Mucochytrium quahogii, also known as QPX or Quahog Parasite Unknown, is the causative agent of QPX disease in the hard clam (Mercenaria mercenaria). Host–pathogen–environment interactions between M. quahogii, the hard clam, and temperature were explored in a microcosm experiment. Hard clams were housed in individual tanks with sterile seawater under two temperature regimes: low (13 °C) temperature, which is thought to be optimal for QPX disease development, and high (20 °C) temperature, which has been shown to promote “healing” of QPX-infected clams. Hard clam tissue, pallial fluid, seawater, and shell biofilms were collected and assayed for M. quahogii. The release of M. quahogii from naturally infected live hard clams into seawater was detected only in the low temperature treatment, suggesting that temperature influences the release of potentially infectious cells. M. quahogii was commonly found in hard clam pallial fluid, even after 9 weeks in the lab, suggesting pallial fluid is a stable reservoir of M. quahogii within its primary host and that M. quahogii is not a transient component of the hard clam microbiota. Overall, results support a host-specific relationship and that M. quahogii is a commensal member of the hard clam microbiota, supporting its classification as an opportunistic pathogen.

Atlantic menhaden are a highly migratory marine species in the Eastern United States that suffer from seasonal chronic mortality. Affected fish show neurologic signs referred to as spinning disease, including circling at the surface and erratic corkscrew swimming before death. We investigated three similar menhaden mortality events consistent with spinning disease in coastal New Jersey and New York between 2020 and 2021 to understand the cause. A unique strain of Vibrio (Listonella) anguillarum (serogroup O3) was detected regularly in high loads, particularly in the brains of moribund fish, by both metagenomics and bacterial isolation. The most common histopathological changes in moribund fish were hemorrhagic meningitis, encephalitis, pyknosis, and karyorrhexis of hematopoietic tissues in the kidney and spleen. Whole genome sequencing of isolates from moribund fish representing a wide spatial and temporal range showed that they were nearly identical clones, suggesting it to be a pathogenic strain circulating in the population. Though V. anguillarum is believed to be the main pathogen associated with spinning disease and mortality, Yersinia ruckeri (serotype O1) was isolated from smaller numbers of fish. Considering the highly migratory nature of Atlantic menhaden throughout the eastern United States and their use as bait for other fisheries, these findings identify potential biosecurity challenges that should be considered in Atlantic salmon aquaculture, fisheries, and emerging marine aquaculture in the region.

The bay scallop, Argopecten irradians, represents a commercially, culturally and ecologically important species found along the United States' Atlantic and Gulf coasts. Since 2019, scallop populations in New York have been suffering large-scale summer mortalities resulting in 90-99% reduction in biomass of adult scallops. Preliminary investigations of these mortality events showed 100% prevalence of an apicomplexan parasite infecting kidney tissues. This study was designed to provide histological, ultrastructural and molecular characteristics of a non-described parasite, member of the newly established Marosporida clade (Apicomplexa) and provisionally named BSM (Bay Scallop Marosporida). Molecular diagnostics tools (quantitative PCR, in situ hybridization) were developed and used to monitor disease development. Results showed that BSM disrupts multiple scallop tissues including kidney, adductor muscle, gill, and gonad. Microscopy observations allowed the identification of both intracellular and extracellular stages of the parasite. Field surveys demonstrated a strong seasonal signature in disease prevalence and intensity, as severe cases and mortality increase as summer progresses. These results strongly suggest that BSM infection plays a major role in the collapse of bay scallop populations in New York. In this framework, BSM may synergistically interact with stressful environmental conditions to impair the host and lead to mortality.

Progress in understanding and managing QPX disease outbreaks in hard clams (Mercenaria mercenaria) has been limited by lack of insight into basic aspects of the biology and ecology of the opportunistic pathogen Mucochytrium quahogii (formerly QPX or Quahog Parasite Unknown). One barrier is that while several methods have been able to detect M. quahogii in seawater and sediment, its abundance was typically too low for reliable quantification by those methods. Here we describe the development and validation of a sensitive, M. quahogii-specific, nested quantitative PCR (nqPCR) assay following the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines. The assay reaches the theoretical limit of detection (LOD) of a PCR assay at 3 copies per reaction with excellent efficiency, linearity, and minimal sample PCR inhibition. The functionality of the assay was evaluated by quantifying M. quahogii in sediment and seawater samples, which revealed that M. quahogii was broadly distributed throughout the marine environment, detected in 75% of samples, with mean estimated abundance of 0.21 cells per mg sediment, 0.55 cells per ml bottom seawater, and 0.02 cells per ml surface seawater. M. quahogii was most prevalent and most abundant in sediment and bottom seawater samples, suggesting that the flocculent layer at the sediment-water interface is an important environmental reservoir where M. quahogii may interact with hard clams. This assay will serve as a valuable tool to better understand QPX disease dynamics and offers a model to guide development of similar assays for other important marine microbes typically present at similarly low abundance.

