Marta Kicia

The lack of mitochondrial AtFtsH4 protease alters Arabidopsis leaf morphology at the late stage of rosette development under short-day photoperiod.

AtFtsH4 is one of four inner membrane-bound mitochondrial ATP-dependent metalloproteases in Arabidopsis thaliana, called AAA proteases, whose catalytic site is exposed to the intermembrane space. In the present study, we used a reverse-genetic approach to investigate the physiological role of the AtFtsH4 protease. We found that loss of AtFtsH4 did not significantly affect Arabidopsis growth under optimal conditions (long days); however, severe morphological and developmental abnormalities in late rosette development occurred under short-day conditions. The asymmetric shape and irregular serration of expanding leaf blades were the most striking features of the ftsh4 mutant phenotype. The severe abnormal morphology of the leaf blades was accompanied by ultrastructural changes in mitochondria and chloroplasts. These abnormalities correlated with elevated levels of reactive oxygen species and carbonylated mitochondrial proteins. We found that two classes of molecular chaperones, Hsp70 and prohibitins, were over-expressed in ftsh4 mutants during late vegetative growth under both short- and long-day conditions. Taken together, our data indicate that lack of AtFtsH4 results in impairment of organelle development and Arabidopsis leaf morphology under short-day conditions.

Pneumocystis jirovecii—from a commensal to pathogen: clinical and diagnostic review

Pneumocystis pneumonia is an opportunistic disease caused by invasion of unicellular fungus Pneumocystis jirovecii. Initially, it was responsible for majority of morbidity and mortality cases among HIV-infected patients, which later have been reduced due to the introduction of anti-retroviral therapy, as well as anti-Pneumocystis prophylaxis among these patients. Pneumocystis pneumonia, however, is still a significant cause of mortality among HIV-negative patients being under immunosuppression caused by different factors, such as transplant recipients as well as oncologically treated ones. The issue of pneumocystosis among these people is particularly emphasized in the article, since rapid onset and fast progression of severe symptoms result in high mortality rate among these patients, who thereby represent the group of highest risk of developing Pneumocystis pneumonia. In contrast, fungal invasion in immunocompetent people usually leads to asymptomatic colonization, which frequent incidence among healthy infants has even suggested the possibility of its association with sudden unexpected infant death syndrome. In the face of emerging strains with different epidemiological profiles resulting from genetic diversity, including drug-resistant genotypes, the colonization phenomenon desires particular attention, discussed in this article. We also summarize specific and sensitive methods, required for detection of Pneumocystis invasion and for distinguish colonization from the disease.

Gastroenteritis Caused by the Cryptosporidium Hedgehog Genotype in an Immunocompetent Man

ABSTRACT The Cryptosporidium hedgehog genotype, which has been reported previously in hedgehogs and horses, was identified as the cause of the diarrheal disease cryptosporidiosis in an immunocompetent man in the Czech Republic. This is the first report of human illness caused by the Cryptosporidium hedgehog genotype.

Concurrent Infection of the Urinary Tract with Encephalitozoon cuniculi and Enterocytozoon bieneusi in a Renal Transplant Recipient

ABSTRACT A urinary tract coinfection, caused by Encephalitozoon cuniculi genotype II and Enterocytozoon bieneusi genotype D, was identified in an HIV-seronegative renal transplant recipient kept under lifelong immunosuppression. To our knowledge, this is the first report describing concurrent infection with these two microsporidia species in organ transplant recipients.

Mitochondrial protease AtFtsH4 protects ageing Arabidopsis rosettes against oxidative damage under short-day photoperiod.

