Michael J. Sweet

Ciliate and bacterial communities associated with White Syndrome and Brown Band Disease in reef-building corals

White Syndrome (WS) and Brown Band Disease (BrB) are important causes of reef coral mortality for which causal agents have not been definitively identified. Here we use culture-independent molecular techniques (DGGE and clone libraries) to characterize ciliate and bacterial communities in these diseases. Bacterial (16S rRNA gene) and ciliate (18S rRNA gene) communities were highly similar between the two diseases. Four bacterial and nine ciliate ribotypes were observed in both diseases, but absent in non-diseased specimens. Only one of the bacteria, Arcobacter sp. (JF831360) increased substantially in relative 16S rRNA gene abundance and was consistently represented in all diseased samples. Four of the eleven ciliate morphotypes detected contained coral algal symbionts, indicative of the ingestion of coral tissues. In both WS and BrB, there were two ciliate morphotypes consistently represented in all disease lesion samples. Morph1 (JN626268) was observed to burrow into and underneath the coral tissues at the lesion boundary. Morph2 (JN626269), previously identified in BrB, appears to play a secondary, less invasive role in pathogenesis, but has a higher population density in BrB, giving rise to the visible brown band. The strong similarity in bacterial and ciliate community composition of these diseases suggests that they are actually the same syndrome.

Evidence of Melanoma in Wild Marine Fish Populations

The increase in reports of novel diseases in a wide range of ecosystems, both terrestrial and marine, has been linked to many factors including exposure to novel pathogens and changes in the global climate. Prevalence of skin cancer in particular has been found to be increasing in humans, but has not been reported in wild fish before. Here we report extensive melanosis and melanoma (skin cancer) in wild populations of an iconic, commercially-important marine fish, the coral trout Plectropomus leopardus. The syndrome reported here has strong similarities to previous studies associated with UV induced melanomas in the well-established laboratory fish model Xiphophorus. Relatively high prevalence rates of this syndrome (15%) were recorded at two offshore sites in the Great Barrier Reef Marine Park (GBRMP). In the absence of microbial pathogens and given the strong similarities to the UV-induced melanomas, we conclude that the likely cause was environmental exposure to UV radiation. Further studies are needed to establish the large scale distribution of the syndrome and confirm that the lesions reported here are the same as the melanoma in Xiphophorus, by assessing mutation of the EGFR gene, Xmrk. Furthermore, research on the potential links of this syndrome to increases in UV radiation from stratospheric ozone depletion needs to be completed.

Algae as reservoirs for coral pathogens.

Benthic algae are associated with coral death in the form of stress and disease. Its been proposed that they release exudates, which facilitate invasion of potentially pathogenic microbes at the coral-algal interface, resulting in coral disease. However, the original source of these pathogens remains unknown. This study examined the ability of benthic algae to act as reservoirs of coral pathogens by characterizing surface associated microbes associated with major Caribbean and Indo-Pacific algal species/types and by comparing them to potential pathogens of two dominant coral diseases: White Syndrome (WS) in the Indo-Pacific and Yellow Band Disease (YBD) in the Caribbean. Coral and algal sampling was conducted simultaneously at the same sites to avoid spatial effects. Potential pathogens were defined as those absent or rare in healthy corals, increasing in abundance in healthy tissues adjacent to a disease lesion, and dominant in disease lesions. Potentially pathogenic bacteria were detected in both WS and YBD and were also present within the majority of algal species/types (54 and 100% for WS and YBD respectively). Pathogenic ciliates were associated only with WS and not YBD lesions and these were also present in 36% of the Indo-Pacific algal species. Although potential pathogens were associated with many algal species, their presence was inconsistent among replicate algal samples and detection rates were relatively low, suggestive of low density and occurrence. At the community level, coral-associated microbes irrespective of the health of their host differed from algal-associated microbes, supporting that algae and corals have distinctive microbial communities associated with their tissue. We conclude that benthic algae are common reservoirs for a variety of different potential coral pathogens. However, algal-associated microbes alone are unlikely to cause coral death. Initial damage or stress to the coral via other competitive mechanisms is most likely a prerequisite to potential transmission of these pathogens.

Review: metal-based nanoparticles; size, function, and areas for advancement in applied microbiology.

