Bateman's principle is reversed in a cooperatively breeding bird.

Bateman's principle is not only used to explain sex differences in mating behaviour, but also to determine which sex has the greater opportunity for sexual selection. It predicts that the relationship between the number of mates and the number of offspring produced should be stronger for males than for females. Yet, it is unclear whether Bateman's principle holds in cooperatively breeding systems where the strength of selection on traits used in intrasexual competition is high in both sexes. We tested Bateman's principle in the cooperatively breeding superb starling (Lamprotornis superbus), finding that only females showed a significant, positive Bateman gradient. We also found that the opportunity for selection was on average higher in females, but that its strength and direction oscillated through time. These data are consistent with the hypothesis that sexual selection underlies the female trait elaboration observed in superb starlings and other cooperative breeders. Even though the Bateman gradient was steeper for females than for males, the year-to-year oscillation in the strength and direction of the opportunity for selection likely explains why cooperative breeders do not exhibit sexual role reversal. Thus, Bateman's principle may not hold in cooperative breeders where both sexes appear to be under mutually strong sexual selection.

Ranavirus (Frog Virus 3) Infection in Free-Living Three-Toed Box Turtles (Terrapene mexicana triunguis) in Missouri, USA.

Frog virus 3 (FV3) and related ranaviruses are emerging infectious disease threats to ectothermic vertebrate species globally. Although the impact of these viruses on amphibian health is relatively well studied, less is understood about their effects on reptile health. We report two cases of FV3 infection, 11 mo apart, in three-toed box turtles (Terrapene mexicana triunguis) from a wildlife rehabilitation center. Case 1 had upper respiratory signs upon intake but had no clinical signs at the time of euthanasia 1 mo later. Case 2 presented for vehicular trauma, had ulcerative pharyngitis and glossitis, and died overnight. In case 1, we detected FV3 nucleic acid with qPCR in oral swabs, kidney, liver, spleen, and tongue. In case 2, we detected FV3 in an oral swab, an oral plaque, heart, kidney, lung, liver, spleen, and tongue. We also detected FV3 nucleic acid with in situ hybridization for case 2. For both cases, FV3 was isolated in cell culture and identified with DNA sequencing. Histopathologic examination of postmortem tissue from case 1 was unremarkable, whereas acute hemorrhagic pneumonia and splenic necrosis were noted in case 2. The difference in clinical signs between the two cases may have been due to differences in the temporal course of FV3 disease at the time of necropsy. Failure to detect this infection previously in Missouri reptiles may be due to lack of surveillance, although cases may also represent a novel spillover to box turtles in Missouri. Our findings reiterate previous suggestions that the range of FV3 infection may be greater than previously documented and that infection may occur in host species yet to be tested.

Health assessments uncover novel viral sequences in five species of Galapagos tortoises.

Emerging infectious diseases (EIDs) have been reported as causes of morbidity and mortality in free-living animal populations, including turtles and tortoises, and they have even resulted in species extinctions, with human activities contributing to the spread of many of these diseases. In the Galapagos, giant tortoises are endangered due to habitat change, invasive species, and other human impacts; however, the impact of EIDs on Galapagos tortoise conservation remains understudied. To fill this gap, we conducted health assessments of five tortoise species from the islands of Santa Cruz, Isabela and Española. We performed health evaluations of 454 animals and PCR testing for pathogens known to be relevant in other tortoise species. We identified two novel sequences of adenoviruses and four of herpesviruses. Based on alignments of the DNA polymerase gene and maximum likelihood phylogenetic analyses, we found both novel adenoviruses to be most closely related to red footed tortoise adenovirus 2, by nucleotide sequence and red footed tortoise adenovirus 1, based on amino acid sequence. Three of the herpesvirus sequences translated into the same deduced amino acid sequence; therefore, they may be considered the same viral species, closely related to terrapene herpesvirus 2. The fourth herpesvirus sequence was highly divergent from any sequence previously detected and is related to an eagle owl herpesvirus based on nucleotide sequence and to loggerhead oro-cutaneous herpesvirus based on amino acids. These novel viruses may be pathogenic for giant tortoises under specific conditions (e.g., stress). Continued screening is crucial to determine if these viruses play a role in tortoise fitness, morbidity and survival. This information allows us to provide recommendations to the Galapagos National Park Directorate and other institutions to improve the management of these unique species both in Galapagos and globally, and for tortoise reintroduction plans throughout the archipelago.

A novel herpesvirus detected in 3 species of chelonians.

