balaenoptera borealis 9768 Sheep Ovis aries 9940 Squirrel Spermophilus richardsonii 37591 Tree shrew Tupaiidae 9393 Trouts, salmons & chars Salmoninae 504568 Turkey Meleagris gallopavo 9103 Vole Microtus ochrogaster 79684 Water buffalo Bubalus bubalis 391902 E. tenella DNA vaccine EtMIC2 VO_0011471 DNA vaccine Research pcDNA vector in ovo in ovo E. tenella microneme 2 (MIC2) DNA vaccine construction The EtMIC2 coding sequence was subcloned from EtMIC2-pGEX into the BamHI/HindIII sites of the pMal4c vector with a NH2-terminal maltose-binding protein tag, expressed in E. coli in TY broth (20 g/l tryptone, 10 g/l yeast extract, 10 g/l NaCl) containing 100 μg/ml ampicillin, the bacteria grown to OD600 = 0.5, induced with 1.0 mM isopropyl-β-d-thiogalactopyranoside for 3 h at 37 °C, collected by centrifugation, and disrupted by sonication on ice (Misonix, Farmingdale, NY). The EtMIC2 protein was isolated on an amylose affinity column (New England Biolabs, Beverly, MA) according to the manufacturer's instructions, digested with Factor Xa to release EtMIC2, and re-passed through the amylose column to remove maltose-binding protein (Ding et al., 2005). To assess anti-EtMIC2 antibody titers and protective immunity to coccidiosis following in ovo vaccination with the EtMIC2 gene, 75 fertile eggs were distributed into five groups (15/group) and non-injected or injected with 100 μl of sterile PBS, 50 μg/egg of the pcDNA empty vector, or 25 or 50 μg/egg of EtMIC2-pcDNA. To determine the effects of post-vaccination boosting with the EtMIC2 gene, 150 fertile eggs were distributed into 10 groups (15/group) and non-injected or injected with 100 μl of sterile PBS, 50 μg/egg of the pcDNA empty vector, or 25 or 50 μg/egg of EtMIC2-pcDNA (Ding et al., 2005). Birds vaccinated with Eimeria tenella microneme recombinant gene (EtMIC2) and encoded protein developed protective immunity against infection by E. tenella as assessed by significantly increased body weight gain and decreased fecal oocyst shedding compared with non-vaccinated controls. Vaccination with the EtMIC2 gene also led to protective immunity against infection by E. acervulina, but not E. maxima (Ding et al., 2005). At day 11 post-hatching, chickens were infected with 10,000 sporulated oocysts of E. tenella. Serum samples were collected at days 1, 10, and 17 post-infection (days 10, 21, and 28 post-hatching) and anti-EtMIC2 antibody titers determined by ELISA (Ding et al., 2005). E. tenella DNA vaccine pcDNA-TA4-IL-2 VO_0011449 DNA vaccine Research pcDNA3.1 [Ref1080:Song et al., 2009] Intramuscular injection (i.m.) Intramuscular injection (i.m.) E. tenella sporozoite TA4 surface antigen DNA vaccine construction PCR amplification of TA4 (without stop codon) was performed using the gene-specific primers with pET28a-TA4 as template. After purification, the amplified TA4 (without stop codon) was digested with BamHI and EcoRI and cloned into the BamHI/EcoRI sites of pcDNA3.1b. The recombinant plasmid pcDNA3.1b-TA4 (without stop codon) was confirmed by PCR amplification and endonuclease cleavage. Using the same method above, amplified chIL-2 product was ligated into pcDNA3.1b-TA4 (without stop codon) and pcDNA4.0c (Invitrogen) to construct pcDNA3.1b-TA4-IL2 and pcDNA4.0c-IL-2 (Song et al., 2009). At 14 days of age, chickens were randomly distributed into seven groups (30/group), group 1 negative control, group 2 positive control, groups 3, 4, 5 and 6 immunized with 25 μg, 50 μg, 100 μg and 200 μg of DNA vaccine, respectively and group 7 vector control. Experimental groups were immunized with corresponding doses of DNA vaccine pcDNA3.1b–TA4–IL-2 by leg intramuscular injection. Control group chickens were injected with sterile TE or 100 μg pcDNA3.1b at the same injection site to the experimental groups. A booster immunization was given by the same method as the first immunization 7 days later (Song et al., 2009). The immunization procedure of DNA vaccine pcDNA-TA4-IL-2 of Eimeria tenella, including route, dose, time of immunization and age of primary immunization of chicken, was