Mosquito Egg Raft Distribution Is Affected by Semiochemicals: Indication of Interspecific Competition.

Numerous species of animals alter their behavior in response to increasing competition. To do so, they must possess the ability to detect the presence and density of interspecific competitors. We studied the role of semiochemicals released by increasing densities of larval Culiseta longiareolata Macquart on female oviposition habitat selection in two field experiments. Similarly to C. longiareolata larvae, subordinate Culex laticinctus Edwards are periphyton grazers who dwell in rain-filled pools in the Mediterranean region. We show that C. laticinctus females oviposited significantly less in mesocosm pools that were treated with crowding signals originating from C. longiareolata larvae. In the second experiment, we placed a similar number of larvae directly inside the 50 L mesocosms. These low-density mesocosms did not affect C. laticinctus oviposition but were attractive to conspecific oviposition. These results increase our understanding of the female ability to detect species-specific signals, indicating increased larval competition.

Fish microbiota repel ovipositing mosquitoes.

The mere presence of predators causes prey organisms to display predation-avoidance strategies. Predator presence is often communicated through predator-released chemical signals. Ovipositing female mosquitoes of several species are repelled by unknown signals released from larvivorous fish. It was previously suggested that in many cases, a predator's microbiota plays an important role in the release of these signals; however, this mechanism is still poorly understood. In this study, we looked into the effects of the microbiota originating from the larvivorous Gambusia affinis (Baird and Girard) on the oviposition behaviour of gravid female mosquitoes. We used fish with altered microbiota and bacterial isolates in a set of outdoor mesocosm experiments to address this aim. We show that interference with the fish microbiota significantly reduces fish's repellent effect. We further show that the bacterium Pantoea pleuroti, isolated from the skin of the fish, repels oviposition of Culex laticinctus Edwards and Culiseta longiareolata Macquart mosquitoes similarly to the way in which live fish repel them. Our results highlight the importance of bacteria in the interspecies interactions of their hosts. Furthermore, this finding may lead to the development of an ecologically friendly mosquito repellent, that may reduce the use of larvivorous fish for mosquito control.

Quantification of bacterial adhesion to tissue in high-throughput kinetics.

Bacterial adhesion to tissue is the starting point for many pathogenic processes and beneficial interactions. The dynamics and speed of adhesion (minutes) make high-resolution temporal kinetic data important, but this capability is absent from the current toolset. We present a high-throughput method with a second-to-minute kinetic resolution, testing the adhesion of Pseudomonas aeruginosa PAO1 wild-type, flagella-, pili-, and quorum-sensing mutants to human embryonic kidney (HEK293) cells. Adhesion rates were in good correlation with HEK293 confluence, and the ways in which various bacterial mutations modified adhesion patterns are in agreement with the published literature. This simple assay can facilitate drug screening and treatment development as well as provide a better understanding of the interactions of pathogenic and probiotic bacteria with tissues, allowing the design of interventions and prevention treatments.

Effect of Drinking Water Salt Content on the Interaction between Surfactants and Bacteria.

Sodium dodecyl sulfate (SDS) is a common surfactant used in various hygienic products. Its interactions with bacteria were studied previously, but the three-way interaction between surfactants, bacteria, and dissolved salts in the context of bacterial adhesion has not been studied. Here, we examined the combined effects of SDS (at concentrations typical of everyday hygienic activities) and salts, sodium chloride, and calcium chloride (at concentrations typically found in tap water) on the adhesion behavior of the common opportunistic pathogen Pseudomonas aeruginosa. We found that bacterial adhesion in the absence of SDS was dependent on the cation concentration rather than the total ionic strength and that combined treatment with several millimolar NaCl and SDS can increase bacterial adhesion. The addition of low concentrations of SDS (2 mM) to tens to hundreds millimolar concentrations of NaCl, typical of systems that suffer seawater incursion, reduced bacterial adhesion dramatically. Combined treatment with Ca+2 (in concentrations typical of those found in hard water) and SDS produced a small increase in total adhesion but a dramatic increase in the strength of adhesion. We conclude that the type and concentration of salts in water can have a considerable effect on the efficacy of soap in reducing bacterial adhesion and should be taken under consideration in critical applications. IMPORTANCE Surface-adhering bacteria are a reoccurring problem in many settings, including households, municipal water systems, food production facilities, and hospitals. Surfactants, and specifically sodium dodecyl sulfate (also known as SDS/SLS), are commonly used to remove bacterial contamination, but data regarding the interaction of SDS with bacteria and especially the effects of water-dissolved salts on this interaction are lacking. Here, we show that calcium and sodium ions can dramatically affect the efficacy of SDS on bacterial adhesion behavior and conclude that salt concentrations and ion species in the water supply should be considered in SDS applications.

