A LigA Three-Domain Region Protects Hamsters from Lethal Infection by Leptospira interrogans

Abstract

The leptospiral LigA protein consists of 13 bacterial immunoglobulin-like (Big) domains and is the only purified recombinant subunit vaccine that has been demonstrated to protect against lethal challenge by a clinical isolate of Leptospira interrogans in the hamster model of leptospirosis. We determined the minimum number and location of LigA domains required for immunoprotection. Immunization with domains 11 and 12 was found to be required but insufficient for protection. Inclusion of a third domain, either 10 or 13, was required for 100% survival after intraperitoneal challenge with Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130. As in previous studies, survivors had renal colonization; here, we quantitated the leptospiral burden by qPCR to be 1.2×103 to 8×105 copies of leptospiral DNA per microgram of kidney DNA. Although renal histopathology in survivors revealed tubulointerstitial changes indicating an inflammatory response to the infection, blood chemistry analysis indicated that renal function was normal. These studies define the Big domains of LigA that account for its vaccine efficacy and highlight the need for additional strategies to achieve sterilizing immunity to protect the mammalian host from leptospiral infection and its consequences.

Oral Immunization with Escherichia coli Expressing a Lipidated Form of LigA Protects Hamsters against Challenge with Leptospira interrogans Serovar Copenhageni

“…Despite the low challenge doses used in this study, culture and qPCR results of kidney tissue from all animals, LigA7-13 immunized or not, were positive for leptospires. This result is consistent with previous studies showing that LigA immunization afforded protection from lethality but not renal infection (26,30). It is worth noting that our qPCR method involved TaqMan quantitation of the number of lipL32 gene copies per gram of kidney tissue, a method that has previously been applied to blood and urine (38,43).…”

“…As in an earlier study, we found that an endpoint criterion of weight loss of Ն10% of the animal's maximum body weight was extremely useful, because a decrease in body weight was the earliest observable sign of clinical leptospirosis (30). Further support for the importance of body weight in animal models of leptospirosis comes from our finding that the weight of animals that survived challenge was also useful as an indication of underlying renal health.…”

“…Effects of LigA7-13 oral immunization on renal infection. As in previous studies examining immunoprotection with LigA (26,30), oral immunization with LigA7-13 improved survival but did not prevent renal infection. In this study, kidney cultures of all hamsters were positive for leptospires, both from survivors and from those that met endpoint criteria early.…”

“…Quantification of LigA7-13 concentrations was performed by densitometry of Western blots of serial dilutions of E. coli and purified LigA7-13 protein with Image J (36). Purified, recombinant LigA7-13 protein was prepared as described previously (30).…”

“…Several groups have reported that immunization with the LigA unique region, consisting of domains 7 to 13 (LigA7-13), confers protection from intraperitoneal (i.p.) challenge in the hamster model of leptospirosis (26)(27)(28)(29)(30). In this study, we examined whether oral immunization with immunoglobulin-like do-mains 7 to 13 of LigA results in an immune response that is protective against intradermal (i.d.)…”