Tom Defoirdt

Alternatives to antibiotics for the control of bacterial disease in aquaculture

The wide and frequent use of antibiotics in aquaculture has resulted in the development and spread of antibiotic resistance. Because of the health risks associated with the use of antibiotics in animal production, there is a growing awareness that antibiotics should be used with more care. This is reflected in the recent implementation of more strict regulations on the prophylactic use of antibiotics and the presence of antibiotic residues in aquaculture products. For a sustainable further development of the aquaculture industry, novel strategies to control bacterial infections are needed. This review evaluates several alternative biocontrol measures that have emerged recently. Most of these methods are still in research phase; few have been tested in real aquaculture settings. It is important to further develop different strategies that could be combined or used in rotation in order to maximise the chance of successfully protecting the animals and to prevent resistance development.

Cinnamaldehyde and cinnamaldehyde derivatives reduce virulence in Vibrio spp. by decreasing the DNA-binding activity of the quorum sensing response regulator LuxR

BackgroundTo date, only few compounds targeting the AI-2 based quorum sensing (QS) system are known. In the present study, we screened cinnamaldehyde and substituted cinnamaldehydes for their ability to interfere with AI-2 based QS. The mechanism of QS inhibition was elucidated by measuring the effect on bioluminescence in several Vibrio harveyi mutants. We also studied in vitro the ability of these compounds to interfere with biofilm formation, stress response and virulence of Vibrio spp. The compounds were also evaluated in an in vivo assay measuring the reduction of Vibrio harveyi virulence towards Artemia shrimp.ResultsOur results indicate that cinnamaldehyde and several substituted derivatives interfere with AI-2 based QS without inhibiting bacterial growth. The active compounds neither interfered with the bioluminescence system as such, nor with the production of AI-2. Study of the effect in various mutants suggested that the target protein is LuxR. Mobility shift assays revealed a decreased DNA-binding ability of LuxR. The compounds were further shown to (i) inhibit biofilm formation in several Vibrio spp., (ii) result in a reduced ability to survive starvation and antibiotic treatment, (iii) reduce pigment and protease production in Vibrio anguillarum and (iv) protect gnotobiotic Artemia shrimp against virulent Vibrio harveyi BB120.ConclusionCinnamaldehyde and cinnamaldehyde derivatives interfere with AI-2 based QS in various Vibrio spp. by decreasing the DNA-binding ability of LuxR. The use of these compounds resulted in several marked phenotypic changes, including reduced virulence and increased susceptibility to stress. Since inhibitors of AI-2 based quorum sensing are rare, and considering the role of AI-2 in several processes these compounds may be useful leads towards antipathogenic drugs.

Quorum Sensing-Disrupting Brominated Furanones Protect the Gnotobiotic Brine Shrimp Artemia franciscana from Pathogenic Vibrio harveyi, Vibrio campbellii, and Vibrio parahaemolyticus Isolates

ABSTRACT Autoinducer 2 (AI-2) quorum sensing was shown before to regulate the virulence of Vibrio harveyi towards the brine shrimp Artemia franciscana. In this study, several different pathogenic V. harveyi, Vibrio campbellii, and Vibrio parahaemolyticus isolates were shown to produce AI-2. Furthermore, disruption of AI-2 quorum sensing by a natural and a synthetic brominated furanone protected gnotobiotic Artemia from the pathogenic isolates in in vivo challenge tests.

Can Bacteria Evolve Resistance to Quorum Sensing Disruption

Traditional treatment of bacterial infections relies heavily on the use of antibacterial compounds that either kill bacteria (bactericidal) or inhibit their growth (bacteriostatic). Typically, the targets for the main conventional antibiotics are essential cellular processes such as bacterial cell wall biosynthesis, bacterial protein synthesis, and bacterial DNA replication and repair. However, resistance to these drugs arises and spreads very rapidly, even to such an extent that bacteria have been identified that are simultaneously resistant to all available antibiotics [1]. The increasing occurrence of resistant bacteria gradually renders antibiotics ineffective in treating infections and has enormous human and economic consequences worldwide. As a result, the identification of novel drug targets and the development of novel therapeutics constitute an important area of current scientific research. An alternative to killing or inhibiting growth of pathogenic bacteria is the specific attenuation of bacterial virulence, which can be attained by targeting key regulatory systems that mediate the expression of virulence factors. One of the target regulatory systems is quorum sensing (QS), or bacterial cell-to-cell communication. QS is a mechanism of gene regulation in which bacteria coordinate the expression of certain genes in response to the presence or absence of small signal molecules (Figure 1). Figure 1 General scheme of a quorum sensing system. Quorum Sensing: Bacterial Cell-to-Cell Communication QS was first discovered in the marine bacterium Vibrio fischeri and was thought to be restricted to only a limited series of species. Later on, similar systems were found to be present in many other Gram-negative bacteria. These Gram-negative bacteria use acylated homoserine lactones (AHLs) as signal molecules (for a review see [2]). AHLs are typically produced by a homolog of V. fischeri LuxI and detected by a homolog of V. fischeri LuxR. In addition to the AHL-mediated systems in Gram-negative bacteria, some Gram-positive bacteria also regulate a variety of processes by QS. The QS systems of Streptococcus pneumoniae, Bacillus subtilis, and Staphylococcus aureus, for instance, have been extensively studied (for a review see [3]). A different kind of QS system is found in vibrios. These bacteria use multichannel QS systems in which different types of signal molecules are produced. The signal molecules are detected at the cell surface by membrane-bound, two-component receptor proteins that feed a common phosphorylation/dephosphorylation signal transduction cascade (for a review on QS in vibrios, see [4]). One of the signals produced by vibrios is the so-called autoinducer 2 (AI-2), a furanosyl borate diester [5]. AI-2 activity has been detected in many different species (Gram-negative as well as Gram-positive), although its function as a signal is not generally accepted for all species (for a detailed discussion see [6]). The language of bacteria seems to be even more diversified as new QS systems, using different types of signal molecules, are still being discovered [7].

