Diversity and community structure of anaerobic gut fungi in the rumen of wild and domesticated herbivores.

The rumen houses a diverse community that plays a major role in the digestion process in ruminants. Anaerobic gut fungi (AGF) are key contributors to plant digestion in the rumen. Here, we present a global amplicon-based survey of the rumen AGF mycobiome by examining 206 samples from 15 animal species, 15 countries, and 6 continents. The rumen AGF mycobiome was highly diverse, with 81 out of 88 currently recognized AGF genera or candidate genera identified. However, only six genera (Neocallimastix, Orpinomyces, Caecomyces, Cyllamyces, NY9, and Piromyces) were present at >4% relative abundance. AGF diversity was higher in members of the families Antilocapridae and Cervidae compared to Bovidae. Community structure analysis identified a pattern of phylosymbiosis, where host family (10% of total variance) and species (13.5%) partially explained the rumen mycobiome composition. As well, diet composition (9%-19%), domestication (11.14%), and biogeography (14.1%) also partially explained AGF community structure; although sampling limitation, geographic range restrictions, and direct association between different factors hindered accurate elucidation of the relative contribution of each factor. Pairwise comparison of rumen and fecal samples obtained from the same subject (n = 13) demonstrated greater diversity and inter-sample variability in rumen versus fecal samples. The genera Neocallimastix and Orpinomyces were present in higher abundance in rumen samples, while Cyllamyces and Caecomyces were enriched in fecal samples. Comparative analysis of global rumen and feces data sets revealed a similar pattern. Our results provide a global view of AGF community in the rumen and identify patterns of AGF variability between rumen and feces in herbivores Gastrointestinal (GI) tract.IMPORTANCERuminants are highly successful and economically important mammalian suborder. Ruminants are herbivores that digest plant material with the aid of microorganisms residing in their GI tract. In ruminants, the rumen compartment represents the most important location where microbially mediated plant digestion occurs, and is known to house a bewildering array of microbial diversity. An important component of the rumen microbiome is the anaerobic gut fungi (AGF), members of the phylum Neocallimastigomycota. So far, studies examining AGF diversity have mostly employed fecal samples, and little is currently known regarding the identity of AGF residing in the rumen compartment, factors that impact the observed patterns of diversity and community structure of AGF in the rumen, and how AGF communities in the rumen compare to AGF communities in feces. Here, we examined the rumen AGF diversity using an amplicon-based survey targeting a wide range of wild and domesticated ruminants (n = 206, 15 different animal species) obtained from 15 different countries. Our results demonstrate that while highly diverse, no new AGF genera were identified in the rumen mycobiome samples examined. Our analysis also indicate that animal host phylogeny, diet, biogeography, and domestication status could play a role in shaping AGF community structure. Finally, we demonstrate that a greater level of diversity and higher inter-sample variability was observed in rumen compared to fecal samples, with two genera (Neocallimastix and Orpinomyces) present in higher abundance in rumen samples, and two others (Cyllamyces and Caecomyces) enriched in fecal samples. Our results provide a global view of the identity, diversity, and community structure of AGF in ruminants, elucidate factors impacting diversity and community structure of the rumen mycobiome, and identify patterns of AGF community variability between the rumen and feces in the herbivorous GI tract.

Integrating Microbiome Analysis, Metabolomics, Bioinformatics, and Histopathology to Elucidate the Protective Effects of Pomegranate Juice against Benzo-alpha-pyrene-Induced Colon Pathologies.

