Multicenter Evaluation of the Clinical Performance and the Neutralizing Antibody Activity Prediction Properties of 10 High-Throughput Serological Assays Used in Clinical Laboratories.

As the coronavirus disease 2019 (COVID-19) pandemic second wave is emerging, it is of the upmost importance to screen the population immunity in order to keep track of infected individuals. Consequently, immunoassays for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with high specificity and positive predictive values are needed to obtain an accurate epidemiological picture. As more data accumulate about the immune responses and the kinetics of neutralizing-antibody (nAb) production in SARS-CoV-2-infected individuals, new applications are forecast for serological assays such as nAb activity prediction in convalescent-phase plasma from recovered patients. This multicenter study, involving six hospital centers, determined the baseline clinical performances, reproducibility, and nAb level correlations of 10 commercially available immunoassays. In addition, three lateral-flow chromatography assays were evaluated, as these devices can be used in logistically challenged areas. All assays were evaluated using the same patient panels in duplicate, thus enabling accurate comparison of the tests. Seven immunoassays examined in this study were shown to have excellent specificity (98 to 100%) and good to excellent positive predictive values (82 to 100%) when used in a low (5%)-seroprevalence setting. We observed sensitivities as low as 74% and as high as 95% at ≥15 days after symptom onset. The determination of optimized cutoff values through receiver operating characteristic (ROC) curve analyses had a significant impact on the diagnostic resolution of several enzyme immunoassays by increasing the sensitivity significantly without a large trade-off in specificity. We found that spike-based immunoassays seem to be better correlates of nAb activity. Finally, the results reported here will add to the general knowledge of the interlaboratory reproducibility of clinical performance parameters of immunoassays and provide new evidence about nAb activity prediction.

Circulating progesterone and obesity in men.

Progesterone can be detected in male plasma and has been considered to originate mainly from the adrenals. We have examined the association between circulating progesterone and obesity in a sample of thirty-eight lean to morbidly obese men aged 44.5 +/- 9.9 years (BMI: 44.3 +/- 12.8 kg/m (2)). Plasma concentrations of progesterone, 17-OH-progesterone as well as androstenedione, testosterone, DHT and DHEA-S were determined. Negative correlations were observed between plasma progesterone levels and body weight (r = - 0.47, p < 0.05), BMI (r = - 0.56, p < 0.001), waist circumference (r = - 0.58, p < 0.001), as well as subcutaneous adipocyte diameter (r = - 0.50, p < 0.05). Plasma levels of 17-OH-progesterone, DHEA-S, androstenedione, testosterone and DHT were also negatively associated with body weight, BMI and waist circumference. However, the ratio of 17-OH-progesterone-to-progesterone and androstenedione-to-17-OH-progesterone were not related to these variables. A positive correlation was found between circulating progesterone and DHEA-S levels (r = 0.50, p < 0.002 after adjustment for age). Accordingly, using multivariate regression analyses, the best steroid predictor of progesterone level was plasma DHEA-S. Waist circumference was the best predictor of progesterone levels in a multivariate model including steroid concentrations as well as waist circumference, BMI and subcutaneous adipocyte diameter. In conclusion, plasma progesterone was negatively associated with markers of obesity such as BMI, waist circumference and subcutaneous adipocyte diameter in this sample of men. Circulating DHEA-S level was the best steroid correlate of plasma progesterone. We suggest that the low progesterone levels observed in obese men may reflect decreased adrenal C(19) steroid production in the adrenal cortex. Further research is needed to confirm this hypothesis.

Airborne microflora in Quebec dairy farms: lack of effect of bacterial hay preservatives.

Pediococcus pentosaceus is a lactic-acid producing bacterium inoculated in hay to prevent hay deterioration. This study sought to verify the effect of this treatment on the barn microenvironment. Air samples were obtained from 19 barns using bacterial hay treatment and from 18 control barns with six-stage Andersen samplers and all-glass impingers. Appropriate culture media were used for the recovery and identification of microorganisms. Endotoxins were measured with chromogenic Limulus amoebocyte lysate assay. Median values (respectively for treated and untreated hay barns) were: 5.28 x 10(5) and 3.84 x 10(5) colony-forming units (CFU)/m3 for total bacteria; 3.18 x 10(6) and 4.5 x 10(6) CFU/m3 for molds; 1.36 x 10(3) and 1.74 x 10(3) endotoxin units/m3 for endotoxin levels; and 1.03 x 10(3) and 3.00 x 10(3) CFU/m3 for Saccharopolyspora rectivirgula. No viable P. pentosaceus were recovered. The presence of S. rectivirgula, the causative agent for farmer's lung, was not influenced by the hay treatment. Since no significant difference was observed in any of the airborne contaminants, this type of hay treatment probably does not protect farmers from the respiratory effect of ambient microbial contaminants.