Mitochondrial AtFtsH4 protease, whose catalytic site is exposed to the intermembrane space, is one of four inner membrane-bound FtsH proteases in Arabidopsis. We found that the loss of AtFtsH4 altered Arabidopsis leaf morphology at the late stage of rosette growth under short-day photoperiod, while such changes were not observed in ftsh4 mutants grown under long days. These morphological changes were correlated with elevated levels of both reactive oxygen species (ROS) and carbonylated proteins, which strongly suggested that ageing ftsh4 plants experienced oxidative stress. This view was supported by the accumulation of electron-dense material, presumably containing aggregated oxidized proteins, in mitochondria of ftsh4 plants with the most strongly malformed leaf blades. Taken together, our data published in the May issue of Plant J 1 suggest a link between the lack of AtFtsH4 protease, oxidative stress and altered leaf morphology at the late rosette stage under short days. Here, we present evidence that the onset of altered leaf morphology in ftsh4 correlates with an increase in the abundance of AtFtsH4 transcript observed in wild-type Arabidopsis growing under the same conditions. We also discuss how the lack of AtFtsH4 may cause oxidative stress towards the end of the vegetative growth in short days.

The mitochondrial protease AtFTSH4 safeguards Arabidopsis shoot apical meristem function.

The shoot apical meristem (SAM) ensures continuous plant growth and organogenesis. In LD 30 °C, plants lacking AtFTSH4, an ATP-dependent mitochondrial protease that counteracts accumulation of internal oxidative stress, exhibit a puzzling phenotype of premature SAM termination. We aimed to elucidate the underlying cellular and molecular processes that link AtFTSH4 with SAM arrest. We studied AtFTSH4 expression, internal oxidative stress accumulation, and SAM morphology. Directly in the SAM we analysed H2O2 accumulation, mitochondria behaviour, and identity of stem cells using WUS/CLV3 expression. AtFTSH4 was expressed in proliferating tissues, particularly during the reproductive phase. In the mutant, SAM, in which internal oxidative stress accumulates predominantly at 30 °C, lost its meristematic fate. This process was progressive and stage-specific. Premature meristem termination was associated with an expansion in SAM area, where mitochondria lost their functionality. All these effects destabilised the identity of the stem cells. SAM termination in ftsh4 mutants is caused both by internal oxidative stress accumulation with time/age and by the tissue-specific role of AtFTSH4 around the flowering transition. Maintaining mitochondria functionality within the SAM, dependent on AtFTSH4, is vital to preserving stem cell activity throughout development.

Prevalence and molecular characteristics of urinary and intestinal microsporidia infections in renal transplant recipients

Transplant recipients have been identified as a new risk group for microsporidia infection. We characterize for the first time the prevalence of microsporidia in intestinal and urinary tracts of renal transplant recipients. Molecular examination of 86 patients showed that 25.5% of them were infected; 86% were confirmed to have pathogens in their urine and 45.5% in stool. Among positive patients, 32% had microsporidia confirmed in both urine and stool. Genotyping revealed Encephalitozoon cuniculi (59%) and Enterocytozoon bieneusi (23%) monoinfections as well as coinfections with both species (18%). Moreover, we found diarrhoea and fever as symptoms significantly associated with microsporidia presence. Our results indicate that microsporidial infection should be considered in the assessment of renal transplant recipients, especially in the urinary tract, even if asymptomatic. Molecular identification of microsporidia species is relevant because of their different susceptibility for treatment.

Pentacyclic triterpenes combined with ciprofloxacin help to eradicate the biofilm formed in vitro by Escherichia coli

