Role of Bacterial Pathogens in Atopic Dermatitis

The skin of atopic dermatitis (AD) patients exhibits a striking susceptibility to colonization and infection with Staphylococcus aureus. This review summarizes our understanding about the role of S. aureus in AD. Indeed, S. aureus colonization is both a cause and a consequence of allergic skin inflammation. The mechanisms that allergic skin inflammation of AD promotes the increase of S. aureus colonization include skin barrier dysfunction, increased synthesis of the extracellular matrix adhesins for S. aureus, and defective innate immune responses due to decreased production of endogenous antimicrobial peptides. On the other hand, the exotoxins secreted by S. aureus are superantigens. Staphylococcal superantigens (SsAgs) may penetrate the skin barrier and contribute to the persistence and exacerbation of allergic skin inflammation in AD through the stimulation of massive T cells, the role of allergens, direct stimulation of antigen-presenting cells and keratinocytes, the expansion of skin-homing cutaneous lymphocyte-associated antigen-positive T cells, and the augmentation of allergen-induced skin inflammation. SsAgs also induce corticosteroid resistance. In therapeutic interventions, anti-inflammatory therapy alone is very effective in reducing S. aureus colonization on the skin, but antibiotic treatment alone is unable to improve the allergic skin inflammation of AD. Therefore, we recommend the combination therapy of anti-inflammatory drugs and antibiotics in the AD patients with secondary bacterial infection, exacerbated AD, or poorly controlled AD. However, when AD is well controlled by anti-inflammatory drugs alone, we do not recommend the antibiotic therapy.     

Bacteriophages: Potential Biocontrol Agents and Treatment Options for Bacterial Pathogens

The global report on antibiotic resistance by the World Health Organization features the burden of antimicrobial resistance and the emerging risk of a post-antibiotic era, where therapeutic options will no longer exist for previously treatable diseases. The search for alternative avenues to combat multidrug-resistant (MDR) pathogens has sparked interest in the use of bacteriophages for the treatment of bacterial infections. The unique ability of bacteriophages to infect and kill bacteria without affecting mammalian cells makes them a promising avenue as biocontrol agents against bacterial pathogens. Phage therapy in both animal and human models has shown promising results. Further, the phage-derived proteins (lytic enzymes), such as endolysins, that are the key weapons used by phages to degrade the bacterial cell have been explored by the scientific community as viable therapeutic options themselves. This review focuses on the prospects of phage therapy; an overview of diverse phages isolated against MDR pathogens; phage-derived proteins as antibacterial agents; phage cocktail development; phage and antibiotic combinational therapy; and the application of phages as biocontrol agents in agriculture, veterinary science, synthetic biology, and aquaculture. The review emphasizes the therapeutic use of phages and how they might be applied on a large scale if certain limitations and challenges can be overcome.     

Tetratricopeptide Repeat Motifs in the World of Bacterial Pathogens: Role in Virulence Mechanisms

The tetratricopeptide repeat (TPR) structural motif is known to occur in a wide variety of proteins present in prokaryotic and eukaryotic organisms. The TPR motif represents an elegant module for the assembly of various multiprotein complexes, and thus, TPR-containing proteins often play roles in vital cell processes. As the TPR profile is well defined, the complete TPR protein repertoire of a bacterium with a known genomic sequence can be predicted. This provides a tremendous opportunity for investigators to identify new TPR-containing proteins and study them in detail. In the past decade, TPR-containing proteins of bacterial pathogens have been reported to be directly related to virulence-associated functions. In this minireview, we summarize the current knowledge of the TPR-containing proteins involved in virulence mechanisms of bacterial pathogens while highlighting the importance of TPR motifs for the proper functioning of class II chaperones of a type III secretion system in the pathogenesis of Yersinia, Pseudomonas, and Shigella.     

Chemical Biology Applied to the Study of Bacterial Pathogens

In recent years, chemical biology and chemical genomics have been increasingly applied to the field of microbiology to uncover new potential therapeutics as well as to probe virulence mechanisms in pathogens. The approach offers some clear advantages, as identified compounds (i) can serve as a proof of principle for the applicability of drugs to specific targets; (ii) can serve as conditional effectors to explore the function of their targets in vitro and in vivo; (iii) can be used to modulate gene expression in otherwise genetically intractable organisms; and (iv) can be tailored to a narrow or broad range of bacteria. This review highlights recent examples from the literature to illustrate how the use of small molecules has advanced discovery of novel potential treatments and has been applied to explore biological mechanisms underlying pathogenicity. We also use these examples to discuss practical considerations that are key to establishing a screening or discovery program. Finally, we discuss the advantages and challenges of different approaches and the methods that are emerging to address these challenges.     