Nanoparticles (NPs) are attracting increased attention in commerce and applied microbiology due to their antimicrobial activity, high electrical conductivity, and optical properties. For example, silver NPs have broad spectrum antimicrobial properties against a wide range of bacteria and fungi, making them ideal for minimizing biofouling. By controlling the size, shape, surface, and agglomeration state of the NPs, specific ion release profiles can be developed for any given application. Currently, NPs are formed in a wide variety of different shapes and sizes including spheres, plates, and wires. This review looks at both commercially and naturally produced NPs with a focus on silver NPs and addresses how these are formed. Furthermore, potential areas for improving these techniques will be highlighted, focusing on advancing shape and structure formation using modern applications. Finally, the review evaluates the feasibility of bioengineering microorganisms to synthesize particles of defined shape and size, by examining genes associated with NP production.

Bacterial assemblages shifts from healthy to yellow band disease states in the dominant reef coral Montastraea faveolata.

Descriptions of microbial diversity in healthy and diseased corals are necessary first steps before further investigating the mechanisms that lead to coral pathology. This is the first study that characterizes the microbial associates from healthy corals to yellow band disease (YBD) lesions using two complementary screening techniques of bacterial 16S rRNA genes [amplified 16S ribosomal DNA restriction analysis (ARDRA) of clone libraries and denaturing gradient gel electrophoresis (DGGE)]. Both these techniques showed similar trends, namely a significant difference in the bacterial community and an increase in diversity from healthy to YBD diseased lesions. There was an increase in the number of sequences retrieved of potentially pathogenic bacteria in diseased tissues compared with healthy samples, most notably from the genus Vibrio. Furthermore, we also detected a number of known pathogenic bacteria within the natural healthy microbiota such as Vibrio carchariae and Vibrio harveyi, a result supporting previous studies, showing healthy corals have the ability to harbour these species.

Experimental antibiotic treatment identifies potential pathogens of white band disease in the endangered Caribbean coral Acropora cervicornis.

Coral diseases have been increasingly reported over the past few decades and are a major contributor to coral decline worldwide. The Caribbean, in particular, has been noted as a hotspot for coral disease, and the aptly named white syndromes have caused the decline of the dominant reef building corals throughout their range. White band disease (WBD) has been implicated in the dramatic loss of Acropora cervicornis and Acropora palmata since the 1970s, resulting in both species being listed as critically endangered on the International Union for Conservation of Nature Red list. The causal agent of WBD remains unknown, although recent studies based on challenge experiments with filtrate from infected hosts concluded that the disease is probably caused by bacteria. Here, we report an experiment using four different antibiotic treatments, targeting different members of the disease-associated microbial community. Two antibiotics, ampicillin and paromomycin, arrested the disease completely, and by comparing with community shifts brought about by treatments that did not arrest the disease, we have identified the likely candidate causal agent or agents of WBD. Our interpretation of the experimental treatments is that one or a combination of up to three specific bacterial types, detected consistently in diseased corals but not detectable in healthy corals, are likely causal agents of WBD. In addition, a histophagous ciliate (Philaster lucinda) identical to that found consistently in association with white syndrome in Indo-Pacific acroporas was also consistently detected in all WBD samples and absent in healthy coral. Treatment with metronidazole reduced it to below detection limits, but did not arrest the disease. However, the microscopic disease signs changed, suggesting a secondary role in disease causation for this ciliate. In future studies to identify a causal agent of WBD via tests of Henle–Kochs postulates, it will be vital to experimentally control for populations of the other potential pathogens identified in this study.

