[Show abstract][Hide abstract] ABSTRACT: A better understanding of the kinetics of accumulated immune cells that are involved in pathophysiology during Mycobacterium tuberculosis (Mtb) infection may help to facilitate the development of vaccines and immunological interventions. However, the kinetics of innate and adaptive cells that are associated with pathogenesis during Mtb infection and their relationship to Mtb virulence are not clearly understood. In this study, we used a mouse model to compare the bacterial burden, inflammation and kinetics of immune cells during aerogenic infection in the lung between laboratory-adapted strains (Mtb H37Rv and H37Ra) and Mtb K strain, a hyper-virulent W-Beijing lineage strain. The Mtb K strain multiplied more than 10- and 3.54-fold more rapidly than H37Ra and H37Rv, respectively, during the early stage of infection (at 28 days post-infection) and resulted in exacerbated lung pathology at 56 to 112 days post-infection. Similar numbers of innate immune cells had infiltrated, regardless of the strain, by 14 days post-infection. High, time-dependent frequencies of F4/80-CD11c+CD11b-Siglec-H+PDCA-1+ plasmacytoid DCs and CD11c-CD11b+Gr-1int cells were observed in the lungs of mice that were infected with the Mtb K strain. Regarding adaptive immunity, Th1 and Th17 T cells that express T-bet and RORγt, respectively, significantly increased in the lungs that were infected with the laboratory-adapted strains, and the population of CD4+CD25+Foxp3+ regulatory T cells was remarkably increased at 112 days post-infection in the lungs of mice that were infected with the K strain. Collectively, our findings indicate that the highly virulent Mtb K strain may trigger the accumulation of pDCs and Gr1intCD11b+ cells with the concomitant down-regulation of the Th1 response and the maintenance of an up-regulated Th2 response without inducing a Th17 response during chronic infection. These results will help to determine which immune system components must be considered for the development of tuberculosis (TB) vaccines and immunological interventions.
[Show abstract][Hide abstract] ABSTRACT: A gradual understanding of the proline-glutamate (PE) and proline-proline-glutamate (PPE) families, which compromise 10% of the coding regions in the Mycobacterium tuberculosis (Mtb) genome, has uncovered unique roles in host-pathogen interactions. However, the immunological function of PE27 (Rv2769c), the largest PE member, remains unclear. Here, we explored the functional roles and related signaling mechanisms of PE27 in the interaction with dendritic cells (DCs) to shape the T cell response. PE27 phenotypically and functionally induces DC maturation by up-regulating CD80, CD86, MHC class I and MHC class II expression on the DC surface to promote the production of TNF-α, IL-1β, IL-6, and IL-12p70 but not IL-10. Additionally, we found that PE27-mediated DC activation requires the participation of mitogen-activated protein kinases (MAPKs) and nuclear factor κB (NF-κB) signaling pathways. Interestingly, PE27-treated DCs directed naïve CD4(+) T cells to secrete IFN-γ and activate T-bet but not GATA-3. PE27 also induced IFN-γ-producing memory T cell responses in Mtb-infected mice, indicating that PE27 contributes to Th1-polarization. Taken together, these findings suggest that PE27 possesses Th1-polarizing potential through DC maturation and could be useful in the design of TB vaccines.
