An opportunistic pathogen isolated from the gut of an obese human causes obesity in germfree mice

Lipopolysaccharide endotoxin is the only known bacterial product which, when subcutaneously infused into mice in its purified form, can induce obesity and insulin resistance via an inflammation-mediated pathway. Here we show that one endotoxin-producing bacterium isolated from a morbidly obese human''s gut induced obesity and insulin resistance in germfree mice. The endotoxin-producing Enterobacter decreased in relative abundance from 35% of the volunteer''s gut bacteria to non-detectable, during which time the volunteer lost 51.4 kg of 174.8 kg initial weight and recovered from hyperglycemia and hypertension after 23 weeks on a diet of whole grains, traditional Chinese medicinal foods and prebiotics. A decreased abundance of endotoxin biosynthetic genes in the gut of the volunteer was correlated with a decreased circulating endotoxin load and alleviated inflammation. Mono-association of germfree C57BL/6J mice with strain Enterobacter cloacae B29 isolated from the volunteer''s gut induced fully developed obesity and insulin resistance on a high-fat diet but not on normal chow diet, whereas the germfree control mice on a high-fat diet did not exhibit the same disease phenotypes. The Enterobacter-induced obese mice showed increased serum endotoxin load and aggravated inflammatory conditions. The obesity-inducing capacity of this human-derived endotoxin producer in gnotobiotic mice suggests that it may causatively contribute to the development of obesity in its human host.     

Isolation and characterization of the emerging foodborn pathogen Arcobacter from human stool

At present, isolation of arcobacters from human specimens is performed by slightly of not modified Campylobacter, Yersinia or Leptospira isolation techniques, and knowledge if arcobacters are part of the human commensal flora is lacking. Therefore, an Arcobacter selective isolation procedure was validated for the examination of human fecal specimens, and the presence and characteristics of Arcobacter in feces of asymptomatic humans was examined in order to assess the clinical relevance of arcobacters in diarrheal stool. With this method, Arcobacter was isolated from seven of 500 (1.4%) stool samples of healthy people with Arcobacter cryaerophilus as the only species present. Seven A. cryaerophilus genotypes were detected and only one genotype was found per person. Neither A. butzleri nor A. skirrowii were isolated, therefore the presence of those latter species in clinical samples requires further attention. Though the pathogenic role and potential virulence factors of arcobacters have to be further examined, the current status of arcobacters as emerging pathogens remains justified.     

Enzymatic characterization of isocitrate dehydrogenase from an emerging zoonotic pathogen Streptococcus suis

Streptococcus suis, a Gram-positive coccus, is an emerging zoonotic pathogen for both humans and pigs, but little is known about the properties of its metabolic enzymes. Isocitrate dehydrogenase (IDH) is a key regulatory enzyme in the citric acid cycle that catalyzes the oxidative decarboxylation of isocitrate yielding α-ketoglutarate and NAD(P)H. Here, we report the overexpression and enzymatic characterization of IDH from S. suis Serotype 2 Chinese highly virulent strain 05ZYH33 (SsIDH). The molecular weight of SsIDH was estimated to be 74 kDa by gel filtration chromatography, suggesting a homodimeric structure. Additionally, SsIDH was divalent cation-dependent and Mg²⁺ was found to be the most effective cation. The optimal pH of SsIDH was 7.0 (Mn²⁺) and 8.5 (Mg²⁺), and the maximum activity was around 30 °C (Mn²⁺) and 50 °C (Mg²⁺), respectively. Heat inactivation studies showed that SsIDH retained 50% activity after 20 min of incubation at 49 °C. Sequence comparison revealed that SsIDH had a significantly homologous identity to bacterial homodimeric IDHs. The recombinant SsIDH displayed a 117-fold (k_cat/K_m) preference for NAD⁺ over NADP⁺ with Mg²⁺, and a 80-fold greater specificity for NAD⁺ than NADP⁺ with Mn²⁺. Therefore, SsIDH has remarkably high coenzyme preference toward NAD⁺. This current work is expected to shed light on the functions of metabolic enzymes in S. suis and provide useful information for SsIDH to be considered as a possible candidate for serological diagnostics and detection of S. suis infection.     

