A Comparison of Diffusion-Conductimetric and Distillation-Titration Methods in Analyzing Ammonium- and Nitrate-Nitrogen in the KCl-Extracts of Georgia Soils

Soil mineral (or inorganic) nitrogen (SMN), which primarily exists as exchangeable and soluble ammonium (NH4⁺) and the nitrate (NO₃^?) ions, represents readily available nitrogen for plant growth. Over the years a 2M potassium chloride (KCl) solution has become the extraction solution of choice for extracting SMN. In the research and service laboratories, either distillation-titration method (DTM) or colorimetric method (CM) is virtually the standard to measure NH₄⁺- and NO₃^?-N in the 2M KCl soil extracts. However, being a time-consuming and labor intensive method, DTM generally has a very low throughput. Likewise, CM is affected by interferences from pH variation, color, turbidity, presence of organic species, and some other constituents in the extracts. In contrast, diffusion conductivity method (DCM) is a less expensive and high throughput one, which is also relatively free from common interferences. In this study, we, therefore, compared the extraction efficiency of various KCl concentrations and performance of diffusion conductivity method (DCM) with DTM in measuring NH₄⁺-N and NO₃^?-N in KCl extracts of 32 agricultural soils of Georgia. A 0.2M KCl solution extracted statistically similar amounts of NH₄⁺-N and NO₃^?-N as did 2M KCl, suggesting that a 10-fold dilute KCl solution than the standard 2M KCl might be good enough to extract and estimate the most of SMN. For the analyses of NH₄⁺- and NO₃^?-N in the KCl extracts, the DCM produced results statistically similar to those produced by DTM. The deviation between the results given by DCM and DTM was no more than ±10%. Thus, DCM appears to be an attractive alternative to the labor intensive and time-consuming DTM for measuring NH₄⁺- and NO₃^?-N in the KCl extract of soils in the research and service laboratories.     

Occurrence and incidence of viruses infecting green beans in south-eastern Spain

A study was conducted to determine the identity and prevalence of viruses in 455 greenhouses in the main Spanish green bean growing area. Directed surveys were conducted in 422 crops from 2000–2004 to collect samples from diseased plants displaying symptoms that could be attributed to viruses. The samples were analysed to detect any virus by means of dsRNA extraction, mechanical inoculation to test plants, as well as ELISA and/or RT-PCR tests to detect potyviruses, geminiviruses and viruses previously known to infect beans in Spain. Random surveys were conducted in the years 2002 and 2005 (in 21 and 12 greenhouses, respectively) to study the actual incidence of known viruses in the area. Symptoms were recorded in 23,108 plants from which 664 plants were collected and analysed by ELISA or RT-PCR. The results of the directed surveys showed that all the analyzed crops carried the cryptic virus Phaseolus vulgaris endornavirus (PVuV), whereas phytopathogenic viruses appeared in smaller percentages of the crops: Tomato yellow leaf curl virus (TYLCV) 20.4%, Southern bean mosaic virus (SBMV) 9.0%, Tomato spotted wilt virus (TSWV) 4.0%, and the new species Bean yellow disorder virus (BnYDV) that broke out in 2004 with occurrence values higher than 34.3% that year. From 2000–2004 an important decrease in TYLCV was observed, along with a slight increase in SBMV and a consistently low occurrence of TSWV. The results of the random surveys confirmed the increased occurrence of virus detected during the directed surveys, and furthermore demonstrated the percentage of incidence for each virus.     

Marine-Derived Penicillium purpurogenum Reduces Tumor Size and Ameliorates Inflammation in an Erlich Mice Model

Background: This study addresses the antitumoral properties of Penicillium purpurogenum isolated from a polluted lagoon in Northeastern Brazil. Methods: Ethyl Acetate Extracellular Extract (EAE) was used. The metabolites were studied using direct infusion mass spectrometry. The solid Ehrlich tumor model was used for antitumor activity. Female Swiss mice were divided into groups (n = 10/group) as follows: The negative control (CTL−), treated with a phosphate buffered solution; the positive control (CTL+), treated with cyclophosphamide (25 mg/kg); extract treatments at doses of 4, 20, and 100 mg/kg; animals without tumors or treatments (Sham); and animals without tumors treated with an intermediate dose (EAE20). All treatments were performed intraperitoneally, daily, for 15 days. Subsequently, the animals were euthanized, and the tumor, lymphoid organs, and serum were used for immunological, histological, and biochemical parameter evaluations. Results: The extract was rich in meroterpenoids. All doses significantly reduced tumor size, and the 20 and 100 mg/kg doses reduced tumor-associated inflammation and tumor necrosis. The extract also reduced the cellular infiltration of lymphoid organs and circulating TNF-α levels. The extract did not induce weight loss or renal and hepatic toxic changes. Conclusions: These results indicate that P. purpurogenum exhibits immunomodulatory and antitumor properties in vivo. Thus, fungal fermentation is a valid biotechnological approach to the production of antitumor agents.     

