A total of 48 strains of thin, filamentous cyanobacteria in Synechococcales were studied by sequencing 16S rRNA and rpoC1 sequence fragments. We also carefully characterized a subset of these by morphology. Phylogenetic analysis of the 16S rRNA gene data using Bayesian inference of a large Synechococcales alignment (345 OTU’s) was in agreement with the phylogeny based on the rpoC1 gene for 59 OTU’s. Both indicated that the large family-level grouping formerly classified as the Leptolyngbyaceae could be further divided into four family-level clades. Two of these family-level clades have been recognized previously as Leptolyngbyaceae and Prochlorotrichaceae. Oculatellaceae fam. nov. and Trichocoleaceae fam. nov. are proposed for the other two families. The Oculatellaceae was studied in greater detail, and six new genera containing 14 species were characterized and named. These new taxa are: Pegethrix botrychoides, P. olivacea, P. convoluta, P. indistincta, Drouetiella lurida, D. hepatica, D. fasciculata, Cartusia fontana, Tildeniella torsiva, T. nuda, Komarkovaea angustata, Kaiparowitsia implicata, Timaviella obliquedivisa, and T. radians.
A total of 27 strains of Oculatella were isolated, characterized and sequenced, and analysed phylogenetically with an additional environmental clone from the Atacama Desert and 10 strains isolated and sequenced by others. The strains were clearly separated based upon phylogenetic analyses conducted with a concatenated alignment of the 16S rRNA and 16S-23S ITS region of the ribosomal operons in the genus Oculatella. Differences in secondary structure of the conserved domains of the ITS region, as well as comparative analysis of P-distance among ITS regions, served to separate the strains into distinct taxonomic units. Seven new species of Oculatella were described, including four from arid to semi-arid soils (O. atacamensis, O. mojaviensis, O. coburnii, O. neakameniensis) and three from more mesic habitats, including a temperate lake (O. hafneriensis), a desert waterfall (O. cataractarum) and a Hawaiian sea cave (O. kauaiensis). The soil forms show statistically significant morphological differences, but the ranges overlap to a degree that they are not diagnosable by morphology, and these four cryptic species are characterized here using molecular characters. The more mesic species, including the type species from Mediterranean hypogea, O. subterranea, are all morphologically distinct from each other and from all four soil taxa. This report is the first to use solely molecular criteria to distinguish cryptic species of cyanobacteria.
Enterococcal spondylitis (ES) is a disease of commercial broiler chickens, with a worldwide distribution. Symmetrical hind limb paralysis typical of ES results from infection of the free thoracic vertebra (FTV) by pathogenic strains of Enterococcus cecorum . To determine the pathogenesis of ES, birds with natural and experimental ES were studied over time. In natural disease, case birds (n = 150) from an affected farm and control birds (n = 100) from an unaffected farm were evaluated at weeks 1-6. In control birds, intestinal colonization by E. cecorum began at week 3. In case birds, E. cecorum was detected in intestine and spleen at week 1, followed by infection of the FTV beginning at week 3. E. cecorum isolates recovered from intestine, spleen, and FTV of case birds had matching genotypes, confirming that intestinal colonization with pathogenic strains precedes bacteremia and infection of the FTV. Clinical intestinal disease was not required for E. cecorum bacteremia. In 1- to 3-week-old case birds, pathogenic E. cecorum was observed within osteochondrosis dissecans (OCD) lesions in the FTV. To determine whether OCD of the FTV was a risk factor for ES, 214 birds were orally infected with E. cecorum, and the FTV was evaluated histologically at weeks 1-7. Birds without cartilage clefts of OCD in the FTV did not develop ES; while birds with OCD scores ≥3 were susceptible to lesion development. These findings suggest that intestinal colonization, bacteremia, and OCD of the FTV in early life are crucial to the pathogenesis of ES.
Enterococcus cecorum, a normal intestinal inhabitant, is increasingly responsible for outbreaks of arthritis and osteomyelitis in chickens worldwide. Enterococcal spondylitis (ES) is a specific manifestation of E. cecorum-associated disease in which increased flock morbidity and mortality result from chronic infection involving the free thoracic vertebra. In this study the genetic relatedness and antimicrobial resistance of isolates recovered from ES-affected flocks in the southeastern United States were determined. ES outbreaks from 2007 to 2011 were investigated in North Carolina (15 flocks, 13 farms, four integrators), South Carolina (one flock, one farm, one integrator) and Alabama (six flocks, six farms, one integrator). From these 22 epidemiologically distinct outbreaks, 326 isolates of E. cecorum were recovered. Isolates from spinal lesions and caeca of affected birds (cases) and caeca of unaffected birds (controls) were genotyped using pulsed-field gel electrophoresis; phenotyped using both GenIII MicroPlate TM (Biolog; Hayward, CA, USA) microbial identification plates and antimicrobial sensitivity testing; and compared with each other. Isolates from spinal lesions were incapable of mannitol metabolism and the majority of these isolates were genetically clonal. In contrast, caecal isolates from control birds varied in their ability to metabolize mannitol and were genetically diverse. Isolates from both case and control birds had high levels of antimicrobial resistance. These findings indicate that the increase in E. cecorum-associated disease in the southeast United States is due to the emergence of new clones with increased pathogenicity and multidrug resistance.
The objective of this cross-sectional study was to compare the prevalence of Salmonella and antimicrobial-resistant Salmonella, as well as investigate the distribution of this pathogen in organic and conventional broiler poultry farms. Fecal (n = 420), feed (n = 140), and drinking water (n = 140) samples were collected from birds at 3 and 8 weeks of age for 2-flock cycles. One house was sampled per farm at three organic and four conventional broiler farms from the same company in North Carolina. All samples were analyzed for the presence of Salmonella using selective enrichment techniques. Further phenotypic (antimicrobial susceptibility) and genotypic (pulsed-field gel electrophoresis [PFGE]) testing were performed. Salmonella prevalences in fecal samples were 5.6% (10/180) and 38.8% (93/240) from organic and conventional farms, respectively. From feed, 5.0% (3/60) and 27.5% (22/80) of the samples were positive for Salmonella from organic and conventional farms, respectively. None of the water samples were positive for Salmonella. Seventy isolates were characterized by antimicrobial susceptibility and PFGE types. The two most common resistance phenotypes were single resistance to streptomycin (36.2% [25/58]: conventional; 25% [3/12] organic), and multidrug resistance to six antimicrobial agents: ampicillin-streptomycin-amoxicillin/clavulanic acid-cephalothin-ceftiofur-cefoxitin (AmStAxChCfFx; 39.7%: conventional only). Genotypic analysis using PFGE showed clonality among isolates within and between the two types of farms. The results of our study suggest that within this poultry company, the prevalence of fecal Salmonella was lower in certified-organic birds than in conventionally raised birds, and the prevalence of antimicrobial-resistant Salmonella was also higher in conventionally raised birds than in certified-organic birds.
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