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Examining the safety of respiratory and intravenous inoculation of Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus in a mouse model.

Shatzkes K, Chae R, Tang C, Ramirez GC, Mukherjee S, Tsenova L, Connell ND, Kadouri DE - Sci Rep (2015)

Bottom Line: We found no reduction in mouse viability after intranasal or intravenous inoculation of B. bacteriovorus 109J, HD100 or M. aeruginosavorus.Intravenous injection caused an increase of IL-6 in the kidney and spleen, TNF in the liver and CXCL-1/KC in the blood at 3 hours post-exposure, returning to baseline levels by 18 hours.Furthermore, qPCR detected predators were cleared from the host quickly and efficiently.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Disease, Department of Medicine, Rutgers New Jersey Medical School, Newark, NJ 07103, USA.

ABSTRACT
Bdellovibrio spp. and Micavibrio spp. are Gram-negative predators that feed on other Gram-negative bacteria, making predatory bacteria potential alternatives to antibiotics for treating multi-drug resistant infections. While the ability of predatory bacteria to control bacterial infections in vitro is well documented, the in vivo effect of predators on a living host has yet to be extensively examined. In this study, respiratory and intravenous inoculations were used to determine the effects of predatory bacteria in mice. We found no reduction in mouse viability after intranasal or intravenous inoculation of B. bacteriovorus 109J, HD100 or M. aeruginosavorus. Introducing predators into the respiratory tract of mice provoked a modest inflammatory response at 1 hour post-exposure, but was not sustained at 24 hours, as measured by RT-qPCR and ELISA. Intravenous injection caused an increase of IL-6 in the kidney and spleen, TNF in the liver and CXCL-1/KC in the blood at 3 hours post-exposure, returning to baseline levels by 18 hours. Histological analysis of tissues showed no pathological changes due to predatory bacteria. Furthermore, qPCR detected predators were cleared from the host quickly and efficiently. This work addresses some of the safety concerns regarding the potential use of predatory bacteria as a live antibiotic.

No MeSH data available.


Related in: MedlinePlus

Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria.qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of cytokines was assessed in the lung at (A) 1, (B) 24, and (C) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at (D) 24 and (E) 48 hours post-inoculation. Fold induction was calculated using the ΔΔCt by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t-test was performed to compare each treated sample to their respective control sample, *P < 0.05; **P < 0.01.
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f2: Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria.qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of cytokines was assessed in the lung at (A) 1, (B) 24, and (C) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at (D) 24 and (E) 48 hours post-inoculation. Fold induction was calculated using the ΔΔCt by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t-test was performed to compare each treated sample to their respective control sample, *P < 0.05; **P < 0.01.

Mentions: As before, none of the total 90 mice that were inoculated with predatory bacteria exhibited any visual adverse effects and all were healthy at the time of sacrifice. At one hour post-inoculation, we observed an increase of IL-1β (9.0- and 12.3-fold), IL-23 (6.3- and 12.6-fold) and TNF (5.0- and 7.9-fold) in the lungs of mice exposed to B. bacteriovorus 109J or M. aeruginosavorus, respectively (Fig. 2A). However, this increased expression was not sustained at 24 or 48 hours post-inoculation (Fig. 2B,C). Conversely, none of the mice exposed to B. bacteriovorus HD100 exhibited a substantial (5-fold or higher) increase in expression of any inflammatory marker gene in the lung or spleen at any time point (Fig. 2). Furthermore, no inflammatory response was detected in the spleens of mice inoculated with either B. bacteriovorus 109J or M. aeruginosavorus at 24 or 48 hours (Fig. 2D,E). Inflammatory protein levels in the lungs of inoculated mice were measured with ELISA to confirm the results obtained from qPCR. We did observe a 4.7-fold increase in CXCL-1/KC protein expression due to B. bacteriovorus 109J at 24 hours. However, no other inflammatory protein showed a substantial fold induction (5-fold or higher) due to inoculation with any of the predatory bacteria (Fig. 3). Additionally, mice examined at five days post-inoculation still exhibited no substantial increases in proinflammatory marker gene expression (data not shown). As before, all mice inoculated with the predators were healthy with no visual adverse effects at any of the examined time points.