Polycyclic aromatic hydrocarbons, e.g., benzo[a]pyrene (BaP), are common dietary pollutants with potential carcinogenic activity, while polyphenols are potential chemopreventive antioxidants. Although several health benefits are attributed to polyphenol-rich pomegranate, little is known about its interaction with BaP. This study integrates histochemical, microbiomic, and metabolomic approaches to investigate the protective effects of pomegranate juice from BaP-induced pathologies. To this end, 48 Sprague-Dawley rats received, for four weeks, either pomegranate, BaP, both, or neither (n = 12 rats per group). Whereas histochemical examination of the colon indicated tissue damage marked by mucin depletion in BaP-fed animals, which was partially restored by administration of pomegranate juice, the fecal microbiome and metabolome retained their resilience, except for key changes related to pomegranate and BaP biotransformation. Meanwhile, dramatic microbiome restructuring and metabolome shift were observed as a consequence of the elapsed time (age factor). Additionally, the analysis allowed a thorough examination of fecal microbiome-metabolome associations, which delineated six microbiome clusters (marked by a differential abundance of Lactobacillaceae and Prevotellaceae, Rumincococcaceae, and Erysipelotrichaceae) and two major metabolome clusters (a sugar- and amino-acids-dominated metabotype vs. a cluster of fatty acids and hydrocarbons), with sugar alcohols maintaining a unique signature. In conclusion, using paired comparisons to minimize inter-individual animal variations allowed the dissection of temporal vs. treatment-derived variations. Microbiome-metabolome association clusters may be further exploited for metabotype prediction and gut-health biomarker discovery.

Immunological characterization of the chemically prepared ghosts of Salmonella Typhimurium as a vaccine candidate.

BACKGROUND: Bacterial ghosts are the evacuated bacterial cellular membranes from most of the genetic and protein contents which preserved their surface characters. Recently, bacterial ghosts exploited for different biomedical applications, for instance, vaccination. The purpose of this study is to measure the immunogenic protective response of bacterial ghosts of Salmonella Typhimurium in animals and to allow future testing this response in humans. The immunologic response was qualitatively, quantitatively, and functionally measured. We have measured the humoral and cellular immune responses, such as immunoglobulins elevation (IgG), increased granulocytes, serum antibacterial activity, clearance of virulence in feces and liver, and the survival rate. RESULTS: The bacterial ghosts' vaccine was able to protect 100% of subcutaneously vaccinated rats and 75% of adjuvant subcutaneously vaccinated rats. The lowest survival rate was in the orally vaccinated group (25%). The maximum level of serum IgG titers, as well as serum and feces bactericidal activity (100% eradication), was exhibited in the subcutaneously vaccinated group with adjuvant vaccines followed by the subcutaneously vaccinated one. Additionally, the highest granulocytes' number was observed in the adjuvant vaccine subcutaneously immunized group. The bacterial load in liver homogenate was eliminated in the subcutaneously vaccinated rats after the virulence challenge. CONCLUSIONS: The bacterial ghosts of Salmonella enterica serovar Typhimurium that prepared by Tween 80 Protocol showed an effective vaccine candidate that protected animals, eliminated the virulence in feces and liver. These findings show that chemically induced bacterial ghosts of Salmonella Typhimurium can be a promising vaccine.

Chlorhexidine leads to the evolution of antibiotic-resistant Pseudomonas aeruginosa.

Antimicrobial resistance is a major public-health concern. We evaluate chlorhexidine role in selection of resistant Pseudomonas aeruginosa mutants and their antibiotic cross-resistance. Mutation frequency and mutation rate after short-term exposure to sub-inhibitory concentrations of chlorhexidine were compared to those after spontaneous chlorhexidine-exposure, in P. aeruginosa PAO1 strain. Chlorhexidine-resistant mutants were generated, either by serial passage in increasing chlorhexidine concentrations or by single exposure to lethal chlorhexidine concentration. The generated mutants were tested for cross-resistance to different antibiotics, by determination of minimum inhibitory concentrations (MIC). The accompanied phenotypic changes in membrane permeability, outer membrane proteins (OMP), and efflux function were evaluated. The effect of exposure to chlorhexidine on MexAB-OprM, MexEF-oprN, and MexXY efflux pumps expression was investigated. No significant change was recorded between the mutation frequencies and mutation rates after short-term exposure to sub-inhibitory concentrations of chlorhexidine and after spontaneous chlorhexidine-exposure, in P. aeruginosa PAO1 strain. Twelve stable mutants, with ≥ eight-fold increase in chlorhexidine MIC, were generated. Several mutants showed increase in the MIC of colistin, cefepime, ceftazidime, meropenem, ciprofloxacin, and amikacin; seven mutants expressed meropenem cross-resistance. This was accompanied by decreased outer membrane permeability and changes in OMP. Using efflux pump inhibitor, chlorhexidine resistance was reverted in most isolates. Exposure to sub-inhibitory concentration of chlorhexidine induced the expression of MexXY efflux pump. Some resistant mutants had overexpressed MexXY efflux pump. Chlorhexidine can select P. aeruginosa strains with antibiotic cross-resistance. This necessitates implementing special protocols for chlorhexidine use and re-evaluation of its benefit versus risk in personal-care products.

