Additional comments on the types and nomina of several North American ratsnakes (Pantherophis obsoletus complex, Colubridae, Serpentes)

Here, we provide updates to our recent paper reviewing the taxonomy and nomenclature of the Eastern ratsnakes (Pantherophis obsoletus complex, Colubridae, Serpentes). Specifically, we clarify that Coluber alleghaniensis Holbrook, 1836 is a subjective, rather than objective, senior synonym of Elaphis holbrookii Duméril, Bibron & Duméril, 1854. Contrary to our statement that USNM 1733–4 were syntypes of Scotophis lindheimerii Baird & Girard, 1853, the former is the holotype and the latter is the paratype. The holotype is lost and the paratype is in poor condition, but no neotype designation is warranted at present. We note that USNM 248870, which we designated as the lectotype of Coluber obsoletus lemniscatus Cope, 1888, was originally cataloged as USNM 4710. This catalog number was shared with the type of the salamander Amblystoma tenebrosum Baird & Girard, 1852, and the snake was re-cataloged as USNM 248870 in 1985. Finally, we originally treated C. reticulatus La Cépède, 1789 and C. reticularis Daudin, 1803 as senior subjective synonyms of C. corais Boie, 1827, but here corroborate recent authors in designating it a senior subjective synonym of C. obsoletus Say in James, 1823. As the Commission suppressed C. reticulatus La Cépède, 1789 (an exoplonym), this subsequently rendered C. reticularis Daudin, 1803 (an exoploneonym) unavailable as well.

A study of the composition of the Obsoletus complex and genetic diversity of Culicoides obsoletus populations in Spain

Background The Culicoides obsoletus species complex (henceforth ‘Obsoletus complex’) is implicated in the transmission of several arboviruses that can cause severe disease in livestock, such as bluetongue, African horse sickness, epizootic hemorrhagic disease and Schmallenberg disease. Thus, this study aimed to increase our knowledge of the composition and genetic diversity of the Obsoletus complex by partial sequencing of the cytochrome c oxidase I (cox1) gene in poorly studied areas of Spain. Methods A study of C. obsoletus populations was carried out using a single-tube multiplex polymerase chain reaction (PCR) assay that was designed to differentiate the Obsoletus complex sibling species Culicoides obsoletus and Culicoides scoticus, based on the partial amplification of the cox1 gene, as well as cox1 georeferenced sequences from Spain available at GenBank. We sampled 117 insects of the Obsoletus complex from six locations and used a total of 238 sequences of C. obsoletus (ss) individuals (sampled here, and from GenBank) from 14 sites in mainland Spain, the Balearic Islands and the Canary Islands for genetic diversity and phylogenetic analyses. Results We identified 90 C. obsoletus (ss), 19 Culicoides scoticus and five Culicoides montanus midges from the six collection sites sampled, and found that the genetic diversity of C. obsoletus (ss) were higher in mainland Spain than in the Canary Islands. The multiplex PCR had limitations in terms of specificity, and no cryptic species within the Obsoletus complex were identified. Conclusions Within the Obsoletus complex, C. obsoletus (ss) was the predominant species in the analyzed sites of mainland Spain. Information about the species composition of the Obsoletus complex could be of relevance for future epidemiological studies when specific aspects of the vector competence and capacity of each species have been identified. Our results indicate that the intraspecific divergence is higher in C. obsoletus (ss) northern populations, and demonstrate the isolation of C. obsoletus (ss) populations of the Canary Islands. Graphical abstract

Field studies on breeding sites of Culicoides Latreille (Diptera: Ceratopogonidae) in agriculturally used and natural habitats

Culicoides are vectors of pathogens mainly of veterinary importance. To establish targeted vector control measures, it is paramount to comprehend the ecological factors determining their distribution. Therefore, we used emergence traps to sample eight biotopes and assess their potential as breeding sites. Part one of the study investigates agricultural habitats, part two compares four biotopes of a forest-dominated area with less anthropogenic influence, including a physicochemical analysis of soil moisture, pH value and organic content. Thirteen culicoid species were collected, with a strong dominance of the Obsoletus Complex on meadows, and with Culicoides punctatus (Meigen), Culicoides pictipennis (Staeger) and the Obsoletus Complex, to be the most abundant species in the natural habitats. Several co-existing species were found, some of them not having been described before. Our results suggest that ungrazed meadows seem unsuitable as breeding sites. Only the influence of livestock creates adequate conditions for certain midge species. The alder on fen site contained most culicoid species with the highest species diversity. Our study clearly indicates that knowledge of species-specific preferences for environmental habitat conditions (choice of breeding site) in connection to soil conditions is crucial to understand the biology and phenology of midges and their role as vectors of pathogens.

