Brucellosis,caused by Brucella,is one of the most common zoonosis.However,there is still no vaccine for human use.Although some live attenuated vaccines have been approved for animals,the protection effect is not idea...Brucellosis,caused by Brucella,is one of the most common zoonosis.However,there is still no vaccine for human use.Although some live attenuated vaccines have been approved for animals,the protection effect is not ideal.In this study,we developed a dual-antigen nanoconjugate vaccine containing both polysaccharide and protein antigens against Brucella.First,the antigenic polysaccharide was covalently coupled to the outer membrane protein Omp19 using protein glycan coupling technology,and then it was successfully loaded on a nano-carrier through the SpyTag/SpyCatcher system.After confirming the efficient immune activation and safety performance of the dual-antigen nanoconjugate vaccine,the potent serum antibody response against the two antigens and remarkable protective effect in non-lethal and lethal Brucella infection models were further demonstrated through different routes of administration.These results indicated that the dual-antigen nanoconjugate vaccine enhanced both T helper 1 cell(Th1)and Th2 immune responses and protected mice from Brucella infection.Furthermore,we found that this protective effect was maintained for at least 18 weeks.To our knowledge,this is the first Brucella vaccine bearing diverse antigens,including a protein and polysaccharide,on a single nanoparticle.Thus,we also present an attractive technology for co-delivery of different types of antigens using a strategy applicable to other vaccines against infectious diseases.展开更多
Objective:To present platelet large cell ratio(P-LCR),reticulocyte,and immature reticulocyte fraction(IRF)values as novel parameters in diagnosis and response to treatment in patients developing sacroiliitis.Methods:S...Objective:To present platelet large cell ratio(P-LCR),reticulocyte,and immature reticulocyte fraction(IRF)values as novel parameters in diagnosis and response to treatment in patients developing sacroiliitis.Methods:Sixty-eight patients with clinical symptoms and Brucella standard tube agglutination(Wright)or Brucella Coombs agglutination test titers≥1:160 were included in the study.Two groups were established,one developing sacroiliitis and another with no sacroiliitis development.P-LCR,reticulocyte,and IRF levels were measured using a Sysmex XN-9000 device(Japan).These were then compared between the two groups.Results:Reticulocyte(P=0.037)and IRF(P=0.026)levels were significantly lower among the patients developing sacroiliitis compared to the non-sacroiliitis group,while P-LCR(P=0.003)levels were significantly higher.P-LCR had the most powerful correlation with sacroiliitis development.Significant negative correlation was observed between reticulocyte,IRF levels and sacroiliitis.Conclusions:Elevated P-LCR levels were observed as a marker of persisting inflammation in patients developing sacroiliitis,while low reticulocyte and IRF levels secondary to bone marrow involvement were detected.These three parameters emerged as highly significant markers in terms of diagnosis and reflecting responses to treatment in organ involvement such as sacroiliitis in brucellosis.These are presented as inexpensive,and easily accessible novel parameters.展开更多
Brucellosis remains one of the most common zoonoses spread worldwide,inducing enormous economic losses to the livestock industry and posing serious health threats to humans.Brucellosis re-emerged in China in the mid-1...Brucellosis remains one of the most common zoonoses spread worldwide,inducing enormous economic losses to the livestock industry and posing serious health threats to humans.Brucellosis re-emerged in China in the mid-1990s and reached a historically high level in 2015.The National Brucellosis Prevention and Control Plan(NBPCP)was initiated from 2016 to 2020.However,the present epidemiological status in livestock has not been elucidated,and whether Brucella variation occurred remains unclear.This study performed an extensive serological investigation in ruminant livestock from 2019 to 2021 in central Gansu Province,China.In total,11,296 samples from 337 farms were collected to detect the specific antibodies of Brucella.The yearly average serological prevalence of Brucella at the flock level and individual level declined from 11.32%to 8.26%and 1.17%to 0.57%,respectively.The apparent individuallevel seroprevalence of small and large ruminants was 0.89%and 0.52%,respectively.The brucellosis distribution has shifted from pastoral areas to agro-pastoral areas.Flock size and gender may be major risks of Brucella infection.Then,the B.melitensis TZ strain was isolated from female Tibetan sheep blood cell lysates.Phonotypical characterization demonstrated that it belongs to B.melitensis.biovar 3,and multilocus sequencing typing results indicated that it belongs to ST8.The whole genome and subsequent phylogenetic analysis demonstrated that the B.melitensis TZ strain is genetically more closely related to the B.melitensis QH61 strain.The B.melitensis TZ strain has similar growth characteristics to the B.melitensis 16 M strain.Overall,our study suggests that after strengthening control and prevention measures based on the NBPCP,there is a very low prevalence or absence of B.melitensis in the central Gansu Province of China,and the genotype of an epidemic strain of Brucella in Northwest China is relatively stable.展开更多
[ Objective] The paper aimed to understand the epidemiological characteristics of Brucella strains in Xinjiang and then provide an available integrated measure to prevent and control brucellosis. [ Method ] Eleven sus...[ Objective] The paper aimed to understand the epidemiological characteristics of Brucella strains in Xinjiang and then provide an available integrated measure to prevent and control brucellosis. [ Method ] Eleven suspected Brucella strains were isolated by traditional methods, which were further identified by AMOS-PCR assay. Conventional biochemical tests were carried out to identify the biological subtype of sheep Brucella. [ Result] Nine strains were all B. meliten- s/s, and biological test indicated that all of them were B. melitensis biotype 3. [ Conclusion] B. melitensis biotype 3 was the predominant strain of Brucella in Xin- jiang, and AMOS-PCR assay could be applied safely and quickly as an assistant tool to detect Brucella. The results of molecular epidemiology laid a foundation for updating prevention and control strategy against brucellosis in Xinjiang.展开更多