Multiple-antibiotic resistance of Enterococcus spp. isolated from commercial poultry production environments

The potential impact of food animals in the production environment on the bacterial population as a result of antimicrobial drug use for growth enhancement continues to be a cause for concern. Enterococci from 82 farms within a poultry production region on the eastern seaboard were isolated to establish a baseline of susceptibility profiles for a number of antimicrobials used in production as well as clinical environments. Of the 541 isolates recovered, Enterococcus faecalis (53%) and E. faecium (31%) were the predominant species, while multiresistant antimicrobial phenotypes were observed among all species. The prevalence of resistance among isolates of E. faecalis was comparatively higher among lincosamide, macrolide, and tetracycline antimicrobials, while isolates of E. faecium were observed to be more frequently resistant to fluoroquinolones and penicillins. Notably, 63% of the E. faecium isolates were resistant to the streptogramin quinupristin-dalfopristin, while high-level gentamicin resistance was observed only among the E. faecalis population, of which 7% of the isolates were resistant. The primary observations are that enterococci can be frequently isolated from the poultry production environment and can be multiresistant to antimicrobials used in human medicine. The high frequency with which resistant enterococci are isolated from this environment suggests that these organisms might be useful as sentinels to monitor the development of resistance resulting from the usage of antimicrobial agents in animal production.     

Corynebacterium pseudodiphtheriticum isolated from relevant clinical sites of infection: a human pathogen overlooked in emerging countries

Aims:  To examine the occurrence of and to determine the antimicrobial susceptibility of Corynebacterium pseudodiphtheriticum among patients with bacterial infections at a teaching hospital.
Methods and Results:  A total of 113 Coryne. pseudodiphtheriticum strains identified by conventional biochemical methods and API-Coryne System were recovered from patients from different age groups: 65·48% adults (18 to ≤59 years old), 9·73% aged (≥60 years old); 14·15% infants (<18 years old); 4·42% newborns (0–7 days). Micro-organisms were mostly related to infections in the urinary (29·2%) and respiratory tracts (27·45%) and intravenous sites (18·6%). Clinical samples were obtained only from 32·7% patients (26 adults, four aged, four infants and three newborns) presenting at least one of the predisposing conditions: end-stage renal disease; renal transplant; AIDS and Mycobacterium tuberculosis infection; cancer, hepatic cirrhosis; haemodialysis and catheter use. Antimicrobial susceptibility tests identified multiresistant phenotypes. Most strains (>50%) were resistant to oxacillin, erythromycin and clindamycin.
Conclusions:  Despite significant differences in age and functional status of patients Coryne. pseudodiphtheriticum may be implicated as a cause of respiratory and nonrespiratory human infections.
Significance and Impact of the Study:  Data are valuable for practitioners indicating the occurrence of multiresistant phenotypes and the possibility of severe infections due to Coryne. pseudodiphtheriticum , a pathogen usually overlooked in emerging countries.     

Vancomycin-resistant Enterococcus spp. isolated from wastewater and chicken feces in the United States

Vancomycin-resistant Enterococcus spp. (VRE) were isolated from sewage and chicken feces but not from other animal fecal sources (dog, cow, and pig) or from surface waters tested. VRE from hospital wastewater were resistant to > or =20 microg of vancomycin/ml and possessed the vanA gene. VRE from residential wastewater and chicken feces were resistant to 3 to 5 microg of vancomycin/ml and possessed the vanC gene.     

