Diagnosis and seroprevalence of leptospirosis in California sea lions from coastal California

The sensitivity and specificity of the microscopic agglutination test (MAT) as a method for detection of exposure to Leptospira spp. in California sea lions (Zalophus californianus) were determined. Sera came from individuals that demonstrated clinical signs of renal disease, had lesions suggestive of leptospirosis at necropsy, and had visible leptospires in silver stained kidney sections as positive controls. Sera from unexposed captive individuals were used as negative controls. The test was 100% sensitive at 1:3,200 for confirming renal infection and 100% specific at negative < 1:100 for detection of Leptospira interrogans scrovar pomona antibodies by MAT in California sea lions. Leptospira interrogans serovar pomona was used as a screening serovar because it has been isolated previously from the kidneys and placentas of California sea lions, and there appears to be cross-reactivity between serovar pomona and other serovars. Sera from 225 free-ranging California sea lions presented to one of three participating California (USA) coastal marine mammal rehabilitation centers in 1996 were then evaluated for antibodies to serovar pomona using the MAT. The overall seroprevalence was 38.2% (86/225), although the prevalence varied among locations from 100% (38/38) in animals at the Marine Mammal Care Center (Fort MacArthur, California, USA) to 0% (0/14) at SeaWorld California (San Diego, California). At The Marine Mammal Center (Sausalito, California) [prevalence 27.8% (48/173)], the majority of seropositive animals were subadults and adults, and males were 4.7 times more likely to be seropositive to serovar pomona than females. When combining results from all three centers, subadult and adult animals were more likely to be seropositive than pups and juvenile sea lions, and the highest proportion of seropositive animals presented during the autumn months. Serum elevations of blood urea nitrogen, creatinine, phosphorus, and/or calcium were associated with seropositivity to serovar pomona. We found no association between potassium or sodium levels and seropositivity.     

Pseudo‐nitzschia blooms,domoic acid,and related California sea lion strandings in Monterey Bay,California

Blooms of the toxin‐producing diatom Pseudo‐nitzschia commonly occur in Monterey Bay, California, resulting in sea lion mortality events. The links between strandings of California sea lions suffering from domoic acid (DA) toxicity, toxic cell numbers, and their associated DA concentration in Monterey Bay and in sea lion feces were examined from 2004 to 2007. While Pseudo‐nitzschia toxic cells and DA concentrations were detectable in the water column most of the time, they were often at low levels. A total of 82 California sea lions were found stranded in the Bay between 2004 and 2007 with acute or chronic signs associated with DA poisoning. The highest number with detectable DA in feces occurred in April 2007 and corresponded with the presence of a highly toxic bloom in the Bay. Higher DA levels occurred in feces from sea lions stranding with acute toxicosis and lower concentrations in feces of sea lions exhibiting signs of chronic DA poisoning or not exhibiting any neurologic signs. Results indicated that sea lions are likely exposed to varying levels of DA through their prey throughout the year, often at sublethal doses that may contribute to a continued increase in the development of chronic neurologic sequelae.     

Identification of two novel coccidian species shed by California sea lions (Zalophus californianus)

Routine fecal examination revealed novel coccidian oocysts in asymptomatic California sea lions (Zalophus californianus) in a rehabilitation facility. Coccidian oocysts were observed in fecal samples collected from 15 of 410 California sea lions admitted to The Marine Mammal Center between April 2007 and October 2009. Phylogenetic analysis using the full ITS-1 region, partial small subunit 18S rDNA sequence, and the Apicomplexa rpoB region identified 2 distinct sequence clades, referred to as Coccidia A and Coccidia B, and placed them in the Sarcocystidae, grouped with the tissue-cyst-forming coccidia. Both sequence clades resolved as individual taxa at ITS-1 and rpoB and were most closely related to Neospora caninum. Coccidia A was identified in 11 and Coccidia B in 4 of 12 sea lion oocyst samples successfully sequenced (3 of those sea lions were co-infected with both parasites). Shedding of Coccidia A oocysts was not associated with age class, sex, or stranding location, but yearlings represented the majority of shedders (8/15). This is the first study to use molecular phylogenetics to identify and describe coccidian parasites shed by a marine mammal.     

