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REPORT Multiple mechanisms of transmission of the Caribbean coral disease white plague E. Clemens1 • M. E. Brandt1 Received: 29 March 2014 / Accepted: 6 July 2015 / Published online: 15 July 2015  Springer-Verlag Berlin Heidelberg 2015 Abstract White plague is one of the most devastating coral diseases in the Caribbean, and yet important aspects of its epidemiology, including how the disease transmits, remain unknown. This study tested potential mechanisms and rates of transmission of white plague in a laboratory setting. Transmission mechanisms including the transport of water, contact with macroalgae, and predation via corallivorous worms and snails were tested on the host species Orbicella annularis. Two of the tested mechanisms were shown to transmit disease: water transport and the corallivorous snail Coralliophila abbreviata. Between these transmission mechanisms, transport of water between a diseased coral and a healthy coral resulted in disease incidence significantly more frequently in exposed healthy corals. Transmission via water transport also occurred more quickly and was associated with higher rates of tissue loss (up to 3.5 cm d-1) than with the corallivorous snail treatment. In addition, water that was in contact with dis- eased corals but was filtered with a 0.22-lm filter prior to being introduced to apparently healthy corals also resulted in the transmission of disease signs, but at a much lower rate than when water was not filtered. This study has pro- vided important information on the transmission potential of Caribbean white plague disease and highlights the need for a greater understanding of how these processes operate in the natural environment. Keywords Coral white plague  Coral disease  Caribbean coral reefs  Disease transmission Introduction Throughout the Caribbean, coral diseases have played a major role in recent coral mortality and widespread decli- nes among reefs (Richardson 1988; Weil 2004; Croquer et al. 2005). Coral diseases have the potential to threaten coral diversity and result in shifts to reef community structure and dynamics (Weil et al. 2003). Over thirty coral diseases have been reported globally, afflicting at least 150 species of coral (Richardson 1988; Green and Bruckner 2000; Jones et al. 2004). Understanding the relationship between coral and disease has proven difficult due to limited knowledge of the etiological agents and many of the fundamental processes associated with disease, including host resistance, pathogen virulence, and mecha- nisms of disease spread (Jones et al. 2004). However, as outbreaks of coral diseases continue to threaten reefs, there is a growing need for understanding of how disease initi- ates and propagates among corals. The coral disease white plague (WP) was one of the first diseases identified (Dustan 1977), and since that time, it has been reported from all parts of the Caribbean, affecting nearly half of all Caribbean coral species (Sutherland et al. 2004). Outbreaks of this disease have resulted in localized but significant declines in coral cover (Nugues 2002; Croquer et al. 2005; Richardson and Voss 2005). Addi- tionally, WP outbreaks following the 2005 mass coral bleaching event in the Caribbean resulted in losses of up to 40 % of coral cover on some reefs (Miller et al. 2009; Eakin et al. 2010). Several other WP outbreaks have been associated with environmental perturbation (Miller and Communicated by Biology Editor Dr. Anastazia Banaszak & E. Clemens eclemens@hawaii.edu 1 Center for Marine and Environmental Studies, University of the Virgin Islands, 2 John Brewers Bay, Saint Thomas, VI 00802, USA 123 Coral Reefs (2015) 34:1179–1188 DOI 10.1007/s00338-015-1327-6 Williams 2007; Miller et al. 2009; Brandt et al. 2013), but the mechanisms that contribute to the spread of disease during these outbreaks often remain unknown. Due to the potential for WP to cause extensive loss of coral in short time frames, there is a definitive need to identify mecha- nisms of disease spread within coral communities. Characteristic signs of WP include lesions that begin at the base or margin of a colony and enlarge rapidly (mm to cm d-1), creating a smooth margin of tissue loss delineating the intersection between living tissue and denuded white skeleton (Fig. 1; Bythell et al. 2004). The name ‘‘white plague’’ encompasses three distinct disease ‘‘types’’ (WP-1, WP-2, and WP-3) with similar disease signs distinguished by their rates of tissue loss. WP-1 progresses slowly (1–10 mm d-1) and is characterized by a smooth margin of tissue loss (Dustan 1977), WP-2 pro- gresses rapidly (up to 2 cm d-1) and occasionally shows a thin band of bleached tissue at the lesion edge (Richardson et al. 1998), and WP-3 progresses rapidly ([2 cm d-1), but only exclusively