Characterization of highly informative cross-species microsatellite panels for the Australian dugong (Dugong dugon) and Florida manatee (Trichechus manatus latirostris) including five novel primers

The Australian dugong (Dugong dugon) and Florida manatee (Trichechus manatus latirostris) are threatened species of aquatic mammals in the order Sirenia. Sirenian conservation and management actions would benefit from a more complete understanding of genetic diversity and population structure. Generally, species-specific microsatellite markers are employed in conservation genetic studies; however, robust markers can be difficult and costly to isolate. To increase the number of available markers, dugong and manatee microsatellite primers were evaluated for cross-species amplification. Furthermore, one manatee and four dugong novel primers are reported. After polymerase chain reaction optimization, 23 (92%) manatee primers successfully amplified dugong DNA, of which 11 (48%) were polymorphic. Of the 32 dugong primers tested, 27 (84%) yielded product in the manatee, of which 17 (63%) were polymorphic. Dugong and manatee primers were compared and the most informative markers were selected to create robust and informative marker-panels for each species. These cross-species microsatellite marker-panels can be employed to assess other sirenian populations and can provide beneficial information for the protection and management of these unique mammals.

Henipaviruses: Emerging Paramyxoviruses Associated with Fruit Bats

Two related, novel, zoonotic paramyxoviruses have been described recently. Hendra virus was first reported in horses and thence humans in Australia in 1994; Nipah virus was first reported in pigs and thence humans in Malaysia in 1998. Human cases of Nipah virus infection, apparently unassociated with infection in livestock, have been reported in Bangladesh since 2001. Species of fruit bats (genus Pteropus ) have been identified as natural hosts of both agents. Anthropogenic changes (habitat loss, hunting) that have impacted the population dynamics of Pteropus species across much of their range are hypothesised to have facilitated emergence. Current strategies for the management of henipaviruses are directed at minimising contact with the natural hosts, monitoring identified intermediate hosts, improving biosecurity on farms, and better disease recognition and diagnosis. Investigation of the emergence and ecology of henipaviruses warrants a broad, cross-disciplinary ecosystem health approach that recognises the critical linkages between human activity, ecological change, and livestock and human health.

Polymorphic microsatellite loci for the zebra shark Stegostoma fasciatum

We report on the development and characterization of 14 polymorphic microsatellite loci in the zebra shark (Stegostoma fasciatum). Five tetranucleotide and nine dinucleotide loci were polymorphic with heterozygosities ranging from 0.400 to 0.967 and from three to 22 alleles per locus. Cross-species amplification of these zebra shark primers on four other species of orectolobid sharks was not successful.

Identification of diverse full-length endogenous betaretroviruses in megabats and microbats

Background: Betaretroviruses infect a wide range of species including primates, rodents, ruminants, and marsupials. They exist in both endogenous and exogenous forms and are implicated in animal diseases such as lung cancer in sheep, and in human disease, with members of the human endogenous retrovirus-K (HERV-K) group of endogenous betaretroviruses (βERVs) associated with human cancers and autoimmune diseases. To improve our understanding of betaretroviruses in an evolutionarily distinct host species, we characterized βERVs present in the genomes and transcriptomes of mega- and microbats, which are an important reservoir of emerging viruses.Results: A diverse range of full-length βERVs were discovered in mega- and microbat genomes and transcriptomes including the first identified intact endogenous retrovirus in a bat. Our analysis revealed that the genus Betaretrovirus can be divided into eight distinct sub-groups with evidence of cross-species transmission. Betaretroviruses are revealed to be a complex retrovirus group, within which one sub-group has evolved from complex to simple genomic organization through the acquisition of an env gene from the genus Gammaretrovirus. Molecular dating suggests that bats have contended with betaretroviral infections for over 30 million years.Conclusions: Our study reveals that a diverse range of betaretroviruses have circulated in bats for most of their evolutionary history, and cluster with extant betaretroviruses of divergent mammalian lineages suggesting that their distribution may be largely unrestricted by host species barriers. The presence of βERVs with the ability to transcribe active viral elements in a major animal reservoir for viral pathogens has potential implications for public health. © 2013 Hayward et al.; licensee BioMed Central Ltd.