Molecular and phenotypic characterization of Pseudomonas spp. isolated from milk

Putative Pseudomonas spp. isolated predominantly from raw and processed milk were characterized by automated ribotyping and by biochemical reactions. Isolates were biochemically profiled using the Biolog system and API 20 NE and by determining the production of proteases, lipases, and lecithinases for each isolate. Isolates grouped into five coherent clusters, predominated by the species P. putida (cluster A), P. fluorescens (cluster B), P. fragi (as identified by Biolog) or P. fluorescens (as identified by API 20 NE) (cluster C), P. fragi (as identified by Biolog) or P. putida (as identified by API 20 NE) (cluster D), and P. fluorescens (cluster E). Isolates within each cluster also displayed similar enzyme activities. Isolates in clusters A, C, and D were generally negative for all three enzyme activities; isolates in cluster B were predominantly positive for all three enzyme activities; and isolates in cluster E were negative for lecithinase but predominantly positive for protease and lipase activities. Thus, only isolates from clusters B and E produced enzyme activities associated with dairy product flavor defects. Thirty-eight ribogroups were differentiated among the 70 isolates. Ribotyping was highly discriminatory for dairy Pseudomonas isolates, with a Simpson's index of discrimination of 0.955. Isolates of the same ribotype were never classified into different clusters, and ribotypes within a given cluster generally showed similar ribotype patterns; thus, specific ribotype fragments may be useful markers for tracking the sources of pseudomonads in dairy production systems. Our results suggest that ribogroups are generally homogeneous with respect to nomenspecies and biovars, confirming the identification potential of ribotyping for Pseudomonas spp.     

E. coli O104:H4: social media and the characterization of an emerging pathogen

The 2011 German E. coli O104:H4 outbreak resulted in thousands of cases of enterohaemorrhagic illness, with approximately 25% of these progressing to develop haemolytic uraemic syndrome (HUS). This high rate of progression to HUS was the first indicator that the bacterial cause of illness was not a typical enterohaemorrhagic E. coli (EHEC) strain. Collaborative bioinformatic analysis while the outbreak was still in progress indicated that the O104:H4 strain was in fact an enteroaggregative E. coli (EAEC) strain which had acquired genes for the production of Shiga - like toxin.     

Modelling an outbreak of an emerging pathogen

To illustrate the usefulness of mathematical models to the microbiology and medical communities, we explain how to construct and apply a simple transmission model of an emerging pathogen. We chose to model, as a case study, a large (>8,000 reported cases) on-going outbreak of community-acquired meticillin-resistant Staphylococcus aureus (CA-MRSA) in the Los Angeles County Jail. A major risk factor for CA-MRSA infection is incarceration. Here, we show how to design a within-jail transmission model of CA-MRSA, parameterize the model and reconstruct the outbreak. The model is then used to assess the severity of the outbreak, predict the epidemiological consequences of a catastrophic outbreak and design effective interventions for outbreak control.     

Preliminary characterization of bacteriocins produced by Enterococcus faecium and Enterococcus faecalis isolated from pig faeces

A total of 92 enterococci, isolated from the faeces of minipigs subjected to an in vivo feeding trial, were screened for the production of antimicrobial substances. Bacteriocin production was confirmed for seven strains, of which four were identified as Enterococcus faecalis and three as Enterococcus faecium, on the basis of physiological and biochemical characteristics. The bacteriocins produced by the Ent. faecalis strains showed a narrow spectrum of activity, mainly against other Enterococcus spp., compared with those from the Ent. faecium strains showing a broader spectrum of activity, against indicator strains of Enterococcus spp., Listeria spp., Clostridium spp. and Propionibacterium spp. The bacteriocins of all seven Enterococcus strains were inactivated by alpha-chymotrypsin, proteinase K, trypsin, pronase, pepsin and papain, but not by lipase, lysozyme and catalase. The bacteriocins were heat stable and displayed highest activity at neutral pH. The molecular weight of the bacteriocins, as determined by tricine SDS-PAGE, was approximately 3.4 kDa. Only the strains of Ent. faecalis were found to contain plasmids. PCR detection revealed that the bacteriocins produced by Ent. faecium BFE 1170 and BFE 1228 were similar to enterocin A, whereas those produced by Ent. faecium BFE 1072 displayed homology with enterocin L50A and B.     

