NMR analysis, cytotoxic activity and theoretical study of a complex between SRPIN340 and p-sulfonic acid calix[6]arene.

Aim: This study aimed to enhance the aqueous dissolution of SRPK inhibitor N-(2-(piperidin-1-yl)-5-(trifluoromethyl)phenyl)isonicotinamide (SRPIN340).Materials & Methods: A complex with p-sulfonic calix[6]arene (Host) and SRPIN340 (Guest) was prepared, studied via 1H nuclear magnetic resonance (NMR) and theoretical calculations and biologically evaluated on cancer cell lines.Results & conclusion: The 1:1 host (H)/guest (G) complex significantly enhanced the aqueous dissolution of SRPIN340, achieving 64.8% water solubility as determined by 1H NMR quantification analysis. The H/G complex reduced cell viability by 75% for HL60, ∼50% for Nalm6 and Jurkat, and ∼30% for B16F10 cells. It exhibited greater cytotoxicity than free SRPIN340 against Jurkat and B16F10 cells. Theoretical studies indicated hydrogen bond stabilization of the complex, suggesting broader applicability of SRPIN340 across diverse biological systems.

[Box: see text].

Chimeric proteins of Mycoplasma hyopneumoniae as vaccine and preclinical model for immunological evaluation.

Mycoplasma hyopneumoniae (M. hyopneumoniae) is a primary agent of porcine enzootic pneumonia, a disease that causes significant economic losses to pig farming worldwide. Commercial vaccines induce partial protection, evidencing the need for a new vaccine against M. hyopneumoniae. In our work, three chimeric proteins were constructed, composed of potentially immunogenic domains from M. hyopneumoniae proteins. We designed three chimeric proteins (Q1, Q2, and Q3) based on bioinformatics analysis that identified five potential proteins with immunogenic potential (MHP418, MHP372, MHP199, P97, and MHP0461). The chimeric proteins were inoculated in the murine model to evaluate the immune response. The mice vaccinated with the chimeras presented IgG and IgG1 against proteins of M. hyopneumoniae. There was induction of IgG in mice immunized with Q3 starting from 30 days post-vaccination, and groups Q1 and Q2 showed induction at 45 days. Mice of the group immunized with Q3 showed the production of IgA. In addition, the mice inoculated with chimeric proteins showed a proinflammatory cytokine response; Q1 demonstrated higher levels of TNF, IL-6, IL2, and IL-17. In contrast, animals immunized with Q2 showed an increase in the concentrations of TNF, IL-6, and IL-4, whereas those immunized with Q3 exhibited an increase in the concentrations of TNF, IL-6, IL-10, and IL-4. The results of the present study indicate that these three chimeric proteins can be used in future vaccine trials with swine because of the promising antigenicity.

A genetic and virulence characterization of Brazilian strains of Mycoplasma hyopneumoniae.

Mycoplasma hyopneumoniae (M. hyopneumoniae) is considered the primary causative agent of porcine enzootic pneumonia (EP), a chronic contagious respiratory disease that causes economic losses. Obtaining new pathogenic isolates and studying the genome and virulence factors are necessary. This study performed a complete sequencing analysis of two Brazilian strains, UFV01 and UFV02, aiming to characterize the isolates in terms of the virulence factors and sequence type. The complete genome analysis revealed the main virulence genes (mhp385, mhp271, MHP_RS03455, p102, p97, p216, MHP_RS00555, mhp107) and ST-123, the presence of three toxin-related genes (tlyC, PLDc_2 and hcnC), and some genetic groups specific to these two isolates. Subsequently, the pathogenicity of the isolates was evaluated via an experimental infection conducted in a swine model. The study was divided into three groups, namely a negative control group (n = 4) and two test groups (n = 8), totaling 20 animals. They were challenged at 35 days of age with 107 CCU (Color Changing Units) M. hyopneumoniae via the intratracheal route. The UFV01 group showed earlier and higher seroconversion (IgG) (100%), while only 50% of the UFV02 group seroconverted. The same trend was observed when analyzing the presence of IgA in the bronchoalveolar lavage fluid (BALF) at 35 days post-infection (dpi). The UFV01 group had a mean macroscopic lesion score of 11.75% at 35 dpi, while UFV02 had 3.125%. Microscopic lesions were more severe in the UFV01 group. Based on laryngeal swab samples evaluated by qPCR, and the detection began at 14 days. The UFV01 group showed 75% positivity at 14 dpi. The UFV02 group also started excreting at 14 dpi, with a positivity rate of 37.5%. The results indicate that the UFV01 isolate exhibits higher virulence than UFV02. These findings may aid in developing new vaccines and diagnostic kits and establishing experimental models for testing.

