The Molecular Basis and Clinical Consequences of Chronic Inflammation in Prostatic Diseases: Prostatitis, Benign Prostatic Hyperplasia, and Prostate Cancer.

Chronic inflammation is now recognized as one of the major risk factors and molecular hallmarks of chronic prostatitis, benign prostatic hyperplasia (BPH), and prostate tumorigenesis. However, the molecular mechanisms by which chronic inflammation signaling contributes to the pathogenesis of these prostate diseases are poorly understood. Previous efforts to therapeutically target the upstream (e.g., TLRs and IL1-Rs) and downstream (e.g., NF-κB subunits and cytokines) inflammatory signaling molecules in people with these conditions have been clinically ambiguous and unsatisfactory, hence fostering the recent paradigm shift towards unraveling and understanding the functional roles and clinical significance of the novel and relatively underexplored inflammatory molecules and pathways that could become potential therapeutic targets in managing prostatic diseases. In this review article, we exclusively discuss the causal and molecular drivers of prostatitis, BPH, and prostate tumorigenesis, as well as the potential impacts of microbiome dysbiosis and chronic inflammation in promoting prostate pathologies. We specifically focus on the importance of some of the underexplored druggable inflammatory molecules, by discussing how their aberrant signaling could promote prostate cancer (PCa) stemness, neuroendocrine differentiation, castration resistance, metabolic reprogramming, and immunosuppression. The potential contribution of the IL1R-TLR-IRAK-NF-κBs signaling molecules and NLR/inflammasomes in prostate pathologies, as well as the prospective benefits of selectively targeting the midstream molecules in the various inflammatory cascades, are also discussed. Though this review concentrates more on PCa, we envision that the information could be applied to other prostate diseases. In conclusion, we have underlined the molecular mechanisms and signaling pathways that may need to be targeted and/or further investigated to better understand the association between chronic inflammation and prostate diseases.

Dietary Chitin Particles Called Mimetic Fungi Ameliorate Colitis in Toll-Like Receptor 2/CD14- and Sex-Dependent Manners.

Chitin is a natural N-acetylglucosamine polymer and a major structural component of fungal cell walls. Dietary chitin is mucoadhesive; anti-inflammatory effects of chitin microparticles (CMPs; 1- to 10-µm diameters) have been demonstrated in models of inflammatory bowel disease (IBD). The goals of this study were to assess (i) whether CMPs among various chitin preparations are the most effective against colitis in male and female mice and (ii) whether host chitin-binding Toll-like receptor 2 (TLR2) and CD14 are required for the anti-inflammatory effect of chitin. We found that colitis in male mice was ameliorated by CMPs and large chitin beads (LCBs; 40 to 70 µm) but not by chitosan (deacetylated chitin) microparticles, oligosaccharide chitin, or glucosamine. In fact, LCBs were more effective than CMPs. In female colitis, on the other hand, CMPs and LCBs were equally and highly effective. Neither sex of TLR2-deficient mice showed anti-inflammatory effects when treated with LCBs. No anti-inflammatory effect of LCBs was seen in either CD14-deficient males or females. Furthermore, an in vitro study indicated that when LCBs and CMPs were digested with stomach acidic mammalian chitinase (AMC), their size-dependent macrophage activations were modified, at least in part, suggesting reduced particle sizes of dietary chitin in the stomach. Interestingly, stomach AMC activity was greater in males than females. Our results indicated that dietary LCBs were the most effective preparation for treating colitis in both sexes; these anti-inflammatory effects of LCBs were dependent on host TLR2 and CD14.

The Invasive Brazilian Pepper Tree (Schinus terebinthifolius) Is Colonized by a Root Microbiome Enriched With Alphaproteobacteria and Unclassified Spartobacteria.