Spatiotemporal Analysis of the Water and Sediment Nile Microbial Community Along an Urban Metropolis.

Assessing microbial identity, diversity, and community structure could be a valuable tool for monitoring the impact of xenobiotics and anthropogenic inputs in rivers, especially in urban and industrial settings. Here, we characterize the Nile River microbial community in water and sediments in summer and winter at five locations that span its natural flow through the Cairo metropolis. 16S rRNA gene datasets were analyzed to identify the role played by sample type (sediment versus water), season, and location in shaping the community, as well as to predict functional potential of the Nile River microbiome. Microbial communities were mostly influenced by sampling type (sediments versus water), while seasonal effects were only observed in water samples. Spatial differences did not represent a significant factor in shaping the community in either summer or winter seasons. Proteobacteria was the most abundant phylum in both water and sediment samples, with the order Betaproteobacteriales being the abundant one. Chloroflexi and Bacteroidetes were also prevalent in sediment samples, while Cyanobacteria and Actinobacteria were abundant in water samples. The linear discriminative analysis effect size (LEfSe) identified the cyanobacterial genus Cyanobium PCC-6307 as the main variable between summer and winter water. Sequences representing human and animal potential pathogens, as well as toxin-producing Cyanobacteria, were identified in low abundance within the Nile microbiome. Functionally predicted metabolic pathways predicted the presence of antibiotic biosynthesis, as well as aerobic xenobiotic degradation pathways in the river microbiome.

Salmonella-innovative targeting carrier: Loading with doxorubicin for cancer treatment.

Cell- based targeted delivery is recently gain attention as a promising platform for delivery of anticancer drug in selective and efficient manner. As a new biotechnology platform, bacterial ghosts (BGs) have novel biomedical application as targeted drug delivery system (TDDS). In the current work, Salmonellas' BGs was utilized for the first time as hepatocellular cancer (HCC) in-vitro targeted delivery system. Successful BGs loading and accurate analysis of doxorubicin (DOX) were necessary steps for testing the applicability of DOX loaded BGs in targeting the liver cancer cells. Loading capacity was maximized to reach 27.5 µg/mg (27.5% encapsulation efficiency), by incubation of 10 mg BGs with 1 mg DOX at pH 9 in constant temperature (25 °C) for 10 min. In-vitro release study of DOX loaded BGs showed a sustained release (182 h) obeying Higuchi sustained kinetic release model. The death rate (tested by MTT assay) of HepG2 reached to 64.5% by using of 4 µg/ml, while it was about 51% using the same concentration of the free DOX (P value < 0.0001 One-way ANOVA analysis). The proliferative inhibitory concentration (IC50) of the DOX combined formula was 1.328 µg/ml that was about one third of the IC50 of the free DOX (3.374 µg/ml). Apoptosis analysis (tested by flow-cytometry) showed more accumulation in early apoptosis (8.3%) and late apoptosis/necrosis (91%) by applying 1 µg/ml BGs combined DOX, while 1 µg/ml free DOX showed 33.4% of cells in early apoptosis and 39.3% in late apoptosis/necrosis, (P value˃ 0.05: one-way ANOVA). In conclusion, DOX loaded Salmonellas' BGs are successfully prepared and tested in vivo with promising potential as hepatocellular cancer (HCC) targeted delivery system.

