Growth inhibition of Staphylococcus aureus and escherichia coli strains by neutralizing IgY antibodies from ostrich egg yolk.

Ostrich raising around the world have some key factors and farming profit depend largely on information and ability of farmers to rear these animals. Non fertilized eggs from ostriches are discharged in the reproduction season. Staphylococcus aureus and Escherichia coli are microorganisms involved in animal and human diseases. In order to optimize the use of sub products of ostrich raising, non fertilized eggs of four selected birds were utilized for development of polyclonal IgY antibodies. The birds were immunized (200ug/animal) with purified recombinant staphylococcal enterotoxin C (recSEC) and synthetic recRAP, both derived from S. aureus, and recBFPA and recEspB involved in E. coli pathogenicity, diluted in FCA injected in the braquial muscle. Two subsequent immunization steps with 21 days intervals were repeated in 0,85% saline in FIA. Blood and eggs samples were collected before and after immunization steps. Egg yolk immunoglobulins were purified by precipitation with 19% sodium sulfate and 20% ammonium sulphate methodologies. Purified IgY 50µL aliquots were incubated in 850µL BHI broth containing 50µL inoculums of five strains of S. aureus and five strains of E.coli during four hours at 37°C. Growth inhibition was evaluated followed by photometry reading (DO550nm). Egg yolk IgY preparation from hiperimmunized birds contained antibodies that inhibited significantly (p<0,05) growth of strains tested. Potential use of ostrich IgY polyclonal antibodies as a diagnostic and therapeutic tool is proposed for diseased animals.

Development of process to produce polyvalent IgY antibodies anti-African snake venom.

Polyvalent anti-Bitis and anti-Naja antivenom IgY antibodies were prepared using B. arietans, B. nasicornis, B. rhinoceros, N. melanoleuca, and N. mossambica venoms to immunize chickens. Blood and eggs were collected before and during the 10-month immunization period; the sera and yolk extracts were then prepared and assayed for the presence of antivenom antibodies by ELISA and Western blot methods. ELISA Antivenom antibody titers, referred to as U-ELISA/ml of serum or egg yolk extracts, absent in pre-immunization sera or yolk, increased sharply during the 4 weeks after immunization, reaching a plateau thereafter. Yolk extracts with high antivenom titers, as detected by ELISA were used to isolate and purify IgY. Purified IgY preparations recognized venom protein bands from 10 to 20 kDa to 60 and 70 kDa, as shown by Western blot. Recovery of antivenom antibodies from the whole yolk was over 80%. Final preparations exhibited high antivenom activity (>100,000 U-ELISA/ml) as well as efficacy in neutralizing venom lethality (1,440 microg of IgY neutralize 62.2 LD(50) of venom), and were free of toxic products, pyrogen or bacterial and fungal contaminations.

The complementarity-determining region sequences in IgY antivenom hypervariable regions.

The data presented in this article are related to the research article entitled "Development of IgY antibodies against anti-snake toxins endowed with highly lethal neutralizing activity" (da Rocha et al., 2017) [1]. Complementarity-determining region (CDR) sequences are variable antibody (Ab) sequences that respond with specificity, duration and strength to identify and bind to antigen (Ag) epitopes. B lymphocytes isolated from hens immunized with Bitis arietans (Ba) and anti-Crotalus durissus terrificus (Cdt) venoms and expressing high specificity, affinity and toxicity neutralizing antibody titers were used as DNA sources. The VLF1, CDR1, CDR2, VLR1 and CDR3 sequences were validated by BLASTp, and values corresponding to IgY VL and VH anti-Ba or anti-Cdt venoms were identified, registered [Gallus gallus IgY Fv Light chain (GU815099)/Gallus gallus IgY Fv Heavy chain (GU815098)] and used for molecular modeling of IgY scFv anti-Ba. The resulting CDR1, CDR2 and CDR3 sequences were combined to construct the three - dimensional structure of the Ab paratope.

Development of IgY antibodies against anti-snake toxins endowed with highly lethal neutralizing activity.

Snakebite envenoming is a major neglected disease related to poverty in developing countries. Treatment involves the administration of a specific antivenom serum and auxiliary therapies, if necessary. The improvement of antibodies is of great importance for the technological advancement of antivenom therapy and to reduce the morbidity and mortality associated with this medical burden. In the present study, adult hens were immunized nine times with 20µg of B. arietans or C. d. terrificus venoms at three-week intervals between immunizations. Developing antibodies presented increasing avidity and affinity to antigenic toxin epitopes along immunization, attaining a plateau after the seventh immunization. Pooled egg yolk-purified IgY antivenom antibodies, subjected to in vitro-in vivo lethality assay using Swiss adult mice, exhibited potent venom lethal neutralizing activity. Taken together, chickens under the described immunization schedule were considered alternative candidates for antivenom production. Lower maintenance costs, a simple antibody manufacturing process and immunization suffering restrictions are additional advantages.