PRRSV2 genetic diversity defined by RFLP patterns in the United States from 2007 to 2019

The genetic diversity of porcine reproductive and respiratory syndrome virus (PRRSV) increases over time. In 1998, restriction-fragment length polymorphism (RFLP) pattern analysis was introduced to differentiate PRRSV wild-type strains from VR2332, a reference strain from which a commercial vaccine (Ingelvac PRRS MLV) was derived. We have characterized here the PRRSV genetic diversity within selected RFLP families over time and U.S. geographic space, using available ISU-VDL data from 2007 to 2019. The 40,454 ORF5 sequences recovered corresponded to 228 distinct RFLPs. Four RFLPs [2-5-2 (21.2%), 1-7-4 (15.6%), 1-4-4 (11.8%), and 1-8-4 (9.9%)] represented 58.5% of all ORF5 sequences and were used for cluster analysis. Over time, there was increased detection of RFLPs 2-5-2, 1-7-4, 1-3-4, 1-3-2, and 1-12-4; decreased detection of 1-4-2, 1-18-4, 1-18-2, and 1-2-2; and different detection trends for 1-8-4, 1-4-4, 1-26-1, 1-22-2, and 1-2-4. An over-time cluster analysis revealed a single cluster for RFLP 2-5-2, supporting that sequences within RFLP 2-5-2 are still relatively conserved. For 1-7-4, 1-4-4, and 1-8-4, there were multiple clusters. State-wise cluster analysis demonstrated 4 main clusters for RFLP 1-7-4 and 1-8-4, and 6 for RFLP 1-4-4. For the other RFLPs, there was a significant genetic difference within them, particularly between states. RFLP typing is limited in its ability to discriminate among different strains of PRRSV. Understanding the magnitude of genetic divergence within RFLPs helps develop PRRSV regional control programs, placement, herd immunization strategies, and design of appropriate animal movements across borders to minimize the risk of PRRSV transmission.     

Comprehensive prediction of plant cytoplasmic and apoplastic effectors underlying Erwinia psidii pathogenicity

Erwinia psidii (Eps) is the causal agent of emerging diseases of eucalypt and guava; however, the mechanisms underlying its pathogenicity are not fully understood. Here, we predicted factors involved in the ability of Eps to cause disease on its host plants. For that, the genomes of four Eps strains exhibiting different virulence on eucalypt were sequenced, and hrp/hrc genes coding for the type III secretion system (T3SS), effectors injected into the plant cell cytoplasm through the T3SS (T3SEs) and their plant subcellular localizations, as well as proteins deployed to the host apoplast, were predicted. It was found that Eps possesses a complete hrp/hrc gene cluster based on comparison with Erwinia amylovora. A total of 18 T3SEs were predicted, 11 of which were shared among all strains, none were exclusive to any strain and seven were absent in at least one strain. No sequence variation among strains was found for five T3SE candidates whereas extensive variation was found for six, suggesting the latter may be determinants of virulence differences. The T3SE candidates are predicted to target the plant cell nucleus, cytoplasm, mitochondrion, chloroplast and peroxisome. The predicted apoplastic effector repertoire common to all four strains was over-represented in proteins of unknown functions or predicted to possess enzymatic activities, among which the most abundant were oxidoreductases and peptidases. Proteins with lytic transglycosylase activity were predicted in strain-specific apoplastic effector repertoires. These results provide an important framework for future research aimed at uncovering the factors underlying Eps pathogenicity.     

