The Intestinal Flora Is Required to Support Antibody Responses to Systemic Immunization in Infant and Germ Free Mice

Abstract

The presence of a complex and diverse intestinal flora is functionally important for regulating intestinal mucosal immune responses. However, the extent to which a balanced intestinal flora regulates systemic immune responses is still being defined. In order to specifically examine whether the acquisition of a less complex flora influences responses to immunization in the pre-weaning stages of life, we utilize a model in which infant mice acquire an intestinal flora from their mothers that has been altered by broad-spectrum antibiotics. In this model, pregnant dams are treated with a cocktail of antibiotics that alters both the density and microbial diversity of the intestinal flora. After challenge with a subcutaneous immunization, the antibiotic altered flora infant mice have lower antigen specific antibody titers compared to control age-matched mice. In a second model, we examined germ free (GF) mice to analyze how the complete lack of flora influences the ability to mount normal antibody responses following subcutaneous immunization. GF mice do not respond well to immunization and introduction of a normal flora into GF mice restores the capacity of these mice to respond. These results indicate that a gastrointestinal flora reduced in density and complexity at critical time points during development adversely impacts immune responses to systemic antigens.

Impact of probiotic supplements on microbiome diversity following antibiotic treatment of mice

“…Ampicillin (Abraxis Pharma), Streptomycin (Abraxis Pharma), and Clindamycin (Pharmacia) were provided into sterile drinking water at a final concentration of 1mg/ml, as previously described. 5 These antibiotics were chosen due to the broad spectrum capacity and well documented impacts on intestinal microbiota. 17 Animals were allowed to drink ad libitum for the duration of the experiment with water replacement at 3 day intervals.…”

“…[1][2][3] The host relies on these microbial denizens for nutrition, regulation of intestinal structures, immune system development, and protection from opportunistic pathogens. [4][5][6] The microbiome plays an essential role in maintaining the host health.…”

Maternal Antibiotic Treatment Impacts Development of the Neonatal Intestinal Microbiome and Antiviral Immunity

“…Although streptomycin is used less often, its activity mirrors gentamicin and is not absorbed from the intestine. This combination of antibiotics appreciably, but not completely, reduces aerobic and anaerobic Gram-positive and Gram-negative microbes in the GIT microbiota of adult mice within 3 d (25) and limits the possible expansion of vancomycin-resistant microbes. Pregnant mice were treated with this antibiotic mixture in their drinking water for 3-5 d prior to birth of a litter and throughout the duration of the suckling/preweaning period when vaccinia infection experiments were conducted (Fig.…”

“…Ceca were homogenized in Buffer ASL (QIAGEN), and DNA was isolated as above. The quantity (16S rRNA gene copies/mg stool or cecal content) of total and specific intestinal bacteria (Bacteroidetes, Firmicutes, Lactobacillus) was measured by quantitative real-time PCR (qPCR) using universal and group-specific 16S rRNA gene primers (25) and the QuantiTect SYBR Green PCR kit (QIAGEN). qPCR was performed on an ABI ViiA 7 realtime PCR system (Applied Biosystems, Foster City, CA).…”

“…We showed previously that maternal antibiotic treatment/treated (MAT) was associated with reduced Ab production following immunization of infant mice ,15 days of age (25). Our goals in this study were to characterize GIT dysbiosis in a model of maternal and perinatal antibiotic treatment and examine its impact on infant antiviral immunity during systemic vaccinia virus infection.…”