Bacterial outer membrane vesicles, a potential vaccine candidate in interactions with host cells based

OMVs secretion is a normal phenomenon during the growth of bacteria. OMVs are spherical, bilayered, membranous nanostructures (shown in Fig. 1). OMVs productions are not consistent in size, varying from approximately 20 nm to 250 nm. The parent bacterial membrane releases them and hence they contain numerous proteins, similar to parent bacteria [1‐4]. Not only that, OMVs are demonstrated in abundance in bacterial components such as DNA [5], RNA [6, 7], lipopolysaccharide (LPS) [8], enzymes [9, 10], peptidoglycan [11] and some molecules [10]. Here, we enumerate some examples about virulence factors carried by OMVs (listed in Table 1). These characteristics of OMVs endow with critical significance in pathogenesis and communication between bacteria and host cells. Recent studies have revealed that OMVs play fundamental roles in activating immune system and facilitate manifest responses against OMVs in the host [12]. Despite of the increasing findings about effects of OMVs on host, innovative strategies based on OMVs have shown great potential in clinical implications and other relevant fields as we will discuss in this review.

OMVs are not described useless as originally thought. Electron microscopy [2‐4, 10] and proteomic analyses [1‐4] have confirmed OMVs are heterogeneous nanostructures packaging various bioactive components. They function as mediators to transmit biological information among different bacteria and host. For example, bacterial OMVs serve as nanovesicles to deliver important biological substances, which can promote the entry of bacterial antigens or even genes. Colitogenic Bacteroides thetaiotaomicron OMVs assist bacterial antigens access host cells in a sulfatase-dependent manner [13]. Bacterial OMVs shoot short RNAs to boost the host-pathogen interaction [6, 7]. In addition, OMVs can act as a delivery system for virulence factors [14] or antibiotic resistence genes [15]. For instance, Bacteroides thetaiotaomicron OMVs carry cephalosporinases to protect gut pathogens against β-lactam antibiotics [16]. Consequently, considering OMVs as effective vehicles to enhance the interactions between host and pathogen, the biosynthesis of OMVs deserve prominent attention to make us better understanding of OMVs as we will discuss below.

Interestingly, although both Gram-positive and Gram-negative bacteria have the capacity of producing OMVs [17], the amounts of OMVs production and the components carried by OMVs are different with each other, even the same bacteria in different environment [18, 19]. Also, heterogeneous sized OMVs contain distinct protein profiles [20].Usually, it is considered that OMVs production as responses to stress around bacteria to make bacteria survive and adapt within host [18]. OMVs released from Vibrio cholerae appear heterogenous in size due to different culture environment and purification methods that affect chemical composition of OMVs as well [21]. Ciprofloxacin-stimulated cultures produced more and larger vesicles which were enriched with cytosolic proteins compared with non-induced condition [22]. Not just ciprofloxacin, antibiotic such as meropenem, fosfomycin, and polymyxin B can also increase production of OMVs [23]. Furthermore, amounts of OMVs produced by cultured clean water bacteria increased upon treatmentwith ultraviolet radiation [24]. Pseudomonas putida secrete OMVs in response to stress caused by cationic surfactants [25]. There is a hypothesis that bacteria produce OMVs as defense mechanisms against external threats, including antibiotics [26], antimicrobial peptides [27], and bacteriophage infection [28], which may enlighten us of the important role of OMVs in bacterial pathopoiesis. There are evidences indicating that Porphyromonas gingivalis (P. gingivalis) OMVs contribute to local immune evasion of P. gingivalis by hindering the host response [29].

With the exception of the findings discussed above, many factors may be responsible for OMVs biosynthesis according to recent study: (1) Loss of some antigens of bacteria may contribute to proteins transformation in OMVs [30] and virulence factor may regulate OMVs biogenesis [31]. (2) Sandro et al., reported that the VacJ/Yrb ABC (ATP-binding cassette) transport system is involved in OMV formation among Gram-negative bacteria [32]. Pseudomonas Quinolone Signal (PQS) is found to modulate OMVs production in Pseudomonas aeruginosa [33]. On the basis of the discovery PQS, Alexander et al., demonstrated reciprocal cross-species can induce OMVs biogenesis via secreted factors in Gamma proteobacteria but not included Alphaproteobacteria [34]. (3) The formation of Vibrio vulnificus OMVs is associated with expression of the capsular polysaccharide [35]. Enterohemorrhagic Escherichia coli OmpT have an impact on the biogenesis, composition, and size of OMVs [36]. In addition, pathogenic or non-pathogenic bacterial OMVs may show great discrepancy in biological activity. Recent study reveals that toxigenic Bacteroides fragilis (B. fragilis) OMVs represent different metabolic activities compared with nonpathogenic B. fragilis OMVs [37]. In addition, emerging evidences indicate that close relationships between bacterial components and OMVs may modulate production of OMVs.Wael et al. declared that LPS remodeling leads to formation of OMVs in Salmonella [38]. Contrarily, Salmonella OMVs can also accelerate LPS remodeling during environmental transitions [39]. Haruyuki et al., found if DNA inversion occur in B. fragilis, it will regulate the formation of OMVs [40], and small RNAs may modulate the composition of outer membrane proteins, which may be critical to OMVs synthesis [41]. In conclusion, these innovative findings have expanded our knowledge of how OMVs generate from parent bacteria. We can speculate from so many evidences about OMVs production that whether OMVs serve as a mirror reflecting the surrounding environment around bacteria and the real time condition of bacteria or not. If it is right, we can evaluate the level of bacterial condition through quantitative analysis of OMVs, which may be a promising idea concerning monitoring the growth of bacteria involved in related fields. However, the mechanisms of OMVs formation have not been explored clearly, no unanimous conclusion can be made in this field. It deserves more attention and more research to find the truth concealed behind the phenomenon.