Rapid behavioural diagnosis of domoic acid toxicosis in California sea lions

Domoic acid is a neurotoxic metabolite of widely occurring algal blooms that has caused multiple marine animal stranding events. Exposure to high doses of domoic acid, a glutamate agonist, may lead to persistent medial temporal seizures and damage to the hippocampus. California sea lions (Zalophus californianus) are among the most visible and frequent mammalian victims of domoic acid poisoning, but rapid, reliable diagnosis in a clinical setting has proved difficult owing to the fast clearance of the toxin from the blood stream. Here, we show that the behavioural orienting responses of stranded sea lions diagnosed with domoic acid toxicosis habituate more slowly to a series of non-aversive auditory stimuli than do those of sea lions with no apparent neurological deficits. A signal detection analysis based on these habituation measures was able to correctly identify 50 per cent of subjects with domoic acid toxicosis while correctly rejecting approximately 93 per cent of controls, suggesting potential diagnostic merit.     

Leptospirosis in northern elephant seals (Mirounga angustirostris) stranded along the California coast

Leptospirosis was identified in six northern elephant seals (Mirounga angustirostris) that were stranded in 1995 along the coast of California (USA). Histologic lesions in all seals included tubulointerstitial nephritis with tubular degeneration and necrosis. Infection was confirmed through identification of spirochetes using an immunohistochemical stain for Leptospira sp. antigens. One affected seal had an elevated titer to Leptospira interrogans serovar pomona. Four of the six seals developed leptospirosis during rehabilitation, and two seals had evidence of exposure in the wild. Potential sources of infection during rehabilitation include other elephant seals, California sea lions (Zalophus californianus), Pacific harbor seals (Phoca vitulina richardsii), or free-ranging wildlife. These results indicate that northern elephant seals are susceptible to leptospirosis and can develop disease both in the natural environment and in a rehabilitation setting.     

加州Santa Monica海湾鳍足类的生态学

研究了加州Santa Monica海湾鳍足类的生态学.从1997-2007年乘船调查了277次,发现海狮(Zalophus californianus)是最常见的动物(89%,见到的次数为1393次),其次是港海豹(Phoca vitulina richardsi,8%,n=131)和北象海豹(Mirounga angustirostris,1%,n=15).在29%的遇见次数(观察到瓶海豚205次)中,发现海狮(偶尔也发现港海豹)与瓶鼻海豚集群(Tursiops truncatus);短喙真海豚(Delphinus delphis)与长喙真海豚(D.capensis)在53% 的遇见次数(遇见真海豚次数n=155)中,发现短喙真海豚(Delphinus delphis)与长喙真海豚(D.capensis)集群;一般在沿岸水域(离岸边距离<500 m)见到海狮和港海豹,但在整个海湾也能见到,表现出这两个物种对海底峡谷的偏爱.北象海豹仅见于近海,主要在海底峡谷附近. 经常看到海狮、港海豹和北象海豹游动(50%,n=728)、进行热调节(14%,n=205)、以及取食(3.2%,n=47),但几乎见不到有社会性活动(0.21%,n=3).     

Isolation of a novel adenovirus from California sea lions Zalophus californianus

Viral hepatitis associated with adenoviral infection has been reported in California sea lions Zalophus californianus admitted to rehabilitation centers along the California coast since the 1970s. Canine adenovirus 1 (CAdV-1) causes viral hepatitis in dogs and infects a number of wildlife species. Attempts to isolate the virus from previous sea lion hepatitis cases were unsuccessful, but as the hepatitis had morphologic features resembling canine infectious hepatitis, and since the virus has a wide host range, it was thought that perhaps the etiologic agent was CAdV-1. Here, we identify a novel adenovirus in 2 stranded California sea lions and associate the infection with viral hepatitis and endothelial cell infection. Phylogenetic analysis confirmed the classification of the sea lion adenovirus in the Mastadenovirus genus with the most similarity to tree shrew adenovirus 1 (TSAdV-1, 77%). However, as the sea lion adenovirus appeared to be equally distant from the other Mastadenovirus species based on phylogenetic analysis, results indicate that it represents an independent lineage and species. Although sequences from this novel virus, otarine adenovirus 1 (OtAdV-1), show some similarity to CAdV-1 and 2, it is clearly distinct and likely the cause of the viral hepatitis in the stranded California sea lions.     

