Aerial surveys of marine mammals are routinely conducted to assess and monitor species’ habitat use and population status. In Australia, dugongs (Dugong dugon) are regularly surveyed and long-term datasets have formed the basis for defining habitat of high conservation value and risk assessments of human impacts. Unmanned aerial vehicles (UAVs) may facilitate more accurate, human-risk free, and cheaper aerial surveys. We undertook the first Australian UAV survey trial in Shark Bay, western Australia. We conducted seven flights of the ScanEagle UAV, mounted with a digital SLR camera payload. During each flight, ten transects covering a 1.3 km2 area frequently used by dugongs, were flown at 500, 750 and 1000 ft. Image (photograph) capture was controlled via the Ground Control Station and the capture rate was scheduled to achieve a prescribed 10% overlap between images along transect lines. Images were manually reviewed post hoc for animals and scored according to sun glitter, Beaufort Sea state and turbidity. We captured 6243 images, 627 containing dugongs. We also identified whales, dolphins, turtles and a range of other fauna. Of all possible dugong sightings, 95% (CI = 90%, 98%) were subjectively classed as ‘certain’ (unmistakably dugongs). Neither our dugong sighting rate, nor our ability to identify dugongs with certainty, were affected by UAV altitude. Turbidity was the only environmental variable significantly affecting the dugong sighting rate. Our results suggest that UAV systems may not be limited by sea state conditions in the same manner as sightings from manned surveys. The overlap between images proved valuable for detecting animals that were masked by sun glitter in the corners of images, and identifying animals initially captured at awkward body angles. This initial trial of a basic camera system has successfully demonstrated that the ScanEagle UAV has great potential as a tool for marine mammal aerial surveys.

The 3.2-kb hepatitis B virus (HBV) genome encodes a single regulatory protein termed HBx. While multiple functions have been identified for HBx in cell culture, its role in virus replication remains undefined. In the present study, we combined an HBV plasmid-based replication assay with the hydrodynamic tail vein injection model to investigate the function(s) of HBx in vivo. Using a greater-than-unit-length HBV plasmid DNA construct (payw1.2) and a similar construct with a stop codon at position 7 of the HBx open reading frame (payw1.2*7), we showed that HBV replication in transfected HepG2 cells was reduced 65% in the absence of HBx. These plasmids were next introduced into the livers of outbred ICR mice via hydrodynamic tail vein injection. At the peak of virus replication, at 4 days postinjection, intrahepatic markers of HBV replication were reduced 72% to 83% in mice injected with HBx-deficient payw1.2*7 compared to those measured in mice receiving wild-type payw1.2. A second plasmid encoding HBx was able to restore virus replication from payw1.2*7 to wild-type levels. Finally, viremia was monitored over the course of acute virus replication, and at 4 days postinjection, it was reduced by nearly 2 logs in the absence of HBx. These studies establish that the role for HBx in virus replication previously shown in transfected HepG2 cells is also apparent in the mouse liver within the context of acute hepatitis. Importantly, the function of HBx can now be studied in an in vivo setting that more closely approximates the cellular environment for HBV replication.Hepatitis B virus (HBV) infection is a major health problem worldwide, with more than 350 million chronically infected individuals who are at risk for developing severe liver disease, including hepatocellular carcinoma (12). The 3.2-kb HBV genome encodes a single regulatory protein termed HBx, but progress towards understanding the role of HBx in virus replication has been hindered by the lack of either a cell culture system or a convenient small animal model. Although initial studies suggested that HBx was not required for virus replication in cell culture (2), experiments with the highly related woodchuck hepatitis virus (WHV) system indicate that the WHV X protein (WHx) is required for virus replication in vivo (23). Other studies have demonstrated that WHVs carrying mutant WHx grow with attenuated kinetics in vivo, depending on the particular mutation (17,22).While many functions have been ascribed to HBx (reviewed in reference 5), most studies have been performed in cell culture and under conditions in which HBx is not required for virus replication. Two recent advances provide new approaches to investigate a role for HBx in the context of virus replication in vivo. The first is a plasmid-based replication assay that utilizes a greater-than-unit-length HBV genome of the ayw subtype (payw1.2) (15) and an identical plasmid containing a stop codon affecting HBx amino acid position 7 (payw1.2*7) (10). These plasmids were used to demonstrate that capsidassoc...

