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We examine the effects of imperfect phase estimation of a reference signal on the bit error rate and mutual information over a communication channel influenced by fading and thermal noise. The Two-Wave Diffuse-Power (TWDP) model is utilized for statistical characterization of propagation environment where there are two dominant line-of-sight components together with diffuse ones. We derive novel analytical expression of the Fourier series for probability density function arising from the composite received signal phase. Further, the expression for the bit error rate is presented and numerically evaluated. We develop efficient analytical, numerical and simulation methods for estimating the value of the error floor and identifying the range of acceptable signal-to-noise ratio (SNR) values in cases when the floor is present during the detection of multilevel phase-shift keying (PSK) signals. In addition, we use Monte Carlo simulations in order to evaluate the mutual information for modulation orders two, four and eight, and identify its dependence on receiver hardware imperfections under the given channel conditions. Our results expose direct correspondence between bit error rate and mutual information value on one side, and the parameters of TWDP channel, SNR and phase noise standard deviation on the other side. The results illustrate that the error floor values are strongly influenced by the phase noise when signals propagate over a TWDP channel. In addition, the phase noise considerably affects the mutual information.

Cryptococcus neoformans (Cn) is a pathogenic yeast that is the leading cause of fungal meningitis in immunocompromised patients. Various Cn virulence factors, such as the enzyme laccase and its product melanin, phospholipase, and capsular polysaccharide have been identified. During a screen of knockout mutants, the gene resistance to aminocholesterol 1 (RTA1) was identified, the function of which is currently unknown in Cn. Rta1 homologs in S. cerevisiae belong to a lipid-translocating exporter family of fungal proteins with transmembrane regions and confer resistance to the antimicrobial agent 7-aminocholesterol when overexpressed. To determine the role of RTA1 in Cn, the knock-out (rta1Δ) and reconstituted (rta1Δ+RTA1) strains were created and phenotypically tested. RTA1 was involved in resistance to 7-aminocholesterol, and also in exocyst complex component 3 (Sec6)-mediated secretion of urease, laccase, and the major capsule component, glucuronoxylomannan (GXM), which coincided with significantly smaller capsules in the rta1Δ and rta1Δ+RTA1 strains compared to the wild-type H99 strain. Furthermore, RTA1 expression was reduced in a secretory 14 mutant (sec14Δ) and increased in an RNAi Sec6 mutant. Transmission electron microscopy demonstrated vesicle accumulation inside the rta1Δ strain, predominantly near the cell membrane. Given that Rta1 is likely to be a transmembrane protein located at the plasma membrane, these data suggest that Rta1 may be involved in both secretion of various fungal virulence factors and resistance to 7-aminocholesterol in Cn.

Devastating fires in Australia over 2019-20 decimated native fauna and flora, including koalas. The resulting population bottleneck, combined with significant loss of habitat, increases the vulnerability of remaining koala populations to threats which include disease. Chlamydia is one disease which causes significant morbidity and mortality in koalas. The predominant pathogenic species, Chlamydia pecorum, causes severe ocular, urogenital and reproductive tract disease. In marsupials, including the koala, gene expansions of an antimicrobial peptide family known as cathelicidins have enabled protection of immunologically naïve pouch young during early development. We propose that koala cathelicidins are active against Chlamydia and other bacteria and fungi. Here we describe ten koala cathelicidins, five of which contained full length coding sequences that were widely expressed in tissues throughout the body. Focusing on these five, we investigate their antimicrobial activity against two koala C. pecorum isolates from distinct serovars; MarsBar and IPTaLE, as well as other bacteria and fungi. One cathelicidin, PhciCath5, inactivated C. pecorum IPTaLE and MarsBar elementary bodies and significantly reduced the number of inclusions compared to the control (p

