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The outbreak caused by SARS-CoV-2 has taken many lives worldwide. Although vaccination has started, the development of drugs to either alleviate or abolish symptoms of COVID-19 is still necessary. Here, four synthetic peptides were assayed regarding their ability to protect Vero E6 cells from SARS-CoV-2 infection and their toxicity to human cells and zebrafish embryos. All peptides had some ability to protect cells from infection by SARS-CoV-2 with the D614G mutation. Molecular docking predicted the ability of all peptides to interact with and induce conformational alterations in the spike protein containing the D614G mutation. PepKAA was the most effective peptide, by having the highest docking score regarding the spike protein and reducing the SARS-CoV-2 plaque number by 50% (EC

Vaccines could be the solution to the current SARS-CoV-2 outbreak. However, some studies have shown that the immunological memory only lasts three months. Thus, it is imperative to develop pharmacological treatments to cope with COVID-19. Here, the in silico approach by molecular docking, dynamic simulations and quantum biochemistry revealed that ACE2-derived peptides strongly interact with the SARS-CoV-2 RBD domain of spike glycoprotein (S-RBD). ACE2-Dev-PepI, ACE2-Dev-PepII, ACE2-Dev-PepIII and ACE2-Dev-PepIV complexed with S-RBD provoked alterations in the 3D structure of S-RBD, leading to disruption of the correct interaction with the ACE2 receptor, a pivotal step for SARS-CoV-2 infection. This wrong interaction between S-RBD and ACE2 could inhibit the entry of SARS-CoV-2 in cells, and thus virus replication and the establishment of COVID-19 disease. Therefore, we suggest that ACE2-derived peptides can interfere with recognition of ACE2 in human cells by SARS-CoV-2 in vivo. Bioinformatic prediction showed that these peptides have no toxicity or allergenic potential. By using ACE2-derived peptides against SARS-CoV-2, this study points to opportunities for further in vivo research on these peptides, seeking to discover new drugs and entirely new perspectives to treat COVID-19. Communicated by Ramaswamy H. Sarma

To assess the effect of vertical sleeve gastrectomy (VSG) and Roux-en-Y gastric bypass (RYGB) on the esophageal and intestinal morphology of western diet (WD)-obese rats and to characterize the stomach histopathology of WD rats submitted to VSG. Male Wistar rats received WD from 2-4 months of age, to induce obesity, before randomly submitting them to pseudo (WD-SHAM), VSG (WD-VSG) or RYGB (WD-RYGB) surgeries. Gastrointestinal histomorphometry was performed at 3-months post-surgery. The upper esophagus of VSG and RYGB rats increased luminal area, while reductions in the keratin layer of the mucosa and the tunica muscularis were observed only in the RYGB animals. In the lower esophagus, both surgeries increased keratin layer thickness, but reduced the mucosal mucus content, while RYGB increased the thickness of the tunica mucosa and muscularis. The glandular region of the stomach of WD-VSG rats exhibited hypotrophy, epithelial erosion, fibrosis and moderate inflammatory infiltration. VSG and RYGB increased the villi height in the ileum, and the thickness of the tunica muscularis in the jejunum and ileum of WD rats; furthermore, RYGB augmented the ileal villi height. Thus both approaches induced histomorphological alterations in the esophagus and intestine and VSG damaged the gastric mucosa, even over the long-term.

The recent outbreak caused by SARS-CoV-2 continues to threat and take many lives all over the world. The lack of an efficient pharmacological treatments are serious problems to be faced by scientists and medical staffs worldwide. In this work, an in silico approach based on the combination of molecular docking, dynamics simulations, and quantum biochemistry revealed that the synthetic peptides RcAlb-PepI, PepGAT, and PepKAA, strongly interact with the main protease (Mpro) a pivotal protein for SARS-CoV-2 replication. Although not binding to the proteolytic site of SARS-CoV-2 Mpro, RcAlb-PepI, PepGAT, and PepKAA interact with other protein domain and allosterically altered the protease topology. Indeed, such peptide-SARS-CoV-2 Mpro complexes provoked dramatic alterations in the three-dimensional structure of Mpro leading to area and volume shrinkage of the proteolytic site, which could affect the protease activity and thus the virus replication. Based on these findings, it is suggested that RcAlb-PepI, PepGAT, and PepKAA could interfere with SARS-CoV-2 Mpro role in vivo. Also, unlike other antiviral drugs, these peptides have no toxicity to human cells. This pioneering in silico investigation opens up opportunity for further in vivo research on these peptides, towards discovering new drugs and entirely new perspectives to treat COVID-19. Communicated by Ramaswamy H. Sarma