Antimicrobial Susceptibility of Bacterial Pathogens in the Oropharynx of Healthy Children

 In a study to determine the prevalence and antimicrobial susceptibility of bacterial pathogens in the oropharynx of healthy children, throat swabs obtained from 1765 children were cultured and the organisms recovered tested by the disk diffusion method and the E test. Six hundred ninety-one children (39.1%) harbored Haemophilus influenzae, 112 (6.3%) Streptococcus pyogenes, 73 (4.1%) Moraxella catarrhalis, 52 (2.9%) Streptococcus pneumoniae, and 50 (2.8%) Neisseria meningitidis in their oropharynx. The rate of penicillin resistance was 2%, 0%, and 12%, respectively, for Streptococcus pneumoniae, Streptococcus pyogenes, and Neisseria meningitidis. Ampicillin resistance was observed in 8.6% of Haemophilus influenzae strains and 78% of Moraxella catarrhalis strains and was associated with the presence of β-lactamase, except in one strain of Haemophilus influenzae. Five (4.4%) isolates of Streptococcus pyogenes were resistant to macrolides. The low level of resistance observed in this area contrasts with the high rates reported in the literature.     

Bacterial Pathogens in Food: Modern Safety Criteria and Methods of Molecular Genetic Analysis

Bulletin of Experimental Biology and Medicine - The review is devoted to the issues of improving the food biosafety system by developing and implementing modern methods of microbiological and...     

Autotransporter and Two-Partner Secretion: Delivery of Large-Size Virulence Factors by Gram-Negative Bacterial Pathogens

A number of protein secretion mechanisms have been identified in gram-negative pathogens. Many of these secretion systems are dependent upon the Sec translocase for protein export from the cytoplasm into the periplasm and then utilize other mechanisms for transport from the periplasm through the outer membrane. In this article, we review secretion similarities between autotransporter and two-partner secretion systems, and we report similarities between the autotransporter secretion mechanism with that of intimin/invasins. Considering that many secreted proteins are virulence factors, a better understanding of their secretion mechanisms will aid in the development of disease treatments and new bacterial vaccines.     

Detection of Bacterial Pathogens in Purulent Clinical Specimens by Immunofluorescence Techniques

A technique is described which may be used to identify Haemophilus influenzae type b, Streptococcus pneumoniae, group A and B streptococci, and Staphylococcus aureus in smears of clinical specimens.     

Occurrence of Immunoglobulin M Antibodies against Several Bacterial and Viral Pathogens in Acute Hantavirus Infection

Elevated levels of immunoglobulin M antibodies against various pathogens, most frequently Epstein-Barr-virus and Coxiella burnetii, were detected by immunoassay in 15 of 48 patients (31.3%) with acute Puumala virus infections. Although the mechanisms leading to this IgM response are not clear yet, polyspecific immunoglobulin M antibodies have to be taken into account to avoid misinterpretation of serological results in acute hantavirus infection.     

Intestinal Epithelial CD98 Directly Modulates the Innate Host Response to Enteric Bacterial Pathogens

CD98 is a type II transmembrane glycoprotein whose expression increases in intestinal epithelial cells (IECs) during intestinal inflammation. Enteropathogenic Escherichia coli (EPEC) is a food-borne human pathogen that attaches to IECs and injects effector proteins directly into the host cells, thus provoking an inflammatory response. In the present study, we investigated CD98 and EPEC interactions in vitro and ex vivo and examined FVB wild-type (WT) and villin-CD98 transgenic mice overexpressing human CD98 in IECs (hCD98 Tg mice) and infected with Citrobacter rodentium as an in vivo model. In vivo studies indicated that CD98 overexpression, localized to the apical domain of colonic cells, increased the attachment of C. rodentium in mouse colons and resulted in increased expression of proinflammatory markers and decreased expression of anti-inflammatory markers. The proliferative markers Ki-67 and cyclin D1 were significantly increased in the colonic tissue of C. rodentium-infected hCD98 Tg mice compared to that of WT mice. Ex vivo studies correlate with the in vivo data. Small interfering RNA (siRNA) studies with Caco2-BBE cells showed a decrease in adherence of EPEC to Caco2 cells in which CD98 expression was knocked down. In vitro surface plasmon resonance (SPR) experiments showed direct binding between recombinant hCD98 and EPEC/C. rodentium proteins. We also demonstrated that the partial extracellular loop of hCD98 was sufficient for direct binding to EPEC/C. rodentium. These findings demonstrate the importance of the extracellular loop of CD98 in the innate host defense response to intestinal infection by attaching and effacing (A/E) pathogens.     

