Pseudoalteromonas piratica strain OCN003 is a coral pathogen that causes a switch from chronic to acute Montipora white syndrome in Montipora capitata

Abstract

Reports of mass coral mortality from disease have increased over the last two decades. Montipora white syndrome (MWS) is a tissue loss disease that has negatively impacted populations of the coral Montipora capitata in Kāne'ohe Bay, Hawai'i. Two types of MWS have been documented; a progressive disease termed chronic MWS (cMWS), that can be caused by Vibrio owensii strain OCN002, and a comparatively faster disease termed acute MWS (aMWS), that can be caused by Vibrio coralliilyticus strain OCN008. M. capitata colonies exhibiting cMWS can spontaneously switch to aMWS in the field. In this study, a novel Pseudoalteromonas species, P. piratica strain OCN003, fulfilled Koch's postulates of disease causation as another etiological agent of aMWS. Additionally, OCN003 induced a switch from cMWS to aMWS on M. capitata in laboratory infection trials. A comparison of OCN003 and Vibrio coralliilyticus strain OCN008, showed that OCN003 was more effective at inducing the cMWS to aMWS switch in M. capitata than OCN008. This study is the first to demonstrate that similar disease signs on one coral species (aMWS on M. capitata) can be caused by multiple pathogens, and describes the first Pseudoalteromonas species that infects coral.

Fig 1. Example of aMWS in Montipora capitata caused by OCN003.

(A) M. capitata before inoculation. (B) M. capitata 21 days post-inoculation with OCN003 displaying aMWS (white dashed line). The white scale bar represents one cm. (C) Kaplan–Meier survival curve of M. capitata fragments exposed to OCN003 (solid line with open black squares) or the control bacterium, OCN004 (dotted line with open circles), at 25°C (McNemar’s test, n = 24 each, p = 0.01). The concentration of bacteria used was 10⁸ CFU/ml of seawater.

Fig 2. Example of OCN003 inducing the switch to aMWS on a Montipora capitata fragment with cMWS.

(A) M. capitata with cMWS (white dashed line) before inoculation. (B) M. capitata three days after inoculation with OCN003, showing progressing tissue loss (aMWS; white dashed line). (C) M. capitata five days after inoculation with OCN003, displaying a complete loss of healthy tissue. The black scale bar represents one cm. (D) Kaplan–Meier survival curve of cMWS-affected M. capitata fragments exposed to OCN003 (solid line with open black squares) or the control bacterium, OCN004 (dotted line with open circles), at 25°C (McNemar’s test: n = 16 for each, p < 0.01). The concentration of bacteria used was 10⁸ CFU/ml of seawater.

Fig 3

Semi-solid (0.15% agar) assay confirming motility of wild type OCN003 cells (A), the lack of motility of the OCN003 ΔfliF::bla mutant cells (B), and the restored motility of the complemented OCN003 ΔfliF::bla mutant cells (C).

Fig 4. Kaplan–Meier survival curve of cMWS-affected M. capitata fragments exposed to OCN003 (solid line with open black squares), OCN003 ΔfliF::bla (solid grey line with open grey circles), the complemented OCN003 ΔfliF::bla mutant (dotted line with open black circles), or the control bacterium, OCN004 (solid grey line with closed grey triangles), at 25°C (n = 8 for each).

The concentration of bacteria used was 10⁸ CFU/ml of seawater.

Fig 5. Neighbor-joining dendrogram showing the estimated phylogenetic relationships between the Pseudoalteromonas piratica strains OCN003 (type strain), OCN050, OCN051, OCN052 (strains isolated during the 2010 aMWS outbreak), and related Pseudoalteromonas spp. based upon a multi-locus sequence analysis.

Analysis was based on the sequences of the housekeeping genes recA, gapA, and ftsZ. Alteromonas macleodii ATCC 27126 was chosen as the outgroup. The scale bar represents five nucleotide substitutions per 100 nucleotides. Bootstrap values >70% (500 replicates) are indicated at nodes.

Fig 6

Kaplan–Meier survival curves of healthy (A) and cMWS-affected (B) Montipora capitata fragments exposed to OCN003 (solid line with open black squares), OCN050 (solid line with open black triangles; Mantel-Cox test: n = 12, p = 0.71 (A); Mantel-Cox test: n = 16, p = 0.50 (B)), or the control bacterium, OCN004 (dotted line with open circles), at 25°C (n = 12 (A) or n = 16 (B) for each). The concentration of bacteria used was 10⁸ CFU/ml of seawater.

Fig 7. Kaplan–Meier survival curve of healthy or cMWS-affected M. capitata fragments exposed to OCN008 or the control bacterium, OCN004, at 25°C.

The solid line with open black rhombi represents healthy coral fragments exposed to OCN008 (n = 22); the dashed line with closed black rhombi represents cMWS-affected coral fragments exposed to OCN008 (n = 13); the dotted line with open circles represents healthy or cMWS-affected coral fragments exposed to OCN004 (n = 22 or n = 13, respectively). The concentration of bacteria used was 10⁸ CFU/ml of seawater.