Development of a real-time TaqMan assay to detect <Emphasis Type="Italic">mendocina</Emphasis> sublineage <Emphasis Type="Italic">Pseudomonas</Emphasis> species in contaminated metalworking fluids

A TaqMan quantitative real-time polymerase chain reaction (qPCR) assay was developed for the detection and enumeration of three Pseudomonas species belonging to the mendocina sublineage (P. oleovorans, P. pseudoalcaligenes, and P. oleovorans subsp. lubricantis) found in contaminated metalworking fluids (MWFs). These microbes are the primary colonizers and serve as indicator organisms of biodegradation of used MWFs. Molecular techniques such as qPCR are preferred for the detection of these microbes since they grow poorly on typical growth media such as R2A agar and Pseudomonas isolation agar (PIA). Traditional culturing techniques not only underestimate the actual distribution of these bacteria but are also time-consuming. The primer–probe pair developed from gyrase B (gyrB) sequences of the targeted bacteria was highly sensitive and specific for the three species. qPCR was performed with both whole cell and genomic DNA to confirm the specificity and sensitivity of the assay. The sensitivity of the assay was 10¹ colony forming units (CFU)/ml for whole cell and 13.7 fg with genomic DNA. The primer–probe pair was successful in determining concentrations from used MWF samples, indicating levels between 2.9 × 10³ and 3.9 × 10⁶ CFU/ml. In contrast, the total count of Pseudomonas sp. recovered on PIA was in the range of <1.0 × 10¹ to 1.4 × 10⁵ CFU/ml for the same samples. Based on these results from the qPCR assay, the designed TaqMan primer–probe pair can be efficiently used for rapid (within 2 h) determination of the distribution of these species of Pseudomonas in contaminated MWFs.     

The analysis of the joint effect of substances on reversion systems and the assessment of antimutagenicity.

The statistical methods for the analysis of mutagenicity and carcinogenicity underwent considerable theoretical-practical development following the need for assessing the mutagenic and carcinogenic potential of substances. Antimutagenicity is investigated through the analysis of respondents in dose-response assays, when two different molecules are administered separately and as a mixture to a respondent system. When the number of respondent units is high, and doses are orthogonal, it is possible to apply simple models such as analysis of variance. This is not always possible or common, and alternative approaches have been developed, based on multiple regression and on tables of proportions. In this work, some of the most frequently used methods for the assessment of joint responses are reviewed, particularly those based on multiple regression, such as the method of Shaeffer et al. and the method of Hass et al. In order to illustrate these methods, joint responses of perylene and cyclopentapyrene, of N-acetylcysteine and dinitropyrene, and of N-acetylcysteine and extracts from diesel exhausts were analyzed. An antagonistic effect of perylene on the action of CPP was detected by the algorithm of Shaeffer et al. The effect is not multiplicative, i.e., it is not proportional to the product of doses. The antimutagenic effect of N-acetylcysteine on dinitropyrene is multiplicative, as detected by the method of Hass et al. The latter reveals that the inhibition by N-acetylcysteine on the mutagenic effect of extracts from diesel exhausts is also multiplicative.     

A simple procedure to obtain one-micrometer sections of routinely embedded paraffin material

By freezing blocks of paraffin-embedded tissues to a convenient temperature it is possible to obtain routinely 1 micron sections that can be further processed as normal thicker sections. Normal and disposable steel knives can be used and the staining time should be increased in most procedures. Gradual freezing of blocks to the temperature of dry ice is the simplest and safest way to obtain an adequate temperature. The best results were obtained using as fixative 4% paraformaldehyde in phosphate buffered saline solution.     

Relationship Between Solute Permeability and Osmotic Remediability in a Galactose-Negative Strain of Saccharomyces cerevisiae

An osmotic remedial allele, gal 7-1, in the galactose pathway of Saccharomyces cerevisiae responds to either penetrating (ethylene glycol and diethylene glycol) or nonpenetrating (KCl, NaCl, and sorbitol) solutes in the growth medium. Extracts from cells grown under restrictive conditions gave no increase in enzyme activity (gal-1-phosphate, uridylyl transferase) when exposed to the penetrating solutes; thus protein synthesis or possibly polymer assembly is proposed as the critical step remedied by the addition of the solutes.     

Haematobia irritans parasitism of F1 yak × beef cattle (Bos grunniens × Bos taurus) hybrids

The horn fly Haematobia irritans (Diptera: Muscidae) is a blood obligate ectoparasite of bovids that causes annual losses to the U.S. beef cattle industry of over US$1.75 billion. Climate warming, the anthropogenic dispersion of bovids and the cross‐breeding of beef cattle with other bovid species may facilitate novel horn fly–host interactions. In particular, hybridizing yaks [Bos grunniens (Artiodactyla: Bovidae)] with beef cows (Bos taurus) for heterosis and carcass improvements may increase the exposure of yak × beef hybrids to horn flies. The present paper reports on the collection of digital images of commingled beef heifers (n = 12) and F1 yak × beef hybrid bovids (heifers, n = 7; steers, n = 5) near Laramie, Wyoming (～ 2200 m a.s.l.) in 2018. The total numbers of horn flies on beef heifers and F1 yak × beef heifers [mean ± standard error (SE): 88 ± 13 and 70 ± 17, respectively] did not differ significantly; however, F1 yak × beef steers had greater total horn fly abundance (mean ± SE: 159 ± 39) than female bovids. The present report of this experiment is the first such report in the literature and suggests that F1 yak × beef bovids are as susceptible as cattle to horn fly parasitism. Therefore, similar monitoring and treatment practices should be adopted by veterinarians, entomologists and producers.     

Segregation of infectious pancreatic necrosis resistance QTL in the early life cycle of Atlantic Salmon (Salmo salar)

In a previous study, three significant quantitative trait loci (QTL) associated with resistance to Infectious Pancreatic Necrosis (IPN) disease were identified by analysing challenge data from one sub-population of Landcatch Atlantic salmon (Salmo salar) smolt. While these QTL were shown to affect the resistance in seawater, their effect in freshwater was unknown. This study investigates the effect of these QTL on IPN resistance in salmon fry in freshwater. Twenty families with intermediate levels of IPN mortality were analysed from a freshwater challenge trial undertaken on a different sup-population of LNS salmon to that studied previously. Only the QTL from linkage group 21 (LG21) appeared to have a significant and large effect on resistance in freshwater; the same QTL was found to have the largest effect in seawater in the previous study. Variance component analysis showed a high heritability for the QTL: 0.45 ± 0.07 on the liability scale and 0.25 ± 0.05 on the observed scale. In a family where both parents were segregating for the QTL, there was a 0% vs. 100% mortality in homozygous offspring for resistant and susceptible QTL alleles. The finding that the same QTL has major effect in both freshwater and seawater has important practical implications, as this will allow the improvement of resistance in both phases through marker assisted selection by targeting this QTL. Moreover, the segregation of the LG21 QTL in a different sub-population gives further evidence of its association with IPN-resistance.