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Trudeau Institute Inc m. tuberculosis strain erdman

WT and B cell-deficient μMT mice were aerogenically infected with 100 CFU of M . <t>tuberculosis</t> <t>Erdman</t> and tissues analyzed by examination of H&E-stained lung sections at 12 weeks (A) and 24 weeks (C) post-infection. Quantification of the number of total lung cells was also carried out at the same time intervals post-inoculation (12 weeks, (B); 24 weeks (D)). These histological studies and enumeration of total lung cell numbers showed that in chronic TB, B cell-deficiency is associated with diminished granulomatous inflammatory response; n = 3–4 mice per group per time interval. The results depicted in (A) and (C) are representative of lung sections from 3 mice examined. The data shown in (B) and (D) denote the mean ± SEM (standard error mean). The results shown in (A, B, C, and D) are representative of 2 experiments. (E) Lung mycobacterial burden as measured by culturable bacilli 4, 12, and 24 weeks after a 100 CFU Erdman aerogenic challenge, demonstrating that bacillary loads in μMT and WT mice are comparable throughout the course of infection; n = 4 to 5 mice per group. The data depicted denote the mean ± SEM. The results shown are representative of two experiments. (F) B cell-deficient μMT mice exhibited increased median survival relative to WT controls after a 100 CFU aerosol Erdman infection (median survival time: 324 days versus 272 days); n = 10 mice per group. The result is demonstrable in three independent experiments.

M. Tuberculosis Strain Erdman, supplied by Trudeau Institute Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice"

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

Journal: PLOS Pathogens

doi: 10.1371/journal.ppat.1011187

WT and B cell-deficient μMT mice were aerogenically infected with 100 CFU of M . tuberculosis Erdman and tissues analyzed by examination of H&E-stained lung sections at 12 weeks (A) and 24 weeks (C) post-infection. Quantification of the number of total lung cells was also carried out at the same time intervals post-inoculation (12 weeks, (B); 24 weeks (D)). These histological studies and enumeration of total lung cell numbers showed that in chronic TB, B cell-deficiency is associated with diminished granulomatous inflammatory response; n = 3–4 mice per group per time interval. The results depicted in (A) and (C) are representative of lung sections from 3 mice examined. The data shown in (B) and (D) denote the mean ± SEM (standard error mean). The results shown in (A, B, C, and D) are representative of 2 experiments. (E) Lung mycobacterial burden as measured by culturable bacilli 4, 12, and 24 weeks after a 100 CFU Erdman aerogenic challenge, demonstrating that bacillary loads in μMT and WT mice are comparable throughout the course of infection; n = 4 to 5 mice per group. The data depicted denote the mean ± SEM. The results shown are representative of two experiments. (F) B cell-deficient μMT mice exhibited increased median survival relative to WT controls after a 100 CFU aerosol Erdman infection (median survival time: 324 days versus 272 days); n = 10 mice per group. The result is demonstrable in three independent experiments.

Figure Legend Snippet: WT and B cell-deficient μMT mice were aerogenically infected with 100 CFU of M . tuberculosis Erdman and tissues analyzed by examination of H&E-stained lung sections at 12 weeks (A) and 24 weeks (C) post-infection. Quantification of the number of total lung cells was also carried out at the same time intervals post-inoculation (12 weeks, (B); 24 weeks (D)). These histological studies and enumeration of total lung cell numbers showed that in chronic TB, B cell-deficiency is associated with diminished granulomatous inflammatory response; n = 3–4 mice per group per time interval. The results depicted in (A) and (C) are representative of lung sections from 3 mice examined. The data shown in (B) and (D) denote the mean ± SEM (standard error mean). The results shown in (A, B, C, and D) are representative of 2 experiments. (E) Lung mycobacterial burden as measured by culturable bacilli 4, 12, and 24 weeks after a 100 CFU Erdman aerogenic challenge, demonstrating that bacillary loads in μMT and WT mice are comparable throughout the course of infection; n = 4 to 5 mice per group. The data depicted denote the mean ± SEM. The results shown are representative of two experiments. (F) B cell-deficient μMT mice exhibited increased median survival relative to WT controls after a 100 CFU aerosol Erdman infection (median survival time: 324 days versus 272 days); n = 10 mice per group. The result is demonstrable in three independent experiments.