Bacterial detoxification of plant defence secondary metabolites mediates the interaction between a shrub and frugivorous birds.

Many plants produce fleshy fruits, attracting fruit-eating animals that disperse the seeds in their droppings. Such seed dispersal results in a conflict between the plant and the animal, as digestion of seeds can be highly beneficial to the animal but reduces plant fitness. The plant Ochradenus baccatus uses the myrosinase-glucosinolates system to protect its seeds. We show that hydrolysis of the O. baccatus fruit glucosinolates by the myrosinase enzyme inhibited digestive enzymes and hampered digestion in naïve individuals of the bird Pycnonotus xanthopygos. However, digestion in birds regularly feeding on O. baccatus fruits was unaffected. We find that Pantoea bacteria, dominating the gut of these experienced birds as well as the fruits, thrive on glucosinolates hydrolysis products in culture. Augmentation of Pantoea protects both naïve birds and plant seedlings from the effects of glucosinolates hydrolysis products. Our findings demonstrate a tripartite interaction, where the plant-bird mutually beneficial interactions are mediated by a communal bacterial tenant.

Pseudomonas aeruginosa Mobbing-Like Behavior against Acanthamoeba castellanii Bacterivore and Its Rapid Control by Quorum Sensing and Environmental Cues.

Mobbing, group attack of prey on predator, is a behavior seen in many animal species in which prey animals use numbers and coordination to counter individually superior predators. We studied attack behavior of Pseudomonas aeruginosa toward the bacterivore Acanthamoeba castellanii. This behavior consists of directed motility toward and specific adhesion to the predator cells, enacted in seconds and responding to both prey and predator population densities. Attack coordination relies on remote sensing of the predator and the use of the Pseudomonas quinolone signal (PQS), a P. aeruginosa species-specific quorum sensing molecule. Mutants unable to produce the PQS show unspecific adhesion and reduced survival, and a corresponding increase in predator population occurs as a result of predation. The addition of an external PQS restored some predator-specific adherence within seconds, suggesting a novel response mechanism to this quorum sensing (QS) signal. Fast behavioral response of P. aeruginosa to PQS is also supported by the rate of signal accumulation in the culture, reaching relevant concentrations within minutes, enabling bacteria response to self population density in these short timescales. These results portray a well-regulated group attack of the bacteria against their predator, reacting within seconds to environmental cues and species-specific signaling, which is analogous in many ways to animal mobbing behavior. IMPORTANCE Pseudomonas aeruginosa was shown previously to attack amoebae and other predators by adhering to them and injecting them with virulent substances. In this work, we show that an active, coordinated group behavior is enacted by the bacteria to utilize these molecular components, responding to both predator and bacterial population density. In addition to their ecological significance, immediate behavioral changes observed in response to PQS suggest the existence of a fast QS signal cascade, which is different from canonical QS that relies on slow-to-respond gene regulation. Similar regulatory circuits may drive other bacterial adaptations and pathogenicity mechanisms and may have important clinical implications.

Bacterial Adhesion Kinetics in a High Throughput Setting in Seconds-minutes Time Resolution.

Bacterial surface adhesion, the first step in many important processes including biofilm formation and tissue invasion, is a fast process that occurs on a time scale of seconds. Adhesion patterns tend to be stochastic and spatially heterogeneous, especially when bacteria are present in low population densities and at early stages of adhesion to the surface. Thus, in order to observe this process, a high degree of temporal resolution is needed across a large surface area in a way that allows several replicates to be monitored. Some of the current methods used to measure bacterial adhesion include microscopy, staining-based microtiter assays, spectroscopy, and PCR. Each of these methods has advantages in assaying aspects of bacterial surface adhesion, but none can capture all features of the process. In the protocol presented here, adapted from Shteindel et al., 2019 , fluorescently-labeled bacteria are monitored in a multi-titer setting using a standard plate fluorimeter and a dye that absorbs light in the fluorophore excitation and emission wavelengths. The advantage of using this dye is that it restricts the depth of the optic layer to the few microns adjacent to the bottom of the microtiter well, eliminating fluorescence originating from unattached bacteria. Another advantage of this method is that this setting does not require any preparatory steps, which enables reading of the sample to be repeated or continuous. The use of a standard multi-titer well allows easy manipulation and provides flexibility in experimental design.