On the validity and taxonomic status of Crypturellus obsoletus griseiventris (Salvadori, 1895) and C. o. hypochraceus (Miranda-Ribeiro, 1938) (Aves, Tinamidae)

The nine currently recognized subspecies in the Brown Tinamou (Crypturellus obsoletus) complex are disjunctly widespread in South America, and at least three of them occur in Brazil. Morphological diagnosis of most of these taxa is imprecise, in contrast with consistent vocal differences described in the literature. We conducted a taxonomic review of two Amazonian taxa, C. o. griseiventris and C. o. hypochraceus, using morphological, morphometric, and vocal characters. Our results indicate that C. o. hypochraceus (Miranda-Ribeiro, 1938) is a junior synonym of C. o. griseiventris (Salvadori, 1895), and that Crypturellus griseiventris (Salvadori, 1895) must be treated as a full species, based on unique and fully diagnosable plumage and vocal patterns.

Post-epidemic Occurrence of Bluetongue Virus Vectors (Culicoides Species) în Iași County

Culicoides is a genus of biting midges in the family Ceratopogonidae, representing a big threat, giving that several species serve as biological vectors of pathogens of veterinary importance. BTV distribution and persistence is thought to be almost entirely dependent upon the presence or absence of Culicoides adults vector. In this study, biting midges were collected during 2015-2016, once a week, from April to December. In the first year, 2617 BTV vector specimens were collected and identified as Culicoides obsoletus (41.2%; 95% CI: 39-43), Culicoides pulicaris (58%; 95% CI: 56-59) and Culicoides nubeculosus (0.8%; 95% CI: 0.4-1.1). In the second year, 165 BTV vector specimens were collected and identified as Culicoides obsoletus (44.25%; 95% CI: 36-51), Culicoides pulicaris (50.3%; 95% CI: 42-57), Culicoides dewulfi (4.84%; 95% CI: 1.5-8.1) and Culicoides nubeculosus (0.61%; 95% CI: 0.5-1.7). Culicoides pulicaris and Culicoides obsoletus complex were especially abundant within the two collection sites in Iași County. Our data on post-epidemic occurrence and composition of BTV vector species in the studied area will facilitate a better understanding of Culicoides vectors distribution and the possible role in bluetongue virus overwintering.

Identifying Spanish Areas at More Risk of Monthly BTV Transmission with a Basic Reproduction Number Approach

Bluetongue virus (BTV) causes a disease that is endemic in Spain and its two major biological vector species, C. imicola and the Obsoletus complex species, differ greatly in their ecology and distribution. Understanding the seasonality of BTV transmission in risk areas is key to improving surveillance and control programs, as well as to better understand the pathogen transmission networks between wildlife and livestock. Here, monthly risk transmission maps were generated using risk categories based on well-known BTV R0 equations and predicted abundances of the two most relevant vectors in Spain. Previously, Culicoides spp. predicted abundances in mainland Spain and the Balearic Islands were obtained using remote sensing data and random forest machine learning algorithm. Risk transmission maps were externally assessed with the estimated date of infection of BTV-1 and BTV-4 historical outbreaks. Our results highlight the differences in risk transmission during April-October, June-August being the period with higher R0 values. Likewise, a natural barrier has been identified between northern and central-southern areas at risk that may hamper BTV spread between them. Our results can be relevant to implement risk-based interventions for the prevention, control and surveillance of BTV and other diseases shared between livestock and wildlife host populations.