Synthesis of new non-natural L-glycosidic flavonoid derivatives and their evaluation as inhibitors of Trypanosoma cruzi ecto-nucleoside triphosphate diphosphohydrolase 1 (TcNTPDase1).

Trypanosoma cruzi is the pathogen of Chagas disease, a neglected tropical disease that affects more than 6 million people worldwide. There are no vaccines to prevent infection, and the therapeutic arsenal is very minimal and toxic. The unique E-NTPDase of T. cruzi (TcNTPDase1) plays essential roles in adhesion and infection and is a virulence factor. Quercetin is a flavonoid with antimicrobial, antiviral, and antitumor activities. Its potential as a partial inhibitor of NTPDases has also been demonstrated. In this work, we synthesized the non-natural L-glycoside derivatives of quercetin and evaluated them as inhibitors of recombinant TcNTPDase1 (rTcNTPDase1). These compounds, and quercetin and miquelianin, a natural quercetin derivative, were also tested. Compound 16 showed the most significant inhibitory effect (94%). Quercetin, miquelianin, and compound 14 showed inhibition close to 50%. We thoroughly investigated the inhibitory effect of 16. Our data suggested a competitive inhibition with a Ki of 8.39 µM (± 0.90). To better understand the interaction of compound 16 and rTcNTPDase1, we performed molecular dynamics simulations of the enzyme and docking analyses with the compounds. Our predictions show that compound 16 binds to the enzyme's catalytic site and interacts with important residues for NTPDase activity. As an inhibitor of a critical T. cruzi enzyme, (16) could be helpful as a starting point in the developing of a future treatment for Chagas disease. Furthermore, the discovery of (16) as an inhibitor of TcNTPDase1 may open new avenues in the study and development of new inhibitors of E-NTPDases.

First evidence of a serine arginine protein kinase (SRPK) in leishmania braziliensis and its potential as therapeutic target.

Leishmaniasis is a parasitic disease found in tropical and subtropical regions around the world, caused by parasites of the genus Leishmania. The disease is a public health concern and presents clinical manifestations that can cause death, disability, and mutilation. The parasite has promastigote (vector) and amastigote (vertebrate host) forms and kinase enzymes are involved in this differentiation process. In the present investigation, we show, for the first time, evidence of a serine/arginine protein kinase in Leshmania braziliensis (LbSRPK). Our results show that amastigotes express more LbSRPK than promastigotes.  Analogues of SRPIN340 (a known inhibitor of SRPK) were evaluated for their leishmanicidal activity and two of them, namely SRVIC22 and SRVIC32 showed important leishmanicidal activity in vitro. SRVIC22 and SRVIC32 were able to reduce the infection rate in macrophages and the number of intracellular amastigotes by 55 and 60%, respectively. Bioinformatics analysis revealed the existence of two different amino acid residues in the active site of LbSRPK compared to their human homologue (Tyr/Leu-and Ser/Tyr), which could explain the absence of leishmanicidal activity of SRPIN340 on infected macrophages. In order to enhance leishmanicidal activity of the analogues, optimizations were proposed in the structures of the ligands, suggesting strong interactions with the catalytic site of LbSRPK. Although the evidence on the action of inhibitors upon LbSRPK is only indirect, our studies not only reveal, for the first time, evidence of a SRPK in Leishmania, but also shed light on a new therapeutic target for drug development.

Leishmania infantum NTPDase1 and NTPDase2 play an important role in infection and nitric oxide production in macrophages.

Leishmania infantum, the causative agent of American Visceral Leishmaniasis (VL), is known for its ability to modulate the host immune response to its own favor. Ecto-nucleoside triphosphate diphosphohydrolase (ENTPDase) represents a family of enzymes that hydrolyze nucleotides and are involved in nucleotide-dependent biological processes. L. infantum has two ENTPDases, namely LiNTPDase1 and LiNTPDase2. Here, we used genetic tools to overexpress or abolish the expression of LiNTPDase1 and -2 to assess their role in parasite growth in culture and macrophage infection. While LiNTPDase1 or 2-overexpressing clones showed no morphological or growth changes in promastigotes, LiNTPDase2 overexpression increased macrophage adhesion and infection by 50% and 30%, respectively. The individual LiNTPDase1 and 2 knockout mutants showed lag in growth profile, which was reversed by the addition of adenine and guanine to the culture media. Moreover, the morphology of the knockout mutants even in supplemented media was changed to an amastigote-like form. The double knockout of both genes was lethal and a mechanism of compensation of deletion of one isoform was detected in these mutants. Correspondingly, the absence of LiNTPDase1 or LiNTPDase2 led to a dramatic reduction in in vitro infection (∼90%). Interestingly, nitric oxide production was decreased in both knockout mutants during infection, which suggests that both LiNTPDases can inhibit macrophage responses against the parasite. Overall, our results show important roles of LiNTPDase1 and -2 concerning in vitro macrophage infection and reinforce their use as potential targets to control Leishmania infections.