A novel protocol for bacterial ghosts' preparation using tween 80.

Bacterial ghosts (BGs) can be prepared by both genetic and chemical means. Genetic method include using lysis gene E. Chemical method include incubation with numerous agents for a short time at their minimum inhibitory or minimum growth concentrations (MIC or MGC). The aim of this study is to prepare the BGs with a new protocol via exposing the bacterial cells to tween 80 for an extended period of time followed by sudden reduction of the surrounding pH. Salmonella enterica serovar typhimurium ATCC 13311 was used for this purpose. The cells were incubated in 7% v/v tween 80 solution in Muller-Hinton broth for 24 h at 37 °C then pH was decreased to 3.6 by adding lactic acid for one hour. The bacterial pellets were separated by high speed centrifugation, and then washed three times by half normal saline solution. High quality BGs were visualized by scanning electron microscopy (SEM) revealing punctured cells with intact outer shells and at least one intramembranous tunnel. The absence of vital cells was confirmed by subculturing. The release of respective amounts of proteins and DNA is another evidence of ghost's production. In addition, the integrity of cells was proved by visualization of Gram-stained cells using light microscopy. In conclusion, this new protocol is simple, economic and feasible for BGs preparation.

Initial bridges between two ribosomal subunits are formed within 9.4 milliseconds, as studied by time-resolved cryo-EM.

Association of the two ribosomal subunits during the process of translation initiation is a crucial step of protein synthesis. The two subunits (30S and 50S) of the bacterial 70S ribosome are held together by 12 dynamic bridges involving RNA-RNA, RNA-protein, and protein-protein interactions. The process of bridge formation, such as whether all these bridges are formed simultaneously or in a sequential order, is poorly understood. To understand such processes, we have developed and implemented a class of microfluidic devices that mix two components to completion within 0.4 ms and spray the mixture in the form of microdroplets onto an electron microscopy grid, yielding a minimum reaction time of 9.4 ms before cryofixation. Using these devices, we have obtained cryo-EM data corresponding to reaction times of 9.4 and 43 ms and have determined 3D structures of ribosomal subunit association intermediates. Molecular analyses of the cryo-EM maps reveal that eight intersubunit bridges (bridges B1a, B1b, B2a, B2b, B3, B7a, B7b, and B8) form within 9.4 ms, whereas the remaining four bridges (bridges B2c, B4, B5, and B6) take longer than 43 ms to form, suggesting that bridges are formed in a stepwise fashion. Our approach can be used to characterize sequences of various dynamic functional events on complex macromolecular assemblies such as ribosomes.

The secretome of Acinetobacter baumannii ATCC 17978 type II secretion system reveals a novel plasmid encoded phospholipase that could be implicated in lung colonization.

Acinetobacter baumannii infections are compounded with a striking lack of treatment options. In many Gram-negative bacteria, secreted proteins play an important early role in avoiding host defences. Typically, these proteins are targeted to the external environment or into host cells using dedicated transport systems. Despite the fact that medically relevant species of Acinetobacter possess a type II secretion system (T2SS), only recently, its significance as an important pathway for delivering virulence factors has gained attention. Using in silico analysis to characterize the genetic determinants of the T2SS, which are found clustered in other organisms, in Acinetobacter species, they appear to have a unique genetic organization and are distributed throughout the genome. When compared to other T2SS orthologs, individual components of the T2SS apparatus showed the highest similarity to those of Pseudomonas aeruginosa. A mutant of Acinetobacter baumannii strain ATCC 17978 lacking the secretin component of the T2SS (ΔgspD), together with a trans-complemented mutant, were tested in a series of in vitro and in vivo assays to determine the role of T2SS in pathogenicity. The ΔgspD mutant displayed decreased lipolytic activity, associated with attenuated colonization ability in a murine pneumonia model. These phenotypes are linked to LipAN, a novel plasmid-encoded phospholipase, identified through mass spectroscopy as a T2SS substrate. Recombinant LipAN showed specific phospholipase activity in vitro. Proteomics on the T2-dependent secretome of ATCC 17978 strain revealed its potential dedication to the secretion of a number of lipolytic enzymes, among others which could contribute to its virulence. This study highlights the role of T2SS as an active contributor to the virulence of A. baumannii potentially through secretion of a newly identified phospholipase.