affects large colonies (3–4 m in diameter) of Colpophyllia natans and Orbicella (formerly Montas- traea) annularis (Richardson et al. 2001). Efforts to identify an etiological agent of the WP types have been ongoing since the first identification. WP-1 was originally associated with bacteria, but Koch’s postulates were not fulfilled (Dustan 1977), and WP-3 has not been associated with any microbial pathogen to our knowledge. Of the three types, WP-2 has been the most comprehen- sively characterized and has a known associated etiological agent. WP-2, originally documented in 1995, was first associated with a bacterium belonging to the group Sph- ingomonas (Richardson et al. 1998), later re-identified as a novel marine bacterium Aurantimonas coralicida (Denner et al. 2003). Nugues et al. (2004) showed that the macroalgae Halimeda opuntia has the potential to be a reservoir and can trigger infection by A. coralicidia. However, similar disease signs in different parts of the Caribbean have not been associated with A. coralicida (Pantos et al. 2003). For instance, Cook et al. (2013) detected A. coralicida in low abundances in samples from both WP affected corals and apparently healthy corals, suggesting it was not the etiological agent of the observed WP signs. However, Gray et al. (2013) identified PCR bias as a potential mechanism preventing the detection of A. coralicida in WP samples. Clearly, the debate over the etiology of WP-2 and other WP types continues, and most recently, viruses have been identified as playing a potential role in WP-1 in the US Virgin Islands (Soffer et al. 2013). Studies of the etiological agent have suggested that WP is an infectious disease caused by a microbial pathogen versus an apoptotic state as has been found in a similar coral disease described from the Pacific (Ainsworth et al. 2007). Understanding how the infection transmits among corals is an important component of characterizing the disease and can also provide an ecological context for etiological agents that are identified. Our understanding of WP transmission in the marine environment has primarily been inferred from spatial patterns of disease. Clustered distributions of diseased individuals typically indicate an infectious nature of a disease (Diggle 1983), and several studies have associated WP with such a distribution (Bor- ger 2003; Voss and Richardson 2006; Brandt and McMa- nus 2009b). However, other studies have not found clustering (Nugues 2002; Borger 2005; Richardson and Voss 2005; Muller and van Woesik 2012), and these con- flicting results may be due to inconsistent sampling regimes and scales. Due to the potential for high connec- tivity within marine environments (McCallum et al. 2003), a clustered distribution of coral disease may indicate transmission through some vector that is limited in its range, such as slow water flow, contact with algae, or via a mobile predator (Brandt and McManus 2009b). In labora- tory experiments, Richardson et al. (1998) showed that the original WP-2 pathogen could spread through water borne transmission within a small tank. Research in the marine environment has demonstrated that WP-2 could be trans- ferred between colonies via the calcareous green macroalga species H. opuntia (Nugues et al. 2004) and in the Virgin Islands WP-1 signs were transferrable via direct contact between two separate pieces of coral that were fixed together (Brandt et al. 2013). Clearly, more work is needed to identify mechanisms of WP transmission and their potential to aid WP propagation in the marine environment. Fig. 1 White plague affecting a colony of Orbicella annularis in Brewers Bay, St. Thomas, USVI. Photograph taken September 23, 2010, 5 m depth 1180 Coral Reefs (2015) 34:1179–1188 123 Multiple mechanisms of transmission have been iden- tified in other disease systems of corals. In studies of black- band disease (BBD) of corals, an infection was able to be transmitted through direct contact (Rutzler et al. 1983) and through a corallivorous butterflyfish vector by oral and fecal transmission (Aeby and Santavy 2006). Other known vectors of disease transmission in corals include the poly- chaete Hermodice carunculata (Sussman et al. 2003), which is both a reservoir and vector for the bacterial bleaching pathogen Vibrio shiloi in the Mediterranean, and the mollusk Coralliophila abbreviata, which was identified as a potential vector for an unknown pathogen causing rapid tissue loss in acroporid corals in the Florida Keys (Williams and Miller 2005). Both H. carunculata and C. abbreviata serve as vectors of infectious agents that may cause disease by feeding on an infected colony and sub- sequently feeding on and infecting a healthy colony. Both H. carunculata and C. abbreviata are found, sometimes abundantly, feeding on corals affected by WP, but these