Structural asymmetry in a trimeric Na+/betaine symporter, BetP, from Corynebacterium glutamicum

The Na⁺-coupled symporter BetP catalyzes the uptake of the compatible solute betaine in the soil bacterium Corynebacterium glutamicum. BetP also senses hyperosmotic stress and regulates its own activity in response to stress level. We determined a three-dimensional (3D) map (at 8 Å in-plane resolution) of a constitutively active mutant of BetP in a C. glutamicum membrane environment by electron cryomicroscopy of two-dimensional crystals. The map shows that the constitutively active mutant, which lacks the C-terminal domain involved in osmosensing, is trimeric like wild-type BetP. Recently, we reported the X-ray crystal structure of BetP at 3.35 Å, in which all three protomers displayed a substrate-occluded state. Rigid-body fitting of this trimeric structure to the 3D map identified the periplasmic and cytoplasmic sides of the membrane. Fitting of an X-ray monomer to the individual protomer maps allowed assignment of transmembrane helices and of the substrate pathway, and revealed differences in trimer architecture from the X-ray structure in the tilt angle of each protomer with respect to the membrane. The three protomer maps showed pronounced differences around the substrate pathway, suggesting three different conformations within the same trimer. Two of those protomer maps closely match those of the atomic structures of the outward-facing and inward-facing states of the hydantoin transporter Mhp1, suggesting that the BetP protomer conformations reflect key states of the transport cycle. Thus, the asymmetry in the two-dimensional maps may reflect cooperativity of conformational changes within the BetP trimer, which potentially increases the rate of glycine betaine uptake.     

Identifying biotic drivers of population dynamics in a benthic–pelagic community

Benthic species and communities are linked to pelagic zooplankton through life‐stages encompassing both benthic and pelagic habitats and through a mutual dependency on primary producers as a food source. Many zooplankton taxa contribute to the sedimentary system as benthic eggs. Our main aim was to investigate the nature of the population level biotic interactions between and within these two seemingly independent communities, both dependent on the pelagic primary production, while simultaneously accounting for environmental drivers (salinity, temperature, and oxygen conditions). To this end, we applied multivariate autoregressive state‐space models to long (1966–2007) time series of annual abundance data, comparing models with and without interspecific interactions, and models with and without environmental variables included. We were not able to detect any direct coupling between sediment‐dwelling benthic taxa and pelagic copepods and cladocerans on the annual scale, but the most parsimonious model indicated that interactions within the benthic community are important. There were also positive residual correlations between the copepods and cladocerans potentially reflecting the availability of a shared resource or similar seasonal dependence, whereas both groups tended to correlate negatively with the zoobenthic taxa. The most notable single interaction within the benthic community was a tendency for a negative effect of Limecola balthica on the amphipods Monoporeia affinis and Pontoporeia femorata which can help explain the observed decrease in amphipods due to increased competitive interference.     

Mustelid herpesvirus-2, a novel herpes infection in northern sea otters (Enhydra lutris kenyoni)

Oral ulcerations and plaques with epithelial eosinophilic intranuclear inclusions were observed in northern sea otters (Enhydra lutris kenyoni) that died or were admitted for rehabilitation after the 1989 Exxon Valdez oil spill (EVOS) in Alaska, USA. Transmission electron microscopy demonstrated the presence of herpesviral virions. Additionally, a serologic study from 2004 to 2005 found a high prevalence of exposure to a herpesvirus in live-captured otters. Tissues from 29 otters after the EVOS and nasal swabs from 83 live-captured otters in the Kodiak Archipelago were tested for herpesviral DNA. Analysis identified a novel herpesvirus in the gamma subfamily, most closely related to Mustelid herpesvirus-1 from badgers. Results indicated that this herpesvirus is associated with ulcerative lesions but is also commonly found in secretions of healthy northern sea otters.     