Little is known about the rhizosphere microbiome of the Brazilian pepper tree (BP) - a noxious category 1 invasive plant inducing an enormous economic and ecological toll in Florida. Some invasive plants have been shown to drastically change the soil microbiome compared to other native plants. The rhizobacteria community structure of BP, two Florida native plants (Hamelia patens and Bidens alba) and bulk soils were characterized across six geographical sites. Although all 19 well-known and 10 poorly described phyla were observed in all plant rhizospheres, BP contained the least total bacterial abundance (OTUs) with a distinct bacteria community structure and clustering patterns differing significantly (pCOA and PERMANOVA) from the natives and bulk soil. The BP rhizosphere community contained the highest overall Proteobacteria diversity (Shannon's diversity 3.25) in spite of a twofold reduction in richness of the Gammaproteobacteria. Remarkably, the invasive BP rhizosphere was highly enriched with Alphaproteobacteria, dominated by Rhizobiales, including Rhodoplanes and Bradyrhizobiaceae. Also, the relative abundance of Spartobacteria under BP rhizosphere was more than twice that of native plants and bulk soil; featuring unique members of the family Chthoniobacteraceae (DA101 genus). The trend was different for the family Pedosphaerae in the phylum Verrucomicrobia where the abundance declined under BP (26%) compared to (33-66%) for the H. patens native plant and bulk soil. BP shared the lowest number of unique phylotypes with bulk soil (146) compared to the other native plants with bulk soil (B. alba - 222, H. patens - 520) suggestive of its capacity to overcome biotic resistance. Although there were no specific biomarkers found, taken together, our data suggests that the occurrence of key bacteria groups across multiple taxonomic ranks provides a somewhat consistent profile of the invasive BP rhizo-community. Furthermore, based on the observed prevalence of a bacteria group (Spartobacteria - Chthoniobacteraceae - DA101); we propose that they have a possible role in BP biology. Our results emphasize the need to further investigate the potential value of "unique phylotypes" in the rhizosphere relative to bulk soil as an ecological tool for monitoring plant-cover/invasion history; or even detecting exotic plants with invasion tendencies.

Tobacco plants expressing the maize nitrate transporter ZmNrt2.1 exhibit altered responses of growth and gene expression to nitrate and calcium.

BACKGROUND: Nitrate uptake is a highly regulated process. Understanding the intricate interactions between nitrate availability and genetically-controlled nitrate acquisition and metabolism is essential for improving nitrogen use efficiency and increasing nitrate uptake capacity for plants grown in both nitrate-poor and nitrate-enriched environments. In this report, we introduced into tobacco (Nicotiana tabacum) the constitutively expressed maize high-affinity transporter ZmNrt2.1 gene that would bypass the tight control for the endogenous nitrate-responsive genes. By using calcium inhibitors and varying levels of NO3-, Ca2+ and K+, we probed how the host plants were affected in their nitrate response. RESULTS: We found that the ZmNrt2.1-expressing plants had better root growth than the wild type plants when Ca2+ was deficient regardless of the nitrate levels. The growth restriction associated with Ca2+-deficiency can be alleviated with a high level of K+. Furthermore, the transgenic plants exhibited altered expression patterns of several endogenous, nitrate-responsive genes, including the high- and low-affinity nitrate transporters, the Bric-a-Brac/Tramtrack/Broad protein BT2 and the transcription factor TGA-binding protein TGA1, in responding to treatments of NO3-, K+ or inhibitors for the calcium channel and the cytosolic Ca2+-regulating phospholipase C, as compared to the wild type plants under the same treatments. Their expression was not only responsive to nitrate, but also affected by Ca2+. There were also different patterns of gene expression between roots and shoots. CONCLUSION: Our results demonstrate the ectopic effect of the maize nitrate transporter on the host plant's overall gene expression of nitrate sensing system, and further highlight the involvement of calcium in nitrate sensing in tobacco plants.

Arbuscular and Ectomycorrhizal Fungi Associated with the Invasive Brazilian Pepper Tree (Schinus terebinthifolius) and Two Native Plants in South Florida.