Herpesviruses are found in free-living and captive chelonian populations, often in association with morbidity and mortality. To date, all known chelonian herpesviruses fall within the subfamily Alphaherpesvirinae. We detected a novel herpesvirus in 3 species of chelonians: a captive leopard tortoise (Stigmochelys pardalis) in western TX, USA; a steppe tortoise (Testudo [Agrionemys] horsfieldii) found near Fort Irwin, CA, USA; and 2 free-living, three-toed box turtles (Terrapene mexicana triunguis) found in Forest Park, St. Louis, MO. The leopard tortoise was coinfected with the tortoise intranuclear coccidian and had clinical signs of upper respiratory tract disease. The steppe tortoise had mucopurulent nasal discharge and lethargy. One of the three-toed box turtles had no clinical signs; the other was found dead with signs of trauma after being observed with blepharedema, tympanic membrane swelling, cervical edema, and other clinical signs several weeks prior to death. Generally, the branching order of the turtle herpesviruses mirrors the divergence patterns of their hosts, consistent with codivergence. Based on phylogenetic analysis, this novel herpesvirus clusters with a clade of viruses that infect emydid hosts and is likely of box turtle origin. Therefore, we suggest the name terrapene alphaherpesvirus 3 (TerAHV3) for the novel virus. This virus also has the ability to host-jump to tortoises, and previously documented herpesviral morbidity tends to be more common in aberrant hosts. The relationship between clinical signs and infection with TerAHV3 in these animals is unclear, and further investigation is merited.

Endogenization of a Prosimian Retrovirus during Lemur Evolution.

Studies of viruses that coevolved with lemurs provide an opportunity to understand the basal traits of primate viruses and provide an evolutionary context for host-virus interactions. Germline integration of endogenous retroviruses (ERVs) are fossil evidence of past infections. Hence, characterization of novel ERVs provides insight into the ancient precursors of extant viruses and the evolutionary history of their hosts. Here, we report the discovery of a novel endogenous retrovirus present in the genome of a lemur, Coquerel's sifaka (Propithecus coquereli). Using next-generation sequencing, we identified and characterized the complete genome sequence of a retrovirus, named prosimian retrovirus 1 (PSRV1). Phylogenetic analyses indicate that PSRV1 is a gamma-type betaretrovirus basal to the other primate betaretroviruses and most closely related to simian retroviruses. Molecular clock analysis of PSRV1 long terminal repeat (LTR) sequences estimated the time of endogenization within 4.56 MYA (± 2.4 MYA), placing it after the divergence of Propithecus species. These results indicate that PSRV1 is an important milestone of lemur evolution during the radiation of the Propithecus genus. These findings may have implications for both human and animal health in that the acquisition of a gamma-type env gene within an endogenized betaretrovirus could facilitate a cross-species jump between vertebrate class hosts.

Investigation of the mechanism by which herpes simplex virus type 1 LAT sequences modulate preferential establishment of latent infection in mouse trigeminal ganglia.

We previously demonstrated that herpes simplex virus type 1 (HSV-1) preferentially establishes latent infection in monoclonal antibody (MAb) A5-positive ganglionic neurons and that a 2.8-kb portion of the HSV-1 genome, corresponding to the 5' end of the LAT (latency-associated transcript) coding region, is responsible for this phenotype (38, 65). In the current study we carried out further genetic mapping of this latency phenotype and investigated some of the mechanisms that might be responsible. Studies with the chimeric virus HSV-1 17syn+/LAT2, an HSV-1 virus engineered to express HSV-2 LAT, demonstrated that this virus exhibited an HSV-2 latency phenotype, preferentially establishing latency in MAb KH10-positive neurons. This result is complementary to that previously described for the chimeric virus HSV-2 333/LAT1 and indicate that the HSV-1 latency phenotype can be changed to that of HSV-2 by substitution of a 2.8-kb piece of complementary viral DNA. Sequential studies in which we evaluated the pattern of HSV-1 latent infection of the mouse trigeminal ganglion following ocular inoculation with viruses with deletions of functional thymidine kinase, glycoprotein E, ICP0, and US9 protein demonstrate that preferential establishment of HSV-1 latent infection in A5-positive neurons is not a consequence of (i) differential access of HSV-1 to A5-positive neurons,(ii) differential cell-to-cell spread of HSV-1 to A5-positive neurons, (iii) differential "round-trip" spread of HSV-1 to A5-positive neurons, or (iv) expression of ICP0. Additional mapping studies with the HSV-1 LAT deletion viruses dLAT371, 17DeltaSty, and 17Delta348 indicate that most of the LAT 5' exon is not required for HSV-1 to preferentially establish latent infection in A5-positive neurons.