Despite the mystery on the mechanisms involved in OMVs formation, the research on the functions of bacterial OMVs especially with host cells keep growing rapidly. Next, we will focus on the correlation between bacterial OMVs and host cells, mainly including the OMVs entry mechanisms of host cells together with the effects of OMVs on host cells.

OMVs possess several inherent characteristics that make them consider as a potential vaccine candidate. (1) OMVs remain intact and steady at different temperatures and treatments [18, 21, 105]. Considering the properties, OMVs are designed as storage containers to protect enzymatic function [106]. (2) they have inanimate activity, non-replicative property and adjuvant effect [105], but they contain substantial immunogenic components [1] associated with parent bacteria, which may elicit potent innate and adaptive immune responses against bacterial infection [73‐79]. Shiga toxin-producing Escherichia coli (STEC) OMVs can be treated as vaccines to protect mice and claves against Hemolytic Uremic Syndrome (HUS) [106]. OMVs derived from Bordetella bronchiseptica can survive mice from sublethal infection [107]. Salmonella enteritidis OMVs provide strong protective efficiency against S. enteritidis infection [108]. In addition, OMVs are multipurpose so that they can be engineered as vehicles to carry any antigens or anything else. Yoshihiro Ojima et al., packaged green fluorescence protein (GFP) into the OMVs of Escherichia coli and succeed to express OmpW-GFP recombinant protein in OMVs [109]. Engineer OMVs deliver surface-associated glycotopes to the immune system and then induce protective antibody [110]. Neisseria meningitidis OMVs modified by genetic technique show stronger immune response than native OMVs [111]. Moreover, they can be modified by special technology to reduce their toxicity and become safe to host. To our knowledge, OMV-based meningococcal vaccine is the only implication of OMVs for clinical trial to control a clonal B outbreak so far [112, 113]. However, because vaccines derived from OMVs are infancy and strategies are immature (shown in Table 2). Vaccines made of OMVs require more time and efforts to be licensed for human use even if amounts of researches have proved their efficiency in animal model (shown in Table 3).

Bacterial OMVs were discovered almost half a century ago, scientists have never desisted from exploring the mechanisms and effects about OMVs. According to the recent study, despite of obscure mechanisms regarding to OMVs formation, we have a better understanding of the mechanisms of OMVs entry, which may enlighten us of how OMVs communicate with host cells. However, a mode of OMVs entry cannot be adopted by all bacteria and we wonder if there is a common way that OMVs can take. Plenty of evidences elucidate the significant effects of OMVs on host cells. We can summarize that OMVs can drive host cells to produce more pro-inflammatory and anti-inflammatory cytokines in response to OMVs challenge. Meanwhile, innate and adaptive immune response can be imitated by exposure to OMVs. In contrast, OMVs have the capacity of inhibiting immune cells responses [69, 85] which may contribute to bacteria survival and spread. It seems that OMVs can play dual role in modulating immune system response against infection (shown in Fig. 3). Together with the responses caused by OMVs, OMVs may play an important role in bacterial pathogenesis and the interactions between pathogen and host cells. There is one report regarding OMVs in indoor dust elucidating the association between asthma with OMVs, which may remind us that OMVs may have a broader effect on inflammatory [114].

Furthermore, OMV-based vaccines show great potential in the development of immunity to diseases caused by bacteria. Innovative technologies make bioengineered OMVs possible to serve as specific vaccines targeted on limited pathogen [115]. Enlightened from the technologies, scientists utilize modified OMVs just as a vehicle to carry drugs or anything else if they want to evaluate the effects on other diseases even cancer, which broad the therapeutic implications and expand the field of OMVs applications, not just restricted to bacterial infection. For instance, bacterial OMVs are engineered as multi-functional nanodevices for biosensing and bioimaging of cancer cells [116]. OMVs decorated with tumor-specific epitopes can induce strong inhibition of tumor growth [117]. Collectively, advances are made to expand our knowledge of exploiting OMVs-associated techniques to benefit the health of human worldwide.