ABSTRACT Intracellular calcium (Ca2+) is crucial for signal transduction in Cryptococcus neoformans, the major cause of fatal fungal meningitis. The calcineurin pathway is the only Ca2+-requiring signaling cascade implicated in cryptococcal stress adaptation and virulence, with Ca2+ binding mediated by the EF-hand domains of the Ca2+ sensor protein calmodulin. In this study, we identified the cryptococcal ortholog of neuronal calcium sensor 1 (Ncs1) as a member of the EF-hand superfamily. We demonstrated that Ncs1 has a role in Ca2+ homeostasis under stress and nonstress conditions, as the ncs1Δ mutant is sensitive to a high Ca2+ concentration and has an elevated basal Ca2+ level. Furthermore, NCS1 expression is induced by Ca2+, with the Ncs1 protein adopting a punctate subcellular distribution. We also demonstrate that, in contrast to the case with Saccharomyces cerevisiae, NCS1 expression in C. neoformans is regulated by the calcineurin pathway via the transcription factor Crz1, as NCS1 expression is reduced by FK506 treatment and CRZ1 deletion. Moreover, the ncs1Δ mutant shares a high temperature and high Ca2+ sensitivity phenotype with the calcineurin and calmodulin mutants (cna1Δ and cam1Δ), and the NCS1 promoter contains two calcineurin/Crz1-dependent response elements (CDRE1). Ncs1 deficiency coincided with reduced growth, characterized by delayed bud emergence and aberrant cell division, and hypovirulence in a mouse infection model. In summary, our data show that Ncs1 has a significant role as a Ca2+ sensor in C. neoformans, working with calcineurin to regulate Ca2+ homeostasis and, consequently, promote fungal growth and virulence. IMPORTANCE Cryptococcus neoformans is the major cause of fungal meningitis in HIV-infected patients. Several studies have highlighted the important contributions of Ca2+ signaling and homeostasis to the virulence of C. neoformans. Here, we identify the cryptococcal ortholog of neuronal calcium sensor 1 (Ncs1) and demonstrate its role in Ca2+ homeostasis, bud emergence, cell cycle progression, and virulence. We also show that Ncs1 function is regulated by the calcineurin/Crz1 signaling cascade. Our work provides evidence of a link between Ca2+ homeostasis and cell cycle progression in C. neoformans.

Free‐space optical (FSO) systems have attracted much attention from both research and application perspectives owing to their many benefits, such as license‐free operation, low‐cost, and high data rates. This paper investigates the ergodic capacity of FSO systems, which is an important metric of system performance. The stochastic temporary laser‐beam blockage, pointing errors, and atmospheric turbulence are simultaneously considered. The results illustrate that the link blockage causes a decreased ergodic capacity. We show that to maximize the ergodic capacity, there is an optimal value of the laser‐beam radius at the waist, which largely depends on pointing errors; however, it is independent of the atmospheric turbulence and the probability of link blockage.

Invasive fungal pathogens cause more than 300 million serious human infections and 1.6 million deaths per year. A clearer understanding of the mechanisms by which these fungi cause disease is needed to identify novel targets for urgently needed therapies. Kinases are key components of the signaling and metabolic circuitry of eukaryotic cells, which include fungi, and kinase inhibition is currently being exploited for the treatment of human diseases. Inhibiting evolutionarily divergent kinases in fungal pathogens is a promising avenue for antifungal drug development. One such group of kinases is the phospholipase C1-dependent inositol polyphosphate kinases (IPKs), which act sequentially to transfer a phosphoryl group to a pre-phosphorylated inositol sugar (IP). This review focuses on the roles of fungal IPKs and their IP products in fungal pathogenicity, as determined predominantly from studies performed in the model fungal pathogen Cryptococcus neoformans, and compares them to what is known in non-pathogenic model fungi and mammalian cells to highlight potential drug targeting opportunities.

In this paper, an average bit error rate (BER) analysis of the free-space optical (FSO) system applying subcarrier intensity modulation with binary differential phase-shift keying is presented. Multiple receiver apertures are considered, when maximal-ratio combining (MRC) is employed. Intensity fluctuations due to atmospheric turbulence are modeled by Melaga (M) distribution, taking the pointing errors effect into account. Novel closed-form average BER expression is derived in terms of Meijer's G-function. Utilizing derived expression, numerical results are presented and confirmed by Monte Carlo simulations. The effects of atmospheric turbulence and pointing errors parameters, as well as the number of photodetectors, on the average BER performance are discussed. Employing multiple receivers with MRC leads to the improvement of the FSO system performance. Numerical results illustrate that this improvement depends on the FSO channel conditions. In addition, it is proved that SNR unbalance scenario can seriously deteriorate system performance.

Cryptococcus neoformans is a human fungal pathogen that adapts its metabolism to cope with limited oxygen availability, nutrient deprivation and host phagocytes. To gain insight into cryptococcal metabolism, we optimized a protocol for the Seahorse Analyzer, which measures extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) as indications of glycolytic and respiratory activities. In doing so we achieved effective immobilization of encapsulated cryptococci, established Rotenone/Antimycin A and 2-deoxyglucose as effective inhibitors of mitochondrial respiration and glycolysis, respectively, and optimized a microscopy-based method of data normalization. We applied the protocol to monitor metabolic changes in the pathogen alone and in co-culture with human blood-derived monocytes. We also compared metabolic flux in wild-type C. neoformans, its isogenic 5-PP-IP5/IP7-deficient metabolic mutant kcs1∆, the sister species of C. neoformans, Cryptococcus deuterogattii/VGII, and two other yeasts, Saccharomyces cerevisiae and Candida albicans. Our findings show that in contrast to monocytes and C. albicans, glycolysis and respiration are tightly coupled in C. neoformans and C. deuterogattii, as no compensatory increase in glycolysis occurred following inhibition of respiration. We also demonstrate that kcs1∆ has reduced metabolic activity that correlates with reduced mitochondrial function. Metabolic inflexibility in C. neoformans is therefore consistent with its obligate aerobe status and coincides with phagocyte tolerance of ingested cryptococcal cells.

