animal mucosal delivery vaccines

487Vet. Res. 37 (2006) 487510 INRA, EDP Sciences, 2006DOI: 10.1051/vetres:2006012

Review article

Mucosal delivery of vaccines in domestic animals

Volker GERDTS*, George K. MUTWIRI, Suresh K. TIKOO, Lorne A. BABIUK

Vaccine and Infectious Disease Organization (VIDO), University of Saskatchewan, 120 Veterinary Rd., Saskatoon, S7N 5E3, Canada

(Received 4 May 2005; accepted 11 October 2005)

Abstract Mucosal vaccination is proving to be one of the greatest challenges in modern vaccinedevelopment. Although highly beneficial for achieving protective immunity, the induction ofmucosal immunity, especially in the gastro-intestinal tract, still remains a difficult task. As a result,only very few mucosal vaccines are commercially available for domestic animals. Here, wecritically review various strategies for mucosal delivery of vaccines in domestic animals. Thisincludes live bacterial and viral vectors, particulate delivery-systems such as polymers, alginate,polyphosphazenes, immune stimulating complex and liposomes, and receptor mediated-targetingstrategies to the mucosal tissues. The most commonly used routes of immunization, strategies fordelivering the antigen to the mucosal surfaces, and future prospects in the development of mucosalvaccines are discussed.

mucosal / vaccine / livestock / animals / review

Table of contents

1. Introduction...................................................................................................................................... 4882. Rationale for mucosal vaccination................................................................................................... 4883. General considerations for mucosal vaccines .................................................................................. 4884. Types of vaccines for mucosal delivery .......................................................................................... 4905. Routes of immunization................................................................................................................... 4916. Strategies for mucosal delivery........................................................................................................ 491

6.1. Live vectors ............................................................................................................................. 4916.1.1. Bacterial vectors .......................................................................................................... 4926.1.2. Viral vectors................................................................................................................. 494

6.2. Particulate delivery systems.................................................................................................... 4966.2.1. Microparticles .............................................................................................................. 496

6.2.1.1. Polymers ........................................................................................................ 4976.2.1.2. Alginate.......................................................................................................... 4986.2.1.3. Polyphosphazenes.......................................................................................... 4986.2.1.4. Other biodegradable polymers....................................................................... 498

6.2.2. ISCOM......................................................................................................................... 4986.2.3. Liposomes .................................................................................................................... 499

* Corresponding author: [email protected]

Article published by EDP Sciences and available at http://www.edpsciences.org/vetres or http://dx.doi.org/10.1051/vetres:2006012

488 V. Gerdts et al.

6.3. Other strategies to enhance vaccine uptake .............................................................................5006.3.1. Lectin-mediated targeting.............................................................................................5006.3.2. Antibody-mediated targeting........................................................................................5006.3.3. Receptor-mediated targeting ........................................................................................500

7. Conclusion .......................................................................................................................................501

1. INTRODUCTIONInfectious diseases remain the major cause

of death and economic losses in animals.One way to reduce this is by vaccination.Although immunization has had a greatimpact on the economics of livestock pro-duction and on animal suffering, todaysvaccines produced by conventional meansare less than optimal in many respectsincluding virulence, safety and efficacy.Moreover, there is a need to develop newvaccines for new emerging diseases. Thus,novel vaccines and alternative routes ofimmunization are needed to meet futurechallenges in the control of human and live-stock diseases. Here we will review currentstrategies for mucosal vaccination and recentprogress in this field.

2. RATIONALE FOR MUCOSAL VACCINATIONThe primary reason for using a mucosal

route of immunization is that most infec-tions affect or initiate the infectious processat the mucosal surfaces, and that in theseinfections, mucosal application of a vaccineis often required to induce a protective immuneresponse [69]. Prime examples of suchinfections include gastrointestinal infectionswith enterotoxigenic E. coli (ETEC), rota-virus or calicivirus, and respiratory infectionswith Mycoplasma, influenza virus or respi-ratory syncytial virus. For most of theseinfections the induction of local immunityat the site of infection is crucial for optimalprotection. The induction of secretory IgA(S-IgA) represents the main effector mech-anism of the local adaptive immune responseand thus, represents the primary goal formost mucosal vaccines. In addition to S-IgAother immunoglobulins, such as IgG aretransudated across the mucosal surface.

This is especially important for respiratoryand genital infections since the transudationof antibodies is more easily facilitated atthese mucosal surfaces than for example inthe intestinal tract [69]. In addition to humoralimmunity, the induction of cytotoxic T cells(CTL) represents another important goal ofmucosal vaccines. However, to date onlyvery few experimental vaccines have beendemonstrated to induce CTL so far, most ofthese vaccines being live attenuated vac-cines. Interestingly, it was recently demon-strated that transcutaneous immunizationresulted in the induction of mucosal CTL inmice [14]. More research will be necessaryto confirm this observation in domestic ani-mals. Other reasons for using mucosalroutes of immunization include the practi-cability of administering the vaccine with-out injections, and therefore the reductionof injection site reactions, and the possibil-ity of vaccination at a very early age of lifein the presence of passively acquired mater-nal antibodies (Tab. I) [15].

3. GENERAL CONSIDERATIONS FOR MUCOSAL VACCINES

Immunity at the mucosal surfaces is medi-ated by the mucosa-associated lymphoid

Table I. Advantages of mucosal routes of immu-nization.

Induces protective immunity at the site of infectionInduces both systemic and mucosal immunityEffective in the presence of maternal antibodiesNo injection site reaction, no needles requiredReadily administered (i.e. oral vaccines combined with feed)

Mucosal vaccines for domestic animals 489

tissues (MALT), which represent the largestimmune compartment within the body. Themain functions of the MALT are to (i) pro-tect the mucosal membranes against infec-tion and colonization with pathogenic micro-organisms, (ii) to tolerate antigens derivedfrom ingested food, airborne matter and com-mensal microorganisms, and (iii) to preventthe development of any potentially harmfulimmune responses against these antigens incase they breach the mucosal lining [69]. Toachieve these functions, the MALT areequipped with highly specific mechanisms,which are discussed in greater detail through-out this special issue of Veterinary Research.However, certain mechanisms are of rele-vance for the development of mucosal vac-cines. For example, it is clear that an earlyinterplay between innate and adaptiveimmunity is essential for effective immu-nity against invading pathogens [68]. Infact, it is the first contact between pathogenand innate immune system, for examplethrough recognition via Toll like receptors(TLR), complement receptors (CR), C-typelectins, or nucleotide-binding oligomeriza-tion domain (NOD) receptors NOD 1 orNOD 2, that determines the outcome of theimmune response [54, 98, 127]. Pattern rec-ognition molecules (PRM) for these highlyconserved pathogen-associated molecularpatterns (PAMP) are found on various typesof immune cells including antigen-present-ing cells, lymphocytes, and epithelial cellswithin the mucosal tissues. Of specialimportance, however, are mucosal den-dritic cells (DC), which express a wide vari-ety of PRM [30, 92, 99]. Signaling throughthese receptors leads to recruitment andactivation of DC, which then can eithersample antigen directly from the externalsurfaces or receive it from highly special-ized M cells [78, 96, 115, 131, 132]. Withinthe lymphoid follicle the matured DC thenpresent the antigen to lymphocytes to inducea mucosal immune response. Depending onthe initial innate stimulus, the DC canimprint the effector cells to selectivelyhome back to certain mucosal sites and shiftthe type of the immune response to a either

Th1 or Th2 type of immune response [30,99]. Interestingly, the ability of intestinalDC to polarize the immune response is con-ditioned by IL-10 and IL-6 secreting epi-thelial cells. In the intestine, this typicallyresults in promotion of a Th2-type of animmune response. Thus, mucosal DC areessential in the early decision making proc-ess at the mucosa and in controlling home-ostasis versus inflammation [132]. Althoughnot demonstrated in domestic animals yet,it is clear that the stimulation of DC by cer-tain components of the vaccine (adjuvant)represents an important issue to consider forfuture vaccine formulations (Tab. II).

Following their priming in lymphoid tis-sues, effector cells leave the site of induc-tion and migrate to the effector site, whichis generally the lamina propria at the mucosalsurfaces. Effector cell trafficking is crucialfor the communication among the variouscompartments of the mucosal immune sys-tem leading to the concept of the commonmucosal immune system. However, it isclear that beyond the common level indeedthe mucosal immune system is highly com-partimentalized. In fact, some of the mucosalcompartments favor the trafficking of effec-tor cells from some inductive sites to certain

Table II. Characteristics of an ideal mucosalvaccine.

Safe, no side effects in adult and newborn animalsStable in gastrointestinal microenvironmentsReadily dissolved and administered (spray, injector, oral), small volumeCross mucosal barrierRetain antigen at the mucosal surface by targeting to mucosal epithelial cellsStimulate both innate and adaptive immunity at the mucosa

Contain delivery-systems that greatly enhance vaccine uptakeEnsure homing of effector cells to the site of infectionInduce long term immunity


effector sites. Cell trafficking between com-partments is facilitated by a complex net-work of interactions mediated by mucosaladdressins, integrins, and chemokines (CC)allowing the tissue-specific migration ofimmune cells from the inductive to theeffector site [85]. Chemokine receptors (CCR)and mucosal homing molecules, such as the47-integrin, are found on effector lym-phocytes, and in fact the expression of thesereceptors is controlled by the imprinting bymucosal DC. For example, in mice only DCderived from Peyers patches and mesentericlymph nodes were able to induce expressionof 47+ and CCR9+ on CD8+ T cells lead-ing to site specific homing of these effectorcells to the small intestinal lamina propria[74, 109]. CCR9, the receptor for the thy-mus-expressed chemokine (TECK, CCL25), is expressed on effector B and T cellsand mediates the selective migration ofthese effector cells to the small intestine inhumans and mice [19, 87, 120]. Interestingly,CCR9/ knockout mice display reducednumbers of plasma cells in the small intes-tinal lamina propria and impaired ability torespond to oral vaccination [119]. Thus,chemokines and their receptors are impor-tant molecules in linking inductive siteswith particular effector sites (compart-mentalization of the mucosal immune sys-tem). As a result, certain routes are morefavorable for inducing immunity at thedesired effector site (Tab. III). This knowl-edge should be taken into consideration forthe design of every novel type of vaccinethat is to be used for mucosal immunization.

4. TYPES OF VACCINES FOR MUCOSAL DELIVERY

Almost all types of current vaccinesincluding live attenuated, inactivated, sub-unit and DNA vaccines have been tested fortheir effectiveness as a mucosal vaccine ina wide variety of species. However, onlyabout 20 vaccines are currently licensed formucosal delivery. Examples are listed inTable IV. Numerous vaccines are currentlyin an experimental stage of development,and it is expected that many of them will hitthe market in the near future. In general,vaccines containing live microorganisms(live attenuated or vector vaccines) aremore effective for mucosal delivery sincethey rely on the viral or bacterial mecha-nisms of invasion. However, because of theperceived lower safety profile of these livevaccines and the chance of reversion to vir-ulence via recombination between vaccineand wild type strains [103, 157], novel typesof vaccines such as DNA vaccines orrecombinant proteins are currently beingtested in animal models. It should beemphasized that genetic deletions in viru-lence genes provides a safer vaccine thanconventional attenuation. However, sincethe effectiveness of these types of vaccinesis expected to be much lower than live vac-cines, potent adjuvants and delivery sys-tems are needed to enhance the uptake of thevaccine antigen and to increase the induc-tion of immunity. Promising candidates formucosal adjuvants include bacterial toxinsand their non-toxic mutants, bacterial cellwall components, CpG oligonucleotides

Table III. Compartmentalization of the mucosal immune system in mice and humans.

