Probiotics have been considered as a promising strategy to prevent various diseases in both humans and animals. This approach has gained interest in recent years as a potential means to control ...bovine mastitis. In a previous study, we found that several L. casei strains, including BL23, were able to inhibit the internalization of S. aureus, a major etiologic agent of mastitis, into bovine mammary epithelial cells (bMEC). This antagonism required a direct contact between L. casei and bMEC or S. aureus, suggesting the inhibition relied on interactions between L. casei cell surface components and bMEC. In this study, we have investigated the impact of some candidates which likely influence bacteria host cell interactions. We have shown that L. casei BL23 fbpA retained its inhibitory potential, indicating that L. casei BL23 antagonism did not rely (solely) on competition between S. aureus and L. casei fibronectin-binding proteins for adhesion to bMEC. We have then investigated the impact of four sortase mutants, srtA1, srtA2, srtC1 and srtC2, and a double mutant (srtA1-srtA2) on L. casei BL23 inhibitory potential. Sortases are responsible for the anchoring on the bacterial cell wall of LPXTG-proteins, which reportedly play an important role in bacteria-host cell interaction. All the srt mutants tested presented a reduced inhibition capacity, the most pronounced effect being observed with the srtA2 mutant. A lower internalization capacity of L. casei srtA2 into bMEC was also observed. This was associated with several changes at the surface of L. casei BL23 srtA2 compared to the wild type (wt) strain, including altered abundance of some LPXTG- and moonlighting proteins, and modifications of cell wall structure. These results strongly support the role of sortase A2 in L. casei BL23 inhibition against S. aureus internalization. Deciphering the contribution of the cell surface components altered in srtA2 strain in the inhibition will require further investigation.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
S. aureus
are among the main bacteria causing problems related to multidrug resistance in nosocomial infections. Therefore, it is necessary to carry out a reliable and rapid diagnosis for the ...identification of the bacteria and characterization of its susceptibility profile, especially vancomycin, which is an alternative treatment against multidrug-resistant (MDR)
S. aureus
. Thus, the goal of this study was to characterize isolates of
S. aureus
regarding the resistance and virulence and to check the susceptibility to vancomycin, through different methods, for comparative purposes. Seventeen antimicrobials were tested to assess the susceptibility profile. It was evaluated the presence of identification (
nuc
), resistance (
mecA
and
blaZ
), biofilm (
icaA
and
icaD
) and siderophore (
sfaD
and
sbnD
) genes. The susceptibility to vancomycin was evaluated by Minimum Inhibitory Concentration (MIC) by broth microdilution (BMD), E-test, commercial panel (Kit), and Phoenix equipment. Most
S. aureus
(93,33%) was classified as MDR. These isolates were 100% positive for
nuc, mecA
,
icaA
,
icaD,
and
sfaD
genes; 96.67% for
sbnD
and 33.33% for
blaZ
. In relation to BMD, all methods correctly classified the susceptibility of the isolates; however, regarding the exact MIC value for vancomycin, Phoenix showed agreement of 63.33%, E-test (33.33%) and Kit (26.66%). In conclusion, most of
S. aureus
was considered MDR. Also, they presented resistance, biofilm production, and siderophores genes, showing the pathogenic potential of these bacteria. Besides, the Phoenix test was considered the most effective, as it presents advantages, such as identification of the microorganism and a greater number of antimicrobials tested at a time.
Hybrid (
×
) table grape cultivars grown in the subtropics often fail to accumulate sufficient anthocyanins to achieve good uniform berry color. Growers of
table grapes in temperate regions generally ...use ethephon and, more recently, (
)-
-abscisic acid (
-ABA) to overcome this problem. The objective of this study was to determine if
-ABA applications at different timings and concentrations have an effect on anthocyanin regulatory and biosynthetic genes, pigment accumulation, and berry color of the Selection 21 cultivar, a new
×
hybrid seedless grape that presents lack of red color when grown in subtropical areas. Applications of
-ABA 400 mg/L resulted in a higher accumulation of total anthocyanins and of the individual anthocyaninsanthocyanins: delphinidin-3-glucoside, cyanidin-3-glucoside, peonidin-3-glucoside, and malvidin-3-glucoside in the berry skin and improved the color attributes of the berries. Treatment with two applications at 7 days after véraison (DAV) and 21 DAV of
-ABA 400 mg/L resulted in a higher accumulation of total anthocyanins in the skin of berries and increased the gene expression of
,
,
, and
and of the
and
transcription factors in the seedless grape cultivar.