Techniques Used: Infection, Staining, Aerosol

WT and μMT mice were infected with 100 CFU M . tuberculosis Erdman delivered via aerosolization. Dilution of CFSE by ex vivo lung cultures taken 5 months after infection. Cells from 3 mice per group were pooled for the analysis. Data in (B and C) show the frequencies of BrdU + CD4 + T cells in the lungs of WT and μMT mice derived from in vivo BrdU labeling studies conducted at 5 months post-infection. The data shown in (B) depicts a representative dot plot targeting BrdU + CD4 + T cells. Results shown in (C) are mean ± SEM of the frequencies of BrdU + CD4 + T cells. Three to 4 mice per group were evaluated. (D) The absolute number of lung BrdU + CD4 + T cells at 5 months after infection upon in vivo BrdU labeling. The data shown depict mean ± SEM of 3 to 4 mice per group. Data shown are representative of 2 to 3 experiments, except for that in (A), which are derived from 1 study.

Figure Legend Snippet: WT and μMT mice were infected with 100 CFU M . tuberculosis Erdman delivered via aerosolization. Dilution of CFSE by ex vivo lung cultures taken 5 months after infection. Cells from 3 mice per group were pooled for the analysis. Data in (B and C) show the frequencies of BrdU + CD4 + T cells in the lungs of WT and μMT mice derived from in vivo BrdU labeling studies conducted at 5 months post-infection. The data shown in (B) depicts a representative dot plot targeting BrdU + CD4 + T cells. Results shown in (C) are mean ± SEM of the frequencies of BrdU + CD4 + T cells. Three to 4 mice per group were evaluated. (D) The absolute number of lung BrdU + CD4 + T cells at 5 months after infection upon in vivo BrdU labeling. The data shown depict mean ± SEM of 3 to 4 mice per group. Data shown are representative of 2 to 3 experiments, except for that in (A), which are derived from 1 study.

Techniques Used: Infection, Ex Vivo, Derivative Assay, In Vivo, Labeling

Mice were infected with 100 CFU of M . tuberculosis Erdman by aerosolization. (A) Representative frequencies of IFN-γ + CD4 + T cells detected by flow cytometry at 5 months after infection; n = 4 mice per group. (B) Absolute number of lung IFN-γ + CD4 + T cells detected 5 months after infection; n = 4 mice per group. The results are expressed as mean ± SEM. Data in this figure is representative of 3 similar experiments. The data show revealed the B cell-deficient μMT mice, relative to WT, display an attenuated Th1 response during the chronic phase of tuberculous infection.

Figure Legend Snippet: Mice were infected with 100 CFU of M . tuberculosis Erdman by aerosolization. (A) Representative frequencies of IFN-γ + CD4 + T cells detected by flow cytometry at 5 months after infection; n = 4 mice per group. (B) Absolute number of lung IFN-γ + CD4 + T cells detected 5 months after infection; n = 4 mice per group. The results are expressed as mean ± SEM. Data in this figure is representative of 3 similar experiments. The data show revealed the B cell-deficient μMT mice, relative to WT, display an attenuated Th1 response during the chronic phase of tuberculous infection.

Techniques Used: Infection, Flow Cytometry

Mice were aerogenically infected with a low dose (100 CFU) of M . tuberculosis Erdman. Ex vivo IL-10 production by lung cells isolated from B cell-deficient μMT and WT control mice was evaluated at 12 weeks (A) and 24 weeks (B) after infection. Single-cell suspension of lung cells were cultured in complete RPMI with or without 10 μg/ml of PPD (Statens Serum Institute, Copenhagen) at 1.0 x 10 7 cells per ml. After 48 hours of culture, supernatants were collected and subjected to detection of IL-10 as described in “Materials and Methods” section. The results revealed that the levels of expression of lung IL-10 in tuberculous μMT mice are enhanced compared to that observed in WT animals. The data are depicted as mean ± SEM. The results shown in this figure are representative of two independent experiments; n = 4 to 5 mice per group per time interval.