The potential role of soil fungi in the invasion of the Brazilian pepper tree (Schinus terebinthifolius-BP) in Florida is not known; although the low biotic resistance of Florida soils is often invoked to explain the prevalence of many invasive species. To gain an initial insight into BP's mycorrhizal associations, this study examined the rhizobiome of BP and two native plants (Hamelia patens and Bidens alba) across six locations. Arbuscular mycorrhizal fungi (AMF) associated with the roots of the target plants and bulk soil was characterized by spore morphotyping. Sequence analysis of metagenomic DNA from lateral roots/rhizosphere of BP (n = 52) and a native shrub H. patens (n = 37) on the same parcel yielded other fungal associates. Overall, the total population of AMF associated with BP was about two folds greater than that of the two native plants (p = 0.0001) growing on the same site. The dominant AMF under Schinus were members of the common Glomus and Rhizophagus spp. By contrast, the most prevalent AMF in the bulk soil and rhizosphere of the two Florida native plants, Acaulospora spp (29%) was sharply diminished (9%) under BP rhizosphere. Analysis of the ITS2 sequences also showed that Schinus rhizosphere had a high relative abundance of ectomycorrhizal fungi (76.5%) compared to the native H. patens (2.6%), with the species Lactifluus hygrophoroides (Basidiomycota) being the most prevalent at 61.5% (p < 0.05). Unlike the native plants where pathogenic fungi like Phyllosticta sp., Phoma sp., and Neofusicoccum andium were present (8.1% for H. patens), only one potentially pathogenic fungal taxon was detected (3.9%) under BP. The striking disparity in the relative abundance of AMF and other fungal types between BP and the native species is quite significant. Fungal symbionts could aide plant invasion via resource-use efficiency and other poorly defined mechanisms of protection from pathogens in their invaded range. This report exposes a potentially significant but previously unappreciated fungal dimension of a complex invasion system and underscores the need to characterize these fungal symbionts, their role and mode of action during invasion; with the goal of devising measures for invasion control and ecological restoration.

Strategies in Ebola virus disease (EVD) diagnostics at the point of care.

Ebola virus disease (EVD) is a devastating, highly infectious illness with a high mortality rate. The disease is endemic to regions of Central and West Africa, where there is limited laboratory infrastructure and trained staff. The recent 2014 West African EVD outbreak has been unprecedented in case numbers and fatalities, and has proven that such regional outbreaks can become a potential threat to global public health, as it became the source for the subsequent transmission events in Spain and the USA. The urgent need for rapid and affordable means of detecting Ebola is crucial to control the spread of EVD and prevent devastating fatalities. Current diagnostic techniques include molecular diagnostics and other serological and antigen detection assays; which can be time-consuming, laboratory-based, often require trained personnel and specialized equipment. In this review, we discuss the various Ebola detection techniques currently in use, and highlight the potential future directions pertinent to the development and adoption of novel point-of-care diagnostic tools. Finally, a case is made for the need to develop novel microfluidic technologies and versatile rapid detection platforms for early detection of EVD.

Advances in Candida detection platforms for clinical and point-of-care applications.

Invasive candidiasis remains one of the most serious community and healthcare-acquired infections worldwide. Conventional Candida detection methods based on blood and plate culture are time-consuming and require at least 2-4 days to identify various Candida species. Despite considerable advances for candidiasis detection, the development of simple, compact and portable point-of-care diagnostics for rapid and precise testing that automatically performs cell lysis, nucleic acid extraction, purification and detection still remains a challenge. Here, we systematically review most prominent conventional and nonconventional techniques for the detection of various Candida species, including Candida staining, blood culture, serological testing and nucleic acid-based analysis. We also discuss the most advanced lab on a chip devices for candida detection.

Emerging Insights on Brazilian Pepper Tree (Schinus terebinthifolius) Invasion: The Potential Role of Soil Microorganisms.

