Assessing the faecal source sensitivity and specificity of ruminant and human genetic microbial source tracking markers in the central Ethiopian highlands

This study tested genetic microbial source tracking (MST) methods for identifying ruminant- (BacR) and human-associated (HF183/BacR287, BacHum) bacterial faecal contaminants in Ethiopia in a newly created regional faecal sample bank (n = 173). BacR performed well, and its marker abundance was high (100% sensitivity (Sens), 95% specificity (Spec), median log₁₀ 8·1 marker equivalents (ME) g⁻¹ ruminant faeces). Human-associated markers tested were less abundant in individual human samples (median: log₁₀ 5·4 and 4·2 (ME + 1) g⁻¹) and were not continuously detected (81% Sens, 91% Spec for BacHum; 77% Sens, 91% Spec for HF183/BacR287). Furthermore, the pig-associated Pig2Bac assay was included and performed excellent (100% Sens, 100% Spec). To evaluate the presence of MST targets in the soil microbiome, representative soil samples were tested during a whole seasonal cycle (n = 60). Only BacR could be detected, but was limited to the dry season and to sites of higher anthropogenic influence (log₁₀ 3·0 to 4·9 (ME + 1) g⁻¹ soil). In conclusion, the large differences in marker abundances between target and non-target faecal samples (median distances between distributions ≥log₁₀ 3 to ≥log₁₀ 7) and their absence in pristine soil indicate that all tested assays are suitable candidates for diverse MST applications in the Ethiopian area.     

Relationship of Vibrio Species Infection and Elevated Temperatures to Yellow Blotch/Band Disease in Caribbean Corals

The bacterial and temperature factors leading to yellow blotch/band disease (YBD), which affects the major reef-building Caribbean corals Montastrea spp., have been investigated. Groups of bacteria isolated from affected corals and inoculated onto healthy corals caused disease signs similar to those of YBD. The 16S rRNA genes from these bacteria were sequenced and found to correspond to four Vibrio spp. Elevating the water temperature notably increased the rate of spread of YBD on inoculated corals and induced greater coral mortality. YBD-infected corals held at elevated water temperatures had 50% lower zooxanthella densities, 80% lower division rates, and a 75% decrease in chlorophyll a and c₂ pigments compared with controls. Histological sections indicated that the algal pyrenoid was fragmented into separate segments, along with a reconfiguration and swelling of the zooxanthellae, as well as vacuolization. YBD does not appear to produce the same physiological response formerly observed in corals undergoing temperature-related bleaching. Evidence indicates that YBD affects primarily the symbiotic algae rather than coral tissue.     

Multiple-antibiotic resistance of Enterococcus spp. isolated from commercial poultry production environments

The potential impact of food animals in the production environment on the bacterial population as a result of antimicrobial drug use for growth enhancement continues to be a cause for concern. Enterococci from 82 farms within a poultry production region on the eastern seaboard were isolated to establish a baseline of susceptibility profiles for a number of antimicrobials used in production as well as clinical environments. Of the 541 isolates recovered, Enterococcus faecalis (53%) and E. faecium (31%) were the predominant species, while multiresistant antimicrobial phenotypes were observed among all species. The prevalence of resistance among isolates of E. faecalis was comparatively higher among lincosamide, macrolide, and tetracycline antimicrobials, while isolates of E. faecium were observed to be more frequently resistant to fluoroquinolones and penicillins. Notably, 63% of the E. faecium isolates were resistant to the streptogramin quinupristin-dalfopristin, while high-level gentamicin resistance was observed only among the E. faecalis population, of which 7% of the isolates were resistant. The primary observations are that enterococci can be frequently isolated from the poultry production environment and can be multiresistant to antimicrobials used in human medicine. The high frequency with which resistant enterococci are isolated from this environment suggests that these organisms might be useful as sentinels to monitor the development of resistance resulting from the usage of antimicrobial agents in animal production.     