Route Effective response in Non-effective response in

Oral Small intestine (proximal), ascending colon, mammary and salivary glands

Distal large intestine, genital mucosa, tonsils

Rectal Rectum Small intestine, proximal colonNasal or tonsilar Upper airway, regional secretions, genital mucosa GutVaginal Genital mucosaSkin Gut (?)


(CpG ODN), and antimicrobial peptides,also called cationic host defense peptides.However, the broad variety of mucosaladjuvants and their mechanisms of actionare discussed in another paper of the specialissue of Veterinary Research [32].

5. ROUTES OF IMMUNIZATION

The observation that the various com-partments of the mucosal immune systemare in permanent contact with each otherresulted in a wide variety of possible routesof immunization including oral, intranasal,rectal, vaginal, and intraocular (Tabs. III andIV). Nevertheless, immunization of live-stock species and poultry is often limited tooral and intranasal immunization for rea-sons of practicability. Companion animals,however, and even dairy cattle could be the-oretically immunized via all of these routes.As outlined above, immunization via someof the routes favors the accumulation ofeffector cells in certain compartments. Forexample, oral vaccination results in strong-est immunity in the small intestinal tract,whereas intranasal immunization results inoptimal immunity in the respiratory tract(Tab. III). Although some of these observa-tions have not been confirmed yet in domes-tic animals, one can assume that the com-partments of the mucosal immune systems

are connected to each other in a similar fash-ion in domestic species. Thus, depending onthe disease and the species to be vaccinated,future mucosal vaccines should be admin-istered according to the site where maximalimmunity is needed, and the practicabilityof vaccination via that specific route.

6. STRATEGIES FOR MUCOSAL DELIVERY

One of the greatest challenges in vacci-nology today is the development of novelmucosal vaccines and vaccine formulationsthat are safe, effective, and yet cost effective(Tab. III). A variety of delivery systemsincluding live vectors, microparticles, andliposomes have been developed for this pur-pose. Combined with novel strategies to tar-get vaccines to the mucosal surfaces theseprovide tremendous opportunities to addressthis challenge. Here we will summarizerecent progress on live vectors, micropar-ticulate delivery systems, and novel strate-gies to target vaccines to the intestinalmucosa.

6.1. Live vectors

One of the most effective ways to delivervaccines to mucosal surfaces is with livevectors that actually infect the mucosal sur-faces. Both viral and bacterial vectors have

Table IV. Currently licensed mucosal vaccines in domestic animals (in North America).

Species Diseases/pathogens

Bovine BHV-1 (i.n.); rotavirus (oral); coronavirus (oral)Ovine Porcine Transmissible gastroenteritis virus (i.n.); rotavirus (oral); Bordetella bronchiseptica (i.n.)Equine Equine influenza virus (i.n.); Streptocoocus equi (i.n.)Canine Canine adenovirus 2 (i.n.); parainfluenza virus (i.n.); Bordetella bronchiseptica (i.n.)Feline Feline calicivirus (i.o.); feline rhinotracheitis (i.o.)Poultry Turkey adenovirus (oral); infectious bronchitis virus (oral, i.n., i.o., spray); Newcastle

virus (oral, i.n., i.o., spray); infectious bursitis virus (oral, i.n., i.o., spray); chicken herpesvirus (oral, i.n., i.o., spray); turkey herpesvirus (oral, i.n., i.o., spray); reovirus (oral, i.n., i.o., spray); Bordetella avium (oral, spray); Pasteurella multocida (oral)


been developed for vaccine delivery inhumans and animals. In general, live vec-tors allow the delivery of recombinant pro-teins expressed within the vector itself, orgenetic information either integrated intothe genome or as plasmid DNA. Genomicand proteomic approaches have helped inunderstanding the structure and function ofa wide variety of infectious agents andthrough the use of genetic engineering, weare now able to generate a select number oflive vaccines that are safer and possiblymore effective than conventional vaccines(Tab. IV). By introducing multiple genedeletion mutations in a directed way in thegenome of an infectious agent, one can vir-tually eliminate the agents ability to causedisease and reversion to virulence, as wellas make room for the insertion of genesencoding multiple vaccine antigens. Oncean agent is attenuated it can be used to carrygenes or plasmid DNA for other pathogensand immunomodulators (e.g. cytokines)

thus making it possible to immunize ani-mals to produce protective immunity to var-ious disease organisms at one time.

An ideal live vector would have the fol-lowing features: (i) non-pathogenic to animalsand humans, (ii) easy to manipulate, (iii) rel-atively easy and cost effective to produce,(iv) contain stable genome, (v) contain welldefined sites for insertion of foreign genes,(vi) easy to deliver, (vii) no integration intothe host genome, (viii) induce both mucosalas well as systemic immune responses whendelivered orally or intranasally. While theseare very stringent criteria one can hope thatbecause of the rapid progress in this fieldthese will be achievable in the near future.Indeed many live vectored vaccines arealready licensed (Tab. V).6.1.1. Bacterial vectors

Live bacterial carrier vaccines also havegreat potential as novel mucosal vaccines,

Table V. Licensed vector vaccines.

Product Backbone Technology Indication Company

Raboral Vaccinia Vector Rabies MERIALTROVAC/NDV Fowlpox Vector NDV MERIALTROVAC/AIV H5 Fowlpox Vector AIV H5 MERIALRecombitek Canarypox Vector Distemper MERIALPurevax Canarypox Vector Rabies MERIALEurifel FeLV Canarypox Vector FeLV MERIALEurifel RCCP FeLV Canarypox Vector Feline Combo MERIALProteqFlu Canarypox Vector Equine Flu MERIALProteqFlu-Te Canarypox Vector Equine Flu/tetanus MERIALRecombitek WNV Canarypox Vector WNV MERIALPurevax FeLV NF Canarypox Vector FeLV (transdermal) MERIALGallivac IBDV Canarypox Vector IBDV MERIALHVT-MDV HVT Vector MDV IntervetHVT-MDV-NDV HVT Vector MDV/NDV IntervetHVT-IBDV HVT Vector IBDV BiomuneMeganVac-1 Salmonella Deletion mutant Salmonella Avant

NDV (Newcastle disease virus); AIV (avian influenza virus); FeLV (feline leukemia virus); WNV (WestNile virus), MDV (Mareks disease virus); IBDV (infectious bursal disease virus), HVT (turkey herpes-virus). This table was presented by MERIAL at the Marker Vaccine Meeting, Sheman Conference Centre,Ames, IA, April 46, 2005. We are thankful to Drs R. Nordgren, J.-C. Audonnet, and H. Hughes, MERIAL.


with attractive advantages over present-dayinjectable vaccines [53, 158]. For example,live attenuated bacteria such as Salmonellaor Shigella can target mucosal tissues anddeliver the antigen specifically to antigen-presenting cells via bacterial secretion sys-tems [4, 55, 57]. Other bacterial vectorshave been demonstrated to carry plasmidDNA across the mucosal surfaces [34, 37,94, 147] and more recently, bacterial ghostswere demonstrated to represent effectivemeans to deliver plasmid DNA [40]. Thefirst live attenuated bacterial vectors such asSalmonella spp. and Shigella spp. were cre-ated by inserting transposon mutations intothe bacterial genomes [151]. These muta-tions resulted in safer bacteria for deliveringa variety of foreign antigens to animals [53,151, 158]. However, since these initialproof of concept-studies various bacterial

delivery systems have been optimized forthe delivery of recombinant antigens as wellas plasmid DNA (Tab. VI) [53].

In general, live recombinant bacteriaappear to be attractive vehicles of vaccinedelivery as they (i) are inexpensive and canbe easily produced economically, (ii) can bedelivered orally or intranasally, (iii) canaccommodate many different foreign genes,thus protecting animals against severalpathogens simultaneously, (iv) can induceboth humoral and cellular immune responsesand (v) adverse effects can be controlledwith antibiotics if necessary. However, oneof the major concerns regarding the use ofbacterial vectors particularly those derivedfrom attenuated strains is the stability of therecombinant phenotype and the potentialreversion to full virulence [101]. In addi-tion, although a number of bacterial vectors

Table VI. Examples of live bacterial vectors.

Vector Pathogen / Antigen Tested in Reference

Salmonella spp. E. tenella / SO7 :TA4E. coli / HL Toxin B

L. monocytogenesT. parva

Shigella spp.Corynebacterium diphteriae

PRV

ChickenMiceMice

CalvesMiceMiceMice

[125][39][33][56]

[152, 164][118][142]

Corynebacterium pseudotuberculosis Anaplasma marginale / ApHDichelobacter nodosus / protease

Babesia bovis / 11C5Tania ovis / 45W

Sheep [107, 108]

Shigella flexneriListeria monocytogenesBacillus anthracis Clostridium perfringens / Iota toxin Mice [146]Lactococcus lactis IBDV / VP2

BRV / NSP4Chicken [38]

[44]Erysipelothrix rhusiopathiae YS-1 Mycoplasma hyponeumoniae / P97 Pigs [144]Bacillus Calmette-Guerin PRRSV / GP5; M

Toxoplasma gondii / GRA1

Mice/pigs

Sheep

[10, 11][153]

E. coli PRV Mice [143]

IBDV (infectious bursal disease virus), BRV (bovine rotavirus), PRRS (porcine respiratory and repro-ductive syndrome virus).


are capable of inducing humoral and cellu-lar immune responses to passenger anti-gens, it still remains to be seen if preexistingimmunity to bacterial vector will affectbooster immunization [101].

Currently, a number of bacterial vectorsare being evaluated for delivering vaccineantigens and DNA plasmids to mucosal sur-faces for inducing potent immune responses(Tab. VI) [33, 34, 37, 158]. These includecommensal microorganisms such as Lacto-bacillus, Lactococcus, Staphylococcus, Strep-tococcus species, and attenuated pathogenicmicroorganisms such as Salmonella, Shig-ella, BCG, Corynebacterium, Bacillus, Yersi-nia, Vibrio, Erysipelothrix and Bordetellaspecies [34, 37, 93, 133, 147, 148, 151].Although significant progress has beenmade in developing and evaluating bacte-rial vectors for inducing protective immuneresponses, there are not many reports of theirapplication to vaccinate veterinary species(Tab. VI). 6.1.2. Viral vectors

Viral vectors represent another potentialstrategy for efficient delivery of vaccineantigens across the mucosal surfaces [140,175]. Today, a number of viruses including

poxviruses, herpesviruses and adenoviruseshave been used as viral vectors for veteri-nary vaccines (Tabs. V and VIIX) [175].Efficacy of a viral vector is mainly deter-mined by its host range and tropism, the abil-ity to replicate in the target animal, and theexpression of the foreign antigen. Both, therequirement of post translational modifica-tions including proper folding and process-ing of the antigen and the detailed charac-terization of viral genomes have led to thedevelopment of genetically modified virusesas vaccine delivery vehicles for use in theveterinary field [140, 175]. However, incontrast to the human field, viral vectors forveterinary application are currently designedonly for the delivery of vaccines, but not fortargeting tumors or delivering genes forgene-therapy. A number of viral vectors arecurrently being evaluated for mucosal deliv-ery including DNA viruses such as adeno-viruses, pox viruses and herpesviruses, andRNA viruses such as Newcastle diseasevirus (NDV), Venezuelan equine encepha-litis (VEE), Semliki forest virus (SFV) anda few retroviruses (Tabs. V and VIIX). Forexample, a recombinant alphaherpesviruspseudorabies (PRV) carrying the E1 glyc-oprotein of the classical swine fever virus

Table VII. Examples of herpesvirus vectors.