Invasive plant species constitute a major ecological and economic problem worldwide, often distorting trophic levels and ecosystem balance. Numerous studies implicate factors ranging from environmental plasticity, competition for nutrient and space, and allelopathy in the success of invasive species in general. The Brazilian Pepper tree (BP) was introduced to the United States in the 1800s and has since become a category one invasive plant in Florida. It has aggressively spread to about 3000 km(2) of terrestrial surface, fueled in part by the prevalence of the hybrid genotypes and environmental perturbations. It displays some of the well-established invasive mechanisms but there is a serious dearth of knowledge on the plant-microbe-soil interactions and whether the rhizobiome plays any roles in the displacement of native flora and the range expansion of BP. Several control measures, including chemical, mechanical, and biological antagonism have been used with limited success while restoration of natives in soils from which BP was removed has proved problematic partly due to a poorly understood phenomenon described as the "BP legacy effect." Emerging evidence suggests that allelopathy, selective recruitment of beneficial soil microbes, disruption of microbial community structure and alteration of nutrient cycling, exhibited by many other invasive plant species may also be involved in the case of BP. This brief review discusses the well-established BP invasion mechanisms and highlights the current understanding of the molecular, below-ground processes. It also points out the gaps in studies on the potential role of microbial interactions in the success of BP invasion. These hitherto poorly studied mechanisms could further explain the aggressive spread of BP and could potentially contribute significantly to effective control measures and enable appropriate strategies for restoring native plants. The review advocates for the use of cutting-edge techniques in advancing the plant microbiome science. Ultimately, comparing metagenomic analyses of the rhizobiome of invasive plants grown in native and non-native soils could lead to a better understanding of the microbial determinants of biotic resistance, potentially empowering environmental managers with some predictive power of future trends of plant invasion.

High numbers of Staphylococcus aureus at three bathing beaches in South Florida.

While the value of Staphylococcus aureus as an indicator for non-enteric diseases is unclear, understanding its prevalence in recreational beaches would prove useful, given its pathogenic potential. Staphylococcus aureus levels were evaluated in sand and seawater at three beaches during one year. To elucidate possible S. aureus sources or colonization trends, distribution in sand was analyzed at Hollywood Beach. Staphylococcus aureus levels fluctuated throughout the study with highest average densities detected in dry sand (3.46 × 105 CFU/g, Hobie Beach), particularly at beaches with high human density. Patchy distribution marked hotspots of human use and/or possible bacterial re-growth. Data from a brief epidemiological survey indicated a very slight association between beach usage and skin conditions; suggesting high S. aureus levels in sand may not necessarily constitute major health risks. Because the possibility of disease transmission exists, particularly to children and immuno-compromised beach-goers, periodic surveying of highly frequented beaches seems warranted.

Co-cultures of Pseudomonas aeruginosa and Roseobacter denitrificans reveal shifts in gene expression levels compared to solo cultures.

Consistent biosynthesis of desired secondary metabolites (SMs) from pure microbial cultures is often unreliable. In a proof-of-principle study to induce SM gene expression and production, we describe mixed "co-culturing" conditions and monitoring of messages via quantitative real-time PCR (qPCR). Gene expression of model bacterial strains (Pseudomonas aeruginosa PAO1 and Roseobacter denitrificans Och114) was analyzed in pure solo and mixed cocultures to infer the effects of interspecies interactions on gene expression in vitro, Two P. aeruginosa genes (PhzH coding for portions of the phenazine antibiotic pathway leading to pyocyanin (PCN) and the RhdA gene for thiosulfate: cyanide sulfurtransferase (Rhodanese)) and two R. denitrificans genes (BetaLact for metallo-beta-lactamase and the DMSP gene for dimethylpropiothetin dethiomethylase) were assessed for differential expression. Results showed that R. denitrificans DMSP and BetaLact gene expression became elevated in a mixed culture. In contrast, P. aeruginosa co-cultures with R. denitrificans or a third species did not increase target gene expression above control levels. This paper provides insight for better control of target SM gene expression in vitro and bypass complex genetic engineering manipulations.

