Biofilm and Helicobacter pylori:From environment to human host

Helicobacter pylori(H.pylori)is a Gram negative pathogen that selectively colonizes the human gastric epithelium.Over 50%of the world population is infected with H.pylori reaching up to 90%of infected individuals in developing countries.Nonetheless the increased impact upon public health care,its reservoir and the transmission pathway of the species has not been clearly established yet.Molecular studies allowed the detection of H.pylori in various aquatic environments,even forming biofilm in tap water distribution systems in several countries,suggesting a role of water as a possible reservoir of the pathogen.The persistence of human infection with H.pylori and the resistance of clinical isolates to commonly used antibiotics in eradication therapy have been related to the genetic variability of the species and its ability to develop biofilm,demonstrated both in vivo and in vitro experiments.Thus,during the last years,experimental work with this pathogen has been focused in the search for biofilm inhibitors and biofilm destabilizing agents.However,only two anti-H.pylori biofilm disrupting agents have been successfully used:Curcumin-a natural dye-and N-acetyl cysteine-a mucolytic agent used in respiratory diseases.The main goal of this review was to discuss the evidences available in the literature supporting the ability of H.pylori to form biofilm upon various surfaces in aquatic environments,both in vivo and in vitro.The results published and our own observations suggest that the ability of H.pylori to form biofilm may be important for surviving under stress conditions or in the spread of the infection among humans,mainly through natural water sources and water distribution systems.

Role of quorum sensing in bacterial infections

Quorum sensing(QS) is cell communication that is widely used by bacterial pathogens to coordinate the expression of several collective traits, including the production of multiple virulence factors, biofilm formation, and swarming motility once a population threshold is reached. Several lines of evidence indicate that QS enhances virulence of bacterial pathogens in animal models as well as in human infections; however, its relative importance for bacterial pathogenesis is still incomplete. In this review, we discuss the present evidence from in vitro and in vivo experiments in animal models, as well as from clinical studies, that link QS systems with human infections. We focus on two major QS bacterial models, the opportunistic Gram negative bacteria Pseudomonas aeruginosa and the Gram positive Staphylococcus aureus, which are also two of the main agents responsible of nosocomial and wound infections. In addition, QS communication systems in other bacterial, eukaryotic pathogens, and even immune and cancer cells are also reviewed, and finally, the new approaches proposed to combat bacterial infections by the attenuation of their QS communication systems and virulence are also discussed.

Current knowledge on the laboratory diagnosis of Clostridium difficile infection

Clostridium difficile (C. difficile) is a spore-forming, toxin-producing, gram-positive anaerobic bacterium that is the principal etiologic agent of antibiotic-associated diarrhea. Infection with C. difficile (CDI) is characterized by diarrhea in clinical syndromes that vary from selflimited to mild or severe. Since its initial recognition as the causative agent of pseudomembranous colitis, C. difficile has spread around the world. CDI is one of the most common healthcare-associated infections and a significant cause of morbidity and mortality among older adult hospitalized patients. Due to extensive antibiotic usage, the number of CDIs has increased. Diagnosis of CDI is often difficult and has a substantial impact on the management of patients with the disease, mainly with regards to antibiotic management. The diagnosis of CDI is primarily based on the clinical signs and symptoms and is only confirmed by laboratory testing. Despite the high burden of CDI and the increasing interest in the disease, episodes of CDI are often misdiagnosed. The reasons for misdiagnosis are the lack of clinical suspicion or the use of inappropriate tests. The proper diagnosis of CDI reduces transmission, prevents inadequate or unnecessary treatments, and assures best antibiotic treatment. We review the options for the laboratory diagnosis of CDI within the settings of the most accepted guidelines for CDI diagnosis, treatment, and prevention of CDI.

