Isolation of <Emphasis Type="Italic">Brucella melitensis</Emphasis> from a RB51-vaccinated seronegative goat

The purpose of this study is to determine the etiology of abortions presented in a goat herd declared as free of brucellosis and vaccinated with RB51 located in Mexico. The serological diagnosis of brucellosis in 33 animals was performed. The study included three goats that aborted in the last third of gestation and 15 goats that gave birth normally; samples of milk and vaginal exudate were subjected to bacteriological study. All animals were negative for serological diagnosis, and isolation of Brucella melitensis was achieved in a single goat from vaginal exudate. However, the particularity is that this goat was negative to the card, indirect ELISA, and radial immunodiffusion tests. Isolation of a field strain was confirmed by biochemical test resistance to rifampicin and PCR. It is concluded that a goat which aborted in the last third of gestation was found spreading B. melitensis through vaginal discharge despite being vaccinated with RB51 and seronegative for brucellosis.     

Isolation and identification of bovine Brucella isolates from Pakistan by biochemical tests and PCR

Brucellosis is endemic in bovines in Pakistan. The Brucella species and biovars involved, however, are unknown. The objectives of the present study were to isolate and characterize brucellae from seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes which had recently aborted. The seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes were collected from the Potohar Plateau, Pakistan. Isolation of brucellae was done on modified Farrell’s serum dextrose agar. Isolates were characterized by conventional biotyping methods, while molecular typing was done by genus (B4/B5) and species-specific (Brucella abortus, Brucella melitensis, Brucella ovis, and Brucella suis) polymerase chain reaction (PCR). A total of 30 isolates were recovered from milk (n?=?5), aborted fetuses (n?=?13), and vaginal swabs (n?=?12). Most isolates were from cattle (56.7 %). All of them were identified as B. abortus biovar 1 based on conventional biotyping methods and genus and species-specific PCR. This preliminary study provides the first report on the prevalence of B. abortus biovar 1 in cattle and buffaloes in Pakistan.     

Use of serology and real time PCR to control an outbreak of bovine brucellosis at a dairy cattle farm in the Nile Delta region,Egypt

Background

Bovine brucellosis remains one of the most prevalent zoonotic infections affecting dairy cattle in developing countries where the applied control programs often fail. We analyzed the epidemiologic pattern of bovine brucellosis in a dairy cattle herd that showed several cases of abortions after regular vaccination with RB51 (B. abortus vaccine). In 2013 thirty dairy cows, from a Holstein-Friesian dairy herd with a population of 600 cattle, aborted five months post vaccination by a regular RB51 vaccine. Blood samples were drawn from milking cows and growing heifers, as well as heifers and cows pregnant up to 6 months. These samples were collected in June 2013 (n?=?257) and May 2014 (n?=?263) and were tested by real time (rt)-PCR as well as serological tests, in particular Rose Bengal Test (RBT), Enzyme-Linked Immunosorbent Assays (ELISA) and Fluorescence Polarization Assay. Tissue specimens were also collected from an aborted fetus and cultured. Isolates were subjected to bacteriological typing tests at the genus and species levels.

Results

Five months post vaccination with RB51 vaccine, Brucella (B.) DNA was detected in blood samples of cows by rt-PCR. The serological tests also revealed the spread of Brucella field strains within the herd in 2013. Four Brucella isolates were recovered from specimens collected from the aborted fetus. These isolates were typed as follows: one B. abortus RB51 vaccine strain and three isolates of B. abortus field strain. The seropositive cows with positive rt-PCR might indicate an infection by the Brucella field strain; while the positive rt-PCR results from seronegative animals may either be due to circulating RB51 vaccine DNA in vaccinated animals or to circulating field strain in infected animals before seroconversion.

Conclusion

The results herein suggest that PCR can be a good supplementary tool in an outbreak situation, if an assay is available that can differentiate vaccine and field strains with a high analytical sensitivity. We recommend using RBT and ELISA in parallel in outbreak situations, to identify as many infected animals as possible during the initial screenings. This test procedure should be repeated for at least three successive negative tests, with one month interval.