Trends in therapeutic and prevention strategies for management of Bovine Mastitis: An overview

Mastitis is one of the most economically significant diseases for the dairy industry for backyard farmers in developing countries and high producing herds worldwide. Two of the major factors impeding reduction in the incidence of this disease is [a] the lack of availability of an effective vaccine capable of protecting against multiple etiological agents and [b] propensity of some of the etiological agents to develop persistent antibiotic resistance in biofilms. This is further complicated by the continuing revolving shift in the predominant etiological agents of mastitis, depending upon a multitude of factors such as variability in hygienic practices on farms, easy access leading to overuse of appropriate or inappropriate antibiotics at suboptimal concentrations, particularly in developing countries, and lack of compliance with the recommended treatment schedules. Regardless, Staphylococcus aureus and Streptococcus uberis followed by Escherichia coli, Streptococcus agalactiae has become the predominant etiological agents of bovine mastitis followed Streptococcus agalactiae, Streptococcus dysagalactiae, Klebsiella pneumonia and the newly emerging Mycoplasma bovis. Current approaches being pursued to reduce the negative economic impact of this disease are through early diagnosis of infection, immediate treatment with an antibiotic found to either inhibit or kill the pathogen(s) in vitro using planktonic cultures and the use of the currently marketed vaccines regardless of their demonstrated effectiveness. Given the limitations of breeding programs, including genetic selection to improve resistance against infectious diseases including mastitis, it is imperative to have the availability of an effective broad-spectrum, preferably cross-protective, vaccine capable of protecting against bovine mastitis for reduction in the incidence of bovine mastitis, as well as interrupting the potential cross-species transmission to humans. This overview highlights the major etiological agents, factors affecting susceptibility to mastitis, and the current status of antibiotic-based therapies and prototype vaccine candidates or commercially available vaccines against bovine mastitis as potential preventative strategies. © 2013 Tiwari JG, et al.

Ecological dynamics of emerging bat virus spillover

Viruses that originate in bats may be the most notorious emerging zoonoses that spill over from wildlife into domestic animals and humans. Understanding how these infections filter through ecological systems to cause disease in humans is of profound importance to public health. Transmission of viruses from bats to humans requires a hierarchy of enabling conditions that connect the distribution of reservoir hosts, viral infection within these hosts, and exposure and susceptibility of recipient hosts. For many emerging bat viruses, spillover also requires viral shedding from bats, and survival of the virus in the environment. Focusing on Hendra virus, but also addressing Nipah virus, Ebola virus, Marburg virus and coronaviruses, we delineate this cross-species spillover dynamic from the within-host processes that drive virus excretion to land-use changes that increase interaction among species. We describe how land-use changes may affect co-occurrence and contact between bats and recipient hosts. Two hypotheses may explain temporal and spatial pulses of virus shedding in bat populations: episodic shedding from persistently infected bats or transient epidemics that occur as virus is transmitted among bat populations. Management of livestock also may affect the probability of exposure and disease. Interventions to decrease the probability of virus spillover can be implemented at multiple levels from targeting the reservoir host to managing recipient host exposure and susceptibility.

Ecological dynamics of emerging bat virus spillover

Viruses that originate in bats may be the most notorious emerging zoonoses that spill over from wildlife into domestic animals and humans. Understanding how these infections filter through ecological systems to cause disease in humans is of profound importance to public health. Transmission of viruses from bats to humans requires a hierarchy of enabling conditions that connect the distribution of reservoir hosts, viral infection within these hosts, and exposure and susceptibility of recipient hosts. For many emerging bat viruses, spillover also requires viral shedding from bats, and survival of the virus in the environment. Focusing on Hendra virus, but also addressing Nipah virus, Ebola virus, Marburg virus and coronaviruses, we delineate this cross-species spillover dynamic from the within-host processes that drive virus excretion to land-use changes that increase interaction among species. We describe how land-use changes may affect co-occurrence and contact between bats and recipient hosts. Two hypotheses may explain temporal and spatial pulses of virus shedding in bat populations: episodic shedding from persistently infected bats or transient epidemics that occur as virus is transmitted among bat populations. Management of livestock also may affect the probability of exposure and disease. Interventions to decrease the probability of virus spillover can be implemented at multiple levels from targeting the reservoir host to managing recipient host exposure and susceptibility.

Coronavirus Infection and Diversity in Bats in the Australasian Region

Following the SARS outbreak, extensive surveillance was undertaken globally to detect and identify coronavirus diversity in bats. This study sought to identify the diversity and prevalence of coronaviruses in bats in the Australasian region. We identified four different genotypes of coronavirus, three of which (an alphacoronavirus and two betacoronaviruses) are potentially new species, having less than 90% nucleotide sequence identity with the most closely related described viruses. We did not detect any SARS-like betacoronaviruses, despite targeting rhinolophid bats, the putative natural host taxa. Our findings support the virus-host co-evolution hypothesis, with the detection of Miniopterus bat coronavirus HKU8 (previously reported in Miniopterus species in China, Hong Kong and Bulgaria) in Australian Miniopterus species. Similarly, we detected a novel betacoronavirus genotype from Pteropus alecto which is most closely related to Bat coronavirus HKU9 identified in other pteropodid bats in China, Kenya and the Philippines. We also detected possible cross-species transmission of bat coronaviruses, and the apparent enteric tropism of these viruses. Thus, our findings are consistent with a scenario wherein the current diversity and host specificity of coronaviruses reflects co-evolution with the occasional host shift.