Figure Legend Snippet: Mice were aerogenically infected with a low dose (100 CFU) of M . tuberculosis Erdman. Ex vivo IL-10 production by lung cells isolated from B cell-deficient μMT and WT control mice was evaluated at 12 weeks (A) and 24 weeks (B) after infection. Single-cell suspension of lung cells were cultured in complete RPMI with or without 10 μg/ml of PPD (Statens Serum Institute, Copenhagen) at 1.0 x 10 7 cells per ml. After 48 hours of culture, supernatants were collected and subjected to detection of IL-10 as described in “Materials and Methods” section. The results revealed that the levels of expression of lung IL-10 in tuberculous μMT mice are enhanced compared to that observed in WT animals. The data are depicted as mean ± SEM. The results shown in this figure are representative of two independent experiments; n = 4 to 5 mice per group per time interval.

Techniques Used: Infection, Ex Vivo, Isolation, Control, Suspension, Cell Culture, Expressing

Mice were treated with the anti-CD20 mAb 5D2 beginning 2 days prior to a low-dose (100 CFU) aerogenic challenge with M . tuberculosis Erdman, as described in

Figure Legend Snippet: Mice were treated with the anti-CD20 mAb 5D2 beginning 2 days prior to a low-dose (100 CFU) aerogenic challenge with M . tuberculosis Erdman, as described in "Materials and Methods". B cell depletion was maintained throughout the duration of the experiment. The control mouse group (WT) received non-specific rat IgG. The lung tissues were examined at 5 months post-infection. The levels of granulomatous inflammation response were analyzed histologically by light microscopy on H&E-stained lung sections (A) and enumeration of total number of lung cells (B). The level of lung CD4 + T cell response was assessed by in vivo BrdU labeling to examine the proliferation capacity of this T cell subset (C), as well as by enumeration of IFN-γ-producing CD4 + T cells (D). Ex vivo evaluation of lung cells for the level of IL-10 production (E) was conducted as described in . Four to 5 mice per group were evaluated per group. Data depicted in B, C, D, and E are presented as means ± SEM. The data shown are representative of two experiments. The results demonstrated that the inflammation, Th1 response, and IL-10 phenotypes observed in the μMT mice are recapitulated in mice depleted for B cells.

Techniques Used: Control, Infection, Light Microscopy, Staining, In Vivo, Labeling, Ex Vivo

C57BL/6 mice were depleted for B cells via administration of 5D2 beginning 2 days prior to infection with a low dose (100 CFU) of M . tuberculosis Erdman delivered by aerosol. B cell depletion was maintained for the duration of the experiment. At 3 months after the infection, IL-10R blockade was initiated using the anti-mouse IL-10R antibody clone 1B1.3A. The control group received non-specific rat IgG. The IL-10R blockade was continued for two months. At five months post-infection (2 months after initiation of IL-10R blockade), mice were sacrificed and analyzed for the levels of inflammation in the lungs, as assessed by histological examination (A) and enumeration of total lung cells (B), CD4 + T cells proliferation via BrdU labeling (C), and Th1 response (D). Data shown are representation of two experiments. Three to four mice were analyzed per group. Data depicted in (B), (C), and (D) denote mean ± SEM.

Figure Legend Snippet: C57BL/6 mice were depleted for B cells via administration of 5D2 beginning 2 days prior to infection with a low dose (100 CFU) of M . tuberculosis Erdman delivered by aerosol. B cell depletion was maintained for the duration of the experiment. At 3 months after the infection, IL-10R blockade was initiated using the anti-mouse IL-10R antibody clone 1B1.3A. The control group received non-specific rat IgG. The IL-10R blockade was continued for two months. At five months post-infection (2 months after initiation of IL-10R blockade), mice were sacrificed and analyzed for the levels of inflammation in the lungs, as assessed by histological examination (A) and enumeration of total lung cells (B), CD4 + T cells proliferation via BrdU labeling (C), and Th1 response (D). Data shown are representation of two experiments. Three to four mice were analyzed per group. Data depicted in (B), (C), and (D) denote mean ± SEM.