With marine diseases on the rise and increased reliance on molecular tools for disease surveillance, validated pathogen detection capabilities are important for effective management, mitigation, and response to disease outbreaks. At the same time, in an era of continual evolution and advancement of molecular tools for pathogen detection, it is critical to regularly reassess previously established assays to incorporate improvements of common practices and procedures, such as the minimum information for publication of quantitative real-time PCR experiments (MIQE) guidelines. Here, we reassessed, re-optimized, and improved the quantitative PCR (qPCR) assay routinely used for Quahog Parasite Unknown (QPX) disease monitoring. We made 19 significant changes to the qPCR assay, including improvements to PCR amplification efficiency, DNA extraction efficiency, inhibition testing, incorporation of linearized standards for absolute quantification, an inter-plate calibration technique, and improved conversion from copy number to number of cells. These changes made the assay a more effective and efficient tool for disease monitoring and pathogen detection, with an improved linear relationship with histopathology compared to the previous version of the assay. To support the wide adoption of validated qPCR assays for marine pathogens, we provide a simple workflow that can be applied to the development of new assays, re-optimization of old or suboptimal assays, or assay validation after changes to the protocol and a MIQE-compliant checklist that should accompany any published qPCR diagnostic assay to increase experimental transparency and reproducibility amongst laboratories.

Mucochytrium quahogii, commonly known as QPX (Quahog Parasite Unknown), is the causative agent of QPX disease in hard clams (Mercenaria mercenaria), but poor understanding of the relationship between host and pathogen has hindered effective management. To address this gap in knowledge, we conducted a two-year study quantifying the distribution and abundance of M. quahogii in hard clam tissue, pallial fluid, and the environment. M. quahogii was broadly distributed in clams and the environment, in areas with and without a known history of QPX disease. M. quahogii in clams was not strongly related to M. quahogii in the environment. M. quahogii was always present in either the tissue or pallial fluid of each clam, with an inverse relationship between the abundance in the two anatomical locations. This study suggests that the sediment–water interface and clam pallial fluid are environmental reservoirs of M. quahogii and that there is a host-specific relationship between M. quahogii and the hard clam, supporting its classification as a commensal, opportunistic pathogen. There appears to be minimal risk of spreading QPX disease to naïve clam populations because M. quahogii is already present and does not appear to be causing disease in hard clam populations in locations unfavorable for pathogenesis.

Quahog Parasite Unknown (QPX) is a facultative parasite of the hard clam, Mercenaria mercenaria. Although it has been observed in clams since the 1960's and cultivated since the 1990's, conflicting reports on important aspects of its biology have prevented its formal description. 18S rRNA gene sequences identify QPX as a thraustochytrid, but its production of copious mucus is atypical for this group. There are also conflicting reports about whether QPX shares common features of thraustochytrids, such as the production of an ectoplasmic net and biflagellate zoospores. This study reaffirms the previous descriptions of zoospore production by QPX in culture, in multiple strains from several geographic locations, and provides detail on how to maintain QPX cultures under conditions that promote the production of zoospores. Furthermore, we describe new aspects of the life cycle not previously observed. Finally, we erect Mucochytrium quahogii gen. nov., sp. nov. to accommodate this unusual thraustochytrid.

The phylum Labyrinthulomycota comprises diverse marine fungus-like protists that are an abundant and widespread component of the marine microbiota. Despite their ubiquity in marine ecosystems, relatively little is known about the ecology of any of the pathogenic species in the Labyrinthulomycota. Most are thought to exist as saprobes, but many species have been documented as pathogens of marine metazoans and metaphytes. The best studied labyrinthulomycotan pathogen in molluscs is Quahog Parasite Unknown (QPX), which causes mortality events in both wild and cultured hard clams, Mercenaria mercenaria. Other labyrinthulomycotan pathogens affect a variety of other molluscs, including juvenile abalone, squid and octopus. Both the ectoplasmic network, a unique feature of the Labyrinthulomycota that attaches vegetative cells to their substrate, and their heterokont zoospores may play important roles in pathogenesis. Most labyrinthulomycotan pathogens appear to be opportunistic, rarely causing disease in healthy hosts but becoming pathogenic when changes in host immunity or other factors alter the host-parasite relationship. All reported labyrinthulomycotan pathogens appear to be host-specific, potentially making Labyrinthulomycota unusual in terms of pathogenesis and virulence, as they seem to be hostspecific like obligate pathogens, but appear to behave and cause disease like opportunistic pathogens.

The widely distributed marine cyanobacterium Synechococcus is thought to exert an influence on the marine silicon (Si) cycle through its high cellular Si relative to organic content. There are few measurements of Si in natural populations of Synechococcus, however, and the degree to which Synechococcus from various oligotrophic field sites and depths accumulate the element is unknown. We used synchrotron x-ray fluorescence to measure Si quotas in individual Synechococcus cells collected during three cruises in the western North Atlantic Ocean in the summer and fall, focusing on cells from the surface mixed layer (SML; <10m) and the deep chlorophyll maximum (DCM). Individual cell quotas varied widely, from 1 to 4700amol Si cell−1, though the middle 50% of quotas ranged between 17 and 119amol Si cell−1. Mean station-specific quotas exhibited an even narrower range of 31–72amol Si cell−1. No significant differences in Si quotas were observed across cruises or among stations, and no effect of ambient silicic acid concentration on quotas was observed within the narrow range of silicic acid concentrations encountered (0.6–1.3μM). Despite this small range in ambient silicic acid, cells collected from the SML had an average of two-fold more Si than cells collected from the DCM. Differences in Si content with depth may be related to observed differences in the dominant Synechococcus clades between the SML and DCM habitats, determined by petB gene sequencing.