The global outbreak of COVID-19 (Coronavirus Disease 2019) caused by SARS-CoV-2 (Severe Acute Respiratory Syndrome caused by Coronavirus 2) began in December 2019. Its closest relative, SARS-CoV-1, has a slightly mutated Spike (S) protein, which interacts with ACE2 receptor in human cells to start the infection. So far, there are no vaccines or drugs to treat COVID-19. So, research groups worldwide are seeking new molecules targeting the S protein to prevent infection by SARS-CoV-2 and COVID-19 establishment. We performed molecular docking analysis of eight synthetic peptides against SARS-CoV-2 S protein. All interacted with the protein, but Mo-CBP3-PepII and PepKAA had the highest affinity with it. By binding to the S protein, both peptides led to conformational alterations in the protein, resulting in incorrect interaction with ACE2. Therefore, given the importance of the S protein-ACE2 interaction for SARS-CoV-2 infection, synthetic peptides could block SARS-CoV-2 infection. Moreover, unlike other antiviral drugs, peptides have no toxicity to human cells. Thus, these peptides are potential molecules to be tested against SARS-CoV-2 and to develop new drugs to treat COVID-19., Highlights • Synthetic peptides bind to SARS-CoV-2 Spike protein. • Synthetic peptides induced conformational changes on SARS-CoV-2 spike protein structure. • Synthetic peptides bind to ACE2 protein but did not affect its structure. • Synthetic peptides induced the wrong interaction of SARS-CoV-2 with ACE2 receptor.

Transitory allergies to cow milk proteins in infants or adults have become a public health problem. Although extensively or partially hydrolyzed cow milk protein formulas are available, these products are costly. Therefore, studies into innovative enzymes to digest cow milk proteins are needed. Danaus plexippus gut peptidases were purified and examined with regard to cow milk protein hydrolysis. The peptidases hydrolyzed caseins and whey proteins. However, after heat treatment, there was a significant improvement in β-lactoglobulin hydrolysis. The hydrolyzed cow milk proteins were not recognized by anti-casein antibodies and only reacted slightly with antibodies against whey proteins. This performance was better than that of partially hydrolyzed formulas and similar to that of an extensively hydrolyzed formula. These results suggest that D. plexippus gut peptidases are suitable and innovative enzymes to produce hypoallergenic cow milk protein formulas.

As proteins isolated from the Rhinella schneideri parotoid gland secretion (RsPP) exhibit anti-inflammatory activity, the goal of this work was to investigate their anti-nociceptive effects using acetic acid-induced writhing, formalin, and hot-plate tests. The intraperitoneal administration of RsPP (2.5 or 5 mg/kg) one hour prior to stimuli significantly reduced the abdominal constrictions induced by acetic acid (73.06 and 72.69% inhibition, respectively) and the inflammatory phase of paw licking time induced by formalin (69.3% inhibition, at 2.5 mg/kg). However, RsPP (1, 2.5 or 5 mg/kg) did not change the latency in response at the hot-plate test. The involvement of inflammatory mediators on the anti-nociceptive effect of RsPP was further demonstrated. RsPP (2.5 mg/kg) significantly inhibited the inflammatory peak of paw edema induced by histamine (44.0%), bradykinin (51.3%), or prostaglandin E2 (53.7%). Our data indicate that RsPP may act on the pain process by inhibiting the effect of inflammatory mediators.