Demographics and spatio‐temporal signature of the biotoxin domoic acid in California sea lion (Zalophus californianus) stranding records

California sea lions (Zalophus californianus) in otherwise good nutritional condition have been consistently affected by the marine biotoxin domoic acid since the late 1990s. In this study we evaluated the temporal and spatial stranding patterns of suspected and confirmed cases of domoic acid intoxicated sea lions from 1998 to 2006, using records of strandings along the California coast obtained from members of the California Marine Mammal Stranding Network. The majority of domoic acid cases were adult females (47%–82% of the total annual domoic acid cases), a contrast to strandings that were not related to domoic acid, which were generally dominated by juveniles and pups. Exposure to this biotoxin led to a 6.67‐fold increase in adult female strandings in 2000, and a 5.44‐fold increase in adult female deaths in 2006, relative to strandings and deaths of adult female not affected by domoic acid. Domoic acid cases have occurred annually since 1998 (except for 1999) between April and August, with clusters centered primarily at Pismo Beach (San Luis Obispo County), as well as at other beaches in San Luis Obispo, Monterey, Santa Cruz, Santa Barbara, Orange, and San Diego counties. The larger ecological and population level implications of increased domoic acid strandings and deaths, particularly among adult female sea lions, warrant further attention and need to be investigated.     

Movement, dive behavior, and survival of California sea lions (Zalophus californianus) posttreatment for domoic acid toxicosis

Domoic acid (DA) is a neuroexcitatory toxin increasingly causing strandings and mortality of marine mammals. The hippocampus of mammalian brains, associated with learning, memory, and spatial navigation, is one of the predominant regions affected by DA exposure. California sea lions stranding from 2003 to 2006 as a result of DA toxicosis were classified as having acute ( n = 12) or chronic neurologic ( n = 22) clinical signs. Chronic neurologic cases were examined by magnetic resonance imaging to determine the extent of brain damage related to DA exposure. Brain damage included hippocampal and parahippocampal atrophy, temporal horn enlargement, and pathological T2 hyperintensity. Posttreatment, animals were fitted with satellite transmitters and their movement and dive behaviors compared with those of a control group. The only significant difference between acute and chronic animals was distance traveled per day. There were, however, significant differences between chronic neurologic cases and controls: chronic neurologic cases dove shallower for shorter durations, traveled further from shore, and spent less time hauled out and more time surface swimming than control animals. There was no relationship between severity of brain damage and behavioral patterns for chronic neurologic cases. Sea lions with chronic neurologic changes had a poor prognosis for survival following release.     

Lateralised swimming behaviour in the California sea lion

Lateralised motor behaviour in the pinnipeds has been subject to little investigation. This study examined the swimming behaviour of seven zoo-housed California sea lions to determine whether they exhibited a directional bias in their motor behaviour. Data were collected on the direction of the animals' swimming patterns from the point of entering a pool of water from dry land. Each animal was studied for 100 episodes of swimming. All seven of the sea lions showed significant (P<0.001) bias in the direction of their swimming, although unidirectional bias was not observed at the level of the population. The direction of the sea lions' swimming patterns varied significantly according to the animals' sex. Males showed a preference at the level of the population for swimming in a clockwise direction, while females showed a population-level counterclockwise swimming preference. Overall, the findings appear to suggest that California sea lions, like other marine mammals, exhibit motor bias in the direction of their swimming patterns, although further work using larger sample sizes is needed before more firm conclusions regarding motor laterality in this species can be reached.     