Hyaline cartilage tissue is formed through the co-culture of passaged human chondrocytes and primary bovine chondrocytes

To circumvent the problem of a sufficient number of cells for cartilage engineering, the authors previously developed a two-stage culture system to redifferentiate monolayer culture-expanded dedifferentiated human articular chondrocytes by co-culture with primary bovine chondrocytes (bP0). The aim of this study was to analyze the composition of the cartilage tissue formed in stage 1 and compare it with bP0 grown alone to determine the optimal length of the co-culture stage of the system. Biochemical data show that extracellular matrix accumulation was evident after 2 weeks of co-culture, which was 1 week behind the bP0 control culture. By 3 to 4 weeks, the amounts of accumulated proteoglycans and collagens were comparable. Expression of chondrogenic genes, Sox 9, aggrecan, and collagen type II, was also at similar levels by week 3 of culture. Immunohistochemical staining of both co-culture and control tissues showed accumulation of type II collagen, aggrecan, biglycan, decorin, and chondroitin sulfate in appropriate zonal distributions. These data indicate that co-cultured cells form cartilaginous tissue that starts to resemble that formed by bP0 after 3 weeks, suggesting that the optimal time to terminate the co-culture stage, isolate the now redifferentiated cells, and start stage 2 is just after 3 weeks.     

The mutational burden of acral melanoma revealed by whole‐genome sequencing and comparative analysis

Acral melanoma is a subtype of melanoma with distinct epidemiological, clinical and mutational profiles. To define the genomic alterations in acral melanoma, we conducted whole‐genome sequencing and SNP array analysis of five metastatic tumours and their matched normal genomes. We identified the somatic mutations, copy number alterations and structural variants in these tumours and combined our data with published studies to identify recurrently mutated genes likely to be the drivers of acral melanomagenesis. We compared and contrasted the genomic landscapes of acral, mucosal, uveal and common cutaneous melanoma to reveal the distinctive mutational characteristics of each subtype.     

Effectiveness of Partial Sedation to Reduce Stress in Captured Mule Deer

Information garnered from the capture and handling of free-ranging animals helps advance understanding of wildlife ecology and can aid in decisions on wildlife management. Unfortunately, animals may experience increased levels of stress, injuries, and death resulting from captures (e.g., exertional myopathy, trauma). Partial sedation is a technique proposed to alleviate stress in animals during capture, yet efficacy of partial sedation for reducing stress and promoting survival post-capture remains unclear. We evaluated the effects of partial sedation on physiological, biochemical, and behavioral indicators of acute stress and probability of survival post-capture for mule deer (Odocoileus hemionus) that were captured via helicopter net-gunning in the eastern Greater Yellowstone Ecosystem, Wyoming, USA. We administered 10–30 mg of midazolam and 15 mg of azaperone intramuscularly (IM) to 32 mule deer in 2016 and 53 mule deer in 2017, and maintained a control group (captured but not sedated) of 38 mule deer in 2016 and 54 mule deer in 2017. To evaluate indicators of acute stress, we measured heart rate, blood-oxygen saturation, body temperature, respiration rate, and levels of serum cortisol. We recorded number of kicks and vocalizations of deer during handling and evaluated behavior during release. We also measured levels of fecal glucocorticoids as an indicator of baseline stress. Midazolam and azaperone did not reduce physiological, biochemical, or behavioral indicators of acute stress or influence probability of survival post-capture. Mule deer that were administered midazolam and azaperone, however, were more likely to hesitate, stumble or fall, and walk during release compared with individuals in the control group, which were more likely to trot, stot, or run without stumbling or falling. Our findings suggest that midazolam (10–30 mg IM) and azaperone (15 mg IM) may not yield physiological or demographic benefits for captured mule deer as previously assumed and may pose adverse effects that can complicate safety for captured animals, including drug-induced lethargy. Although we failed to find efficacy of midazolam and azaperone as a method for reducing stress in captured mule deer, the efficacy of midazolam and azaperone or other combinations of partial sedatives in reducing stress may depend on the dose of tranquilizer, study animal, capture setting, and how stress is defined. © 2020 The Wildlife Society.     