BHV-1 PRV / gC FMDV / VP1, Cp-epitope

BVDV / E2 BRSV / G

Swine, Cattle CattleCattleCattle

[84][82, 83]

[86][138, 156]

HVT NDV / HN;FMDV /

IBDV / VP2

ChickenChickenChicken

[130][130]

[35, 162] PRV HCV / E1 Swine [167]FHV FCV / prCapsid

T. gondii / ROP2CatsCats

[176][106]

BHV (bovine herpesvirus)-1, HVT (herpesvirus of turkey), PRV (pseudorabies virus), FHV (feline her-pesvirus of turkey), FMDV (foot and mouth disease virus), BVDV (bovine viral diarrhea virus), BRSV(bovine respiratory syncytial virus), NDV (New castle disease virus), MDV (Mareks disease virus), IBDV(infectious bursal disease virus), HCV (hog cholera virus), FCV (feline calcivirus), T. gondii (Toxoplasmagondii).


Table VIII. Examples of poxvirus vectors.

Vector Pathogen / Antigen Tested in ReferenceCanary pox virus CDV / HA; F

WNV / PrM; E

DogsSiberian polecat

FerretsHorses

[121][172][171][105]

Modified vaccinia Virus Ankara

FCV / MEIV / HA; NP

T. gondii / ROP2

CatsHorses

Cats

[65][21][134]

Capri pox Pdp ruminants / HA Goats [36]Fowl pox IBDV / VP2

HEVT / hexonBVDV / E2

AIV / H7,H1

ChickenTurkeys

MiceChicken

[22, 25, 66][41][20]

Swinepox virus PRV Pigs [166]CDV (canine distemper virus), WNV (West Nile virus), FCV (feline coronavirus), EIA ( equine influenzavirus), T. gondii (Toxoplasma gondii), Pdp ruminants (peste des petits ruminants), IBDV (infectious bursaldisease virus), BVDV (bovine viral diarrhea virus), HEVT (hemorrhagic enteritis virus of turkey).

Table IX. Examples of adenovirus vectors.

Vector Pathogen / Antigen Tested in ReferenceBAdV-3 BHV-1 / gD

BVDV / E2 Cattle, Cotton rats

Cotton rats [129, 178][12, 177]

OAV Tinea ovis / 45W Sheep [135]PAdV-3

PAdV-5

CSFV / E2PRV / gDTGEV / S

SwineSwineSwine

[62] [63][163]

CAdV-2 CDV / HA; F Dogs [47]CELO IBDV / VP2 Chicken [49]FAdV-8 FAdV-10 IBV / S1

IBDV / VP2ChickenChicken

[75][141]

HAdV-5 PRCV / STGE / S

Rabies / GRabies / GBCV / HEFIV / envPRV / gD PRV / gD

BVDV / E2BHV-1 / gD

PRRSV / GP5

SwineSwineSkunkDogs

Cotton ratsCats

SwineRabbit / Mouse

CattlePigsPigs

[23][161][174][126][5] [60][2]

[43][42][59][52]

BAdV (bovine adenovirus)-3, PAdV (porcine adenovirus)-3;-5, CAdV (canine adenovirus) -2, CELO (cellassociated lethal orphan), FAdV (fowl adenovirus)-8;-10, HAdV (human adenovirus)-5, BHV (bovine her-pesvirus)-1, BVDV (bovine viral diarrhea virus), BRSV (bovine respiratory syncytial virus), CSFV (clas-sical swine fever virus), PRV (pseudorabies virus), TGEV (transmissible gastroenteritis virus), CDV(canine distempervirus), IBDV (infectious bursal disease virus), IBV (infectious bronchitis virus), PRCV(porcine respiratory coronavirus), BCV (bovine coronavirus), FIV (feline immuno deficiency virus), PRV(pseudorabies virus), BVDV (bovine viral diarrhea virus), BHV (bovine herpesvirus)-1, PRRSV (porcinerespiratory and reproductive syndrome virus).


(CSFV) was used by van Zijl et al. [167] toprotect pigs against challenge infectionwith CSFV. Kit et al. [82] used the bovineherpesvirus type 1 (BHV-1) to deliver theviral protein (VP) 1 of foot-and-mouth dis-ease virus (FMDV) to young calves, andKweon et al. [86] used BHV-1 as a vectorfor delivering the glycoprotein E2 of thebovine virus diarrhea virus (BVDV) tocalves. A recombinant BHV-1 vector wasalso used by Schrijver et al., who deliveredthe bovine respiratory virus glycoprotein Gto calves [138]. Other examples for herpes-viral vectors include the herpesvirus of tur-keys (HVT), which has been used to deliverglycoproteins from Newcastle disease virus(NDV) and Marek disease virus (MDV)[130] and the infectious bursal virus(IBDV) VP2 [35] [162]. The feline herpes-virus 1 (FHV) was used for delivering anti-gens from the feline calicivirus (FCV;[176]) or Toxoplasma gondii [106]. Poxvi-ruses represent another type of promisingviral vectors. Indeed, canary pox based vac-cines have been licensed already or are inthe process of getting licensed (Tab. V).Thus, live viral vectors represent powerfultools for delivering vaccine antigens acrossmucosal surfaces. However, concernsregarding their use are related to safety andstability of the vaccines, the applicability,as well as their effectiveness in the presenceof pre-existing immunity. Thus, other strat-egies for inducing protective immunity atthe mucosal surfaces such as particulate

delivery systems may prove to be very com-plementary for vaccinating livestock.

6.2. Particulate delivery systemsIn general, non-replicating antigens such

as proteins and killed vaccines are poorlyimmunogenic when given mucosally. Thishas been attributed to degradation of anti-gen or inefficient uptake by APC in themucosae. Studies in laboratory animals haveclearly demonstrated that particulate deliv-ery systems can significantly improve theimmunogenicity of mucosally administeredantigens. However, there are only limitedstudies in domestic animals and the poten-tial of mucosal immunization with killedantigen remains largely unexploited in vet-erinary medicine. We will briefly reviewthe delivery of antigens to mucosae usingparticulate delivery systems (microparticles,immune stimulating complex (ISCOM) andliposomes), and where possible highlightinvestigations in domestic animals or whereantigens from pathogens of veterinaryimportance have been used.

6.2.1. Microparticles

The enhanced immune responses observedwith the use of microparticles for mucosaldelivery is related to protecting the antigenfrom degradation and increasing antigenuptake into specialized mucosa-associatedlymphoid tissues [116]. Also, microparti-cles facilitate presentation by APC via both

Table X. Examples of other viral vectors.


VEE EAV / G(L); MAnthrax / PA

HorsesMice

[8][88]

SFV LIV / ME; NSIBDV / VP2; VP2,4,3

SheepChickens

[112][124]

NDV IBDV / VP2 Chicken [71] FFV FCV / capsid Pr Cats [139]

VEE (Venezuelan equine encephalitis), SFV (Semliki forest virus), NDV (Newcastle disease virus),FFV (feline foamy virus), EAV (equine arteritis virus), LIV (Looping ill virus), IBDV (infectious bursaldisease virus), FCV (feline calcivirus).


MHC class I and MHC class II restrictedprocessing and presentation pathways [102].Uptake of microparticles occurs predomi-nantly by M cells in the follicle-associatedepithelium overlying the Peyers patches inmice and domestic animals [13, 81]. Inter-estingly, DC extend their dendrites betweenepithelial cells to sample antigen in thelumen [131], but the significance of thispathway in particle uptake is unclear. How-ever, the poor uptake of microparticles is amajor limitation of all microparticle deliv-ery systems which results in suboptimalresponses. Approaches such as targetingmicroparticles to M cells using lectins [48]as well as incorporating adjuvants such asCpG ODN can improve the potency ofmicroparticles as delivery vehicles.

6.2.1.1. Polymers

A variety of polymers can be made intomicroparticles including Poly-lactide-co-glycolide (PLG), alginate, polyphosphazenesand starch. PLG is a biodegradable aliphaticpolyester used in humans as suture materi-als and has been extensively investigatedfor the delivery of micro-encapsulated vac-cines. A variety of antigens have beenencapsulated in PLG microparticles withsuccessful induction of protective immuneresponses [117, 165]. Interestingly, oralimmunization of mice with ovalbumin(OVA) encapsulated in PLG microparticlesinduced serum IgG and intestinal IgAresponses to a level similar to that inducedwith cholera toxin (CT) adjuvants [117].This is significant given that CT is the mostpotent and widely investigated mucosaladjuvant. Mucosal immunization with anti-gens encapsulated in PLG microparticlesprotected animals against challenge withmucosal pathogens. Mice orally immunizedwith antigen in PLG microparticles werebetter protected against oral challenge withS. typhimurium, than those immunized intra-peritoneally (IP) with antigen in Freundsadjuvant [3, 45]. Interestingly, a single oralimmunization with fimbriae from Borde-tella pertussis encapsulated in PLG micro-

particles protected mice from intranasalchallenge with the bacteria [76], indicatingthat the microparticle delivery systemeffectively stimulated the common mucosalimmune system to protect against infectionat a distant mucosal site. PLG microparti-cles have also been evaluated with DNAvaccines. Mice orally immunized with rota-virus VP4, VP6 and VP7 DNA vaccines inPLG microparticles elicited both rotavirus-specific antibodies and protection againstchallenge with rotavirus [28, 67]. Sinceencapsulation of plasmid DNA causes sig-nificant damage to the DNA as a result ofshear, adsorption of DNA on the surface ofPLG microparticles can circumvent thisproblem. Intranasal (IN) immunization ofmice with DNA encoding HIV-1 gagadsorbed on the surface of cationic PLGmicroparticles induced antibody and cell-mediated immune responses locally andsystemically, but no responses were seenwith naked DNA [145]. A single IN immu-nization of mice with synthetic malariaSPf66 vaccine in PLG microparticles greatlyimproved and sustained systemic Th1 andTh2 responses over conventional alumadjuvant [24]. This suggests that mucosalimmunization with microparticles has thepotential to deliver the next generation ofvaccines against many different diseases.Besides mice, PLG microparticles havealso been evaluated in other species. Threeof ten adult humans immunized via an intes-tinal tube with enterotoxigenic E. coli (ETEC)antigen (CFA/II) in PLG microparticleswere protected, but all ten unimmunizedcontrols developed clinical disease [154].In contrast, oral immunization of pigs withETEC antigens in PLG microparticles didnot induce any significant antibody responseor reduction in E. coli shedding, perhapsreflecting differences in immunization pro-cedures [46]. Torche et al. [159, 160] dem-onstrated phagocytosis of PLG-particles byporcine macrophages in vitro, and assessedtheir uptake in the intestine via M cells. Asingle oral immunization in 2-week-old chick-ens with formalin-inactivated S. enteritidisin PLG microparticles elicited a significant


intestinal S-IgA antibody response and pro-tected birds against oral and intramuscularchallenge [91]. The use of PLG micropar-ticles may be limited by the fact that PLGis soluble only in organic solvents, whichmay denature critical epitopes required forinducing protection. Poly--caprolactone(PEC) microparticles, like PLG, are biode-gradable polyesters. Oral immunization ofmice with an antigen from Brucella ovis inPEC microparticles protected mice againstIP challenge while antigens in PLG did not[114]. However, a study by [9] using Schis-tosoma mansonii antigens did not supportthis conclusion.