In this paper, we study the outage performance of a complex system consisting of a free-space optical (FSO)/radio-frequency (RF) link. Using radio over free-space optics technology, the FSO link carries Worldwide Interoperability for Microwave Access (WiMAX) signals from a core network to a WiMAX base station, delivering traffic to multiple end RF users. A novel closed-form analytical expression for overall outage probability is derived when M-ary phase-shift keying (M-PSK) and M-ary quadrature amplitude modulation (M-QAM) are applied. The analysis is performed when the FSO link is under the influence of the Gamma-Gamma turbulence, path loss, and misalignment between the transmitter and receiver apertures, and the RF part is influenced by the Gamma-shadowed Nakagami-m multipath fading. To illustrate the usefulness of the derived expressions, we present some numerical results that enable us to estimate the effects of different transceiver and channel parameters on the outage probability. The results are used for optimizing the transmitter laser beam radius at the waist to achieve the minimal overall outage probability in the case of different conditions over the RF part. The numerical and simulation results show that multipath fading severity and shadowing spread over the RF part have a significant effect on the optimal value of the laser beam waist and can decrease the overall outage probability for several orders of magnitude.

Influenza A virus (IAV) infects millions of people annually and predisposes to secondary bacterial infections. Inhalation of fungi within the Cryptococcus complex causes pulmonary disease with secondary meningo-encephalitis. Underlying pulmonary disease is a strong risk factor for development of C. gattii cryptococcosis though the effect of concurrent infection with IAV has not been studied. We developed an in vivo model of Influenza A H1N1 and C. gattii co-infection. Co-infection resulted in a major increase in morbidity and mortality, with severe lung damage and a high brain fungal burden when mice were infected in the acute phase of influenza multiplication. Furthermore, IAV alters the host response to C. gattii, leading to recruitment of significantly more neutrophils and macrophages into the lungs. Moreover, IAV induced the production of type 1 interferons (IFN-α4/β) and the levels of IFN-γ were significantly reduced, which can be associated with impairment of the immune response to Cryptococcus during co-infection. Phagocytosis, killing of cryptococci and production of reactive oxygen species (ROS) by IAV-infected macrophages were reduced, independent of previous IFN-γ stimulation, leading to increased proliferation of the fungus within macrophages. In conclusion, IAV infection is a predisposing factor for severe disease and adverse outcomes in mice co-infected with C. gattii.

ABSTRACT Phosphate acquisition by fungi is regulated by the phosphate-sensing and acquisition (PHO) signaling pathway. Cryptococcus neoformans disseminates from the lung to the brain and is the commonest cause of fungal meningitis worldwide. To investigate the contribution of PHO signaling to cryptococcal dissemination, we characterized a transcription factor knockout strain (hlh3Δ/pho4Δ) defective in phosphate acquisition. Despite little similarity with other fungal Pho4 proteins, Hlh3/Pho4 functioned like a typical phosphate-responsive transcription factor in phosphate-deprived cryptococci, accumulating in nuclei and triggering expression of genes involved in phosphate acquisition. The pho4Δ mutant strain was susceptible to a number of stresses, the effect of which, except for alkaline pH, was alleviated by phosphate supplementation. Even in the presence of phosphate, the PHO pathway was activated in wild-type cryptococci at or above physiological pH, and under these conditions, the pho4Δ mutant had a growth defect and compromised phosphate uptake. The pho4Δ mutant was hypovirulent in a mouse inhalation model, where dissemination to the brain was reduced dramatically, and markedly hypovirulent in an intravenous dissemination model. The pho4Δ mutant was not detected in blood, nor did it proliferate significantly when cultured with peripheral blood monocytes. In conclusion, dissemination of infection and the pathogenesis of meningitis are dependent on cryptococcal phosphate uptake and stress tolerance at alkaline pH, both of which are Pho4 dependent. IMPORTANCE Cryptococcal meningitis is fatal without treatment and responsible for more than 500,000 deaths annually. To be a successful pathogen, C. neoformans must obtain an adequate supply of essential nutrients, including phosphate, from various host niches. Phosphate acquisition in fungi is regulated by the PHO signaling cascade, which is activated when intracellular phosphate decreases below a critical level. Induction of phosphate acquisition genes leads to the uptake of free phosphate via transporters. By blocking the PHO pathway using a Pho4 transcription factor mutant (pho4Δ mutant), we demonstrate the importance of the pathway for cryptococcal dissemination and the establishment of brain infection in murine models. Specifically, we show that reduced dissemination of the pho4Δ mutant to the brain is due to an alkaline pH tolerance defect, as alkaline pH mimics the conditions of phosphate deprivation. The end result is inhibited proliferation in host tissues, particularly in blood. Podcast: A podcast concerning this article is available.

