Poaching and illegal trade of Danube sturgeons.

A market study on sturgeon products in the Lower Danube countries (Bulgaria, Romania, Serbia and Ukraine) resulted in a total of 149 samples of caviar and meat, which we subjected to genetic-isotope analyses. The samples comprised 31 samples (21%) of illegal wild-caught origin, 17 samples (11.4%) sold in violation of CITES and EU regulations, and 47 cases (32%) of consumer deception. Although poaching and illegal wildlife trade are often considered a problem in developing countries, these findings bear evidence that a high ratio of poached sturgeon products originates from EU and accession candidate states. Therefore, caviar and sturgeon trade urgently needs improvement to ensure that sturgeon populations will have a future.

The Irrawaddy dolphin, Orcaella brevirostris from the Mekong river Cambodia: Preliminary health and toxicological investigations.

The subpopulation of the Irrawaddy dolphin (Orcaella brevirostris) living in the Mekong River, Cambodia, is considered to be critically endangered. The aim of the investigation was to gain information about the genetic variation, health status and exposure to toxic compounds of these dolphins. Tissue samples from 27 Irrawaddy river dolphins found dead along the Mekong River between 2004 and 2009 were analysed with regards to genetics, pathology and ecotoxicology. Genetic maternal lineage detection, based on polymorphisms of the mitochondrial d-loop sequences, was performed. Data indicate a genetic separation of the Mekong dolphins from both the coastal population and the Mahakam dolphins. Pathological investigations revealed acute moderate multifocal suppurative bronchopneumonia, moderate periportal hepatic lipidosis, moderate diffuse hepatic atrophy and acute severe diffuse suppurative leptomeningitis. Residue levels of organochlorines and polybrominated diphenyl ethers in Irrawaddy dolphins from the Mekong River were lower than the concentrations reported for other cetaceans in the coastal and riverine waters of Asia, except for Dichlorodiphenyltrichloroethane. A high percentage of organic mercury compared to the immuno-toxic methylmercury was observed. Due to numerous confounding factors, it is not possible to relate levels of pollutants to observed morphological lesions. However, it is likely that chemical contaminants do adversely impact on the health of the Irrawaddy dolphins at present, and have also affected previous generations.

An Agouti-Signaling-Protein Mutation is Strongly Associated with Melanism in European Roe Deer (Capreolus capreolus).

Although the European roe deer (Capreolus capreolus) population of North-West Germany has a remarkable number of melanistic specimens between 10% and 25%, the underlying genetic mutation-causing melanism is still unknown. We used a gene targeting approach focusing on MC1R and ASIP as important genes of coat coloration. Overall, 1384 bp of MC1R and 2039 bp of ASIP were sequenced in 24 specimens and several SNPs were detected. But only the ASIP-SNP c.33G>T completely segregated both phenotypes leading to the amino acid substitution p.Leu11Phe. The SNP was further evaluated in additional 471 samples. Generally, all black specimens (n = 33) were homozygous TT, whereas chestnut individuals were either homozygote GG (n = 436) or heterozygote GT (n = 26). Considering the fact that all melanistic animals shared two mutated alleles of the strongly associated SNP, we concluded that melanism is inherited in a recessive mode in European roe deer.

Demographic collapse and low genetic diversity of the Irrawaddy dolphin population inhabiting the Mekong River.