[Show abstract][Hide abstract] ABSTRACT: Mycobacterium bovis bacillus Calmette-Guerin (BCG), the only licensed vaccine, shows limited protection efficacy against pulmonary tuberculosis (TB), particularly hypervirulent Mycobacterium tuberculosis (Mtb) strains, suggesting that a logistical and practical vaccination strategy is urgently required. Boosting the BCG-induced immunity may offer a potentially advantageous strategy for advancing TB vaccine development, instead of replacing BCG completely. Despite the improved protection of the airway immunization by using live BCG, the use of live BCG as an airway boosting agent may evoke safety concerns. Here, we analyzed the protective efficacy of γ-irradiated BCG as a BCG-prime boosting agent for airway immunization against a hypervirulent clinical strain challenge with Mycobacterium tuberculosis HN878 in a mouse TB model. After the aerosol challenge with the HN878 strain, the mice vaccinated with BCG via the parenteral route exhibited only mild and transient protection, whereas BCG vaccination followed by multiple aerosolized boosting with γ-irradiated BCG efficiently maintained long-lasting control of Mtb in terms of bacterial reduction and pathological findings. Further immunological investigation revealed that this approach resulted in a significant increase in the cellular responses in terms of a robust expansion of antigen (PPD and Ag85A)-specific CD4+ T cells concomitantly producing IFN-γ, TNF-α, and IL-2, as well as a high level of IFN-γ-producing recall response via both the local and systemic immune systems upon further boosting. Collectively, aerosolized boosting of γ-irradiated BCG is able to elicit strong Th1-biased immune responses and confer enhanced protection against a hypervirulent Mycobacterium tuberculosis HN878 infection in a boosting number-dependent manner.
[Show abstract][Hide abstract] ABSTRACT: Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, is an outstanding pathogen that modulates the host immune response. This inconvenient truth drives the continual identification of antigens that generate protective immunity, including Th1-type T cell immunity. Here, the contribution of methylmalonate semialdehyde dehydrogenase (MmsA, Rv0753c) of Mtb to immune responses was examined in the context of dendritic cell (DC) activation and T cell immunity both in vitro and in vivo. The results showed that MmsA induced DC activation by activating the MAPK and NF-κB signaling pathways. Additionally, MmsA-treated DCs activated naïve T cells, effectively polarized CD4(+) and CD8(+) T cells to secrete IFN-γ and IL-2, and induced T cell proliferation. These results indicate that MmsA is a novel DC maturation-inducing antigen that drives the Th1 immune response. Thus, MmsA was found to potentially regulate immune responses via DC activation toward Th1-type T cell immunity, enhancing our understanding of Mtb pathogenesis.
No preview · Article · Oct 2015 · Cellular Immunology
[Show abstract][Hide abstract] ABSTRACT: The latency and reactivation of Mycobacterium tuberculosis infection has been well studied. However, there have been few studies of the latency and reactivation of Mycobacterium avium complex (MAC), the most common etiological non-tuberculous Mycobacterium species next to M. tuberculosis in humans worldwide. We hypothesized that latent MAC infections can be reactivated following immunosuppression after combination chemotherapy with clarithromycin and rifampicin under experimental conditions. To this end, we employed a modified Cornell-like murine model of tuberculosis and investigated six strains consisting of two type strains and four clinical isolates of M. avium and M. intracellulare. After aerosol infection of each MAC strain, five to six mice per group were euthanized at 2, 4, 10, 18, 28 and 35 weeks post-infection, and lungs were sampled to analyze bacterial burden and histopathology. One strain of each species maintained a culture-negative state for 10 weeks after completion of 6 weeks of chemotherapy, but was reactivated after 5 weeks of immunosuppression in the lungs with dexamethasone (three out of six mice in M. avium infection) or sulfasalazine (four out of six mice in both M. avium and M. intracellulare infection). The four remaining MAC strains exhibited decreased bacterial loads in response to chemotherapy; however, they remained at detectable levels and underwent regrowth after immunosuppression. In addition, the exacerbated lung pathology demonstrated a correlation with bacterial burden after reactivation. In conclusion, our results suggest the possibility of MAC reactivation in an experimental mouse model, and experimentally demonstrate that a compromised immune status can induce reactivation and/or regrowth of MAC infection.