New microsatellite loci for Carcharhinid sharks (Carcharhinus tilstoni and C. sorrah) and their cross-amplification in other shark species

Twelve microsatellite DNA markers were isolated in the spot-tail shark (Carcharhinus sorrah) and nine were isolated in Australian black-tip shark (Carcharhinus tilstoni). These loci plus 18 others developed for sharks from the genera Negaprion, Ginglymostoma, Carcharodon and Isurus were tested for amplification success on four species of Carcharhinus (including C. sorrah and C. tilstoni) and four other species representing three diverse families. Cross-amplification was most common within families. Five loci were subsequently tested for polymorphism on 50 C. sorrah and 60 C. tilstoni. The number of alleles per locus was two to 24 and the average heterozygosity was 0.54 (range 0.16-0.87) for C. sorrah and 0.64 (range 0.44-0.78) for C. tilstoni. These loci may be useful tools for genetic analyses of the Carcharhinidae.

Validation of in silico-predicted genic SNPs in white clover (Trifolium repens L.), an outbreeding allopolyploid species

White clover (Trifolium repens L.) is an obligate outbreeding allotetraploid forage legume. Gene-associated SNPs provide the optimum genetic system for improvement of such crop species. An EST resource obtained from multiple cDNA libraries constructed from numerous genotypes of a single cultivar has been used for in silico SNP discovery and validation. A total of 58 from 236 selected sequence clusters (24.5%) were fully validated as containing polymorphic SNPs by genotypic analysis across the parents and progeny of several two-way pseudo-testcross mapping families. The clusters include genes belonging to a broad range of predicted functional categories. Polymorphic SNP-containing ESTs have also been used for comparative genomic analysis by comparison with whole genome data from model legume species, as well as Arabidopsis thaliana. A total of 29 (50%) of the 58 clusters detected putative ortholoci with known chromosomal locations in Medicago truncatula, which is closely related to white clover within the Trifolieae tribe of the Fabaceae. This analysis provides access to translational data from model species. The efficiency of in silico SNP discovery in white clover is limited by paralogous and homoeologous gene duplication effects, which are resolved unambiguously by the transmission test. This approach will also be applicable to other agronomically important cross-pollinating allopolyploid plant species.

Congruence in QTL for adventitious rooting in Pinus elliottii x Pinus caribaea hybrids resolves between and within-species effects

Targeting between-species effects for improvement in synthetic hybrid populations derived from outcrossing parental tree species may be one way to increase the efficacy and predictability of hybrid breeding. We present a comparative analysis of the quantitative trait loci (QTL) which resolved between from within-species effects for adventitious rooting in two populations of hybrids between Pinus elliottii and P. caribaea, an outbred F1 (n=287) and an inbred-like F2 family (n=357). Most small to moderate effect QTL (each explaining 2-5% of phenotypic variation, PV) were congruent (3 out of 4 QTL in each family) and therefore considered within-species effects as they segregated in both families. A single large effect QTL (40% PV) was detected uniquely in the F2 family and assumed to be due to a between-species effect, resulting from a genetic locus with contrasting alleles in each parental species. Oligogenic as opposed to polygenic architecture was supported in both families (60% and 20% PV explained by 4 QTL in the F 2 and F1 respectively). The importance of adventitious rooting for adaptation to survive water-logged environments was thought in part to explain oligogenic architecture of what is believed to be a complex trait controlled by many hundreds of genes.

Mapping species differences for adventitious rooting in a Corymbia torelliana x Corymbia citriodora subspecies variegata hybrid

Quantitative trait loci (QTL) detection was carried out for adventitious rooting and associated propagation traits in a second-generation outbred Corymbia torelliana x Corymbia citriodora subspecies variegata hybrid family (n=186). The parental species of this cross are divergent in their capacity to develop roots adventitiously on stem cuttings and their propensity to form lignotubers. For the ten traits studied, there was one or two QTL detected, with some QTL explaining large amounts of phenotypic variation (e.g. 66% for one QTL for percentage rooting), suggesting that major effects influence rooting in this cross. Collocation of QTL for many strongly genetically correlated rooting traits to a single region on linkage group 12 suggested pleiotropy. A three locus model was most parsimonious for linkage group 12, however, as differences in QTL position and lower genetic correlations suggested separate loci for each of the traits of shoot production and root initiation. Species differences were thought to be the major source of phenotypic variation for some rooting rate and root quality traits because of the major QTL effects and up to 59-fold larger homospecific deviations (attributed to species differences) relative to heterospecific deviations (attributed to standing variation within species) evident at some QTL for these traits. A large homospecific/heterospecific ratio at major QTL suggested that the gene action evident in one cross may be indicative of gene action more broadly in hybrids between these species for some traits.