Ramularia collo-cygni: the biology of an emerging pathogen of barley.

Ramularia collo-cygni is now recognized as an important pathogen of barley in Northern Europe and New Zealand. It induces necrotic spotting and premature leaf senescence, leading to loss of green leaf area in crops, and can result in substantial yield losses. The fungus produces a number of anthraquinone toxins called rubellins, which act as host nonspecific toxins with photodynamic activity. These toxins induce lipid peroxidation and are possibly the cause of the chlorosis and necrosis observed in leaves infected with R. collo-cygni. The fact that the fungus can remain latent in barley plants until flowering, coupled with its very slow growth in vitro, makes it difficult to detect in crops. As a result, the epidemiology of this pathogen remains poorly understood. However, the recent development of rapid and reliable PCR methods for specific detection of R. collo-cygni offers the prospect of increased understanding of its epidemiology and improved disease control.     

Molecular characterization of Enterococcus faecium isolated from hospitalized patients in Korea

AIMS: Multilocus sequence typing (MLST) was performed for vancomycin-resistant Enterococcus faecium (VREF) from diverse geographical areas in Korea to obtain insights into the genetic relationships with other molecular profiles. To understand the diversity of lineages, vancomycin-susceptible E. faecium (VSEF) were included. METHODS AND RESULTS: A total of 60 E. faecium isolates were analysed by MLST and esp profile. Molecular typing of Tn1546 of 30 VREF strains was evaluated by overlapping PCR of Tn1546 and DNA sequencing. Seven sequence types (ST) were found among 30 VSEF isolates, and four STs were found among 30 VREF isolates. The types most frequently encountered were ST 78 (26 isolates) and ST 203 (16 isolates). Of the 60 E. faecium isolates, 35 isolates were positive for the esp gene. On molecular typing of Tn1546, all VREF isolates were divided into four main types. Strains with the same ST showed divergence in Tn1546 types and strains with the same Tn1546 type represented different STs. CONCLUSIONS: An association between Tn1546 typing and MLST was not found. SIGNIFICANCE AND IMPACT OF THE STUDY: These results suggest that the horizontal spread of Tn1546 between strains plays a major role in the dissemination of vancomycin resistance in Korea.     

Identification and characterization of an RTX toxin in the emerging pathogen Kingella kingae

Kingella kingae is an emerging bacterial pathogen that is increasingly recognized as the causative agent of a variety of pediatric diseases, including septic arthritis and osteomyelitis. The pathogenesis of K. kingae disease is believed to begin with colonization of the upper respiratory tract. In the present study, we examined interactions between K. kingae and cultured respiratory epithelial cells and observed potent cytotoxicity, detected by both microscopy and lactic acid dehydrogenase (LDH) release assays. Experiments with synovial and macrophage cell lines revealed cytotoxicity for these cell types as well. Using mariner mutagenesis and a screen for loss of cytotoxicity, a genetic locus encoding an RTX toxin system was identified. Disruption of the K. kingae RTX locus resulted in a loss of cytotoxicity for respiratory epithelial, synovial, and macrophage cell lines. DNA sequence analysis demonstrated that the RTX locus is flanked by insertion elements and has a reduced G+C content compared to that of the whole genome. Two relatively less invasive Kingella species, K. oralis and K. denitrificans, were found to be noncytotoxic and to lack the RTX region, as determined by LDH release assays and Southern blotting. We concluded that K. kingae expresses an RTX toxin that has wide cellular specificity and was likely acquired horizontally. The possible roles for this toxin in the pathogenesis of K. kingae disease include breaching of the epithelial barrier and destruction of target tissues, such as synovium (joint lining).     

