Candida krusei
attracts attention from medical professionals mainly for its intrinsic resistance to fluconazole and the limited number of drugs available to treat
C. krusei
vulvovaginal candidiasis. ...Miltefosine was demonstrated to have good antifungal activity both
in vitro
and
in vivo
. Here, we determined the susceptibility profiles of 57 clinical
C. krusei
isolates from vulvovaginal candidiasis patients and assessed the antifungal activity of miltefosine against
C. krusei
. All isolates were susceptible to voriconazole and itraconazole, whereas 1.8% of the isolates were of non-wild-type phenotype to amphotericin B. In contrast, miltefosine showed low MICs against all
C. krusei
isolates with fungicidal activity. The checkerboard assay showed that the synergistic effect of miltefosine in combination with amphotericin B was observed in 25% of the tested planktonic
C. krusei
isolates and 18.8% of the tested preformed biofilms, whereas miltefosine in combination with fluconazole showed indifferent interaction for all tested planktonic isolates. The presence of sorbitol in the broth microdilution assay did not influence the MIC values of miltefosine against
C. krusei
, but the presence of ergosterol increased the MIC values. Visible changes in cell content in cells treated with miltefosine were observed. We found that cells treated with miltefosine showed decreased cell viability and chromatin condensation under PI staining, which indicates that miltefosine may induce apoptosis-like cell death in
C. krusei
. In conclusion, we found miltefosine has a good activity against
C. krusei
isolates and exerts its fungicidal effect by binding to ergosterol in the cell membrane and inducing apoptosis.
is a common cause of
infections. In our present study, we investigated the antifungal susceptibility and molecular epidemiology of vaginal and non-vaginal
isolates. Seventy-six vaginal
strains ...isolated from patients with vulvovaginal candidiasis and 57 non-vaginal
isolates were collected at two hospitals in Shanghai, China. Antifungal susceptibility was examined using a broth microdilution method. Multilocus sequence typing was used for genotyping. Overall, 28 (21.1%), 28 (21.1%), and 29 (21.8%)
isolates were resistant to fluconazole, itraconazole, and voriconazole, respectively. Briefly, 18 (23.7%), 18 (23.7%), and 19 (25%) vaginal strains were resistant to fluconazole, itraconazole, and voriconazole. While the resistance to these antifungals were all 17.5% (10/57) in non-vaginal strains. All isolates retained susceptibility to amphotericin B, and only four non-vaginal isolates were caspofungin resistant. Genotyping identified 17 ST patterns. In non-vaginal samples, the same genotypes appear as in the vaginal samples, except for one genotype (ST-182), while in the vaginal samples more genotypes appear (ST8, ST19, ST45, ST55, ST66, ST80, ST138, and ST17). The most common genotype was ST7 (81 strains), followed by ST10 (14 strains) and ST15 (11 strains). The majority of resistant phenotype strains (25/30, 83.3%) correlated to the predominant genotype (ST7), and the rest belonged to ST3 (2/30, 6.7%), ST10 (1/30, 3.3%), ST19 (1/30, 3.3%), and ST45 (1/30, 3.3%). Our survey revealed cross-resistance in vaginal and non-vaginal
isolates. Moreover, there is no genotype associated with the resistance phenotype.
is an opportunistic fungal pathogen with low susceptibility to current antifungal drugs. Here, we tested the
susceptibility of 8 drugs against 42 clinical
isolates. All isolates showed high MICs to ...voriconazole (MIC
>16 μg/ml), itraconazole (MIC
>16 μg/ml), posaconazole (MIC
>16 μg/ml), isavuconazole (MIC
>16 μg/ml), amphotericin B (MIC
>16 μg/ml), and terbinafine (MIC
>64 μg/ml) and high minimum effective concentrations (MECs) to micafungin (MEC
>8 μg/ml), with the exception of miltefosine showing an MIC
value of 4 μg/ml. We examined six different
drug combinations and found that the combination of voriconazole and terbinafine achieved the most synergistic effort against
We then annotated the
whole genome and located its
and
genes. We completely sequenced the two genes to determine if any mutation would be related to azole and echinocandin resistance in
We found no amino acid changes in Cyp51 protein and no tandem repeats in the 5' upstream region of the
gene. However, we identified three intrinsic amino acid residues (G138S, M220I, and T289A) in the Cyp51 protein that were linked to azole resistance. Likewise, two intrinsic amino acid residues (F639Y, W695F) that have reported to confer echinocandin resistance were found in Fks1 hot spot regions. In addition, three new amino acid alterations (D440A, S634R, and H1245R) were found outside Fks1 hot spot regions, and their contributions to echinocandin resistance need future investigation. Overall, our findings support the notion that
is intrinsically resistant to azoles and echinocandins.