Acinetobacter baumannii universal stress protein A plays a pivotal role in stress response and is essential for pneumonia and sepsis pathogenesis.

Acinetobacter baumannii is one of the most significant threats to global public health. This threat is compounded by the fact that A. baumannii is rapidly becoming resistant to all relevant antimicrobials. Identifying key microbial factors through which A. baumannii resists hostile host environment is paramount to the development of novel antimicrobials targeting infections caused by this emerging pathogen. An attractive target could be a molecule that plays a role in the pathogenesis and stress response of A. baumannii. Accordingly, the universal stress protein A (UspA) was chosen to be fully investigated in this study. A platform of A. baumannii constructs, expressing various levels of the uspA gene ranging from zero to thirteen folds of wild-type level, and a recombinant E. coli strain, were employed to investigate the role of UspA in vitro stress and in vivo pathogenesis. The UspA protein plays a significant role in protecting A. baumannii from H(2)O(2), low pH, and the respiratory toxin 2,4-DNP. A. baumannii UspA protein plays an essential role in two of the deadliest types of infection caused by A. baumannii; pneumonia and sepsis. This distinguishes A. baumannii UspA from its closely related homolog, the Staphylococcus aureus Usp2, as well as from the less similar Burkholderia glumae Usps. Heterologous and overexpression experiments suggest that UspA mediates its role via an indirect mechanism. Our study highlights the role of UspA as an important contributor to the A. baumannii stress and virulence machineries, and polishes it as a plausible target for new therapeutics.

Structure-Activity Relationships of Antimicrobial Gallic Acid Derivatives from Pomegranate and Acacia Fruit Extracts against Potato Bacterial Wilt Pathogen.

Bacterial wilts of potato, tomato, pepper, and or eggplant caused by Ralstonia solanacearum are among the most serious plant diseases worldwide. In this study, the issue of developing bactericidal agents from natural sources against R. solanacearum derived from plant extracts was addressed. Extracts prepared from 25 plant species with antiseptic relevance in Egyptian folk medicine were screened for their antimicrobial properties against the potato pathogen R. solancearum by using the disc-zone inhibition assay and microtitre plate dilution method. Plants exhibiting notable antimicrobial activities against the tested pathogen include extracts from Acacia arabica and Punica granatum. Bioactivity-guided fractionation of A. arabica and P. granatum resulted in the isolation of bioactive compounds 3,5-dihydroxy-4-methoxybenzoic acid and gallic acid, in addition to epicatechin. All isolates displayed significant antimicrobial activities against R. solanacearum (MIC values 0.5-9 mg/ml), with 3,5-dihydroxy-4-methoxybenzoic acid being the most effective one with a MIC value of 0.47 mg/ml. We further performed a structure-activity relationship (SAR) study for the inhibition of R. solanacearum growth by ten natural, structurally related benzoic acids.

Gas-Assisted Annular Microsprayer for Sample Preparation for Time-Resolved Cryo-Electron Microscopy.