Management of Helicobacter pylori infection in Latin America: A Delphi technique-based consensus

AIM: To optimize diagnosis and treatment guidelines for this geographic region, a panel of gastroenterologists, epidemiologists, and basic scientists carried out a structured evaluation of available literature.METHODS: Relevant questions were distributed among the experts, who generated draft statements for consideration by the entire panel. A modified three-round Delphi technique method was used to reach consensus. Critical input was also obtained from representatives of the concerned medical community. The quality of the evidence and level of recommendation supporting each statement was graded according to United States Preventive Services Task Force criteria.RESULTS: A group of ten experts was established. The survey included 15 open-ended questions that were distributed among the experts, who assessed the articles associated with each question. The levels of agreement achieved by the panel were 50% in the first round, 73.3% in the second round and 100% in the third round. Main consensus recommendations included:(1) when available, urea breath and stool antigen test(HpSA) should be used for non-invasive diagnosis;(2) detect and eradicate Helicobacter pylori(H. pylori) in all gastroscopy patients to decrease risk of peptic ulcer disease, prevent o retard progression in patients with preneoplastic lesions, and to prevent recurrence in patients treated for gastric cancer;(3) further investigate implementation issues and health outcomes of H. pylorieradication for primary prevention of gastric cancer in high-risk populations;(4) prescribe standard 14-d triple therapy or sequential therapy for first-line treatment;(5) routinely assess eradication success post-treatment in clinical settings; and(6) select second- and third-line therapies according to antibiotic susceptibility testing.CONCLUSION: These achievable steps toward better region-specific management can be expected to improve clinical health outcomes.

Controllable Biosynthesis of Small Silver Nanoparticles Using Fungal Extract

The search for reliable and eco-friendly methods for the production of nanoscale materials is an important aspect of nanotechnology. Silver nanoparticles (AgNPs) are of special interest because of their antimicrobial properties, especially those of small size. In this work, AgNPs were produced under different conditions of temperature and pH using the extract from the fungus Neurospora crassa as reducing agent. Mainly quasi-spherical particles were obtained at all incubation conditions. However, optimum conditions to produce small sizes in the range of 2 - 9 nm were at 4&degC and pH 3, also particles of 2 - 22 nm were obtained at 25&degC with unmodified pH (6.5) and pH 10. Nevertheless, only particles synthesized at 25&degC and pH 10 maintained the same size range after storage of 10 months. In summary, optimal incubation conditions for the synthesis of silver nanoparticles of small size range are reported. This improves the storage time of particles without losing its original size and without going into aggregation or agglomeration.

Preliminary Evaluation of Biocontrol Agents against Maize Pathogens <i>Exserohilum turcicum</i>and <i>Puccinia sorghi</i>in Field Assays

Exserohilum turcicum and Puccinia sorghi cause foliar fungal diseases that affect maize crop in Argentina. These diseases, northern leaf blight and common rust respectively, are presented each year with different levels of severity affecting significantly the yield in susceptible hybrids. Disease control usually consists in the use of resistant cultivars and chemical control. Biological control as a preventive method is a viable alternative to evaluate. The aims of this study were to evaluate the natural incidence of both foliar diseases in maize after application of two antagonists, to determine the survival of the antagonists in the maize phyllosphere and to evaluate the effect of inoculation on grain yield at harvest. Plants treated with both biological control agents showed significant reductions in the incidence of both foliar diseases. In northern leaf blight the reduction was higher than 50% during 40 days in plants treated with Bacillus spp. Moreover, grain yield was significantly higher as compared to control treatments.

Revealing Toxin Signatures in Cyanobacteria: Report of Genes Involved in Cylindrospermopsin Synthesis from Saxitoxin-Producing <i>Cylindrospermopsis raciborskii</i>

Cyanotoxins are distinctive molecules in Cyanobacteria whose evolutionary origin, radiation and ecological role are still controversial. The cyanobacterium Cylindrospermopsis raciborskii is alternately capable of producing two types of potent toxins, cylindrospermopsin (CYN) or saxitoxin and analogues (SAX). It has been proposed that this species spread to all continents early in its evolutionary history and biogeographical differences in toxin production are found between populations. Most reports indicate that American strains are able to produce SAX but not CYN, while Australian strains are described to produce CYN but not SAX. Here we describe the presence of three genes belonging to the cylindrospermopsin cluster (cyr), cyrA, cyrB and cyrC, in two SAX producing South American C. raciborskii strains, MVCC14 and MVCC19, which due to their differences in morphology, growth preferences, SAX production and genetic context are defined as different ecotypes. No CYN production was detected in either strain (by ELISA) after growth under nitrogen replete or nitrogen-free nutrient conditions. Phylogenetic analyses of cyrA, cyrB and cyrC partial sequences from both strains showed high similarity (>99%) with CYN genes belonging to C. raciborskii strains from Australia and Germany and to Aphanizomenon strains. This is the first report of the presence of cyr genes in strains known to produce only SAX.