Objective:To design a duplex PCR for rapid and simultaneous detection of Brucella species, in human blood samples.Methods:Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis.Fo...Objective:To design a duplex PCR for rapid and simultaneous detection of Brucella species, in human blood samples.Methods:Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis.Following DNA extraction,PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals.Results:Of the 52 peripheral bloods samples tested, 25 sample(48%) showed positive reactions in PCR.Twelve samples were positive for Brucella abortus(B.abortus)(23%,13 for Brucella melUensis(B.melUensis)(25%) and 0 for Brucella ovis (6.ovis)(Ow.Conclusions:This work de=monstrates dial in case where specific primers were utilized,duplex PCR has proved to be a simple,fast,and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples.展开更多
Objective:To evaluate simultaneous detection and differentiates of Brucella abortus(B.abortus) and Brucella melitensis(B.melitensis) through the combinatorial POR method.Methods:This study was designed using three pri...Objective:To evaluate simultaneous detection and differentiates of Brucella abortus(B.abortus) and Brucella melitensis(B.melitensis) through the combinatorial POR method.Methods:This study was designed using three primers that could simultaneously identify and differentiate two major species of pathogenic Brucella in humans and animals.Identification and differentiation of each species using the size of the PCR product were determined.To determine the specifieity of the method,bacteria close to the genus Brucella were used.Finally,to confirm PCR products.In addition to the products sequence,RFLP was performed on PCR products using restriction enzymes.Results:The method of optimized combinatorial PCR in this study could simultaneously detect and differentiate B.abortus and B.melitensis with high specificity and sensitivity in clinical samples.Differentiation of species is based on the resulting bands: therefore,the band 494 bp for B.abortus and 733 bp for B.melitensis were obtained.RFLP and sequencing results confirmed PCR results.Conclusions:The results of this study shows that without routine diagnostic methods such as culture and serology tests,using the molecular method of combinatorial PCR,important species of Brucella can be simultaneously identified and differentiated in clinical samples.展开更多
Brucellosis is a bacterial anthropozoonosis usually caused by Brucella abortus, Brucella melitensis, Brucella suis and Brucella canis. Brucella suis, the causative agent of swine brucellosis, is classified into five b...Brucellosis is a bacterial anthropozoonosis usually caused by Brucella abortus, Brucella melitensis, Brucella suis and Brucella canis. Brucella suis, the causative agent of swine brucellosis, is classified into five biovars and preferentially infects different animal hostsIll. In China, brucellosis is a national notifiable communicable disease both in animals and in human. In 2009, 35 816 brucellosis cases were reported. The annual incidence was 2.7 per 100 000 population.展开更多
Objective:To clone the virB12 gene in pET28a expression vector for production of recombinant protein to be used as antigenic component for future serological test development.Methods: Brucella melitensis(B.melitensis)...Objective:To clone the virB12 gene in pET28a expression vector for production of recombinant protein to be used as antigenic component for future serological test development.Methods: Brucella melitensis(B.melitensis) 16M strain was cultured and bacterial DNA was extracted by Bioneer AccuPrep~ Genomic DNA Extraction Kit.Oligonucleotide primer pair was designed based on Brucella virB12 gene sequence with BamHI and HindIII restriction site at 5’ end of the forward and reverse primers,respectively.DNA amplification was performed using PrimSTAR~ HS DNA polymerase and the PCK product was purified by DNA AccuPrepGel Purification Kit.Purified DNA was cloned into pJET1.2 cloning vector.VirB12 gene fragment was excised from pJET1.2 asing BamHI/HindIII and subsequendy subcloned into pET28a(+).Results:Brucella virB12 gene was successfully cloned in pJET1.2 and then in pET28a(+) plasmids.PCR and restriction enzyme digestion confirms the procedure.Conclusion:We cloned and expressed the Brucella virB12 gene which could be used as antigenic component for specific serological assay development.展开更多
Objective:To explore the antibiotic resistance of Brucella melitensis and instruct rational use of antimicrobial agents in clinical treatment of Brucella infection.Methods:Bacteria were cultured and identified by BACT...Objective:To explore the antibiotic resistance of Brucella melitensis and instruct rational use of antimicrobial agents in clinical treatment of Brucella infection.Methods:Bacteria were cultured and identified by BACTEC9120 and VTTEK Ⅱ automicrobic system.E-test was used to detect the minimal inhibitory concentration(MIC) of antimicrobial agents in the drug susceptivity experiment.Results:A total of 19 brucella strains(all Brucella melitensis) were isolated from 19 patients,who had fever between January 2010 and June 2012,and 17 samples were blood,one was bone marrow,the other sample was cerebrospinal fluid.The MIC range of ceftazidime was 2.0-8.0 mg/L,rifampicin was 0.06-2.0 mg/L,amikacin was 4.0-12.0 mg/L,levofloxacin was 2.0-8.0 mg/ L,doxycycline was 8.0-32.0 mg/L,sulfamethoxazole-trimethoprim was 4.0-16.0 mg/L,ampicillin was 1.5-2.0 mg/L and gentamicin was 0.50-0.75 mg/L.Conclusions:The drugs used in this experiment cover common drugs for treating Brcella.Meanwhile,the results are consistent with clinical efficacy.It is suggested personalized regimen according to patients' status in treatment of Brucella.展开更多
