Tick-Borne Pathogens Associated with Medically Important Ticks in Alabama: A Four-Year Survey.

Background: Tick-borne diseases (TBDs) represent a significant threat to human health in the United States. Based on reported cases of notifiable TBDs to the Centers for Disease Control and Prevention (CDC) the state of Alabama is no exception, yet previously there has been no active surveillance program in place to comprehensively assess the presence and prevalence of tick vectors and their associated TBD pathogens in Alabama. Here we summarize initial findings from a 4-year survey to address this unmet need. Materials and Methods: Beginning in 2018 and proceeding through 2021, ticks were collected throughout the state of Alabama and pooled before being screened for a panel of TBD pathogens known to circulate in the United States. Results: Consistent with previously reported cases, TBD pathogens associated with anaplasmosis, babesiosis, ehrlichiosis, and spotted fever rickettsiosis were detected in ticks of Alabama. Causative agents for tularemia and Lyme disease were not detected despite previously reported human disease cases. There was also no evidence of Heartland virus despite recent reports of the virus being detected in ticks in Northwestern counties. Conclusions: While these results serve to provide some insights into TBD pathogens associated with ticks in Alabama, they also raise many questions that highlight the need for additional studies and continued surveillance to fully understand the TBD threat to human health in Alabama.

The general architecture of black fly-parasite interactions: Parasitism in lotic systems at a continental scale.

We examined the general architecture of interactions between stream-dwelling larval black flies (Diptera: Simuliidae) and their common parasites in 1736 collections across North America. Mermithid nematodes (family Mermithidae), microsporidia (phylum Microsporidia), and the fungus Coelomycidium simulii Debaisieux (phylum Blastocladiomycota) infected larval black flies. We found similar continental distributions for these three parasite taxa across North America. At least one of these taxa was represented in 42.2% of all black fly collections. Species interactions in ecological networks typically imply that each link between species is equally important. By employing quantitative measures of host susceptibilities and parasite dependencies, we provide a more complete structure for host-parasite networks. The distribution of parasite dependencies and host susceptibilities were right-skewed, with low values indicating that most dependencies (parasites) and susceptibilities (hosts) were weak. Although regression analysis between host frequency and parasite incidence were highly significant, frequency analysis suggested that the distributions of parasites differ significantly among the four most common and closely related (same subgenus) species of hosts. A highly significant pattern of nestedness in our bipartite host-parasite network indicated that specialized parasites (i.e., those that interact with few host species) tend to occur as subsets of the most common hosts.

The influence of habitat heterogeneity and latitude on gamma diversity of the Nearctic Simuliidae, a ubiquitous group of stream-dwelling insects.

Among the most prominent, large-scale patterns of species richness are the increases in richness with decreasing latitude and with increasing habitat heterogeneity. Using the stream-dwelling larval and pupal stages of North American black flies (Diptera: Simuliidae), we address 3 broad questions about species richness: (i) Does a significant latitude-richness relationship exist? (ii) How does habitat heterogeneity influence gamma diversity? (iii) What is the sign (positive or negative) of the latitude-richness and the heterogeneity-richness relationships? We found no evidence that habitat heterogeneity influences gamma diversity. The estimated peak species richness for black flies in North America was at 50-53°N, which also corresponds with peak generic richness. All plesiomorphic, extant lineages of the Simuliidae in the Western Hemisphere are found in cool mountainous environments of North America, suggesting that peak richness at 50-53°N might be a signature of this phylogenetic pattern and a reflection of underlying historical processes.

Biodiversity of Simulium metallicum sensu lato (Diptera: Simuliidae), a complex of Neotropical vectors associated with human onchocerciasis.

The polytene chromosomes of 130 larvae of the Neotropical Simulium metallicum complex from Brazil, Costa Rica, and Ecuador revealed five cytoforms, including three ('M', 'N', and 'O') that are new and two ('B' and 'J') that represent range extensions of up to 850km. The discovery of three new cytoforms brings the total number in the complex to 17. Cytoforms 'B', 'J', and 'N' are reproductively isolated from one another, and their species status is corroborated by morphological evidence. None of the three new cytoforms is known from current or historical onchocerciasis foci, although 'M' inhabits the periphery of the former Ecuadorian Santiago onchocerciasis focus a mere 30km to the west. The number of fixed chromosomal differences, as many as 24, separating some members of the S. metallicum complex far exceeds that known between members of any other simuliid species complex. Two distinct groupings can be diagnosed within the S. metallicum complex, based on at least eight fixed chromosomal rearrangements and structural characters in the larval stage. Consequently, a recommendation is made to recognize the S. horacioi complex and the S. metallicum complex sensu stricto. Recognition of two separate complexes provides potential phylogenetic content with predictive power for understanding biological phenomena such as vector potential.