In threatened wildlife populations, it is important to determine whether observed low genetic diversity may be due to recent anthropogenic pressure or the consequence of historic events. Historical size of the Irrawaddy dolphin (Orcaella brevirostris) population inhabiting the Mekong River is unknown and there is significant concern for long-term survival of the remaining population as a result of low abundance, slow reproduction rate, high neonatal mortality, and continuing anthropogenic threats. We investigated population structure and reconstructed the demographic history based on 60 Irrawaddy dolphins samples collected between 2001 and 2009. The phylogenetic analysis indicated reciprocal monophyly of Mekong River Orcaella haplotypes with respect to haplotypes from other populations, suggesting long-standing isolation of the Mekong dolphin population from other Orcaella populations. We found that at least 85% of all individuals in the two main study areas: Kratie and Stung Treng, bore the same mitochondrial haplotype. Out of the 21 microsatellite loci tested, only ten were polymorphic and exhibited very low levels of genetic diversity. Both individual and frequency-based approaches suggest very low and non-significant genetic differentiation of the Mekong dolphin population. Evidence for recent bottlenecks was equivocal. Some results suggested a recent exponential decline in the Mekong dolphin population, with the current size being only 5.2% of the ancestral population. In order for the Mekong dolphin population to have any potential for long-term survival, it is imperative that management priorities focus on preventing any further population fragmentation or genetic loss, reducing or eliminating anthropogenic threats, and promoting connectivity between all subpopulations.

The Value of Molecular vs. Morphometric and Acoustic Information for Species Identification Using Sympatric Molossid Bats.

A fundamental condition for any work with free-ranging animals is correct species identification. However, in case of bats, information on local species assemblies is frequently limited especially in regions with high biodiversity such as the Neotropics. The bat genus Molossus is a typical example of this, with morphologically similar species often occurring in sympatry. We used a multi-method approach based on molecular, morphometric and acoustic information collected from 962 individuals of Molossus bondae, M. coibensis, and M. molossus captured in Panama. We distinguished M. bondae based on size and pelage coloration. We identified two robust species clusters composed of M. molossus and M. coibensis based on 18 microsatellite markers but also on a more stringently determined set of four markers. Phylogenetic reconstructions using the mitochondrial gene co1 (DNA barcode) were used to diagnose these microsatellite clusters as M. molossus and M. coibensis. To differentiate species, morphological information was only reliable when forearm length and body mass were combined in a linear discriminant function (95.9% correctly identified individuals). When looking in more detail at M. molossus and M. coibensis, only four out of 13 wing parameters were informative for species differentiation, with M. coibensis showing lower values for hand wing area and hand wing length and higher values for wing loading. Acoustic recordings after release required categorization of calls into types, yielding only two informative subsets: approach calls and two-toned search calls. Our data emphasizes the importance of combining morphological traits and independent genetic data to inform the best choice and combination of discriminatory information used in the field. Because parameters can vary geographically, the multi-method approach may need to be adjusted to local species assemblies and populations to be entirely informative.

Tracing the maternal roots of the domestic Red Mountain Cattle.

The Red Mountain Cattle (RHV) is an important native ancient breed from the lower mountain ranges of Central Europe, which was originally raised for milk and meat production and as draught animal. In the 1980s, the RHV was close to extinction and only the sperm of a single purebreed bull and a few cows were available for breed formation. In this study the mitogenomes were sequenced of RHV from six maternal founder lineages. We observed six novel mitogenomes which have not been found in any other cattle breed so far. The RHV mitogenomes are grouped phylogenetically in the T-haplogroup indicating a South European origin and supporting their primitive position within the taurine breeds.

Fertility of a spontaneous hexaploid male Siberian sturgeon, Acipenser baerii.