Coral diseases in aquaria and in nature

Many reef coral diseases have been described affecting corals in the wild, several of which have been associated with causal agents based on experimental inoculation and testing of Kochs postulates. In the aquarium industry, many coral diseases and pathologies are known from the grey literature but as yet these have not been systematically described and the relationship to known diseases in the wild is difficult to determine. There is therefore scope to aid the maintenance and husbandry of corals in aquaria by informing the field of the scientifically described wild diseases, if these can be reliably related. Conversely, since the main driver to identifying coral diseases in aquaria is to select an effective treatment, the lessons learnt by aquarists on which treatments work with particular syndromes provides invaluable evidence for determining the causal agents. Such treatments are not commonly sought by scientists working in the natural environment due the cost and potential environmental impacts of the treatments. Here we review both wild and aquarium diseases and attempt to relate the two. Many important aquarium diseases could not be reconciled to those in the wild. In one case, however, namely that of the ciliate Helicostoma sp. as a causal agent of brown jelly syndrome in aquarium corals, there may be similarities with pathogenic agents of the wild coral diseases, such as white syndrome and brown band syndrome. We propose that Helicostoma is actually a misnomer, but improved understanding of this pathogen and others could benefit both fields. Improved practices in aquarium maintenance and husbandry would also benefit natural environments by reducing the scale of wild harvest and improving the potential for coral culture, both for the aquarium industry and for rehabilitation programmes.

Silver nanoparticles: a microbial perspective.

Silver nanoparticles (NPs) are used for a wide range of commercial reasons to restrict microbial growth. The increasing use of silver NPs in modern materials ensures they will find their way into environmental systems. The mode of action which makes them desirable as an antimicrobial tool could also pose a severe threat to the natural microbial balance existing in these systems. Research into the potential environmental threats of silver NPs has mainly focused on particular areas, such as their influence in rivers and estuaries or their effect on organisms such as earthworms and plants. There is a need to focus studies on all aspects of the microbial world and to highlight potential risks and methods of overcoming problems before significant damage is done. This review focuses on the antimicrobial uses, mechanisms of toxicity, and effects on the environment (mainly soil) of silver NPs, illustrating gaps in current knowledge.

Development of Bacterial Biofilms on Artificial Corals in Comparison to Surface-Associated Microbes of Hard Corals

Numerous studies have demonstrated the differences in bacterial communities associated with corals versus those in their surrounding environment. However, these environmental samples often represent vastly different microbial micro-environments with few studies having looked at the settlement and growth of bacteria on surfaces similar to corals. As a result, it is difficult to determine which bacteria are associated specifically with coral tissue surfaces. In this study, early stages of passive settlement from the water column to artificial coral surfaces (formation of a biofilm) were assessed. Changes in bacterial diversity (16S rRNA gene), were studied on artificially created resin nubbins that were modelled from the skeleton of the reef building coral Acropora muricata. These models were dip-coated in sterile agar, mounted in situ on the reef and followed over time to monitor bacterial community succession. The bacterial community forming the biofilms remained significantly different (R = 0.864 p<0.05) from that of the water column and from the surface mucus layer (SML) of the coral at all times from 30 min to 96 h. The water column was dominated by members of the α-proteobacteria, the developed community on the biofilms dominated by γ-proteobacteria, whereas that within the SML was composed of a more diverse array of groups. Bacterial communities present within the SML do not appear to arise from passive settlement from the water column, but instead appear to have become established through a selection process. This selection process was shown to be dependent on some aspects of the physico-chemical structure of the settlement surface, since agar-coated slides showed distinct communities to coral-shaped surfaces. However, no significant differences were found between different surface coatings, including plain agar and agar enhanced with coral mucus exudates. Therefore future work should consider physico-chemical surface properties as factors governing change in microbial diversity.

Age-Related Shifts in Bacterial Diversity in a Reef Coral.

This study investigated the relationship between microbial communities in differently sized colonies of the massive coral Coelastrea aspera at Phuket, Thailand where colony size could be used as a proxy for age. Results indicated significant differences between the bacterial diversity (ANOSIM, R = 0.76, p = 0.001) of differently sized colonies from the same intertidal reef habitat. Juvenile and small colonies (<6cm mean diam) harboured a lower bacterial richness than medium (~10cm mean diam) and large colonies (>28 cm mean diam). Bacterial diversity increased in a step-wise pattern from juveniles<small<medium colonies, which was then followed by a slight decrease in the two largest size classes. These changes appear to resemble a successional process which occurs over time, similar to that observed in the ageing human gut. Furthermore, the dominant bacterial ribotypes present in the tissues of medium and large sized colonies of C. aspera, (such as Halomicronema, an Oscillospira and an unidentified cyanobacterium) were also the dominant ribotypes found within the endolithic algal band of the coral skeleton; a result providing some support for the hypothesis that the endolithic algae of corals may directly influence the bacterial community present in coral tissues.