     

Monitoring clinical outcomes,pathological changes and shedding of Chlamydophila abortus following experimental challenge of periparturient ewes utilizing the natural route of infection

Enzootic abortion of ewes (EAE) caused by Chlamydophila abortus is an important disease resulting in significant lamb loss in most sheep producing countries. Ewes are considered to be naturally infected with C. abortus via the oral–nasal route and may become persistent carriers, shedding during subsequent oestrous cycles and at lambing. The aim of this study was to monitor the clinical outcomes, pathological changes and shedding of C. abortus in 18 periparturient orally infected sheep for two breeding seasons. In the first season, C. abortus was detected by real-time PCR (rt-PCR) in 13/18 conjunctival swabs at oestrus. Three out of the 15 pregnant ewes gave birth to 1 live and 1 dead lamb, and 2 of them aborted. Following parturition/abortion, C. abortus was detected in 12/15 vaginal swabs and in all the collected foetal membranes. However, only those membranes containing high copy numbers of the bacterium displayed the EAE typical lesions. In the second season, none of the 13 pregnant ewes aborted, and 5 of them gave birth to dead or weak lambs. C. abortus was not detected in conjunctival or vaginal swabs at oestrus or parturition. The bacterium was detected at low levels in 36% of the foetal membranes, but with no evidence of histopathological lesions. These results indicate that C. abortus can be detected in a large proportion of animals during the first pregnancy after oral infection. However, this proportion is reduced at the subsequent breeding season, confirming the occurrence of a chronic low level persistent infection in post-abortion/lambing ewes.     

The first report in new zealand of Lymnaea columella say (Mollusca: gastropoda) an intermediate host of the liver fluke Fasciola hepatica L

Six herds of dairy cattle, in which infection had persisted after measures had been employed to eradicate brucellosis, were investigated in detail. One hundred and two animals out of 700 (14.6%) had evidence of the disease from cultural or serological tests. Only 4 infected animals aborted; the remaining 98 animals with brucellosis exhibited no clinical sign of the disease, and 52 were heifers calving for the first time. Sixty-five of the 700 animals (9.3%) produced brucella-infected vaginal discharges at the end of a normal period of gestation, and another 17 (2.4%) were detected only by the brucellosis (Brewer) card test (BCT) or complement fixation test (CFT). All the infected animals were removed from the herds immediately after detection.

About 2 weeks after all the cows had calved, 20 of the remain-ing 510 animals (3.9%) in 4 of the herds became positive to one or more tests and 8 excreted brucellae in their milk. The pregnancy of three 2-year-old primiparous heifers terminated normally and one aborted. All four discharged brucellae until they were slaughtered 20 to 35 days after parturition.

Twenty-five of the 102 infected animals were detected by bacteriological, and 29 by serological, means only. A compari-son was made of the results of the testsfrom 73 culturally positive animals; 4.1% of the sera were CFT positive but BCT negative, and 5.5% were BCT positive but CFT negative. An attempt has been made to explain the large numbers (34.2%) of infected animals that were serologically negative.     

The role of TREM-2 in internalization and intracellular survival of Brucella abortus in murine macrophages

Triggering receptor expressed on myeloid cells-2 (TREM-2) is a cell surface receptor primarily expressed on macrophages and dendritic cells. TREM-2 functions as a phagocytic receptor for bacteria as well as an inhibitor of Toll like receptors (TLR) induced inflammatory cytokines. However, the role of TREM-2 in Brucella intracellular growth remains unknown. To investigate whether TREM-2 is involved in Brucella intracellular survival, we chose bone marrow derived macrophages (BMDMs), in which TREM-2 is stably expressed, as cell model. Colony formation Units (CFUs) assay suggests that TREM-2 is involved in the internalization of Brucella abortus (B. abortus) by macrophages, while silencing of TREM-2 decreases intracellular survival of B. abortus. To further study the underlying mechanisms of TREM-2-mediated bacterial intracellular survival, we examined the activation of B. abortus-infected macrophages through determining the kinetics of activation of the three MAPKs, including ERK, JNK and p38, and measuring TNFα production in response to lipopolysaccharide (LPS) of Brucella (BrLPS) or B. abortus stimulation. Our data show that TREM-2 deficiency promotes activation of Brucella-infected macrophages. Moreover, our data also demonstrate that macrophage activation promotes killing of Brucella by enhancing nitric oxygen (NO), but not reactive oxygen species (ROS) production, macrophage apoptosis or cellular death. Taken together, these findings provide a novel interpretation of Brucella intracellular growth through inhibition of NO production produced by TREM-2-mediated activated macrophages.     