optimized. results illustrated that 25 microg was the optimal dose and intramuscular injection was the most effective route to induce protective immunity in Chinese Yellow chickens (Song et al., 2009). At 28 days of age, the chickens were weighed one-by-one and challenged with 5 × 10^4 sporulated oocysts of E. tenella JS except the negative control group. The chickens were weighed one-by-one and slaughtered 7 days post challenge (Song et al., 2009). E. tenella DNA vaccine proIE VO_0011469 DNA vaccine Research proVAX Intramuscular injection (i.m.) Intramuscular injection (i.m.) Eimeria tenella antigen gene (3-1E) and chicken interferon gamma gene (ChIFN-gamma) DNA vaccine construction The Eimeria tenella antigen gene (3-1E) and chicken interferon gamma gene (ChIFN-gamma) were subcloned into the mammalian expression vector proVAX forming the plasmids proE and prol, and then linked by splicing overlap extension by polymerase chain reaction to construct the chimeric plasmid prolE; the chimeric protein (rlE) was expressed in Escherichia coli harboring the constructed plasmid pGEX/IE (Xu et al., 2006). Broiler Broilers were administered two intramuscular injections with the constructed DNA vaccines (50 microg); in the protein booster groups 100 microg of the rlE were given following the proIE prime (Xu et al., 2006). The Eimeria tenella antigen gene (3-1E) and chicken interferon gamma gene (ChIFN-gamma) were subcloned into the mammalian expression vector proVAX forming the plasmids proE and prol. After challenge the proIE-vaccinated chickens showed protective immunity as demonstrated by significantly reduced oocyst shedding (Xu et al., 2006). One week following the booster immunization, all chickens except for the unchallenged control group were orally challenged with 5 x 10^4 sporulated E. tenella GS oocysts (Xu et al., 2006). E. tenella GX3262 protein vaccine VO_0011470 Subunit vaccine Research Subcutaneous injection Freund complete adjuvant Subcutaneous injection E. tenella sporozoite antigen GX3262 Recombinant protein preparation To produce the 3-galactosidase-GX3262 fusion protein (Pgal-GX3262), recombinant E. coli cells. grown overnight in Luria-Bertani broth with ampicillin and induced with IPTG, were lysed by sonication and centrifuged, and the insoluble cell pellet was washed extensively with PBS (Miller et al., 1989). Hubbard broiler The basic experimental design involved immunizing birds subcutaneously or orally with partially purified recombinant antigen preparations, heatkilled bacterins. live E. coli cells, or control materials (Miller et al., 1989). A cDNA clone derived from sporulated oocysts of Eimeria tenella and encoding the expression product GX3262 was identified using a monoclonal antibody raised against Eimeria acervulina sporozoites. After challenge with an experimental E. tenella infection, the greatest degree of protection in immunized broiler chickens was seen after only a single immunization of 2-day-old birds with a live recombinant E. coli preparation (Miller et al., 1989). Broilers were challenged with E. tendell/i oocysts; and then evaluated for cecal lesions (Miller et al., 1989). E. tenella vaccine rBCG pMV261-Rho VO_0011472 Recombinant vector vaccine Research BCG Intravenous injection (i.v.) Intravenous injection (i.v.) Eimeria tenella rhomboid gene (Rho) Recombinant vector construction The rhomboid gene segment (GenBank accession no. DQ323509) was generated from the cDNA of E. tenella strains by PCR with primers QF (5’-CTGACTGCAGATGTCGGACATCGAATCCCAGAG-3′) containing PstI restriction site (underlined) and QR (5’-GACTATCGATTTATGCGCATCCCATGGGCAAAGG-3’) containing ClaI restriction site (underlined), and cloned into the pMD18-T vector. The 774-bp rhomboid gene fragment was subcloned into the PstI/ClaI sites of pMV261. The plasmid pMV261-Rho was electrotransfected into BCG and selected by kanamycin (Sigma) (Wang et al., 2009). Chickens were randomly assigned to 4 groups (40 birds in each group) and immunized intranasal (i.n.) with 10^6 CFU/200 μl of BCG or rBCG strains expressing rhomboid