Latency-associated transcript (LAT) exon 1 controls herpes simplex virus species-specific phenotypes: reactivation in the guinea pig genital model and neuron subtype-specific latent expression of LAT.

Herpes simplex virus 1 (HSV-1) and HSV-2 cause similar acute infections but differ in their abilities to reactivate from trigeminal and lumbosacral dorsal root ganglia. During latency, HSV-1 and HSV-2 also preferentially express their latency-associated transcripts (LATs) in different sensory neuronal subtypes that are positive for A5 and KH10 markers, respectively. Chimeric virus studies showed that LAT region sequences influence both of these viral species-specific phenotypes. To further map the LAT region sequences responsible for these phenotypes, we constructed the chimeric virus HSV2-LAT-E1, in which exon 1 (from the LAT TATA to the intron splice site) was replaced by the corresponding sequence from HSV-1 LAT. In intravaginally infected guinea pigs, HSV2-LAT-E1 reactivated inefficiently relative to the efficiency of its rescuant and wild-type HSV-2, but it yielded similar levels of viral DNA, LAT, and ICP0 during acute and latent infection. HSV2-LAT-E1 preferentially expressed the LAT in A5+ neurons (as does HSV-1), while the chimeric viruses HSV2-LAT-P1 (LAT promoter swap) and HSV2-LAT-S1 (LAT sequence swap downstream of the promoter) exhibited neuron subtype-specific latent LAT expression phenotypes more similar to that of HSV-2 than that of HSV-1. Rescuant viruses displayed the wild-type HSV-2 phenotypes of efficient reactivation in the guinea pig genital model and a tendency to express LAT in KH10+ neurons. The region that is critical for HSV species-specific differences in latency and reactivation thus lies between the LAT TATA and the intron splice site, and minor differences in the 5' ends of chimeric sequences in HSV2-LAT-E1 and HSV2-LAT-S1 point to sequences immediately downstream of the LAT TATA.

Human herpesvirus 6-A, 6-B, and 7 in vitreous fluid samples.

Human herpesvirus 6 and 7 (HHV-6, HHV-7) have been associated with several neurologic syndromes and have been detected in nervous tissue from healthy persons; however, only two cases of HHV-6A have been reported to be associated with intraocular inflammatory disease. Vitreous fluid was tested from 101 patients, including 69 samples from patients with ocular inflammation including CMV retinitis, idiopathic retinitis, iritis, and vitritis, for HHV-6A, HHV-6B, and HHV-7 DNA by PCR. HHV-6A DNA (4,950 copies per ml) was detected in vitreous fluid from one patient with CMV retinitis, HHV-6B DNA (10,140 copies per ml) was detected in vitreous fluid from one patient with idiopathic ocular inflammation in the absence of CMV DNA, and HHV-7 was not detected in any of the vitreous samples. HHV-6A, HHV-6B, and HHV-7 DNA are detectable in less than 2% of vitreous samples in patients with ocular inflammation.

Pathogen Transmission from Humans to Great Apes is a Growing Threat to Primate Conservation.

All six great ape species are listed as endangered or critically endangered by the IUCN and experiencing decreasing population trends. One of the threats to these non-human primates is the transmission of pathogens from humans. We conducted a literature review on occurrences of pathogen transmission from humans to great apes to highlight this often underappreciated issue. In total, we found 33 individual occurrences of probable or confirmed pathogen transmission from humans to great apes: 23 involved both pathogen and disease transmission, 7 pathogen transmission only, 2 positive antibody titers to zoonotic pathogens, and 1 pathogen transmission with probable disease. Great ape populations were categorized into captive, semi-free-living, and free-living conditions. The majority of occurrences involved chimpanzees (Pan troglodytes) (n = 23) or mountain gorillas (Gorilla beringei beringei) (n = 8). These findings have implications for conservation efforts and management of endangered great ape populations. Future efforts should focus on monitoring and addressing zoonotic pathogen and disease transmission between humans, great ape species, and other taxa to ensure the health of humans, wild and domestic animals, and the ecosystems we share.

Perkinsus mediterraneus n. sp., a protistan parasite of the European flat oyster Ostrea edulis from the Balearic Islands, Mediterranean Sea.

