Immune correlates of protection against intracellular bacterial pathogens are largely thought to be cell-mediated, although a reasonable amount of data supports a role for antibody-mediated protection. To define a role for antibody-mediated immunity against an intracellular pathogen, Rhodococcus equi, that causes granulomatous pneumonia in horse foals, we devised and tested an experimental system relying solely on antibody-mediated protection against this host-specific etiologic agent. Immunity was induced by vaccinating pregnant mares 6 and 3 weeks prior to predicted parturition with a conjugate vaccine targeting the highly conserved microbial surface polysaccharide, poly-N-acetyl glucosamine (PNAG). We ascertained antibody was transferred to foals via colostrum, the only means for foals to acquire maternal antibody. Horses lack transplacental antibody transfer. Next, a randomized, controlled, blinded challenge was conducted by inoculating at ~4 weeks of age ~106 cfu of R. equi via intrabronchial challenge. Eleven of 12 (91%) foals born to immune mares did not develop clinical R. equi pneumonia, whereas 6 of 7 (86%) foals born to unvaccinated controls developed pneumonia (P = 0.0017). In a confirmatory passive immunization study, infusion of PNAG-hyperimmune plasma protected 100% of 5 foals against R. equi pneumonia whereas all 4 recipients of normal horse plasma developed clinical disease (P = 0.0079). Antibodies to PNAG mediated killing of extracellular and intracellular R. equi and other intracellular pathogens. Killing of intracellular organisms depended on antibody recognition of surface expression of PNAG on infected cells, along with complement deposition and PMN-assisted lysis of infected macrophages. Peripheral blood mononuclear cells from immune and protected foals released higher levels of interferon-γ in response to PNAG compared to controls, indicating vaccination also induced an antibody-dependent cellular release of this critical immune cytokine. Overall, antibody-mediated opsonic killing and interferon-γ release in response to PNAG may protect against diseases caused by intracellular bacterial pathogens.
There is currently no licensed vaccine that protects foals against Rhodococcus equi–induced pneumonia. Oral administration of live, virulent R. equi to neonatal foals has been demonstrated to protect against subsequent intrabronchial challenge with virulent R. equi. Electron beam (eBeam)-inactivated R. equi are structurally intact and have been demonstrated to be immunogenic when administered orally to neonatal foals. Thus, we investigated whether eBeam inactivated R. equi could protect foals against developing pneumonia after experimental infection with live, virulent R. equi. Foals (n = 8) were vaccinated by gavaging with eBeam-inactivated R. equi at ages 2, 7, and 14 days, or gavaged with equal volume of saline solution (n = 4), and subsequently infected intrabronchially with live, virulent R. equi at age 21 days. The proportion of vaccinated foals that developed pneumonia following challenge was similar among the vaccinated (7/8; 88%) and unvaccinated foals (3/4; 75%). This vaccination regimen did not appear to be strongly immunogenic in foals. Alternative dosing regimens or routes of administration need further investigation and may prove to be immunogenic and protective.
Background
The bacterium
Rhodococcus equi
can cause severe pneumonia in foals. The absence of a licensed vaccine and limited effectiveness of commercial
R. equi
hyperimmune plasma (RE‐HIP) create a great need for improved prevention of this disease.
Hypothesis
Plasma hyperimmune to the capsular polysaccharide poly‐
N
‐acetyl glucosamine (PNAG) would be significantly more effective than RE‐HIP at mediating complement deposition and opsonophagocytic killing (OPK) of
R. equi
.
Animals
Venipuncture was performed on 9 Quarter Horses.
Methods
The ability of the following plasma sources to mediate complement component 1 (C1) deposition onto either PNAG or
R. equi
was determined by ELISA: (1) PNAG hyperimmune plasma (PNAG‐HIP), (2) RE‐HIP, and (3) standard non‐hyperimmune commercial plasma (SP). For OPK, each plasma type was combined with
R. equi
, equine complement, and neutrophils isolated from horses (n = 9); after 4 hours, the number of
R. equi
in each well was determined by quantitative culture. Data were analyzed using linear mixed‐effects regression with significance set at
P
< .05.
Results
The PNAG‐HIP and RE‐HIP were able to deposit significantly (
P
< .05) more complement onto their respective targets than the other plasmas. The mean proportional survival of
R. equi
opsonized with PNAG‐HIP was significantly (
P
< .05) less (14.7%) than that for SP (51.1%) or RE‐HIP (42.2%).
Conclusions and Clinical Importance
Plasma hyperimmune to PNAG is superior to RE‐HIP for opsonizing and killing
R. equi
in vitro. Comparison of these 2 plasmas in field trials is warranted because of the reported incomplete effectiveness of RE‐HIP.
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