Rational:Brucellosis is a globally prevalent zoonotic disease.Any part of the body can be affected by active brucellosis but osteoarticular involvement are the most common symptoms which was reported to vary from 10%t...Rational:Brucellosis is a globally prevalent zoonotic disease.Any part of the body can be affected by active brucellosis but osteoarticular involvement are the most common symptoms which was reported to vary from 10%to 85%.The spine is the most common site of brucellosis in the bones.However,noncontiguous brucellar spondylitis is rare,only few cases have been reported in the literature.Patient concerns:A 62-year-old woman with brucellar spondylitis presented with lower back pain and pain in the right lower extremity for six months.Diagnosis:Brucella agglutination test(1:320)and the result of polymerase chain reaction(PCR)confirmed the diagnosis of noncontiguous brucellar spondylitis.Intervention:During hospital stay,the women received intravenous treatment for brucellosis(A combination of doxycycline 200 mg/d,rifampicin 900 mg/d,levofloxacin 0.5 g/QD,and ceftriaxone 2 g/QD was administered for 1 week),The L4-S1 vertebral body was fixed by posterior lumbar debridement.Outcome:Six months after discharge,the follow-up radiographic images showed stable vertebral height and good lumbar stability.She complained no discomfort.Lessons:Multi-level involvement is an exceptional form of brucellar spondylitis.To the best of our knowledge,only few similar cases have been reported.PCR and bacterial culture is necessary for confirmed diagnosis.展开更多
Brucella spp. are pathogenic to humans and domestic animal. Nowadays,there is no effective vaccine and control strategy in China. So,it is necessary to research effective vaccines for prevention and treatment of this ...Brucella spp. are pathogenic to humans and domestic animal. Nowadays,there is no effective vaccine and control strategy in China. So,it is necessary to research effective vaccines for prevention and treatment of this disease. In order to deal with these,we isolated and identified the type of Brucella in Darhan Muminggan Joint Banner and Siziwang Banner of Inner Mongolia. Totally 26 samples of sheep blood which were positive in serological test,one sample of spleen from aborted and one sample of secretion from birth canal were isolated,and the 16 S r DNA genes of positive samples were sequenced. Phylogenetic analysis proved that there were four isolates similar to B. melitensis. As an important diagnostic antigens of B. melitensis,the BP26 gene was amplified. The BP26 gene was cloned into vector p ET24a( +) and conducted sequence analysis. The BP26 gene was 900 bp,with an open reading frame of 753 bp. The homology of BP26 gene with the vaccine strain M5 was 100%,and that with S2 and A19 vaccine strains was 99. 99%. These finding supported the development of BP26-based specific serodiagnostic test and vaccine for B. melitensis in China.展开更多
Background: Brucellosis in male goats is characterized by arthritis, orchitis and epididymitis, which may induce infertility. Nevertheless, these lesions were categorized as chronic while acute lesions had not been de...Background: Brucellosis in male goats is characterized by arthritis, orchitis and epididymitis, which may induce infertility. Nevertheless, these lesions were categorized as chronic while acute lesions had not been described. This study investigates the histopathological and immuno histochemistry reactions in organs of bucks acutely infected by Brucella melitensis. Results: Only testis and prepuce of acutely infected bucks showed significantly severe histological lesions. Other internal organs had mild to moderate lesions. However, positive immunohistochemistry stainings were observed in organs except the bulbourethral gland. There was a significant positive correlation between the distribution of B. melitensis and IHC intensity but no significant correlation between the IHC intensity and histopathology lesions. Conclusion: The results indicate that acute brucellosis did not lead to clinical presentation, although B. melitensis was well distributed in various organs of bucks.展开更多
Importance of urease activity on pathogenic differences among Brucella species was evaluated. In cell-free extracts, the B. suis urease showed 12 times greater specific activity than the B. melitensis urease. When Fis...Importance of urease activity on pathogenic differences among Brucella species was evaluated. In cell-free extracts, the B. suis urease showed 12 times greater specific activity than the B. melitensis urease. When Fisher-344 rats were inoculated intraperitoneally (IP), at 1 week post-inoculation (PI), B. melitensis wild type 16 M was recovered from spleens and livers in greater numbers than B. suis wild type 1330. At 8 weeks PI, spleens were clear of B. melitensis, whereas B. suis remained. The wild type and the urease deficient strains of B. suis did not differ from each other in terms of recovery from spleen or liver. Our observations suggest that B. melitensis induces greater acute infectivity in Fisher-344 rats, whereas B. suis causes chronic infectivity;and urease activity has no influence on Brucella infection using an IP route.展开更多
Objective:To explore immunochemical characterization of antigens of Brucella canis(B. canis),and the use in seroprevalence study of canine brucellosis.Methods:External hot phosphate buffer saline extract(HPBSE) and in...Objective:To explore immunochemical characterization of antigens of Brucella canis(B. canis),and the use in seroprevalence study of canine brucellosis.Methods:External hot phosphate buffer saline extract(HPBSE) and internal sonicated(SA) antigens were prepared from B.canis strain MEX 51 and imniunochemically characterized.These antigens were used to test 527 serum samples of dogs by 2-mercaptoethanol-tubc agglutination test(2 ME-TAT), agar gel immunodiffusion test(AGID).dot-ELISA and indirect enzyme-linked immunosorbent assay(I-ELISA) to assess the seroprevalence of canine brucellosis.Results:The protein content of HPBSE and SA antigens was 0.387 mg/ml.and 0.195 mg/mL,respectively,whereas carbohydrate content was 0.174 mg/mL and 0.150 mg/mL,respectively.The sodium dodecyl sulfate-polyacrylamide gel electrophoresis(12.5%) of HPBSE and SA,revealed 6 and 8 visible peptide bands ranging from 18-80 kDa and 12-45 kDa,respectively.Western blot analysis showed immunodominant bands of MW 12.28.39 and 45 kl)a for HPBSE and 20-24 kl)a for SA. The AGII) revealed HPBSE as more specific antigen than SA but both I-ELISA and dot-ELISA indicated SA antigen to be more specific and reliable than HPBSE.The seroprevalence of canine brucellosis was 2.27%by 2ME-TAT.1.5%by AGID.3.03%by dot-ELISA and 16.12%by I-ELISA. Conclusions:On the basis of the results of present study,we concluded that HPBSE is suitable antigen for AGID,which is more specific:whereas SA antigen is suitable for I-ELISA,which is highly sensitive.Therefore,initial screening of serum samples should be carried out by I-ELISA followed by confirmation with AGID.展开更多