Serology of a Neospora abortion outbreak on a dairy farm in New Zealand: a case study

AIM: To describe the kinetics of serological titres after an abortion outbreak in April-May 1995 due to Neospora caninum affected 17 dairy cows in a herd of 320. METHODS: Thirty-five cows, that had either aborted, carried mummified calves, were not pregnant or calved normally were: bled several times at regular intervals and the sera tested for Neospora antibodies in the indirect fluorescence antibody test (IFAT). RESULTS: Maximal IFAT titres of up to 1:4000 occurred within 6 weeks of the abortion outbreak, decreased over the next 2 months to < or = 1:200 and remained at this level until the next scheduled bleed a further 2 months later. A rise in titres was subsequently observed in the cows that had aborted or were not pregnant (at the time of the abortions) or had carried mummified foetuses. Seroconversion was also observed in some of the control cows, which had, up until then, remained seronegative. A dog and cat in contact with the cows in the herd investigated were, however, negative in the IFAT. CONCLUSIONS: Maximal serological titres in Neospora abortions are observed within weeks of the abortion event and then quickly return to very low levels. Subsequently, a recrudescence of titres can be observed in infected cows during the next pregnancy, without it being associated with repeat abortions.     

Prevalence and molecular identification of Chlamydia abortus in commercial dairy goat farms in a hot region in Mexico

The aim of this study was to determine the seroprevalence and presence of Chlamydia abortus in Saanen breed female goats from commercial dairy goat farms under intensive production in the municipality of Guanajuato, Mexico. Sera were collected to determine the prevalence of anti-C. abortus IgG antibodies using recombinant enzyme-linked immunosorbent assay (rELISA) and cell culture. Polymerase chain reaction (PCR) was used to prove the presence of the pathogen in swab samples collected from the vagina and rectum of selected animals. Additionally, foetal tissue samples from a sudden abortion were collected. C. abortus prevalence in female goats of commercial milking farms sampled in Guanajuato, Mexico, was 4.87 % (n?=?246). Seropositive animals were found in six out of nine (66.6 %) dairy goat farms sampled, and prevalence among animals in individual farms ranged between 3.44 and 13.51 %. C. abortus was detected using PCR in spleen tissue from the aborted foetus. PCR-based detection, as well as isolation from vaginal and rectal swabs, was not possible in the present study. Isolation through cell culture was also unsuccessful from aborted foetal tissue samples. In conclusion, the results from rELISA and PCR show that C. abortus is present in dairy goat farms in the state of Guanajuato, Mexico.     

Investigation of abortions in brucellosis tested herds

The mandatory reporting and investigation of bovine abortions provides a system for the surveillance of the progress of the Brucellosis Eradication Scheme and a warning of resurgence of brucellosis in tested herds. Before the scheme was introduced in 1971, about a quarter of all abortions were caused by Brucella abortus. By 1976, the percentage of abortions due to Brucella in herds under test for brucellosis had been reduced to 3.4%. Most Brucella abortions occurred during the seventh and eighth months of gestation. Cows in the 4- to 6-year-old age group were at greater risk of aborting from Brucella infection than heifers. Approximately 94% of all aborting cows were in herds which experienced three or fewer abortions. Less than 4% of herds experienced four or more abortions annually. In most cases of abortion due to brucellosis, more than one diagnostic test yielded positive results. However, 13.3% of cases were recognised by positive bacteriological results alone.     

Investigation of bovine brucellosis in the Northeastern Turkey

Bovine brucellosis, caused by Brucella abortus, is a significant problem for both public and animal health in Turkey. This study was conducted on the calving seasons between 2001 and 2006. A total of 626 serum samples of cattle obtained from 27 herds with a history of abortions was examined for Brucella antibodies by RBPT, SAT and ELISA. Of the cattle sera analysed, 221 (35,30%) and 206 (32, 92%) and 247 (39,45%) were found to be positive by RBPT , SAT and ELISA, respectively. B. abortus was isolated from 48 (32,21%) of 149 lung samples and stomach contents of the aborted fetuses. Based on the biochemical tests and the agglutination tests with monospecific A and M antisera, only 3 of the isolates were found to be B. abortus biotype 1 and the remaining 45 were biotype 3. This study also revealed that the dominant biotype of B. abortus was biotype 3 in this region. The determination of the agents responsible for bovine brucellosis and serosurvey of this disease are expected to help better understanding of this zoonotic infection in this region and neighbouring countries.     