The roles of abiotic factors, dispersal, and species interactions in structuring stream assemblages of black flies (Diptera: Simuliidae).

BACKGROUND: The patterns and drivers of species assemblages represent the core of community ecology. We focus on the assemblages of a single family of ubiquitous lotic insects, the Simuliidae (black flies), of which the larvae play a critical role in resource turnover in steams. We use Mantel tests and null models to tease out the potential influence of abiotic stream conditions, species interactions, and dispersal on the assemblage patterns of larval black flies over two spatial scales (within and across ecoregions) and two seasons (spring and summer). RESULTS: When stream sites were considered across ecoregions in the spring, stream conditions and dispersal were correlated significantly with species similarity; however, within ecoregions in the spring, dispersal was important only in the Piedmont and Sandhills and abiotic factors only in the Mountains. In contrast, results of the summer analyses within and across ecoregions were congruent; assemblage similarity was significantly correlated with stream conditions both across and within ecoregions. Null models suggested that patterns of species segregation in the spring were consistent with a community structured by competition, whereas patterns in the summer were consistent with species assemblages influenced by abiotic factors. CONCLUSIONS: Species composition of black flies at streams sites is correlated with dispersal factors and stream conditions, but results vary over spatial and temporal scales. Communities of black flies can be viewed within a metacommunity context; local assemblages are consistent with species sorting and mass effects. Given that black flies have a terrestrial stage, with females deciding where to place the eggs, a full understanding of the processes that determine local aquatic assemblages will require integration of the dynamics of the aquatic immature stages and the terrestrial adults.

Predicting occurrence of the fungal symbiote Harpella colonizing black fly larvae in coastal streams of Alabama and Mississippi, USA.

The environmental conditions governing symbioses are poorly known in aquatic systems. Stream conditions associated with the distribution of the black fly (Simuliidae) midgut symbiote Harpella were investigated in southern Alabama and Mississippi streams. Stream conditions that were most useful in predicting the distribution of Harpella spp. in the study area were dissolved oxygen and water temperature. Presence of Harpella species in streams was associated with higher dissolved oxygen and decreased water temperature compared to streams where Harpella spp. was absent. Stream conditions associated with the distribution of Harpella spp. in other regions of the world vary according to conditions other than those elucidated here, indicating that geography, host species, and stream conditions play important roles in the spatial distribution of Harpella species.

Morphological differences of symbiotic fungi Smittium culisetae (Harpellales: Legeriomycetaceae) in different Dipteran hosts.

Harpellales (Legeriomycetaceae, Zygomycota) or 'trichomycetes' are fungi that inhabit the digestive tracts of arthropods such as insects, millipedes, and crustaceans. In the current study we examined changes in 5 morphological characters of Smittium culisetae (Harpellales: Legeriomycetaceae) between the two dipteran (mosquito, black fly) hosts reared under 3 different temperatures (17, 22, 30 degrees C). Both host and temperature had a pervasive effect on the linear dimension of trichospores, their generative cells and hyphae width. At 30 degrees C the mean size of all 5 morphological characters were consistently larger in fungus taken from the mosquito host than from the black fly host. At 17 degrees C and 22 degrees C, however, there were no consistent patterns. The effect of host was so pronounced that it could be accurately determined which host S. culisetae colonised based on differences in linear morphology. Such changes in fungal morphology between hosts have important ramifications for the morphologically based taxonomy of this group.

Do different species of Smittium (Harpellales, Legeriomycetaceae) influence each other in the host gut?