Impaired expression of serine/arginine protein kinase 2 (SRPK2) affects melanoma progression.

Melanoma is one of the most aggressive tumors, and its lethality is associated with the ability of malignant cells to migrate and invade surrounding tissues to colonize distant organs and to generate widespread metastasis. The serine/arginine protein kinases 1 and 2 (SRPK1 and SRPK2) are classically related to the control of pre-mRNA splicing through SR protein phosphorylation and have been found overexpressed in many types of cancer, including melanoma. Previously, we have demonstrated that the pharmacological inhibition of SRPKs impairs pulmonary colonization of metastatic melanoma in mice. As the used compounds could target at least both SRPK1 and SRPK2, here we sought to obtain additional clues regarding the involvement of these paralogs in melanoma progression. We analyzed single-cell RNA sequencing data of melanoma patient cohorts and found that SRPK2 expression in melanoma cells is associated with poor prognosis. Consistently, CRISPR-Cas9 genome targeting of SRPK2, but not SRPK1, impaired actin polymerization dynamics as well as the proliferative and invasive capacity of B16F10 cells in vitro. In further in vivo experiments, genetic targeting of SRPK2, but not SRPK1, reduced tumor progression in both subcutaneous and caudal vein melanoma induction models. Taken together, these findings suggest different functional roles for SRPK1/2 in metastatic melanoma and highlight the relevance of pursuing selective pharmacological inhibitors of SRPK2.

Immunomodulatory activity of trifluoromethyl arylamides derived from the SRPK inhibitor SRPIN340 and their potential use as vaccine adjuvant.

The serine/arginine-rich protein kinases (SRPK) specifically phosphorylate their substrates at RS-rich dipeptides, which are abundantly found in SR splicing factors. SRPK are classically known for their ability to affect the splicing and expression of gene isoforms commonly implicated in cancer and diseases associated with infectious processes. Non-splicing functions have also been attributed to SRPK, which highlight their functional plasticity and relevance as therapeutic targets for pharmacological intervention. In this sense, different SRPK inhibitors have been developed, such as the well-known SRPIN340 and its derivatives, with anticancer and antiviral activities. Here we evaluated the potential immunomodulatory activity of SRPIN340 and three trifluoromethyl arylamide derivatives. In in vitro analysis with RAW 264.7 macrophages and primary splenocytes, all the compounds modulated the expression of immune response mediators and antigen-presentation molecules related to a tendency for M2 macrophage polarization. Immunization experiments were carried out in mice to evaluate their potential as vaccine immunostimulants. When administrated alone, the compounds altered the expression of immune factors at the injection site and did not produce macroscopic or microscopic local reactions. In addition, when prepared as an adjuvant with inactivated EHV-1 antigens, all the compounds increased the anti-EHV-1 neutralizing antibody titers, a change that is consistent with an increased Th2 response. These findings demonstrate that SRPIN340 and its derivatives exhibit a noticeable capacity to modulate innate and adaptative immune cells, disclosing their potential to be used as vaccine adjuvants or in immunotherapies.

The SRPK inhibitor N-(2-(piperidin-1-yl)-5-(trifluoromethyl)phenyl) isonicotinamide (SRPIN340) increases the immune response against metastatic melanoma in mice.