Prevalence and distribution of fecal indicator organisms in South Florida beach sand and preliminary assessment of health effects associated with beach sand exposure.

Fecal indicator levels in nearshore waters of South Florida are routinely monitored to assess microbial contamination at recreational beaches. However, samples of sand from the surf zone and upper beach are not monitored which is surprising since sand may accumulate and harbor fecal-derived organisms. This study examined the prevalence of fecal indicator organisms in tidally-affected beach sand and in upper beach sand and compared these counts to levels in the water. Since indicator organisms were statistically elevated in sand relative to water, the study also considered the potential health risks associated with beach use and exposure to sand. Fecal coliforms, Escherichia coli, enterococci, somatic coliphages, and F(+)-specific coliphages were enumerated from sand and water at three South Florida beaches (Ft. Lauderdale Beach, Hollywood Beach, and Hobie Beach) over a 2-year period. Bacteria were consistently more concentrated in 100g samples of beach sand (2-23 fold in wet sand and 30-460 fold in dry sand) compared to 100ml samples of water. Somatic coliphages were commonly recovered from both sand and water while F(+)-specific coliphages were less commonly detected. Seeding experiments revealed that a single specimen of gull feces significantly influenced enterococci levels in some 3.1m(2) of beach sand. Examination of beach sand on a micro-spatial scale demonstrated that the variation in enterococci density over short distances was considerable. Results of multiple linear regression analysis showed that the physical and chemical parameters monitored in this study could only minimally account for the variation observed in indicator densities. A pilot epidemiological study was conducted to examine whether the length of exposure to beach water and sand could be correlated with health risk. Logistic regression analysis results provided preliminary evidence that time spent in the wet sand and time spent in the water were associated with a dose-dependent increase in gastrointestinal illness.

Use of peptide nucleic acid probes for rapid detection and enumeration of viable bacteria in recreational waters and beach sand.

Environmental monitoring and public health risk assessments require methods that are rapid and quantitative with defined sensitivity and specificity thresholds. Although several molecular techniques have been developed to rapidly detect bacteria in complex matrices, the challenge to simultaneously detect and enumerate only viable cells remains a limiting factor to their routine application. This chapter describes the use of peroxidase-labeled peptide nucleic acid (PNA) probes to simultaneously detect and count live Staphylococcus aureus, a human pathogen in sea water and beach sand. Mixed bacteria from the environmental sample were immobilized on polyvinylidene difluoride membrane filters and allowed to form microcolonies during a 5-h incubation on Tryptic soy agar plates. PNA probes targeting species-specific regions of the 16S rRNA sequences of S. aureus were then used to hybridize the target bacteria in situ. Probes were detected by capturing chemiluminiscence on instant (e.g., Polaroid) films. Each viable cell (i.e., rRNA producing) is detected as a light spot from its microcolony on the film after scanning the image into a computer. This rapid in situ hybridization technique is simple and highly sensitive and could be developed into portable kits for monitoring pathogens and indicators in the environment.

The application of peptide nucleic acid probes for rapid detection and enumeration of eubacteria, Staphylococcus aureus and Pseudomonas aeruginosa in recreational beaches of S. Florida.