Retrospective and prospective studies of transmissible viral proventriculitis in broiler chickens in Brazil

We investigated the occurrence and pathologic findings of transmissible viral proventriculitis (TVP) associated with the chicken proventricular necrosis virus (CPNV) in commercial broiler chickens in southeastern Brazil. Seventy-three broilers, 25–36 d old, with a history of reduced growth, were referred to our veterinary pathology services from 2013 to 2017. Broilers were clinically examined, weighed, and euthanized for postmortem examination. Broilers of different ages with proventricular histologic lesions were positive for CPNV by RT-PCR; however, the intensity of histologic lesions was higher among 33-d-old animals, and viral RNA detection was more frequent among those that were 28 d old. In the proventriculi of 35 of 73 (48%) broilers, lesions were characterized by glandular epithelial necrosis, lymphoplasmacytic and histiocytic infiltrates, and metaplasia of glandular epithelium to ductal epithelium. In 24 of 73 (36%) broilers with histologic TVP-compatible lesions, CPNV was detected by RT-PCR for the viral protein 1 (VP1) gene. Broilers with histologic lesions were lighter than expected compared to the Cobb 500 standard weight. TVP has not been reported previously in broiler chickens in Brazil, to our knowledge.     

Prolactin stimulates the internalization of Staphylococcus aureus and modulates the expression of inflammatory response genes in bovine mammary epithelial cells

The incidence of mastitis in dairy cattle is highest at the drying off period and parturition, which are characterized by high levels of the lactogenic hormone prolactin (PRL). One of the most frequently isolated contagious pathogens causing mastitis is Staphylococcus aureus. However, the role of PRL on S. aureus infection in mammary epithelium has not been studied. In this work we evaluated the effect of bovine PRL (bPRL) on S. aureus internalization in a primary culture of bovine mammary epithelial cells (bMEC) and on the expression of cytokine and innate immune response genes. Our data show that 5ng/mL bPRL enhances approximately 3-fold the internalization of S. aureus (ATCC 27543) into bMEC. By RT-PCR analysis, we showed that bPRL is able to up-regulate the expression of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and inducible nitric oxide synthase (iNOS) mRNAs. However, bPRL together with S. aureus did not modify the expression of TNF-alpha and iNOS mRNAs, while it down-regulated the expression of beta-defensin and IL-1beta mRNAs, as well as nitric oxide production, suggesting that infection and bPRL together can inhibit elements of the host immune response. To our knowledge, this is the first report that shows a role of bPRL during the internalization of S. aureus into bMEC.     

Protein Hydrolysate Fractions from Sea Cucumber (Isostichopus badionotus) Inhibit Angiotensin-Converting Enzyme

Angiotensin-converting enzyme (ACE) inhibitory capacity was evaluated in hydrolysates from sea cucumber Isostichopus badionotus. The pepsin-pancreatin and Alcalase®-Flavorzyme® sequential systems were used. Hydrolysates were characterized in terms of degree of hydrolysis (DH), electrophoretic profile, and inhibitory activity. The Alcalase®-Flavorzyme® system produced hydrolysates with the highest DH, with that produced at 90 min exhibiting the highest ACE inhibitory action (86%). This hydrolysate was chosen for purification by ultrafiltration with five molecular weight cut-offs (< 1, 1–3, 3–5, 5–10, and > 10). The > 10-kDa fraction exhibited the highest ACE inhibitory action (80.7%) and was fractionated by gel fractionation chromatography. Of the 11 resulting fractions, IX had the highest ACE inhibitory action (73.4%) and lowest IC₅₀ value (2-μg prot/mL). Hydrolysis of sea cucumber I. badionotus with the Alcalase®-Flavorzyme® system produced hydrolysates with ACE inhibitory action. These contained a peptide with an IC₅₀ value comparable to those of synthetic ACE inhibitors.     

Potential Utilization of Green Tide-Forming Macroalgae from Patos Lagoon,Rio Grande-RS,Brazil

ABSTRACT

This study was conducted to evaluate the biochemical and nutritional characteristics of naturally occurring estuarine macroalgae in the Patos Lagoon (Rio Grande, Brazil). The study of these macroalgae, dominated by Ulva and Cladophora species, contributes to current research on the potential utilization of green tide-forming algae. Although high levels of carotenoids (378 ± 0.02 µg g^?1) and antioxidant activity (75.0% for DPPH) were found in the naturally occurring macroalgae, these macroalgae cannot be used in the food industry due to the presence of toxic levels of arsenic (exceeding the thresholds of both French and Brazilian legislation for food). Regarding the potential of the studied macroalgae as a biofuel source, the low lipid and very high carbohydrate contents suggest a possible use in the fermentation of ethanol, butanol, or methane.