Cancers have a strong relationship with immune cells in their microenvironment, which significantly influences tumor proliferation and progression. Thus, pharmacological strategies that stimulate the immune system to combat tumor cells are promising for better therapeutic efficacy. Deregulated expression of the splicing regulatory serine arginine protein kinases (mostly SRPK1 and SRPK2) has been found in different cancer types, leading to the expression of isoforms related to tumor growth and metastasis. The microenvironment of melanoma exhibits a strong presence of immune cells, which significantly influences tumor progression, and around 50% of cutaneous melanoma patients benefit from targeted immunotherapy. Here, we analyzed human malignant melanoma single-cell gene expression data and observed that SRPK1/2 overexpression correlates with immune system pathway alterations. In further analysis, we observed an increased presence of immune cells in biopsies from mice bearing metastatic melanoma treated with SRPIN340, a well-known SRPK1/2 pharmacological inhibitor. Local treatments increased the expression of proinflammatory cytokines at the tumor lesions and the activity of the spleen, accompanied by reduced pulmonary metastasis foci, edema formation, and alveolar congestion. In in vitro assays, SRPIN340 also potentiated immunological susceptibility, by increasing the expression of the antigen presenting MHCI and MHCII molecules and by increasing the ability of B16F10 cells to attract splenic cells in transwell assays. Taken together, these results reveal that the antimetastatic effect of SRPIN340 can also involve an increased immune response, which suggests additional functional clues for SRPKs in tumor biology.

Urea, salts, and Tween 20 influence on adsorption of IgG and Leishmania rNTPDase2 to nitrocellulose.

Lateral flow immunochromatography is a widely used technique for immunological assays. Construction of test and control lines is mostly done by antigen adsorption to nitrocellulose membranes, a process not fully understood. This study aimed to evaluate the influence of urea, salts, and Tween 20, on adsorption. The performance of canine IgG in water and in buffer containing urea and salts (pH 8.3) were compared to observe if the interferents would lead to protein stripping when challenged with increasing concentrations of Tween 20 in the lateral flow buffer. Immobilization of the rLiNTPDase2, an antigen for Canine Leishmaniasis diagnosis, was evaluated and compared to the rLbNTPDase2 by the same method. There were no differences between adsorption coefficients of IgG in water and in buffer, but high salt and urea concentrations seems to stabilize and enhance IgG immobilization. Adsorption performance between canine IgG and rNTPDases had different patterns, but was highly similar between rNTPDases, indicating that protein identity may have an important role. Also, low concentrations of Tween 20 in the flow solution may aid the maintenance of rNTPDase2 on the strips. Our results bring insights about protein adsorption and perspectives about the influence of urea, salts and Tween 20 on this process.

Synthesis of cinnamic acid ester derivatives with antiproliferative and antimetastatic activities on murine melanoma cells.

Melanoma is the most aggressive skin cancer, and its incidence has continued to rise during the past decades. Conventional treatments present severe side effects in cancer patients, and melanoma can be refractory to commonly used anticancer drugs, which justify the efforts to find new potential anti-melanoma drugs. An alternative to promote the discovery of new pharmacological substances would be modifying chemical groups from a bioactive compound. Here we describe the synthesis of seventeen compounds derived from cinnamic acid and their bioactivity evaluation against melanoma cells. The compound phenyl 2,3-dibromo-3-phenylpropanoate (3q) was the most effective against murine B16-F10 cells, as observed in cytotoxicity and cell migration assays. Simultaneously, this compound showed low cytotoxic activity on non-tumor cells. At the highest concentration, the compound 3q was able to trigger apoptosis, whereas, at lower concentrations, it affected the cell cycle and melanoma cell proliferation. Furthermore, cinnamate 3q impaired cell invasion, adhesion, colonization, and actin polymerization. In conclusion, these results highlight the antiproliferative and antimetastatic potential of cinnamic acid derivatives on melanoma.

Apoptosis in the late replication phase of Bovine alphaherpesvirus 1 in experimentally infected calves.

Bovine alphaherpesvirus 1 (BoHV-1) is a pathogen causing respiratory and reproductive clinical signs in cattle. Infected animals may develop rhinotracheitis, vulvovaginitis, balanoposthitis, and abortion. Viral latency is generally established in neuronal ganglia simultaneously to a decrease in both genes or genome expression and viral replication. Under stressful conditions, infection is reactivated leading to viral replication and the manifestation of clinical signs. In this study, we evaluated both viral reactivation and apoptosis in trigeminal ganglia cells as BoHV-1 progressed from the latent to the acute phase of infection after dexamethasone administration in experimentally infected calves. To test ganglia cell death as a consequence of BoHV-1 infection, we stained the BoHV-1 samples with TUNEL after the viral shedding by the calves. RT-qPCR of apoptotic genes was also performed, showing the upregulation of the caspase 8 gene in the trigeminal ganglia from cattle experimentally infected with BoHV-1. These results showed the occurrence of apoptosis in ganglion cells of calves infected by BoHV-1.

Genetic diversity of porcine circovirus 3 strains and the first detection of two different PCV3 strains coinfecting the same host in Minas Gerais, Brazil.