This paper presents the study of the effects of imperfect reference signal recovery on the bit error rate (BER) performance of dual-branch switch-and-stay combining (SSC) and multibranch selection combining (SC) receivers in a generalized - fading channel. The average BER of binary and quaternary phase shift keying (BPSK and QPSK) is derived under the assumption that the reference carrier signal is extracted from the received modulated signal. For SSC receiver the optimal switching threshold (in a minimum BER sense) is numerically evaluated. Hereby we determine and discuss the simultaneous influence of the average signal-to-noise ratio (SNR) per bit, fading severity, product phase-locked loop (PLL) bandwidthbit duration (B_LT_b), switching threshold of SSC and diversity order of SC on BER performance. The influence of B_LT_b in different channel conditions and modulation formats is pointed out. The numerical results are confirmed by computer simulations.

ABSTRACT Inositol pyrophosphates (PP-IPs) comprising inositol, phosphate, and pyrophosphate (PP) are essential for multiple functions in eukaryotes. Their role in fungal pathogens has never been addressed. Cryptococcus neoformans is a model pathogenic fungus causing life-threatening meningoencephalitis. We investigate the cryptococcal kinases responsible for the production of PP-IPs (IP7/IP8) and the hierarchy of PP-IP importance in pathogenicity. Using gene deletion and inositol polyphosphate profiling, we identified Kcs1 as the major IP6 kinase (producing IP7) and Asp1 as an IP7 kinase (producing IP8). We show that Kcs1-derived IP7 is the most crucial PP-IP for cryptococcal drug susceptibility and the production of virulence determinants. In particular, Kcs1 kinase activity is essential for cryptococcal infection of mouse lungs, as reduced fungal burdens were observed in the absence of Kcs1 or when Kcs1 was catalytically inactive. Transcriptome and carbon source utilization analysis suggested that compromised growth of the KCS1 deletion strain (Δkcs1 mutant) in the low-glucose environment of the host lung is due to its inability to utilize alternative carbon sources. Despite this metabolic defect, the Δkcs1 mutant established persistent, low-level asymptomatic pulmonary infection but failed to elicit a strong immune response in vivo and in vitro and was not readily phagocytosed by primary or immortalized monocytes. Reduced recognition of the Δkcs1 cells by monocytes correlated with reduced exposure of mannoproteins on the Δkcs1 mutant cell surface. We conclude that IP7 is essential for fungal metabolic adaptation to the host environment, immune recognition, and pathogenicity. IMPORTANCE Cryptococcus neoformans is responsible for 1 million cases of AIDS-associated meningitis and ~600,000 deaths annually. Understanding cellular pathways responsible for pathogenicity might have an impact on new drug development. We characterized the inositol polyphosphate kinases Kcs1 and Asp1, which are predicted to catalyze the production of inositol pyrophosphates containing one or two diphosphate moieties (PP-IPs). Using gene deletion analysis and inositol polyphosphate profiling, we confirmed that Kcs1 and Asp1 are major IP6 and IP7 kinases, respectively. Kcs1-derived IP7, but not Asp1-derived IP8, is crucial for pathogenicity. Global expression profiling and carbon source utilization testing suggest that IP7-deficient cryptococci cannot adapt their metabolism to allow growth in the glucose-poor environment of the host lung, and consequently, fungal burdens are significantly reduced. Persistent asymptomatic Δkcs1 mutant infection correlated with decreased mannoprotein exposure on the Δkcs1 mutant surface and reduced phagocytosis. We conclude that IP7 is crucial for the metabolic adaptation of C. neoformans to the host environment and for pathogenicity.

BACKGROUND:Cryptococcus neoformans is the commonest cause of fungal meningitis, with a substantial mortality despite appropriate therapy. Quantitative culture of cryptococci in cerebrospinal fluid (CSF) during antifungal therapy is of prognostic value and has therapeutic implications, but is slow and not practicable in many resource-poor countries. METHODS:We piloted two rapid techniques for quantifying viable cryptococci using mixtures of live and heat-killed cryptococci cultured in vitro: (i) quantitative microscopy with exclusion staining using trypan blue dye, and (ii) flow cytometry, using the fluorescent dye 2'-7'-Bis-(2-carboxyethyl)-5-(6)-carboxyfluorescein, acetoxymethyl ester (BCECF-AM). Results were compared with standard quantitative cryptococcal cultures. Quantitative microscopy was also performed on cerebrospinal fluid (CSF) samples. RESULTS:Both microscopy and flow cytometry distinguished between viable and non-viable cryptococci. Cell counting (on log scale) by microscopy and by quantitative culture were significantly linearly associated (p