Alpha and beta-diversity

Alpha-diversity is an indicator of microbial species diversity, which considers the variety of species (richness) and the evenness of species abundance [48]. Variations in this parameter have been observed in several diseases [49]. Most of the studies in mice included in this review assessed alpha-diversity, but the results were heterogeneous. In six-week-old MRL/lpr mice [14], a lower alpha-diversity was found, while in MRL+/+ mice, the decrease was not significant [14]. On the other hand, in NZB/NZWF1 mice, the diversity of the microbiota increased after SLE onset, approximately at 20 weeks of age, as well as in NZB/W F1 mice induced by HCMVpp65_422–439, a viral peptide of human cytomegalovirus [17], suggesting that the disease progression influences the microbiota diversity [42].

Differences in the microbial community composition (beta-diversity) were also found by several authors [14, 16, 18–20, 42]. One study only identified changes in the beta-diversity of 16-week-old SNF1 mice housed littermates but not in 4-week-old mice [18], showing, once again, that disease progression affects the gut microbiota.

F/B ratio

The F/B ratio has been considered a marker of gut homeostasis. Thus a higher or lower ratio has been considered a dysbiosis marker associated with several diseases [50]. Nevertheless, it should be considered together with other markers of gut microbiota diversity and composition [49]. The heterogeneous results between studies highlight the variation in the gut microbiota of lupus mouse models. An increased F/B ratio was found by some authors [17], but others found an opposite trend, with a decreased ratio in 6-week-old MRL/lpr mice [14] or a trend toward a decreased ratio in 30-week-old segmented filamentous bacteria (SFB) positive mice without statistical significance [22].

Within the phyla Firmicutes and Bacteroidetes, several bacteria may contribute to an altered F/B ratio. In 6-week-old MRL/lpr mice, the lower F/B ratio was attributed to a decreased abundance in Peptostreptococcaceae and Lactobacillaceae (phylum Firmicutes) and increased abundance in Rikenellaceae (phylum Bacteroidetes) [14]. However, there was no correlation with advanced lupus activity, suggesting that a lower ratio may contribute to early disease onset [14].

Gut bacterial composition

The gut microbiota dysbiosis may exacerbate SLE manifestations rather than trigger the disease [19, 24, 25], and the translocation of pathobionts is one of the possible mechanisms involved [22, 37, 51]. In SNF1 mice, the depletion of the gut microbiota with an antibiotic resulted in decreased expression of pro-inflammatory cytokines while not decreasing immune regulatory ones, such as interleukin 10 (IL-10) [18]. Moreover, after antibiotic use, only 40% of the female mice developed severe nephritis, while, without antibiotics, 80% of the mice developed this manifestation [18]. Interestingly, the microbiota depletion in male mice did not alter the disease incidence, immune phenotype, disease stage, or renal involvement [18]. Moreover, the gut microbiota composition in male and female littermates was significantly different only at adult ages [18]. Furthermore, castrated male SNF1 mice had different microbial communities at the genus level than non-castrated mice and showed higher lupus incidence and earlier onset (although not statistically significant) [18]. This work highlights that the gut microbiota in SLE may have a different influence in females and in males, which is reinforced by the results obtained with fecal transfer from male to female SNF1 mice, showing a slower lupus progression, lower proteinuria, and a modest decrease in anti-dsDNA and anti-nucleohistone antibodies than female mice that did not receive the fecal transplant [18].

Other authors also identified different variations in the microbial communities of female and male mice. Increased levels of Lachnospiraceae [16], Bacteroidetes, and Parabacteroides, and decreased Dysgonomonas [18] were found in females, while higher levels of Bifidobacterium were found in males [16].

Besides gender bias, the disease’s duration and characteristics may also contribute to the identified variability in the gut microbiota. In female and castrated male lpr mice, fecal transplant with Lactobacillus spp., particularly an uncultured Lactobacillus spp. and L. reuteri, starting before the disease onset, improved renal function and survival [21]. These alterations were associated with the increased expression of tight-junction proteins, meaning an improved gut barrier function and a decreased endotoxemia induced by Lactobacillus spp. treatment [21]. The improvement of the gut barrier integrity, and consequent decrease in bacterial translocation and activation of immune responses, was corroborated by the reduction in T cell migration to the gut lamina propria [21]. Moreover, Lactobacillus spp. also increased the systemic production of IL-10, meaning that these bacteria may have a systemic anti-inflammatory effect [21]. Furthermore, treatment with Lactobacillus spp. increased the influx of CD8⁺ T cells and forkhead box P3 (Foxp3)⁺ T reg cells to the kidney while decreasing T helper 17 (Th17) cells, attenuating LN, and improving the Treg-Th17 balance in castrated mice (that showed a similar response to female mice), but not in non-castrated ones. These results suggest that androgens attenuate Lactobacillus spp. effects, and in this mouse model, the control of LN induced by the gut microbiota was sex hormone-dependent [21].