protein, respectivelly. Negative control was immunized i.n. with 200 μl PBS only. 2 weeks after the first vaccination, a second immunization was carried out with the same amounts of each sample (Wang et al., 2009). All the recombinant BCG immunized chickens developed specific immune responses, and there was a significant increases of the percentages of CD4(+) and CD8(+) cells compared to the control (P<0.05). Challenge experiments demonstrated that the two rBCG strains could provide significant protection against E. tenella challenge (Wang et al., 2009). All groups were challenged with 3 × 10^4 sporulated oocysts of E. tenella per bird 2 weeks after the final immunization (Wang et al., 2009). E. tenella vaccine rBCG pMV361-Rho VO_0011448 Recombinant vector vaccine Research BCG Intranasal Intranasal E. tenella rhomboid gene (Rho) Recombinant vector construction The rhomboid gene segment (GenBank accession no. DQ323509) was generated from the cDNA of E. tenella strains by PCR with primers QF (5’-CTGACTGCAGATGTCGGACATCGAATCCCAGAG-3′) containing PstI restriction site (underlined) and QR (5’-GACTATCGATTTATGCGCATCCCATGGGCAAAGG-3’) containing ClaI restriction site (underlined), and cloned into the pMD18-T vector. The 774-bp rhomboid gene fragment was subcloned into the PvuII/ClaI site of pMV361. The plasmid pMV361-Rho was electrotransfected into BCG and selected by kanamycin (Sigma) (Wang et al., 2009). Chickens were randomly assigned to 4 groups (40 birds in each group) and immunized intranasal (i.n.) with 10^6 CFU/200 μl of BCG or rBCG strains expressing rhomboid protein, respectivelly. Negative control was immunized i.n. with 200 μl PBS only. 2 weeks after the first vaccination, a second immunization was carried out with the same amounts of each sample (Wang et al., 2009). All the recombinant BCG immunized chickens developed specific immune responses, and there was a significant increases of the percentages of CD4(+) and CD8(+) cells compared to the control (P<0.05). Challenge experiments demonstrated that the two rBCG strains could provide significant protection against E. tenella challenge (Wang et al., 2009). All groups were challenged with 3 × 10^4 sporulated oocysts of E. tenella per bird 2 weeks after the final immunization (Wang et al., 2009). Livacox Livacox BIOPHARM VO_0001177 Live, attenuated vaccine Clinical trial NA Precocious and egg-passaged lines(Williams, 2002) NA 3-1E Eimeria tenella VO_0011066 126362934 CDD:320770 CDD:238085 5802 ? sporozoite antigen 3-1E 3.96 17761.05 237 Profilin binds actin monomers, membrane polyphosphoinositides such as PI(4,5)P2, and poly-L-proline. Profilin can inhibit actin polymerization into F-actin by binding to monomeric actin (G-actin) and terminal F-actin subunits, but - as a regulator of the...; cl00123 >ABO10495.1 sporozoite antigen 3-1E, partial [Eimeria tenella] MGEEADTQAWDTSVKEWLVDTGKVYAGGIASIADGCRLFGAAIDNGEDAWSQLVKTGYQIEVLQEDGSST QEDCDEAETLRQAIVDGRAPNGVYIGGIKYKLAEVKRDFTYNDQNYDVAILGKNKGGGFLIKTPNDNVVI ALYDEEKEQNKADALTTALAFAEYLYQGGF Protective antigen The Eimeria tenella antigen gene (3-1E) and chicken interferon gamma gene (ChIFN-gamma) were subcloned into the mammalian expression vector proVAX forming the plasmids proE and prol. After challenge the proIE-vaccinated chickens showed protective immunity as demonstrated by significantly reduced oocyst shedding [Ref1081:Xu et al., 2006]. Protective antigen GX3262 Eimeria tenella VO_0011067 102304 5802 ? sporozoite antigen 9.06 11721.58 176 >pir||A60111 sporozoite antigen - Eimeria tenella (fragment) LLLLLLLLLQGAECLLRSSKLALEALLEGARVAATRGLLLVESSKDTVLRSIPHTQEKLAQAYSSFLRGY QGAAAGRSLGYGAPAAAYGQQQQPSSYGAPPASSQQPSGFFW Protective antigen A cDNA clone derived from sporulated oocysts of Eimeria tenella and encoding the expression product GX3262 was identified using a monoclonal antibody raised against Eimeria acervulina sporozoites. After challenge with an experimental E. tenella infection, the greatest degree of protection in immunized broiler chickens was seen after only a single immunization of 2-day-old birds with a live recombinant