Effect of Domestic Wastewater as Co-Substrate on Biological Stain Wastewater Treatment Using Fungal/Bacterial Consortia in Pilot Plant and Greenhouse Reuse

In this study, a pilot wastewater treatment plant was used to evaluate the co-treatment of biological-staining residues and domestic wastewater under non-sterile conditions. A novel microbial consortia formed by Trametes versicolor, Trametes sp, Pleurotus ostreatus, Pseudomonas fluorescens, Pseudomonas azotoformans, Pseudomonas sp, Enterobacter xianfangensis and Bacillus subtillis was inoculated in an extended aeration type bio-reactor. The treatment units were operated during three consecutive cycles during a period of 147 h. After the last operating cycle, the concentrations of Chemical Oxygen Demand, Biochemical Oxygen Demand, Color Units, Total suspended solids, and the pH value were 1695 mg/L, 105 mg/L, 106 CU, 5), 1367 (CU), 566 mg/L (TSS) and 7.0 (pH) respectively. The reduction of pollutants load was related with the ratio of the two types of wastewater (3.5:0.5) combined to increase biodegradability, the concentration of fungi and bacteria used in the consortia (30 × 103 - 55 × 106 CUF/mL Total Fungi and 70 × 107 - 83 × 108 CFU/mL of Total Bacteria) and ligninolytic enzymes production, Laccase (13 - 96 U/L), MnP (9.8 - 39 U/L) and LiP (0.3 - 5.3 U/L). The post-treated effluent was used as irrigation water. Lolium perenne plants were watered during 60 days with post-treated effluent. The results of root weight showed that there are significant differences between the initial water and the effluent obtained after the operational cycles (p = 0.00470). The highest root weights (1 - 1.12 g) were found in plants irrigated with water obtained from the last treatment cycle.

First Characterization of <i>Escherichia coli</i>Strains Isolated from Wildlife Griffon Vulture (<i>Gyps fulvus</i>) in the Southeast of Spain

The aim of the present study was to characterize a collection of Escherichia coli strains isolated from asymptomatic griffon vulture (Gyps fulvus) during a reintroduction program in the southeast of Spain, in order to establish if griffon vulture could play a role in the spread of resistant or potentially pathogenic E. coli strains. For this purpose, 14 E. coli strains obtained from 10 griffon vulture were studied to establish their serotypes, phylogroups, virulencegene profiles and antimicrobial resistances. High heterogeneity was observed within the 14 strains isolated which belonged to three phylogroups (A, B1 and D), 8 serogroups (O2, O21, O29, O60, 073, O78, O103 and O141) and 13 different serotypes. Out of 34 genes screened, we have detected eight virulence genes that are typical of extraintestinal pathogenic E. coli (ExPEC) (fimH, fimAvMT78, iroN, iucD, cvaC, iss, traT and tsh);however, none of the studied strains showed the ExPEC status. The 14 strains were also analyzed for the production of extended-spectrum beta-lactamases (ESBLs) and for antimicrobial resistances. None of the 14 strains were ESBL-producing E. coli, but high resistance-prevalences to ampicillin and cotrimoxazole were detected. To our knowledge, this is the first characterization of E. coli strains isolated from griffon vulture and although they did not show high virulencegene scores, they showed cotrimoxazole resistance.

Biotechnological Alternatives for the Utilization of Dairy Industry Waste Products

The industrial obtainment and manufacturing of food cause high levels of pollution because of the generation of waste byproducts. Over the past few years, there has been a significantly increased interest in preserving, restoring and establishing an ecological balance during food production. Many investigators propose biotechnological solutions to the treatment of industrial wastes, especially waste from the dairy industry. The aim of this review is to present biotechnological approaches to the treatment and utilization of wastes from the dairy industry, specifically for milk whey and also discussed biotechnological methods to reduce environmental pollution and obtain chemical compounds with potential applications in the industry.