Objective: To study Brucella(B.) abortus strains isolated in the Russian Federation, in order to identify their detailed position in the phylogenetic structure of the species global population as well as to determine ...Objective: To study Brucella(B.) abortus strains isolated in the Russian Federation, in order to identify their detailed position in the phylogenetic structure of the species global population as well as to determine genetic relationships for isolates from different geographical areas.Methods: Based on Bayesian method, the whole genome singlenucleotide polymorphism(SNP) analysis of 258 B. abortus strains from different geographical areas of the world including 20 B. abortus strains isolated in Russia was carried out. Results: The core genome SNP analysis of the B. abortus isolates allowed describing the main genetic lineages. The Russian strains entered two separate clades, including the basal branch and the C1 branch that is widely spread in Eurasia. The data on the isolation time was used for the dating of phylogenetic tree, and also the estimated time frame for the B. abortus genotype diversification was determined. There were sets of specific SNPs identified, which defined each of the genotypes and sub-genotypes.Conclusions: A significant genetic diversity of the brucellosis pathogen strains from Russia has been proven. The sets of unique specific SNPs described in our study may become one of the elements within a bio-informational analysis algorithm to be used for epidemiological study of brucellosis outbreaks, including those caused by new(atypical) genetic variants of B. abortus.展开更多
Brucellosis is a worldwide zoonosis caused by Brucella, which poses great threat to human health. Totally 234 milk samples from a scale dairy farm were amplified by nested PCR, and the pathogens were further separated...Brucellosis is a worldwide zoonosis caused by Brucella, which poses great threat to human health. Totally 234 milk samples from a scale dairy farm were amplified by nested PCR, and the pathogens were further separated. The results showed that a DNA band of 419 bp was amplified from 18 out ot＂234 milk samples. Among 18 positive milk samples, nine samples amplified the DNA band of 535 bp, which were B. su/s ; one sample amplified the DNA bands of 535 bp and 285 bp, which was the mixture of B. su/s and B. bov/s. The results laid a foundation for prevention and control of brucellosis.展开更多
[ Objective] To investigate the occurrence and prevalence of Brucella infection in Hulunbiur region. [ Method] A total of 906 serum samples were collected from sheep in Hulunbiur region and detected by the Rose Bengal...[ Objective] To investigate the occurrence and prevalence of Brucella infection in Hulunbiur region. [ Method] A total of 906 serum samples were collected from sheep in Hulunbiur region and detected by the Rose Bengal plate agglutination test to determine the positive rate of Brucella antibodies. [Result] The positive rates of Brucella antibodies in lambs, ewes and mutton sheep were 5.28%, 5.68% and 4.22%, respectively. The total positive rate of Brucella antibodies was 4.86% in sheep. [ Condusionl The positive rate of Brucella antibodies was high in sheep in Hulunblur region.展开更多
[ Objective] In order to clone and express the formyltransferase of Brucella abortus in E. coli, purify the expressed protein and detect its immunogenicity. EMethods] A gene recoding formyltransferase 27 -35 ku （Wbkc...[ Objective] In order to clone and express the formyltransferase of Brucella abortus in E. coli, purify the expressed protein and detect its immunogenicity. EMethods] A gene recoding formyltransferase 27 -35 ku （Wbkc） was amplified from the genomic DNA of Brucella abortus PCR. The amplified fragments were digested with BamH I and Sal I, and then cloned to the vector pET28a. The constructed recombinant plasmid pET28a-Wbkc was transformed to E. coil BL21 and was induced to express the fusion protein. Then the protein was purified by histidine-binding res- in column chromatography, and the immunogenicity was detected by Western blot. The PCR product of Wbkc gene was cloned to the vector pET28a, and the recombinant vector was confirmed by colony PCR identification, recombinant vector digested identification and sequencing analy- sis. [ Results] It was successfully expressed in E. coil BL21 as a fusion protein with histidine at the presence of IPTG, and a specific protein band of 29 ku was found when identified by SDS-PAGE. Western blot showed good immunoreactivity of the expressed product. Wbkc was successfully cloned and expressed, and the purified fusion protein had immunogenicity. This study provides a solid foundation for the further study on the function of proteins and brucellosis diagnostic antigens. [ Conclusion] Amplification of formyltransferase gene of Brucella abortus and prokaryotic expression of WbkC_ indicatino the formvItransferase has specific immune reactivitv.展开更多