In another study of two strains of lupus mouse models with different disease characteristics, distinct bacterial communities were identified [14]. Six-week-old MRL/lpr mice had a lower abundance of Muribaculaceae and Anaeroplasmataceae than 6-week-old MRL+/+ mice, while in 18-week-old MRL+/+ mice, there was an enrichment in Akkermansiaceae [14]. The authors suggested that this change may contribute to immune-mediated responses preceding the disease progression, favoring altered gut permeability and mucosal immune activation [14]. Lactobacillaceae showed a decreasing trend between 6-week-old and 18-week-old MRL/lpr mice, meaning that the inflammatory responses of the gut mucosa may have originated in a prolonged decrease in Lactobacillaceae [14]. Other authors found a similar reduction in Lactobacillaceae in the same mouse model [16]. Akkermansiaceae was significantly increased in 18-week-old MRL+/+ mice (more aggressive disease) compared with 6-week-old, but lower in MRL/lpr mice, suggesting that these bacteria can have different roles in different lupus phenotypes [14]. Other authors have also found that Akkermansiaceae and R. were negatively correlated with the serum levels of complement component 3 (C3) [33].

The bacterial composition of the gut microbiota varies between mouse studies. In 30-week-old NZM2410 mice inoculated with SFB, higher Prevotellaceae and Rikenellaceae and reduced Lachnospiraceae were present in severe glomerulonephritis [22]. In NZB/W F1 mice, Odoribacter, Desulfovibrio, and Roseburia genera were positively correlated with lupus-like effects, and the creatinine level, anti-dsDNA IgG titer, and glomerulonephritis were positively correlated with Candidatus Saccharimonas. These four genera are linked to the regulation of the host immune system, and immune-mediated diseases [17], demonstrating that the gut microbiota alterations are associated with lupus-like symptoms in this mouse model. Moreover, the authors also identified increased levels of the proinflammatory cytokines IL-6 and IL-17A related to the observed alterations in the gut microbiota [17].

In another study Bacteroidetes, Succinivibrio, Bilophila, and Parabateroides were positively correlated with IL-17, tumor necrosis factor (TNF)-related weak inducer of apoptosis (TWEAK), IL-2 receptor (IL-2R), IL-21, IL-35, IL-10, and interferon (IFN)-γ, while Dialister and Gemmiger were negatively associated with IL-17, IL-2R, and IL-3 [26], meaning that interfering with the production of these inflammatory cytokines, may be one of the mechanisms by which the gut microbiota influences SLE pathogenesis [37].

Finally, the influence of genetic variations on the composition and modulation of the gut microbiota is a new focus of research [52]. Several polymorphisms have been associated with the diversity of the gut microbiota, and some with specific bacteria, such as the well-described LCT gene association with Bifidobacterium [53]. Recently, in a transgenic mouse model, defective T cell receptor (TCR) signaling altered the gut microbiota and promoted systemic autoimmunity by driving Th17 cell differentiation [25]. Once again, this study showed the connection between genome, microbiota, and autoimmunity.

The connection between gut microbiota and lupus phenotype was further reinforced by the observation that a colony of MRL/lpr mice with a milder disease phenotype had an altered gut microbiota profile [23].

Further alterations in the gut microbiota in lupus mouse models can be found in Table S1.

Alpha and beta-diversity

Similarly to the results in mouse studies, in humans the alpha-diversity alterations were also heterogeneous, varying between lower alpha-diversity [20, 26, 32, 33, 35, 37, 39, 41, 42], increased alpha-diversity [30] or no alterations in this marker compared with HC [16, 19, 27, 31, 34]. No differences were found between the newly onset patients and other patients, nor between patients with and without LN [35]. The same authors did not find differences in alpha-diversity between patients with high and low SLEDAI scores [35]. However, other authors found a trend toward an inverse correlation between alpha-diversity and higher disease activity (SLEDAI ≥ 8) [37]. When assessing differences between active and in remission patients, one study did not find global differences but detected an increase in Bifidobacterium [32]. Some authors identified that, despite decreasing disease activity, SLE treatment further decreased the gut bacterial diversity [41].

The beta-diversity was altered between SLE patients and controls [16, 27, 30, 32, 35, 37–39, 41], but not in all studies [31]. Although no association was found with medication [32], some authors found that after PPI use, there was an approximation between the microbiota profile of SLE patients with HC [39]. Beta-diversity also varied between patients with low and high disease activity [37, 38], although not consensually [35], and before and after disease onset [22, 42].