Abstract. Aerial surveys are conducted for various fauna to assess abundance, distribution, and habitat use over large spatial scales. They are traditionally conducted using light aircraft with observers recording sightings in real time. Unmanned Aerial Vehicles (UAVs) offer an alternative with many potential advantages, including eliminating human risk. To be effective, this emerging platform needs to provide detection rates of animals comparable to traditional methods. UAVs can also acquire new types of information, and this new data requires a reevaluation of traditional analyses used in aerial surveys; including estimating the probability of detecting animals. We conducted 17 replicate UAV surveys of humpback whales (Megaptera novaeangliae) while simultaneously obtaining a 'census' of the population from land-based observations, to assess UAV detection probability. The ScanEagle UAV, carrying a digital SLR camera, continuously captured images (with 75% overlap) along transects covering the visual range of land-based observers. We also used ScanEagle to conduct focal follows of whale pods (n = 12, mean duration = 40 min), to assess a new method of estimating availability. A comparison of the whale detections from the UAV to the land-based census provided an estimated UAV detection probability of 0.33 (CV = 0.25; incorporating both availability and perception biases), which was not affected by environmental covariates (Beaufort sea state, glare, and cloud cover). According to our focal follows, the mean availability was 0.63 (CV = 0.37), with pods including mother/calf pairs having a higher availability (0.86, CV = 0.20) than those without (0.59, CV = 0.38). The follows also revealed (and provided a potential correction for) a downward bias in group size estimates from the UAV surveys, which resulted from asynchronous diving within whale pods, and a relatively short observation window of 9 s. We have shown that UAVs are an effective alternative to traditional methods, providing a detection probability that is within the range of previous studies for our target species. We also describe a method of assessing availability bias that represents spatial and temporal characteristics of a survey, from the same perspective as the survey platform, is benign, and provides additional data on animal behavior.

Hepatitis B virus (HBV) is a small (3.2-kb) DNA virus that causes acute and chronic inflammation of the liver, and the latter is a risk factor for the development of hepatocellular carcinoma (HCC) (39). Worldwide, an estimated 350 million people have chronic HBV and are at risk for severe liver disease (39). New insight into the virus-host interactions underlying chronic virus replication was provided with the demonstration that HBV infection fails to activate the innate immune response in chimpanzees (52). This observation was recently confirmed in acutely infected humans (10) and primary human hepatocytes exposed to HBV (17). Several studies have clearly demonstrated that HBV replication is controlled by an activated adaptive immune response (4,14,36,53,54), suggesting that HBV has evolved a strategy to dampen activation of the innate immune response.The sole HBV regulatory protein, the 17-kDa HBx protein, plays an essential role in virus replication in HepG2 cells (3) and in hydrodynamically injected mice (22). Given the abundance of properties attributed to HBx (reviewed in reference 2), it is likely that HBx has more than one function during the virus life cycle. These functions may be mediated, in part, through HBx interactions with cellular proteins. Indeed, screening protocols such as the yeast two-hybrid assay have been used to identify over 30 HBx-interacting proteins (reviewed in reference 13). The biologic importance of these virus-host protein interactions is difficult to assess due to a paucity of virus replication assays. In the related woodchuck virus replication model, it was demonstrated that the HBx interaction with cellular DDB1 is critical for virus replication (42), and this was confirmed in plasmid-transfected HepG2 cells (27). However, the role of other HBx binding partners in virus replication remains unknown. Cells respond to virus infection through the recognition of viral pathogen-associated molecular patterns (PAMPs). Several cytoplasmic host pattern recognition receptors (PRRs) are responsible for this, including the Toll-like receptors (TLRs) and retinoic acid-inducible gene I (RIG-I) and other RIG-Ilike receptors (RLRs), such as MDA-5 and LGP2 (43; also reviewed in reference 55). Following recognition of the viral DNA or RNA, the PRRs undergo conformational changes that activate downstream pathways, ultimately leading to the induction of type I interferon (IFN) and proinflammatory cytokines. A key adaptor protein in this process is the beta interferon promoter stimulator 1 (IPS-1) protein (21), also known as mitochondrial antiviral signaling protein (MAVS) (40), VISA (58), and Cardiff (35; also reviewed in reference 20). Upon its activation, IPS-1 recruits kinases that phosphorylate latent transcription factors required for the production of 21,25). Interestingly, IPS-1 is targeted for interaction by several viral proteins, effectively inactivating the innate antiviral immune response (43).The goal of the present study was to identify HBx-interacting proteins in the liver of the H...

Robust hepatitis B virus (HBV) replication is stimulated by the regulatory HBx protein. HBx binds the cellular protein DDB1; however, the importance of this interaction for HBV replication remains unknown. We tested whether HBx binding to DDB1 was required for HBV replication using a plasmid based replication assay in HepG2 cells. Three DDB1 binding-deficient HBx point mutants (HBx69, HBx90/91, HBxR96E) failed to restore wildtype levels of replication from an HBx-deficient plasmid, which established the importance of the HBx-DDB1 interaction for maximal HBV replication. Analysis of overlapping HBx truncation mutants revealed that both the HBx-DDB1 binding domain and the carboxyl region are required for maximal HBV replication both in vitro and in vivo, suggesting the HBx-DDB1 interaction recruits regulatory functions critical for replication. Finally we demonstrate that HBx localizes to the Cul4A-DDB1 complex, and discuss the possible implications for models of HBV replication.