Environmental Drivers of Adult Seasonality and Abundance of Biting Midges Culicoides (Diptera: Ceratopogonidae), Bluetongue Vector Species in Spain

Bluetongue is a viral disease affecting wild and domestic ruminants transmitted by several species of biting midges Culicoides Latreille. The phenology of these insects were analyzed in relation to potential environmental drivers. Data from 329 sites in Spain were analyzed using Bayesian Generalized Linear Mixed Model (GLMM) approaches. The effects of environmental factors on adult female seasonality were contrasted. Obsoletus complex species (Diptera: Ceratopogonidae) were the most prevalent across sites, followed by Culicoides newsteadi Austen (Diptera: Ceratopogonidae). Activity of female Obsoletus complex species was longest in sites at low elevation, with warmer spring average temperatures and precipitation, as well as in sites with high abundance of cattle. The length of the Culicoides imicola Kieffer (Diptera: Ceratopogonidae) female adult season was also longest in sites at low elevation with higher coverage of broad-leaved vegetation. Long adult seasons of C. newsteadi were found in sites with warmer autumns and higher precipitation, high abundance of sheep. Culicoides pulicaris (Linnaeus) (Diptera: Ceratopogonidae) had longer adult periods in sites with a greater number of accumulated degree days over 10°C during winter. These results demonstrate the eco-climatic and seasonal differences among these four taxa in Spain, which may contribute to determining sites with suitable environmental circumstances for each particular species to inform assessments of the risk of Bluetongue virus outbreaks in this region.

Comparing Different Larval Food Sources and Temperature Regimes for the Rearing of Culicoides obsoletus/scoticus Complex Midges, the Predominant Bluetongue, and Schmallenberg Virus Vectors in Northern Europe

During the last decade, Europe was confronted with the emergence of two Culicoides-borne viruses, bluetongue virus and Schmallenberg virus. Both diseases caused economic losses in cattle, sheep, and goats. Despite their importance, only limited knowledge is available on the developmental stages of Culicoides obsoletus complex midges, and a lab-reared colony has not yet been established. Therefore, this experimental study aims to compare the emergence patterns from field-collected C. obsoletus complex from egg to adult that were exposed to different combinations of temperature and larval substrates (T1: 1% liquid broth (LB) agar at 24 °C; T2: 1% LB agar with dung patches at 24 °C; T3: 1% LB agar at 28 °C). The rearing dishes were kept in an environmental chamber under an 8 h/16 h dark–light regime and 80%–85% humidity. Algae and nematodes were provided as a food source. The average period required to develop from egg to adult was 24 days and varied strongly in each of the conditions (T1: 15–47 days; T2: 14–35 days; T3: 20–37 days). A bias of sex ratio was observed towards the production of males in all three treatments with only 23.9% (T1), 23.5% (T2), and 0% (T3) females. Larval mortality was highest in T3 (25.9%), followed by T2 (8.6%), and T1 (1.8%). All pupae produced adults within 1 to 10 days, and the emergence rates for pupae varied strongly between treatments: 49.5% (T1), 71.6% (T2), and 38.5% (T3). This study shows that the C. obsoletus complex can be reared under laboratory conditions from blood-fed, wild-caught females to emerged progeny. More larval substrates and food sources should be tested with the aim of obtaining a 1:1 sex ratio to bring us one step closer to a viable lab-reared colony.

Resolving spatial complexities of hybridization in the context of the gray zone of speciation in North American ratsnakes (Pantherophis obsoletus complex)

Inferring the history of divergence between species in a framework that permits the presence of gene flow has been crucial for characterizing the gray zone of speciation, which is the period of time where lineages have diverged but have not yet achieved strict reproductive isolation. However, estimates of both divergence times and rates gene flow often ignore spatial information, for example the formation and shape of hybrid zones. Using population genomic data from the eastern ratsnake complex (Pantherophis obsoletus), we infer phylogeographic groups, gene flow, changes in demography, the timing of divergence, and hybrid zone widths. We examine the spatial context of diversification by linking migration and timing of divergence to the location and widths of hybrid zones. Artificial neural network approaches are applied to understand how landscape features and past climate have influenced population genetic structure among these lineages prior to hybridization. Rates of migration between lineages are associated with the width and shape of hybrid zones. Timing of divergence is not related to migration rate across species pairs and is therefore a poor proxy for inferring position in the gray zone. However, timing of divergence is related to the number of loci weakly introgressing through hybrid zones.