[ Objective ] This study aimed to clone and express formyltransferase （Wbkc） gene from Brucella abortus in E. coli, purify the expressed protein and analyze its immunogenicity. [Method] A gene encoding 27 -35 ku for...[ Objective ] This study aimed to clone and express formyltransferase （Wbkc） gene from Brucella abortus in E. coli, purify the expressed protein and analyze its immunogenicity. [Method] A gene encoding 27 -35 ku formyltransferase （Wbkc） was amplified from the genomic DNA of BruceUa abortus by PCR. The amplified fragments were digested with BamH I and Sal I, and inserted into pET28a vector. The constructed recombinant plasmid pET 28a-Wbkc was trans- formed into E. coli BL21 and was induced to express the fusion protein. Subsequently, the protein was purified by histidine-binding resin column chromatography, and the immunogenicity was detected by Western blot assay. The recombinant plasmid was identified by colony PCR, double digestion and sequencing analysis. [ Result] Wbkc was successfully cloned and expressed in E. coli. A specific protein band of 29 ku was detected by SDS-PAGE. Western blot showed specific im- munoreactivity of the purified fusion protein. [ Conclusion] This study provided a solid foundation for further investigating diagnostic proteins for brucellosis and developing Brucella gene-deletion vaccines.展开更多
基金supported by the National Key Research and Development Program of China(2021YFC2102100)the National Natural Science Foundation of China(U20A20361,32271507,81930122,and 82171819)the Beijing Postdoctoral Research Foundation(2021-ZZ-035)。
文摘Brucellosis,caused by Brucella,is one of the most common zoonosis.However,there is still no vaccine for human use.Although some live attenuated vaccines have been approved for animals,the protection effect is not ideal.In this study,we developed a dual-antigen nanoconjugate vaccine containing both polysaccharide and protein antigens against Brucella.First,the antigenic polysaccharide was covalently coupled to the outer membrane protein Omp19 using protein glycan coupling technology,and then it was successfully loaded on a nano-carrier through the SpyTag/SpyCatcher system.After confirming the efficient immune activation and safety performance of the dual-antigen nanoconjugate vaccine,the potent serum antibody response against the two antigens and remarkable protective effect in non-lethal and lethal Brucella infection models were further demonstrated through different routes of administration.These results indicated that the dual-antigen nanoconjugate vaccine enhanced both T helper 1 cell(Th1)and Th2 immune responses and protected mice from Brucella infection.Furthermore,we found that this protective effect was maintained for at least 18 weeks.To our knowledge,this is the first Brucella vaccine bearing diverse antigens,including a protein and polysaccharide,on a single nanoparticle.Thus,we also present an attractive technology for co-delivery of different types of antigens using a strategy applicable to other vaccines against infectious diseases.
文摘Objective:To present platelet large cell ratio(P-LCR),reticulocyte,and immature reticulocyte fraction(IRF)values as novel parameters in diagnosis and response to treatment in patients developing sacroiliitis.Methods:Sixty-eight patients with clinical symptoms and Brucella standard tube agglutination(Wright)or Brucella Coombs agglutination test titers≥1:160 were included in the study.Two groups were established,one developing sacroiliitis and another with no sacroiliitis development.P-LCR,reticulocyte,and IRF levels were measured using a Sysmex XN-9000 device(Japan).These were then compared between the two groups.Results:Reticulocyte(P=0.037)and IRF(P=0.026)levels were significantly lower among the patients developing sacroiliitis compared to the non-sacroiliitis group,while P-LCR(P=0.003)levels were significantly higher.P-LCR had the most powerful correlation with sacroiliitis development.Significant negative correlation was observed between reticulocyte,IRF levels and sacroiliitis.Conclusions:Elevated P-LCR levels were observed as a marker of persisting inflammation in patients developing sacroiliitis,while low reticulocyte and IRF levels secondary to bone marrow involvement were detected.These three parameters emerged as highly significant markers in terms of diagnosis and reflecting responses to treatment in organ involvement such as sacroiliitis in brucellosis.These are presented as inexpensive,and easily accessible novel parameters.
基金the National Key Research and Development Program(2021YFC2600204 and 2021YFD1800403)the Science and Technology Program of Gansu,China(20YF8NH153).
文摘Brucellosis remains one of the most common zoonoses spread worldwide,inducing enormous economic losses to the livestock industry and posing serious health threats to humans.Brucellosis re-emerged in China in the mid-1990s and reached a historically high level in 2015.The National Brucellosis Prevention and Control Plan(NBPCP)was initiated from 2016 to 2020.However,the present epidemiological status in livestock has not been elucidated,and whether Brucella variation occurred remains unclear.This study performed an extensive serological investigation in ruminant livestock from 2019 to 2021 in central Gansu Province,China.In total,11,296 samples from 337 farms were collected to detect the specific antibodies of Brucella.The yearly average serological prevalence of Brucella at the flock level and individual level declined from 11.32%to 8.26%and 1.17%to 0.57%,respectively.The apparent individuallevel seroprevalence of small and large ruminants was 0.89%and 0.52%,respectively.The brucellosis distribution has shifted from pastoral areas to agro-pastoral areas.Flock size and gender may be major risks of Brucella infection.Then,the B.melitensis TZ strain was isolated from female Tibetan sheep blood cell lysates.Phonotypical characterization demonstrated that it belongs to B.melitensis.biovar 3,and multilocus sequencing typing results indicated that it belongs to ST8.The whole genome and subsequent phylogenetic analysis demonstrated that the B.melitensis TZ strain is genetically more closely related to the B.melitensis QH61 strain.The B.melitensis TZ strain has similar growth characteristics to the B.melitensis 16 M strain.Overall,our study suggests that after strengthening control and prevention measures based on the NBPCP,there is a very low prevalence or absence of B.melitensis in the central Gansu Province of China,and the genotype of an epidemic strain of Brucella in Northwest China is relatively stable.