A new species, Perkinsus mediterraneus, a protistan parasite of the European oyster Ostrea edulis (L.), farmed along the coast of the Balearic Islands, Mediterranean Sea, is described. Morphological examinations with light and transmission electron microscopy, DNA sequence-analysis and enlargement in Ray's fluid thioglycollate medium (RFTM) confirmed that this parasite belongs to the genus Perkinsus. Specific morphological and genetic characteristics indicated that it should be considered a new species in the genus. Sequencing of the small subunit ribosomal (ssu rRNA) gene confirmed that the parasite belongs to the genus Perkinsus, and sequences of the internal transcribed spacer (ITS) were distinct from any Perkinsus ITS sequences previously published and/or deposited in the GenBank. Phylogenetic analysis revealed that the ITS sequences of the new species formed a monophyletic group comprising a sister clade to the P. atlanticus/olseni group. In addition, morphological differences were observed between the new species and the other described Perkinsus spp.. After incubation in RFTM for 1 wk, the prezoosporangium had reached an extremely large size (97.4 +/- 1.99 microm) (mean +/- SE), and after 2 wk incubation had again almost doubled in size (167.1 +/- 8.09 microm). The discharge-tube length was one sixth the diameter of the zoosporangium, i.e. a ratio of 17.36:97.38, the lowest ratio observed for any Perkinsus species. At the ultrastructural level, zoosporangia and zoospores exhibited some differences compared to other Perkinsus species.

LAT region factors mediating differential neuronal tropism of HSV-1 and HSV-2 do not act in trans.

After HSV infection, some trigeminal ganglion neurons support productive cycle gene expression, while in other neurons the virus establishes a latent infection. We previously demonstrated that HSV-1 and HSV-2 preferentially establish latent infection in A5+ and KH10+ sensory neurons, respectively, and that exchanging the latency-associated transcript (LAT) between HSV-1 and HSV-2 also exchanges the neuronal preference. Since many viral genes besides the LAT are functionally interchangeable between HSV-1 and HSV-2, we co-infected HSV-1 and HSV-2, both in vivo and in vitro, to determine if trans-acting viral factors regulate whether HSV infection follows a productive or latent pattern of gene expression in sensory neurons. The pattern of HSV-1 and HSV-2 latent infection in trigeminal neurons was no different following co-infection than with either virus alone, consistent with the hypothesis that a trans-acting viral factor is not responsible for the different patterns of latent infection of HSV-1 and HSV-2 in A5+ and KH10+ neurons. Since exchanging the LAT regions between the viruses also exchanges neuronal preferences, we infected transgenic mice that constitutively express 2.8 kb of the LAT region with the heterologous viral serotype. Endogenous expression of LAT did not alter the pattern of latent infection after inoculation with the heterologous serotype virus, demonstrating that the LAT region does not act in trans to direct preferential establishment of latency of HSV-1 and HSV-2. Using HSV1-RFP and HSV2-GFP in adult trigeminal ganglion neurons in vitro, we determined that HSV-1 and HSV-2 do not exert trans-acting effects during acute infection to regulate neuron specificity. Although some neurons were productively infected with both HSV-1 and HSV-2, no A5+ or KH10+ neurons were productively infected with both viruses. Thus, trans-acting viral factors do not regulate preferential permissiveness of A5+ and KH10+ neurons for productive HSV infection and preferential establishment of latent infection.

Spontaneous reactivation of herpes simplex virus type 1 in latently infected murine sensory ganglia.

Careful studies of mouse trigeminal ganglia (TG) latently infected with herpes simplex virus type 1 (HSV-1) indicate the presence of productive cycle viral gene products and persistent immune response, suggesting ongoing spontaneous viral reactivation in these tissues. In the present study we set out to determine whether infectious virus is present in murine TG latently infected with HSV-1 (KOS). At 37 days after ocular inoculation we found a small amount of infectious virus in ca. 6% of latently infected murine TG. Furthermore, the amount of infectious virus that we detected (PFU per viral antigen-positive neuron) was similar to that detected in acutely infected ganglia. We conclude that spontaneous reactivation of infectious HSV-1 occurs in the mouse TG and is likely the principle cause of viral protein expression in these tissues. We next examined the role of latency-associated transcript (LAT) in spontaneous ganglionic reactivation by examining ganglia latently infected with KOS dlLAT1.8, a LAT deletion virus. Through the use of immunocytochemistry we found that KOS dlLAT1.8 had a rate of spontaneous ganglionic reactivation very similar to that of HSV-1 (KOS). Studying spontaneous ganglionic reactivation of HSV in the mouse TG allows a direct study of viral reactivation from latently infected neurons without the potential confounders and complicating downstream events that accompany the study of viral reactivation by explantation or peripheral viral shedding. Since most cases of human viral shedding and reactivation are not associated with a known trigger, spontaneous ganglionic reactivation of HSV-1 may be a better model of human disease than existing models.