BACKGROUND: Evolution of sturgeons and paddlefishes (order Acipenseriformes) is inherently connected with polyploidization events which resulted in differentiation of ploidy levels and chromosome numbers of present acipenseriform species. Moreover, allopolyploidization as well as autopolyploidization seems to be an ongoing process in these fishes and individuals with abnormal ploidy levels were occasionally observed within sturgeon populations. Here, we reported occurrence of Siberian sturgeon (Acipenser baerii) male with abnormal ploidy level for this species, accessed its ploidy level and chromosome number and investigate its potential sterility or fertility in comparison with normal individuals of sterlet (A. ruthenus), Russian sturgeon (A. gueldenstaedtii) and Siberian sturgeon (A. baerii). RESULTS: Acipenser ruthenus possessed 120 chromosomes, exhibiting recent diploidy (2n), A. gueldenstaedtii and A. baerii had ~245 chromosomes representing recent tetraploidy (4n), and A. baerii male with abnormal ploidy level had ~ 368 chromosomes, indicating recent hexaploidy (6n). Genealogy assessed from the mtDNA control region did not reveal genome markers of other sturgeon species and this individual was supposed to originate from spontaneous 1.5 fold increment in number of chromosome sets with respect to the number most frequently found in nature for this species. Following hormone stimulation, the spontaneous hexaploid male produced normal sperm with ability for fertilization. Fertilization of A. baerii and A. gueldenstaedtii ova from normal 4n level females with sperm of the hexaploid male produced viable, non-malformed pentaploid (5n) progeny with a ploidy level intermediate to those of the parents. CONCLUSION: This study firstly described occurrence of hexaploid individual of A. baerii and confirmed its autopolyploid origin. In addition to that, the first detailed evidence about fertility of spontaneous hexaploid sturgeon was provided. If 1.5 fold increment in number of chromosome sets occurring in diploids, resulted triploids possess odd number of chromosome sets causing their sterility or subfertility due to interference of gametogenesis. In contrast, 1.5 fold increment in number of chromosome sets in naturally tetraploid A. baerii resulted in even number of chromosome sets and therefore in fertility of the hexaploid specimen under study.

Personality assessment and its association with genetic factors in captive Asian and African elephants.

Elephants live in a complex society based on matrilineal groups. Management of captive elephants is difficult, partly because each elephant has a unique personality. For a better understanding of elephant well being in captivity, it would be helpful to systematically evaluate elephants' personalities and their underlying biological basis. We sent elephant' personality questionnaires to keepers of 75 elephants. We also used 196 elephant DNA samples to search for genetic polymorphisms in genes expressed in the brain that have been suggested to be related to personality traits. Three genes, androgen receptor (AR), fragile X related mental retardation protein interacting protein (NUFIP2), and acheate-scute homologs 1 (ASH1) contained polymorphic regions. We examined the association of personality with intraspecific genetic variation in 17 Asian and 28 African elephants. The results suggest that the ASH1 genotype was associated with neuroticism in Asian elephants. Subjects with short alleles had lower scores of neuroticism than those with long alleles. This is the first report of an association between a genetic polymorphism and personality in elephants.

Experimental superinfection of a Lesser Malayan mousedeer (Tragulus javanicus) persistently infected with bovine viral diarrhea virus.

A Lesser Malayan mousedeer (Tragulus javanicus), persistently infected with noncytopathogenic bovine viral diarrhea virus (BVDV) type 1f, was experimentally superinfected with a cytopathogenic isolate of BVDV type 1c, which antigenically partially matched the endogenous strain. Within the observational period of 125 days after superinfection, the animal did not demonstrate any clinical signs of the disease and/or significant changes in blood values. Neutralizing antibodies were detected at 35 and 42 days postinfection. The isolate causing the superinfection was found in feces, nasal swabs, and saliva starting from day 29 and at various times postchallenge. Macroscopic or histologic examination did not reveal mucosal disease-like lesions, despite the detection of the cytopathogenic isolate in the salivary gland, rumen, abomasum, kidney, and superficial prescapular lymph node. Results indicate that the cytopathogenic BVDV strain, which was used in the superinfection, persisted in the viremic animal without causing disease within the observation period.

A Hepatozoon species genetically distinct from H. canis infecting spotted hyenas in the Serengeti ecosystem, Tanzania.