The emergence of antibiotic-resistant microbes has stimulated research worldwide seeking new biologically active molecules. In this respect, synthetic antimicrobial peptides (SAMPs) have been suggested to overcome this problem. Although there are some online servers that provide putative SAMPs from protein sequences, the choice of the best peptide sequences for further analysis is still difficult. Therefore, the goal of this paper is not to launch a new tool but to provide a friendly workflow to characterize and predict potential SAMPs by employing existing tools. Using this proposed workflow, two peptides (PepGAT and PepKAA) were obtained and extensively characterized. These peptides damaged microbial membranes and cell walls, and induced overproduction of reactive oxygen species (ROS). Both peptides were found to assume random coil secondary structure in aqueous solution, organic solvent, and upon binding to negatively charged lipid systems. Peptides were also able to degrade formed biofilms but not to prevent biofilm formation. PepGAT was not resistant to proteolysis, whereas PepKAA was resistant to pepsin but not to pancreatin. Furthermore, both presented no hemolytic activity against red blood cells, even at a 10-fold higher concentration than the antimicrobial concentration. The pipeline proposed here is an easy way to design new SAMPs for application as alternatives to develop new drugs against human pathogenic microorganisms.

Osmotin- and thaumatin-like proteins (OLPs and TLPs) have been associated with plant defense responses to different biotic stresses. In the present work, several in silico sequences from OLPs and TLPs were investigated by means of bioinformatics tools aiming to prospect for antimicrobial peptides. The peptide sequences chosen were further synthesized and characterized, and their activities and action mechanisms were assayed against some phytopathogenic fungi, bacteria and yeasts of clinical importance. From this survey approach, four peptide sequences (GDCKATSC, CPRALKVPGGCN, IVGQCPAKLKA, and CAADIVGQCPAKLK) were selected considering some chemical parameters commonly attributed to antimicrobial peptides. Antimicrobial assays showed that these peptides were unable to inhibit mycelial growth of phytopathogenic fungi and they did not affect bacterial cell growth. Nevertheless, significant inhibitory activity was found for CPRALKVPGGCN and CAADIVGQCPAKLK against Candida albicans and Saccharomyces cerevisiae. Fluorescence and scanning electron microscopy assays suggested that CAADIVGQCPAKLK did not damage the overall cell structure, or its activity was negligible on yeast membrane and cell wall integrity. However, it induced the production of reactive oxygen species (ROS) and apoptosis. Molecular docking analysis showed that CAADIVGQCPAKLK had strong affinity to interact with specific plasma membrane receptors of C. albicans and S. cerevisiae, which have been described as promoting the induction of apoptosis. The results indicate that CAADIVGQCPAKLK can be a valuable target for the development of a desired antimicrobial agent against the pathogen C. albicans.

The production of recombinant human granulocyte-colony stimulating factor (hG-CSF) for therapeutic purposes relies on its expression in selected clones of transfected mammalian cells. Alternatively, this protein can be produced by targeted secretion in the milk of transgenic goats. Thus, the aim of this study was to produce founder transgenic goats expressing hG-CSF and to propagate a transgenic production herd. After DNA microinjection of in vivo-produced pronuclear embryos was performed, two founders were obtained (one male and one female, named 10 M and 12 F, respectively). The mean level of hG-CSF secreted in the milk from the 12 F goat, which was measured during a period of induced lactation, was 620.92 ± 179.93 μg/ml. In addition, the recombinant protein presented in vitro biological activity on differentiation of human umbilical stem cells to neutrophil granulocyte series. A total of nine kids (six from 10 M and three from 12 F) that carried the hG-CSF transgene were generated by outbreeding of the founders. In summary, we produced two transgenic goats with a stably integrated hG-CSF gene that were capable of secreting recombinant hG-CSF from the lactating mammary gland without causing any harm to the animals’ health. Additionally, the founders proved to be fertile and capable of transmitting the hG-CSF gene to first generation progeny of each line. Additional investigations of the phenotypic and genotypic characteristics of the 10 M and 12 F lines are warranted.