potential vectors have not been tested as mechanisms of WP transmission. In the US Virgin Islands (USVI), WP has been identified affecting corals in all monitored reef habitats for at least a decade (Smith et al. 2008). This disease had a devastating impact on many reef communities following the 2005 mass bleaching event (Miller et al. 2009) and continues to affect coral populations throughout the region (Smith et al. 2013). Recent targeted studies of WP in this region have shown that signs most closely match those described for WP-1 (Brandt et al. 2013). In addition to its association with mass bleach- ing, this disease was also associated with storm-related fragmentation and contact with sediment (Brandt et al. 2013). Its spatial distribution suggests infectious transmis- sion (Brandt et al. 2013), and viruses may play an important role in the etiology of this disease (Soffer et al. 2013). In this study, we examined multiple potential mecha- nisms of transmission of the Virgin Islands WP-1 in a controlled laboratory setting. Experiments consisted of using fragments of the susceptible reef-building coral O. annularis exposed to various vectors and media with the potential to transmit the unknown pathogen causing WP. We expected that multiple mechanisms would transmit the unknown pathogen causing disease, but that one would be the most effective. By identifying transmission mecha- nisms and targeting them for further study, we aimed to provide better understanding of the determinants behind the dynamics and ultimately the impact of this disease. This work may also lead to the identification of important aspects of the transmission mechanisms that could be tar- geted for intervention or in an effort to prevent future outbreaks and impacts. Methods Study region All research took place in St. Thomas, USVI, at the Center for MarineandEnvironmentalStudies(CMES)attheUniversityof the Virgin Islands (UVI) from November 1 to December 22, 2012. Collection of experimental animals occurred at two locations on the south side of St. Thomas separated by a dis- tance of 4.65 km, and both sites typically experience low dis- ease prevalence (\1 %; Brandt pers. obs.) These sites (Perseverance Bay: 1820053.4500N 6459033.0900W and Saba Island: 1818019.7000N 6459055.2700W) are characterized by shallow reefs (\10 m) dominated by the coral species O. an- nularis and O. faveolata. Specimen collection and acclimation Experimental corals were exposed to treatments in three separate trials that took place in 2012 from 1 to 22 November, 12 November to 3 December, and 1 to 22 December. For each of the trials, fragments of O. annularis were collected from sites by divers on SCUBA. From each site, 2–3 coral fragments of\30 cm in maximum diameter were collected from 4 to 5 colonies that were separated by at least 5 m. The growth form of O. annularis is such that the colony grows in columns with living tops and dead sides. Fragments collected for the study consisted of the living tops of columns that were broken off at the dead area using a hammer and chisel. Large fragments were further fragmented such that a total of 25 pieces of apparently healthy O. annularis were used in each trial. Average fragment size had a length of 6.7 cm ± 0.2 SE, width of 5.4 cm ± 0.2, and area of 26.7 cm2 ± 1.4. Following collection, fragments were immediately transferred in seawater to the laboratory where they were placed in running seawater tables and allowed to acclimate for 7 d before being used in various treatments. The colony of origin was recorded and tracked for each coral fragment, and all fragments were photographed and monitored daily during a 7-d acclimation period. The sea water tables are part of a flow-through system located at CMES, which sits next to Brewer’s Bay. The system consists of seawater that is pumped from Brewer’s Bay into a head tank located physically higher than the elevation of the Marine Science building. Seawater flows by gravity to the culture tanks below via a 3 Sch. 40 PVC main supply line. Seawater temperatures in the water tables remained at ambient levels found on the reef during the same time period (Trial 1: 28.7–29.0 C, Trial 2: 28.3–28.8 C, Trial 3: 27.8–28.3 C). Coral Reefs (2015) 34:1179–1188 1181 123 TwopiecesofWP-diseasedcoral werecollected separately and kept in a separate seawater table. Six additional fragments of apparently healthy O. annularis were directly exposed to these pieces, developed signs of WP, and were subsequently used in experiments as inoculant corals. Therefore, a total of eight diseased fragments were used in inoculant experiments in each trial. Each diseased fragment was assessed each day to determine whether the lesion was still progressing and active before being used in any experiments. The mobile vectors (H. carunculata and C. abbreviata) were