Intrapulmonary Delivery of CpG-ODN Microdroplets Provides Protection AgainstEscherichia coliSepticemia in Neonatal Broiler Chickens

Abstract

Synthetic oligodeoxynucleotides (ODN) containing unmethylated cytosine phosphodiester guanine (CpG) motifs (CpG-ODN) are effective immunostimulatory agents against a variety of viral, bacterial, and protozoan diseases in different animals including poultry. We have recently demonstrated that in ovo injection of CpG-ODN confers protection in neonatal chickens against bacterial septicemias. The objective of this study was to investigate the effectiveness of needle-free intrapulmonary (IPL) delivery of CpG-ODN microdroplets against Escherichia coli infection in neonatal chicks. In the present study, we used 880 chicks in total keeping 40 chicks per group. Chicks were delivered CpG-ODN or saline by IPL at the day 1 of hatch. Three days later, chicks were challenged with two doses (1 × 10 CFU, n = 20 or 1 × 10 CFU, n = 20) of E. coli. Chicks treated with CpG-ODN by the IPL route had significantly lower clinical signs and bacterial load compared to the group treated with saline ( P < 0.05). CpG-ODN-treated groups were significantly protected against E. coli septicemia. We observed dose- and exposure time-dependent immunoprotective effects of IPL CpG-ODN in chicks. We found that IPL delivery of CpG-ODN can induce protective immunity as early as 6 hr that remains effective at least until day 5 post-treatment. Moreover, there were no adverse effects of IPL delivery of CpG-ODN on growth or mortality up to 42 days of age. Based on these findings, it can be suggested that CpG-ODN delivery by IPL route can be a promising alternative to antibiotics for inducing protective immunity in chicks during the critical first week of neonatal life.

Mucosal delivery of CpG-ODN mimicking bacterial DNA via the intrapulmonary route induces systemic antimicrobial immune responses in neonatal chicks

“…Aliquots of bacteria were stored at −80 °C in 50% brain-heart infusion broth (Difco, Detroit, MI) supplemented with 25% (w/v) glycerol (VWR Scientific, Inc., Montreal, Quebec). Bacterial culture was prepared as previously described 27 . Briefly, bacteria used for the challenge were cultured on 5% Columbia sheep blood agar plates for 18-24 hr at 37 °C.…”

“…The E. coli animal challenge studies were carried out following the procedure as described previously 27 . Briefly, 2 d post-treatment, birds were challenged subcutaneously in the neck with either 1 × 10 5 or 1 × 10 6 cfu of E. coli.…”

“…Birds that received a clinical score of 2 were euthanized by cervical dislocation. At the end of the trial, each bird was given a cumulative clinical score (CCS) as a sum of daily clinical scores as previously described 7,27 . Chicks that were found dead or euthanized were subjected to immediate necropsy.…”

“…The temperature was maintained at 28-30 °C in the acrylic chamber during the administration of CpG-ODN or saline. The CpG-ODN delivery technique and the duration of nebulization is based on our previous published 27 and unpublished data to facilitate uniform distribution of CpG-ODN in chicks. www.nature.com/scientificreports www.nature.com/scientificreports/ E. coli challenge.…”

“…www.nature.com/scientificreports www.nature.com/scientificreports/ E. coli challenge. Two days post IPL delivery of CpG-ODN (day-2 post-hatch), birds were challenged with either 1 × 10 5 (n = 20/group) or 1 × 10 6 cfu (n = 20/group) of a virulent strain of E. Coli by subcutaneous injection in the neck, following the previosuly published challenge model 27 . Data were collected on mortality, clinical signs, pathology and bacterial isolations from the air sacs for 7 days following challenge with E. coli.…”