Time-resolved cryo electron microscopy (TRCEM) has emerged as a powerful technique for transient structural characterization of isolated biomacromolecular complexes in their native state within the time scale of seconds to milliseconds. For TRCEM sample preparation, microfluidic device [9] has been demonstrated to be a promising approach to facilitate TRCEM biological sample preparation. It is capable of achieving rapidly aqueous sample mixing, controlled reaction incubation, and sample deposition on electron microscopy (EM) grids for rapid freezing. One of the critical challenges is to transfer samples to cryo-EM grids from the microfluidic device. By using microspraying method, the generated droplet size needs to be controlled to facilitate the thin ice film formation on the grid surface for efficient data collection, while not too thin to be dried out before freezing, i.e., optimized mean droplet size needs to be achieved. In this work, we developed a novel monolithic three dimensional (3D) annular gas-assisted microfluidic sprayer using 3D MEMS (MicroElectroMechanical System) fabrication techniques. The microsprayer demonstrated dense and consistent microsprays with average droplet size between 6-9 µm, which fulfilled the above droplet size requirement for TRCEM sample preparation. With droplet density of around 12-18 per grid window (window size is 58×58 µm), and the data collectible thin ice region of >50% total wetted area, we collected ~800-1000 high quality CCD micrographs in a 6-8 hour period of continuous effort. This level of output is comparable to what were routinely achieved using cryo-grids prepared by conventional blotting and manual data collection. In this case, weeks of data collection process with the previous device [9] has shortened to a day or two. And hundreds of microliter of valuable sample consumption can be reduced to only a small fraction.

Insertion domain within mammalian mitochondrial translation initiation factor 2 serves the role of eubacterial initiation factor 1.

Mitochondria have their own translational machineries for the synthesis of thirteen polypeptide chains that are components of the complexes that participate in the process of oxidative phosphorylation (or ATP generation). Translation initiation in mammalian mitochondria requires two initiation factors, IF2(mt) and IF3(mt), instead of the three that are present in eubacteria. The mammalian IF2(mt) possesses a unique 37 amino acid insertion domain, which is known to be important for the formation of the translation initiation complex. We have obtained a three-dimensional cryoelectron microscopic map of the mammalian IF2(mt) in complex with initiator fMet-tRNA(iMet) and the eubacterial ribosome. We find that the 37 amino acid insertion domain interacts with the same binding site on the ribosome that would be occupied by the eubacterial initiation factor IF1, which is absent in mitochondria. Our finding suggests that the insertion domain of IF2(mt) mimics the function of eubacterial IF1, by blocking the ribosomal aminoacyl-tRNA binding site (A site) at the initiation step.

Detection, characterization, and molecular typing of human Mycoplasma spp. from major hospitals in Cairo, Egypt.

Mycoplasmas are fastidious slow growing organisms lacking a cell wall and mostly isolated from the mucosal surfaces of the respiratory and genitourinary tracts. There is a dearth of information regarding clinical Mycoplasma spp. isolates among Egyptian patients. A total of 170 samples were collected from patients and apparently healthy personnel in local public hospitals in Cairo, Egypt. Isolation of Mycoplasma spp. was carried out using appropriate culture media and further identification was carried out by biochemical tests followed by serotyping using specific antisera. Confirmation was done by PCR for detection of different Mycoplasma spp. using genus-specific primers targeting 16S ribosomal RNA gene. Characterization of the antibiotic resistance and sensitivity pattern against different antimicrobials was carried out using disc diffusion test. The results indicated the presence of six Mycoplasma spp. in 22.94% of the samples. Mycoplasmas were detected more frequently in throat swabs than sputum. Mycoplasma pneumoniae was highly sensitive to macrolides and quinolones but less sensitive to aminoglycosides and tetracyclines. Molecular techniques were found to be of more rapid, highly sensitive, able to detect nonviable organisms, and cost effective. These results shed light on difficulties of Mycoplasma detection and the superiority of molecular techniques over culture.

A molecular investigative approach to an outbreak of acute hemorrhagic conjunctivitis in Egypt, October 2010.

