How do gut bacterial residents fight whooping cough after taking antibiotics ?

Author: Ziying yuan & Nicolas de Azevedo

Introduction

Bordetella pertussis is a Gram-negative, aerobic, pathogenic bacterium, meaning it possesses a complex cell envelope of the plasma membrane, peptidoglycan cell wall and an outer membrane. It survives and grows in an oxygenated environment, capable of infecting the respiratory system and causing a disease called “whooping cough” in humans. B. pertussis is notorious for severe and uncontrollable coughing that can make it hard to breath or even deadly for babies. It is spread by airborne droplets and infects humans by colonizing the lung cells, then toxin is produced by the bacterium to prevent human cilia from clearing debris from the lungs. The lungs have to cough very hard to expel the debris and bacteria out of the body into the air, which can then infect other people. B. pertussis can affect people of all ages; however, infants, young children, pregnant women and the unvaccinated population remain the most susceptible to pertussis-related morbidity and mortality. Babies at the highest risk of serious disease, are more likely to need hospitalization or even die from whooping cough; about one in every 200 babies under 6 months old who catch whooping cough dies from pneumonia or brain damage.

Vaccine for whooping cough has been widely used since the end of the 20th century. Early administration of antibiotics and corticosteroids treatment immediately after diagnosis can limit the course of the disease, and the therapy is most effective during the incubation period. The side effects of antibiotics varied from case to case.

Findings

In this study, the researchers intervened to change the microbiota in mice using antibiotics treatment, and then measured their resistance against B. pertussis or whooping cough by infecting them. They found out that an intact and balanced microbiota inhibits the colonization of B. pertussis in the lungs, while antibiotics-treated mice showed higher susceptibility to bacterial infection.

In the beginning, the mice were given a slew of four antibiotics for three weeks so that their microbiota could be manipulated as the exposed group. Three days after the antibiotic treatment was done, a sufficient amount of B. pertussis was introduced into the exposed and unexposed groups of mice to mimic the natural bacterial infection in the nose. Then, the exposed group and the unexposed group were compared in terms of their development of symptoms and change in biochemical markers in the body for whooping cough.

Three days after the antibiotics treatment and before B. pertussis being introduced, the lung samples and feces from the antibiotics-exposed mice were compared to those from the non-exposed mice. They found out that the relative abundance of ten main bacteria living in the mice’s feces had drastically changed due to previous antibiotics administration. Noticeably, the antibiotics-exposed mice showed an 87.65% increase in Proteobacteria. Other bacterial populations were affected to different extents, such as the absence of Saccharibacteria. The lungs, however, showed no signs of obvious changes in their bacterial composition, leading to the scientists believing that the oral delivery of antibiotics would not affect the lung microbiota. This inconsistency of population change between the gut and lung tissue could potentially be due to the small sample size used in this experiment. The conclusion drawn from a small sample size is not statistically robust regarding the cause-effect relationship, and the general pattern could be easily overlooked. In other words, the generalizability of the findings in this study to the target population can be a challenge. Moreover, the ten types of gut bacterial residents studied were based on relative abundance so percentage. Without an accurate population count for the microbiota in lungs after antibiotics administration, the conclusion lacks internal validity and reliability. 

 The presence of these bacteria in the feces and lungs is indicative of the gut and the lung’s microbiota (also known as the diverse microorganisms that reside within the aforementioned sections of the body). When the bacteria within the microbiota, also called commensals, were killed, a lack of diversity was found in the fecal matter and thus the gut microbiota. This became a case of dysbiosis, which can be defined as “a compositional and functional alteration in the microbiota that is driven by a set of environmental and host-related factors that perturb the microbial ecosystem to an extent that exceeds its resistance and resilience capabilities”. Multiple studies had shown that the antibiotic treatments could have profound and rapid changes in gut microbiota, some of which could last for years and dysbiosis can be a stable state depending on individuals.  

Figure 1 is an illustration explaining the concept of dysbiosis. The colored bacteria are commensals and the black one is the pathogen. Source: Nicolas de Azevedo, 2021

The next experiment the researchers conducted was to determine if an intact microbiota plays a role in the resistance against whooping cough infection.

Figure 2 is a graph showing the mean of the log₁₀ number of CFU per lung of each mouse across multiple time points post infection of B. pertussis in antibiotic-treated mice (Ab-BP) and non-antibiotic treated mice (BP). CFU means Colony Forming Unit, which is a unit used to describe the concentration of a microorganism or in this case, the concentration of B. pertussis per lung. Source: Adapted from Commensal Microbes Affect Host Humoral Immunity to Bordetella pertussis Infection, by Y. Zhang, 2019, American society for microbiology.