collected from intertidal areas around the dock at the Marine Science Building on Brewer’s Bay. For each trial, three individuals of H. carunculata and three of C. abbre- viata were used and allowed to acclimate in separate 2-L buckets with individual bubblers and daily water changes for 3 d. H. carunculata that were collected ranged in size from 7 to 9 cm long. The animals were not fed before being used in experiments, and during the experiment, all H. carunculata and C. abbreviata were documented feed- ing on experimental corals. Clumps of Dictyota menstru- alis separated by 5 m were also collected from the dock area and dabbed dry before they were allowed to acclimate in separate 2-L buckets with bubblers and daily water changes for 3 d before being used in experiments. Experimental design After acclimation, coral fragments were removed from seawater tables and placed into numbered 2-L buckets with an individual bubbler and daily water changes. Coral fragments were then randomly assigned to be used as an inoculant, treatment, or control. Five sets of experiments explained below were performed, including (1) unfiltered water, (2) filtered water, (3) Dictyota, (4) C. abbreviata, and (5) H. carunculata (Fig. 2). Each experiment included a set of controls, and all buckets were washed with alcohol before each trial. In the unfiltered water experiment, treatment corals received 1/2 L of water from a bucket with a diseased inoculation coral during their daily water changes. Corresponding control corals received 1/2 L of water from a bucket with a healthy coral. The treatment and control corals were photographed and monitored each day for signs of disease. An identical design was used in the filtered water experiment, but water being transferred between buckets was first filtered using a 0.22-lm filter (Millipore Express, 47 mm, polyethersulfone, hydrophilic). For both the unfiltered and filtered water experiments, treatment and control corals were subjected to transferred water daily for the entirety of a 2-week study period. Fig. 2 Experimental design of transmission experiments. Replicates for the water transmission experiment included a treatment replicates where 0.5 L of filtered or unfiltered water was transferred from a 2-L bucket containing a diseased coral to a 2-L bucket containing a healthy coral, and b control replicates for the water transmission experiment consisted of the same design except water was transferred from a 2-L bucket containing a healthy coral. Replicates for the vector transmission experiments included c treatment replicates where Coralliophila abbreviata or Hermodice carunculata were allowed to feed on a diseased coral or Dictyota was placed in contact with a diseased coral and then transferred to a bucket with a healthy coral, and d control replicates where the same design was used, but the initial coral was a healthy coral. Red ovals represent diseased corals, and green ovals represent apparently healthy corals 1182 Coral Reefs (2015) 34:1179–1188 123 Buckets with inoculation corals used as a source of water were refilled and allowed to sit for 24 h before being sampled again. Therefore, water that was used in the treatments had been in contact with either diseased or healthy corals for at least 24 h but no more than 36 h before being transferred to the treatment corals. For H. carunculata, C. abbreviata, and Dictyota experiments, each suspected vector was placed in either a diseased or healthy coral bucket and allowed to feed for 2 d before being transferred into a treatment or control bucket. All buckets received individual bubblers and daily water changes. Each coral fragment was then photographed and monitored for signs of disease and mortality daily. Inocu- lation corals used for water experiments were not used in vector experiments. Each trial lasted 2 weeks, and coral fragments were monitored until they died from disease or survived until the trial period was over. For each of the three trial periods, there were two controls and four treatments per trial for each of the filtered and unfiltered water experiments. This resulted in a total of N = 12 treatments and N = 6 controls for each experiment. For the Dictyota, H. carunculata, and C. abbreviata experiments, there was one control and three treatments per trial, resulting in a total of N = 9 treatments and N = 3 controls for each experiment. Low sample sizes were limited by logistical and permit constraints, and sequential trials were performed due to the scattered availability of diseased corals located in the natural environment. Following previous descriptions (Remily and Richard- son 2006), a WP lesion was defined as a smooth and undulating tissue margin adjacent to freshly denuded skeleton and exposed coral skeleton that continued expanding across the coral for at least 2 d. In