A novel chemiluminescent in situ hybridization technique using peptide nucleic acids (PNA) was adapted for the detection of bacteria in beach sand and recreational waters in South Florida. The simultaneous detection and enumeration of eubacteria and the novel indicators, Staphylococcus aureus and Pseudomonas aeruginosa, was achieved within 6-8 h of processing. Following 5 h of incubation on TSA, soybean peroxidase-labeled peptide nucleic acid probes (Boston Probes, Boston, MA) targeting species-specific 16S rRNA sequences of P. aeruginosa and S. aureus were used to hybridize microcolonies of the target species in-situ. In addition, a universal probe for 16S rRNA sequences was used to target the eubacteria. Probes were detected after a light generating reaction with a chemiluminescent substrate and their presence recorded on Polaroid film. The probes showed limited cross-reactivity with mixed indigenous bacteria extracted from seawater and sand by shaking with phosphate-buffered saline (PBS). Specificity and cross-reactivity was tested on the reference bacterial genera Pseudomonas, Staphylococcus, Vibrio, Shigella, Salmonella, Acinetobacter, Enterobacter, Escherichia and Citrobacter. These tests confirmed that the probes were specific for the microorganisms of interest and were unaffected by high salt levels. The results of the PNA chemiluminescent in situ hybridization were compared with traditional plate count methods (PCM) for total 'freshwater' eubacteria, S. aureus and P. aeruginosa. Counts of eubacteria and S. aureus were comparable with numbers obtained from traditional plate counts but levels of P. aeruginosa were higher with PNA than with PCM. It is possible that PNA is more sensitive than PCM because it can detect microcolonies on the agar surface that never fully develop with the plate count method. We conclude that the in situ hybridization technique used here represents an important potential tool for the rapid monitoring of novel indicator organisms in beaches and recreational waters.

An assessment of the in vitro antimicrobial effects of two antiepileptic drugs--sodium valproate and phenytoin.

An incidental observation led to the evaluation of the antimicrobial properties of valproate and phenytoin. In vitro inhibition of Mycobacterium smegmatis, Candida albicans, and other standard test organisms by these two antiepileptic drugs was assessed using the broth microdilution procedure. Fluorescence microscopy (Viability Stains, Molecular Probes Inc.) and cultural techniques were employed to distinguish between microbicidal and microbistatic effects. Phenytoin showed no inhibitory activity against the microbes tested. Sodium valproate, on the other hand, was selectively potent against the yeast strains in a dose-dependent manner (MIC = 10-20 microg ml(-1)). In vitro activity against Mycobacterium smegmatis (70% growth inhibition by 81 microg ml(-1)) was moderate to low while Staphylococcus aureus. Pseudomonas aeruginosa and Escherichia coli were not significantly affected. Viability data from fluorescent microscopy and plate cultures correlated well with absorbance (A620nm) growth index, and showed that valproate was microbicidal against susceptible organisms. The mode of action may include blockage of calcium channels and perturbation of membrane potential. This report opens up yet another opportunity for further enquiry into the fundamental mechanisms of drug action and microbial resistance.

Antibiotic resistance in soil and water environments.

Seven locations were screened for antibiotic-resistant bacteria using a modified agar dilution technique. Isolates resistant to high levels of antibiotics were screened for r plasmids. Low-level resistance (25 micro g x ml(-1)) was widespread for ampicillin, penicillin, tetracycline, vancomycin and streptomycin but not for kanamycin. Resistant populations dropped sharply at high antibiotic levels, suggesting that intrinsic non-emergent mechanisms were responsible for the multiple drug resistance exhibited at low doses. Dairy farm manure contained significantly (P < 0.01) more (%) resistant bacteria than the other sites. Bacteria isolated from a dairy water canal, a lake by a hospital and a residential garden (fertilized by farm manure) displayed resistance frequencies of 77, 75 and 70%, respectively. Incidence of tetracycline resistance was most prevalent at 47-89% of total bacteria. Out of 200 representative isolates analyzed, Pseudomonas, Enterococcus-like bacteria, Enterobacter and Burkholderia species constituted the dominant reservoirs of resistance at high drug levels (50-170 micro g x ml(-1)). Plasmids were detected in only 29% (58) of these bacteria with tetracycline resistance accounting for 65% of the plasmid pool. Overall, resistance trends correlated to the abundance and type of bacterial species present in the habitat. Environmental reservoirs of resistance include opportunistic pathogens and constitute some public health concern.