Porcine circovirus 3 (PCV3) is a recently emerged circovirus discovered in 2016 that has drawn the attention of the swine industry worldwide. In this study, we evaluated the genetic diversity of PCV3 strains on pig farms. A total of 261 samples from sows, weaning pigs, growing pigs, and stillborn/mummified fetuses were analyzed by quantitative real-time PCR. The results revealed that at least two main lineages of PCV3 are circulating in Brazil. For the first time, it was possible to detect the presence of two different PCV3 strains in the same host.

Structural and molecular bases of angiotensin-converting enzyme inhibition by bovine casein-derived peptides: an in silico molecular dynamics approach.

The angiotensin-converting enzyme (ACE) plays a key role in blood pressure regulation process, and its inhibition is one of the main drug targets for the treatment of hypertension. Though various peptides from milk proteins are well-known for their ACE-inhibitory capacity, research devoted to understand the molecular bases of such property remain scarce, specifically for such peptides. Therefore, in this work, computational molecular docking and molecular dynamics calculations were performed to enlighten the intermolecular interactions involved in ACE inhibition by six different casein-derived peptides (FFVAPFPEVFGK, FALPQYLK, ALNEINQFYQK, YLGYLEQLLR, HQGLPQEVLNENLLR and NAVPITPTLNR). Two top ranked docking poses for each peptide (one with N- and the other C-terminal peptide extremity oriented towards the ACE active site) were selected for dynamic simulations (50 ns; GROMOS53A6 force field), and the results were correlated to in vitro ACE inhibition capacity. Two molecular features appeared to be essential for peptides to present high ACE inhibition capacity in vitro: i) to interact with the S1 active site residues (Ala354, Glu384, and Tyr523) by hydrogen bonds; ii) to interact with Zn2+ coordinated residues (His383, His387, and Glu411) by short-lenght hydrogen bonds, as observed in the cases of ALNEINQFYQK (IACE = 80.7%), NAVPITPTLNR (IACE = 80.7%), and FALPQYLK (IACE = 79.0%). Regardless of the temporal stability of these strong interactions, they promoted some disruption of Zn2+ tetrahedral coordination during the molecular dynamics trajectories, and were pointed as the main reason for the greatest ACE inhibition by these peptides. On the other hand, peptides with intermediate inhibition capacity (50% < IACE < 45%) interacted mainly by weaker interactions (e.g.: electrostatic and hydrophobic) with the Zn2+ coordinated residues, and were not able to change significantly its tetrahedral coordination structure. These findings may: i) assist the discrimination in silico of "good" and "bad" ACE-inhibitory peptides from other food sources, and/or ii) aid in designing de novo new molecules with ACE-inhibitory capacity. Communicated by Ramaswamy Sarma.

Detection and partial molecular characterization of Picobirnavirus in swine from the state of Minas Gerais, Brazil.

Picobirnavirus (PBV) is a small two-segmented double-stranded RNA (dsRNA) virus that has been identified in diarrheic feces of a large range of animal hosts, including humans. For this reason, PBV has been recognized as an opportunistic agent of gastrointestinal disease. Even under these circumstances, there is a lack of studies regarding this pathogen. Not outstanding, in Brazil, the single description of the PBV occurrence in pigs was provided in the 1980s. Hence, this study aimed to verify the PBV occurrence in Brazilian swine farms and to perform molecular characterization of the identified strains. High genetic variability was found in the analyzed sequences. Further studies comprehending the infection of swine by PBV in Brazilian herds should be performed to provide more accurate information on its epidemiology and to discuss the role of the virus in gastrointestinal diseases.

Antimicrobial susceptibility and genetic profile of Mycoplasma hyopneumoniae isolates from Brazil.

Mycoplasma hyopneumoniae is the etiologic agent of porcine enzootic pneumonia, responsible for major production losses worldwide. The bacteria have a limited metabolism and need to obtain molecules from the growth environment, which causes multiple difficulties for in vitro culture. These limitations have a negative influence on the ability to carry out research for the development of the rational use of antimicrobials and vaccines. The objective of this investigation was to evaluate the genetic profile and in vitro susceptibility of field isolates of M. hyopneumoniae to different antimicrobials. All 16 isolates obtained from the samples presented 100% of identity in the partial sequence of 16S rRNA gene when compared to M. hyopneumoniae. A dendrogram was created using the PCR results of the genes related to pathogenicity, and the isolates were distributed into four clusters, suggesting genetic variability among four different isolates circulating on the same farm. The minimum inhibitory concentration of the isolates was higher for the antimicrobials tylosin (< 0.001-16 mg/L) and spiramycin (< 0.001-16 mg/L) than for enrofloxacin (< 0.001-0.125 mg/L) and tiamulin (< 0.001-0.125 mg/L). Our results demonstrate the genetic variability among M. hyopneumoniae isolates from pigs of the same farm, with differences in their susceptibility to antimicrobial agents.