Human-induced eutrophication enhances reproductive success through effects on parenting ability in sticklebacks

Human-induced processes are altering habitats at an unprecedented rate and scale. This has changed the biodiversity and biomass in many areas, but also led to phenotypic and genetic alterations of populations. Here we investigated the effects of the ongoing eutrophication in the Baltic Sea on the reproductive success of threespine stickleback males Gasterosteus aculeatus , through effects on reproductive behaviour and parenting ability. We allowed males to complete breeding cycles in a competitive setting under increased macro algae cover or increased turbidity caused by phytoplankton growth. Both environmental factors improved the parenting ability of the males and enhanced reproductive output. Increased alga growth and turbidity reduced aggressive interactions between males during the parental phase, probably due to reduced visibility, which slowed down a deterioration of condition. This increased the reproductive lifespan of the males and enabled them to complete more breeding cycles, as found when males were allowed to complete as many breeding cycles as they could under increased algae cover. In addition, increased turbidity improved oxygen conditions, which enhanced hatching success and reduced the need for vigorous fanning behaviour. Increased turbidity, however, relaxed selection on male size. Together with earlier results on relaxed sexual selection under changed environmental conditions, this suggests that the effect of eutrophication on stickleback populations is complex. It increases the reproductive output of populations, since more individuals are spawning within eutrophicated areas and their hatching success is increased, but it relaxes sexual and natural selection at the reproductive stage. Whether this will shift selection and population regulation to other life stages, such as the juvenile stage, deserves further investigations.     

Correlative Analysis of miRNA Expression and Oncotype Dx Recurrence Score in Estrogen Receptor Positive Breast Carcinomas

Altered expression of miRNAs has been observed in many types of cancer, including breast cancer, and shown to contribute to cancer growth, aggressiveness, and response to therapies. In this pilot study, we investigated the possible correlation of miRNAs with risk of recurrence of estrogen receptor positive, lymph node-negative mammary carcinomas as determined by the Oncotype DX^® Breast Cancer assay. To accomplish this, we extracted RNA from a collection of breast carcinomas that had previously been analyzed by Oncotype DX^®. Multiple Let-7 family members were negatively correlated with the recurrence score (RS), which is consistent with their tumor suppressor properties. Additional miRNAs were found to positively correlate with RS, including miR-377-5p, miR-633b, miR-548t and miR-3648. Pathway analysis of putative and validated targets suggests that these miRNAs may have a diverse range of functions that may contribute to tumor recurrence. Taken together, these findings provide evidence that a miRNA expression signature can be developed to aid existing methods to determine the risk of recurrence for women with estrogen receptor positive breast cancers treated with endocrine therapy.     

A Novel Rhabdovirus Associated with Acute Hemorrhagic Fever in Central Africa

Deep sequencing was used to discover a novel rhabdovirus (Bas-Congo virus, or BASV) associated with a 2009 outbreak of 3 human cases of acute hemorrhagic fever in Mangala village, Democratic Republic of Congo (DRC), Africa. The cases, presenting over a 3-week period, were characterized by abrupt disease onset, high fever, mucosal hemorrhage, and, in two patients, death within 3 days. BASV was detected in an acute serum sample from the lone survivor at a concentration of 1.09×10⁶ RNA copies/mL, and 98.2% of the genome was subsequently de novo assembled from ∼140 million sequence reads. Phylogenetic analysis revealed that BASV is highly divergent and shares less than 34% amino acid identity with any other rhabdovirus. High convalescent neutralizing antibody titers of >1∶1000 were detected in the survivor and an asymptomatic nurse directly caring for him, both of whom were health care workers, suggesting the potential for human-to-human transmission of BASV. The natural animal reservoir host or arthropod vector and precise mode of transmission for the virus remain unclear. BASV is an emerging human pathogen associated with acute hemorrhagic fever in Africa.     

Effect of Estuarine Wetland Degradation on Transport of Toxoplasma gondii Surrogates from Land to Sea