Health monitoring of spotted hyenas (Crocuta crocuta) in the Serengeti ecosystem, Tanzania, revealed Hepatozoon infection in all of 11 immature individuals examined following death from natural causes. Hepatozoon infection was probably an important factor contributing to mortality in two cases that exhibited clinical signs of ataxia, lethargy, ocular discharge, retching, and labored breathing before death. Whether Hepatozoon infection contributed to six deaths from fire, probable lion predation and unknown causes could not be determined. Four deaths from infanticide and starvation were unlikely to be associated with Hepatozoon infection. Histologic examination revealed lung tissue infected with cyst-like structures containing protozoan stages in all eight cases examined and interstitial pneumonia in most cases. Systemic spread of infection to several organs was found in three cases. Alignment of a 426 bp sequence from the parasite's 18s rRNA gene revealed a Hepatozoon species identical to that recently described from two domestic cats in Spain and only 7 bp substitutions when a 853 bp sequence was aligned to this cat Hepatozoon species. Previous reports of infection of wild carnivores in eastern and southern Africa with an unspecified Hepatozoon species similar in appearance to H. canis may have involved the species described in this study.

Detection of Equid herpesvirus 9 DNA in the trigeminal ganglia of a Burchell's zebra from the Serengeti ecosystem.

Equid herpesvirus 9 (EHV-9) was isolated from a herd of Thomson's gazelles affected by encephalitis. The natural host of EHV-9 is unknown, but zebras are suspected to be the source of infection in gazelles. To prove this hypothesis, we analyzed 43 sera from Burchell's zebras (Equus burchelli) and 21 Thomson's gazelles (Gazella thomsoni) from the Serengeti ecosystem for neutralizing antibodies. Seven zebra sera were positive for EHV-1, EHV-9 and EHV-1 from Grevy's zebra strains T965 and T616. The trigeminal ganglia of 17 other Burchell's zebras and one Thomson's gazelle were tested by EHV-9 gB and EHV-1 ICP0-specific nested polymerase chain reaction (PCR). PCR sequencing confirmed that one zebra ganglion was positive for EHV-9. These results suggest that the Burchell's zebras were exposed to EHV-9 and latently infected.

New variants of European brown hare syndrome virus strains in free-ranging European brown hares (Lepus europaeus) from Slovakia.

Investigations regarding European brown hare syndrome virus (EBHSV) in European brown hares (Lepus europaeus) in Slovakia were undertaken in order to detect the possible presence of EBHSV and to evaluate its phylogenetic position. Liver and/or serum samples were obtained from 135 European brown hares shot by hunters in eight regional hunting areas. From 36 animals corresponding liver and serum samples were available; from the remaining 49 and 50 animals only liver or serum samples were examined, respectively. Samples were tested for antibodies against EBHSV and for viral RNA by reverse transcriptase-polymerase chain reaction (RT-PCR) and RT-PCR products were subsequently sequenced. Additionally, matrilinear hare haplotypes were analyzed in order to detect potential familial susceptibility to EBHSV. Sixty-three of 86 sera contained antibodies against EBHSV, whereas 15 of 85 liver samples were PCR positive. Of the latter, 14 were sequenced, revealing three new strains of EBHSV. Fifteen different matrilinear haplotypes were identified, but no correlation was found between haplotype and susceptibility to EBHSV infection. Our findings confirmed the existence of EBHSV in Slovakia and reinforce the need for determining EBHSV status when importing hares for restocking.

Characterization of equid herpesvirus 1 (EHV-1) related viruses from captive Grevy's zebra and blackbuck.

Equid herpes virus 1 (EHV-1) related isolates from a captive blackbuck (strain Ro-1) and Grevy's zebra (strain T965) behaved similarly to EHV-1 and EHV-9 in respect to their host cell range. Restriction enzyme analysis and a phylogenetic tree confirmed that Ro-1 and T965 were identical and more closely related to EHV-1 than to EHV-9. Differences from EHV-1 became obvious firstly, by amino acid alignments revealing two unique substitutions in the gB protein of Ro-1 and T965. Secondly, an EHV-1 type-specific monoclonal antibody did not detect its antigen on Ro-1, T965 or EHV-9 infected cells by immunohistochemistry. The results support the view that Ro-1 and T965 isolates represent a distinct, previously unrecognized species of equid herpesviruses.

Severe Streptococcus infection in spotted hyenas in the Ngorongoro Crater, Tanzania.