Techniques Used: Infection, Aerosol, Control, Labeling

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Journal: PLOS Pathogens

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

doi: 10.1371/journal.ppat.1011187

Figure Lengend Snippet: WT and B cell-deficient μMT mice were aerogenically infected with 100 CFU of M . tuberculosis Erdman and tissues analyzed by examination of H&E-stained lung sections at 12 weeks (A) and 24 weeks (C) post-infection. Quantification of the number of total lung cells was also carried out at the same time intervals post-inoculation (12 weeks, (B); 24 weeks (D)). These histological studies and enumeration of total lung cell numbers showed that in chronic TB, B cell-deficiency is associated with diminished granulomatous inflammatory response; n = 3–4 mice per group per time interval. The results depicted in (A) and (C) are representative of lung sections from 3 mice examined. The data shown in (B) and (D) denote the mean ± SEM (standard error mean). The results shown in (A, B, C, and D) are representative of 2 experiments. (E) Lung mycobacterial burden as measured by culturable bacilli 4, 12, and 24 weeks after a 100 CFU Erdman aerogenic challenge, demonstrating that bacillary loads in μMT and WT mice are comparable throughout the course of infection; n = 4 to 5 mice per group. The data depicted denote the mean ± SEM. The results shown are representative of two experiments. (F) B cell-deficient μMT mice exhibited increased median survival relative to WT controls after a 100 CFU aerosol Erdman infection (median survival time: 324 days versus 272 days); n = 10 mice per group. The result is demonstrable in three independent experiments.

Article Snippet: Bacterial stock of M . tuberculosis strain Erdman (Dr. Frank Collins, Trudeau Institute, Saranac Lake, NY) was prepared by passage through mice to maintain virulence, expanded once by culture in 7H9 liquid medium (Difco), and stored in 3x10 8 bacilli/ml aliquots at -80°C until use.

Techniques: Infection, Staining, Aerosol

Journal: PLOS Pathogens

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

doi: 10.1371/journal.ppat.1011187

Figure Lengend Snippet: WT and μMT mice were infected with 100 CFU M . tuberculosis Erdman delivered via aerosolization. Dilution of CFSE by ex vivo lung cultures taken 5 months after infection. Cells from 3 mice per group were pooled for the analysis. Data in (B and C) show the frequencies of BrdU + CD4 + T cells in the lungs of WT and μMT mice derived from in vivo BrdU labeling studies conducted at 5 months post-infection. The data shown in (B) depicts a representative dot plot targeting BrdU + CD4 + T cells. Results shown in (C) are mean ± SEM of the frequencies of BrdU + CD4 + T cells. Three to 4 mice per group were evaluated. (D) The absolute number of lung BrdU + CD4 + T cells at 5 months after infection upon in vivo BrdU labeling. The data shown depict mean ± SEM of 3 to 4 mice per group. Data shown are representative of 2 to 3 experiments, except for that in (A), which are derived from 1 study.

Techniques: Infection, Ex Vivo, Derivative Assay, In Vivo, Labeling

Journal: PLOS Pathogens

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

doi: 10.1371/journal.ppat.1011187

Figure Lengend Snippet: Mice were infected with 100 CFU of M . tuberculosis Erdman by aerosolization. (A) Representative frequencies of IFN-γ + CD4 + T cells detected by flow cytometry at 5 months after infection; n = 4 mice per group. (B) Absolute number of lung IFN-γ + CD4 + T cells detected 5 months after infection; n = 4 mice per group. The results are expressed as mean ± SEM. Data in this figure is representative of 3 similar experiments. The data show revealed the B cell-deficient μMT mice, relative to WT, display an attenuated Th1 response during the chronic phase of tuberculous infection.

Techniques: Infection, Flow Cytometry

Journal: PLOS Pathogens

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

doi: 10.1371/journal.ppat.1011187

Figure Lengend Snippet: Mice were aerogenically infected with a low dose (100 CFU) of M . tuberculosis Erdman. Ex vivo IL-10 production by lung cells isolated from B cell-deficient μMT and WT control mice was evaluated at 12 weeks (A) and 24 weeks (B) after infection. Single-cell suspension of lung cells were cultured in complete RPMI with or without 10 μg/ml of PPD (Statens Serum Institute, Copenhagen) at 1.0 x 10 7 cells per ml. After 48 hours of culture, supernatants were collected and subjected to detection of IL-10 as described in “Materials and Methods” section. The results revealed that the levels of expression of lung IL-10 in tuberculous μMT mice are enhanced compared to that observed in WT animals. The data are depicted as mean ± SEM. The results shown in this figure are representative of two independent experiments; n = 4 to 5 mice per group per time interval.