Equilibrium model in an in vitro poly(ADP-ribose) turnover system.

Poly(ADP-ribose) metabolism plays an important role in numerous DNA-related functions. This homopolymer is synthesized by poly(ADP-ribose) polymerase and is degraded mainly by the poly(ADP-ribose) glycohydrolase. The activities of these two enzymes in the nucleus are closely coordinated. To better understand the interactions between these enzymes, we designed an in vitro system in which both enzymes are present at the same time. In this work, we report a model describing the synthesis and degradation of the poly(ADP-ribose) in turnover conditions. Because the half-life of the polymer in the cell is close to 1 min, we studied the very early kinetic interactions of these two enzymes.

Biochemical properties and function of poly(ADP-ribose) glycohydrolase.

We describe here the latest observations on poly(ADP-ribose) glycohydrolase. There is now extensive evidence that this nuclear enzyme is an endo-exoglycosidase which has a key role to perform in the removal of polymers which interact with proteins through covalent and non-covalent interactions. Also, we have developed a zymogram which will permit the isolation of the various isoforms of the glycohydrolase and the eventual cloning of this enzyme. Finally, we have evidence that very short oligomers and even monomers of ADP-ribose covalently bound to proteins can be removed by poly(ADP-ribose) glycohydrolase.

Mode of action of poly(ADP-ribose) glycohydrolase.

The turnover of the homopolymer of ADP-ribose, which is known to be involved in many DNA-related functions, is controlled by 2 principal enzymes. Poly(ADP-ribose) polymerase (EC 2.4.2.30) synthesizes the polymer from NAD, and poly(ADP-ribose) glycohydrolase (PARG) is the major enzyme responsible for its catabolism (Thomassin et al. (1992) Biochim. Biophys. Acta 1137, 171-181). In vivo, poly(ADP-ribose) polymers constitute a heterogeneous population of branched polymers attaining sizes of 200-400 residues. They are rapidly degraded by PARG, displaying variable kinetic parameters as a function of polymer size. Several studies have suggested that PARG acts exoglycosidically on its substrate but others observed that it could act endo/exo-glycosidically. We analysed the mode of action of PARG under conditions most suitable for expression of all the activities of PARG, using HPLC purified long free polymer and very pure PARG. We conclusively show that on large free polymers, PARG exhibits endoglycosidic activity along with exoglycosidic activity. This endoglycosidic activity could have a significant role during cellular response to DNA damage.

Purification of poly(ADP-ribose) glycohydrolase and detection of its isoforms by a zymogram following one- or two-dimensional electrophoresis.

Poly(ADP-ribosyl)ation metabolism, a post-translational modification, involves two nuclear enzymes. Poly(ADP-ribose) polymerase (PARP) and poly(ADP-ribose) glycohydrolase (PARG) are responsible for the anabolism and catabolism of poly(ADP-ribose) polymer, respectively. PARG, despite being less abundant than PARP, is a crucial determinant of polymer metabolism which is known to be implicated in DNA repair and other cellular processes. Here, we describe modifications to improve the purification of PARG from calf thymus, in terms of both quantity and quality, which would allow biochemical and immunological studies. We also developed a zymogram to identify functional polypeptides exhibiting PARG activity. Purified and crude enzyme preparations from calf thymus were electrophoresed in two-dimensional gels. Samples were resolved on sodium dodecyl sulfate-polyacrylamide gel electrophoresis containing the polymer substrate in the form of automodified PARP after a nonequilibrium pH gradient electrophoresis. After renaturation of PARG in the gel, four isoforms of activity were clearly detected in the purified enzyme preparation. Even in the crude extract of the tissue, we could observe the major isoform of PARG. This technique will permit a better understanding of poly(ADP-ribose) catabolism and better characterization of PARG isoforms.

Classification of Acinetobacter phages.