6.2.1.2.Alginate Alginate is a naturally occurring carbo-

hydrate (kelp) which polymerizes into par-ticles upon contact with divalent cations.Oral immunization of cattle with the modelantigen OVA in alginate microparticlesenhanced IgA- and IgG-immune responsesin the respiratory tract, providing evidencethat oral immunization with microparticlescan stimulate the common mucosal immunesystem in a large animal [17]. Calves immu-nized IN with porcine serum albumin (PSA)in alginate microparticles developed highlevel of anti-PSA IgG1 antibodies in serum,nasal secretions and saliva [128]. In con-trast, oral immunization did not induce anysignificant responses, suggesting that INwas more efficient for induction of nasaland serum responses. Rabbits orally immu-nized with P. multocida antigens encapsu-lated in alginate microparticles developedsignificantly higher nasal IgA responses[18]. Increased protection against P. haemo-lytica and Streptococcus pneumoniae chal-lenge in mice was demonstrated by encap-sulating antigens in alginate microparticlesfollowing oral immunization [80]. Thesestudies clearly show that alginate micropar-ticles are effective for mucosal delivery ofvaccines in small and large animals.

6.2.1.3.Polyphosphazenes Polyphosphazenes are synthetic biode-

gradable and water-soluble polymers (no

need for organic solvents) with potential asvaccine delivery systems. Procedures forthe preparation of poly[di(carboxylatophe-noxy)phosphazene] (PCPP) microparticlesappear to be relatively simple [122]. INimmunization of mice with tetanus toxoidor influenza antigens in PCPP microparti-cles induced significant increases in serumIgG in mice [122, 123]. Considering thatPCPP microparticles are relatively easy toproduce in an aqueous environment, and thepolymer has adjuvant activity as well, inves-tigations in large animals are warranted.

6.2.1.4.Other biodegradable polymersAntigens from Mycoplasma hyopneu-

moniae and Actinobacillus pleuropneumo-niae coated with acrylic resins (celluloseacetate phthalate and methacrylic acid) toprotect them from the low pH of the stom-ach and released in intestines have beentested and shown efficacy when givenorally to swine [89, 90]. Fimbriae of ETECin a cellulose-based delivery system reduceddisease in experimentally infected pigs [150].

6.2.2. ISCOM

ISCOM is a small 40 nm nanoparticlecomposed of saponin (adjuvant), lipids andantigen, and has been described as an anti-gen delivery system because it not only hasadjuvant activity and but also the ability totarget APC [111]. ISCOM is a versatiledelivery system that allows incorporation ofadditional adjuvants and targetting mole-cules. The mechanisms whereby ISCOMare so effective in inducing immunity isrelated to the ability to stimulate innateimmunity by producing cytokines such asIL-12 and modulation of antigen uptake viarecruitment of APC [51, 149]. Furthermore,ISCOM target antigens to both the endo-somal and cytosolic pathways resulting inboth MHC class I and MHC class IIrestricted immune responses [110]. Numer-ous studies indicate that ISCOM enhanceimmune responses to a variety of vaccineantigens in laboratory and domestic ani-mals, and ISCOM are now included in


injectable commercial veterinary vaccines[18, 111]. However, there is also evidencethat ISCOM have great potential in thedelivery of antigens to the mucosae. Mowatet al. [97, 113] showed OVA incorporatedinto ISCOM given orally induced serumantibodies, mucosal IgA, Th1 and Th2 CD4T cell responses and MHC I restricted CTLactivity. Mice fed OVA in ISCOM prior tofeeding with tolerogenic doses of OVA hadelevated OVA-specific antibody responses,indicating that ISCOM may deviate immuneresponse in favor of immunity rather thantolerance [97, 113]. Additionally, OVA inISCOM induced the recruitment of DCand macrophages into the mesenteric lymphnodes and recruitment of macrophagesand B cells in Peyers patch tissues [51].Lovgren et al. [95] showed that mice immu-nized IN with influenza ISCOM were pro-tected against subsequent challenge withinfluenza virus.

Intranasal immunization with respira-tory syncytial virus (RSV) antigens inISCOM induced high levels of IgA in theupper respiratory tract and in the lungs ofmice [70]. Antigens from Mycoplasmamycoides subspecies mycoides, a cause ofsevere lung disease in cattle, were incorpo-rated in ISCOM and induced mucosal andsystemic IgG1, IgG2a and IgG2b antibodyresponses in mice [1]. Dogs immunized INwith Echinococcus granulosus antigens inISCOM induced high mucosal IgA responsesbut no systemic antibody responses [26].Based on these reports it appears thatISCOM fulfill some important attributes ofan effective mucosal delivery system; itcontains an adjuvant (saponin) and allowsincorporation of additional adjuvants, stim-ulates the innate immune system andrecruits and targets antigen to DC. By incor-porating targeting molecules into the ISCOMit should be possible to further enhance theirutility as mucosal delivery systems [111].Further investigations in domestic animalsusing relevant antigens are warranted tofully exploit the potential of this technologyin veterinary medicine.

6.2.3. Liposomes

Liposomes are phospholipid vesicleswhich enhance immune responses prima-rily by increasing antigen uptake and pres-entation [61]. Liposomes have been usedextensively in oral and IN delivery of anti-gens in mice [104]. Since uptake of lipo-somes by the GALT following oral deliveryis a major concern [179], some recent inves-tigations have focused on improving uptakeof liposomes and the co-delivery of adju-vants as a way of improving mucosalresponses. In this regard, conjugation ofliposomes with recombinant B subunit ofcholera toxin (rCTB) significantly enhancedmucosal IgA and serum IgG compared toresponses in mice immunized orally withunconjugated liposomes [64]. Similarly,the presence of IgA on liposomal surfacesincreased uptake of liposomes into the PPmucosa, and the local rectal and colonic IgAresponses to ferritin following rectal immu-nization of mice was about 5-fold higherthan those induced by uncoated liposomes[180]. The addition of CT to a liposome-antigen formulation also enhanced responsesto ferritin [180]. Addition of the adjuvantmonophosphoryl lipid A to VTEC antigenformulated in liposomes induced signifi-cant systemic and mucosal antibodyresponses in mice immunized orally [155].IN immunization of mice with a commer-cial inactivated Yersinia pestis vaccine inliposomes resulted in significant enhance-ment of IgG and IgA titers in lung and nasalwashes of immunized mice [7]. Interest-ingly, IN immunization of mice with lipo-some-encapsulated plasmid DNA encodinginfluenza hemagglutinin conferred com-plete protection while all mice immunizedwith naked plasmid died [169]. Chickensimmunized by intraoccular, oral and INdelivery with liposome-associated wholecell extract of Salmonella enterica serovarenteritidis developed significant serum andintestinal IgG and IgA antibody responseswhereas those immunized with antigenalone had no detectable antibody responses.Interestingly, intraoccular immunization of


chickens elicited better responses than bothIN and oral immunization [50]. Apparentlyintraocular immunization circumvents acids,enzymes and dilution effects encounteredby antigen in the respiratory and gastroin-testinal tracts.

Liposomes have been successfully usedto co-deliver antigen and IL-12 by IN deliv-ery [6]. An alternative to co-deliveringcytokines is to use liposomes containingcytokine-inducing immune modulators suchas CpG ODN. This would reduce the poten-tial toxicity often associated with large dosesof cytokines. Indeed, the adjuvant activityof CpG ODN was augmented by liposomaldelivery in mice immunized IN againstinfluenza and hepatitis B viruses [77].

6.3. Other strategies to enhance vaccine uptake

6.3.1. Lectin-mediated targeting

The efficacy of mucosal vaccines is cur-rently limited by very inefficient delivery ofthe antigen to the mucosal surfaces. Thus,various strategies to selectively target anti-gens to M cells or enterocytes have beendeveloped in small rodents [73]. One of themost promising current strategies is the useof lectins that specific recognize sugar-res-idues on the apical surfaces of M cells. Forexample, the Ulex europeus 1 (UEA1), alectin specifically for -L-fucose residues,binds almost exclusively to the apical sur-face of mouse Peyers patch M cells. UEA1-coated antigens (OVA) were specificallytargeted to the M cells in murine Peyerspatches after administration into ligatedintestinal loops [48], and resulted inenhanced immunity against OVA. Simila-rily, incorporation of wheat germ agglutininlectin (WGA) also enhanced uptake of pol-ymerised liposomes by Peyers patch, albeitat lower levels than UEA1 [27]. Giannascaet al. reported that an M cell selective lectinadministered via the intranasal route tar-geted to and was endocytosed by respira-tory hamster M cells in vivo [58] and Roth-

Walter et al. showed that lectins from Aleu-ria aurantia can be used to target M cell fororal allergen immunotherapy [136, 137].Novel strategies for targeting DNA vac-cines to intestinal and respiratory M cellsare based on using the M cell specific reo-virus protein 1 as a carrier [170, 173].Yersinia pseudotuberculosis invades via Mcells in vivo and this is mediated by inter-action between bacterial invasin with cellsurface 1-integrins on M cells [29]. Hus-sain and Florence mimicked this microbialstrategy and showed that Yersinia adhesionprotein invasion could be used to improveuptake of nanoparticles [72]. However, itremains to be formerly proven whether thisstrategy can result in significant enhance-ment of immune responses.

6.3.2. Antibody-mediated targeting

Other strategies for targeting the mucosalsurfaces include antibody-mediated target-ing to mucosal homing molecules such asthe mucosal-addressin cellular adhesionmolecule (MAdCAM-1). Mc Kenzie et al.vaccinated mice with OVA and demon-strated that even after parenteral immuni-zation the targeting to MadCAM enhancedOVA-specific antibody responses in gutand serum [100]. Bonifaz et al. [16] recentlyreported that antibody-mediated targetingof antigens to maturing dendritic cells viathe DEC-205 receptor increased the effi-ciency of mucosal and systemic immunityin mice.

6.3.3. Receptor-mediated targeting

Another strategy of targeting intestinalsurfaces is to use antigen-specific receptors.For example, the E. coli fimbriae protein(F4) binds to the F4 receptor (F4R) on thesurface of porcine intestinal cells [31].Interaction between ligand and receptor isrequired for pathogenesis, and only F4R+pigs develop disease, and interestingly alsoimmunity against E. coli. Thus, a possiblestrategy for mucosal vaccines would be tofuse a vaccine antigen to the F4 in order toretain the antigen within the intestinal tract


and facilitate antigen uptake via M cells anddendritic cells. In fact, Verdonck et al.recently demonstrated that chemically con-jugated F4 to human serum albumin (HAS)resulted in enhanced HSA-specific IgA-and IgG responses [168]. Finally, it has alsobeen possible to express a fragment of thenon-glycosylated E2 antigen of classicalswine fever in E. coli and administer theinclusion bodies orally to mice. All vacci-nated mice developed IgG and IgA antibod-ies against the E2 [79]. This is an interestingobservation since systemic immunizationwith inclusion bodies generally is ineffec-tive in inducing immune responses. It ispossible that the proteolytic cleavage in theintestine combined with the particulatenature of the inclusion bodies allows uptakeand processing of the antigen suitable forinduction of immunity.