contrast, feeding scars from the mobile vectors and other lesions due to fragmentation were distinguished from lesion signs based on their ragged edges the fact that the lesion did not enlarge following feeding or contact. In order to calculate lesion enlargement rates, the area of recently denuded skeleton left by the lesion was traced using ImageJ software (NIH) for each daily photograph of fragments that developed lesions. Each lesion was mea- sured anywhere from 3 to 6 times throughout the lesion’s enlargement. The length of time for the disease signs to appear was recorded as the number of days between initial exposure to the treatment and start of tissue loss. Statistical analyses Rates of lesion appearance after exposure to inoculant corals were calculated as the number of days from the start of the trial until a WP lesion was observed. Where lesions appeared on multiple corals, a nonparametric Wilcoxon test was performed to test for a significant effect on the rate of lesion appearance between the two experiment types. Tissue loss rate data were calculated per lesion as the area (cm2) of tissue loss per day. These data were first square root transformed to approximate normality and then tested using a nested ANOVA with experiment type as the main factor and coral of origin as a nested factor. Once coral of origin was determined not to affect lesion pro- gression rates, a repeated-measures ANOVA was used to test for an effect of time and experiment type on lesion progression rate. To determine whether a relationship existed between the experiment type and the number of corals that became diseased, a contingency table analysis was used. A separate contingency table analysis tested for a relationship between the number of corals that became diseased and coral of origin. Results White plague disease signs were transmitted between dis- eased and healthy corals, but were never transmitted in any control treatments. Eleven of the 42 (26 %) treatment fragments became affected by WP during the study period (Fig. 3). The most effective mechanism of transmission was unfiltered water, which transmitted the disease 8/12 times, followed by filtered water (2/12) and C. abbreviata (1/6). Signs of disease transmission were not observed in either H. carunculata or Dictyota experiments; however, feeding was observed on all H. carunculata treatments. However, Dictyota caused partial bleaching in 2/3 controls and 5/6 experimental treatments. Fragments in the Fig. 3 Percent of experimental colonies that became affected by disease in each experiment. N are indicated in parentheses Coral Reefs (2015) 34:1179–1188 1183 123 unfiltered water experiment showed faster rates of lesion appearance than those in the filtered water experiments (v2 = 4.06, df = 1, p \ 0.05; Fig. 4). The average days until tissue loss began after exposure for unfiltered water, C. abbreviata, and filtered water were 2, 3, and 4.5 d, respectively (Fig. 4). Overall, new tissue losses were observed between 1 and 5 d after exposure to the different treatments. Overall, average lesion enlargement rate on experi- mental corals was 0.8 cm2 d-1 ± 0.2 (SEM), with a range of 0.007–3.6 cm2 d-1. Most lesions resulted in total mor- tality, where total mortality was defined as the complete loss of tissue from the coral fragment. Average length of time until total mortality was 6.7 d (±1.5 SEM) after the first initiation of the lesion. Figure 5 shows an example of a lesion’s initiation and tissue loss progression to completely denuded skeleton. The nested ANOVA revealed no sig- nificant effect of coral of origin (F = 8.2637, df = 8, p [ 0.05). When tested without coral of origin as a factor, there was no significant difference in progression rates between experiments (Kruskal–Wallis test: v2 = 0.75, df = 2, p [ 0.05). However, throughout the lesion’s enlargement, the new tissue loss rates at each time of measurement increased (repeated-measures ANOVA: F = 9.23, df = 3.6, p \ 0.05; Fig. 6). The results of the contingency table analyses indicated that the distribution of colonies that became diseased ver- sus remaining healthy was significantly related to experi- mental mechanism, but this distribution was not significantly related to the colony of origin (Table 1). However, low sample sizes for both of these analyses indicate a cautious interpretation of the Chi-square statistic. Fig. 4 Average rates of lesion appearance (±SEM) within treatments where disease transmission was identified. N are indicated in parentheses Fig. 5 Expansion of a white plague lesion in Trial 1 where the coral fragment was exposed to a Coralliophila abbreviata that had been allowed to feed on a separate diseased coral fragment. a Day 2 after exposure showing a feeding scar on the bottom right of the fragment, b day 3 showing the development of a smooth