The photocatalytic degradation of textile effluent was investigated using TiO
2
, ZnO, and Nb
2
O
5
catalysts under solar irradiation. The procedures were carried out at ambient conditions in April ...2014, with pH 3.0 and catalyst concentration of 0.250 g L
−1
. The photocatalytic activity of the oxides was evaluated by means of kinetic efficiency (rate constant and half-life time), chemical oxygen demand reduction, and absorbance reduction at 228, 254, 284, 310, 350, 500, and 660 nm (λ
máx
). Mineralization in terms of the formation of inorganic ions and toxicity reduction using bioassays with
Artemia salina
were performed. TiO
2
reduced the absorbance at 660 nm (λ
max
) after 300 min of solar irradiation around 94 and 93%; and 68 and 60% of COD, respectively. ZnO showed lower photocatalytic activity giving 64 and 42% of absorbance and COD reduction, respectively. The photocatalytic activity of Nb
2
O
5
was very close to TiO
2
-P25. In this sense, Nb
2
O
5
becomes a promising alternative to replace the commercial TiO
2
-P25. Bioassays confirmed the efficacy of treatment, increasing the lethal concentration of 27.59 (
in natura
) to 131.95% in the presence of Nb
2
O
5
.
Corynebacterium pseudotuberculosis, a facultative intracellular bacterial pathogen, is the etiological agent of caseous lymphadenitis (CLA), an infectious disease that affects sheep and goats and it ...is responsible for significant economic losses. The disease is characterized mainly by bacteria-induced caseous necrosis in lymphatic glands. New vaccines are needed for reliable control and management of CLA. Thus, the putative virulence factors SpaC, SodC, NanH, and PknG from C. pseudotuberculosis FRC41 may represent new target proteins for vaccine development and pathogenicity studies.
SpaC, PknG and NanH presented better vaccine potential than SodC after in silico analyses. A total of 136 B and T cell epitopes were predicted from the four putative virulence factors. A cluster analysis was performed to evaluate the redundancy degree among the sequences of the predicted epitopes; 57 clusters were formed, most of them (34) were single clusters. Two clusters from PknG and one from SpaC grouped epitopes for B and T-cell (MHC I and II). These epitopes can thus potentially stimulate a complete immune response (humoral and cellular) against C. pseudotuberculosis. Several other clusters, including two from NanH, grouped B-cell epitopes with either MHC I or II epitopes. The four target proteins were expressed in Escherichia coli. A purification protocol was developed for PknG expression.
In silico analyses show that the putative virulence factors SpaC, PknG and NanH present good potential for CLA vaccine development. Target proteins were successfully expressed in E. coli. A protocol for PknG purification is described.
The present study elucidates the mechanical performance of different designs of resin-bonded fixed dental prostheses made of lithium disilicate simulating masticatory loads of anterior or canine ...guidance. A three-dimensional model of maxilla was constructed containing central incisor and canine teeth, with edentulous space of the lateral incisor. Three designs of prosthesis were created: retained in central incisor (1-I), retained in canine (1-C) and fixed in both teeth (2-IC). The computational analysis was performed for load in canine and central incisor separately (100N, 45°). The tensile and shear stresses were calculated for the resin-bonded fixed dental prosthesis, bonding surface of each retainer and cement layer using 3D finite element analysis. The 20 highest stress values were analyzed using two-way ANOVA and post-hoc Tukey test, all with α = 5%. The computational analysis showed that 2-retainer resin-bonded fixed dental prosthesis presented the worst prognosis regardless of the mandibular movement. ANOVA showed that Mandibular movement*Retainer interaction influenced on the tensile and shear stresses values (p < 0.01). Higher stresses were observed in the connector region for all groups (13–82.2 MPa; 11–70.2 MPa). In order to reduce the stress concentration in the resin-bonded fixed dental prosthesis and the retainer made of lithium disilicate, the occlusion may serve as the selection criteria of the unitary abutment for better sustainability.
Sugarcane (Saccharum officinarum, Poaceae) is cultivated on a large scale in (sub)tropical regions such as Brazil and has considerable economic value for sugar and biofuel production. The plant is a ...rich substrate for endo- and epiphytic fungi. Black yeasts in the family Herpotrichiellaceae (Chaetothyriales) are colonizers of human-dominated habitats, particularly those rich in toxins and hydrocarbon pollutants, and may cause severe infections in susceptible human hosts. The present study assessed the diversity of Herpotrichiellaceae associated with sugarcane, using in silico identification and selective isolation. Using metagenomics, we identified 5833 fungal sequences, while 639 black yeast-like isolates were recovered in vitro. In both strategies, the latter fungi were identified as members of the genera Cladophialophora, Exophiala, and Rhinocladiella (Herpotrichiellaceae), Cyphellophora (Cyphellophoraceae), and Knufia (Trichomeriaceae). In addition, we discovered new species of Cladophialophora and Exophiala from sugarcane and its rhizosphere. The first environmental isolation of Cladophialophora bantiana is particularly noteworthy, because this species up to now is exclusively known from the human host where it mostly causes fatal brain disease in otherwise healthy patients.