The researchers took samples from the lung tissues to determine the amount of B. pertussis colonizing the Ab-BP mice and BP mice as time went on. The main trend for both series of data points is an increase in B. pertussis from 3hrs to D7 after infection, followed by a gradual clearance of the bacteria from the lungs until D17. However, the Ab-BP demonstrated more serious bacterial colonization than the BP had until D17, notably at 3hrs and D3. This means that previous antibiotic exposure could have a role in lessening resistance to whooping cough in mice, and by extension potentially be caused by the lack of commensal microbes and gut dysbiosis.

The researchers wanted to see if antibiotics could affect the production of antibodies in mice: ten days after B. pertussis infection, the number of antibodies in the mice’s serum (liquid component of blood without clotting factors) was compared between antibiotic treated mice, non-antibiotic treated mice, and control non-infected mice. They observed that in general, a larger quantity of antibodies was collected in the non-antibiotic treated mice, meaning stronger immune response against bacterial infection.

They repeated the same experiment seventeen days after the infection, only to find that the non-antibiotic treated mice have at least more than twice the antibodies compared to their antibiotic treated counterparts. Taken together, these experiments allude to the idea that antibiotics can cause dysbiosis, increase susceptibility to B. pertussis and lower the number of antibodies produced yet the exact mechanisms behind this event are unknown.

Importance of the findings

In this era of increasing demand for antibiotics treatment, the fact that the oral antibiotics can impair the gut health and cause profound and long-term disruption in the composition of gut bacterial population remains a thorny issue because it can lead to increased susceptibility to B. pertussis infection and perturb normal immune response. This research had underscored the importance of the gut microbiome in human immune responses to respiratory infection, which means that by having a better and healthier gut bacterial population, people may have more resistance to whooping cough. Future research should be done to explore novel strategies to recover or optimize the microbiota for robust immune defense. Isolation of specific gut bacteria that are protective against B. pertussis pathogenesis is of great importance to counteract the side effect of oral antibiotics treatment.

Noticeably, cutting down the use of antibiotics is not a simple thing to do in current society. People are becoming more dependent on this fast and simple way of treating a variety of infections. The consequences are a disrupted microbiota and a vulnerable immune system to fight disease. To prevent this, the selection of antibiotics that are less likely to have a long-term effect on the gut microbiota is important. Figuring out the exact cellular and molecular mechanism by which the human microbiota manipulates the immune response to B. pertussis infection is the future focus of the investigation. Hills et al. proposed that the healthy gut flora exert their beneficial effects through the fermentation of dietary fiber to produce short-chain fatty acids (SCFA), so eating a diverse diet rich in whole foods may help build a balanced microbiome.  SCFA promotes immunity and suppress inflammatory process in the intestine and other organs. By consuming dietary fibre which is not digested but fermented by commensal bacteria in the small intestine, SCFA will be released to support the growth of beneficial bacteria in the gut and help fight possible invaders.

Reference

Australian government department of health. (2020, May 27). Whooping cough (pertussis). Retrieved from https://www.health.gov.au/health-topics/whooping-cough-pertussis.

K. L. Sealey, T. Belcher & A. Preston (2016, June 1). Bordetella pertussis epidemiology and evolution in the light of pertussis resurgence. Infection, Genetics and Evolution: journal of molecular epidemiology and evolutionary genetics in infectious disease. 40, pp. 136-143.

N. Shin, T. Wong & J. Bae. (2015, July 22). Proteobacteria: microbial signature of dysbiosis in gut microbiota. Trends in biotechnology, 33(9), pp496-503

S Black (1997, April 16). Epidemiology of pertussis. The Pediatric infectious disease journal, pp. 85-89.

Silvana Bettiol, K. W. (2012). Symptomatic treatment of the cough in whooping cough. Retrieved from https://doi.org/10.1002/14651858.CD003257.pub4.

Y. Zhang, Z. Ran, M. Tian et al (2019, December 19). Commensal Microbes Affect Host Humoral Immunity to Bordetella pertussis Infection. Infection and Immunity, pp. e00421-19. Retrieved from https://journals.asm.org/doi/10.1128/iai.00421-19?permanently=true.

M. Levy, A. Kolodziejczyk, C. Thaiss & E. Elinac. (2017). Dysbiosis and the immune system. Nature reviews. Immunology, 17(4), pp219-232.

L. Dethlefsen & D. A. Relman. (2011). Incomplete recovery and individualized responses of the human distal gut microbiota to repeated antibiotic perturbation. Proceedings of the National Academy of Science of the United States of America, 108 suppl1, pp4554-4561.

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