[Show abstract][Hide abstract] ABSTRACT: Purpose: To develop strategies for the control of hospital-acquired methicillin-resistant Staphylococcus aureus (HA-MRSA) which is a serious threat to burns patients with the aid of molecular studies.Methods: Staphylococcus aureus strains were collected from the Burns Unit of Khyber Teaching Hospital (KTH) Peshawar, Pakistan from July - December 2011. Antibiotic resistance was determined according to the recommendations of Clinical Laboratory Standard Institute (CLSI). Molecular epidemiology of the S. aureus strains were determined by pulse field gel electrophoresis (PFGE).Results: PFGE identified 14 clusters which included 29 different pulso-types prevailing in the Burns Unit. Of the 29 types, 11 contained two or more strains of the same pulso-type. These MRSA isolates were highly resistant to various kinds of penicillin and cephalosporin (85 – 100 %). Among the important anti-staphylococal agents tested, 17 % of the isolates were resistant to fusidic acid and linezolid. All the 54 strains were susceptible to vancomycin.Conclusion: Several of the same pulso-types prevail in the Burns Unit of KTH. Furthermore, 29 pulsotypes mong the 54 strains suggest the diversity of the MRSA strains collected from burns patients.
No preview · Article · Jan 2015 · Tropical Journal of Pharmaceutical Research
[Show abstract][Hide abstract] ABSTRACT: In Korea, the addition of antibiotics to feed was banned in July 2011; since then, reports of outbreaks of porcine pleuropneumonia have increased. Therefore, knowledge of the characteristics of clinical isolates emerging in recent years is needed. This study analysed the serotype and antimicrobial susceptibility of 102 Actinobacillus pleuropneumoniae isolates from pigs with pleuropneumonia in Korea from 2006 to 2010. Serotype 5 was the most frequently isolated but the incidence of serotype 1 significantly increased in 2010. The susceptibility to florfenicol decreased considerably from 2006 to 2010, as did that of a minor percentage of isolates to ampicillin and amoxicillin. Of 11 tetracycline resistance genes, tet (B) was predominant (62 per cent), followed by tet (H) and tet (O). The florfenicol resistance (flo R) gene (34 per cent) and beta-lactam antibiotic resistance (bla ROB-1) gene (15 per cent) were also detected. The expression levels of the tetracycline repressor gene and resistance gene were not significantly correlated with the minimum inhibitory concentration values of tetracycline-resistant strains. Interspecies transferability of the resistance genes (flo R, tet (B)/bla ROB-1) was observed with frequencies of 5.7x10(-3) and 3.5x10(-2), respectively. In analysing DNA patterns using pulsed field gel electrophoresis, there was a slight positive correlation with serotypes but not with isolated location and year. This study acknowledges the emergence of new serotype and antimicrobial resistance of A pleuropneumoniae based on the current situation in Korea. This knowledge may be useful in controlling the disease caused by A pleuropneumoniae.
[Show abstract][Hide abstract] ABSTRACT: Tuberculosis (TB) is the second leading infectious cause of mortality worldwide with about two million deaths per year. The only licensed TB vaccine, Mycobacterium bovis bacillus Calmette-Guerin (BCG) shows limited protection efficacy suggesting an improved vaccination strategy is required. Recently, several TB vaccine candidates have entered clinical trials. These vaccine candidates are live mycobacterial vaccines designed to replace BCG or subunit vaccines designed to boost immunity induced by BCG. Vaccines with different strategy such as therapeutic vaccines, which can also be used in combination with drug therapy, are in the early stages of development to resolve latent TB or reactivation from the latent state. In this review, we discuss about development of BCG and BCG-based vaccines and further studies necessary for novel TB vaccine development to sterilize tuberculosis.
Preview · Article · Jan 2014 · Journal of Bacteriology and Virology
[Show abstract][Hide abstract] ABSTRACT: Swine hepatitis E virus (HEV) is widespread throughout pigs in both developing and industrialized countries. This virus is an important zoonotic agent and a public concern worldwide. Infected pigs are asymptomatic, so diagnosing swine HEV relies on detection of the virus or antibodies against the virus. However, several obstacles need to be overcome for effective and practical serological diagnosis. In this study, we developed an enzyme-linked immunosorbent assay (ELISA) that used a purified recombinant capsid protein of swine HEV. The potential clinical use of this assay was evaluated by comparing it with a commercial kit (Genelabs Technologies, Diagnostics, Singapore). Results of the ELISA were highly correlated with those of the commercial kit with a sensitivity of 97% and specificity of 95%. ROC (receiving operator characteristic) analysis of the ELISA data produced a value of 0.987 (95% CI, 0.977~0.998, p < 0.01). The cut-off value for the ELISA was also determined using negative pig sera. In summary, the HEV-specific ELISA developed in the present study appears to be both practical and economical.