Interannual variation in the at‐sea behavior of California sea lions (Zalophus californianus)

To be successful, marine predators must alter their foraging behavior in response to changes in their environment. To understand the impact and severity of environmental change on a population it is necessary to first describe typical foraging patterns and identify the underlying variability that exists in foraging behavior. Therefore, we characterized the at‐sea behavior of adult female California sea lions (n = 32) over three years (2003, 2004, and 2005) using satellite transmitters and time‐depth recorders and examined how foraging behavior varied among years. In all years, sea lions traveled on average 84.7 ± 11.1 km from the rookery during foraging trips that were 3.2 ± 0.3 d. Sea lions spent 42.7% ± 1.9% of their time at sea diving and displayed short (2.2 ± 0.2 min), shallow dives (58.5 ± 8.5 m). Among individuals, there was significant variation in both dive behavior and movement patterns, which was found in all years. Among years, differences were found in trip durations, distances traveled, and some dive variables (e.g., dive duration and bottom time) as sea lions faced moderate variability in their foraging habitat (increased sea‐surface temperatures, decreased upwelling, and potential decreased prey abundance). The flexibility we found in the foraging behavior of California sea lions may be a mechanism to cope with environmental variability among years and could be linked to the continuing growth of sea lion populations.     

Age‐ and sex‐specific survival of California sea lions (Zalophus californianus) at San Miguel Island,California

We conducted a mark‐recapture study of California sea lions (Zalophus californianus) using pups branded on San Miguel Island, California, from 1987 to 2014, and annual resightings from 1990 to 2015. We used the Burnham model (Burnham 1993), an extension of the Cormack‐Jolly‐Seber mark‐recapture model, which includes recoveries of dead animals, to analyze age, sex, and annual patterns in survival. Generally, females had higher survival than males. For female pups, the average annual survival was 0.600 and for male pups it was 0.574. Yearling survival was 0.758 and 0.757 for females and males, respectively. Peak annual survival was at age 5 and was 0.952 for females and 0.931 for males. Pups with larger mass at branding had higher survival as pups and yearlings, but the effect was relative within each cohort because of large between‐cohort variability in survival. Annual variability in sea surface temperature (SST) affected survival. For each 1°C increase in SST, the odds of survival decreased by nearly 50% for pups and yearlings; negative SST anomalies yielded higher survival. Annual variation in male survival was partly explained by leptospirosis outbreaks. Our study provides a unique view of one demographic parameter that contributed to the successful recovery of the California sea lion population.     

The fecal viral flora of California sea lions

California sea lions are one of the major marine mammal species along the Pacific coast of North America. Sea lions are susceptible to a wide variety of viruses, some of which can be transmitted to or from terrestrial mammals. Using an unbiased viral metagenomic approach, we surveyed the fecal virome in California sea lions of different ages and health statuses. Averages of 1.6 and 2.5 distinct mammalian viral species were shed by pups and juvenile sea lions, respectively. Previously undescribed mammalian viruses from four RNA virus families (Astroviridae, Picornaviridae, Caliciviridae, and Reoviridae) and one DNA virus family (Parvoviridae) were characterized. The first complete or partial genomes of sapeloviruses, sapoviruses, noroviruses, and bocavirus in marine mammals are reported. Astroviruses and bocaviruses showed the highest prevalence and abundance in California sea lion feces. The diversity of bacteriophages was higher in unweaned sea lion pups than in juveniles and animals in rehabilitation, where the phage community consisted largely of phages related to the family Microviridae. This study increases our understanding of the viral diversity in marine mammals, highlights the high rate of enteric viral infections in these highly social carnivores, and may be used as a baseline viral survey for comparison with samples from California sea lions during unexplained disease outbreaks.     

An unusual genotype of Toxoplasma gondii is common in California sea otters (Enhydra lutris nereis) and is a cause of mortality

Toxoplasma gondii-associated meningoencephalitis is a significant disease of California sea otters (Enhydra lutris nereis), responsible for 16% of total mortality in fresh, beachcast carcasses. Toxoplasma gondii isolates were obtained from 35 California otters necropsied between 1998 and 2002. Based on multi-locus PCR-restriction fragment length polymorphism and DNA sequencing at conserved genes (18S rDNA, ITS-1) and polymorphic genes (B1, SAG1, SAG3 and GRA6), two distinct genotypes were identified: type II and a novel genotype, here called type x, that possessed distinct alleles at three of the four polymorphic loci sequenced. The majority (60%) of sea otter T. gondii infections were of genotype x, with the remaining 40% being of genotype II. No type I or III genotypes were identified. Epidemiological methods were used to examine the relationship between isolated T. gondii genotype(s) and spatial and demographic risk factors, such as otter stranding location and sex, as well as specific outcomes related to pathogenicity, such as severity of brain inflammation on histopathology and T. gondii-associated mortality. Differences were identified with respect to T. gondii genotype and sea otter sex and stranding location along the California coast. Localised spatial clustering was detected for both type II (centred within Monterey Bay) and x (centred near Morro Bay)-infected otters. The Morro Bay cluster of type x-infected otters overlaps previously reported high-risk areas for sea otter infection and mortality due to T. gondii. Nine of the 12 otters that had T. gondii-associated meningoencephalitis as a primary cause of death were infected with type x parasites.     