Positional relationships between photoperiod response QTL and photoreceptor and vernalization genes in barley

Winterhardiness has three primary components: photoperiod (day length) sensitivity, vernalization response, and low temperature tolerance. Photoperiod and vernalization regulate the vegetative to reproductive phase transition, and photoperiod regulates expression of key vernalization genes. Using two barley mapping populations, we mapped six individual photoperiod response QTL and determined their positional relationship to the phytochrome and cryptochrome photoreceptor gene families and the vernalization regulatory genes HvBM5A, ZCCT-H, and HvVRT-2. Of the six photoreceptors mapped in the current study (HvPhyA and HvPhyB to 4HS, HvPhyC to 5HL, HvCry1a and HvCry2 to 6HS, and HvCry1b to 2HL), only HvPhyC coincided with a photoperiod response QTL. We recently mapped the candidate genes for the 5HL VRN-H1 (HvBM5A) and 4HL VRN-H2 (ZCCT-H) loci, and in this study, we mapped HvVRT-2, the barley TaVRT-2 ortholog (a wheat flowering repressor regulated by vernalization and photoperiod) to 7HS. Each of these three vernalization genes is located in chromosome regions determining small photoperiod response QTL effects. HvBM5A and HvPhyC are closely linked on 5HL and therefore are currently both positional candidates for the same photoperiod effect. The coincidence of photoperiod-responsive vernalization genes with photoperiod QTL suggests vernalization genes should also be considered candidates for photoperiod effects.     

Environmental maternal influences on body composition in mice selected for body weight

Summary The effect of the postnatal maternal environment, simulated by rearing mice in litters of three, six or nine, on body weight and body composition was investigated in three lines of mice differing widely in growth rate. The lines were selected for high (H₆) and low (L₆) 6-week body weight while the control line was maintained by random selection. Body weight and weights and percentages of ether extract, water, ash and protein at 21, 42, 63 and 84 days were recorded. With few exceptions, there were positive correlated responses to selection in body weight and in weights of body components. At 21 and 42 days the correlated responses were larger in L₆ mice than in H₆ mice. Body weight and weights of body components were larger for mice reared in litters of three than for those reared in litters of nine. Also, mice reared in litters of six were intermediate in body weight and weights of some of the body components between those reared in litters of three and nine. Differences in body weight and weights of body components due to postnatal maternal environment were small by comparison with differences due to genetic line. There were significant line by maternal environment interactions in body weight at 21 days and in ether extract weight at 21 and 63 days. Line and maternal environment differences in percentages of body components did not follow any consistent trend. The results for percentages of body components were further complicated by line x maternal environment interactions. In general, both line and postnatal maternal environmental differences in percentages of body components diminished with age.Paper No. 5670 of the Journal Series of the North Carolina Agricultural Experiment Station, Raleigh, North Carolina 27650. The use of trade names in this publication does not imply endorsement by the North Carolina Agricultural Experiment Station of the products named, nor criticism of similar ones not mentioned     

Polyadenylation accelerates degradation of chloroplast mRNA.

The expression of chloroplast genes is regulated by several mechanisms, one of which is the modulation of RNA stability. To understand how this regulatory step is controlled during chloroplast development, we have begun to define the mechanism of plastid mRNA degradation. We show here that the degradation petD mRNA involves endonucleolytic cleavage at specific sites upstream of the 3' stem-loop structure. The endonucleolytic petD cleavage products can be polyadenylated in vitro, and similar polyadenylated RNA products are detectable in vivo. PCR analysis of the psbA and psaA-psaB-rps14 operons revealed other polyadenylated endonucleolytic cleavage products, indicating that poly(A) addition appears to be an integral modification during chloroplast mRNA degradation. Polyadenylation promotes efficient degradation of the cleaved petD RNAs by a 3'-5' exoribonuclease. Furthermore, polyadenylation also plays an important role in the degradation of the petD mRNA 3' end. Although the 3' end stem-loop is usually resistant to nucleases, adenylation renders the secondary structure susceptible to the 3'-5' exoribonuclease. Analysis of 3' ends confirms that polyadenylation occurs in vivo, and reveals that the extent of adenylation increases during the degradation of plastid mRNA in the dark. Based on these results, we propose a novel mechanism for polyadenylation in the regulation of plastid mRNA degradation.