Protection efficacy of the Brucella abortus ghost vaccine candidate lysed by the N-terminal 24-amino acid fragment (GI24) of the 36-amino acid peptide PMAP-36 (porcine myeloid antimicrobial peptide 36) in murine models

Bovine brucellosis, caused by a gram-negative facultative intracellular pathogen, Brucellaabortus, is an important zoonotic disease worldwide [2, 26, 34]. Brucellosis in cattle causes infertility, abortion, and production of contagious milk, thus resulting in major economic losses [5, 15]. It also affects humans and causes headache, fever, arthritis, and chronic fatigue; however, this infection does not spread among humans [16, 17, 50]. A number of strategies, including promotion of awareness, improvement of hygiene standards and the use of vaccines, have been implemented to prevent the spread of bovine brucellosis, because of its high socioeconomic impact [2, 34]. Currently, only live attenuated vaccines are available for prevention of brucellosis in livestock. Among them, B. abortus strain S19 and B. abortus strain RB51 (strain RB51) are widely used [2, 34]. B. abortus strain 19 has been effective in preventing abortion and controlling brucellosis in adult cattle. It also helped to decrease the prevalence of the disease in a herd [2, 35]. However, B. abortus strain 19 does not discriminate between infected and vaccinated animals. In addition, there is a low risk of abortion in livestock [2, 34, 35]. The live attenuated strain RB51 is an alternative to the B. abortus strain 19 vaccine. The strain RB51 vaccine is less abortifacient and virulent. Furthermore, it does not induce an antibody response in the standard serological diagnostic tests. It is also safe to use in calves elder than 3 months [2, 34, 35]. Nevertheless, vaccination of pregnant cows with strain RB51 carries a low risk of abortion or premature birth. Thus, it is recommended to be used with caution in pregnant cattle [2, 26, 29, 34, 35]. Although a live attenuated vaccine is a common practice for prevention of brucellosis, it also poses high risks due to the potential ability to revert to virulence and to cause abortion and because of shedding in milk, urine, semen or fecal matter, thus infecting the humans coming into contact with the animals. Hence, many different approaches, such as killed vaccines, subunit vaccines, recombinant proteins and vector vaccines, have been tried against brucellosis with varying degrees of success [2, 29]. In the past few years, bacterial lysates have emerged as an effective inactivated nonliving vaccine against a wide variety of gram-negative bacteria. Bacterial cell lysates constitute empty, nonliving bacterial envelops of gram-negative bacteria with intact cellular morphology, including cell surface structures, but lacking cytoplasmic content [27]. Host defense peptides (HDPs) or antimicrobial peptides (AMPs) are a part of the innate immune system [8, 9]. These peptides have a diverse range of activities against gram-positive as well as gram-negative bacteria [45], parasites [19], and enveloped viruses [12]. The mechanism of action of HDPs is disruption of membrane barrier function by pore formation or induction of membrane permeabilization, without disturbing integrity of the membrane [18, 20, 46, 47]. Until now, 11 porcine AMPs have been reported [40]. Porcine myeloid antimicrobial peptide 36 (PMAP-36) has the highest positive charge among all the porcine cathelicidins. It may be advantageous, because PMAP-36’s binding to the bacterial cell membrane is mediated by the positive charge of the peptide and the negatively charged molecules at the surface of the bacterial cell membrane through electrostatic interactions [3]. In particular, in the 36-amino acid (aa) sequence of PMAP-36, the N-terminal α-helical domain consists of 24 aa (GI24), and GI24 can also penetrate the bacterial membrane like PMAP-36 can [3, 12, 19]. The aim of the present study was to compare the ability of bacteria lysed by GI24 to induce a cellular immune response and a humoral immune response between mice immunized orally and mice immunized intraperitoneally (ip). Another objective of this study was to compare the protection efficacy of the B.abortus vaccine candidate constructed via lysis of B. abortus biotype 1 isolate from Korean cattle by means of GI24 with that of Brucellaabortus strain RB51 vaccine in a mouse model.