7. CONCLUSION

The advent of molecular biology and ourunderstanding of the critical antigens involvedin inducing protection from infectious dis-eases has allowed researchers and compa-nies to identify and produce large quantitiesof putative protective antigens from almostany pathogen. However, the major chal-lenge to effective vaccination now lies informulation and delivery of these antigens.Our thesis is that even with the best antigenidentification production systems, the vac-cines will not reach their full potentialunless they are formulated and deliveredproperly. Thus, economic losses and animalsuffering will continue.

Since over 90% of all pathogens, regard-less of which species, enter and initiateinfection at mucosal surfaces, the best targetfor effective vaccines is the mucosal surfaceto reduce the ability of the pathogen to getestablished. Thus, the mucosal surface shouldbe our site for immunization since systemicimmunization rarely induces mucosalresponses. As a result, systemic immunity,if effective, can only prevent disease buthave little impact on the initial infection

process. Even if mucosal immunizationdoes not totally eliminate infection, mucosalantibody limits the degree of replication andshedding of the pathogen, thereby, reducingthe pathogen load in the environment andconsequently dramatically reducing the rateof herd infection and transmission of dis-ease through the herd.

The recent advances in mucosal immu-nization, especially in rodents are very encour-aging since they demonstrate that mucosalimmunization is feasible. Whether all vac-cines will be given mucosally is debateablesince some management systems will makeintranasal immunization very difficult.However, one could envisage initial sys-temic immunization followed by intranasalboosting, thereby, achieving the full benefitof mucosal immunization with only half thechallenge of intranasal immunization. Asecond challenge is the duration of immu-nity at mucosal surfaces. Since mucosalimmunization induces both mucosal andsystemic immunity even if mucosal immu-nity wanes upon exposure to a mucosalpathogen, there will be a rapid responseboth systemically and mucosally.

Possibly the most exciting developmentin the past few years is the knowledge thatthe most effective antigen-presenting cellshave specific cell markers. This is allowingus to directly link antigens to antibodies orlectins that specifically bind to the dendriticcell, thereby ensuring that the quantity ofantigen needed to induce immunity isextremely small. This makes the productionof expensive antigens feasible for livestockvaccine due to the extremely low quantitiesof antigen acquired (antigen sparing). Weenvisage that such approaches will becomecommon place in livestock vaccines beingdeveloped over the next decade. Thus, wewill not only be able to target the vaccinesto mucosal surfaces, but ensure that thesevaccines are further targeted to the antigen-presenting cells that are monitoring theevents occurring at these mucosal surfaces.It is interesting that such targeting can occurwith live vectored vaccines, as well as with


subunit and even DNA-based vaccines.Furthermore, by designing formulationdelivery systems which focus the immuneresponse to either give a balanced immuneresponse or one skewed to either Th1 or Th2depending on the pathogen of interest, wecan target the response as needed for max-imum protection and reduce the conse-quences of infection from most pathogens.As a result of the recent advances, we areon the brink of unprecedented opportunitiesto reduce animal suffering, improve theeconomics of livestock production, andreduce the spread of many infectious dis-eases from animals to humans. This latterfactor is becoming extremely important asthe livestock industry finds itself at thecrossroads with our continued urbaniza-tion. Historically, over 50% of all humandiseases were considered zoonotic, how-ever, currently, a full 80% of the newemerging diseases are zoonotic. As our glo-bal population increases, there will be morepressure on the agricultural industry toreduce all types of contaminants arisingfrom livestock including infectious diseases.

ACKNOWLEDGEMENTS

Funding in the investigators laboratories wasprovides by the Canadian Institute for HealthResearch (CIHR), the National Science andEngineering Research Council of Canada(NSERC), the Sakatchewan Health ResearchFoundation (SHRF), and the Agriculture Devel-opment Fund Saskatchewan (ADF Saskatch-ewan). Published with permission of theDirector of the Vaccine and Infectious DiseaseOrganization as article number 406.

REFERENCES[1] Abusugra I., Morein B., Iscom is an efficient

mucosal delivery system for Mycoplasmamycoides subsp. mycoides (MmmSC) anti-gens inducing high mucosal and systemicantibody responses, FEMS Immunol. Med.Microbiol. 23 (1999) 512.

[2] Adam M., Lepottier M.F., Eloit M., Vaccina-tion of pigs with replication-defective aden-ovirus vectored vaccines: the example of

pseudorabies, Vet. Microbiol. 42 (1994)205215.

[3] Allaoui-Attarki K., Pecquet S., Fattal E.,Trolle S., Chachaty E., Couvreur P.,Andremont A., Protective immunity againstSalmonella typhimurium elicited in mice byoral vaccination with phosphorylcholineencapsulated in poly(DL-lactide-co-glycol-ide) microspheres, Infect. Immun. 65 (1997)853857.

[4] Autenrieth I.B., Schmidt M.A., Bacterialinterplay at intestinal mucosal surfaces:implications for vaccine development,Trends Microbiol. 8 (2000) 457464.

[5] Baca-Estrada M.E., Liang X., Babiuk L.A.,Yoo D., Induction of mucosal immunity incotton rats to haemagglutinin-esterase glyco-protein of bovine coronavirus by recom-binant adenovirus, Immunology 86 (1995)134140.

[6] Baca-Estrada M.E., Foldvari M., Snider M.,Induction of mucosal immune responses byadministration of liposome-antigen formula-tions and interleukin-12, J. InterferonCytokine Res. 19 (1999) 455462.

[7] Baca-Estrada M.E., Foldvari M.M., SniderM.M., Harding K.K., Kournikakis B.B.,Babiuk L.A., Griebel P.P., Intranasal immu-nization with liposome-formulated Yersiniapestis vaccine enhances mucosal immuneresponses, Vaccine 18 (2000) 22032211.

[8] Balasuriya U.B., Heidner H.W., Davis N.L.,Wagner H.M., Hullinger P.J., Hedges J.F.,Williams J.C., Johnston R.E., David WilsonW., Liu I.K., James MacLachlan N., Alpha-virus replicon particles expressing the twomajor envelope proteins of equine arteritisvirus induce high level protection againstchallenge with virulent virus in vaccinatedhorses, Vaccine 20 (2002) 16091617.

[9] Baras B., Benoit M.A., Dupre L., Poulain-Godefroy O., Schacht A.M., Capron A.,Gillard J., Riveau G., Single-dose mucosalimmunization with biodegradable micropar-ticles containing a Schistosoma mansoniantigen, Infect. Immun. 67 (1999) 26432648.

[10] Bastos R.G., Dellagostin O.A., Barletta R.G.,Doster A.R., Nelson E., Osorio F.A., Con-struction and immunogenicity of recom-binant Mycobacterium bovis BCG express-ing GP5 and M protein of porcinereproductive respiratory syndrome virus,Vaccine 21 (2002) 2129.

[11] Bastos R.G., Dellagostin O.A., Barletta R.G.,Doster A.R., Nelson E., Zuckermann F.,Osorio F.A., Immune response of pigs inoc-ulated with Mycobacterium bovis BCGexpressing a truncated form of GP5 and M


protein of porcine reproductive and respira-tory syndrome virus, Vaccine 22 (2004) 467474.

[12] Baxi M.K., Deregt D., Robertson J., BabiukL.A., Schlapp T., Tikoo S.K., Recombinantbovine adenovirus type 3 expressing bovineviral diarrhea virus glycoprotein E2 inducesan immune response in cotton rats, Virology278 (2000) 234243.

[13] Beier R., Gebert A., Kinetics of particleuptake in the domes of Peyers patches, Am.J. Physiol. 275 (1998) G130G137.

[14] Belyakov I.M., Hammond S.A., Ahlers J.D.,Glenn G.M., Berzofsky J.A., Transcutaneousimmunization induces mucosal CTLs andprotective immunity by migration of primedskin dendritic cells, J. Clin. Invest. 113(2004) 9981007.

[15] Bjarnarson S.P., Jakobsen H., Del GiudiceG., Trannoy E., Siegrist C.A., Jonsdottir I.,The advantage of mucosal immunization forpolysaccharide-specific memory responsesin early life, Eur. J. Immunol. 35 (2005)10371045.

[16] Bonifaz L.C., Bonnyay D.P., CharalambousA., Darguste D.I., Fujii S., Soares H.,Brimnes M.K., Moltedo B., Moran T.M.,Steinman R.M., In vivo targeting of antigensto maturing dendritic cells via the DEC-205receptor improves T cell vaccination, J. Exp.Med. 199 (2004) 815824.

[17] Bowersock T.L., HogenEsch H., TorregrosaS., Borie D., Wang B., Park H., Park K., Induc-tion of pulmonary immunity in cattle by oraladministration of ovalbumin in alginate micro-spheres, Immunol. Lett. 60 (1998) 3743.

[18] Bowersock T.L., HogenEsch H., Suckow M.,Guimond P., Martin S., Borie D., TorregrosaS., Park H., Park K., Oral vaccination of ani-mals with antigens encapsulated in alginatemicrospheres, Vaccine 17 (1999) 18041811.

[19] Bowman E.P., Kuklin N.A., YoungmanK.R., Lazarus N.H., Kunkel E.J., Pan J.,Greenberg H.B., Butcher E.C., The intestinalchemokine thymus-expressed chemokine(CCL25) attracts IgA antibody-secretingcells, J. Exp. Med. 195 (2002) 269275.

[20] Boyle D.B., Selleck P., Heine H.G., Vacci-nating chickens against avian influenza withfowlpox recombinants expressing the H7haemagglutinin, Aust. Vet. J. 78 (2000) 4448.

[21] Breathnach C.C., Rudersdorf R., Lunn D.P.,Use of recombinant modified vacciniaAnkara viral vectors for equine influenzavaccination, Vet. Immunol. Immunopathol.98 (2004) 127136.

[22] Butter C., Sturman T.D., Baaten B.J., DavisonT.F., Protection from infectious bursal dis-

ease virus (IBDV)-induced immunosuppres-sion by immunization with a fowlpox recom-binant containing IBDV-VP2, Avian Pathol.32 (2003) 597604.

[23] Callebaut P., Enjuanes L., Pensaert M., Anadenovirus recombinant expressing the spikeglycoprotein of porcine respiratory coronavi-rus is immunogenic in swine, J. Gen. Virol.77 (1996) 309313.

[24] Carcaboso A.M., Hernandez R.M., IgartuaM., Rosas J.E., Patarroyo M.E., Pedraz J.L.,Potent, long lasting systemic antibody levelsand mixed Th1/Th2 immune response afternasal immunization with malaria antigenloaded PLGA microparticles, Vaccine 22(2004) 14231432.

[25] Cardona C.J., Reed W.M., Witter R.L., SilvaR.F., Protection of turkeys from hemorrhagicenteritis with a recombinant fowl poxvirusexpressing the native hexon of hemorrhagicenteritis virus, Avian Dis. 43 (1999) 234244.

[26] Carol H., Nieto A., A mucosal IgA response,but no systemic antibody response, is evokedby intranasal immunisation of dogs withEchinococcus granulosus surface antigensiscoms, Vet. Immunol. Immunopathol. 65(1998) 2941.

[27] Chen H., Torchilin V., Langer R., Lectin-bearing polymerized liposomes as potentialoral vaccine carriers, Pharm. Res. 13 (1996)13781383.