E. coli preparation [Ref1082:Miller et al., 1989]. Protective antigen MIC2 Eimeria tenella VO_0011063 225579627 CDD:115333 5802 ? microneme 2 4.13 32481.77 390 Microneme protein Etmic-2; pfam06670 >ACN93990.1 microneme 2 [Eimeria tenella] MARALSLVALGLLFSLPPSSAVRTRVPGEDSFSPESGVLSGTDAPERRPIVPGLVEGNCGRLTVRNGPSV DETIKVTSAGWTKSERDFIVSLVADETRKVVQLRESEGASGASGPGPAPAEKPPSGQGSAEEAPKGEGGQ EKPSVPLIAVRIHGSGGDKGESAPQSAVLLYGNDESEPTEVPLETAAGPTTPLMVLITQQNPKEVEVRVL AWISTDATTGKGSWKENSVVVGSSLSGRDLTVNLGDCGPSSLRVYGSASADLVTVKEGMCEADDPELIAL TRPHTSAASPLPAEEGDVAQDAQQSAGAQQEAEAQEVGEPQQEAAAAEQGSSAAESDTQQSS Protective antigen Birds vaccinated with Eimeria tenella microneme recombinant gene (EtMIC2) and encoded protein developed protective immunity against infection by E. tenella as assessed by significantly increased body weight gain and decreased fecal oocyst shedding compared with non-vaccinated controls. Vaccination with the EtMIC2 gene also led to protective immunity against infection by E. acervulina, but not E. maxima [Ref1078:Ding et al., 2005]. Protective antigen Rho Eimeria tenella VO_0011064 84043169 CDD:304416 5802 ? rhomboid-like protein 8.07 26568.3 314 Rhomboid family; cl21536 >ABC50099.1 rhomboid-like protein [Eimeria tenella] MSDIESQRVLGAGIPRTGRLIDVAFPNISLDKSIVVITLAQIAVYIMSCAMSSAFEPTERVLYQLGATYG PGIRHLQAWRLIMPVFLHVGIVHLLFNVLFILHMGLDKEIKYGRTNFLILYFSSALIGNMFTVLMRPCSL AVGASTAGFGLVGSILAEILIVWHKLDERTRNMYTLDMTVFGALMILLSYGQTVDIWGHLGGFVCGFAVT CEFNKNIRDLPHWFDLAKNGTRTICFGVVALTLARVFLGLPLPMGCA Protective antigen Two recombinant Mycobacterium bovis BCG (rBCG) strains carrying the Eimeria tenella rhomboid gene (Rho) delivered by extrachromosomal vector pMV261 and integrative vector pMV361 were evaluated for their ability to protect chickens against E. tenella challenge. Chickens were immunized intranasal with BCG, rBCG pMV261-Rho, or rBCG pMV361-Rho twice at a 2-week interval. All the recombinant BCG immunized chickens developed specific immune responses, and there was a significant increases of the percentages of CD4(+) and CD8(+) cells compared to the control (P<0.05). Challenge experiments demonstrated that the two rBCG strains could provide significant protection against E. tenella challenge [Ref1079:Wang et al., 2009]. Protective antigen TA4 Eimeria tenella VO_0011065 84783094 CDD:287966 5802 ? TA4 antigen protein 4.47 22659.37 293 Sporozoite TA4 surface antigen; pfam11054 >ABC61818.1 TA4 antigen protein, partial [Eimeria tenella] DYPTAVTLDCKEAMNKMRKAAGLPAFEDAVGDTFVLPAYSHEESRAAPVAETLWKTEICPKVLGGGRSRN VTEAVRLTGNFAYYPVTDGKKECSDAVEYWKGGLSQFNDTIPPTFQALNDPVVYNDRAVSFVALYNPKTS PVVSCVLLQCPNAGVGGRRLAAGTTDAVICLTNPAPLEARSQPFDDEQWKKIVDSLSLSEEEEEKGGVSP VVPSVALISAAVISAFALF Protective antigen The immunization procedure of DNA vaccine pcDNA-TA4-IL-2 of Eimeria tenella, including route, dose, time of immunization and age of primary immunization of chicken, was optimized. results illustrated that 25 microg was the optimal dose and intramuscular injection was the most effective route to induce protective immunity in Chinese Yellow chickens [Ref1080:Song et al., 2009]. Protective antigen Ding et al., 2005 journal Ding X, Lillehoj HS, Dalloul RA, Min W, Sato T, Yasuda A, Lillehoj EP 2005 23 28 3733-3740 Vaccine Lai et al., 2011 journal Lai L, Bumstead J, Liu Y, Garnett J, Campanero-Rhodes MA, Blake DP, Palma AS, Chai W, Ferguson DJ, Simpson P, Feizi T, Tomley FM, Matthews S 2011 7 10 e1002296 PLoS pathogens Miller et al., 1989 journal Miller GA, Bhogal BS, McCandliss R, Strausberg RL, Jessee EJ, Anderson AC, Fuchs CK, Nagle J, Likel MH, Strasser JM 1989 57 7 2014-2020 Infection and immunity Song et al., 2009 journal Song X, Xu L, Yan R, Huang X, Shah MA, Li X 2009 159 1 30-36 Veterinary parasitology Wang et al., 2009 journal Wang Q, Li J, Zhang X, Liu Q, Liu C, Ma G, Cao L, Gong P, Cai Y, Zhang G 2009 160 3-4 198-203 Veterinary parasitology Wiki: Eimeria tenella website http://en.wikipedia.org/wiki/Eimeria_tenella Williams, 2002 journal Williams RB 2002 46 4 775-802 Avian diseases Xu et al., 2006 journal Xu SZ, Chen T, Wang M 2006 50 4 579-585 Avian diseases