Efficacy of strain RB51 vaccine in protecting infection and vertical transmission against Brucella abortus in Sprague-Dawley rats

Immunizing animals in the wild against Brucella (B.) abortus is essential to control bovine brucellosis because cattle can get the disease through close contact with infected wildlife. The aim of this experiment was to evaluate the effectiveness of the B. abortus strain RB51 vaccine in protecting infection as well as vertical transmission in Sprague-Dawley (SD) rats against B. abortus biotype 1. Virgin female SD rats (n = 48) two months of age were divided into two groups: one group (n = 24) received RB51 vaccine intraperitoneally with 3 × 10¹⁰ colony forming units (CFU) and the other group (n = 24) was used as non-vaccinated control. Non-vaccinated and RB51-vaccinated rats were challenged with 1.5 × 10⁹ CFU of virulent B. abortus biotype 1 six weeks after vaccination. Three weeks after challenge, all rats were bred. Verification of RB51-vaccine induced protection in SD rats was determined by bacteriological, serological and molecular screening of maternal and fetal tissues at necropsy. The RB51 vaccine elicited 81.25% protection in SD rats against infection with B. abortus biotype 1. Offspring from rats vaccinated with RB51 had a decreased (p < 0.05) prevalence of vertical transmission of B. abortus biotype 1 compared to the offspring from non-vaccinated rats (20.23% and 87.50%, respectively). This is the first report of RB51 vaccination efficacy against the vertical transmission of B. abortus in the SD rat model.     

Evaluation of PCR methods for detection of <Emphasis Type="Italic">Brucella</Emphasis> strains from culture and tissues

The genus Brucella causes significant economic losses due to infertility, abortion, stillbirth or weak calves, and neonatal mortality in livestock. Brucellosis is still a zoonosis of public health importance worldwide. The study was aimed to optimize and evaluate PCR assays used for the diagnosis of Brucella infections. For this aim, several primers and PCR protocols were performed and compared with Brucella cultures and biological material inoculated with Brucella. In PCR assays, genus- or species-specific oligonucleotide primers derived from 16S rRNA sequences (F4/R2, Ba148/928, IS711, BruP6-P7) and OMPs (JPF/JPR, 31ter/sd) of Brucella were used. All primers except for BruP6-P7 detected the DNA from reference Brucella strains and field isolates. In spiked blood, milk, and semen samples, F4-R2 primer-oriented PCR assays detected minimal numbers of Brucella. In spiked serum and fetal stomach content, Ba148/928 primer-oriented PCR assays detected minimal numbers of Brucella. Field samples collected from sheep and cattle were examined by bacteriological methods and optimized PCR assays. Overall, sensitivity of PCR assays was found superior to conventional bacteriological isolation. Brucella DNA was detected in 35.1, 1.1, 24.8, 5.0, and 8.0% of aborted fetus, blood, milk, semen, and serum samples by PCR assays, respectively. In conclusion, PCR assay in optimized conditions was found to be valuable in sensitive and specific detection of Brucella infections of animals.     

Molecular identification of chlamydial cause of abortion in small ruminants in Jordan

Chlamydophila abortus (Ch. abortus) is the etiological agent of ovine enzootic abortion (OEA) and one of the most common infectious agents of abortion in small ruminants worldwide. RFLP-PCR analysis of the outer membrane protein gene (OMP2 gene) was used for diagnosis and characterization of chlamydial causes of abortion in small ruminants in Jordan. Sixty-six placental tissues and 15 vaginal swabs were collected from aborted ewes and does to identify cause of abortion in Jordan. Thirty-eight placental samples (58 %) and 13 vaginal swabs (87 %) were positive for chlamydial DNA. Shedding of bacteria in vaginal swabs was detected within 7 days after abortion. The results of this study showed that chlamydiosis is one of the important causes of abortion in small ruminants in Jordan. In addition, vaginal swab is an excellent sample for molecular diagnosis of chlamydiosis. DNA sequencing and RFLP analysis of the OMP2 reveal that all chlamydial cause of abortion in small ruminants in Jordan are due to Ch. abortus. While, Ch. pecorum was not detected in any sample. OMP2 gene of the isolated Jordanian strain was identical (100 %) to Ch. abortus FAS strain. In conclusion, Ch. abortus is an important cause of abortion in Jordan; vaginal swab within 7 days of abortion can be used for molecular diagnosis of chlamydiosis in small ruminants.     