In a population of spotted hyenas (Crocuta crocuta) monitored between 1996 and 2005 in the Ngorongoro Crater, Tanzania, 16 individuals from five of eight social groups displayed clinical signs of an infection, including severe unilateral swelling of the head followed by abscess formation at the mandibular angle, respiratory distress, mild ataxia, and lethargy. Two (12.5%) of these 16 individuals died within days of developing signs. Clinical signs in hyenas were first noted in 2001, and most cases occurred between September 2002 and February 2003, suggesting an outbreak of infection during this period. Histopathological examination of internal organs from one hyena that died with signs revealed morphological changes consistent with severe bacterial infection. Phenotypic examination and phylogenetic analysis of the 16S rRNA gene of the causative agent of infection revealed a Lancefield group C Streptococcus with a high level of homology to S. equi subsp. ruminatorum, a subspecies of S. equi recently described in domestic sheep (Ovis aries) and goats (Capra hircus) with mastitis in Spain. Strains similar to this bacterium were also isolated from two hyenas without obvious clinical signs, suggesting that hyenas may be 'carriers' of this bacterium, and from a sympatric Burchell's zebra (Equus burchelli), a herbivore species often consumed by hyenas. To our knowledge this is the first report of a Streptococcus infection in these two wildlife species. The high genetic similarity between the hyena and zebra isolates indicates that inter-specific transmission may occur, possibly when hyenas consume infected zebra carcasses.

Phylogenetic position of the Dalmatian genus Phoxinellus and description of the newly proposed genus Delminichthys (Teleostei: Cyprinidae).

The Dalmatian cyprinid genus Phoxinellus is characterized by reductive characters most likely associated with the environmental conditions of small karstic streams, where all species of this genus occur. Based on 33 morphological traits, nuclear and mtDNA sequences Phoxinellus was found to be paraphyletic and included three not closely related monophyletic units. The scientific name Phoxinellus should therefore be restricted to species having plain coloration, small or absent postcleithrum, no genital papilla and an almost entirely naked body such as P. alepidotus, P. dalmaticus, and P. pseudalepidotus. Species that also have a small or absent postcleithrum and no genital papilla but display a dark stripe from the head to the caudal peduncle, and are entirely covered by distinct, not overlapping scales should be positioned closely to Telestes. Thus, we suggest inclusion of Phoxinellus croaticus, P. fontinalis and Paraphoxinus metohiensis in the genus Telestes. The Phoxinellus species that have a irregularly spotted color pattern, a large postcleithrum, an increased number of precaudal anal-fin pterygiophores, and a large genital papilla in females represent its own evolutionary line closely related to the Balkan species of Pseudophoxinus. For this monophyletic group, we propose to introduce a new genus: Delminichthys. This genus includes the species D. adspersus, D. ghetaldii, D. krbavensis and D. jadovensis.

Detection of a newly described pestivirus of Pyrenean chamois (Rupicapra pyrenaica pyrenaica) in France.

A pestivirus was detected and characterized in chamois (Rupicapra pyrenaica pyrenaica) originating from the French part of the Pyrenees. Phylogenetic analysis of the pestivirus was done on the basis of a fragment from the 5' noncoding region including 22 published nucleotide sequences of different pestivirus strains. Our strain was grouped within the clade of border disease viruses (BDV). However, it had an intermediate position between clade BDV and classical swine fever viruses representing a basal position to BDV strains of domestic sheep. Our strain was grouped as a sister unit to a novel pestivirus (Chamois-1) recently described from chamois in Spain. Therefore, we postulate that this virus occurs in the entire population of Pyrenean chamois. On the basis of the phylogenetic grouping of this isolate, a postulated cross-species transmission of pestivirus from domestic sheep to chamois via shared pastures seems to be unlikely.

Molecules and morphology: evidence for introgression of mitochondrial DNA in Dalmatian cyprinids.