The efficiency of the transrectal real-time ultrasonography (RTU) scanning technique to monitor the number of corpora haemorraghica (CH) of goat embryo donors was compared to results obtained by a reference method (laparotomy). The ovaries of 24 Saanen goats were observed by RTU, approximately 72 h after superovulatory treatment, followed by laparotomy. The diameter of the four largest CHs was measured using image analysis software. The use of ultrasonographic scanning allowed in determining if a goat had ovulated or not in all the females. In all the ovaries examined the total number of CHs counted was lower ( p r 2 = 0.71; p

The effect of two successive superovulatory treatments was assessed in 15 Santa Ines ewes synchronized with 60 mg medroxyprogesterone acetate sponges for 14 days and superovulated with 200 IU of porcine FSH (pFSH) in decreasing doses at 12 h intervals during days 12–14 of progestagen treatment. Estrus was observed from 12 h after sponge removal and the ewes in estrus were hand mated with Santa Ines rams of proven fertility. All embryos were recovered by laparotomy 6–7 days after estrus. Treated ewes showed estrus from 24 to 72 h after sponge removal. The time of onset of estrus after sponge removal and the length of estrus were not affected (P>0.05) by the repetition of treatment. Ewes that failed to superovulate in the first and the second treatment (P>0.05) were 27 and 40%, respectively. The mean ovulation rate for those ewes responding to treatment was similar (P>0.05) between the first (9.9±1.1) and the second (11.3±1.9) treatment. There was no significant difference in fertility between the first (53%) and second (40%) treatment. Among those fertilized, viable embryos were detected for 78% in the first treatment and 64% in the second treatment (P>0.05). It was concluded that Santa Ines ewes respond to superovulatory regimes with pFSH and the treatment may be repeated for at least two times.

The objective of this study was to examine the effect of donor breed on pronuclear-stage embryo yield to be used for DNA microinjection in a transgenesis goat program. Twelve Caninde and twelve Saanen goats were heat synchronized using a progestagen-cloprostenol treatment. Forty-eight hours before the sponge removal, superovulation was induced with a total administration of 4.4 mg/kg bodyweight NIH-FSH-P1, given twice daily in decreasing doses over 3 days. In addition, goats received 100 μg of GnRH and they were hand-mated at 36 and 48 h after progestagen removal. Embryo recovery was performed by oviduct flushing at 72 h after sponge removal. Embryos were microinjected with a DNA construct and noticeable swelling of the nuclei was the criterion for successful microinjection. The total diameter, cytoplasm diameter, zona pellucida thickness and pronuclei diameter were measured for each microinjected embryo. A higher (p < 0.05) percentage of fertilized ova was observed in Caninde (89.9%) than Saanen (36.2%) goats. In addition, Caninde donors produced a higher percentage of pronuclear embryos when compared with Saanen: 72.5% vs 20.6% (p < 0.05), respectively. Successful microinjection was verified in 96.7% and 73.3% of times in Caninde and Saanen embryos, respectively (p < 0.05). Significant differences were observed for all morphometric parameters except for cytoplasm diameter. In conclusion, under our study experimental conditions, Caninde were more efficient than Saanen goats concerning the pronuclear embryo yield and manipulation. The use of Caninde goats in transgenesis programs could be increase the interest in their breeding and could be contribute to saving them from extinction.

Fifteen Anglo-Nubian and 14 Saanen female kids were used to evaluate the age at puberty when raised in a semi-arid environment. The kids were monitored from 90 days of age until all females showed, at least, one (first) oestrous period. The kids were weighed at 15-day intervals and the occurrence of oestrus was checked twice a day. Blood samples were taken weekly and assayed for progesterone in order to detect luteal function. The age at puberty was defined as the date of first oestrus followed by luteal function. After puberty, the oestrous cycles were observed and classified as short ( 25 days) in length. The initial and final body weights were, respectively, 14.6±1.2 and 34.0±2.0 kg for Anglo-Nubian kids, and 17.4±1.2 and 40.1±2.5 kg for Saanen kids. From 150 days of age, the Saanen goats were heavier (P