基金Supported by Special Fund for Agro-scientific Research in the Public Interest of the Ministry of Agriculture"Animal Disease Prevention and Control Technology System in Border Areas"(201103008)National Key Technology R&D Program"Integration and Demonstration of Production-Life-Ecosystem Safeguard Technique in Xinjiang Desert Arid Oasis Steppe Region"(2012BAD13B03)
文摘[ Objective] The paper aimed to understand the epidemiological characteristics of Brucella strains in Xinjiang and then provide an available integrated measure to prevent and control brucellosis. [ Method ] Eleven suspected Brucella strains were isolated by traditional methods, which were further identified by AMOS-PCR assay. Conventional biochemical tests were carried out to identify the biological subtype of sheep Brucella. [ Result] Nine strains were all B. meliten- s/s, and biological test indicated that all of them were B. melitensis biotype 3. [ Conclusion] B. melitensis biotype 3 was the predominant strain of Brucella in Xin- jiang, and AMOS-PCR assay could be applied safely and quickly as an assistant tool to detect Brucella. The results of molecular epidemiology laid a foundation for updating prevention and control strategy against brucellosis in Xinjiang.
基金supported by Molecular Biology Research Center, Baqiyatallah University of Medical Sciences,with grant number BMSU/MBRC-90-001
文摘Objective:To design a duplex PCR for rapid and simultaneous detection of Brucella species, in human blood samples.Methods:Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis.Following DNA extraction,PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals.Results:Of the 52 peripheral bloods samples tested, 25 sample(48%) showed positive reactions in PCR.Twelve samples were positive for Brucella abortus(B.abortus)(23%,13 for Brucella melUensis(B.melUensis)(25%) and 0 for Brucella ovis (6.ovis)(Ow.Conclusions:This work de=monstrates dial in case where specific primers were utilized,duplex PCR has proved to be a simple,fast,and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples.
基金supported by Molecular Biology Research Center,Baqiyatallah University of Medical Sciences,with grant number BMSU/MBRC- 89-009
文摘Objective:To evaluate simultaneous detection and differentiates of Brucella abortus(B.abortus) and Brucella melitensis(B.melitensis) through the combinatorial POR method.Methods:This study was designed using three primers that could simultaneously identify and differentiate two major species of pathogenic Brucella in humans and animals.Identification and differentiation of each species using the size of the PCR product were determined.To determine the specifieity of the method,bacteria close to the genus Brucella were used.Finally,to confirm PCR products.In addition to the products sequence,RFLP was performed on PCR products using restriction enzymes.Results:The method of optimized combinatorial PCR in this study could simultaneously detect and differentiate B.abortus and B.melitensis with high specificity and sensitivity in clinical samples.Differentiation of species is based on the resulting bands: therefore,the band 494 bp for B.abortus and 733 bp for B.melitensis were obtained.RFLP and sequencing results confirmed PCR results.Conclusions:The results of this study shows that without routine diagnostic methods such as culture and serology tests,using the molecular method of combinatorial PCR,important species of Brucella can be simultaneously identified and differentiated in clinical samples.
文摘Brucellosis is a bacterial anthropozoonosis usually caused by Brucella abortus, Brucella melitensis, Brucella suis and Brucella canis. Brucella suis, the causative agent of swine brucellosis, is classified into five biovars and preferentially infects different animal hostsIll. In China, brucellosis is a national notifiable communicable disease both in animals and in human. In 2009, 35 816 brucellosis cases were reported. The annual incidence was 2.7 per 100 000 population.
基金supported by grant from Tehran University of Medical Sciences
文摘Objective:To clone the virB12 gene in pET28a expression vector for production of recombinant protein to be used as antigenic component for future serological test development.Methods: Brucella melitensis(B.melitensis) 16M strain was cultured and bacterial DNA was extracted by Bioneer AccuPrep~ Genomic DNA Extraction Kit.Oligonucleotide primer pair was designed based on Brucella virB12 gene sequence with BamHI and HindIII restriction site at 5’ end of the forward and reverse primers,respectively.DNA amplification was performed using PrimSTAR~ HS DNA polymerase and the PCK product was purified by DNA AccuPrepGel Purification Kit.Purified DNA was cloned into pJET1.2 cloning vector.VirB12 gene fragment was excised from pJET1.2 asing BamHI/HindIII and subsequendy subcloned into pET28a(+).Results:Brucella virB12 gene was successfully cloned in pJET1.2 and then in pET28a(+) plasmids.PCR and restriction enzyme digestion confirms the procedure.Conclusion:We cloned and expressed the Brucella virB12 gene which could be used as antigenic component for specific serological assay development.
基金This project was supported by the scientific Research Foundation of Shaanxi Province.China(Grant No.2009JQ4011)
文摘Objective:To explore the antibiotic resistance of Brucella melitensis and instruct rational use of antimicrobial agents in clinical treatment of Brucella infection.Methods:Bacteria were cultured and identified by BACTEC9120 and VTTEK Ⅱ automicrobic system.E-test was used to detect the minimal inhibitory concentration(MIC) of antimicrobial agents in the drug susceptivity experiment.Results:A total of 19 brucella strains(all Brucella melitensis) were isolated from 19 patients,who had fever between January 2010 and June 2012,and 17 samples were blood,one was bone marrow,the other sample was cerebrospinal fluid.The MIC range of ceftazidime was 2.0-8.0 mg/L,rifampicin was 0.06-2.0 mg/L,amikacin was 4.0-12.0 mg/L,levofloxacin was 2.0-8.0 mg/ L,doxycycline was 8.0-32.0 mg/L,sulfamethoxazole-trimethoprim was 4.0-16.0 mg/L,ampicillin was 1.5-2.0 mg/L and gentamicin was 0.50-0.75 mg/L.Conclusions:The drugs used in this experiment cover common drugs for treating Brcella.Meanwhile,the results are consistent with clinical efficacy.It is suggested personalized regimen according to patients' status in treatment of Brucella.