Smittium (Harpellales, Legeriomycetaceae) belongs to a cosmopolitan group of filamentous fungi (Trichomycetes, Zygomycota) that live as obligate commensals in the digestive tract of various marine, freshwater, and terrestrial arthropods. The outcome of the paired introductions of three species of Smittium was investigated within the individual hosts of the mosquito Aedes aegypti (Culicidae: Diptera). In the first set of experiments, the host was inoculated with a single species of Smittium to determine whether hyphae location within the host was species specific. In the second experiment the host was exposed to two species of Smittium to determine whether hyphae showed positional displacement when two species of fungi co-inhabited the same host gut. Single species introductions of Smittium resulted in 80-85% of hosts with hyphae present only in the rectum. In all three paired combinations of Smittium species examined, only 40-65% of host larvae had hyphae restricted to the rectum. This is first study to experimentally demonstrate that the microdistribution of Harpellaceae hyphae can be influenced by the presence of a second species of Harpellaceae, suggesting a competition of the symbionts within the host.

The effect of temperature and host species on the development of the trichomycete Smittium culisetae (Zygomycota).

We examined the growth and development of the trichomycete Smittium culisetae (Harpellales: Legeriomycetaceae) in the larval hosts Simulium vittatum Zetterstedt (Diptera: Simuliidae) and Aedes aegypti (L.) (Diptera: Culicidae) at three temperatures, 17, 22 and 30 C. Trichospore maturation of Sm. culicetae external to the host as well as the ability of these trichospores to colonize new hosts also was investigated. Although the development of Sm. culisetae varied with both temperature and host there was a pattern of maximum trichospore production at 48-72 h postinoculation. In addition thalli under laboratory conditions are capable of spore production after extraction from a host and these trichospores can colonize new hosts. Furthermore this was noted to occur in both host species. These results suggest that synchrony between host and symbiont development is not as tightly coupled as previously assumed.

In situ studies of algal biomass in relation to physicochemical characteristics of the Salt Plains National Wildlife Refuge, Oklahoma, USA.

This is the first in a series of experiments designed to characterize the Salt Plains National Wildlife Refuge (SPNWR) ecosystem in northwestern Oklahoma and to catalogue its microbial inhabitants. The SPNWR is the remnant of an ancient ocean, encompassing approximately 65 km2 of variably hypersaline flat land, fed by tributaries of the Arkansas River. Relative algal biomass (i.e., chlorophyll concentrations attributed to Chlorophyll-a-containing oxygenic phototrophs) and physical and chemical parameters were monitored at three permanent stations for a one-year period (July 2000 to July 2001) using a nested block design. Salient features of the flats include annual air temperatures that ranged from -10 to 40 degrees C, and similar to other arid/semi-arid environments, 15-20-degree daily swings were common. Shade is absent from the flats system; intense irradiance and high temperatures (air and sediment surface) resulted in low water availability across the SPNWR, with levels of only ca. 15 % at the sediment surface. Moreover, moderate daily winds were constant (ca. 8-12 km h-1), sometimes achieving maximum speeds of up to 137 km h-1. Typical of freshwater systems, orthophosphate (PO(4)3-) concentrations were low, ranging from 0.04 to <1 microM; dissolved inorganic nitrogen levels were high, but spatially variable, ranging from ca. 250-600 microM (NO(3)- + NO(2)-) and 4-166 microM (NH(4)+). Phototroph abundance was likely tied to nutrient availability, with high-nutrient sites exhibiting high Chl-a levels (ca. 1.46 mg m-2). Despite these harsh conditions, the phototrophic microbial community was unexpectedly diverse. Preliminary attempts to isolate and identify oxygenic phototrophs from SPNWR water and soil samples yielded 47 species from 20 taxa and 3 divisions. Our data indicate that highly variable, extreme environments might support phototrophic microbial communities characterized by higher species diversity than previously assumed.

Laboratory investigations of trichomycete prevalence, abundance and fecundity in a Smittium-simuliid model.

Smittium, the most speciose genus of the "gut fungi" (Zygomycota: Trichomycetes), is found attached to the hindgut cuticle of larval aquatic Diptera. Smittium spp. colonize several host families (e.g., Smittium culisetae in Chironomidae, Culicidae and Simuliidae), but some species appear to be specific to a single host family (e.g., Smittium morbosum Sweeney in Culicidae). The specificity of Smittium spp. within a host family has been difficult to resolve. This research presents evidence that certain Smittium spp. differentially colonize particular species of black fly (Diptera: Simuliidae) hosts as measured by differences in prevalence, abundance and fecundity. Reasons for this differential occurrence and fecundity in hosts are unclear but might include fungal responses to variations in host morphology, physiology, distribution or behavior. Variable fitness of Smittium spp., within a suite of available hosts, could be a factor in the diversity of this fungal group.