Examining the safety of respiratory and intravenous inoculation of Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus in a mouse model.

Shatzkes K, Chae R, Tang C, Ramirez GC, Mukherjee S, Tsenova L, Connell ND, Kadouri DE - Sci Rep (2015)

Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria.qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of cytokines was assessed in the lung at (A) 1, (B) 24, and (C) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at (D) 24 and (E) 48 hours post-inoculation. Fold induction was calculated using the ΔΔCt by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t-test was performed to compare each treated sample to their respective control sample, *P < 0.05; **P < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4528228&req=5

f2: Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria.qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of cytokines was assessed in the lung at (A) 1, (B) 24, and (C) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at (D) 24 and (E) 48 hours post-inoculation. Fold induction was calculated using the ΔΔCt by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t-test was performed to compare each treated sample to their respective control sample, *P < 0.05; **P < 0.01.
Mentions: As before, none of the total 90 mice that were inoculated with predatory bacteria exhibited any visual adverse effects and all were healthy at the time of sacrifice. At one hour post-inoculation, we observed an increase of IL-1β (9.0- and 12.3-fold), IL-23 (6.3- and 12.6-fold) and TNF (5.0- and 7.9-fold) in the lungs of mice exposed to B. bacteriovorus 109J or M. aeruginosavorus, respectively (Fig. 2A). However, this increased expression was not sustained at 24 or 48 hours post-inoculation (Fig. 2B,C). Conversely, none of the mice exposed to B. bacteriovorus HD100 exhibited a substantial (5-fold or higher) increase in expression of any inflammatory marker gene in the lung or spleen at any time point (Fig. 2). Furthermore, no inflammatory response was detected in the spleens of mice inoculated with either B. bacteriovorus 109J or M. aeruginosavorus at 24 or 48 hours (Fig. 2D,E). Inflammatory protein levels in the lungs of inoculated mice were measured with ELISA to confirm the results obtained from qPCR. We did observe a 4.7-fold increase in CXCL-1/KC protein expression due to B. bacteriovorus 109J at 24 hours. However, no other inflammatory protein showed a substantial fold induction (5-fold or higher) due to inoculation with any of the predatory bacteria (Fig. 3). Additionally, mice examined at five days post-inoculation still exhibited no substantial increases in proinflammatory marker gene expression (data not shown). As before, all mice inoculated with the predators were healthy with no visual adverse effects at any of the examined time points.

Bottom Line: We found no reduction in mouse viability after intranasal or intravenous inoculation of B. bacteriovorus 109J, HD100 or M. aeruginosavorus.Intravenous injection caused an increase of IL-6 in the kidney and spleen, TNF in the liver and CXCL-1/KC in the blood at 3 hours post-exposure, returning to baseline levels by 18 hours.Furthermore, qPCR detected predators were cleared from the host quickly and efficiently.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Disease, Department of Medicine, Rutgers New Jersey Medical School, Newark, NJ 07103, USA.

ABSTRACT
Bdellovibrio spp. and Micavibrio spp. are Gram-negative predators that feed on other Gram-negative bacteria, making predatory bacteria potential alternatives to antibiotics for treating multi-drug resistant infections. While the ability of predatory bacteria to control bacterial infections in vitro is well documented, the in vivo effect of predators on a living host has yet to be extensively examined. In this study, respiratory and intravenous inoculations were used to determine the effects of predatory bacteria in mice. We found no reduction in mouse viability after intranasal or intravenous inoculation of B. bacteriovorus 109J, HD100 or M. aeruginosavorus. Introducing predators into the respiratory tract of mice provoked a modest inflammatory response at 1 hour post-exposure, but was not sustained at 24 hours, as measured by RT-qPCR and ELISA. Intravenous injection caused an increase of IL-6 in the kidney and spleen, TNF in the liver and CXCL-1/KC in the blood at 3 hours post-exposure, returning to baseline levels by 18 hours. Histological analysis of tissues showed no pathological changes due to predatory bacteria. Furthermore, qPCR detected predators were cleared from the host quickly and efficiently. This work addresses some of the safety concerns regarding the potential use of predatory bacteria as a live antibiotic.

No MeSH data available.


Related in: MedlinePlus