Majoranolide, a butanolide isolated from the nonpolar fraction of an ethanol extract of Mezilaurus crassiramea (Lauraceae) fruits, is being reported for the first time in this genus and the third ...time in plants. Structurally identified from 1D and 2D NMR and HRESIMS data, majoranolide proved cytotoxic against cancer cells—MCF-7 and MDA-MB-231 (breast), HT-29 (colon), PC-3 (prostate), 786-0 (renal), and HL-60 (leukemia)—inhibiting growth in HL-60 cells (GI50 = 0.21 μM) and exhibiting higher selectivity for this line than for nonneoplastic NIH/3T3 murine fibroblasts. Effects on the cell cycle, caspase-3 activation, and plasma membrane integrity were evaluated by flow cytometry. Expression of genes related to apoptotic pathways (BAX, BCL2, BIRC5, and CASP8) was investigated using RT-qPCR. At 50 μM, majoranolide induced cell cycle arrest at G1 in 24 h increased the sub-G1 population in 48 h and increased caspase-3 activation in a time-dependent manner. The compound upregulated BAX and CASP8 transcription (proapoptotic genes) and downregulated BIRC5 (antiapoptotic). Loss of plasma membrane integrity in 30% of cells occurred at 48 h, but not at 24 h, characterizing gradual, programmed death. The results suggest that majoranolide cytotoxicity involves apoptosis induction in HL-60 cells, although other mechanisms may contribute to this cell death.
Although dysbiosis in the gut microbiota is known to be involved in several inflammatory diseases, whether any specific bacterial taxa control host response to inflammatory stimuli is still elusive. ...Here, we hypothesized that dysbiotic indigenous taxa could be involved in modulating host response to inflammatory triggers. To test this hypothesis, we conducted experiments in germ-free (GF) mice and in mice colonized with dysbiotic taxa identified in conventional (CV) mice subjected to chemotherapy-induced mucositis. First, we report that the absence of microbiota decreased inflammation and damage in the small intestine after administration of the chemotherapeutic agent 5-fluorouracil (5-FU). Also, 5-FU induced a shift in CV microbiota resulting in higher amounts of Enterobacteriaceae, including E. coli, in feces and small intestine and tissue damage. Prevention of Enterobacteriaceae outgrowth by treating mice with ciprofloxacin resulted in diminished 5-FU-induced tissue damage, indicating that this bacterial group is necessary for 5-FU-induced inflammatory response. In addition, monocolonization of germ-free (GF) mice with E. coli led to reversal of the protective phenotype during 5-FU chemotherapy. E. coli monocolonization decreased the basal plasma corticosterone levels and blockade of glucocorticoid receptor in GF mice restored inflammation upon 5-FU treatment. In contrast, treatment of CV mice with ciprofloxacin, that presented reduction of Enterobacteriaceae and E. coli content, induced an increase in corticosterone levels. Altogether, these findings demonstrate that Enterobacteriaceae outgrowth during dysbiosis impacts inflammation and tissue injury in the small intestine. Importantly, indigenous Enterobacteriaceae modulates host production of the anti-inflammatory steroid corticosterone and, consequently, controls inflammatory responsiveness in mice.
Probiotics have been adopted to treat and prevent various diseases in humans and animals. They were notably shown to be a promising alternative to prevent mastitis in dairy cattle. This inflammation ...of the mammary gland is generally of infectious origin and generates extensive economic losses worldwide. In a previous study, we found that Lactobacillus casei BL23 was able to inhibit the internalisation of Staphylococcus aureus, one of the major pathogens involved in mastitis, into bovine mammary epithelial cells (bMEC). In this study, we further explored the capacity of this strain to modulate the innate immune response of bovine mammary epithelial cells during S. aureus infection. L. casei BL23 was able to decrease the expression of several pro-inflammatory cytokines, including interleukins 6, 8, 1α and 1β and tumour necrosis factor alpha, in S. aureus-stimulated bMEC, 8 h post-infection. On the other hand, L. casei did not impair the induction of defensins, such as lingual antimicrobial peptide and defensin β1 in the presence of S. aureus, and even slightly increased the induction of tracheal antimicrobial peptide during S. aureus infection. Finally, this strain did not alter the expression of the pattern recognition receptor nucleotide-binding oligomerisation domain proteins (NOD2). This study demonstrates that L. casei BL23 displayed anti-inflammatory properties on S. aureus-stimulated bMEC. These results open the way to further characterisation of the BL23 probiotic potential in a bovine mammary gland context and to a better understanding of how all these beneficial properties combine in vivo to combat mastitis pathogens.
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