Identification of quantitative trait loci for resistance to two species of root-lesion nematode (Pratylenchus thornei and P. neglectus) in wheat

Pratylenchus thornei and P. neglectus are two species of root-lesion nematode that cause substantial yield losses in wheat. No commercially available wheat variety has resistance to both species. A doubled-haploid population developed from a cross between the synthetic hexaploid wheat line CPI133872 and the bread wheat Janz was used to locate and tag quantitative trait loci (QTLs) associated with resistance to both P. thornei and P. neglectus. Wheat plants were inoculated with both species of nematode in independent replicated glasshouse trials repeated over 2 years. Known locations of wheat microsatellite markers were used to construct a framework map. After an initial single-marker analysis to detect marker-trait linkages, chromosome regions associated with putative QTLs were targetted with microsatellite markers to increase map density in the chromosome regions of interest. In total, 148 wheat microsatellite markers and 21 amplified fragment length polymorphism markers were mapped. The codominant microsatellite marker Xbarc183 on the distal end of chromosome 6DS was allelic for resistance to both P. thornei and P. neglectus. The QTL were designated QRlnt.lrc-6D.1 and QRlnn.lrc-6D.1, for the 2 traits, respectively. The allele inherited from CPI133872 explained 22.0-24.2% of the phenotypic variation for P. thornei resistance, and the allele inherited from Janz accounted for 11.3-14.0% of the phenotypic variation for P. neglectus resistance. Composite interval mapping identified markers that flank a second major QTL on chromosome 6DL (QRlnt.lrc-6D.2) that explained 8.3-13.4% of the phenotypic variation for P. thornei resistance. An additional major QTL associated with P. neglectus resistance was detected on chromosome 4DS (QRlnn.lrc-4D.1) and explained a further 10.3-15.4% of the phenotypic variation. The identification and tagging of nematode resistance genes with molecular markers will allow appropriate allele combinations to be selected, which will aid the successful breeding of wheat with dual nematode resistance.

Pathogenic variation of Alternaria species associated with leaf blotch and fruit spot of apple in Australia

Four Alternaria species groups (A. longipes, A. arborescens, A. alternata/A. tenuissima and A. tenuissima/A. mali) are associated with leaf blotch and fruit spot of apple in Australia. There is no information on the variability of pathogenicity among the species and isolates within each species causing leaf blotch or fruit spot. We used a detached leaf assay and an in planta fruit inoculation assay to determine the pathogenicity and virulence of the four Alternaria species. Our results showed that isolates within the same species were not specific to either leaf or fruit tissue and showed great variability in pathogenicity and virulence, indicating cross-pathogenicity, which may be isolate dependent rather than species dependent. Generally, virulence of A. tenuissima and A. alternata isolates on leaf and fruit was higher than other species. Isolates of all species groups were pathogenic on leaves of different cultivars, but pathogenicity on fruit of different cultivars varied among isolates and species. Implications of our findings on prevalence of the diseases in different apple-producing regions in Australia and the development of targeted disease management of the diseases are discussed

Susceptibility to sulfuryl fluoride and lack of cross-resistance to phosphine in developmental stages of the red flour beetle, Tribolium castaneum (Coleoptera: Tenebrionidae)

BACKGROUND Our aim was to ascertain the potential of sulfuryl fluoride (SF) as an alternative fumigant to manage phosphine-resistant pests. We tested the susceptibility of all life stages of red flour beetle, Tribolium castaneum (Herbst), to SF and assessed the presence of cross-resistance to this fumigant in phosphine-resistant strains of this species. RESULTS Analysis of dose–response data indicated that the egg was the stage most tolerant to SF under a 48 h exposure period. At LC50, eggs were 29 times more tolerant than other immature stages and adults, and required a relatively high concentration of 48.2 mg L−1 for complete mortality. No significant differences in tolerance to SF were observed among the three larval instars, pupae and adults, and all of these stages were controlled at a low concentration of 1.32 mg L−1. Phosphine-resistant strains did not show cross-resistance to SF. CONCLUSION Our research concluded that the current maximum registered rate of SF, 1500 gh m−3, is adequate to control all the post-embryonic life stages of T. castaneum over a 48 h fumigation period, but it will fail to achieve complete mortality of eggs, indicating the risk of some survival of eggs under this short exposure period. As there is no cross-resistance to SF in phosphine-resistant insects, it will play a key role in managing phosphine resistance in stored-grain insect pests. © 2014 Commonwealth of Australia. Pest Management Science © 2014 Society of Chemical Industry