Pyruvate kinase is necessary for Brucella abortus full virulence in BALB/c mouse

Brucellosis, caused by a facultative intracellular pathogen Brucella, is one of the most prevalent zoonosis worldwide. Host infection relies on several uncanonical virulence factors. A recent research hotpot is the links between carbon metabolism and bacterial virulence. In this study, we found that a carbon metabolism-related pyruvate kinase (Pyk) encoded by pyk gene (locus tag BAB_RS24320) was associated with Brucella virulence. Determination of bacterial growth curves and resistance to environmental stress factors showed that Pyk plays an important role in B. abortus growth, especially under the conditions of nutrition deprivation, and resistance to oxidative stress. Additionally, cell infection assay showed that Pyk is necessary for B. abortus survival and evading fusion with lysosomes within RAW264.7 cells. Moreover, animal experiments exhibited that the Pyk deletion significantly reduced B. abortus virulence in a mouse infection model. Our results elucidated the role of the Pyk in B. abortus virulence and provided information for further investigation of Brucella virulence associated carbon metabolism.     

Protective effects of recombinant Brucella abortus Omp28 against infection with a virulent strain of Brucella abortus 544 in mice

The outer membrane proteins (OMPs) of Brucella (B.) abortus have been extensively studied, but their immunogenicity and protective ability against B. abortus infection are still unclear. In the present study, B. abortus Omp28, a group 3 antigen, was amplified by PCR and cloned into a maltose fusion protein expression system. Recombinant Omp28 (rOmp28) was expressed in Escherichia coli and was then purified. Immunogenicity of rOmp28 was confirmed by Western blot analysis with Brucella-positive mouse serum. Furthermore, humoral- or cell-mediated immune responses measured by the production of IgG1 or IgG2a in rOmp28-immunized mice and the ability of rOmp28 immunization to protect against B. abortus infection were evaluated in a mouse model. In the immunogenicity analysis, the mean titers of IgG1 and IgG2a produced by rOmp28-immunized mice were 20-fold higher than those of PBS-treated mice throughout the entire experimental period. Furthermore, spleen proliferation and bacterial burden in the spleen of rOmp28-immunized mice were approximately 1.5-fold lower than those of PBS-treated mice when challenged with virulent B. abortus. These findings suggest that rOmp28 from B. abortus is a good candidate for manufacturing an effective subunit vaccine against B. abortus infection in animals.     

Milk yield and reproductive performance of brucellosis-vaccinated but seropositive Holstein cows

The aim of this research was to study if seropositivity for brucellosis in vaccinated cows against this disease hampers reproductive performance and milk production in high-yielding Holstein cows. For this purpose 1,026 healthy cows and 372 cows seropositive for brucellosis were enrolled in this study. Cows positive to card test and subsequently to the rivanol test were further subjected to the radial immunodiffusion (RID) test. It was found that only 11 % of the presumably infected cows by brucellosis screening tests were really infected with this disease. The reproductive performance of the group of cows with 11 % Brucella-infected animals was not impaired; overall pregnancy rate did not differ between seropositive and healthy cows (30.9 vs. 29.6 %). The abortion rates were similar between seropositive cows (5.3 %) and seronegative animals (6.9 %). Cows in the herd with 11 % Brucella-infected animals produced significantly more milk than unaffected cows over a 305-day lactation (10,684?±?1,720 vs. 10,345?±?1,736; mean ± SD; P?<?0.05). It was concluded that in dairy herds vaccinated against brucellosis with both 19 and RB51 strains, supplemental tests such as RID need to be conducted on all reactors in order to maintain diagnostic accuracy. These results also indicate that 11 % animal prevalence of brucellosis did not exert a detrimental effect on 305-day milk yield and reproductive performance in high milk-yielding Holstein cows.     