[28] Chen S.C., Jones D.H., Fynan E.F., FarrarG.H., Clegg J.C., Greenberg H.B., HerrmannJ.E., Protective immunity induced by oralimmunization with a rotavirus DNA vaccineencapsulated in microparticles, J. Virol. 72(1998) 57575761.

[29] Clark M.A., Hirst B.H., Jepson M.A., M-cellsurface beta1 integrin expression and inva-sin-mediated targeting of Yersinia pseudotu-berculosis to mouse Peyers patch M cells,Infect. Immun. 66 (1998) 12371243.

[30] Colonna M., Trinchieri G., Liu Y.J., Plasma-cytoid dendritic cells in immunity, Nat.Immunol. 5 (2004) 12191226.

[31] Cox E., Van der Stede Y., Verdonck F.,Snoeck V., Van den Broeck W., GoddeerisB., Oral immunisation of pigs with fimbrialantigens of enterotoxigenic E. coli: an inter-esting model to study mucosal immunemechanisms, Vet. Immunol. Immunopathol.87 (2002) 287290.

[32] Cox E., Verdonck F., Goddeeris B., Adjuvantsmodulating mucosal immune responses ordirecting systemic responses towards themucosa, Vet. Res. (2006) 511539.


[33] Darji A., Guzman C.A., Gerstel B., WachholzP., Timmis K.N., Wehland J., ChakrabortyT., Weiss S., Oral somatic transgene vacci-nation using attenuated S. typhimurium, Cell91 (1997) 765775.

[34] Darji A., zur Lage S., Garbe A.I.,Chakraborty T., Weiss S., Oral delivery ofDNA vaccines using attenuated Salmonellatyphimurium as carrier, FEMS Immunol.Med. Microbiol. 27 (2000) 341349.

[35] Darteil R., Bublot M., Laplace E., BouquetJ.F., Audonnet J.C., Riviere M., Herpesvirusof turkey recombinant viruses expressinginfectious bursal disease virus (IBDV) VP2immunogen induce protection against anIBDV virulent challenge in chickens, Virol-ogy 211 (1995) 481490.

[36] Diallo A., Minet C., Berhe G., Le Goff C.,Black D.N., Fleming M., Barrett T., GrilletC., Libeau G., Goat immune response tocapripox vaccine expressing the hemaggluti-nin protein of peste des petits ruminants,Ann. NY Acad. Sci. 969 (2002) 8891.

[37] Dietrich G., Spreng S., Favre D., Viret J.F.,Guzman C.A., Live attenuated bacteria asvectors to deliver plasmid DNA vaccines,Curr. Opin. Mol. Ther. 5 (2003) 1019.

[38] Dieye Y., Hoekman A.J., Clier F., Juillard V.,Boot H.J., Piard J.C., Ability of Lactococcuslactis to export viral capsid antigens: a crucialstep for development of live vaccines, Appl.Environ. Microbiol. 69 (2003) 72817288.

[39] Dougan G., Hormaeche C.E., Maskell D.J.,Live oral Salmonella vaccines: potential useof attenuated strains as carriers of heterolo-gous antigens to the immune system, ParasiteImmunol. 9 (1987) 151160.

[40] Ebensen T., Paukner S., Link C., Kudela P.,de Domenico C., Lubitz W., Guzman C.A.,Bacterial ghosts are an efficient delivery sys-tem for DNA vaccines, J. Immunol. 172(2004) 68586865.

[41] Elahi S.M., Shen S.H., Talbot B.G., MassieB., Harpin S., Elazhary Y., Induction ofhumoral and cellular immune responsesagainst the nucleocapsid of bovine viraldiarrhea virus by an adenovirus vector withan inducible promoter, Virology 261 (1999)17.

[42] Elahi S.M., Shen S.H., Talbot B.G., MassieB., Harpin S., Elazhary Y., Recombinant ade-noviruses expressing the E2 protein of bovineviral diarrhea virus induce humoral and cel-lular immune responses, FEMS Microbiol.Lett. 177 (1999) 159166.

[43] Eloit M., Gilardi-Hebenstreit P., Toma B.,Perricaudet M., Construction of a defectiveadenovirus vector expressing the pseudora-bies virus glycoprotein gp50 and its use as a

live vaccine, J. Gen. Virol. 71 (1990) 24252431.

[44] Enouf V., Langella P., Commissaire J., CohenJ., Corthier G., Bovine rotavirus nonstruc-tural protein 4 produced by Lactococcus lac-tis is antigenic and immunogenic, Appl.Environ. Microbiol. 67 (2001) 14231428.

[45] Fattal E., Pecquet S., Couvreur P.,Andremont A., Biodegradable microparti-cles for the mucosal delivery of antibacterialand dietary antigens, Int. J. Pharm. 242(2002) 1524.

[46] Felder C.B., Vorlaender N., Gander B.,Merkle H.P., Bertschinger H.U., Microen-capsulated enterotoxigenic Escherichia coliand detached fimbriae for peroral vaccina-tion of pigs, Vaccine 19 (2000) 706715.

[47] Fischer L., Tronel J.P., Pardo-David C., TannerP., Colombet G., Minke J., Audonnet J.C.,Vaccination of puppies born to immune damswith a canine adenovirus-based vaccine pro-tects against a canine distemper virus chal-lenge, Vaccine 20 (2002) 34853497.

[48] Foster N., Clark M.A., Jepson M.A., HirstB.H., Ulex europaeus 1 lectin targets micro-spheres to mouse Peyers patch M-cells invivo, Vaccine 16 (1998) 536541.

[49] Francois A., Chevalier C., Delmas B.,Eterradossi N., Toquin D., Rivallan G.,Langlois P., Avian adenovirus CELO recom-binants expressing VP2 of infectious bursaldisease virus induce protection against bursaldisease in chickens, Vaccine 22 (2004)23512360.

[50] Fukutome K., Watarai S., Mukamoto M.,Kodama H., Intestinal mucosal immuneresponse in chickens following intraocularimmunization with liposome-associated Sal-monella enterica serovar enteritidis antigen,Dev. Comp. Immunol. 25 (2001) 475484.

[51] Furrie E., Smith R.E., Turner M.W., StrobelS., Mowat A.M., Induction of local innateimmune responses and modulation of antigenuptake as mechanisms underlying themucosal adjuvant properties of immune stim-ulating complexes (ISCOMS), Vaccine 20(2002) 22542262.

[52] Gagnon C.A., Lachapelle G., Langelier Y.,Massie B., Dea S., Adenoviral-expressedGP5 of porcine respiratory and reproductivesyndrome virus differs in its cellular matura-tion from the authentic viral protein but main-tains known biological functions, Arch.Virol. 148 (2003) 951972.

[53] Garmory H.S., Leary S.E., Griffin K.F.,Williamson E.D., Brown K.A., Titball R.W.,The use of live attenuated bacteria as a deliv-ery system for heterologous antigens, J. DrugTarget. 11 (2003) 471479.


[54] Gasque P., Complement: a unique innateimmune sensor for danger signals, Mol.Immunol. 41 (2004) 10891098.

[55] Gentschev I., Mollenkopf H., Sokolovic Z.,Hess J., Kaufmann S.H., Goebel W., Devel-opment of antigen-delivery systems, basedon the Escherichia coli hemolysin secretionpathway, Gene 179 (1996) 133140.

[56] Gentschev I., Glaser I., Goebel W., McKeeverD.J., Musoke A., Heussler V.T., Delivery ofthe p67 sporozoite antigen of Theileria parvaby using recombinant Salmonella dublin:secretion of the product enhances specificantibody responses in cattle, Infect. Immun.66 (1998) 20602064.

[57] Gentschev I., Dietrich G., Goebel W., The E.coli alpha-hemolysin secretion system and itsuse in vaccine development, Trends Micro-biol. 10 (2002) 3945.

[58] Giannasca P.J., Boden J.A., Monath T.P.,Targeted delivery of antigen to hamster nasallymphoid tissue with M-cell-directed lectins,Infect. Immun. 65 (1997) 42884298.

[59] Gogev S., Vanderheijden N., Lemaire M.,Schynts F., DOffay J., Deprez I., Adam M.,Eloit M., Thiry E., Induction of protectiveimmunity to bovine herpesvirus type 1 in cat-tle by intranasal administration of replica-tion-defective human adenovirus type 5expressing glycoprotein gC or gD, Vaccine20 (2002) 14511465.

[60] Gonin P., Fournier A., Oualikene W., MoraillonA., Eloit M., Immunization trial of cats witha replication-defective adenovirus type 5expressing the ENV gene of feline immuno-deficiency virus, Vet. Microbiol. 45 (1995)393401.

[61] Gregoriadis G., Immunological adjuvants: arole for liposomes, Immunol. Today 11(1990) 8997.

[62] Hammond J.M., McCoy R.J., Jansen E.S.,Morrissy C.J., Hodgson A.L., Johnson M.A.,Vaccination with a single dose of a recom-binant porcine adenovirus expressing theclassical swine fever virus gp55 (E2) geneprotects pigs against classical swine fever,Vaccine 18 (2000) 10401050.

[63] Hammond J.M., Jansen E.S., Morrissy C.J.,van der Heide B., Goff W.V., WilliamsonM.M., Hooper P.T., Babiuk L.A., Tikoo S.K.,Johnson M.A., Vaccination of pigs with arecombinant porcine adenovirus expressingthe gD gene from pseudorabies virus, Vac-cine 19 (2001) 37523758.

[64] Harokopakis E., Hajishengallis G., MichalekS.M., Effectiveness of liposomes possessingsurface-linked recombinant B subunit ofcholera toxin as an oral antigen delivery sys-tem, Infect. Immun. 66 (1998) 42994304.

[65] Hebben M., Duquesne V., Cronier J., RossiB., Aubert A., Modified vaccinia virusAnkara as a vaccine against feline coronavi-rus: immunogenicity and efficacy, J. FelineMed. Surg. 6 (2004) 111118.

[66] Heine H.G., Boyle D.B., Infectious bursaldisease virus structural protein VP2expressed by a fowlpox virus recombinantconfers protection against disease in chick-ens, Arch. Virol. 131 (1993) 277292.

[67] Herrmann J.E., Chen S.C., Jones D.H.,Tinsley-Bown A., Fynan E.F., GreenbergH.B., Farrar G.H., Immune responses andprotection obtained by oral immunizationwith rotavirus VP4 and VP7 DNA vaccinesencapsulated in microparticles, Virology 259(1999) 148153.

[68] Hoebe K., Janssen E., Beutler B., The inter-face between innate and adaptive immunity,Nat. Immunol. 5 (2004) 971974.

[69] Holmgren J., Czerkinsky C., Mucosal immu-nity and vaccines, Nat. Med. 11 (2005) S45S53.

[70] Hu K.F., Elvander M., Merza M., AkerblomL., Brandenburg A., Morein B., The immu-nostimulating complex (ISCOM) is an effi-cient mucosal delivery system for respiratorysyncytial virus (RSV) envelope antigensinducing high local and systemic antibodyresponses, Clin. Exp. Immunol. 113 (1998)235243.

[71] Huang Z., Elankumaran S., Yunus A.S.,Samal S.K., A recombinant Newcastle dis-ease virus (NDV) expressing VP2 protein ofinfectious bursal disease virus (IBDV) pro-tects against NDV and IBDV, J. Virol. 78(2004) 1005410063.