Background & objectives: Ciprofloxacin is commonly used in clinical practice for the treatment of recurrent urinary tract infections caused by Escherichia coli. However, very often these recurrent infections are due to a failure in a complete eradication of the microorganisms colonizing the urinary tract, especially in catheterized patients. To enhance the bactericidal activity of ciprofloxacin against biofilm-forming uropathogenic E. coli (UPECs), we examined its effect in combination with two pentacyclic triterpenes – asiatic and ursolic acids. Methods: The anti-biofilm activity of ciprofloxacin and pentacyclic triterpenes - asiatic acid (AA) and ursolic acid (UA), as well as their synergistic effect were tested on two types of surfaces - polystyrene microtiter plates and silicone catheters. It was investigated using the time-killing and biofilm assays. Results: Anti-biofilm activity of ciprofloxacin was not observed on microtiter plates or on the catheters. Ciprofloxacin combined with ursolic acid inhibited the biofilm formation on microtitre plates. This mixture, however, did not express such a strong activity against the synthesis of biofilm on the surface of catheters. Ciprofloxacin combined with asiatic acid had very weak inhibiting effect on the synthesis of biofilm mass on microtitre plates as well as on the catheters. Despite this, both mixtures – ciprofloxacin and asiatic acid, as well as ciprofloxacin and ursolic acid, exhibited strong and significant impact on the eradication of mature biofilm (P < 0.05). Interpretation & conclusions: Although ciprofloxacin is recommended in the treatment of urinary tract infections caused by UPECs, but its efficacy is arguable. Subinhibitory concentrations of ciprofloxacin did not inhibit the formation of biofilm. Pentacyclic triterpenes used in combination with ciprofloxacin enhanced its anti-biofilm effectiveness. However, this anti-biofilm activity was found to depend on the type of surface on which biofilm was formed.

Comparison of the effects of subinhibitory concentrations of ciprofloxacin and colistin on the morphology of cardiolipin domains in Escherichia coli membranes

Membrane domains characterized by unique protein and lipid composition allow for compartmentalization and regulation of various biological processes. In Escherichia coli cardiolipin domains play a key role in the dynamic organization of bacterial membranes, and their distribution depends on the stage of the cell cycle. We studied the influence of subinhibitory concentrations of ciprofloxacin and colistin on the morphology and distribution of E. coli cardiolipin domains. Using the fluorescent dye 10-N-nonyl acridine orange we found that exposure of bacteria to ciprofloxacin significantly increased the percentage of filamentous cells with altered morphology of the cardiolipin domains, while colistin did not induce any significant changes. These results allow us to conclude that inhibition of DNA gyrase causes effects even at the bacterial membrane level and those changes can be easily visualized using 10-N-nonyl acridine orange.

Diversity of Pneumocystis jirovecii Across Europe: A Multicentre Observational Study

Pneumocystis jirovecii is an airborne human-specific ascomycetous fungus responsible for Pneumocystis pneumonia (PCP) in immunocompromised patients, affecting > 500,000 patients per year (www.gaffi.org). The understanding of its epidemiology is limited by the lack of standardised culture. Recent genotyping data suggests a limited genetic diversity of P. jirovecii. The objective of the study was to assess the diversity of P. jirovecii across European hospitals and analyse P. jirovecii diversity in respect to clinical data obtained from the patients. Genotyping was performed using six already validated short tandem repeat (STR) markers on 249 samples (median: 17 per centre interquartile range [11 − 20]) from PCP patients of 16 European centres. Mixtures of STR markers (i.e., ≥ 2 alleles for ≥ 1 locus) were detected in 67.6% (interquartile range [61.4; 76.5]) of the samples. Mixture was significantly associated with the underlying disease of the patient, with an increased proportion in HIV patients (78.3%) and a decreased proportion in renal transplant recipients (33.3%) (p < 0.001). The distribution of the alleles was significantly different (p < 0.001) according to the centres in three out of six markers. In analysable samples, 201 combinations were observed corresponding to 137 genotypes: 116 genotypes were country-specific; 12 in two; six in three; and two in four and one in five countries. Nine genotypes were recorded more than once in a given country. Genotype 123 (Gt123) was significantly associated with France (14/15, p < 0.001) and Gt16 with Belgium (5/5, p < 0.001). More specifically, Gt123 was observed mainly in France (14/15/16 patients) and in renal transplant patient (13/15). Our study showed the wide population diversity across Europe, with evidence of local clusters of patients harbouring a given genotype. These data suggest a specific association between genotype and underlying disease, with evidence of a different natural history of PCP in HIV patients and renal transplant recipients.