In one population of Scardinius dergle, mitochondrial DNA was observed originating from Squalius tenellus. Scardinius dergle shared all diagnostic morphological characters and similarities at a highly variable nuclear region with the genus Scardinius. While crosses and backcrosses most likely resulted in the loss of morphologically diagnostic Squalius-features, maternal inheritance of mtDNA fixed their diagnostic substitutions. Anthropogenic influences resulting from dam building in 1962 are suspected to be the initial force for the hybridization events. However, because hybridization took place despite both lineages being only very distantly related (p=11.2%), we conclude that introgressive hybridization events can be seen as generally possible among leuciscine cyprinids.

When the American sea sturgeon swam east.

The two species of Atlantic sea sturgeon on either shore of the North Atlantic, Acipenser sturio in Europe and A. oxyrinchus in North America, probably diverged with the closure of the Tethys Sea and the onset of the North Atlantic Gyre 15-20 million years ago, and contact between them was then presumably precluded by geographic distance. Here we present genetic, morphological and archaeological evidence indicating that the North American sturgeon colonized the Baltic during the Middle Ages and replaced the native sturgeon there, before recently becoming extinct itself in Europe as a result of human activities. In addition to representing a unique transatlantic colonization event by a fish that swims upriver to spawn, our findings have important implications for projects aimed at restocking Baltic waters with the European sturgeon.

Phylogenetic relationships and ancestral areas of the bustards (Gruiformes: Otididae), inferred from mitochondrial DNA and nuclear intron sequences.

The taxonomy of the bustards is poorly understood phylogenetically and has not been extensively evaluated using molecular methods. We sequenced part of the mitochondrial cytochrome b gene, the control region (central domain II), and an intron-exon crossing fragment of the nuclear chromo-helicase-DNA binding gene (CHD1) in 27 bustard taxa (including multiple subspecies) representing 11 genera and four gruiform outgroup species. Molecular datings suggest a Miocene origin for the family. Inferred phylogenetic relationships include the following: (i) the basal polytomy consists of 10 branches (mostly consistent with traditional genera), suggesting a rapid early radiation; (ii) sister relationships between several couplets of genera include Ardeotis with Neotis, Afrotis with Eupodotis (excluding E. rueppellii), Otis with Chlamydotis, and Houbaropsis with Sypheotides; (iii) the genus Eupodotis may be polyphyletic; and (iv) the currently delimited genera Ardeotis and Neotis do not form independent monophyletic lineages. Molecular evidence for the Afro-tropical origin of the Otididae is provided.

An exceptional case of historical outbreeding in African sable antelope populations.

Empirical investigations of intraspecific outbreeding and subsequent introgressive hybridization in natural populations are rare, particularly among conspecific populations of large mammals. Using mitochondrial DNA data [partial control region (496 basepairs - bp) and cytochrome b gene (343 bp) sequences analysed from 95 individuals representing 17 sampling locations scattered through the African miombo (Brachystegia) woodland ecosystem] and phylogeographical statistical procedures (gene genealogy, nested cladistic and admixture proportion analyses), we (i) give a detailed dissection of the geographical genetic structure of Hippotragus niger; (ii) infer the processes and events potentially involved in the population history; and (iii) trace extensive introgressive hybridization in the species. The present-day sable antelope population shows a tripartite pattern of genetic subdivision representing West Tanzanian, Kenya/East Tanzanian and Southern Africa locations. Nested clade analysis revealed that past allopatric fragmentation, caused probably by habitat discontinuities associated with the East African Rift Valley system, together with intermediary episodic long-distance colonization and restricted, recurrent gene flow have played an predominant role in shaping the extent of maternal genetic diversity (10.4%) and population structure. An extensive (average rate of admixture = 20.0%), but geographically circumscribed and unidirectional hybridization event in the past was inferred, resulting in an extreme (the highest discovered so far in mammals) intraspecific difference of 18.2% among morphologically monotypic sable antelopes from West Tanzania. The results are used to provide an evolutionary framework within which taxonomic implications and conservation decisions can be evaluated.