The flux of terrestrially derived pathogens to coastal waters presents a significant health risk to marine wildlife, as well as to humans who utilize the nearshore for recreation and seafood harvest. Anthropogenic changes in natural habitats may result in increased transmission of zoonotic pathogens to coastal waters. The objective of our work was to evaluate how human-caused alterations of coastal landscapes in California affect the transport of Toxoplasma gondii to estuarine waters. Toxoplasma gondii is a protozoan parasite that is excreted in the feces of infected felids and is thought to reach coastal waters in contaminated runoff. This zoonotic pathogen causes waterborne toxoplasmosis in humans and is a significant cause of death in threatened California sea otters. Surrogate particles that mimic the behavior of T. gondii oocysts in water were released in transport studies to evaluate if the loss of estuarine wetlands is contributing to an increased flux of oocysts into coastal waters. Compared to vegetated sites, more surrogates were recovered from unvegetated mudflat habitats, which represent degraded wetlands. Specifically, in Elkhorn Slough, where a large proportion of otters are infected with T. gondii, erosion of 36% of vegetated wetlands to mudflats may increase the flux of oocysts by more than 2 orders of magnitude. Total degradation of wetlands may result in increased Toxoplasma transport of 6 orders of magnitude or more. Destruction of wetland habitats along central coastal California may thus facilitate pathogen pollution in coastal waters with detrimental health impacts to wildlife and humans.Estuaries are recognized as being critically endangered worldwide. Pollution of estuarine waters is a significant threat to the health of aquatic life, as well as to humans who depend on coastal habitats (23). Contamination of nearshore waters with terrestrially derived, zoonotic pathogens has received little attention in the field of marine water pollution, which has primarily focused on chemical and nutrient pollutants (22, 42, 46, 55). Yet, studies have documented the presence of fecal pathogens from terrestrial animals in coastal waters and filter-feeding shellfish (7, 37, 48), as well as infections and deaths in aquatic wildlife and humans who become exposed through recreation activities or seafood (4, 18, 39). The zoonotic parasite Toxoplasma gondii is emerging as an important waterborne pathogen in both human and marine wildlife populations (2, 3, 6, 11, 15, 38). Consumption of raw oysters, clams, or mussels has recently been determined to be a risk factor for human exposure to T. gondii (24). Moreover, this parasite is an important cause of death in threatened Southern sea otters (Enhydra lutris nereis) (10, 29). Sea otter infection appears most likely to result from ingestion of environmentally resistant T. gondii oocysts that reach coastal waters in contaminated freshwater runoff (35, 36). These oocysts are shed in the feces of infected wild and domestic felids, with an individual cat capable of shedding up to 1 billion oocysts over several days postinfection (12).Elkhorn Slough, within Monterey Bay in California, is one of the high-risk sites for sea otter infection with T. gondii, with seroprevalence rates of 79% in otters sampled in this area (35). To date, the reasons for the high sea otter prevalence of infections with T. gondii at this site remain unknown. This estuarine habitat has been extensively altered by human activities and is listed as an impaired body of water by the State of California (9). Specifically, extensive degradation has been observed in the slough, with over one-third of vegetated wetlands converted to mudflats due to erosion (49). While the effect of this landscape alteration on the transport of waterborne pathogens is not currently known, such degradation may facilitate contamination of nearshore waters with T. gondii.Wetland habitats provide valuable ecosystem services, including improvement of effluent water quality characteristics through removal of a variety of pollutants (28, 50, 57). Artificially constructed wetlands are now used globally in water treatment facilities to remove nutrients, chemical pollutants, and fecal pathogens from contaminated waters before discharge into receiving water bodies (8, 17, 21, 26, 27). However, compared with freshwater and constructed wetlands, significantly less research has focused on the effects of natural, estuarine wetlands on water quality. In the few studies that investigated the impact of saltwater marshes on marine water quality, these habitats were shown to reduce concentrations of chemicals and nutrients that reach coastal waters in contaminated overland runoff (5, 51). In addition, the percentage of watershed-impervious surface coverage and reduction of natural coastal habitats due to anthropogenic changes has been associated with increased coastal water pollution (33, 34). Despite previous research suggesting a link between wetland degradation and coastal pathogen pollution (5, 33, 34, 51), the role estuarine wetlands play in the transport of terrestrial pathogens from land to sea has not been previously investigated.The overall goal of our research was to evaluate the effect of coastal wetland degradation on contamination of estuarine and coastal waters with terrestrially derived, zoonotic pathogens. Specifically, the objective of this study was to measure T. gondii oocyst transport through vegetated estuarine wetlands and nonvegetated mudflats to quantify the effect of vegetation loss on the flux of this zoonotic pathogen to coastal waters. Due to the biohazard risks associated with the release of environmentally resistant oocysts, experiments used previously validated surrogate microspheres and a specially designed flume that was deployed in vegetated and mudflat (nonvegetated) estuarine wetland habitats. The flume-in-field study design allowed for replication of experiments using specific hydrological parameters while conducting the study within a natural estuarine environment with in situ vegetation, substrate, and water. The two autofluorescent microspheres used in this study have similar physical and surface chemistry properties to T. gondii oocysts and have been previously evaluated as surrogate particles for this protozoan parasite (44). Our results provide novel insights into the consequences of changes in coastal habitat on the ecology of zoonotic infectious disease organisms in coastal marine ecosystems.