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T19:00:48Z No. of bitstreams: 0 Made available in DSpace on 2013-08-22T19:00:48Z (GMT). No. of bitstreams: 0 Previous issue date: 1998-01-01 Made available in DSpace on 2013-09-30T19:59:50Z (GMT). No. of bitstreams: 0 Previous issue date: 1998-01-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T15:15:23Z No. of bitstreams: 0 Made available in DSpace on 2014-05-20T15:15:23Z (GMT). No. of bitstreams: 0 Previous issue date: 1998-01-01 Fez-se um estudo de campo sobre a competição e a dinâmica de ocupação de sítios de nodulação de estirpes de Bradyrhizobium japonicum naturalizadas ou inoculadas em sementes. O ensaio foi instalado em Latossolo Vermelho Escuro utilizando blocos ao acaso, com 8 repetições dos seguintes tratamentos: A) controle sem inoculante; B) controle sem inoculante, com adubação nitrogenada; C) inoculante com cerca de 107 rizóbios/g (8g/kg de semente); D) inoculação com cerca de 1010 rizóbios/g (8g/kg de semente). Os inoculantes foram preparados com as estirpes recomendadas na ocasião, SMS - 314 (= SEMIA 587) e SMS - 463 (= SEMIA 5019 = 29W). A identificação das estirpes formadoras dos nódulos foi realizada em amostras coletadas aos 10, 42 e 70 dias após a germinação. Realizou-se também a colheita dos grãos para avaliação da produção e de N-total. A tipificação sorológica foi feita com antissoro preparado a partir de antígenos constituídos pelas seguintes estirpes de Bradyrhizobium japonicum: SEMIA 587, SEMIA 5019 e SEMIA 5052 (= USDA 6), utilizando-se da técnica immunodot. A caracterização das estirpes dos nódulos revelou um pequeno aumento (6,2 a 8,7%), na ocorrência de nódulos formados pelas estirpes recomendadas, nas três épocas de amostragem. Houve expressiva participação da estirpe 5052 (= sorogrupo USDA 6) na formação dos nódulos. Verificou-se que as estirpes naturalizadas podem contribuir de forma diferente na formação dos nódulos durante o ciclo, dependendo da amostragem. A field study on the competion and on the dynamics of occupation of nodulation sites by naturalized and seed inoculated strains of Bradyrhizobium japonicum was conducted on a Dark Red Latosol. The experimental design consisted of randomized blocks, with eight replications of the following treatments: A) control without inoculation; B) control without inoculation, with nitrogen fertilizer; C) inoculation with about 107 rhizobia/g (8g/kg of seed); D) inoculation with about 1010 rhizobia/g (8g/kg of seed). The inoculant was prepared with the strains SMS-314 (= SEMIA-587) and SMS-463 (=SEMIA-5019 = 29W), officially recommended for inoculant production. The identification of the strains which formed nodules was made in samples collected at 10, 42 and 70 days after seed germination. The grains were harvested to evaluate production and nitrogen content. Sorological typification was made with antiserum prepared from antigens constituted by B. japonicum strains SEMIA-587, SEMIA-5019 and SEMIA-5052 (=USDA-6), using the immunodot method. The characterization of the strains in nodules revealed a small increase (6,2 to 8,7%) in the occurrence of nodules formed by the recommended strains for the three sampling dates. There was an expressive participation of the strain 5052 (serogroup USDA-6) in the formation of the nodules. It was also observed that the naturalized strains may contribute differently to the formation of the nodules during the cycle, depending on the time of sampling. IAC UNESP UNESP

Foram induzidos sintomas de carência de macronutrientes no milho, var. Piranão, exceto no caso do enxofre. Os níveis foliares de N, P, K, Ca, Mg e S foram determinados nas plantas sujeitos à nutrição completa e às deficiências de macronutrientes. Corn plants, Piranão variety, were grown in nutrient solution, complete and with omission of one macronutrient each time, in order to gain information on the effects of treatments on growth, symptoms of deficiency, and mineral composition of the leaves. The following symptoms of deficiency were obtained: Nitrogen -- plants underdeveloped; "V" shaped chlorosis beginning in the older leaves followed by drying and necrosis; Phosphorus - older leaves dark green with a purple hve; Potassium - plants underdeveloped; marginal chlorisis and necrosis in the older leaves; iron induced chlorosis in the younger leaves; Calcium - plants very underdeveloped due to death of apical merystem; younger leaves showing tip and marginal chlorosis and afterwards necrosis and break down of the tissue; wide whitish areas in the younger leaves; Magnesium - plants very underdeveloped; internerval chlorosis in older leaves first, proceeding to the young ones; Sulfur - absence of marked symptoms, except for a slight chlorosis in the younger leaves. Dry matter production obeyed the following decreasing order: Complete, minus S, minus P, minus K, minus Mg, minus N and minus Ca. Ears were obtained only in treatments complete and minus S. Top/root ratio decreased according to this order: minus S, complete, minus K, minus Mg, minus N, minus P and minus Ca. The macronutrient content for the treatment complete was the following respectively for lower and upper, leaves: N - 2.76 and 2.52%; P - 0.59 and 0.53%; K - 2.25 and 2.26%;Ca - 2.17 and 1.85;Mg - 0.82 and0.66;S - 0.38 and 0.32. The values of the element corresponding to each treatment were: N - 1.84 and 1.22 (that is: lower leaves in the minus N plants had 1 84% N, whereas the upper leaves in same treatment had 1.22% N; P - 0.14 and 0.17%; K - 0.29 and 0.78%; Ca - 0.29 and 0.05; Mg -0.09 and 0.05%; S- 0.31 and 0.26%.