First findings of Cryptosporidium and Giardia in California sea lions (Zalophus californianus)

We report the detection and identification of Cryptosporidium and Giardia from 1 of 3 species of pinnipeds. Fecal samples were collected from Pacific harbor seal (Phoca vitulina richardsi), northern elephant seal (Mirounga angustirostris), and California sea lion (Zalophus californianus) in the northern California coastal area. By means of fluorescently labeled monoclonal antibodies, Cryptosporidium oocysts were detected in 3 samples from California sea lions, 1 of which also contained Giardia cysts. Oocysts of Cryptosporidium and cysts of Giardia were morphologically indistinguishable from oocysts of C. parvum and cysts of G. duodenalis from other animal origins. Oocysts and cysts were then purified using immunomagnetic separation techniques and identified by polymerase chain reaction (PCR), from which species-specific products were obtained. Sequence analysis revealed that the 452-bp and 358-bp PCR products of Cryptosporidium isolated from California sea lion had identities of 98% with sequences of their template fragments of C. parvum obtained from infected calves. Based on morphological, immunological, and genetic characterization, the isolates were identified as C. parvum and G. duodenalis, respectively. The findings suggested that California sea lions could serve as reservoirs in the environmental transmission of Cryptosporidium and Giardia.     

Monitoring glucocorticoid response to rehabilitation and research procedures in California and Steller sea lions

We used serum and fecal corticosteroid analysis to study the physiological response to a range of invasive and non-invasive procedures in sea lions. Four experimental groups of California sea lions (Zalophus californianus; Group A: restraint only [n=9], Group B: gas anesthesia without surgery [n=10], Group C: minimally invasive surgery [n=10], and Group D: invasive surgery [n=5]) were monitored for adrenal response. Feces were collected opportunistically from 72 hr before 72 hr post procedure for corticosterone analysis. All experimental groups showed substantial individual variation and no significant change in corticosterone levels after the procedures. Additional fecal and serum corticoid samples were collected from six free-ranging Steller sea lions (Eumetopias jubatus) in temporary captivity undergoing abdominal implantation of long-term telemetry devices. Only one sea lion exhibited a significant fecal corticosterone increase in response to the surgery. Capture and restraint appear to elicit a greater glucocorticoid response than invasive or non-invasive procedures. J. Exp. Zool. 309A:73-82, 2008. (c) 2008 Wiley-Liss, Inc.     

Isolation by distance among California sea lion populations in Mexico: redefining management stocks

Understanding the spatial structure of a population is critical for effective assessment and management. However, direct observation of spatial dynamics is generally difficult, particularly for marine mammals. California sea lions ( Zalophus californianus ) are polygynous pinnipeds distributed along the Pacific coast of North America. The species' range has been subdivided into three management stocks based on differences in mitochondrial DNA, but to date no studies have considered nuclear genetic variation, and thus we lack a comprehensive understanding of gene flow patterns among sea lion colonies. In light of recent population declines in the Gulf of California, Mexico, it is important to understand spatial structure to determine if declining sea lion colonies are genetically isolated from others. To define population subdivision and identify sex biases in gene flow, we analysed a 355-bp sequence of the mitochondrial DNA control region and 10 polymorphic microsatellite loci from 355 tissue samples collected from six colonies distributed along Mexican waters. Using a novel approach to estimate sex biases in gene flow, we found that male sea lions disperse on average 6.75 times more frequently than females. Analyses of population subdivision strongly suggest a pattern of isolation by distance among colonies and challenge current stock definitions. Based on these results, we propose an alternative classification that identifies three Mexican management units: Upper Gulf of California, Southern Baja Peninsula, and Upper Pacific Coast of Baja. This revised classification should be considered in future assessment and management of California sea lion populations in Mexican waters.     

