Pulmonary Aspergillus fumigatus infection in rats affects gastrointestinal homeostasis

Abstract

Microbiota inhabiting mucosal tissues is involved in maintenance of their immune homeostasis. Growing body of evidence indicate that dysbiosis in gut influence immune responses at distal sites including lungs. There are also reports concerning gut involvement with pulmonary injury/inflammation in settings of respiratory viral and bacterial infections. The impact of infections with other microorganisms on gut homeostasis is not explored. In this study, the rat model of sublethal pulmonary infection with Aspergillus fumigatus was used to investigate the effect of fungal respiratory infection on gut immune-mediated homeostasis. Signs of intestinal damage, intestinal and gut-draining lymphoid tissue cytokine responses and gut bacterial microbiota diversity were examined. Intestinal injury, inflammatory cell infiltration, as well as increased levels of intestinal interferon-γ (IFN-γ) and interleukin-17 (IL-17) (as opposed to unchanged levels of anti-inflammatory cytokine IL-10) during the two-week period depict intestinal inflammation in rats with pulmonary A. fumigatus infection. It could not be ascribed to the fungus as it was not detected in the intestine of infected rats. Increased production of pro-inflammatory cytokines by major gut-draining mesenteric lymph nodes point to these lymphoid organs as places of generation of cytokine-producing cells. No changes in spleen or systemic cytokine responses was observed, showing lack of the effects of pulmonary A. fumigatus infection outside mucosal immune system. Drop of intestinal bacterial microbiota diversity (disappearance of several bacterial bands) was noted early in infection with normalization starting from day seven. From day three, appearance of new bacterial bands (unique to infected individuals, not present in controls) was seen, and some of them are pathogens. Alterations in intestinal bacterial community might have affected intestinal immune tolerance contributing to inflammation. Disruption of gut homeostasis during pulmonary infection might render gastrointestinal tract more susceptible to variety of physiological and pathological stimuli. Data which showed for the first time gut involvement with pulmonary infection with A. fumigatus provide the baseline for future studies of the impact of fungal lung infections to gut homeostasis, particularly in individuals susceptible to these infections.

Introduction

At late seventies the term “common mucosal immunological system” (CMIS) was coined to describe similarities in lymphoid tissues associated with bronchus and intestine (McDermott and Bienenstock, 1979). From then mechanisms underlying immune responses to pathogens in the gastrointestinal and respiratory tract have been established (Mowat and Agace, 2014; Sato and Kiyono, 2012). Growing body of evidence suggests that two mucosal sites communicate and interact in health and disease what brought suggestion that mucosal immune system is system wide-organ (Gill et al., 2010). Pulmonary involvement seen in patients with chronic gastrointestinal diseases such as inflammatory bowel disease and irritable bowel syndrome as well as association of gastrointestinal symptoms observed in patients with pulmonary infections depict cross-talk between gastrointestinal and pulmonary defences (Samuelson et al., 2015; Tulic et al., 2016).

Much interest in investigation of gut and lung communication was given to gut microbiota (Dickson et al., 2013). The use of animals treated with antibiotics or which differ in intestinal microbiota composition showed that gut microbiota is essential for promoting optimal pulmonary response to allergens (Kim et al., 2014), against viruses (Abt et al., 2012; Ichinohe et al., 2011; Wu et al., 2013) and bacteria (Chen et al., 2011; Fagundes et al., 2012; Gauguet et al., 2015; Schuijt et al., 2016).

Changes in gut microbiota were also found to be involved in settings of pulmonary immune responses. Infection of mice with Influenza A (IA) (Deriu et al., 2016; Groves et al., 2018; Wang et al., 2014; Yildiz et al., 2018; Zhang et al., 2015) or respiratory syncytial (RSV) viruses (Groves et al., 2018) resulted in altered diversity of gut bacterial microbiota and was associated with intestinal injury and inflammation. Presence of the virus in the intestine was responsible for tissue injury only in one study (Zhang et al., 2015). Intestinal injury is thought to be induced by virus-specific T cells recruited from the lung (Wang et al., 2014), though innate pulmonary responses might be responsible judging by data which showed that administration of agents which induce pulmonary inflammation such are poly I:C (Deriu et al., 2016) and LPS (Sze et al., 2014) also affected gut microbiota. Alterations of gut microbiota were detected in patients (Luo et al., 2017) and mice with pulmonary M. tuberculosis infection (Winglee et al., 2014).

Gut microbiota has the potential to influence host response to fungi as well. Treatment of mice with antibiotics and oral C. albicans thereafter, prime mice for allergic T-cell based response to Aspergillus (Noverr et al., 2004). More recently, the influence of intestinal bacterial microbiota on the generation of pulmonary IL-17 response to A. fumigatus was noted in mice (McAleer et al., 2016). There are, however no data concerning the effects of pulmonary fungal infections on gut homeostasis.

In the view of above cited studies which showed the impact of pulmonary viral and bacterial infections on gut homeostasis, we hypothesized that pulmonary immune response to fungi might affect gut homeostasis as well. Among variety of rat models of aspergillosis (Chandenier et al., 2009; Desoubeaux and Cray, 2017) we used rat model of pulmonary Aspergillus fumigatus infection in immunocompetent rats (El-Muzghi et al., 2013; Mirkov et al., 2015) to investigate the effect of A. fumigatus infection on gut homeostasis. Signs of intestinal damage, intestinal and gut-associated lymphoid tissue cytokine responses as well as gut bacterial microbiota diversity were examined at different time points following infection, shown previously by us to be accompanied with marked changes in fungal load resulting from inflammatory/immune activity in the lungs (El-Muzghi et al., 2013; Mirkov et al., 2015). Data that were obtained showed for the first time that lung infection with A. fumigatus is associated with intestinal damage and pro-inflammatory cytokine response in small intestine and major-gut draining (mesenteric) lymph nodes. Decrease in diversity of intestinal bacterial microbiota was observed during pulmonary infection, with appearance of new bacterial species, not noted in non-infected controls, what might have contributed to intestinal inflammation. Disruption of gut homeostasis during pulmonary infection might render gastrointestinal tract more susceptible to variety of physiological and pathological stimuli.