lesion line, c day 4 where the lesion has expanded across the colony fragment, and d day 5 where total mortality has occurred 1184 Coral Reefs (2015) 34:1179–1188 123 Discussion Disease transmission The results of this study demonstrate that the causal agent of WP disease can be transmitted through multiple mech- anisms, including water transport and snail predation. Transmission via water transport was the most effective mechanism, supporting the early observations by Richardson et al. (1998) that WP can be transmitted among colonies through water transport in laboratory conditions. While the experiments lasted 2 weeks, signs of disease generally occurred within the first 1–2 d for the unfiltered water treatment and 5–6 d for the filtered water and snail treatments, suggesting that transmission can occur rapidly. This is in contrast to field experiments in the same reef system where direct tissue-to-tissue transmission was not observed until after 3 weeks (Brandt et al. 2013). The discrepancy in timing was likely the result of laboratory conditions; laboratory coral fragments were kept in 2-L buckets with circulation limited to bubblers and daily water changes, while experimental corals in the field had natural circulation conditions and were constantly flushed. This may have allowed for potential pathogenic agents to be removed from field corals, while laboratory corals were in constant contact with materials that could have resulted in infection. Alternatively, laboratory conditions may have affected the resistance of experimental corals to disease and increased their susceptibility to infection. Follow-up experiments should investigate the timing of disease transmission in the field and in laboratory experiments that reflect more natural conditions. In this study, water filtered to remove bacteria[0.22 lm was also able to induce disease signs, although at a highly reduced rate. Recent work has shown that viruses may play an important role in the etiology of WP in the Virgin Islands, and the transmission of disease using 0.2-lm fil- tered water supports a potentially viral infectious agent (Soffer et al. 2013). However, the low transmission rate of disease in the filtered water compared with unfiltered water treatments (2/12 vs. 8/12, respectively) suggests that the process of infection is more rapid when large microbial components such as bacteria are present. This could indi- cate a role of a filterable factor in the initiation of disease, similar to what has been demonstrated in WP signs from other regions (Barash et al. 2005). However, it is possible that these results are the result of contamination or incomplete efficiency, although all precautions were taken to reduce the risk of contamination and methods were applied equally to all treatments. The microbial commu- nities found within a coral are diverse and include bacteria, viruses, and Archaea (Rosenberg et al. 2007). While less is known about the role of viruses and Archaea, bacteria have been noted to work as pathogen defense mechanisms and play a role in nutrient cycling (Lesser et al. 2007). Shifts in the overall microbial communities of corals can result in decreased resistance to disease (Ritchie 2006), and poten- tially increased virulence of pathogens (Vega-Thurber et al. 2012). Therefore, it would not be surprising if WP results from changes in multiple components of the microbial community and is not just the results of infection by one etiological agent. The mobile vectors C. abbreviata and H. carunculata showed very low (1/6) and no transmission of the disease, respectively. Field studies have shown that feeding of C. abbreviata resulted in rapid tissue loss in A. cervicornis (Williams and Miller 2005) and have reported H. caruncu- lata feeding on disease lesions (Miller and Williams 2007). Since these vectors have limited mobility and are not likely to travel between reefs, it is far more likely that water transmission may be the effective mechanism of transmis- sion in the natural environment. However, as these predators feed on and negatively impact the coral’s health, they may cause the coral to expend extra energy to heal feeding scars. This would stress the coral resulting in potential negative effects to the corals’ immune system and possibly making it more susceptible to disease. Williams and Miller (2005) also hypothesized that once disease is introduced into a system, mobile predators that feed on one colony with disease can carry it to another, intensifying disease spread within the reef system. Although predators such as these may not be the primary mechanism of pathogen transmission within an ecosystem, they may have the capability of increasing the coral population’s chances of infection or transfer the pathogen directly through feeding. Fig. 6 Average rates of lesion expansion (±SEM) at successive time points across the lesion’s enlargement. Letters indicate significant groups (p \ 0.05) as determined by post hoc tests following a significant repeated-measures ANOVA. N = 12 at each measurement time point Coral Reefs (2015) 34:1179–1188 1185 123 Although no Dictyota treatments were found to transmit disease, the presence of Dictyota caused mild paling to complete bleaching of several experimental corals, indi- cating a negative interaction between this macroalga and the corals in this study. Macroalgae can negatively affect coral health (Nugues et al. 2004), and Dictyota negatively affects coral by causing hypoxia (Barott et al. 2012) or emitting dissolved compounds (Smith et al. 2006). This macroalgal species group has also been associated with bacterial shifts in coral tissue (Morrow et al. 2012). This may provide insight into the correlation Brandt et al. (2012) reported between increased Dictyota cover and the expansion of WP disease when there were no other phys- ical or biological factors influencing the outbreak of dis- ease. The bleaching of coral tissue and shifts in the bacterial communities due to the interaction with the macroalgae may increase the risk of infection. Brandt and McManus (2009a) found that the incidence of WP increased in Orbicella after a bleaching event, and similar patterns have been identified in different coral disease systems (Harvell et al. 2002; Weil et al. 2006; Muller et al. 2008). Although most disease lesions in this study appeared within the first 4 d, in the case of Dictyota, more time may have been needed to see the full effects of this macroalga on disease incidence. Enlargement rates Interestingly, in this study the enlargement rates of disease on individual corals increased through time. Although these results derive from laboratory experiments, it is possible that this trend also exists in the field and that measuring the rate of WP lesion progression at only one or two points in time (compared to the entirety of the lesion’s life) may cause incorrect classification of WP types. The change in enlargement rates on individual corals through time could be explained by progressive changes in the microbial community, which is known to change within the coral holobiont in response to diseases and other stressors (Ritchie 2006). In some instances, bacteria within the holobiont coordinate their behavior by regulating gene expression and creating a community wide response. This phenomenon is known as quorum sensing (QS), which can take place when density dependent cell-to-cell communi- cation occurs among bacteria within the coral (Fuqua et al. 1994). QS has also been noted to regulate bacteria’s response to virulence (Bandara et al. 2012). For bacterial pathogens, QS molecules are used in the establishment of an infection and help regulate a switch from a non- pathogenic to a pathogenic state. These regulated systems have the potential to suppress the coral’s immune response to pathogens until an adequate number of bacteria have accumulated to have a successful infection (Bassler 1999). Recent studies have found a QS signal in isolates from coral diseases such as BBD (Goldberg et al. 2011). In this study, QS could be causing the large range of progression rates by delaying responses or by increasing the virulence of a pathogen. However, if QS is disrupted, it can block production of antibiotics or even reduce the virulence of a pathogen, which may also explain the wide ranges of progression rates recorded in the field (Teplitski and Ritchie 2009). In summary, WP is one of the most important sources of coral mortality on Caribbean reefs, yet ambiguity remains regarding its emergence and distribution in the natural environment. In this study, we have provided insight into the potential pathogen transmission mechanisms and dynamics of this disease, although our studies were limited to the laboratory environment. Future research should investigate transmission dynamics in wild populations, Table 1 Observed and expected (in parentheses) frequencies of colonies that became diseased or remained healthy (not diseased) with respect to treatment types with the results of the tests of independence Diseased Not diseased Statistical results Treatment v2 = 15.283, df = 4, p = 0.0042 Dictyota 0 (1.57) 9 (4.42) (Significant) Filtered water 2 (3.14) 16 (8.86) Fireworm 0 (1.57) 9 (4.42) Snail 1 (1.57) 8 (4.42) Unfiltered water 8 (3.14) 8 (8.86) Coral of origin v2 = 3.613, df = 6, p = 0.7289 2 1 (1.78) 5 (4.21) (Not significant) 3 3 (2.37) 5 (5.62) 20 3 (1.48) 2 (3.51) 1 1 (1.48) 4 (3.51) 27 1 (0.89) 2 (2.11) 7 1 (1.46) 4 (3.51) 12 1 (1.46) 4 (3.51) 1186 Coral Reefs (2015) 34:1179–1188 123 especially in light of the severity and increasing number of reports of WP outbreaks. 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