Full-text · Article · Dec 2013 · Journal of veterinary science (Suwŏn-si, Korea)
[Show abstract][Hide abstract] ABSTRACT: Field efficacy of enterotoxigenic Escherichia coli-specific phage () as a feed additive was evaluated in weaning piglets. Fifty-four piglets at 3-4 weeks old were allocated in three different groups and two of them were fed with bacteriophage at different concentrations ( feed and feed, respectively) for 30 days. Body weight and feed intake were measured at 10 days interval and body condition and fecal score were inspected every day. Based on the measurement, feed conversion rate (FCR) and average daily gain (ADG) of each group during 30 days were analyzed. The analysis suggests that the bacteriophage may help the improvement of FCR and ADG at of bacteriophage feeding group in 30 days. A result from analysis of fecal score indicates that the bacteriophage also may help to relieve the intermittent diarrhea in post-weaning stage. Those results suggest that bacteriophage might help the growth of piglets in post-weaning stage.
Full-text · Article · Jun 2013 · Korean Journal of Veterinary Research
[Show abstract][Hide abstract] ABSTRACT: Brucella abortus is an intracellular zoonotic pathogen which causes undulant fever, endocarditis, arthritis and osteomyelitis in human and abortion and infertility in cattle. This bacterium is able to invade and replicate in host macrophage instead of getting removed by this defense mechanism. Therefore, understanding the interaction between virulence of the bacteria and the host cell is important to control brucellosis. Previously, we generated internalization defective mutants and analyzed the envelope proteins. The present study was undertaken to evaluate the changes in early transcriptional responses between wild type and internalization defective mutants infected mouse macrophage, RAW 264.7.
Both of the wild type and mutant infected macrophages showed increased expression levels in proinflammatory cytokines, chemokines, apoptosis and G-protein coupled receptors (Gpr84, Gpr109a and Adora2b) while the genes related with small GTPase which mediate intracellular trafficking was decreased. Moreover, cytohesin 1 interacting protein (Cytip) and genes related to ubiquitination (Arrdc3 and Fbxo21) were down-regulated, suggesting the survival strategy of this bacterium. However, we could not detect any significant changes in the mutant infected groups compared to the wild type infected group.
In summary, it was very difficult to clarify the alterations in host cellular transcription in response to infection with internalization defective mutants. However, we found several novel gene changes related to the GPCR system, ubiquitin-proteosome system, and growth arrest and DNA damages in response to B. abortus infection. These findings may contribute to a better understanding of the molecular mechanisms underlying host-pathogen interactions and need to be studied further.
[Show abstract][Hide abstract] ABSTRACT: Paratuberculosis (PTB) or Johne's disease (JD) is one of the most serious chronic debilitating diseases of ruminants worldwide that is caused by Mycobacterium avium subsp. paratuberculosis (MAP). MAP is a slow-growing bacterium that has very long latent periods resulting in difficulties in diagnosing and controlling the disease, especially regarding the diagnosis of fecal shedders of MAP without any clinical signs. Based on this situation, our attempts were made to identify biomarkers which were shown early responses to M. paratuberculosis infection in professional macrophage cell line, RAW264.7. In response to the infection with the bacterium, a lot of genes were turn on and/or turn off in the cells. Of the altered genes, three different categories were identified based on the time-dependent gene expression patterns. Those genes were considered as possible candidates for biomarkers of M. paratuberculosis infection after confirmation by quantitative RT-PCR analysis. To the best of our knowledge, this is the first attempt at discovering the host transcriptomic biomarkers of PBT, although it will be required further investigation to determine whether these biomarker candidates are associated within the natural host.