Accumulation of the betaine and ectoine in osmotic stress adaptation of biocontrol agents against <i>Fusarium verticillioides</i>in maize

The aims of this study was to examine the effect of the osmotic (NaCl) water stress on, growth and accumulation of endogenous compatible solutes of Bacillus amyloliquefaciens and Microbacterium oleovorans, biocontrol agents against Fusarium verticillioides. Triptic soya broth media were modified ionically to 0.99;0.98;0.97 and 0.96 aw with NaCl. The endogenous contents of the intracellular compatible solutes glycine-betaine and ectoine were quantified. Cells grown under ionic solute stress showed accumulation of significant amounts of both amino acids in all treatments. The growth rate of F. verticillioides was decreased significantly by interaction with B. amyloliquefaciens and M. oleovorans physiologically adapted at 0.96 aw. This study have demonstrated the ability to synthesize betaine and ectoine under high-osmolality conditions of B. amyloliquefaciens and M. oleovorans cells. Such ecophysiological manipulation, especially to water stress, may increase the potential for biological control of F. verticillioides at field under wider aw conditions.

Studies on Possible Modes of Action and Tolerance to Environmental Stress Conditions of Different Biocontrol Agents of Foliar Diseases in Maize

The present study evaluates the possible modes of action of antagonistic bacteria and their tolerance to UV radiation, temperature and osmotic stress. The partial 16S-23S rRNA gene sequencing of eight antagonistic bacteria had a high match with three bacterial genera: Curtobacterium, Pantoea and Bacillus. In this study, the three Bacillus isolates showed the most relevant production of enzymes, volatile organic compounds and antibiosis against Exserohilum turcicum. Respect to UV radiation and temperature Pantoea and Bacillus isolates were more tolerant;whereas the eight isolates were tolerant to osmotic stress in varying degree. The three Bacillus isolates have the greatest potential as biocontrol agents for foliar diseases in maize. The antagonistic action could be explained through different modes of action such as enzymes, volatile organic compounds and/or direct antibiosis by other secondary metabolites. Bacillus isolates tolerance to environmental stresses including UV radiation, temperature and osmotic stress is relevant for survival and persistence on the leaf surface. This work provides new information about the mode of action of antagonistic bacteria with proven efficacy against maize leaf pathogens. In addition, it provides information about the tolerance of antagonistic bacteria against different stress conditions. The data of the present study could contribute to the development of a successful foliar biofungicide.

Ultrasound Promoted Synthesis and Antimicrobial Evaluation of Novel Seven and Eight-Membered 1,3-Disubstituted Cyclic Amidinium Salts

Series of 1,3-dibenzyl-1H-4,5,6,7-tetrahydro-1,3-diazepinium and 1,4,5,6,7,8-hexahydro-1,3-diazocinium salts derivatives were efficiently synthesized in excellent yields by dehydrogenation of the corresponding N,N'-dibenzyl aminals employing N-bromosuccinimide (NBS) as dehydrogenating agent under ultrasound irradiation. The present methodology has proven to be simple, efficient and environmentally benign. All novel compounds were identified and characterized by 1H and 13C NMR spectra. The synthesized compounds were screened for their antimicrobial activities.

Nitrogen Source Affects Glycolipid Production and Lipid Accumulation in the Phytopathogen Fungus <i>Ustilago maydis</i>

When cultured in medium limited of nitrogen sources, the phytopathogen Ustilago maydis produces two amphipathic glycolipids: Ustilagic acid (UA) and Mannosylerythritol lipid (MEL), which in addition to the hydrophilic moiety, contain dior tri-hydroxylated C16 fatty acids (UA), or C8 and C16 saturated fatty acids (MEL). We compared the growth and morphology of cells in YPD and in minimum media containing glucose and nitrogen sources such as nitrate or urea and those deprived of nitrogen. Nitrogen-starved cells showed a dramatic accumulation of internal lipids identified as lipid droplets when stained with the hydrophobic probe BODIPY;these lipid droplets were enriched in unsaturated fatty acids. Fatty acids in YPD or medium containing nitrate as nitrogen source showed a combination of saturated/unsaturated lipids, but when urea was the nitrogen source, cells only contained saturated fatty acids. The glycolipid profiles produced in the presence or absence of nitrogen showed preferences towards the production of one kind of glycolipid: cells in media containing nitrate or urea produced different proportions of UA/MEL, but under nitrogen starvation cells contained only UA. The emulsification capacity of the glycolipids produced in media with or without nitrogen was similar (72% - 76%). HPLC of the glycolipids allowed the separation of fractions with different emulsifying characteristics. Our results indicate that U. maydis accumulates lipid droplets when deprived of nitrogen source and confirm that UA is not under nitrogen control, but rather that MEL and lipid droplets are produced and oppositely regulated by nitrogen.