Correction to: Genetic variation of Mycoplasma hyopneumoniae from Brazilian field samples.

Following publication of the original article [1], we have been notified that two of the author names were incomplete or incorrect.

Genetic variation of Mycoplasma hyopneumoniae from Brazilian field samples.

BACKGROUND: Porcine enzootic pneumonia is a worldwide problem in swine production. The infected host demonstrates a respiratory disease whose etiologic agent is Mycoplasma hyopneumoniae (Mhp). A total of 266 lung samples with Mycoplasma-like lesions were collected from two slaughterhouses. We analyzed the genetic profile of Mhp field samples using 16 genes that encode proteins involved in the mechanisms of bacterial pathogenesis and/or the immune responses of the host. Bioinformatic analyses were performed to classify the Mhp field samples based on their similarity according to the presence of the studied genes. RESULTS: Our results showed variations in the frequency of the 16 studied genes among different Mhp field samples. It was also noted that samples from the same farm were genetically different from each other and samples from different regions could be genetically similar, which is evidence of the presence of different genetic profiles among the Mhp field strains that circulate in Brazilian swine herds. CONCLUSION: This work demonstrated the genetic diversity of several Mhp field strains based on 16 selected genes related to virulence and/or immune response in Brazil. Our findings demonstrate the difference between Mhp field strains could influence the virulence, and we hypothesize that the most frequent genes in Mhp field strains could possibly be used as vaccine candidates. Based on our results, we suspect that Mhp genetic variability may be associated with the frequency of genes among the field strains and we have demonstrated that some Mhp field samples could not have many important genes described in the literature.

Synthesis of cinnamic acid derivatives and leishmanicidal activity against Leishmania braziliensis.

Leishmania braziliensis is one of the pathogenic agents of cutaneous and mucocutanoeous leishmaniasis. There are no validated vaccines to prevent the infection and the treatment relies on drugs that often present severe side effects, which justify the efforts to find new potential antileishmanial drugs. An alternative to promote the discovery of new drugs would be the association of different chemical groups of bioactive compounds. Here we describe the synthesis and bioactivity evaluation against L. braziliensis of cinnamic acid derivatives possessing isobenzofuranone and 1,2,3-triazole functionalities. We tested 25 compounds at 10 µM concentration against extracellular promastigotes and intracellular amastigotes during macrophage infection. Most compounds were more active against amastigotes than to promastigotes. The derivatives (E)-3-oxo-1,3-dihydroisobenzofuran-5-yl-(3,4,5-trimethoxy) cinnamate (5c), (1-(3,4-difluorobenzyl)-1H-1,2,3-triazol-4-yl)methyl cinnamate (9g), and (1-(2-bromobenzyl)-1H-1,2,3-triazol-4-yl)methyl cinnamate (9l) were the most effective presenting over 80% toxicity on L. braziliensis amastigotes. While compound 5c is a cinnamate with an isobenzofuranone portion, 9g and 9l are triazolic cinnamic acid derivatives. The action of these compounds was comparable to amphotericin B used as positive control. Ultrastructural analysis revealed that 5c-treated parasites showed impaired cytokinesis and apoptosis triggering. Taken together, these results highlight the potential of cinnamic acid derivatives in development of novel anti-leishmanial drugs.

Feline coronavirus isolates from a part of Brazil: insights into molecular epidemiology and phylogeny inferred from the 7b gene.

The Feline coronavirus (FCoV) can lead to Feline infectious peritonitis (FIP), which the precise cause is still unknown. The theory of internal mutation suggests that a less virulent biotype of FCoV (FECV) would lead to another more pathogenic biotype (FIPV) capable of causing FIP. In this work, the 7b gene was amplified from 51 domestic cat plasma samples by semi-nested PCR and tested through phylogenetic and phylogeographical approaches. The 7b gene of Brazilian isolates displayed high conservation, a strong correlation between the geographic origin of the viral isolates and their genealogy, and its evolution was possibly shaped by a combination of high rates of nucleotide substitution and purifying selection.