Um experimento foi conduzido em Campinas, em latossolo vermelho escuro, barrento, por dois anos consecutivos, para testar o efeito dos herbicidas trifluralin, vernolate e alachlor e duas doses de nitrogênio, na nodulação e produção de soja, CV santa rosa, inoculada. No primeiro ano, os herbicidas não influenciaram a nodulação aos 35, 58 e 98 dias após o plantio. A aplicação de nitrogênio diminuiu o peso dos nódulos secos observados aos 58 dias, e seu número e peso, aos 98 dias. A produção de grãos foi semelhante para todos os tratamentos. No segundo ano, a nodulação das plantas inoculadas não foi influenciada pela aplicação de nitrogênio, porém nos tratamentos com nodulação espontânea (não inoculados), o peso dos nó dulos secos diminuiu significativamente, nas três épocas, devido à aplicação daquele nutriente. Os tratamentos inoculados produziram maiores quantidades de sementes em comparação com os tratamentos não inoculados. Houve interação significativa de inoculação x adubação nitrogenada, sendo que o aumento devido à adubação só foi verificado no tratamento sem inoculação. Nos canteiros em que se fez a aplicação de herbicidas, a produção de grãos foi maior; não foi , entretanto, possível atribuir esse efeito à aplicação dos herbicidas. A field trial was performed on a loamy soil, for two consecutive years, to study the effect of trifluralin, vernolate and alachlor and two rates of nitrogen, on nodulation and yield of inoculated soybeans. The herbicides did not affect the nodulation or yield during the crop cycle, in any of the experiments. The N application depressed the weight of dry nodules at 58 days and its number and weight at 98 days in the first year. The yield of the different treatmens showed no difference. In the second year the nitrogen application did not affect the nodulation of inoculated plants, but the weight of dry nodules of the treatments with spontaneous nodulation was significantly reduced in the three periods of sampling. This year the inoculated treatments showed higher yields when compared with those not inoculated, and the herbicide treated plots yielded more than the checks without herbicide.

O milho, var. Piranão, foi cultivado em solução nutritiva com níveis crescentes de N, R e K. Houve resposta linear à adição de N e assintótica às doses de R e de K. A determinação da atividade da reductase de nitrato se correlacionou melhor com a produção da matéria seca que a do N total nas folhas. O teor de potássio total nas folhas, por sua vez refletiu melhor o estado nutricional que a determinação de putrescina nas folhas. Maize plants, Piranão cultivar, were grown in nutrient solution with 5 levels each of N, R and K till harvest. Nitrate reductase activity and putrescine level were determined in leaves of plants in the N and K series, respectively, at silking stage. Upper and lower leaves were analysed for N, R and K in the corresponding treatments at the end of the life cycle. The main conclusions were the following : 5.1 There was a linear effect of level of N in the substrate on dry matter production. 5.2. High activities of nitrate reductase suggest that under natural conditions the variety should be very responsive to N fertilization. 5.3. An asymptotic response curve was found in the treatments with increasing levels of R in the medium, as well as in the case of the K treatments. 5.4. Curves of response of roots (dry matter) showed a tendency to level of as a function of increasing levels of element at lower concentrations than the components aeral part of the plant (leaves, stems, ears). 5.5. Determination of leaf putrescine did not of prove a better indicator of the K status leaf K. 5.6. The variety under study seems to be relatively more efficient in the utilization of N for yield components; the efficiency for utilization of K, is rather low and that for R is intermediate, findings that should have a bearing on the fertilization in field conditions.