BACKGROUND: During October 2010, Egypt reported an outbreak of acute hemorrhagic conjunctivitis (AHC). A total of 1831 cases were reported from three governorates; 1703 cases in El Daqahliya, 92 cases in Port Said, and 36 in Damietta. The purpose of this study was to identify and characterize the causative agent associated with this outbreak. METHODS: The U.S. Naval Medical Research Unit No.3 (NAMRU-3) was contacted by the Egyptian Ministry of Health and Population to perform diagnostic laboratory testing on eighteen conjunctival swabs from patients with conjunctivitis from El Daqahliya Governorate. Conjunctival swabs were tested by molecular methods for human adenovirus (HAdV) and enteroviruses (EV). Virus isolation was performed; the isolated virus was further characterized by molecular typing and phylogenetic analysis. RESULTS: The majority of the samples (17/18) were positive for enterovirus and all were negative for HAdV. Molecular typing and sequencing of the isolated virus revealed the presence of coxsackievirus A24 variant. Phylogenetic analysis based on the VP1 and 3C regions demonstrated that the Egyptian viruses belonged to Genotype IV and are closely related to coxsackievirus A24 variant, reported in a similar outbreak in China in August 2010. CONCLUSIONS: This study strongly suggests that coxsackievirus A24 variant was associated with the acute hemorrhagic conjunctivitis outbreak reported in Egypt in October 2010. There is a possibility that the same strain of CV-A24v was implicated in the AHC outbreaks in both China and Egypt in 2010.

Advanced approaches for endotoxin detection and removal from snake antivenoms.

A comprehensive assessment of the literature on strategies for the detection and removing endotoxin from biotechnological preparations was conducted. This study highlighted the brief history of endotoxin. After that, a review of endotoxin's chemical and physical features, as well as its pathophysiological consequences when the body is exposed to LPS excessively or systemically, is presented. The procedures for determining endotoxin and the interaction of endotoxin with proteins are also discussed, considering both known approaches and cutting-edge technology in this sector. This review presented the endotoxin detection and removal approaches from antisera with an economical approach using several processes documented in the literature (e.g., adsorption, ultrafiltration, and chromatography). Different methods with relatively high protein recoveries are mentioned. This review concludes that heat activation at 70 °C-80 °C for 10 min and rehydration of the LAL reagent with endotoxin-specific buffer solution is the best technique to control the enhancement problem when testing polyvalent snake venom antiserum samples by the LAL method. The most efficient method for eliminating endotoxins has proven to be affinity resin-based chromatography.

Patterns and determinants of the global herbivorous mycobiome.

Despite their role in host nutrition, the anaerobic gut fungal (AGF) component of the herbivorous gut microbiome remains poorly characterized. Here, to examine global patterns and determinants of AGF diversity, we generate and analyze an amplicon dataset from 661 fecal samples from 34 mammalian species, 9 families, and 6 continents. We identify 56 novel genera, greatly expanding AGF diversity beyond current estimates (31 genera and candidate genera). Community structure analysis indicates that host phylogenetic affiliation, not domestication status and biogeography, shapes the community rather than. Fungal-host associations are stronger and more specific in hindgut fermenters than in foregut fermenters. Transcriptomics-enabled phylogenomic and molecular clock analyses of 52 strains from 14 genera indicate that most genera with preferences for hindgut hosts evolved earlier (44-58 Mya) than those with preferences for foregut hosts (22-32 Mya). Our results greatly expand the documented scope of AGF diversity and provide an ecologically and evolutionary-grounded model to explain the observed patterns of AGF diversity in extant animal hosts.

Viral etiologies of lower respiratory tract infections among Egyptian children under five years of age.