Minimum Core Genome Sequence Typing of Bacterial Pathogens: a Unified Approach for Clinical and Public Health Microbiology

Bacterial pathogens impose a heavy health burden worldwide. In the new era of high-throughput sequencing and online bioinformatics, real-time genome typing of infecting agents, and in particular those with potential severe clinical outcomes, holds promise for guiding clinical care to limit the detrimental effects of infections and to prevent potential local or global outbreaks. Here, we sequenced and compared 85 isolates of Streptococcus suis, a zoonotic human and swine pathogen, wherein we analyzed 32 recognized serotypes and 75 sequence types representing the diversity of the species and the human clinical isolates with high public health significance. We found that 1,077 of the 2,469 genes are shared by all isolates. Excluding 201 common but mobile genes, 876 genes were defined as the minimum core genome (MCG) of the species. Of 190,894 single-nucleotide polymorphisms (SNPs) identified, 58,501 were located in the MCG genes and were referred to as MCG SNPs. A population structure analysis of these MCG SNPs classified the 85 isolates into seven MCG groups, of which MCG group 1 includes all isolates from human infections and outbreaks. Our MCG typing system for S. suis provided a clear separation of groups containing human-associated isolates from those containing animal-associated isolates. It also separated the group containing outbreak isolates, including those causing life-threatening streptococcal toxic shock-like syndrome, from sporadic or less severe meningitis or bacteremia-only isolates. The typing system facilitates the application of genome data to the fields of clinical medicine and epidemiology and to the surveillance of S. suis. The MCG groups may also be used as the taxonomical units of S. suis to define bacterial subpopulations with the potential to cause severe clinical infections and large-scale outbreaks.     

Parallel Detection of Bacterial Pathogens in Cerebrospinal Fluid with 16S rRNA Probe Microarray

The most commonly used method for detection ofpathogenic bacteria in cerebrospinal fluid (CSF) speci mens of clinical laboratories is isolation and identificationof the causative agents by cultural method, biochemicaland serological t…     

16S Ribosomal DNA Typing for Identification of Pathogens in Patients with Bacterial Keratitis

The identification of pathogens in patients with bacterial keratitis remains problematic because standard diagnostic tests are negative for 40 to 60% of patients. A cross-sectional study was undertaken to determine if PCR and sequence analysis of 16S ribosomal DNA (rDNA) could be used to detect bacterial pathogens in patients with keratitis. Corneal specimens were collected for culture and rDNA typing. Variable segments of each rDNA specimen were amplified by PCR, sequenced, and aligned with the sequences in GenBank. Eleven patients had microbiologically documented bacterial keratitis, while 17 patients had keratitis due to other causes. Nine (82%) of 11 bacterial keratitis patients were PCR positive; each sequencing result matched the culture results. Seventeen (100%) patients with nonbacterial keratitis were PCR negative. Our data suggest that 16S rDNA typing holds promise as a rapid alternative to culture for identifying pathogens in patients with bacterial keratitis.     

Comparison of the EntericBio Multiplex PCR System with Routine Culture for Detection of Bacterial Enteric Pathogens