F/B ratio

Five studies found a decrease in the F/B ratio in SLE patients compared to HC [26–28, 31, 38], while others only found a decreasing trend [22, 32], two did not find differences in this variable [33, 42] and one detected an increase [27]. An inverse correlation between disease activity and F/B ratio was found in some SLE patients [28]. Paradoxically, in active disease an increased F/B ratio was also reported [17, 26].

Gut bacterial composition

Several authors identified gut microbiota changes in SLE [14, 17, 32, 38, 39], but the bacteria involved varied between studies. One explanation may be the disease duration since variations in bacterial composition have been observed with disease progression [16, 42] and severity. Some authors found differences in the gut bacterial composition of patients with higher and lower disease activity [28, 33, 37], but not all [26], and some found a correlation between certain bacteria and disease activity, such as Streptococcus anginosus and Veillonella [32, 37], Acholeplasma, Capnocytophaga and Leptotrichia [33]. Some bacteria were associated with SLE risk (Bacilli, Eggerthella, and Lactobacillales), while others were found to be protective (Coprobacter, Bacillales, and Lachnospira) [29]. Interestingly, Lactobacillales were increased in active SLE patients but not in patients in remission [32].

R. gnavus has been recognized as a possible pathobiont contributing to renal involvement in SLE patients, as a 5-fold increase in this species, and a decrease in bacteria with beneficial roles, such as B. uniformis, were identified in this population [41]. Azzouz and collaborators [37], in 2019, mentioned that SLE patients showed decreased bacterial diversity, which was inversely correlated with disease activity and altered dynamics between species. When assessing the bacterial abundance, R. gnavus, a Gram-positive bacteria, was significantly increased and correlated with lupus disease activity [41]. Moreover, when LN was present, the increase in R. gnavus was even higher [41]. Other authors reported similar findings, encountering an increased abundance of R. gnavus in SLE patients with high disease activity [37]. SLE patients, especially with LN, showed increased gut permeability, leading to increased IgG anti-rat glioma model (RG) 2 strain antibodies, which correlated with disease activity [41]. R. gnavus seems, therefore, to contribute to renal damage [41]. These results were not replicated in other studies. Strikingly, some SLE patients with active disease showed a decreased abundance of R. gnavus compared to patients in remission [32]. Differences between populations related to genetics and diet and other lifestyle factors should be considered to explain these differences. Further studies assessing the variability of R. gnavus will help clarify these observations.

An association between gut microbiota composition and immune factors seems to exist [31]. Certain strains of B. fragilis, which are increased in some SLE patients [41], produce a toxin that binds to a colonic epithelial cell receptor and alters gut permeability [56]. However, in murine models, B. fragilis have beneficial effects, preventing colitis [56]. The immunomodulatory characteristics of this bacterial species are due to polysaccharide A (PSA) that can induce IL-10 secretion, by Treg cells, reducing gut inflammation [57]. It can also have a protective effect against pathogenic infections and systemic immune-mediated diseases [58]. Despite these different results, the gut microbiota and the host immune system certainly crosstalk, shaping the chronic activation of the immune system.

Akkermansia muciniphila showed immunogenic and inflammatory potential in a group of SLE patients with increased levels of this bacterium in the gut. In these patients, a bacterial peptide, which mimics an extracellular part of the human FAS, can bind IgG and positively correlates with serologic markers of inflammation [41]. The genus Lactobacillus was lower in SLE patients than in HC [28], although Lactobacillus abundance has not been steady through the different studies [32, 33], which may be due to the different species included [28].

Phylum Bacteroidetes [26, 38] and Proteobacteria [30], and genus B. and Alistipes were found to be increased in SLE patients [38], although there are inconsistent results between studies [33]. Bacteroidetes species are glycan-degrading and short-chain fatty acid-producing bacteria, usually considered beneficial for the host. However, simultaneously B. theta has been considered a potential gut pathobiont, enhancing lupus-like disease [38], and Proteobacteria, a marker of dysbiosis [31].

Despite the variations between the bacterial communities of SLE patients and lupus-mouse models, some shared alterations were also found. R. torques and Blautia genera were increased, and the Desulfovibrio genera were decreased in SLE patients and MRL/lpr mice. However, the role of the mucin degrader R. torques in SLE warrants a better understanding [41].

Streptococcus anginosus and Streptococcus intermedius, two species belonging to the oral and gut microbiota, were increased in SLE metagenome [35], reinforcing the concept of an oral-gut interaction. Nevertheless, the gut bacterial composition is not always consistent with the oral bacterial composition [33].

Although some studies included both male and female participants, the differences in the gut microbiota composition between genders were not assessed, possibly due to the low number of male participants, considering that SLE is mainly a female disease. Further changes in the gut microbiota can be found in Table S1.