Sloths host Anhanga virus‐related phleboviruses across large distances in time and space

Sloths are genetically and physiologically divergent mammals. Phleboviruses are major arthropod‐borne viruses (arboviruses) causing disease in humans and other animals globally. Sloths host arboviruses, but virus detections are scarce. A phlebovirus termed Anhanga virus (ANHV) was isolated from a Brazilian Linnaeus's two‐toed sloth (Choloepus didactylus) in 1962. Here, we investigated the presence of phleboviruses in sera sampled in 2014 from 74 Hoffmann's two‐toed (Choloepus hoffmanni, n = 65) and three‐toed (Bradypus variegatus, n = 9) sloths in Costa Rica by broadly reactive RT‐PCR. A clinically healthy adult Hoffmann's two‐toed sloth was infected with a phlebovirus. Viral load in this animal was high at 8.5 × 10⁷ RNA copies/ml. The full coding sequence of the virus was determined by deep sequencing. Phylogenetic analyses and sequence distance comparisons revealed that the new sloth virus, likely representing a new phlebovirus species, provisionally named Penshurt virus (PEHV), was most closely related to ANHV, with amino acid identities of 93.1%, 84.6%, 94.7% and 89.0% in the translated L, M, N and NSs genes, respectively. Significantly more non‐synonymous mutations relative to ANHV occurred in the M gene encoding the viral glycoproteins and in the NSs gene encoding a putative interferon antagonist compared to L and N genes. This was compatible with viral adaptation to different sloth species and with micro‐evolutionary processes associated with immune evasion during the genealogy of sloth‐associated phleboviruses. However, gene‐wide mean dN/dS ratios were low at 0.02–0.15 and no sites showed significant evidence for positive selection, pointing to comparable selection pressures within sloth‐associated viruses and genetically related phleboviruses infecting hosts other than sloths. The detection of a new phlebovirus closely‐related to ANHV, in sloths from Costa Rica fifty years after and more than 3,000 km away from the isolation of ANHV confirmed the host associations of ANHV‐related phleboviruses with the two extant species of two‐toed sloths.     

Incubation of frozen-thawed llama sperm with seminal plasma

Seminal plasma is intimately connected to sperm physiology and particularly in South American Camelids, has demonstrated to be involved in multiple physiological reproductive events. Different percentages of seminal plasma (0%, 10% and 50%) were added to thawed llama semen samples with the objective of evaluating the interaction with cryopreserved sperm over time (0, 1.5 and 3 hr at 37°C). A total of 20 ejaculates from five adult llama males (n = 5; r = 4) were evaluated. A significant decrease in sperm motility, membrane function and live sperm was observed in all thawed samples (0%, 10% and 50%) at 0 hr when compared to raw semen. Neither morphology nor chromatin condensation was altered in all thawed samples (p > .05), but a significant increase in the percentage of spermatozoa with fragmented DNA was observed after thawing all samples versus raw semen. When evaluating thawed samples over time, a significant decrease of motility and membrane function was observed, while the percentages of total live sperm were preserved over the 3 hr of incubation in all final concentrations evaluated. To conclude, the addition of 10% or 50% of seminal plasma was incapable of preserving motility or membrane function of frozen-thawed llama sperm during 3 hr of incubation.     

Origins and antiquity of X-linked triallelic color vision systems in New World monkeys

It is known that the squirrel monkey, marmoset, and other related New World (NW) monkeys possess three high-frequency alleles at the single X-linked photopigment locus, and that the spectral sensitivity peaks of these alleles are within those delimited by the human red and green pigment genes. The three alleles in the squirrel monkey and marmoset have been sequenced previously. In this study, the three alleles were found and sequenced in the saki monkey, capuchin, and tamarin. Although the capuchin and tamarin belong to the same family as the squirrel monkey and marmoset, the saki monkey belongs to a different family and is one of the species that is most divergent from the squirrel monkey and marmoset, suggesting the presence of the triallelic system in many NW monkeys. The nucleotide sequences of these alleles from the five species studied indicate that gene conversion occurs frequently and has partially or completely homogenized intronic and exonic regions of the alleles in each species, making it appear that a triallelic system arose independently in each of the five species studied. Nevertheless, a detailed analysis suggests that the triallelic system arose only once in the NW monkey lineage, from a middle wavelength (green) opsin gene, and that the amino acid differences at functionally critical sites among alleles have been maintained by natural selection in NW monkeys for >20 million years. Moreover, the two X-linked opsin genes of howler monkeys (a NW monkey genus) were evidently derived from the incorporation of a middle (green) and a long wavelength (red) allele into one chromosome; these two genes together with the (autosomal) blue opsin gene would immediately enable even a male monkey to have trichromatic vision.     

Molecular and Insecticidal Characterization of a Novel Cry-Related Protein from Bacillus Thuringiensis Toxic against Myzus persicae

This study describes the insecticidal activity of a novel Bacillus thuringiensis Cry-related protein with a deduced 799 amino acid sequence (~89 kDa) and ~19% pairwise identity to the 95-kDa-aphidicidal protein (sequence number 204) from patent US 8318900 and ~40% pairwise identity to the cancer cell killing Cry proteins (parasporins Cry41Ab1 and Cry41Aa1), respectively. This novel Cry-related protein contained the five conserved amino acid blocks and the three conserved domains commonly found in 3-domain Cry proteins. The protein exhibited toxic activity against the green peach aphid, Myzus persicae (Sulzer) (Homoptera: Aphididae) with the lowest mean lethal concentration (LC₅₀ = 32.7 μg/mL) reported to date for a given Cry protein and this insect species, whereas it had no lethal toxicity against the Lepidoptera of the family Noctuidae Helicoverpa armigera (Hübner), Mamestra brassicae (L.), Spodoptera exigua (Hübner), S. frugiperda (J.E. Smith) and S. littoralis (Boisduval), at concentrations as high as ~3.5 μg/cm². This novel Cry-related protein may become a promising environmentally friendly tool for the biological control of M. persicae and possibly also for other sap sucking insect pests.     

Serological Positivity against Selected Flaviviruses and Alphaviruses in Free-Ranging Bats and Birds from Costa Rica Evidence Exposure to Arboviruses Seldom Reported Locally in Humans

Arboviruses have two ecological transmission cycles: sylvatic and urban. For some, the sylvatic cycle has not been thoroughly described in America. To study the role of wildlife in a putative sylvatic cycle, we sampled free-ranging bats and birds in two arbovirus endemic locations and analyzed them using molecular, serological, and histological methods. No current infection was detected, and no significant arbovirus-associated histological changes were observed. Neutralizing antibodies were detected against selected arboviruses. In bats, positivity in 34.95% for DENV-1, 16.26% for DENV-2, 5.69% for DENV-3, 4.87% for DENV-4, 2.43% for WNV, 4.87% for SLEV, 0.81% for YFV, 7.31% for EEEV, and 0.81% for VEEV was found. Antibodies against ZIKV were not detected. In birds, PRNT results were positive against WNV in 0.80%, SLEV in 5.64%, EEEV in 8.4%, and VEEV in 5.63%. An additional retrospective PRNT analysis was performed using bat samples from three additional DENV endemic sites resulting in a 3.27% prevalence for WNV and 1.63% for SLEV. Interestingly, one sample resulted unequivocally WNV positive confirmed by serum titration. These results suggest that free-ranging bats and birds are exposed to not currently reported hyperendemic-human infecting Flavivirus and Alphavirus; however, their role as reservoirs or hosts is still undetermined.     

Response to: Human papillomavirus (HPV) vaccine safety concerning POTS,CRPS and related conditions

Clinical Autonomic Research -