[72] Hussain N., Florence A.T., Utilizing bacte-rial mechanisms of epithelial cell entry: inva-sin-induced oral uptake of latex nanoparti-cles, Pharm. Res. 15 (1998) 153156.

[73] Jepson M.A., Clark M.A., Foster N., MasonC.M., Bennett M.K., Simmons N.L., HirstB.H., Targeting to intestinal M cells, J. Anat.189 (1996) 507516.

[74] Johansson-Lindbom B., Svensson M., WurbelM.A., Malissen B., Marquez G., Agace W.,Selective generation of gut tropic T cells ingut-associated lymphoid tissue (GALT):requirement for GALT dendritic cells andadjuvant, J. Exp. Med. 198 (2003) 963969.

[75] Johnson M.A., Pooley C., Ignjatovic J.,Tyack S.G., A recombinant fowl adenovirusexpressing the S1 gene of infectious bronchi-tis virus protects against challenge withinfectious bronchitis virus, Vaccine 21(2003) 27302736.

[76] Jones D.H., McBride B.W., Thornton C.,OHagan D.T., Robinson A., Farrar G.H.,


Orally administered microencapsulated Bor-detella pertussis fimbriae protect mice fromB. pertussis respiratory infection, Infect.Immun. 64 (1996) 489494.

[77] Joseph A., Louria-Hayon I., Plis-Finarov A.,Zeira E., Zakay-Rones Z., Raz E., HayashiT., Takabayashi K., Barenholz Y., Kedar E.,Liposomal immunostimulatory DNA sequence(ISS-ODN): an efficient parenteral andmucosal adjuvant for influenza and hepatitisB vaccines, Vaccine 20 (2002) 33423354.

[78] Kelsall B.L., Rescigno M., Mucosal dendriticcells in immunity and inflammation, Nat.Immunol. 5 (2004) 10911095.

[79] Kesik M., Saczynska V., Szewczyk B.,Plucienniczak A., Inclusion bodies fromrecombinant bacteria as a novel system fordelivery of vaccine antigen by the oral route,Immunol. Lett. 91 (2004) 197204.

[80] Kidane A., Guimond P., Ju T.R., Sanchez M.,Gibson J., Bowersock T.L., The efficacy oforal vaccination of mice with alginate encap-sulated outer membrane proteins of Pas-teurella haemolytica and One-Shot, Vaccine19 (2001) 26372646.

[81] Kim B., Bowersock T., Griebel P., KidaneA., Babiuk L.A., Sanchez M., Attah-PokuS., Kaushik R.S., Mutwiri G.K., Mucosalimmune responses following oral immuniza-tion with rotavirus antigens encapsulated inalginate microspheres, J. Control. Release 85(2002) 191202.

[82] Kit M., Kit S., Little S.P., Di Marchi R.D.,Gale C., Bovine herpesvirus-1 (infectiousbovine rhinotracheitis virus)-based viral vec-tor which expresses foot-and-mouth diseaseepitopes, Vaccine 9 (1991) 564572.

[83] Kit S., Kit M., DiMarchi R.D., Little S.P.,Gale C., Modified-live infectious bovine rhi-notracheitis virus vaccine expressing mono-mer and dimer forms of foot-and-mouthdisease capsid protein epitopes on surfaceof hybrid virus particles, Arch. Virol. 120(1991) 117.

[84] Kit S., Otsuka H., Kit M., Expression of por-cine pseudorabies virus genes by a bovineherpesvirus-1 (infectious bovine rhinotra-cheitis virus) vector, Arch. Virol. 124 (1992)120.

[85] Kunkel E.J., Butcher E.C., Plasma-cell hom-ing, Nat. Rev. Immunol. 3 (2003) 822829.

[86] Kweon C.H., Kang S.W., Choi E.J., KangY.B., Bovine herpes virus expressing enve-lope protein (E2) of bovine viral diarrheavirus as a vaccine candidate, J. Vet. Med. Sci.61 (1999) 395401.

[87] Lazarus N.H., Kunkel E.J., Johnston B., WilsonE., Youngman K.R., Butcher E.C., A com-mon mucosal chemokine (mucosae-associ-

ated epithelial chemokine/CCL28) selec-tively attracts IgA plasmablasts, J. Immunol.170 (2003) 37993805.

[88] Lee J.S., Hadjipanayis A.G., Welkos S.L.,Venezuelan equine encephalitis virus-vec-tored vaccines protect mice against anthraxspore challenge, Infect. Immun. 71 (2003)14911496.

[89] Liao C.W., Chiou H.Y., Yeh K.S., Chen J.R.,Weng C.N., Oral immunization using forma-lin-inactivated Actinobacillus pleuropneu-moniae antigens entrapped in microsphereswith aqueous dispersion polymers preparedusing a co-spray drying process, Prev. Vet.Med. 61 (2003) 115.

[90] Lin J.H., Weng C.N., Liao C.W., Yeh K.S.,Pan M.J., Protective effects of oral microen-capsulated Mycoplasma hyopneumoniaevaccine prepared by co-spray drying method,J. Vet. Med. Sci. 65 (2003) 6974.

[91] Liu W., Yang Y., Chung N., Kwang J., Induc-tion of humoral immune response and pro-tective immunity in chickens against Salmo-nella enteritidis after a single dose of killedbacterium-loaded microspheres, Avian Dis.45 (2001) 797806.

[92] Liu Y.-J., IPC: Professional type i interferon-producing cells and plasmacytoid dendriticcell precursors, Ann. Rev. Immunol. 23(2005) 275306.

[93] Locht C., Live bacterial vectors for intranasaldelivery of protective antigens, Pharm. Sci.Technol. Today 3 (2000) 121128.

[94] Loessner H., Weiss S., Bacteria-mediatedDNA transfer in gene therapy and vaccina-tion, Exp. Opin. Biol. Ther. 4 (2004) 157168.

[95] Lovgren K., Kaberg H., Morein B., An exper-imental influenza subunit vaccine (ISCOM):induction of protective immunity to chal-lenge infection in mice after intranasal orsubcutaneous administration, Clin. Exp.Immunol. 82 (1990) 435439.

[96] MacPherson G., Milling S., Yrlid U., CousinsL., Turnbull E., Huang F.P., Uptake of anti-gens from the intestine by dendritic cells,Ann. NY Acad. Sci. 1029 (2004) 7582.

[97] Maloy K.J., Donachie A.M., Mowat A.M.,Induction of Th1 and Th2 CD4+ T cellresponses by oral or parenteral immunizationwith ISCOMS, Eur. J. Immunol. 25 (1995)28352841.

[98] McGreal E.P., Martinez-Pomares L., GordonS., Divergent roles for C-type lectinsexpressed by cells of the innate immune sys-tem, Mol. Immunol. 41 (2004) 11091121.

[99] McKenna K., Beignon A.S., Bhardwaj N.,Plasmacytoid dendritic cells: linking innateand adaptive immunity, J. Virol. 79 (2005)1727.


[100] McKenzie B.S., Corbett A.J., Johnson S.,Brady J.L., Pleasance J., Kramer D.R., BoyleJ.S., Jackson D.C., Strugnell R.A., LewA.M., Bypassing luminal barriers, deliveryto a gut addressin by parenteral targetingelicits local IgA responses, Int. Immunol. 16(2004) 16131622.

[101] Medina E., Guzman C.A., Use of live bacte-rial vaccine vectors for antigen delivery:potential and limitations, Vaccine 19 (2001)15731580.

[102] Men Y., Audran R., Thomasin C., Eberl G.,Demotz S., Merkle H.P., Gander B., Corra-din G., MHC class I- and class II-restrictedprocessing and presentation of microencap-sulated antigens, Vaccine 17 (1999) 10471056.

[103] Meurens F., Keil G.M., Muylkens B., GogevS., Schynts F., Negro S., Wiggers L., ThiryE., Interspecific recombination between tworuminant alphaherpesviruses, bovine her-pesviruses 1 and 5, J. Virol. 78 (2004) 98289836.

[104] Michalek S., OHagan D., Gould-FogeriteS., Rimmelzwaan G., Osterhaus A., Antigendelivery systems: Non-living microparticles,liposomes, cochleates and ISCOMs, Aca-demic Press, San Diego, 1999, pp. 759778.

[105] Minke J.M., Siger L., Karaca K., Austgen L.,Gordy P., Bowen R., Renshaw R.W., Loos-more S., Audonnet J.C., Nordgren B.,Recombinant canarypoxvirus vaccine carry-ing the prM/E genes of West Nile virus pro-tects horses against a West Nile virus-mos-quito challenge, Arch. Virol. Suppl. (2004)221230.

[106] Mishima M., Xuan X., Yokoyama N., IgarashiI., Fujisaki K., Nagasawa H., Mikami T.,Recombinant feline herpesvirus type 1expressing Toxoplasma gondii ROP2 anti-gen inducible protective immunity in cats,Parasitol. Res. 88 (2002) 144149.

[107] Moore R.J., Rothel L., Krywult J., RadfordA.J., Lund K., Hodgson A.L., Foreign geneexpression in Corynebacterium pseudotu-berculosis: development of a live vaccinevector, Vaccine 18 (1999) 487497.

[108] Moore R.J., Stewart D.J., Lund K., HodgsonA.L., Vaccination against ovine footrotusing a live bacterial vector to deliver basicprotease antigen, FEMS Microbiol. Lett. 194(2001) 193196.

[109] Mora J.R., Bono M.R., Manjunath N.,Weninger W., Cavanagh L.L., RosemblattM., Von Andrian U.H., Selective imprintingof gut-homing T cells by Peyers patch den-dritic cells, Nature 424 (2003) 8893.

[110] Morein B., Villacres-Eriksson M., Lovgren-Bengtsson K., Iscom, a delivery system forparenteral and mucosal vaccination, Dev.Biol. Stand. 92 (1998) 3339.

[111] Morein B., Hu K.F., Abusugra I., Currentstatus and potential application of ISCOMsin veterinary medicine, Adv. Drug Deliv.Rev. 56 (2004) 13671382.

[112] Morris-Downes M.M., Sheahan B.J., FleetonM.N., Liljestrom P., Reid H.W., Atkins G.J.,A recombinant Semliki Forest virus particlevaccine encoding the prME and NS1 pro-teins of louping ill virus is effective in asheep challenge model, Vaccine 19 (2001)38773884.

[113] Mowat A.M., Smith R.E., Donachie A.M.,Furrie E., Grdic D., Lycke N., Oral vaccina-tion with immune stimulating complexes,Immunol. Lett. 65 (1999) 133140.

[114] Murillo M., Grillo M.J., Rene J., MarinC.M., Barberan M., Goni M.M., Blasco J.M.,Irache J.M., Gamazo C., A Brucella ovisantigenic complex bearing poly-epsilon-caprolactone microparticles confer protec-tion against experimental brucellosis inmice, Vaccine 19 (2001) 40994106.

[115] Niess J.H., Brand S., Gu X., Landsman L.,Jung S., McCormick B.A., Vyas J.M., BoesM., Ploegh H.L., Fox J.G., Littman D.R.,Reinecker H.C., CX3CR1-mediated den-dritic cell access to the intestinal lumen andbacterial clearance, Science 307 (2005) 254258.

[116] OHagan D.T., The intestinal uptake of par-ticles and the implications for drug and anti-gen delivery, J. Anat. 189 (1996) 477482.