Serological study of a dairy herd with a recent history of Neospora abortion

HERD HISTORY: A cluster of 19 abortions occurred in April and May 1994 on a seasonal dairy farm of 277 cows in Taranaki. Lesions consistent with neosporosis were seen in several foetuses. The age of the aborting cows ranged from 2 to 12 years. Pregnant rising 2-year-old heifers had been grazed off the main farm on a run-off' until returning in May, when they were replaced on the run-off by the rising l-year-old calves. None of these pregnant heifers aborted. METHODS: Sera were obtained from all calves (n = 32), 32 of 33 pregnant heifers, 59 of 60 3-4 year-old cows and 101 of 150 older cows, plus all cows that had aborted. Sera were tested using an indirect fluorescent antibody assay. Non-aborting cattle sera were screened at 1:400, and further dilutions were tested from 15 aborted cows. RESULTS: The percentages of seropositive cattle were: 32% of the calves, 3% of the heifers, 31% of the younger cows and 27% of the older cows. The proportion of seropositive heifers was significantly lower (p< 0.01) than in the other age classes. There was no significant difference in the proportion of seropositive animals between these other age classes (p >0.05). Of the 15 cows that aborted, and for which further dilutions were tested, nine had titres of 1: 12,800, four had titres of 1:6400, one had a titre of 1: 1600 and one had a titre of 1:400. Twenty cows were the dams of pregnant heifers. Six of these cows were seropositive but all their offspring were seronegative. Fifteen aborting cows were held over to the following year when 13 became pregnant and calved normally. There were only two to three abortions in the whole herd in this following year. CONCLUSION: These results suggest that either a point-infection occurred sometime in early 1994 with an infective period short enough so that the heifers did not get infected when they returned to the main farm, or some factor precipitated a recrudescence of latent infection with a subsequent rise in titres in about 29% of the animals on. the main farm only. Although neither hypothesis can be proven or disproven in this observational study, the former hypothesis seems more likely.     

A20 promotes Brucella intracellular growth via inhibition of macrophage cell death and activation

The zinc-finger protein A20 has crucial physiological functions as a dual inhibitor of macrophage activation and apoptosis in tumor necrosis factor receptor1 (TNFR1) signaling pathway. Brucella infection can induce A20 expression in macrophages. Here, we hypothesize that A20 promotes Brucella intracellular growth via inhibition of activation and apoptosis of macrophages. To test this hypothesis, we stably incorporated mouse A20-shRNA into the RAW264.7 cells by lentiviral gene transfer to successfully knockdown A20. A20-deficient RAW264.7 cells were subsequently challenged with Brucella abortus and colony formation units (CFUs) of bacteria, TNFα production, NF-kB activation, macrophages apoptosis and cell death were evaluated. The A20 knockdown was shown to effectively promote B. abortus-stimulated TNFα release, NF-kB activation and macrophage cell death, which suppressed B. abortus intracellular replication. Unexpectedly, deficiency of A20 failed to lead to B. abortus-induced macrophage apoptosis. A20 deficiency coupled NF-kB inhibition promoted caspase-8 dependent B. abortus-induced macrophage apoptosis. These findings provide a novel mechanism by which Brucella intracellular growth within macrophages occurs through up-regulation of A20 thereby limiting activation and macrophages cell death.     

Use of Brucella abortus vaccine strain RB51 in pregnant cows after calfhood vaccination with strain 19 in Argentina

One hundred and seven pregnant cows, which had been calfhood vaccinated with Brucella abortus strain 19 (S-19) were revaccinated with either S-19 or strain RB51 (S-RB51). All S-19-revaccinated animals seroconverted, while none of the RB51-revaccinated animals seroconverted. Two out of 25 (8%) S-19-revaccinated animals aborted, while none of the 57 RB51-revaccinated group aborted. Four of the S-19-revaccinated animals shed S-19 in the milk for at least 7 days, while only 1 cow shed S-RB51 for at least 3 days (but <7 days) post-parturition. Revaccination of strain 19 calfhood-vaccinated, pregnant cattle with S-RB51 appears to be a safe procedure with no diagnostically negative consequences.     

Immunogenicity of adenovirus and DNA vaccines co-expressing P39 and lumazine synthase proteins of <Emphasis Type="Italic">Brucella abortus</Emphasis> in BALB/c mice

Brucella poses a great threat to animal and human health. Vaccination is the most promising strategy in the effort to control Brucella abortus (B. abortus) infection, but the currently used live vaccines interfere with diagnostic tests and could potentially result in disease outbreak. Therefore, new subunit vaccines and combined immunization strategies are currently under investigation. In this study, immunogenicity and protection ability of a recombinant adenovirus and plasmid DNA vaccine co-expressing P39 and lumazine synthase proteins of B. abortus were evaluated based on the construction of the two molecular vaccines. Four immunization strategies (single adenovirus, single DNA, adenovirus/DNA, DNA/adenovirus) were investigated. The results showed that the immunization strategy of DNA priming followed by adenovirus boosting induced robust humoral and cellular immune responses, and it significantly reduced the numbers of B. abortus in a mouse model. These results suggest that it could be a potential antigen candidate for development of a new subunit vaccine against B. abortus infection.