Eight phage species and type viruses are proposed. They belong to the Myoviridae, Siphoviridae, and Podoviridae families of tailed phages and are characterized by a combination of morphological and physicochemical properties. An unusual siphovirus species has an elongated head and transverse tail disks.

Zebrin II immunoreactivity in the rat and in the weakly electric teleost Eigenmannia (gymnotiformes) reveals three modes of Purkinje cell development.

Monoclonal antibody (mab) anti-zebrin II recognizes a single 36-kD polypeptide in Purkinje cells in the rat and fish cerebellum. In the adult rat, zebrin II+ Purkinje cells form, in each hemicerebellum, seven parasagittal bands interposed by zebrin II- bands. We show that, in rats, immunoreactivity first appears caudally at postnatal day 5 and spreads; all Purkinje cells are labelled by postnatal day 12. Subsequently, immunoreactivity is selectively lost so that by day 18 the adult pattern of zebrin II+/-immunoreactive bands is created. This pattern indicates two types of Purkinje cells according to developmental trajectory, zebrin II-/+/-. In the adult gymnotiform teleost Eigenmannia, Purkinje cells in the corpus cerebelli (CCb), lateral valvula cerebelli (VCbl), and eminentia granularis anterior (EGa) are zebrin II+. Purkinje cells in the eminentia granularis posterior (EGp) and medialis (EGm) and the medial valvula cerebelli (VCbm) are zebrin II-. Zebrin II antigenicity is first present at 6 days postspawning (P6) in the EGa and at P8 in the CCb. In the valvula, labelling does not appear until P29. Immunoreactivity in the CCb, VCBl, and the EGa persists in the adult, whereas in the VCbm Purkinje cells become zebrin II- before reaching adulthood. These developmental histories (zebrin II-/+ and zebrin II-/+/-) correspond to the patterns of Purkinje cell development in mammals. Additionally, Eigenmannia has a third class of Purkinje cells, in the EGp and EGm, that never express zebrin II immunoreactivity, indicating that zebrin II expression is not an obligatory feature of Purkinje cell development in all vertebrates.

Occupational asthma caused by exposure to neurospora in a plywood factory worker.

A 24 year old man developed severe asthma two years after starting to work in a plywood plant. Four years later the patient had to stop working because of the increasing severity of his asthma. Three months after leaving his job, the patient's asthma was greatly improved. His job consisted of placing plywood sheets into a drying machine. The plywood sheets had stayed outside in wet conditions for at least four to six weeks and were usually covered with moulds. Drying the plywood sheets changed the mould into a fine orange powder. Exposure to this in the laboratory induced an isolated immediate asthmatic reaction. The same reaction was seen when the patient was challenged with an extract of the mould powder at a 0.1% w/v concentration. Skin prick test with the mould extract induced a weal and flare reaction and IgE antibodies against the dry mould powder were identified. A control patient with the same degree of bronchial hyperreactivity did not have any asthmatic reaction when challenged with the same mould extract. Culture of the dry mould powder on Sabouraud agar plates grew pure Neurospora sp. This mould has not been previously reported as a cause of occupational asthma. The immunological mechanism is probably related to an IgE mediated mast cell allergy.

Airborne microbial contents in two types of swine confinement buildings in Quebec.

Airborne microorganisms were isolated with a sampler in two types of swine confinement buildings (farrowing units and fattening units). Respirable (particles less than 5 microns) and total dust fractions were obtained. Samplings were repeated every 2 weeks for a total of 6 samplings per unit between January and April. The predominant microorganisms isolated were bacteria (up to 1.25 x 10(6) CFU/m3) with an important fraction in the respirable size range (up to 0.5 x 10(6) CFU/m3). Only small quantities of gram-negative bacteria, yeasts, and molds were found. Identification of the colonies isolated revealed a great diversity of microorganisms present in the air of the different buildings. Enterobacter agglomerans, Moraxella, Acinetobacter calcoaceticus, and Pseudomonas were the most frequently identified bacteria. Scopulariopsis, Aspergillus, Penicillium, and Candida were the most numerous fungi. Faenia rectivirgula, the causative agent of farmer's lung, was not a major contaminant. The results show some differences in airborne microbial contamination between farrowing and fattening units; the distinction, however, is not clear-cut and was observed only for the total bacteria. The level of airborne microbial contamination in swine units does not significantly vary as a function of the outside temperature. Some species of bacteria and fungi isolated in this study are known to induce extrinsic allergic alveolitis. Other fungi are known to be potentially pathogenic for man. The air of swine confinement buildings is highly contaminated with bacteria, yeasts, and molds at a level up to 1200 time higher than so-called "normal air."