Approximately 10% of humans with anophthalmia (absent eye) or severe microphthalmia (small eye) show haploid insufficiency due to mutations in SOX2, a SOXB1-HMG box transcription factor. However, at ...present, the molecular or cellular mechanisms responsible for these conditions are poorly understood. Here, we directly assessed the requirement for SOX2 during eye development by generating a gene-dosage allelic series of Sox2 mutations in the mouse. The Sox2 mutant mice display a range of eye phenotypes consistent with human syndromes and the severity of these phenotypes directly relates to the levels of SOX2 expression found in progenitor cells of the neural retina. Retinal progenitor cells with conditionally ablated Sox2 lose competence to both proliferate and terminally differentiate. In contrast, in Sox2 hypomorphic/null mice, a reduction of SOX2 expression to <40% of normal causes variable microphthalmia as a result of aberrant neural progenitor differentiation. Furthermore, we provide genetic and molecular evidence that SOX2 activity, in a concentration-dependent manner, plays a key role in the regulation of the NOTCH1 signaling pathway in retinal progenitor cells. Collectively, these results show that precise regulation of SOX2 dosage is critical for temporal and spatial regulation of retinal progenitor cell differentiation and provide a cellular and molecular model for understanding how hypomorphic levels of SOX2 cause retinal defects in humans.
Carbapenem-resistant
Klebsiella pneumoniae
(CRKP), a pathogen that causes severe nosocomial infections and yields a high mortality rate, poses a serious threat to global public health due to its high ...antimicrobial resistance. Bacteriophages encode polysaccharide-degrading enzymes referred to as depolymerases that cleave the capsular polysaccharide (CPS), one of the main virulence factors of
K. pneumoniae
. In this study, we identified and characterized a new capsule depolymerase K19-Dpo41 from
K. pneumoniae
bacteriophage SH-KP156570. Our characterization of K19-Dpo41 demonstrated that this depolymerase showed specific activities against K19-type
K. pneumoniae
. K19-Dpo41-mediated treatments promoted the sensitivity of a multidrug-resistant K19-type
K. pneumoniae
strain to the bactericidal effect of human serum and significantly increased the survival rate of
Galleria mellonella
infected with K19-type
K. pneumoniae
. Our results provided strong primary evidence that K19-Dpo41 was not only effective in capsular typing of K19-type
K. pneumoniae
but promising in terms of developing new alternative therapeutic strategies against K19-type CRKP infections in the future.
Adrenomedullin (AM) is a multifunctional peptide vasodilator that signals through a G-protein-coupled receptor when the receptor, called calcitonin receptor-like receptor (CL), is associated with a ...receptor activity-modifying protein 2 (RAMP2). We demonstrated previously that haploinsufficieny for each of these genes led to reduced maternal fertility, and that even a modest genetic reduction of AM peptide caused maternal defects in implantation, placentation, and fetal growth. Here, we further demonstrate that Adm⁺/⁻ female mice displayed reduced pregnancy success rates that were not caused by defects in folliculogenesis, ovulation, or fertilization. The poor fertility of Adm⁺/⁻ female mice could not be rescued by transfer of wild-type blastocysts, which suggested an underlying defect in uterine receptivity. In fact, we found that Adm, Calcrl, and Ramp2 gene expressions are tightly and spatiotemporally regulated in the luminal epithelial cells of the uterus during the estrus cycle and the peri-implantation period. RAMP3, which also generates an AM receptor when associated with CL, had a diametrically opposite expression pattern than that of Adm, Calcrl, and Ramp2 and was most robustly induced in the stroma of the uterus. Finally, we discovered that Adm⁺/⁻ female mice have a substantially reduced number of pinopodes on the uterine luminal epithelial surface, which is indicative and possibly causative of the poor uterine receptivity. Taken together, our studies identify a new class of pharmacologically tractable proteins that are involved in establishing uterine receptivity through the regulation of pinopode formation.
Recent studies have demonstrated that the topography of thalamocortical (TC) axon projections is initiated before they reach the cortex, in the ventral telencephalon (VTel). However, at this point, ...the molecular mechanisms patterning the topography of TC projections in the VTel remains poorly understood. Here, we show that a long-range, high-rostral to low-caudal gradient of Netrin-1 in the VTel is required in vivo for the topographic sorting of TC axons to distinct cortical domains. We demonstrate that Netrin-1 is a chemoattractant for rostral thalamic axons but functions as a chemorepulsive cue for caudal thalamic axons. In accordance with this model, DCC is expressed in a high-rostromedial to low-caudolateral gradient in the dorsal thalamus (DTh), whereas three Unc5 receptors (Unc5A-C) show graded expression in the reverse orientation. Finally, we show that DCC is required for the attraction of rostromedial thalamic axons to the Netrin-1-rich, anterior part of the VTel, whereas DCC and Unc5A/C receptors are required for the repulsion of caudolateral TC axons from the same Netrin-1-rich region of the VTel. Our results demonstrate that a long-range gradient of Netrin-1 acts as a counteracting force from ephrin-A5 to control the topography of TC projections before they enter the cortex.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Carbapenem-resistant Klebsiella pneumoniae (CRKP) has caused wide dissemination among pediatric patients globally and thus has aroused public concern. Here, we investigated the clinical ...epidemiological characteristics of 140 nonreplicate clinical K. pneumoniae strains isolated from pediatric patients between January and December 2021. Of all isolates, 16.43% (23 of 140) were CRKP strains, which predominantly contained KPC carbapenemase.