[117] OHagan D.T., Microparticles and polymersfor the mucosal delivery of vaccines, Adv.Drug Deliv. Rev. 34 (1998) 305320.

[118] Orr N., Galen J.E., Levine M.M., Expressionand immunogenicity of a mutant diphtheriatoxin molecule, CRM(197), and its frag-ments in Salmonella typhimurium vaccinestrain CVD 908-htrA, Infect. Immun. 67(1999) 42904294.

[119] Pabst O., Ohl L., Wendland M., WurbelM.A., Kremmer E., Malissen B., Forster R.,Chemokine receptor CCR9 contributes tothe localization of plasma cells to the smallintestine, J. Exp. Med. 199 (2004) 411416.

[120] Papadakis K.A., Prehn J., Nelson V., ChengL., Binder S.W., Ponath P.D., Andrew D.P.,Targan S.R., The role of thymus-expressedchemokine and its receptor CCR9 on lym-phocytes in the regional specialization of themucosal immune system, J. Immunol. 165(2000) 50695076.


[121] Pardo M.C., Bauman J.E., Mackowiak M.,Protection of dogs against canine distemperby vaccination with a canarypox virusrecombinant expressing canine distempervirus fusion and hemagglutinin glycopro-teins, Am. J. Vet. Res. 58 (1997) 833836.

[122] Payne L.G., Andrianov A.K., Protein releasefrom polyphosphazene matrices, Adv. DrugDeliv. Rev. 31 (1998) 185196.

[123] Payne L.G., Jenkins S.A., Andrianov A.,Roberts B.E., Water-soluble phosphazenepolymers for parenteral and mucosal vaccinedelivery, Pharm. Biotechnol. 6 (1995) 473493.

[124] Phenix K.V., Wark K., Luke C.J., SkinnerM.A., Smyth J.A., Mawhinney K.A., ToddD., Recombinant Semliki Forest virus vectorexhibits potential for avian virus vaccinedevelopment, Vaccine 19 (2001) 31163123.

[125] Pogonka T., Klotz C., Kovacs F., Lucius R.,A single dose of recombinant Salmonellatyphimurium induces specific humoralimmune responses against heterologousEimeria tenella antigens in chicken, Int. J.Parasitol. 33 (2003) 8188.

[126] Prevec L., Campbell J.B., Christie B.S.,Belbeck L., Graham F.L., A recombinanthuman adenovirus vaccine against rabies, J.Infect. Dis. 161 (1990) 2730.

[127] Raulet D.H., Interplay of natural killer cellsand their receptors with the adaptive immuneresponse, Nat. Immunol. 5 (2004) 9961002.

[128] Rebelatto M.C., Guimond P., BowersockT.L., HogenEsch H., Induction of systemicand mucosal immune response in cattle byintranasal administration of pig serum albu-min in alginate microparticles, Vet. Immu-nol. Immunopathol. 83 (2001) 93105.

[129] Reddy P.S., Idamakanti N., Pyne C.,Zakhartchouk A.N., Godson D.L., Papp Z.,Baca-Estrada M.E., Babiuk L.A., MutwiriG.K., Tikoo S.K., The immunogenicity andefficacy of replication-defective and replica-tion-competent bovine adenovirus-3 express-ing bovine herpesvirus-1 glycoprotein gD incattle, Vet. Immunol. Immunopathol. 76(2000) 257268.

[130] Reddy S.K., Sharma J.M., Ahmad J., ReddyD.N., McMillen J.K., Cook S.M., WildM.A., Schwartz R.D., Protective efficacy ofa recombinant herpesvirus of turkeys as an inovo vaccine against Newcastle and Mareksdiseases in specific-pathogen-free chickens,Vaccine 14 (1996) 469477.

[131] Rescigno M., Identification of a new mech-anism for bacterial uptake at mucosal sur-

faces, which is mediated by dendritic cells,Pathol. Biol. 51 (2003) 6970.

[132] Rescigno M., Chieppa M., Gut-level deci-sions in peace and war, Nat. Med. 11 (2005)254255.

[133] Roland K.L., Tinge S.A., Killeen K.P.,Kochi S.K., Recent advances in the develop-ment of live, attenuated bacterial vectors,Curr. Opin. Mol. Ther. 7 (2005) 6272.

[134] Roque-Resendiz J.L., Rosales R., Herion P.,MVA ROP2 vaccinia virus recombinant as avaccine candidate for toxoplasmosis, Parasi-tology 128 (2004) 397405.

[135] Rothel J.S., Boyle D.B., Both G.W., PyeA.D., Waterkeyn J.G., Wood P.R., Light-owlers M.W., Sequential nucleic acid andrecombinant adenovirus vaccination induceshost-protective immune responses againstTaenia ovis infection in sheep, ParasiteImmunol. 19 (1997) 221227.

[136] Roth-Walter F., Scholl I., Untersmayr E.,Fuchs R., Boltz-Nitulescu G., WeissenbockA., Scheiner O., Gabor F., Jensen-Jarolim E.,M cell targeting with Aleuria aurantia lectinas a novel approach for oral allergen immu-notherapy, J. Allergy Clin. Immunol. 114(2004) 13621368.

[137] Roth-Walter F., Scholl I., Untersmayr E.,Ellinger A., Boltz-Nitulescu G., Scheiner O.,Gabor F., Jensen-Jarolim E., Mucosal target-ing of allergen-loaded microspheres byAleuria aurantia lectin, Vaccine 23 (2005)27032710.

[138] Schrijver R.S., Langedijk J.P., Keil G.M.,Middel W.G., Maris-Veldhuis M., VanOirschot J.T., Rijsewijk F.A., Immunizationof cattle with a BHV1 vector vaccine or aDNA vaccine both coding for the G proteinof BRSV, Vaccine 15 (1997) 19081916.

[139] Schwantes A., Truyen U., Weikel J., WeissC., Lochelt M., Application of chimericfeline foamy virus-based retroviral vectorsfor the induction of antiviral immunity incats, J. Virol. 77 (2003) 78307842.

[140] Sheppard M., Viral vectors for veterinaryvaccines, Adv. Vet. Med. 41 (1999) 145161.

[141] Sheppard M., Werner W., Tsatas E., McCoyR., Prowse S., Johnson M., Fowl adenovirusrecombinant expressing VP2 of infectiousbursal disease virus induces protectiveimmunity against bursal disease, Arch.Virol. 143 (1998) 915930.

[142] Shiau A.L., Chen Y.L., Liao C.Y., HuangY.S., Wu C.L., Prothymosin alpha enhancesprotective immune responses induced byoral DNA vaccination against pseudorabies


delivered by Salmonella choleraesuis, Vac-cine 19 (2001) 39473956.

[143] Shiau A.L., Chu C.Y., Su W.C., Wu C.L.,Vaccination with the glycoprotein D gene ofpseudorabies virus delivered by nonpatho-genic Escherichia coli elicits protectiveimmune responses, Vaccine 19 (2001)32773284.

[144] Shimoji Y., Oishi E., Kitajima T., MunetaY., Shimizu S., Mori Y., Erysipelothrix rhu-siopathiae YS-1 as a live vaccine vehicle forheterologous protein expression and intrana-sal immunization of pigs, Infect. Immun. 70(2002) 226232.

[145] Singh M., Vajdy M., Gardner J., Briones M.,OHagan D., Mucosal immunization withHIV-1 gag DNA on cationic microparticlesprolongs gene expression and enhances localand systemic immunity, Vaccine 20 (2001)594602.

[146] Sirard J.C., Weber M., Duflot E., PopoffM.R., Mock M., A recombinant Bacillusanthracis strain producing the Clostridiumperfringens Ib component induces protec-tion against iota toxins, Infect. Immun. 65(1997) 20292033.

[147] Sizemore D.R., Branstrom A.A., Sadoff J.C.,Attenuated Shigella as a DNA delivery vehi-cle for DNA-mediated immunization, Sci-ence 270 (1995) 299302.

[148] Sizemore D.R., Branstrom A.A., Sadoff J.C.,Attenuated bacteria as a DNA delivery vehi-cle for DNA-mediated immunization, Vac-cine 15 (1997) 804807.

[149] Smith R.E., Donachie A.M., Grdic D., LyckeN., Mowat A.M., Immune-stimulating com-plexes induce an IL-12-dependent cascadeof innate immune responses, J. Immunol.162 (1999) 55365546.

[150] Snoeck V., Huyghebaert N., Cox E., VermeireA., Vancaeneghem S., Remon J.P., GoddeerisB.M., Enteric-coated pellets of F4 fimbriaefor oral vaccination of suckling pigletsagainst enterotoxigenic Escherichia coliinfections, Vet. Immunol. Immunopathol. 96(2003) 219227.

[151] Stocker B.A., Aromatic-dependent salmo-nella as anti-bacterial vaccines and as pre-senters of heterologous antigens or of DNAencoding them, J. Biotechnol. 83 (2000)4550.

[152] Su G.F., Brahmbhatt H.N., de Lorenzo V.,Wehland J., Timmis K.N., Extracellularexport of Shiga toxin B-subunit/haemolysinA (C-terminus) fusion protein expressed inSalmonella typhimurium aroA-mutant andstimulation of B-subunit specific antibody

responses in mice, Microb. Pathog. 13(1992) 465476.

[153] Supply P., Sutton P., Coughlan S.N., Bilo K.,Saman E., Trees A.J., Cesbron Delauw M.F.,Locht C., Immunogenicity of recombinantBCG producing the GRA1 antigen from Toxo-plasma gondii, Vaccine 17 (1999) 705714.

[154] Tacket C.O., Reid R.H., Boedeker E.C.,Losonsky G., Nataro J.P., Bhagat H., Edel-man R., Enteral immunization and challengeof volunteers given enterotoxigenic E. coliCFA/II encapsulated in biodegradablemicrospheres, Vaccine 12 (1994) 12701274.

[155] Tana Watarai S., Isogai E., Oguma K.,Induction of intestinal IgA and IgG antibod-ies preventing adhesion of verotoxin-pro-ducing Escherichia coli to Caco-2 cells byoral immunization with liposomes, Lett.Appl. Microbiol. 36 (2003) 135139.

[156] Taylor G., Rijsewijk F.A., Thomas L.H.,Wyld S.G., Gaddum R.M., Cook R.S.,Morrison W.I., Hensen E., van Oirschot J.T.,Keil G., Resistance to bovine respiratorysyncytial virus (BRSV) induced in calves bya recombinant bovine herpesvirus-1 express-ing the attachment glycoprotein of BRSV, J.Gen. Virol. 79 (1998) 17591767.

[157] Thiry E., Meurens F., Muylkens B., McVoyM., Gogev S., Thiry J., Vanderplasschen A.,Epstein A., Keil G., Schynts F., Recombina-tion in alphaherpesviruses, Rev. Med. Virol.15 (2005) 89103.

[158] Thole J.E., van Dalen P.J., Havenith C.E.,Pouwels P.H., Seegers J.F., Tielen F.D., vander Zee M.D., Zegers N.D., Shaw M., Livebacterial delivery systems for developmentof mucosal vaccines, Curr. Opin. Mol. Ther.2 (2000) 9499.

[159] Torche A.M., Jouan H., Le Corre P., AlbinaE., Primault R., Jestin A., Le Verge R., Exvivo and in situ PLGA microspheres uptakeby pig ileal Peyers patch segment, Int.

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