Zebrin II: a polypeptide antigen expressed selectively by Purkinje cells reveals compartments in rat and fish cerebellum.

Monoclonal antibody mab-zebrin II was generated against a crude homogenate of cerebellum and electrosensory lateral line lobe from the weakly electric fish Apteronotus leptorhynchus. On Western blots of fish cerebellar proteins, mab-zebrin II recognizes a single polypeptide antigen of apparent molecular weight 36 kD. Immunocytochemistry of apteronotid brains reveals that zebrin II immunoreactivity is confined exclusively to Purkinje cells in the corpus cerebelli, lateral valvula cerebelli, and the eminentia granularis anterior. Other Purkinje cells, in the medial valvula cerebelli and eminentia granularis posterior, are not zebrin II immunoreactive. Immunoreactive Purkinje cells are stained completely, including dendrites, axons, and somata. The antigen seems to be absent only from the nucleus. A similar distribution is seen in catfish, goldfish, and a mormyrid fish. Zebrin II immunoreactivity is also found in the rat cerebellum. Western blotting of rat cerebellar proteins reveals a single immunoreactive polypeptide, with apparent molecular weight 36 kD, as in the fish. Also as in the fish, staining in the adult rat cerebellum is confined to a subset of Purkinje cells. Peroxidase reaction product is deposited throughout the immunoreactive Purkinje cells with the exception of the nucleus. No other cells in the cerebellum express zebrin II. At higher antibody concentrations, a weak glial cross reactivity is seen in most other brain regions: we believe that this is probably nonspecific. Zebrin II+ Purkinje cells are clustered together to form roughly parasagittal bands interposed by similar nonimmunoreactive clusters. In all there are 7 zebrin II+ and 7 zebrin II- compartments in each hemicerebellum. One immunoreactive band is adjacent to the midline; two others are disposed laterally to each side in the vermis; there is a paravermal band; and finally three more bands are identified in each hemisphere. Both in number and position, these compartments correspond precisely to the bands revealed by using another antibody, mabQ113 (anti-zebrin I). In both fish and rat the compartmentation revealed by zebrin II immunocytochemistry is related to the organization of cerebellar afferent and efferent projections and may provide clues as to the fundamental architecture of the vertebrate cerebellum.

Concentrations of fusidic acid, cloxacillin, and cefamandole in sera and atrial appendages of patients undergoing cardiac surgery.

The concentrations of cefamandole, cloxacillin and fusicid acid were measured in the serum and heart tissue of 100 recipients of these drugs before cardiac surgery. During cardiopulmonary bypass, mean (+/- standard deviation) peak concentrations in serum of all patients were 63.0 +/- 34.0 micrograms of cefamandole per ml, 30.8 +/- 17.7 micrograms of cloxacillin per ml, and 32.4 +/- 10.8 micrograms of fusidic acid per ml. Mean (+/- standard deviation) concentrations in atrial appendages taken 1 h (+/- 15 min) after infusion were 21.3 +/- 11.0 micrograms of cefamandole per g, 23.8 +/- 17.3 micrograms of cloxacillin per g, and 10.7 +/- 4.2 micrograms of fusidic acid per g. No cloxacillin could be detected in 5 of 39 heart specimens. Mean tissue-to-serum ratios at 1 h for cefamandole, cloxacillin, and fusidic acid were respectively 0.35, 0.73, and 0.33. Fusidic acid, a drug which is highly effective in vitro against both methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis, was detectable in heart tissue in concentrations which were 12 times higher than the MICs of this agent against these resistant microorganisms.

Short-term antibiotherapy for peritonitis: prospective, randomized trial comparing cefotaxime-metronidazole and clindamycin-tobramycin.

The combination of cefotaxime and metronidazole has been suggested for the treatment of peritonitis. We compared their effectiveness with that of tobramycin and clindamycin. Since antibiotics have most of their beneficial effect within a few days a four day course was used and a randomized trial was undertaken. The effectiveness of the 4-day course was 86% and no difference was seen between the two groups of the study.