The EntericBio system uses a multiplex PCR assay for the simultaneous detection of Campylobacter spp., Salmonella enterica, Shigella spp., and Escherichia coli O157 from feces. It combines overnight broth enrichment with PCR amplification and detection by hybridization. An evaluation of this system was conducted by comparing the results obtained with the system with those obtained by routine culture, supplemented with alternative PCR detection methods. In a study of 773 samples, routine culture and the EntericBio system yielded 94.6 and 92.4% negative results, respectively. Forty-two samples had positive results by culture, and all of these were positive with the EntericBio system. This system detected an additional 17 positive samples (Campylobacter spp., n = 12; Shigella spp., n = 1; E. coli O157, n = 4), but the results for 5 samples (Campylobacter spp., n = 2; Shigella spp., n = 1; E. coli O157, n = 2) could not be confirmed. The target for Shigella spp. detected by the EntericBio system is the ipaH gene, and the molecular indication of the presence of Shigella spp. was investigated by sequence analysis, which confirmed that the ipaH gene was present in a Klebsiella pneumoniae isolate from the patient. The sensitivity, specificity, positive predictive value, and negative predictive value were 100%, 99.3%, 91.5%, and 100%, respectively. Turnaround times were significantly reduced with the EntericBio system, and a result was available between 24 and 32 h after receipt of the sample in the laboratory. In addition, the amount of laboratory waste was significantly reduced by use of this system. In summary, the EntericBio system proved convenient to use, more sensitive than the conventional culture used in this study, and highly specific; and it generated results significantly faster than routine culture for the pathogens tested.Infectious gastroenteritis is a leading cause of morbidity and mortality worldwide (3), particularly in developing countries (2). There are also a significant economic and social costs associated with gastroenteritis (14), in addition to increased morbidity and mortality (5).Conventional culture methods remain the norm for the isolation of bacterial enteric pathogens in clinical laboratories. A major advantage of molecular methods is the reduced time to detection (13). Faster diagnostic outputs allow earlier epidemiological investigations and infection control interventions. Furthermore, the use of molecular methods highlights that conventional methods for the isolation of Campylobacter are less sensitive than PCR (15). Molecular methods suitable for use in the detection of fecal enteric pathogens have not been routinely available to clinical laboratories, until recently. Here we describe a new multiplex PCR method for clinical diagnostic use. The method combines an overnight enrichment step with PCR and hybridization detection by use of a line blot assay for the simultaneous detection of Campylobacter spp., Salmonella enterica, Shigella spp., and Escherichia coli O157 from feces. The application of the system in a clinical laboratory is also described.     

Utility of a Commercially Available Multiplex Real-Time PCR Assay To Detect Bacterial and Fungal Pathogens in Febrile Neutropenia

Infection is the main treatment-related cause of mortality in cancer patients. Rapid and accurate diagnosis to facilitate specific therapy of febrile neutropenia is therefore urgently warranted. Here, we evaluated a commercial PCR-based kit to detect the DNA of 20 different pathogens (SeptiFast) in the setting of febrile neutropenia after chemotherapy. Seven hundred eighty-four serum samples of 119 febrile neutropenic episodes (FNEs) in 70 patients with hematological malignancies were analyzed and compared with clinical, microbiological, and biochemical findings. In the antibiotic-naïve setting, bacteremia was diagnosed in 34 FNEs and 11 of them yielded the same result in the PCR. Seventy-three FNEs were negative in both systems, leading to an overall agreement in 84 of 119 FNEs (71%). During antibiotic therapy, positivity in blood culture occurred only in 3% of cases, but the PCR yielded a positive result in 15% of cases. In six cases the PCR during antibiotic treatment detected a new pathogen repetitively; this was accompanied by a significant rise in procalcitonin levels, suggestive of a true detection of infection. All patients with probable invasive fungal infection (IFI; n = 3) according to the standards of the European Organization for Research and Treatment of Cancer had a positive PCR result for Aspergillus fumigatus; in contrast there was only one positive result for Aspergillus fumigatus in an episode without signs and symptoms of IFI. Our results demonstrate that the SeptiFast kit cannot replace blood cultures in the diagnostic workup of FNEs. However, it might be helpful in situations where blood cultures remain negative (e.g., during antimicrobial therapy or in IFI).While systemic infection is the most common cause of a febrile neutropenia episode (FNE) with significant effects on morbidity and mortality, only 30% of blood cultures taken at the onset of fever are positive (11, 15). Nonetheless, patients with FNEs are treated with broad-spectrum antimicrobial agents regardless of the result of their blood culture (7) because potentially life-threatening infections need early treatment to ensure better clinical outcome. Noninfective causes of a systemic reaction culminating in a rise in temperature such as tumor fever, drug fever, or transfusion reactions complicate the diagnostic challenge in cancer patients. In addition, the etiology of a deterioration of an FNE during antimicrobial therapy is often difficult to elucidate, since blood cultures are infrequently positive once effective antimicrobial therapy has started (4). Pathogens such as molds which are rarely found in blood cultures are not uncommon in patients with FNEs, particularly if they suffer from hematological malignancies. For these reasons, FNE is one of the conditions where new diagnostic tools to distinguish an infection from a nonmicrobial cause for fever or to identify rare pathogens are most urgently needed. In the past, raised levels of indirect markers such as procalcitonin (PCT) and interleukin 6 (3, 16) have been shown to be associated with bacteremia. Ideally, though, the cause should be identified directly and improvements in the detection of pathogens in the bloodstream should be made. In addition to refinements of the classical blood culture systems, attempts have been made to detect pathogen DNA by means of PCR. Initially, this involved conventional PCR techniques (9) detecting the gene for the 16S subunit of bacterial rRNA for the presence of bacterial DNA. Specification was then carried out by sequencing the PCR product. Later, more rapid methods were developed when real-time PCR became available (1, 14, 17). PCR results are more readily available, and the method also detects remnants of bacteria, which might make it more robust to the influence of antibiotic treatment, while potentially detecting pathogens which do not grow in the blood cultures. However, the main disadvantage, apart from higher costs, a potential for false-positive results during transient bacteremia/fungemia (e.g., during brushing of teeth), and laboratory workload, is the restriction of the spectrum of species detected. In addition, because most PCR methods use smaller sample volumes (commonly 1 to 4 ml [9, 17]), this method depends on a higher concentration of bacteria than that for the blood culture, which theoretically can reveal positive results after one living and propagating bacterial cell has been injected into the culture bottle.A new commercially available kit (SeptiFast) to detect DNA from 20 clinically relevant pathogens has recently been evaluated in a small cohort of neutropenic patients (10) with promising results. The aim of our study was to evaluate the usefulness of the SeptiFast kit in a larger cohort of patients with febrile neutropenia after chemotherapy for hematological malignancies. Also, we sought to determine a correlation between SeptiFast results and clinical findings. Altogether 784 samples from 119 FNEs in 70 patients were analyzed and compared with clinical, microbiological, and biochemical findings in the antibiotic-naïve setting and during antimicrobial therapy.     