Techniques: Infection, Ex Vivo, Isolation, Control, Suspension, Cell Culture, Expressing

Journal: PLOS Pathogens

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

doi: 10.1371/journal.ppat.1011187

Figure Lengend Snippet: Mice were treated with the anti-CD20 mAb 5D2 beginning 2 days prior to a low-dose (100 CFU) aerogenic challenge with M . tuberculosis Erdman, as described in "Materials and Methods". B cell depletion was maintained throughout the duration of the experiment. The control mouse group (WT) received non-specific rat IgG. The lung tissues were examined at 5 months post-infection. The levels of granulomatous inflammation response were analyzed histologically by light microscopy on H&E-stained lung sections (A) and enumeration of total number of lung cells (B). The level of lung CD4 + T cell response was assessed by in vivo BrdU labeling to examine the proliferation capacity of this T cell subset (C), as well as by enumeration of IFN-γ-producing CD4 + T cells (D). Ex vivo evaluation of lung cells for the level of IL-10 production (E) was conducted as described in . Four to 5 mice per group were evaluated per group. Data depicted in B, C, D, and E are presented as means ± SEM. The data shown are representative of two experiments. The results demonstrated that the inflammation, Th1 response, and IL-10 phenotypes observed in the μMT mice are recapitulated in mice depleted for B cells.

Techniques: Control, Infection, Light Microscopy, Staining, In Vivo, Labeling, Ex Vivo

Journal: PLOS Pathogens

Article Title: B cells promote granulomatous inflammation during chronic Mycobacterium tuberculosis infection in mice

doi: 10.1371/journal.ppat.1011187

Figure Lengend Snippet: C57BL/6 mice were depleted for B cells via administration of 5D2 beginning 2 days prior to infection with a low dose (100 CFU) of M . tuberculosis Erdman delivered by aerosol. B cell depletion was maintained for the duration of the experiment. At 3 months after the infection, IL-10R blockade was initiated using the anti-mouse IL-10R antibody clone 1B1.3A. The control group received non-specific rat IgG. The IL-10R blockade was continued for two months. At five months post-infection (2 months after initiation of IL-10R blockade), mice were sacrificed and analyzed for the levels of inflammation in the lungs, as assessed by histological examination (A) and enumeration of total lung cells (B), CD4 + T cells proliferation via BrdU labeling (C), and Th1 response (D). Data shown are representation of two experiments. Three to four mice were analyzed per group. Data depicted in (B), (C), and (D) denote mean ± SEM.

Techniques: Infection, Aerosol, Control, Labeling

Mice were infected with M. tuberculosis strain Erdman and treated with drugs starting 18 to 21 days after aerosol infection; with isoniazid (INH), rifampin (RIF), ethambutol (EMB), gatifloxacin (Gati) or moxifloxacin (Moxi). Sacrifice times were at 2, 5 and 7 days of drug treatment. Data points represent mean log 10 viable bacilli +/− standard error present in whole lung homogenates.

Journal: PLoS ONE

Article Title: Location of Intra- and Extracellular M. tuberculosis Populations in Lungs of Mice and Guinea Pigs during Disease Progression and after Drug Treatment

doi: 10.1371/journal.pone.0017550

Figure Lengend Snippet: Mice were infected with M. tuberculosis strain Erdman and treated with drugs starting 18 to 21 days after aerosol infection; with isoniazid (INH), rifampin (RIF), ethambutol (EMB), gatifloxacin (Gati) or moxifloxacin (Moxi). Sacrifice times were at 2, 5 and 7 days of drug treatment. Data points represent mean log 10 viable bacilli +/− standard error present in whole lung homogenates.

Article Snippet: Mice were exposed to a low-dose aerosol infection with the M. tuberculosis strain Erdman ( TMC 107; ATCC 35801) in a Glas-Col inhalation exposure system (Glas-Col Inc., Terre Haute, IN) .