No preview · Article · Jun 2013 · Journal of Microbiology and Biotechnology
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to applicate and evaluate a SYBR Green real-time PCR for the specific detection of Salmonella spp. Specificity of the PCR method was confirmed with 48 Salmonella spp. and 5 non-Salmonella strains using invA gene primer. The average threshold cycle () of Salmonella spp. was while non-Salmonella spp. was . Correlation coefficients of standard curves constructed using versus copy number of Salmonella Enteritidis ATCC 13076 showed good linearity (; slope = 3.563). Minimum level of detection with the method was > colony forming units (CFU)/mL. These results suggested that the SYBR Green real-time PCR might be applicable for the specific detection of Salmonella spp. isolates.
Full-text · Article · Mar 2013 · Korean Journal of Veterinary Research
[Show abstract][Hide abstract] ABSTRACT: Key words: Bovine herpesvirus-1 Calves Germanium biotite Immune stimulator Mannheimia haemolytica Serotype A1 Germanium biotite, a natural mineral, is comprised of mainly silicate. This mineral showed activities of increase in feed efficiency and non-specific immunostimulation in previous studies. The aims of the present study were to evaluate the prophylactic effects of germanium biotite against respiratory diseases in calves as a feed supplement and investigate the possibilities of the substitution of antibiotics with germanium biotite as feed additive. To achieve these purposes, bovine herpesvirus-1 (BHV-1) and Mannheimia haemolytica serotype A1 were experimentally inoculated into the calves. After challenge, germanium biotite showed a lower cumulative clinical score (CCS) than the control group. In accordance with these clinical results, enhanced clearance of BHV-1, a low infection rate of Mannheimia haemolytica serotype A1, tempered superficial lesions, and moderated histopathological signs were observed in the germanium biotite group, compared with the control group. The results of the present study indicated that germanium biotite had prophylactic effects against bovine respiratory disease and could be a candidate for a new alternative feed supplement in calves, through its effects as a non-specific immune stimulator., 2012. The effects of germanium biotite supplement as a prophylactic agent against respiratory infection in calves. Pak Vet J, 32(3): 319-324.
[Show abstract][Hide abstract] ABSTRACT: We evaluated effect of enterotoxigenic Escherichia coli (ETEC) specific lytic phage CJ12 in ETEC infected pigs. Phage was mixed with feed at a ratio of 1:1,000 (0.1%). One week after initially providing phage mixed feed, pigs were challenged orally with 10(11) CFU of ETEC and body weight, diarrhea score, bacterial CFU and phage PFU in the feces were measured. Pigs of phage treated groups C (10(6) PFU/g) and D (10(8) PFU/g) showed more resistance to diarrhea due to ETEC infection compared to positive control group B on the third day after the initial challenge. Moreover, during the quantitation of ETEC in feces, both groups C and D showed approximately 63.92 and 60.73% reduced ETEC compared to positive control group B. Phages were successfully isolated from feces in both groups C and D during the experiment without any adverse effects, suggesting the possibility of using CJ12 as a feed additive.
No preview · Article · Mar 2012 · Journal of Veterinary Medical Science
[Show abstract][Hide abstract] ABSTRACT: Germanium biotite, a natural mineral, is comprised of mainly silicate. This mineral showed activities of increase in feed efficiency and non-specific immunostimulation in previous studies. The aims of the present study were to evaluate the prophylactic effects of germanium biotite against respiratory diseases in calves as a feed supplement and investigate the possibilities of the substitution of antibiotics with germanium biotite as feed additive. To achieve these purposes, bovine herpesvirus-1 (BHV-1) and Mannheimia haemolytica serotype A1 were experimentally inoculated into the calves. After challenge, germanium biotite showed a lower cumulative clinical score (CCS) than the control group. In accordance with these clinical results, enhanced clearance of BHV-1, a low infection rate of Mannheimia haemolytica serotype A1, tempered superficial lesions, and moderated histopathological signs were observed in the germanium biotite group, compared with the control group. The results of the present study indicated that germanium biotite had prophylactic effects against bovine respiratory disease and could be a candidate for a new alternative feed supplement in calves, through its effects as a non-specific immune stimulator.