肝硬化合并自发性细菌性腹膜炎患者病原菌分布及药物敏感性分析

目的 研究肝硬化合并自发性细菌性腹膜炎(spontaneous bacterial peritonitis,SBP)腹水细菌培养阳性患者的病原菌分布及药物敏感性,为SBP患者合理使用抗生素提供理论依据.方法 回顾性分析2011～ 2014年在首都医科大学附属北京地坛医院住院肝硬化伴有腹水症状且送检腹水细菌培养患者资料,研究对象为其中腹水细菌培养阳性患者.结果 肝硬化并有腹水症状且送检腹水细菌培养患者共638例,其中细菌培养阳性者67例(10.50％).细菌培养阳性患者中共培养39株菌,其中革兰阴性杆菌19株(49％),革兰阳性球菌20株(51％).阳性率前5位的细菌依次是肠球菌7株(17.9％)、链球菌7株(17.9％)、葡萄球菌6株(15.4％)、大肠埃希菌6株(15.4％)、铜绿假单胞菌3株(7.7％),其它菌种均为革兰阴性杆菌,所占比例较小(共占25.7％).革兰阴性杆菌对碳青霉烯类、氨基糖苷类等的敏感率较高,均超过83％;革兰阳性球菌对万古霉素敏感率为100％.结论 肝硬化并发SBP患者细菌感染以革兰阳性球菌为主,其中革兰阳性球菌以屎肠球菌为主,革兰阴性杆菌以大肠埃希菌为主.临床治疗肝硬化并发SBP应在经验用药的基础上根据药敏结果合理应用抗生素.     

Evaluation of Phytochemical and Antibacterial Potential of Helicteres Isora L. Fruits Against Enteric Bacterial Pathogens

Antibacterial activities of aqueous, acetone, ethanol and methanol extracts of fruits of Helicteres isora (Mororphali) were studied. The fruit aqueous extracts of H. isora showed prominent antibacterial activities against E.coli, Staphylococcus epidermidis, Salmonella typhimurium and Proteus vulgaris; moderate activity against Enterobacter aerogenes, Staphylococcus aureus, Salmonella typhi and least activity against Pseudomonas aeruginosa. The aqueous extract showed maximal, the ethanol and methanol extract moderate and acetone extracts least antibacterial activities. Phytochemical screening revealed the presence of carbohydrates, anthraquinon glycosides, proteins, tannin and phenolic compounds and steroids These antibacterial properties supports its traditional use of fruits of H. isora in the treatment of enteric or diarrhoeal infections.