Stage-Specific Changes on <i>Plasmodium yoelii yoelii </i>Following Treatment with <i>Hintonia latiflora</i>Stem Bark Extract and Phytochemical-Antioxidant Evaluation

Malaria endemic zones are mostly located on third world countries, where antimalarials are not easily found or patients cannot afford them, and in consequence, they must turn toward natural products or phytomedicines. In the present study, the effect of Hinotnia latiflora (Hl) methanolic stem bark extract (HlMeOHe) on the ultrastructure of the asexual intraerythrocytic stages of Plasmodium yoelii yoelii (Pyy) after a Peters’ four-day oral treatment was assessed by transmission electron microscopy (TEM), as well as the parasite development on blood smears, analyzed by light microscopy. Likewise, extract was subjected to qualitative tests adopting standard procedures for identification of phytoconstituents;its antioxidant activity was evaluated according to the method of Brand-Williams and by the radical cation decolorization assay. Results showed higher percentage of rings and lower percentage of trophozoites and schizonts in the treated animals, in comparison with those of the control groups, which demonstrated lower percentage of rings and trophozoites, and schizonts in higher number. Images of TEM showed in some treated parasites, mild parasite membranes, organelle swelling and ribosomal depletion. The phytochemical profile demonstrated that the extract contains alkaloids, tannis, steroids, terpenoids, flavonoids, phenolics and saponins. The obtained values of the half maximal inhibitory concentration (IC50) in μg/mL, for both antioxidant assays were of 423.83 and 202.95 respectively. It is concluded that HlMeOHe altered the development of the intraerythrocytic asexual stages and the ultrastructure of Pyy, and due to its phytochemical constituents, showed an in vitro antioxidant activity.

Detection of Antibodies against Mycoplasma mycoides subsp. capri in Goats with the Complement Fixation Test

Mycoplasma mycoides subsp. capri is the causative agent of severe and acute respiratory problems in goats, which spreads rapidly and represents high mortality. The serological profile of the goat population, from nine regions in seven states of Mexico, was screened by the Complement Fixation test (CF) in sera from asymptomatic goats and animals with mild respiratory symptoms. Sera and nasal swabs of 827 goats were collected for the isolation of the organism. An antiserum was prepared against a previously isolated field strain of Mycoplasma mycoides subsp. capri. CF Antibody titers were associated with the results of the isolates to determine the cutoff point. The CF was considered as positive if its result was ≥1/16. The CF registered 251 positive goats (30.35%) and 576 (69.65%) negative;the test showed high sensitivity (93.33%) and specificity (72.27%). In the specific case of diagnosis for mycoplasmosis associated with respiratory problems in goats, the CF proved to be a good diagnosis test, this study determined that 30% of the goat population showed antibody titers against Mycoplasma mycoides subsp. capri and revealed those animals who have had contact with this microorganism during their lives regardless of the presence or absence of respiratory symptoms.

Relevant Frequency of Multiple Infections with High- and Low-Risk HPV Genotypes among Mexican Women Attending a Tertiary Care Hospital

Aims: To assess the frequency and the main HPV genotypes circulating among a group of women attending at a third level Hospital in Mexico City. Methods: A cross-sectional and descriptive study was performed in a group of 143 female outpatients of the Gynecology and Obstetrics Service at the National Institute of Perinatology of Mexico. Cervical swabs were taken from participants and subjected to simultaneous detection/genotyping of HPV by Linear Array Genotyping Test (Roche Molecular Systems). Mann-Whitney U, median and/or Square Chi tests were used to compare socio-demographical features between HPV-infected and uninfected women. Results: A total of 66 women (46.2%) had HPV infection. Overall, 112 genotypes were detected either as single infections (45.5%) or multiple genotype infections (54.5%). The cumulated frequency of multiple infections with high-/low- and high-/high-risk HPV genotypes was 63.9 %. The most frequent high-risk genotypes were HPV52 HPV58 and HPV51, whereas the most frequent low-risk genotypes were HPV6, HPV53 and HPV84. Infected women were significantly younger and have less stable partner relationships than uninfected women (p < 0.05). Conclusion: A relevant frequency of mixed infections with high- and low-risk HPV genotypes, other than those considered most prevalent worldwide, was observed. Most circulating high-risk genotypes among the women of this study are not covered by commercial vaccine formulations.