Quorum sensing and quorum quenching in Vibrio harveyi: lessons learned from in vivo work

Luminescent vibrios, bacteria belonging to the species Vibrio harveyi and closely related species, are important pathogens in aquaculture that can affect almost all types of cultured animals. Due to large-scale use of antibiotics, many luminescent vibrios have acquired (multiple) resistance, which render antibiotic treatments ineffective. One of the alternative strategies that has recently been developed to control infections caused by antibiotic-resistant bacteria is the disruption of quorum sensing, bacterial cell-to-cell communication. The quorum sensing system of V. harveyi has been studied quite intensively in vitro. Recent studies have been directed towards understanding the impact of quorum sensing and quorum sensing disruption on the virulence of luminescent vibrios towards different host organisms in vivo. This mini-review aims at discussing the current knowledge of quorum sensing in luminescent vibrios in vivo. Subsequently, quorum quenching by halogenated furanones is discussed and finally, some directions for further research are presented.

Short-chain fatty acids and poly-β-hydroxyalkanoates: (New) Biocontrol agents for a sustainable animal production.

Because of the risk of antibiotic resistance development, there is a growing awareness that antibiotics should be used more carefully in animal production. However, a decreased use of antibiotics could result in a higher frequency of pathogenic bacteria, which in its turn could lead to a higher incidence of infections. Short-chain fatty acids (SCFAs) have long been known to exhibit bacteriostatic activity. These compounds also specifically downregulate virulence factor expression and positively influence the gastrointestinal health of the host. As a consequence, there is currently considerable interest in SCFAs as biocontrol agents in animal production. Polyhydroxyalkanoates (PHAs) are polymers of beta-hydroxy short-chain fatty acids. Currently, PHAs are applied as replacements for synthetic polymers. These biopolymers can be depolymerised by many different microorganisms that produce extracellular PHA depolymerases. Interestingly, different studies provided some evidence that PHAs can also be degraded upon passage through the gastrointestinal tract of animals and consequently, adding these compounds to the feed might result in biocontrol effects similar to those described for SCFAs.

Quorum sensing inhibitors: how strong is the evidence?

Because of its promising effect as an alternative to antibiotics, quorum sensing disruption is an intensively studied field, and there are many studies that describe the quorum sensing inhibitory activity of natural and synthetic compounds. In this opinion article, we present an overview of recent literature with respect to quorum sensing inhibitors. Most of this research is based on experiments with quorum sensing signal molecule reporter strains. However, these experiments are prone to bias due to other effects compounds may have on reporter strains. We argue that researchers should perform adequate control experiments and should carefully assess toxicity of the compounds in the bacterial species they are working with in order to confirm that what they observe really is quorum sensing inhibition.

Early mortality syndrome outbreaks: a microbial management issue in shrimp farming?

A recent disease of farmed Penaeid shrimp, commonly referred to as “early mortality syndrome” (EMS) or more technically known as “acute hepatopancreatic necrosis disease” (AHPND), was first reported in southern China in 2010 and subsequently in Vietnam, Thailand, and Malaysia [1]. The EMS/AHPND disease typically affects shrimp postlarvae within 20–30 days after stocking and frequently causes up to 100% mortality. The Global Aquaculture Alliance [2] has estimated that losses to the Asian shrimp culture sector amount to USD 1 billion. The causative agent of EMS/AHPND has been reported to be a bacterium—more specifically a pathogenic Vibrio belonging to the Harveyi clade, presumably Vibrio parahaemolyticus [3]. So far, this has been the only description of a bacterial isolate capable of causing EMS/AHPND. Strategies to remedy this disease are urgently needed. However, as long as it remains unclear whether or not all incidences of EMS/AHPND are caused by one or more specific V. parahaemolyticus strains, approaches that focus on controlling the presence or activity of vibrios in general have the highest chance of decreasing the risk of EMS/AHPND outbreaks.

Disruption of Bacterial Cell-to-Cell Communication by Marine Organisms and its Relevance to Aquaculture

Bacterial disease is one of the most critical problems in commercial aquaculture. Although various methods and treatments have been developed to curb the problem, yet they still have significant drawbacks. A novel and environmental-friendly approach in solving this problem is through the disruption of bacterial communication or quorum sensing (QS). In this communication scheme, bacteria regulate their own gene expression by producing, releasing, and sensing chemical signals from the environment. There seems to be a link between QS and diseases through the regulation of certain phenotypes and the induction of virulence factors responsible for pathogen–host association. Several findings have reported that numerous aquatic organisms such as micro-algae, macro-algae, invertebrates, or even other bacteria have the potential to disrupt QS. The mechanism of action varies from degradation of signals through enzymatic or chemical inactivation to antagonistic as well as agonistic activities. This review focuses on the existing marine organisms that are able to interfere with QS with potential application for aquaculture as bacterial control.

The application of bioflocs technology to protect brine shrimp (Artemia franciscana) from pathogenic Vibrio harveyi

Aims:  To study the potential biocontrol activity of bioflocs technology.