基金funded by science foundation of Beijing Ditan Hospital Capital Medical University(No.DTQL201803)
文摘Rational:Brucellosis is a globally prevalent zoonotic disease.Any part of the body can be affected by active brucellosis but osteoarticular involvement are the most common symptoms which was reported to vary from 10%to 85%.The spine is the most common site of brucellosis in the bones.However,noncontiguous brucellar spondylitis is rare,only few cases have been reported in the literature.Patient concerns:A 62-year-old woman with brucellar spondylitis presented with lower back pain and pain in the right lower extremity for six months.Diagnosis:Brucella agglutination test(1:320)and the result of polymerase chain reaction(PCR)confirmed the diagnosis of noncontiguous brucellar spondylitis.Intervention:During hospital stay,the women received intravenous treatment for brucellosis(A combination of doxycycline 200 mg/d,rifampicin 900 mg/d,levofloxacin 0.5 g/QD,and ceftriaxone 2 g/QD was administered for 1 week),The L4-S1 vertebral body was fixed by posterior lumbar debridement.Outcome:Six months after discharge,the follow-up radiographic images showed stable vertebral height and good lumbar stability.She complained no discomfort.Lessons:Multi-level involvement is an exceptional form of brucellar spondylitis.To the best of our knowledge,only few similar cases have been reported.PCR and bacterial culture is necessary for confirmed diagnosis.
基金Supported by Natural Science Foundation of Inner Mongolia(2014MS0356)
文摘Brucella spp. are pathogenic to humans and domestic animal. Nowadays,there is no effective vaccine and control strategy in China. So,it is necessary to research effective vaccines for prevention and treatment of this disease. In order to deal with these,we isolated and identified the type of Brucella in Darhan Muminggan Joint Banner and Siziwang Banner of Inner Mongolia. Totally 26 samples of sheep blood which were positive in serological test,one sample of spleen from aborted and one sample of secretion from birth canal were isolated,and the 16 S r DNA genes of positive samples were sequenced. Phylogenetic analysis proved that there were four isolates similar to B. melitensis. As an important diagnostic antigens of B. melitensis,the BP26 gene was amplified. The BP26 gene was cloned into vector p ET24a( +) and conducted sequence analysis. The BP26 gene was 900 bp,with an open reading frame of 753 bp. The homology of BP26 gene with the vaccine strain M5 was 100%,and that with S2 and A19 vaccine strains was 99. 99%. These finding supported the development of BP26-based specific serodiagnostic test and vaccine for B. melitensis in China.
文摘Background: Brucellosis in male goats is characterized by arthritis, orchitis and epididymitis, which may induce infertility. Nevertheless, these lesions were categorized as chronic while acute lesions had not been described. This study investigates the histopathological and immuno histochemistry reactions in organs of bucks acutely infected by Brucella melitensis. Results: Only testis and prepuce of acutely infected bucks showed significantly severe histological lesions. Other internal organs had mild to moderate lesions. However, positive immunohistochemistry stainings were observed in organs except the bulbourethral gland. There was a significant positive correlation between the distribution of B. melitensis and IHC intensity but no significant correlation between the IHC intensity and histopathology lesions. Conclusion: The results indicate that acute brucellosis did not lead to clinical presentation, although B. melitensis was well distributed in various organs of bucks.
文摘Importance of urease activity on pathogenic differences among Brucella species was evaluated. In cell-free extracts, the B. suis urease showed 12 times greater specific activity than the B. melitensis urease. When Fisher-344 rats were inoculated intraperitoneally (IP), at 1 week post-inoculation (PI), B. melitensis wild type 16 M was recovered from spleens and livers in greater numbers than B. suis wild type 1330. At 8 weeks PI, spleens were clear of B. melitensis, whereas B. suis remained. The wild type and the urease deficient strains of B. suis did not differ from each other in terms of recovery from spleen or liver. Our observations suggest that B. melitensis induces greater acute infectivity in Fisher-344 rats, whereas B. suis causes chronic infectivity;and urease activity has no influence on Brucella infection using an IP route.
基金The financial support provided to the first author in the form of Junior Fellowship by ICAR,New Delhi is duly acknowledged
文摘Objective:To explore immunochemical characterization of antigens of Brucella canis(B. canis),and the use in seroprevalence study of canine brucellosis.Methods:External hot phosphate buffer saline extract(HPBSE) and internal sonicated(SA) antigens were prepared from B.canis strain MEX 51 and imniunochemically characterized.These antigens were used to test 527 serum samples of dogs by 2-mercaptoethanol-tubc agglutination test(2 ME-TAT), agar gel immunodiffusion test(AGID).dot-ELISA and indirect enzyme-linked immunosorbent assay(I-ELISA) to assess the seroprevalence of canine brucellosis.Results:The protein content of HPBSE and SA antigens was 0.387 mg/ml.and 0.195 mg/mL,respectively,whereas carbohydrate content was 0.174 mg/mL and 0.150 mg/mL,respectively.The sodium dodecyl sulfate-polyacrylamide gel electrophoresis(12.5%) of HPBSE and SA,revealed 6 and 8 visible peptide bands ranging from 18-80 kDa and 12-45 kDa,respectively.Western blot analysis showed immunodominant bands of MW 12.28.39 and 45 kl)a for HPBSE and 20-24 kl)a for SA. The AGII) revealed HPBSE as more specific antigen than SA but both I-ELISA and dot-ELISA indicated SA antigen to be more specific and reliable than HPBSE.The seroprevalence of canine brucellosis was 2.27%by 2ME-TAT.1.5%by AGID.3.03%by dot-ELISA and 16.12%by I-ELISA. Conclusions:On the basis of the results of present study,we concluded that HPBSE is suitable antigen for AGID,which is more specific:whereas SA antigen is suitable for I-ELISA,which is highly sensitive.Therefore,initial screening of serum samples should be carried out by I-ELISA followed by confirmation with AGID.