Laribacter hongkongensis: an emerging pathogen of infectious diarrhea

Laribacter hongkongensis is relatively a new name in the list of bacterial pathogens for gastroenteritis and travelers’ diarrhea. Addition of another name increases burden on the enteric infections as a whole. L. hongkongensis belongs to Neisseriaceae family of β subclass Proteobacteria. L. hongkongensis was initially isolated in Hong Kong from blood and empyema of an alcoholic cirrhotic patient in 2001, followed by reports from Korea and China, representing a total of 38 articles in PubMed until April 2013. As of now, there is no report from Indian subcontinent where infectious diarrhea is very much prevalent and a major burden. This review provides information about the microbiological characteristics, consideration of an emerging pathogen, relative pathogenicity, genome and proteome content, resistance toward multiple antibiotics, adaptability to different stress, and other features since its time of discovery. Investigation for this bacterium may avoid misidentification as other microbial flora. Further studies like the geographical distribution, type of infection, disease burden, pathogenicity, or genomic exploration of this bacterium will be useful in characterizing them properly. This bacterium may possibly be the emerging threat to public health.     

Genomic description and characterization of Nigeribacterium massiliense gen. nov., sp. nov., isolated from the human gut

Strain Marseille-P1302 was isolated from the stool of a 2-year-old Nigerian boy suffering from Kwashiorkor, a form of severe acute malnutrition. The strain grows in aerobic atmosphere and bacterial cells are Gram-positive cocci ranging in diameter from 0.8 to 1 μm. Among species with standing in nomenclature, strain Marseille-P1302 exhibited a highest 16S rRNA sequence similarity of 94.97% with Brevilactibacter flavus strain VG341^T, but was phylogenetically-closest to Brevilactibacter sinopodophylli strains KCTC 33808^T. The draft genome of strain Marseille-P1302 was 2,934,258-bp-long with a 70.38% G + C content, and contained 2704 protein-coding genes and 55 RNAs that included 9 rRNA genes. On the basis of these data, we propose the creation of the new genus Nigeribacterium gen. nov., with strain Marseille-P1302^T (= CSUR P1302 = DSM 29084) being the type strain of the new species Nigeribacterium. massiliense gen. nov., sp. nov.     

Genome sequence of Diplorickettsia massiliensis, an emerging Ixodes ricinus-associated human pathogen

Diplorickettsia massiliensis is a gammaproteobacterium in the order Legionellales and an agent of tick-borne infection. We sequenced the genome from strain 20B, isolated from an Ixodes ricinus tick. The genome consists of a 1,727,973-bp chromosome but no plasmid and includes 2,269 protein-coding genes and 42 RNA genes, including 3 rRNA genes.     

Complete genome sequence of Rickettsia heilongjiangensis, an emerging tick-transmitted human pathogen

Rickettsia heilongjiangensis is an emerging tick-transmitted human pathogen causing far-Eastern spotted fever. Here we report the complete sequence and the main features of the genome of R. heilongjiangensis (strain 054).     

Genotypic characterization of Brochothrix spp. isolated from meat,poultry and fish

Aim: To study genotypic diversity of isolates of Brochothrix thermosphacta recovered from meat, poultry and fish. Methods and Results: A total of 27 bacteria isolated from 19 samples of meat, poultry and fish were identified phenotypically and genotypically using PCR amplification of 16S‐23S rDNA intergenic transcribed spacer (ITS‐PCR), repetitive sequence‐based PCR (rep‐PCR) and 16S rDNA sequencing. Using ITS‐PCR, all bacteria showed the same DNA profile as the reference strains of Br. thermosphacta, allowing typing of the isolates at species level. Using 16S rDNA sequencing, all isolates were identified, at genus and species level, as Br. thermosphacta. Identification as Br. campestris was observed with a lower, but very close, level of similarity. Rep‐PCR was more discriminatory than ITS‐PCR and allowed differentiation of four subgroups among the isolates. Conclusion: Minor genotypic differences among Br. thermosphacta strains from meat, poultry and fish were observed. Significance and Impact of the Study: A rudimentary exploration of genotypic differences of Br. thermosphacta from meat, poultry and fish resulted in preliminary confirmation of the suitability of ITS‐PCR for typing Br. thermosphacta and confirmed the value of rep‐PCR fingerprinting to discriminate between Br. thermosphacta strains.