BACKGROUND: Lower respiratory tract infections (LRTI) are responsible for a considerable number of deaths among children, particularly in developing countries. In Egypt and the Middle East region, there is a lack of data regarding the viral causes of LRTI. In this study, we aimed to identify the relative prevalence of various respiratory viruses that contribute to LRTIs in young children. Although, nucleic acid-based methods have gained importance as a sensitive tool to determine the viral infections, their use is limited because of their prohibitive cost in low-income countries. Therefore, we applied three different laboratory methods, and presented the different virus prevalence patterns detected by each method. METHODS: We collected nasopharyngeal aspirate samples, demographic data and, clinical data from 450 children under five years of age who presented with LRTI at Abou El Reesh hospital in Cairo during a one-year period. To identify the viral causes of the LRTI we used direct fluorescence assay, real-time reverse-transcriptase polymerase chain reaction (rt-RT-PCR), and shell vial culture. We tested for eight major respiratory viruses. RESULTS: Two hundred sixty-nine patients (59.9%) had a viral infection, among which 10.8% had a co-infection with two or more viruses. By all three methods, respiratory syncytial virus (RSV) was the most predominant, and parainfluenza virus type 2 (HPIV-2), influenza B virus (FLUBV) were the least predominant. Other viral prevalence patterns differed according to the detection method used. The distribution of various viruses among different age groups and seasonal distribution of the viruses were also determined. CONCLUSIONS: RSV and human adenovirus were the most common respiratory viruses detected by rt-RT-PCR. Co-infections were found to be frequent among children and the vast majority of co-infections were detected by nucleic acid-based detection assays.

Phenotype-Genotype Characterization and Antibiotic-Resistance Correlations Among Colonizing and Infectious Methicillin-Resistant Staphylococcus aureus Recovered from Intensive Care Units.

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) presents a profound hazard to public health. MRSA colonizing skin, mucous membranes, and the anterior nares without clinical symptoms is termed "colonizing MRSA". Upon manifestation of clinical symptoms, it is termed "infectious MRSA". Here, we characterize and differentiate colonizing and infectious MRSA, and analyze the phenotypic-genotypic and antibiotic susceptibility correlations. METHODOLOGY: Clinical MRSA isolates were recovered from intensive care units (ICUs) of two major Egyptian hospitals and their biofilm formation ability was tested. Antibiograms against 16 antibiotics were determined, in addition to the minimum inhibitory concentrations (MICs) of vancomycin and linezolid. The entire collection was typed by enterobacterial repetitive intergenic consensus (ERIC)-PCR, as well as multi-locus sequence typing (MLST). Representative resistance and virulence genes were detected by PCR amplification. RESULTS: Forty-nine isolates were confirmed as MRSA, of which 30 isolates were infectious and 19 were colonizing. Versatile resistance patterns were observed in both groups of isolates. We report a higher tendency for biofilm-formation and borderline minimum inhibitory concentrations among infectious isolates. A Positive antibiotic correlation was observed between susceptibility to protein synthesis inhibitors and cell wall inhibitors. Positive correlations were observed between isolation site and rifampicin resistance: nasal samples were enriched in rifampicin-resistant isolates, while urine and blood samples were enriched in susceptible ones. Furthermore, biofilm formation ability was slightly associated with amikacin resistance, and an association between teicoplanin resistance and the presence of the Panton-Valentine leukocidin gene was the only significant phenotype-genotype correlation observed. Finally, ERIC typing and MLST had congruent results. CONCLUSION: Linezolid and vancomycin are still the most convenient choice for MRSA treatment. ERIC PCR and MLST show promising typing combination that could be easily used periodically for tracking the genotypic changes of MRSA, especially within the healthcare facilities. Several correlations were established between groups of antibiotics and the genotypes/phenotypes of the selected isolates.

Draft genome sequence of a prodigiosin-hyperproducing Serratia marcescens strain isolated from Cairo, Egypt.

Serratia marcescens is a Gram-negative bacterium with both environmental and host-associated strains. Pigmentation is reportedly inversely correlated with infection frequency, and prodigiosin is one of Serratia pigments that has medical and industrial applications. Here, we report the draft genome sequence of prodigiosin-hyperproducing Serratia marcescens strain N2, isolated from Cairo, Egypt. The sequence is assembled into 142 contigs, with a combined size of 5,570,793 bp. The assembled genome carries typical S. marcescens genes, with potential prodigiosin-biosynthesizing genes detected.