Techniques: Infection

Efficacy against M. tuberculosis aerosol challenge of boosting BCG-primed mice with a combination of recombinant attenuated L. monocytogenes vaccines expressing 5 M. tuberculosis antigens. (a) Efficacy of boosting with 5Ag recombinant attenuated L. monocytogenes vaccines i.m. once versus twice. BALB/c mice ( n = 8/group) were immunized i.d. with PBS or with BCG at week 0. BCG-primed mice were either not boosted or boosted intramuscularly (i.m.) or subcutaneously (s.c.) once (×1) at week 18 with L. monocytogenes vector (LmV) or the combination of rLm4Ag (expressing M. tuberculosis fusion protein Mpt64-TB10.4-ESAT6-CFP10) and rLm30. An additional group was boosted twice (×2) at weeks 14 and 18 with rLm4Ag plus rLm30. The mice were then challenged with aerosolized M. tuberculosis Erdman (average of 19 CFU delivered to the lungs of each animal, as assayed at day 1 postchallenge to 2 mice) at week 22 and euthanized at week 32 (top). Lungs (bottom left) and spleens (bottom right) of mice were assayed for organ bacterial burden. Shown are means ± SEM. One symbol represents one animal. Organ log 10 CFU were analyzed by one-way ANOVA. *, P < 0.05; ****, P < 0.0001 by Tukey’s multiple-comparison test; #, P < 0.05 by Fisher’s LSD test (Prism 9.2.0). (b) Efficacy of 5Ag recombinant attenuated L. monocytogenes vaccines by route of administration. BALB/c mice (8/group) were immunized i.d. with PBS (sham) or 5 × 10 5 CFU BCG at week 0. Mice immunized i.d. with BCG were either not boosted or boosted i.m., i.d., subcutaneously (s.c.), or intravenously (i.v.) once at week 18 with 2 × 10 6 CFU of the combination of rLm4Ag (expressing Mpt64-TB10.4-ESAT6-CFP10) and rLm30. The mice were then challenged at week 22 with aerosolized M. tuberculosis Erdman at the same time as the companion experiment shown in panel a (average of 19 CFU delivered to the lungs of each animal) and euthanized at week 26 (top). Afterward, lungs (bottom left) and spleens (bottom right) were removed and assayed for bacillus burdens. Shown are means ± SEM. Each symbol represents one mouse. *, P < 0.05; ***, P < 0.001; ****, P < 0.0001 by one-way ANOVA with Fisher’s LSD test (Prism 9.2.0).

Journal: mBio

Article Title: Listeria -Vectored Multiantigenic Tuberculosis Vaccine Enhances Protective Immunity against Aerosol Challenge with Virulent Mycobacterium tuberculosis in BCG-Immunized C57BL/6 and BALB/c Mice

doi: 10.1128/mbio.00687-22

Figure Lengend Snippet: Efficacy against M. tuberculosis aerosol challenge of boosting BCG-primed mice with a combination of recombinant attenuated L. monocytogenes vaccines expressing 5 M. tuberculosis antigens. (a) Efficacy of boosting with 5Ag recombinant attenuated L. monocytogenes vaccines i.m. once versus twice. BALB/c mice ( n = 8/group) were immunized i.d. with PBS or with BCG at week 0. BCG-primed mice were either not boosted or boosted intramuscularly (i.m.) or subcutaneously (s.c.) once (×1) at week 18 with L. monocytogenes vector (LmV) or the combination of rLm4Ag (expressing M. tuberculosis fusion protein Mpt64-TB10.4-ESAT6-CFP10) and rLm30. An additional group was boosted twice (×2) at weeks 14 and 18 with rLm4Ag plus rLm30. The mice were then challenged with aerosolized M. tuberculosis Erdman (average of 19 CFU delivered to the lungs of each animal, as assayed at day 1 postchallenge to 2 mice) at week 22 and euthanized at week 32 (top). Lungs (bottom left) and spleens (bottom right) of mice were assayed for organ bacterial burden. Shown are means ± SEM. One symbol represents one animal. Organ log 10 CFU were analyzed by one-way ANOVA. *, P < 0.05; ****, P < 0.0001 by Tukey’s multiple-comparison test; #, P < 0.05 by Fisher’s LSD test (Prism 9.2.0). (b) Efficacy of 5Ag recombinant attenuated L. monocytogenes vaccines by route of administration. BALB/c mice (8/group) were immunized i.d. with PBS (sham) or 5 × 10 5 CFU BCG at week 0. Mice immunized i.d. with BCG were either not boosted or boosted i.m., i.d., subcutaneously (s.c.), or intravenously (i.v.) once at week 18 with 2 × 10 6 CFU of the combination of rLm4Ag (expressing Mpt64-TB10.4-ESAT6-CFP10) and rLm30. The mice were then challenged at week 22 with aerosolized M. tuberculosis Erdman at the same time as the companion experiment shown in panel a (average of 19 CFU delivered to the lungs of each animal) and euthanized at week 26 (top). Afterward, lungs (bottom left) and spleens (bottom right) were removed and assayed for bacillus burdens. Shown are means ± SEM. Each symbol represents one mouse. *, P < 0.05; ***, P < 0.001; ****, P < 0.0001 by one-way ANOVA with Fisher’s LSD test (Prism 9.2.0).