sequencing demonstrated that KL47 (65.22%, 15 of 23) was the most frequent capsular type, followed by KL64 (17.39%, 4 of 23). A total of 23 CRKP strains were classified into three different O-genotypes, including OL101 (65.22%, 15 of 23), O1 (26.09%, 6 of 23), and O3 (8.7%, 2 of 23). Interestingly, KL47 strains were strongly associated with OL101, while KL64 strains were all linked with O1. Some capsule-deficient strains were identified by serological typing, phage-typing, depolymerase-typing, and uronic acid assay. In this study, compared with healthy children, higher titers of anti-capsular polysaccharides (CPS) IgG were first detected in the sera of K47 and K64 K. pneumoniae-infected children, which had the effective bactericidal activity against corresponding serotype K. pneumoniae strains. These findings will facilitate the development of novel therapeutic and vaccine strategies against K. pneumoniae infection in children.
The emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) strains resistant to numerous antibiotics and the limited therapeutic options available have become an urgent health threat to the immunocompromised pediatric population. Vaccines and antibodies, especially those targeting capsular polysaccharides, may be novel and effective prevention and treatment options. Thus, it is important to understand the spread of CRKP in pediatric populations. This research presents OL101:KL47 and O1:KL64 as the predominant combinations among CRKP strains in children in Shanghai, China. The primary carbapenemase gene is KPC in CRKP strains. Additionally, this study found elevated levels of anti-CPS IgG against K47 and K64 K. pneumoniae strains in pediatric patients for the first time. The significant bactericidal activity of these anti-CPS IgGs was confirmed.
Caspase-3 is a major cell death effector protease in the adult and neonatal nervous system. We found a greater number and higher density of cells in the cortex of caspase-3-/-adult mice, consistent ...with a defect in developmental cell death. Caspase-3-/-mice were also more resistant to ischemic stress both in vivo and in vitro. After 2 h of ischemia and 48 h of reperfusion, cortical infarct volume was reduced by 55%, and the density of terminal deoxynucleotidyl-transferase-mediated dUTP nick end labeling-positive cells was decreased by 36% compared with wild type. When subjected to oxygen-glucose deprivation (2 h), cortical neurons cultured from mice deficient in caspase-3 expression were also more resistant to cell death by 59%. Mutant brains showed caspase-specific poly-(ADP-ribose) polymerase cleavage product (85-kDa fragment) in vivo and in vitro, suggesting redundant mechanisms and persistence of caspase-mediated cell death. In the present study, we found that caspase-8 mediated poly(ADP-ribose) polymerase cleavage in caspase-3-/-neurons in vivo and in vitro. In addition, mutant neurons showed no evidence of compensatory activation by caspase-6 or caspase-7 after ischemia. Taken together, these data extend the pharmacological evidence supporting an important role for caspase-3 and caspase-8 as cell death mediators in mammalian cortex and indicate the potential advantages of targeting more than a single caspase family member to treat ischemic cell injury.
Parkinson's disease (PD) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity are both associated with dopaminergic neuron death in the substantia nigra (SN). Apoptosis has been ...implicated in this cell loss; however, whether or not it is a major component of disease pathology remains controversial. Caspases are a major class of proteases involved in the apoptotic process. To evaluate the role of caspases in PD, we analyzed caspase activation in MPTP-treated mice, in cultured dopaminergic cells, and in postmortem PD brain tissue. MPTP was found to elicit not only the activation of the effector caspase-3 but also the initiators caspase-8 and caspase-9, mitochondrial cytochrome c release, and Bid cleavage in the SN of wild-type mice. These changes were attenuated in transgenic mice neuronally expressing the general caspase inhibitor protein baculoviral p35. These mice also displayed increased resistance to the cytotoxic effects of the drug. MPTP-associated toxicity in culture was found temporally to involve cytochrome c release, activation of caspase-9, caspase-3, and caspase-8, and Bid cleavage. Caspase-9 inhibition prevented the activation of both caspase-3 and caspase-8 and also inhibited Bid cleavage, but not cytochrome c release. Activated caspase-8 and caspase-9 were immunologically detectable within MPP(+)-treated mesencephalic dopaminergic neurons, dopaminergic nigral neurons from MPTP-treated mice, and autopsied Parkinsonian tissue from late-onset sporadic cases of the disease. These data demonstrate that MPTP-mediated activation of caspase-9 via cytochrome c release results in the activation of caspase-8 and Bid cleavage, which we speculate may be involved in the amplification of caspase-mediated dopaminergic cell death. These data suggest that caspase inhibitors constitute a plausible therapeutic for PD.
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