First report of a trichomycete fungus (Zygomycota: Trichomycetes) inhabiting larvae of Simulium ochraceum sensu lato Walker (Diptera: Simuliidae) from the Galapagos Islands.

A collection of the larval black fly Simulium ochraceum sensu lato Walker was made at six lotic habitats on San Cristobal Island, Galapagos Islands, Ecuador, during May 2003. Sixty-eight larvae were assayed for the presence of gut fungi (Zygomycota: Trichomycetes). A trichomycete, designated as Smittium sp. was found inhabiting the hindgut of 17.6% of S. ochraceum s.l. larvae. Implications of this finding with respect to both host and trichomycete biogeography are discussed.

Prevalence of the trichomycete fungus Harpella melusinae (Harpellales: Harpellaceae) in larval black flies (Diptera: Simuliidae) across a heterogeneous environment.

A total of 2063 mid- to late-instar larval black flies were collected from 64 stream sites in South Carolina and screened for the presence of the trichomycete fungus Harpella melusinae. Sixteen of 18 host species were colonized by H. melusinae on at least one occasion. Prevalence of H. melusinae in larvae of Simulium tuberosum cytospecies "A" was highest in acidic streams with low conductivity, whereas H. melusinae colonized larvae of Simulium verecundum most frequently in slower-moving streams. Ecological conditions, therefore, can serve as predictors of the prevalence of H. melusinae. Prevalence in host larvae was significantly lower in the Piedmont ecoregion than in the Mountain ecoregion. Prevalence did not differ in the host species S. verecundum across ecoregions, suggesting that different prevalences among host species might indicate some host preference. The prevalence of H. melusinae differed significantly between two univoltine host species (Simulium venustum and Prosimulium magnum) at the same site but not between two multivoltine host species (S. tuberosum cytospecies "FG" and S. tuberosum cytospecies "CDE"), suggesting that host life history could be important in determining fungal prevalence.

The microdistribution of the trichomycete Smittium culisetae in the hindgut of the black fly host Simulium vittatum.

We examined the distribution of hyphae of the trichomycete fungus Smittium culisetae (Harpellales: Legeriomycetaceae) in the hindgut of a larval black fly (Simulium vittatum, cytospecies IS-7) by analyzing its prevalence and relative abundance. Hyphal prevalence was highest in the posterior colon (93.1%) and rectum (86.3%), with low prevalence (12.0%) in the anterior colon. Relative abundance of hyphae was highest in the posterior colon, followed by the rectum; relative abundance of hyphae in the anterior colon was lower. Hyphae of S. culisetae were not observed in the pylorus. We used a novel method of quantifying the relative abundance of S. culisetae in the host hindgut. The hindgut was observed with an ocular grid, and abundance was expressed as the ratio of grids occupied by hyphae to the number of grids occupied by hindgut.

Scale, time, space, and predictability: species distributions of preimaginal black flies (Diptera: Simuliidae).

This study examines species-specific distributions of preimaginal black flies (Diptera: Simuliidae) over two large spatial scales (within and across ecoregions) and two seasons (spring and summer). The study area in the western two-thirds of South Carolina, United States, was divided into three ecoregions (Mountains, Piedmont, Sandhills). The mean correct classification of species distributions among stream sites was 81.3%. Predictability of species distributions varied with spatial scale, location, and time. Species occurrence was not independent of ecoregion; distributions of 12 of the 13 species examined at this scale were predictable on the basis of changes in water column (temperature, percentage dissolved oxygen) and channel characters (velocity, streambed-particle size). However, with the exception of the Mountains during the summer, predicting species distributions within ecoregions was far less successful than predicting distributions across these regions; predictability was particularly poor in the Sandhills. Seasonal effects on predictability were most pronounced in the Mountains. As stream sites became more homogeneous, predictive capability declined. Species-level identifications, aided by cytogenetic techniques, were of paramount importance in recognizing the association between species distributions and stream conditions, emphasizing that considerable ecological information is lost when species are not considered. This study emphasizes the critical role of taxonomic resolution, linked with spatial and temporal scale of observation, in elucidating patterns of species assemblages. It also demonstrates that predictability of species distributions at a fixed scale can vary with geographic location and time.