Article Snippet: M. tuberculosis Erdman strain (ATCC 35801) was harvested from infected outbred guinea pigs to verify virulence, cultured on 7H11 agar, subjected to gentle sonication to obtain a single-cell suspension, and frozen at −80°C for use in animal challenge experiments.

Techniques: Aerosol, Recombinant, Vaccines, Expressing, Plasmid Preparation, Comparison

Efficacy against M. tuberculosis aerosol challenge of priming mice with BCG and boosting them with recombinant attenuated L. monocytogenes expressing 5 M. tuberculosis antigens. (a) Experimental schedule. BALB/c mice (8/group) were immunized i.d. with PBS (sham) or 5 × 10 5 CFU of BCG at week 0. Mice immunized i.d. with BCG were either not boosted or boosted once at week 18 or twice at weeks 14 and 18 (group G2 only) with 2 × 10 6 CFU of rLm5Ag vaccine candidates. At week 22, the mice were challenged with aerosolized M. tuberculosis Erdman strain (average of 19 CFU delivered to the lungs of each animal, as assayed at day 1 postchallenge to mice), and at week 32, 10 weeks postchallenge, mice were euthanized. (b) Organ bacterial burden. Lungs (top) and spleens (bottom) of mice in the vaccinated and challenge groups (described in the table to the right of the graphs) were removed and assayed for bacillus burdens. Each symbol represents one mouse, and the means ± SEM are shown as bars. Group designations are listed beneath the horizontal axis. The pink-colored boxes indicate group B, BCG-primed only. The log 10 CFU in the lungs and spleens were compared to group A (sham) and group B (BCG i.d.) by one-way ANOVA with Fisher’s LSD test. The significant P values to group B (BCG i.d.) in the lungs and spleens are shown in the graph; **, P < 0.01; ***, P < 0.001; and ****, P < 0.0001. The significant P values between sham group (A) and all other groups are listed in the table to the right.

Journal: mBio

doi: 10.1128/mbio.00687-22

Figure Lengend Snippet: Efficacy against M. tuberculosis aerosol challenge of priming mice with BCG and boosting them with recombinant attenuated L. monocytogenes expressing 5 M. tuberculosis antigens. (a) Experimental schedule. BALB/c mice (8/group) were immunized i.d. with PBS (sham) or 5 × 10 5 CFU of BCG at week 0. Mice immunized i.d. with BCG were either not boosted or boosted once at week 18 or twice at weeks 14 and 18 (group G2 only) with 2 × 10 6 CFU of rLm5Ag vaccine candidates. At week 22, the mice were challenged with aerosolized M. tuberculosis Erdman strain (average of 19 CFU delivered to the lungs of each animal, as assayed at day 1 postchallenge to mice), and at week 32, 10 weeks postchallenge, mice were euthanized. (b) Organ bacterial burden. Lungs (top) and spleens (bottom) of mice in the vaccinated and challenge groups (described in the table to the right of the graphs) were removed and assayed for bacillus burdens. Each symbol represents one mouse, and the means ± SEM are shown as bars. Group designations are listed beneath the horizontal axis. The pink-colored boxes indicate group B, BCG-primed only. The log 10 CFU in the lungs and spleens were compared to group A (sham) and group B (BCG i.d.) by one-way ANOVA with Fisher’s LSD test. The significant P values to group B (BCG i.d.) in the lungs and spleens are shown in the graph; **, P < 0.01; ***, P < 0.001; and ****, P < 0.0001. The significant P values between sham group (A) and all other groups are listed in the table to the right.

Techniques: Aerosol, Recombinant, Expressing

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