文摘Objective: To study Brucella(B.) abortus strains isolated in the Russian Federation, in order to identify their detailed position in the phylogenetic structure of the species global population as well as to determine genetic relationships for isolates from different geographical areas.Methods: Based on Bayesian method, the whole genome singlenucleotide polymorphism(SNP) analysis of 258 B. abortus strains from different geographical areas of the world including 20 B. abortus strains isolated in Russia was carried out. Results: The core genome SNP analysis of the B. abortus isolates allowed describing the main genetic lineages. The Russian strains entered two separate clades, including the basal branch and the C1 branch that is widely spread in Eurasia. The data on the isolation time was used for the dating of phylogenetic tree, and also the estimated time frame for the B. abortus genotype diversification was determined. There were sets of specific SNPs identified, which defined each of the genotypes and sub-genotypes.Conclusions: A significant genetic diversity of the brucellosis pathogen strains from Russia has been proven. The sets of unique specific SNPs described in our study may become one of the elements within a bio-informational analysis algorithm to be used for epidemiological study of brucellosis outbreaks, including those caused by new(atypical) genetic variants of B. abortus.
文摘Brucellosis is a worldwide zoonosis caused by Brucella, which poses great threat to human health. Totally 234 milk samples from a scale dairy farm were amplified by nested PCR, and the pathogens were further separated. The results showed that a DNA band of 419 bp was amplified from 18 out ot＂234 milk samples. Among 18 positive milk samples, nine samples amplified the DNA band of 535 bp, which were B. su/s ; one sample amplified the DNA bands of 535 bp and 285 bp, which was the mixture of B. su/s and B. bov/s. The results laid a foundation for prevention and control of brucellosis.
基金funded by the Colleges and Universities Research Project of Inner Mongolia Autonomous Region(NJzy08165)
文摘[ Objective] To investigate the occurrence and prevalence of Brucella infection in Hulunbiur region. [ Method] A total of 906 serum samples were collected from sheep in Hulunbiur region and detected by the Rose Bengal plate agglutination test to determine the positive rate of Brucella antibodies. [Result] The positive rates of Brucella antibodies in lambs, ewes and mutton sheep were 5.28%, 5.68% and 4.22%, respectively. The total positive rate of Brucella antibodies was 4.86% in sheep. [ Condusionl The positive rate of Brucella antibodies was high in sheep in Hulunblur region.
基金funded by the Natural Science Foundation o China(31060352)
文摘[ Objective] In order to clone and express the formyltransferase of Brucella abortus in E. coli, purify the expressed protein and detect its immunogenicity. EMethods] A gene recoding formyltransferase 27 -35 ku （Wbkc） was amplified from the genomic DNA of Brucella abortus PCR. The amplified fragments were digested with BamH I and Sal I, and then cloned to the vector pET28a. The constructed recombinant plasmid pET28a-Wbkc was transformed to E. coil BL21 and was induced to express the fusion protein. Then the protein was purified by histidine-binding res- in column chromatography, and the immunogenicity was detected by Western blot. The PCR product of Wbkc gene was cloned to the vector pET28a, and the recombinant vector was confirmed by colony PCR identification, recombinant vector digested identification and sequencing analy- sis. [ Results] It was successfully expressed in E. coil BL21 as a fusion protein with histidine at the presence of IPTG, and a specific protein band of 29 ku was found when identified by SDS-PAGE. Western blot showed good immunoreactivity of the expressed product. Wbkc was successfully cloned and expressed, and the purified fusion protein had immunogenicity. This study provides a solid foundation for the further study on the function of proteins and brucellosis diagnostic antigens. [ Conclusion] Amplification of formyltransferase gene of Brucella abortus and prokaryotic expression of WbkC_ indicatino the formvItransferase has specific immune reactivitv.
基金Supported by National Natural Science Foundation of China(31260608)Key Science and Technology Project for Colleges and Universities in Inner Mongolia Autonomous Region(NJZZ12117)Scientific and Technological Cooperation Project between Tongliao City and Universities in Inner Mongolia Autonomous Region(SXZD2012131)
文摘[ Objective ] This study aimed to clone and express formyltransferase （Wbkc） gene from Brucella abortus in E. coli, purify the expressed protein and analyze its immunogenicity. [Method] A gene encoding 27 -35 ku formyltransferase （Wbkc） was amplified from the genomic DNA of BruceUa abortus by PCR. The amplified fragments were digested with BamH I and Sal I, and inserted into pET28a vector. The constructed recombinant plasmid pET 28a-Wbkc was trans- formed into E. coli BL21 and was induced to express the fusion protein. Subsequently, the protein was purified by histidine-binding resin column chromatography, and the immunogenicity was detected by Western blot assay. The recombinant plasmid was identified by colony PCR, double digestion and sequencing analysis. [ Result] Wbkc was successfully cloned and expressed in E. coli. A specific protein band of 29 ku was detected by SDS-PAGE. Western blot showed specific im- munoreactivity of the purified fusion protein. [ Conclusion] This study provided a solid foundation for further investigating diagnostic proteins for brucellosis and developing Brucella gene-deletion vaccines.