Animals as sources of food-borne pathogens: A review

Food-producing animals are the major reservoirs for many foodborne pathogens such as Campylobacter species, non-Typhi serotypes of Salmonella enterica, Shiga toxin-producing strains of Escherichia coli, and Listeria monocytogenes. The zoonotic potential of foodborne pathogens and their ability to produce toxins causing diseases or even death are sufficient to recognize the seriousness of the situation. This manuscript reviews the evidence that links animals as vehicles of the foodborne pathogens Salmonella,Campylobacter, Shiga toxigenic E. coli, and L. monocytogenes, their impact, and their current status. We conclude that these pathogenic bacteria will continue causing outbreaks and deaths throughout the world, because no effective interventions have eliminated them from animals and food.

A Comprehensive Review on Human Aichi Virus

Although norovirus,rotavirus,adenovirus and Astrovirus are considered the most important viral agents transmitted by food and water,in recent years other viruses,such as Aichi virus(AiV),have emerged as responsible for gastroenteritis outbreaks associated with different foods.AiV belongs to the genus Kobuvirus of the family Picornaviridae.It is a virus with icosahedral morphology that presents a single stranded RNA genome with positive sense(8280 nucleotides)and a poly(A)chain.AiV was first detected from clinical samples and in recent years has been involved in acute gastroenteritis outbreaks from different world regions.Furthermore,several studies conducted in Japan,Germany,France,Tunisia and Spain showed a high prevalence of AiV antibodies in adults(between 80%and 99%),which is indicative of a large exposure to this virus.The aim of this review is to bring together all the discovered information about the emerging pathogen human Aichi virus(AiV),discussing the possibles routes of transmission,new detection techniques and future research.Although AiV is responsible for a low percentage of gastroenteritis outbreaks,the high seroprevalence shown by human populations indicates an evident role as an enteric agent.The low percentage of AiV detection could be explained by the fact that the pathogen is more associated to subclinical infections.Further studies will be needed to clarify the real impact of AiV in human health and its importance as a causative gastroenteritis agent worldwide.

A Saccharomyces cerevisiae RC016-based feed additive reduces liver toxicity,residual aflatoxin B1 levels and positively influences intestinal morphology in broiler chickens fed chronic aflatoxin B1-contaminated diets

The present study was conducted to investigate the ability of Saccharomyces cerevisiae RC016(Sc)-based feed additive to reduce liver toxicity,residual aflatoxin B1(AFB1)levels and influence intestinal structure in broiler chickens fed chronic aflatoxin B1-contaminated diets.A total of 100 one-day-old male commercial line(Ross)broiler chickens were divided into 4 treatments,with 5 pens per treatment and 5 broiler chickens per pen.Birds were randomly assigned to 4 treatments,which were namely treatment 1(T1),control diet(CD);T2,CD+Sc at 1 g/kg;T3,CD+AFB1 at 100μg/kg;T4,CD+Sc at 1 g/kg+AFB1 at100μg/kg.The liver histopathology of broiler chickens fed diets with AFB1 showed diffused microvacuolar fatty degeneration.The addition of Sc showed normal hepatocytes similar to the control.The small intestine villi from AFB1 group showed atrophy,hyperplasia of goblet cells,prominent inflammatory infiltrate and oedema.In contrast,the small intestine villi from birds that received the yeast plus AFB1 showed an absence of inflammatory infiltrate,and atrophy;moreover,a lower number of goblet cells compared to the groups with AFB1 was observed.The morphometric intestine studies showed that a significant decrease(P<0.05)in the crypt depth values when Sc was applied to AFB1-contaminated diets.Although the intestinal villus height and apparent adsorption area did not show significant differences(P>0.05),there was a tendency to improve these parameters.The residual levels of AFB1 in livers were significantly reduced(P<0.05)in the presence of the yeast.The present work demonstrated that the addition of Sc alone or in combination with AFB1 in the broiler chicken diets had a beneficial effect in counteracting the toxic effects of AFB1 in livers besides improving the histomorphometric parameters and modulating the toxic effect of AFB1 in the intestine.