Ancient DNA analysis of the Japanese sea lion (Zalophus californianus japonicus Peters, 1866): preliminary results using mitochondrial control-region sequences

In this study, we successfully extracted ancient DNA from skeletal remains of the Japanese sea lion-a species that is practically extinct-from archaeological sites and determined a partial sequence of its mitochondrial DNA control region. A molecular phylogenetic tree constructed by the neighbor-joining (NJ) method showed that the sequences from Japanese sea lions clustered together, with a high bootstrap value, and that this cluster was closest to the California sea lion cluster. The distinctly divergent cluster of Japanese sea lions reflected the morphological classification of these animals as a distinct species of the genus Zalophus; however, proximity to the California sea lion cluster simultaneously implied conformation with the traditional classification of these animals as a subspecies of Zalophus californianus. The average amount of nucleotide substitution between the Japanese and California sea lions was 7.02%. The Japanese and California sea lions were estimated to have diverged 2.2 million years ago, i.e., in the late Pliocene Epoch. This is the first report on a genetic analysis of the Japanese sea lion.     

Evidence of Leptospira interrogans infection in California sea lion pups from the Gulf of California

Forty-two urine and 96 blood and serum samples were obtained from California sea lion (Zalophus californianus) pups from the Gulf of California during the 2000 reproductive season. Antibody prevalence to 13 serovars of Leptospira interrogans was determined by microagglutination tests (MAT); presence of pathogenic leptospires was detected by polymerase chain reaction (PCR). Samples with antibody titers > or = 1:25 or 115 bp fragments on ethidium bromidestained 1.5% agarose gels were considered positive. Antibody prevalence was 54% overall with highest prevalence against serovar cynopteri (50% of all positive reactions). Highest antibody titers (1:50) were detected against serovars cynopteri and pomona. Polymerase chain reaction products were observed in two of 42 urine samples, six of 96 blood samples, and one of 96 serum samples. Presence of PCR products in blood and serum was demonstrated in pups that were seronegative. Kruskall-Wallis tests and corresponding post hoc Tukey tests (alpha = 0.05) showed that prevalence of leptospirosis was significantly different among all rookeries. The high seroprevalence (54%), low antibody titers (maximum 1:50), absence of pups showing clinical signs indicative of the disease, and lack of recent reports of increased mortality of sea lions in the Gulf of California are suggestive of the presence of enzootic host-adapted serovars. Crowding in rookeries as well as the presence of bats and rodents on some of the islands may explain infection by L. interrogans (sensu lato) and some of the differences in seroprevalence among reproductive rookeries.     

Aerial hearing thresholds and detection of hearing loss in male California sea lions (Zalophus californianus) using auditory evoked potentials

Auditory evoked potential (AEP) measurements are useful for describing the variability of hearing among individuals in marine mammal populations, an important consideration in terms of basic biology and the design of noise mitigation criteria. In this study, hearing thresholds were measured for 16 male California sea lions at frequencies ranging from 0.5 to 32 kHz using the auditory steady state‐response (ASSR), a frequency‐specific AEP. Audiograms for most sea lions were grossly similar to previously reported psychophysical data in that hearing sensitivity increased with increasing frequency up to a steep reduction in sensitivity between 16 and 32 kHz. Average thresholds were not different from AEP thresholds previously reported for male and female California sea lions. Two sea lions from the current study exhibited abnormal audiograms: a 26‐yr‐old sea lion had impaired hearing with a high‐frequency hearing limit (HFHL) between 8 and 16 kHz, and an 8‐yr‐old sea lion displayed elevated thresholds across most tested frequencies. The auditory brainstem responses (ABRs) for these two individuals and an additional 26‐yr‐old sea lion were aberrant compared to those of other sea lions. Hearing loss may have fitness implications for sea lions that rely on sound during foraging and reproductive activities.