No preview · Article · Jan 2012 · Pakistan Veterinary Journal
[Show abstract][Hide abstract] ABSTRACT: Germanium biotite, a natural mineral, has been used as a feed supplement to reinforce innate immune ability. The aim of the present study was to evaluate the effects of germanium biotite on the adsorptive and inhibition of growth abilities against Escherichia (E.) coli and Salmonella spp. in vitro. Two strains of enterotoxigenic E. coli and four strains of two Salmonella serotypes (Salmonella Derby and Salmonella Typhimurium), major bacterial diarrheal pathogens, were used for this experiment. The absorptive ability of germanium biotite against most Salmonella used in present experiment was observed weakly. The germanium biotite, however, showed significant effect of bacterial growth inhibition in most experiment bacteria. These results suggest that the use of the germanium biotite as feed supplement could alleviate diarrhea following inhibition of bacteria growth. It is also presumed that antibiotics usage for farm animals, considered as causes of antibiotic residue in meat and emerging antibiotic resistance, could be reduced through the use of germanium biotite as a feed supplement, in place of antibiotics used for the prevention of diarrhea.
No preview · Article · Jan 2012 · Korean Journal of Veterinary Research
[Show abstract][Hide abstract] ABSTRACT: Brucella abortus is a facultative intracellular bacteria that replicates within a macrophage without producing any classical virulence factors. It can become internalized to cells by zipper-like and/or swimming internalization mechanisms. However, the bacterial proteins involved in internalization remain unclear. To define these bacterial proteins, random insertion mutants of B. abortus were generated by the Tn5 transposome complexes. In all, 132 mutants were screened, cellular internalization-defective mutants were selected, and these genomic and envelope proteomic features were identified. The transposon insertion sites were ccmC,ppk and BruAb2_0168 for the mutant C10, C29 and D7, respectively. Mutant C10 showed a deficiency in internalization without any changes in expression of the cell envelope proteins; however, mutant C29 showed a reduced expression of OMP25, and a mutant D7 also showed reduced expression of OMP25, OMP28 and Porin2b. These results suggest OMP25 is not an essential factor, but might be involved in host cellular internalization. We identified the ppk gene and BruAb2_0168 locus which are associated to expression of OMP25, OMP28 and Porin2b as well as pleiotropic effects of ccmC gene.
Full-text · Article · Nov 2011 · FEMS Immunology & Medical Microbiology
[Show abstract][Hide abstract] ABSTRACT: Actinobacillus pleuropneumoniae is the etiologic agent of porcine pleuropneumonia, a highly contagious pulmonary disease in pigs with major economic losses for pig producers worldwide. Whereas A. pleuropneumoniae isolates are divided into 15 serotypes, the isolates secrete 4 types of exotoxins (ApxI, ApxII, ApxIII, and ApxIV), which are known as major virulence factors. In the current study, the ApxIA, ApxIIA, and ApxIIIA genes were amplified and their recombinant proteins expressed in Escherichia coli M15 cells. The antigenicity of each recombinant protein was demonstrated by Western blot and enzyme-linked immunosorbent assay (ELISA) using sera from pigs vaccinated with a subunit vaccine. When ELISAs using the recombinant antigens were optimized and then applied to sera from 320 randomized pigs in Korea, an observed increase in seroprevalence was found among sows in comparison with weaned piglets and growing pigs, indicating an age-dependent seroprevalence. The results obtained in the study suggest that the developed ELISAs may be useful for A. pleuropneumoniae vaccination strategy as a screening tool for pig herds as well as for detection of specific antibodies to Apx exotoxins.
Full-text · Article · Jul 2011 · Journal of veterinary diagnostic investigation: official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc