Staphylococcus aureus may be a direct pathogenetic factor in granulomatosis with polyangiitis (GPA). Chronic bacterial colonization or chronic infections of the upper respiratory tract have been proposed as trigger of IgA vasculitis and IgA nephropathy. Interventions to remove major lymphoid organs, such as tonsillectomy, have shown conflicting results but may be an option in IgA vasculitis. Interestingly no clear clinical benefit despite similar local colonization with bacterial strains has been detected in patients with IgA nephropathy. In systemic lupus erythematosus injection of bacterial lipopolysaccharide induced progressive lupus nephritis in mouse models. The aim of this review is to discuss and summarize the knowledge of microbial antigens in autoimmune renal disease. Novel methods may provide insight into the involvement of microbial antigens in the onset, progression, and prognosis of autoimmune kidney disorders.A better understanding of the pathophysiology of autoimmune disorders is desired to allow tailored interventions. Despite increased scientific interest a direct pathogenic factor in autoimmune renal disease has been described only in a minority like membranous nephropathy or ANCA-associated vasculitis. Nonetheless the initial step leading to the formation of these antibodies is still obscure. In this review we will focus on the possible role of microbial factors in this context.  Over the last decades several investigations have focused on the pathogenesis of immunologically mediated kidney diseases. Besides the detection of novel aspects responsible for the onset of particular diseases, such as the phospholipase A2 (PLA2) receptor  and moreAbnormalities in the immune reactions can induce kidney damage, either by provoking an autoimmune phenomenon or induction of molecular mimicry. The immune response comprises the innate as well as the more orchestrated adaptive immunity. The former acts as the first line of defence and is mediated by monocytes/macrophages, neutrophils, natural killer cells, complement activation, and cytokines. Pathogen-associated small molecules recognized by pattern recognition receptors such as toll-like receptors (TLR), C-type lectin receptors, and complement activation via the mannose-binding lectin pathway initiate innate recognition of bacteria and thus activation of inflammation and coagulation. Once persistent, the antigenic stimulus leads to the activation of the adaptive immunity triggering a T-cell and B-cell response. An autoimmune response occurs whenever the immune system activation leads to a response against self-antigens. Several factors are implicated in the loss of immune tolerance including genetic, hormonal, and environmental factors as well as a defective clearance of apoptotic cells –5.Characterization of the human microbiome  offers nBacterial infection-related glomerulonephritis has attracted attention for a long period of time, for example, with streptococcal infections as leading cause of postinfectious glomerulonephritis in children and staphylococcal infection in adults . In this∗ OR microbiology OR microbio∗ OR microbiome OR microbial OR microorganism OR microbiota) AND (glomerulonephritis OR nephrotic syndrome).” Additional studies were identified by examining the bibliography of the retrieved articles by forward search. The search was limited to articles published in English and reports on postinfectious and IgA-dominant acute staphylococcal associated glomerulonephritis forms were excluded from further analysis.A systematic literature search of the MEDLINE database was conducted, using the key words: “(bacteria OR bactern = 2196) was excluded, since these records reported on treatment regimens, antibody findings (such as anti-neutrophil cytoplasmic antibodies), and patients with virus-related diseases (such as hepatitis C-associated membranoproliferative glomerulonephritis or human immunodeficiency virus-associated kidney disease) or were single case reports/case series. After access to full text articles we excluded another 244 articles, which were not reporting on bacterial antigens or bacteria leading to autoimmune kidney diseases but rather again reported on poststreptococcal or poststaphylococcal glomerulonephritis or viral antigens, leading to a number of 68 articles for further full text analysis. Through forward search of the retrieved bibliography we identified another 34 records, which were also analyzed. Of these, one article was not indexed in common electronic databases. After exclusion of one article reporting the same results a total of 100 records were finally included  overall resulted in 2508 records. A large number of articles (uded see .The group of anti-neutrophil cytoplasmic antibody- (ANCA-) associated vasculitis (AAV) consists of four distinct diseases, namely, granulomatosis with polyangiitis , microscopic polyangiitis (MPA), eosinophilic granulomatosis with polyangiitis , and renal limited vasculitis. The extent of kidney involvement varies between and within each clinical entity being around 90–100% in MPA, 50–80% in GPA, and, depending on the ANCA status, 4–51% in EGPA . Several Staphylococcus aureus obtained from nasal swab cultures was identified as an independent risk factor for relapse of GPA when compared to noncarriers. Chronic Staphylococcus aureus carriers were defined as a positive culture in ≥75% of the specimen  examined [ Staphylococcus aureus may be responsible for this beneficial effect, although nasal swabs were not performed. In addition, a significant reduction of respiratory and nonrespiratory tract infections was reported in actively treated patients [ Staphylococcus aureus (positivity in ≥80% of samples obtained) was furthermore confirmed as independent risk factor for relapse in biopsy-proven GPA in another cohort [ Staphylococcus aureus superantigen, which can lead to an antigen-independent T-cell activation. In general, the presence of Staphylococcus aureus was accompanied by a higher risk for relapse, which however was modulated by the type of superantigen present, with mostly tsst-1 being associated with disease relapse [ Staphylococcus aureus or related superantigens are capable of T-cell expansion were performed. T-cell expansion was present at a significantly higher rate in patients with GPA than in healthy individuals but was neither associated with the presence of Staphylococcus aureus nor associated with its superantigens [ Staphylococcus aureus presence in vasculitis, staphylococcal acid phosphatase (SAcP) and its binding ability were studied in human umbilical vein endothelial cells. SAcP was capable of binding to endothelial cells in a concentration-dependent manner and endothelial cell-bound SAcP was recognized by sera of patients with GPA [ Staphylococcus aureus (n = 7) led to the development of pauci-immune glomerulonephritis in one rat [Bacterial antigens potentially implicated in the pathogenesis of AAV have been studied for three decades. In a prospective longitudinal cohort the chronic presence ofexamined . A subsepatients . Chronicr cohort . A retro relapse . Furtherantigens . In ordewith GPA . Immuniz one rat .41–49 to the amino acids 72–80 of mature bacterial adhesin FimH, which is present on the tip of fimbriae crucial for attachment to host epithelia of Gram negative bacteria such as Escherichia coli, Klebsiella pneumonia, and Proteus mirabilis. A retrospective survey revealed a high proportion of Gram negative infections prior to the onset of disease activity and autoantibodies to hLAMP-2 were detected in nearly all patients with renal involvement and underlying AAV in one study. The same authors reported that hLAMP-2 autoantibodies, when injected into rats, were causative of developing focal necrotizing glomerulonephritis and a monoclonal antibody directed against hLAMP-2 induced apoptosis of human microvascular endothelium in vitro [ Escherichia coli immunization of rats led to induction of pauci-immune glomerulonephritis and detectable ANCA in one out of eight animals [In patients with pauci-immune glomerulonephritis, gp130 was detected in about 90% of patients in their active phase and frequently coappeared with myeloperoxidase and proteinase 3 . Gp 130 in vitro . Howeverin vitro . Nonethe animals  and anot animals   is a small vessel vasculitis with predominantly deposition of IgA1 within the capillaries, venules, or arterioles of affected tissues. The disease affects the skin and the gastrointestinal tract in a majority of patients and glomerulonephritis, indistinguishable from IgA nephropathy on kidney biopsy, is present  in appro Staphylococcus aureus has been proposed in IgA vasculitis when it was shown that the percentage of T-cell receptor- (TCR-) Vβ-(5.2 + 5.3) and Vβ 8-positive cells, members of the T-cell repertoire with marked expression after stimulation with staphylococcal enterotoxin, was significantly increased in patients with active disease when compared to healthy individuals whereas specific TCR-Vβ was not present when infection was controlled [Evidence for a potential microbial influence on the pathogenesis stems from the observation that onset of vasculitis is frequently preceded by an upper respiratory tract infection. It was proposed that the tonsils play an especially important role and chronic infections of the oropharynx and the middle ear can be detected in a majority of children with IgA vasculitis with apical periodontitis, rhinosinusitis, and otitis media being present in 53%, 48%, and 10%, respectively. In one report eradication of the infectious focus  resulted in a permanent remission without recurrence of disease in 31 out of 40 patients . Nephritntrolled . Helicobacter pylori antigen in 100% of samples investigated and the positive staining was restricted to glomerular structures [ Haemophilus parainfluenzae antigen was detected in 35% of kidney biopsy samples but only in 4% in other kidney diseases. Moreover, significantly higher plasma IgA1 antibody levels against Haemophilus parainfluenzae were present in IgA vasculitis than in patients with other renal diseases [Additional microbes have been alleged to contribute to the pathogenesis of IgA vasculitis. Immunohistochemistry of kidney specimen revealed presence ofructures . Mesangidiseases   shortly after an upper respiratory tract or gastrointestinal infection, whereas in contrast adult patients commonly present with microscopic hematuria, proteinuria, and hypertension. Microbes have been implicated in etiopathogenesis for a long time as it was assumed that an infection may facilitate synthesis of anti-glycan antibodies cross-reacting with galactose-deficient IgA1. For the nephritogenicity of galactose-deficient IgA1 formation of immune complexes is critical .In line with this idea, a pivotal role of the tonsils has been proposed. Bacterial strains in patients with chronic tonsillitis and IgA nephropathy showed a similar distribution and alpha streptococci were most abundantly present in both groups . When co Haemophilus segnis and Campylobacter rectus in about half of the cases examined, significantly more frequent when compared to controls with chronic tonsillitis, while Treponema sp. could be detected in 24% of patients with IgA nephropathy and in 7% of controls (no significant difference). Remission of proteinuria in IgA nephropathy subjects was achieved more often when Campylobacter rectus and Treponema sp. were present [ Helicobacter pylori in their palatine tonsils and half of them had coexistence with Actinomyces israelii even though the latter finding did not differ for patients with recurrent pharyngotonsillitis only [ Helicobacter pylori in all samples and cytotoxin-associated antigen A (CagA) was produced by a majority of the strains [ Helicobacter pylori in palatine glands in patients with IgA nephropathy compared to those with recurrent pharyngotonsillitis only [Analysis of tonsil specimens obtained from 68 patients with IgA nephropathy revealed presence of periodontal bacteria like present . Examinatis only . In anot strains . In bothtis only , 38. Haemophilus parainfluenzae antigens (OMHP) by sera obtained from patients with IgA nephropathy revealed homology with amino acid sequences from outer membrane protein P6 precursor, P5, and P2 porin protein of Haemophilus influenzae [Blood samples of 21 IgA nephropathy patients indicated significantly higher antibody titers to the IgA-binding region (BR) of streptococcal M proteins M4 and M60 when compared to healthy controls . Recognifluenzae .Analysis of 35 patients with IgA nephropathy showed presence of IgA antibodies against four of five investigated pneumococcal polysaccharides , the titers of which were significantly increased when compared to patients with other glomerulopathies, whereas IgG antibodies did not differ between both groups . Moreove Helicobacter pylori infection showed a greater rate of IgA anti-Helicobacter pylori seropositivity and a more pronounced IgA and IgG anti-Helicobacter pylori response compared to patients without renal disease, the latter predominantly existing of polymeric IgA1, IgG2, and IgG3 in the diseased group [Moreover, patients with IgA nephropathy anded group . Haemophilus parainfluenzae significantly correlated with the degree of glomerular injury especially in patients with episodes of macroscopic hematuria [As already pointed out earlier significantly higher levels of IgA and IgG antibodies directed against OMHP have been observed in patients with IgA nephropathy compared to patients with other glomerular diseases  and the ematuria . Haemophilus influenzae could be demonstrated in three out of 5 kidney biopsies but the same phenomenon was also observed in two out of 6 non-IgA nephropathy renal specimens [ Escherichia coli and Haemophilus influenzae revealed significantly higher values against both bacteria in IgA nephropathy which correlated with total serum IgA and IgM [Reactivity of IgA eluted from kidney tissues againstpecimens . Measure and IgM . Staphylococcus aureus were also increased in patients with IgA nephropathy compared to healthy controls. Moreover, the avidity of these antibodies against Staphylococcus aureus was significantly lower in patients compared to controls [ Staphylococcus aureus cell envelope antigen in the glomeruli of patients with IgA nephropathy and colocalisation thereof with glomerular deposits of IgA [IgA class antibodies againstcontrols . Interess of IgA .β in PBMC supernatants by 3-fold compared to controls. In addition, the proliferation capacity of lymphocytes was higher when compared to the one observed in patients with nonproliferative glomerulonephritis [ Helicobacter pylori-associated CagA. Stimulation of a B-cell line with CagA resulted in cell proliferation and dose-dependently increased production of IgA1 and CagA led to the stimulation of underglycosylated IgA1 in vitro [Streptococcal M5 isolated from peripheral blood mononuclear cells (PBMC) of IgA nephropathy patients increased surface IgA-positive B cells by 1.6-fold and induced transforming growth factor-ephritis  and tonsephritis . An immuin vitro . Haemophilus parainfluenzae antigens compared to lymphocytes from patients with chronic tonsillitis [ Haemophilus parainfluenzae antigens are capable of stimulating tonsillar T and B cells in patients with IgA nephropathy. Tonsillar mononuclear cells of patients with IgA nephropathy exhibited a higher capability to produce transforming growth factor-β and interleukin-10 along with total IgA after stimulation with OMHP antigens than those from patients with chronic tonsillitis in vitro [ Haemophilus parainfluenzae-specific IgA is produced by Haemophilus parainfluenzae and other bacteria and viruses possessing specific CpG motifs in tonsillar mononuclear cells [ Haemophilus parainfluenzae is enhancing the expression of TCR-Vβ-6 in vitro and further experiments indicated that the TCR-Vβ-6 was used more frequently in tonsillar T cells of patients with IgA nephropathy and the proportion of these cells in peripheral blood decreased significantly following tonsillectomy compared to controls, suggesting a selective expansion of T cells in IgA nephropathy [Lymphocytes obtained from palatine tonsils of patients with IgA nephropathy showed a significantly higher stimulation index to along with a higher level of IgA and IgA1 antibodies againstsillitis . These rsillitis , suggestin vitro . Moreovear cells . Stimulahropathy . Haemophilus parainfluenzae indicated a direct nephritogenic effect. Mice either ingesting orally or receiving intraperitoneally OMHP antigens develop glomerular IgA deposition and mesangial expansion, similar to the findings in IgA nephropathy [Administration of specific pneumococcal C-polysaccharide (PnC) led to deposition of IgA-PnC within the GBM and nuclear factor-kappa B transcription factor was activated early and progressively increased in response to glomerular IgA-PnC deposits in a mouse model . Moreovehropathy . Staphylococcus aureus antigens [ Staphylococcus aureus was also capable of inducing an IgA nephropathy like phenotype. Furthermore, anti-20-peptide antibodies labelled glomeruli of patients with IgA nephropathy [Induction of glomerulonephritis resembling an IgA nephropathy phenotype was observed in Balb/c mice after immunization withantigens . Immunizhropathy  , and minimal change disease (MCD). In idiopathic membranous nephropathy the target antigen has been identified with PLA2 receptor antibody positivity in approximately 70% of patients , with th Helicobacter pylori as detected by a high-molecular-weight cell-associated protein test was found to be significantly higher (66%) when compared to an age-matched control group without history of kidney disease (44%). When eradication was achieved in four patients, remission was reported in three of them. However, the disease course may have been influenced by concomitant glucocorticoid treatment as well [ Helicobacter pylori antigen in about two-thirds of the patients with membranous nephropathy [In membranous nephropathy, the infection rate of as well . Examinahropathy , a numbehropathy .Conflicting results regarding the frequency of urinary tract infections preceding onset of nephrotic syndrome in children have been reported. While in an Indian cohort urinary tract infections were common in patients with non-steroid responsive nephrotic syndrome or during relapse with approximately 40% , only on Helicobacter pylori antigen deposition in renal biopsy specimens and evidence of infection in blood [Microbial antigens may play a role in membranous nephropathy as was shown by an abundance ofin blood , 68. Furin blood . Costimuin blood . Thus, mSystemic lupus erythematosus is a remarkable complex autoimmune disorder with considerable heterogeneity in the onset of symptoms, the presentation of organ involvement, and therapeutic response towards immunomodulatory as well as immunosuppressive medication. Renal involvement, present in about 50% of patients within the first year after diagnosis, is one of the most important hallmarks necessitating more intensive immunosuppression. Antibodies directed against nuclear antigens play a pivotal role in the development and monitoring of the disease . Helicobacter pylori in kidney biopsy specimens from lupus nephritis patients (classes I–V) was observed in 18 out of 27 patients in one study [Presence ofne study . However in vitro experiments. Cross-reactivity of anti-pneumococcal antibodies obtained from a patient after vaccination has been shown with foreign- and self-antigens. Moreover, these antibodies exerted the capability to bind anti-double-stranded DNA antibodies, indicating a potential role of molecular mimicry [ Escherichia coli DNA and CpG-oligodeoxynucleotides increased DNA antibodies in lupus mice, accompanied by progression of mild to crescentic glomerulonephritis, interstitial fibrosis, and heavy proteinuria [ Vibrio cholerae, promoted autoantibody production and onset of glomerulonephritis in lupus prone mice [More evidence for an impact of microbes in SLE largely derives from animal models and mimicry . Further mimicry . Effectsteinuria . Choleraone mice Several investigations in mice highlighted that injection of bacterial lipopolysaccharide induced anti-double-stranded DNA antibodies –81. More Helicobacter pylori antigen could be detected in two-thirds of the kidney biopsies examined [In systemic lupus erythematosus, at least mouse models indicate a role of bacteria in the development and progression of nephritis, associated with an increase in immune complexes, polyclonal B-cell activation, increasing proteinuria, and irreversible kidney damage. Furthermore,examined . However Staphylococcus aureus exerts a direct pathogenic role in nasal epithelia in GPA. Other bacteria may not be directly related to the development of autoimmune kidney disorders, but molecular mimicry or deposition of immune complexes secondary to infections may contribute to renal involvement or damage.Bacterial antigens are implicated in the onset and in the progression of many autoimmune kidney disorders. There is evidence that Haemophilus parainfluenzae in IgA nephropathy. Nevertheless, both proposed implications rely on robust data. Moreover, some of the investigations have focused on detection of antigens in kidney biopsies or eluting antigens from kidney biopsy samples. There is a strong need to corroborate such associative findings in more convincing pathomechanistic experimental models. The lack of proof of causality is also a problem with studies in which an abundance of bacterial strains was detected in tonsils of patients with IgA nephropathy, as no satisfying effect of tonsillectomy to induce clinical remission was demonstrated in a recent multicenter (AE) trial even though the operation was able to mitigate proteinuria [However there are several limitations when assessing data about the role of microbial agents in autoimmune kidney disorders. Some of the most interesting findings have been proposed by single research groups, like the findings of hLAMP-2 in pauci-immune crescentic glomerulonephritis or the role ofteinuria .More investigations with novel methods  will yield a way to better define the role of bacteria in these diseases. Several sites of interest such as the sinuses in GPA, the tonsils and the intestine in IgA vasculitis, and IgA nephropathy may show significant alterations in bacterial colonization diversity, which in turn may contribute to an immunologic imbalance  leading to local inflammation and provoking the onset or the recurrence of disease. Since studies of the human microbiome already offered insights into several other autoimmune disorders, we are convinced that more profound analyses with longitudinal sample collection  will also clarify at least in part etiopathogenesis of some autoimmune kidney disorders. Furthermore, results obtained from microbiome analysis may enable us to prescribe tailored therapeutic measures aiming to eliminate abundant strains, such as trimethoprim-sulfamethoxazole treatment in limited GPA or eradication of infectious foci in IgA vasculitis, potentially restoring microbial and epithelial barrier imbalance.


Clostridium difficile enterocolitis, especially in children. We review the 6 pediatric cases published in the English and non-English literature and discuss their clinical presentation, outcome, treatment, and pathophysiology. We also report the seventh case of Clostridium difficile reactive arthritis in a 6-year-old boy who was treated with amoxicillin-clavulanate for 10 days because of an upper respiratory infection. After the antibiotic course, the child developed at the same time diarrhea with positive stool culture for Clostridium difficile and an asymmetric polyarthritis. Nonsteroidal anti-inflammatory drugs and metronidazole completely resolved the pain, joint swelling, and diarrhea. After twelve months of follow-up there has been no recurrence. This report confirms the self-limiting course of Clostridium difficile reactive arthritis. Clostridium difficile testing in children with gastrointestinal symptoms and acute onset of joint pain should be always considered.Reactive arthritis is a rare complication of Salmonella, Shigella, Yersinia, and Campylobacter. It is often associated with the human leucocyte antigen- (HLA-) B27 and usually considered in adult classification as a subtype of spondyloarthropathies. The criteria currently used to diagnose ReA are those of the 1995 Berlin Third International Workshop: (1) the presence of a predominantly lower limb asymmetrical oligoarthritis and (2) clinical laboratory evidence of a preceding infection [Reactive arthritis (ReA) is an aseptic acute inflammatory arthritis which develops after an extra-articular infection triggered by arthritogenic bacteria, more frequently from Clostridium difficile (CD) colitis. Although Clostridium difficile ReA is largely described in adults, there are few pediatric case reports probably due to CD asymptomatic colonization in newborns and infants. However, the incidence of CD among hospitalized children has increased in the last few years [A particular type of ReA has been described in connection withew years . To the ew years –7. We reA 6-year-old boy was admitted to our department because of pain in the right knee and both ankles started five days before admission. Three weeks earlier he had completed a 10-day course of oral amoxicillin-clavulanate. Off antibiotics, he developed watery diarrhea and abdominal pains.On admission the child presented with a profuse diarrhea; his left knee and both ankles were swollen, warm, and painful with significant limitation of motion. He could neither stand nor walk because of severe pain. All his other joints were normal. He had no fever. The remaining general examination was unremarkable; in particular he had no rash, mucositis, pharyngitis, urethritis, or conjunctivitis. Initial investigations showed a normal white cells count with a mild increase of C-reactive protein  and of the erythrocyte sedimentation rate ; electrolytes were normal. Haemoglobin concentration, platelet count, and fibrinogen were all in the normal range, as antistreptolysin-O (ASO) level . Streptococcus, Borrelia, Cytomegalovirus, Epstein-Barr-Virus, andParvovirus B19) were also negative.Three days later the arthritis slowly resolved spontaneously in the left knee and ankles but appeared in the right shoulder and homolateral hip with a migratory pattern. This polyarthritis was accompanied, at that time, with unremitting and not elevated fever (38°C at the maximum). Ultrasonography showed a mild effusion in the right knee, ankles, and right hip joint. X-rays were normal. Both synovial fluid and blood cultures were negative. Tests for immunoglobulin and complement were normal. Autoantibodies  were absent and HLA-B27 antigen was negative. Eyes did not show any sign of iridocyclitis on the slit lamp. A cardiac evaluation, including echocardiogram, ruled out any evidence of carditis. Serological tests for arthritis related infections  were negative but CD and CD toxin A and toxin B were detected by ELISA technique. Fecal calprotectin was not increased.Our patient's diarrhea, which started following antibiotics, gradually resolved during his hospital stay. Stool cultures for enteric pathogens  for three weeks and metronidazole therapy (7.5 mg/kg every eight hours) for ten days.Over that period, diarrhea and the polyarthritis rapidly improved. After three weeks of naproxen therapy his symptoms completely resolved. Joint pain abated, the mobility recuperated, and no other joint involvement was noted. Inflammatory parameters turn out to be negative, within seven days after starting treatment. Stool culture after 4 weeks was negative. Fecal calprotectin remained negative.Our little patient was clinically followed up for 12 months. By then he had completely recovered.ReA is a nonseptic acute inflammatory arthritis occurring after an intercurrent infection in which the causative organism is outside from the joint. The most frequent causes of ReA are intestinal and genital bacterial infections. ReA is typically characterized by asymmetrical oligo or polyarthritis predominantly in the lower limbs large joints  , 9. SympSeveral patients present a migratory pattern of polyarticular arthritis, but a few have monoarticular or oligoarticular arthritis. Acute arthritis is characterized by severe pain and sometimes erythema over the affected joints but some children may show only light or moderate joint pain and swelling. Enthesitis or tenosynovitis may occur alone or with arthritis. Extra-articular symptoms included systemic signs such as fever, weight loss, fatigue, and skin, eyes, and mucous membranes disease  . There iSalmonella, Shigella, Yersinia enterocolitica, and Campylobacter jejuni), pharyngitis caused by group A Streptococcus, or more rarely a genitourinary tract infection (Chlamydia trachomatis).ReA can occur at any age, most frequently in young children after enteric infections .No reasonable alternative diagnosis for arthritis or diarrhea .In order to perform a diagnosis of ReA associated with CD enterocolitis, a history of diarrhea after a course of antibiotic or the exclusion of other causes of gastroenteritis or noninfectious arthritis is essential.CD infections are generally more severe in certain populations, including patients receiving chemotherapy or with chronic gastrointestinal diseases . AccordiOur case fulfills the above criteria by Putterman and Rubinow . MicrobiIn addition, antibiotic therapy was previously administered in our patient. There was no evidence of antecedent group A streptococcal infection (ASO title was not raised) and our case did not meet the modified Jones criteria, therefore excluding the diagnosis of acute rheumatic fever or poststreptococcal reactive arthritis. An inflammatory bowel disease (IBD) was considered as it is frequently associated with CD colonization but the dosage of fecal calprotectin, which has a high negative predictive value for IBD, was negative . MoreoveClostridium difficile”, “arthritis”, and “child” to identify all papers reporting CD enterocolitis-associated reactive arthritis in childhood was performed. In addition, the search was extended to all relevant articles in the reference lists. No year or language restriction was applied to our search. Two reviewers performed a first stage selection of the total identified studies based on the following inclusion and exclusion criteria (using the abstract or the full text article when required).A literature search of MEDLINE, Google Scholar, and Cochrane Reviews computerized databases using the keywords “All published papers were included in the present review only if they met the following inclusion criteria: (1) appearance of acute arthritis and/or tenosynovitis between 4 weeks before and 12 weeks after the onset of diarrhea and/or colitis; (2) diarrhea appearing some time after a course of systemic antimicrobial therapy; (3) microbiologic proof of CD involvement (either positive stool culture or assay for toxin); (4) no reasonable alternative diagnosis for arthritis or diarrhea ; (5) negative synovial fluid cultures (if obtained); and (6) age of patient between 1 and 18 years.Six reports 5 papers) –7 met ou papers –In agreement with the other reports, our case confirms the good outcome of ReA following CD infection with a self-limiting course (3 weeks). Only one case documented a child who developed an acute abdomen and underwent a laparotomy for presumed peritonitis .The treatment of ReA following CD infection included the discontinuation of antimicrobial agents and the use of oral NSAIDs coupled with metronidazole therapy for 7–10 days in children and adolescents with mild-to-moderate colitis or vancomycin in more severe diseases. All of previous cases were treated with vancomycin or metronidazole; only the case described by Finger and Neubauer  was resoThe CD infection should be suspected in children presenting with an acute inflammatory arthritis following an episode of diarrhea especially when stool culture for common enteric bacterial infection turns out to be negative and the patient has received antibiotic therapy before the onset of diarrhea. Therefore, we suggest performing tests for CD toxin in children with these characteristics because CD infection associated with ReA can be unrecognized.


Inflammatory and protein mediators  play an important, but still not completely understood, role in the morbidity and mortality of intra-abdominal sepsis/injury. We therefore systematically reviewed preclinical and clinical studies of mediators in intra-abdominal sepsis/injury in order to evaluate their ability to: (1) function as diagnostic/prognostic biomarkers; (2) serve as therapeutic targets; and (3) illuminate the pathogenesis mechanisms of sepsis or injury-related organ dysfunction.We searched MEDLINE, PubMed, EMBASE and the Cochrane Library. Two investigators independently reviewed all identified abstracts and selected articles for full-text review. We included original studies assessing mediators in intra-abdominal sepsis/injury.Among 2437 citations, we selected 182 studies in the scoping review, including 79 preclinical and 103 clinical studies. Serum procalcitonin and C-reactive protein appear to be useful to rule out infection or monitor therapy; however, the diagnostic and prognostic value of mediators for complications/outcomes of sepsis or injury remains to be established. Peritoneal mediator levels are substantially higher than systemic levels after intra-abdominal infection/trauma. Common limitations of current studies included small sample sizes and lack of uniformity in study design and outcome measures. To date, targeted therapies against mediators remain experimental.Whereas preclinical data suggests mediators play a critical role in intra-abdominal sepsis or injury, there is no consensus on the clinical use of mediators in diagnosing or managing intra-abdominal sepsis or injury. Measurement of peritoneal mediators should be further investigated as a more sensitive determinant of intra-abdominal inflammatory response. High-quality clinical trials are needed to better understand the role of inflammatory mediators.The online version of this article (doi:10.1186/s13054-015-1093-4) contains supplementary material, which is available to authorized users. Sepsis is a syndrome that results from the interactions between the host and insults  leading to the production/release of biochemical mediators and the triggering of inflammatory cascades . Tissue Intra-abdominal infection and injury are common causes of sepsis in surgical intensive care unit (ICU) patients, with high rates of mortality –9. EarlyGiven that both intra-abdominal infection and injury can incite hyperinflammatory status and sepsis, and the inflammatory response to injury is often clinically indistinguishable from sepsis, many studies have sought to identify biomarkers or mediators to aid in the diagnosis and management of sepsis , 12. TheWe developed a protocol according to the Arksey and O’Malley methodological framework for conducting scoping reviews with modifications . ScopingTwo investigators  created a preliminary search strategy that was subsequently refined by a medical librarian with extensive systematic review experience (HLR). Relevant articles were identified by searching the following databases from the first date available until August 2014: Ovid MEDLINE; Ovid EMBASE; PubMed; Cochrane Database of Systematic Reviews, and Cochrane Central Register of Controlled Trials. Combinations of the following search terms were used  independently screened the titles and abstracts of all identified citations. We included all articles that were original studies  assessing inflammatory or protein mediators in intra-abdominal sepsis or injury. Participants included patients (10 or more) admitted for surgery with peritonitis or abdominal injury, or animal models of abdominal sepsis/injury. Participants required measurement of mediator(s) from blood and/or peritoneal fluid for inclusion. Articles were excluded if not reporting original data, such as reviews, letters, and conference abstracts (no full text available). Disagreements between the two assessors were resolved by consensus.The full texts of the remaining articles were independently reviewed by the same two authors. The two investigators extracted data independently using a predesigned electronic data extraction form. Assessors were not blinded to author or journal name . We extrResults were first mapped based on their characteristics, such as study design, sample size, preclinical or clinical studies. Owing to the significant variability in study design, sample size, analytical technique and outcome measures, it was impossible to perform a full synthesis analysis of the findings. Therefore, studies were presented in tables based on thematic characteristics of the articles. In addition, the level of evidence for a mediator as a biomarker was discussed based on the “best-evidence synthesis” guideline used in review studies  and the search from reference lists identified an additional 25 articles. The review of abstracts led to retrieval of 333 full-text articles for assessment. We selected 182 articles for inclusion of this review  explored PCT as an indicator to diagnose infection, predict outcomes, or guide treatment of abdominal sepsis or injury , 101. Sel sepsis . The resl sepsis . Similar surgery . Four st surgery , 93, 95. surgery , 78. Imp surgery , 101.Fifty-five of 86 studies examined IL-6, alone, or in conjunction with other cytokines . IL-6 levels in plasma are rapidly dynamic and peaking at maximum from wound closure to POD1. IL-6 generally declined on POD1, and was back to baseline on POD3 , 79, 90.. demonstrated high mobility group box protein 1 (HMGB1) was elevated in plasma early after injury and shock. Levels of HMGB1 correlated with injury severity and morbidity in trauma patients [. in surgical patients [Recent studies have reported that DAMPs can act as proinflammatory mediators. Cohen et alpatients . Similarpatients . HMGB1 lpatients , 99. Mitpatients .All studies that examined both systemic (blood) and peritoneal levels of mediators demonstrated that peritoneal mediator levels were 10–1000 times higher than systemic values in patients with peritonitis , 89, 102. carried out a small case-control study, and indicated a significant increase in blood IL-6 after reperfusion in aortic abdominal aneurysm surgery patients, which was followed by a significant reduction of lung function [. reported blood IL-6 levels were statistically higher in males than females after injury [Escherichia coli bacteria was significantly depressed when compared to patients without septic shock [Table S4 in Additional file function . Sperry r injury . In patiic shock . Moreoveic shock .. compared surgical to endoscopic necrosectomy for the treatment of infected necrotizing pancreatitis [Table S5 in Additional file reatitis . From threatitis .. performed a RCT looking at mediator levels in 45 patients with intra-abdominal sepsis or injury who needed open abdomen management, to determine whether active negative pressure peritoneal therapy with the ABThera device reduces systemic or peritoneal inflammation [P = 0.04) was observed among patients randomized to the ABThera (21.7 %) versus Barker’s vacuum pack (50 %). Interestingly, there was no significant difference in clearance of the peritoneal or plasma IL-6 (or a number of other cytokines) between 24 and 48 hours among patients randomized to the ABThera versus Barker’s vacuum pack.Kirkpatrick et alammation . In thisTable S6 in Additional file Studies also demonstrated that local  inflammatory mediator levels were not accurately reflected by systemic (blood) mediator levels. Generally, peritoneal mediator levels were much higher than the systemic levels, and thus might be a truer or better indicator of the local inflammatory response and outcomes , 163. ThKnockout animal studies demonstrated IL-10 was a potent anti-inflammatory mediator, resulting in the attenuation of lung neutrophil infiltration and injury after visceral ischemia . ICAM-1-. reported that systemic and peritoneal inflammation was reduced with less organ damage in pigs with peritonitis treated by negative pressure therapy [Table S7 in Additional file  therapy . AlthougThis scoping review focused on examining the reported role of inflammatory and protein mediators in intra-abdominal sepsis or injuries. In surgical ICU patients, traumatic injuries and septic conditions can be simultaneously or subsequently present. Each can be associated with profound and dynamic production of bioactive mediators that are at the present poorly understood, particularly in terms of their kinetics and overall interaction with the host inflammatory response. This inflammatory response is a complex and multifaceted process. Traumatic injuries induce overwhelming reactions in the immunological and neurohormonal systems. Innate immunocytes are activated by hypoxia stress and endogenous signals (DAMPs) released by damaged tissues , 6, 203.Although animal studies suggest that mediators play a critical role in both the pathogenesis and potential management in intra-abdominal sepsis/injury, the clinical evidence to support measuring mediators as biomarkers to discriminate SIRS from sepsis is conflicting.Due to the heterogeneity of included studies, we discussed the overall results based on the “best-evidence synthesis” rating, which is widely used in narrative and systematic reviews . ModeratFurthermore, although the serum PCT value appears to predict severity of septic complications , 64, it Sepsis further increased IL-6 levels over the already elevated levels from the original injury. Although moderate level of evidence supports IL-6 to be a useful indicator for sepsis or severity , 87, 91,It appeared to be futile to specifically target a single inflammatory or protein mediator in an effort to mitigate disease and improve outcomes. It is more likely that identifying promising biomarkers will only be possible when a number of mediators are analyzed as a profile , 205.It is currently unknown if the peritoneal cavity functions to restrain the inflammatory mediators or to act as a reservoir of mediators. Evidence has shown that peritoneal fluid functions as a priming and activating stimulus for neutrophils both in the peritoneum and in remote organs  after injury, enhancing the incidence of MOF upon subsequent infection , 163. MoFinally, based on the evidence from animal models that injury followed by a second insult  substantially enhanced the inflammatory response and organ damage , 190, itThis review has limitations. First, despite the search of multiple databases using comprehensive search strategies with the assistance of a medical librarian, our search has excluded studies assessing extra-abdominal sepsis or injury. Moreover, limited abdominal trauma studies were available; the majority of which focused on surgical injuries. Importantly, the exclusion of studies assessing other kinds of sepsis can present a selection bias. Second, due to the inconsistencies in study design, it proved difficult to extract accurate data from all studies, even with the assistance of predefined data abstraction tools. Furthermore, only a small number of the included studies actually examined biomarkers for both sensitivity and specificity.Systemic inflammatory response syndrome is the expected immunologic response to both infection and traumatic injuries, which makes the distinction between infectious sepsis and sterile SIRS exceedingly difficult. Although animal studies suggest that mediators play an important role, the clinical use of mediators as biomarkers in discriminating SIRS from sepsis in intra-abdominal injury or sepsis is conflicting. Persistent elevation of CRP or PCT levels in blood appears to be correlated with septic complications. The role of IL-6 as a biomarker in intra-abdominal sepsis is controversial. Limited evidence suggests endogenous molecules may be candidate biomarkers of sepsis or injury. Importantly, peritoneal mediator levels cannot be proportionately reflected by the systemic levels. Victims subjected to trauma followed by infection show more excessive inflammatory response and organ damage than those to trauma only. To date, therapies targeting a specific inflammatory or protein mediator in abdominal sepsis have shown poor results. Interventions aiming to remove peritoneal fluid/mediators and improve outcomes are still investigational. Large sample size randomized controlled trails are desirable to identify the “ideal” biomarker(s) and understand the roles of mediators in intra-abdominal sepsis and injury.Preclinical studies demonstrate that intra-abdominal injury/ischemia, especially with subsequent infection, induces an excessive inflammatory/protein mediator production and uncontrolled inflammation in the peritoneum and remote organs.Both preclinical and clinical studies have shown the peritoneal mediator levels are much higher than the blood levels, and the release of these mediators (mostly via the lymphatic pathway) into the systemic circulation may precipitate the deleterious effects of sepsis and multiple organ dysfunction.Normal postoperative clearance kinetics of CRP, PCT, or IL-6 may help rule out infection or complications.The clinical role of measuring inflammatory/protein mediators as biomarkers to diagnose abdominal infection or predict outcomes of abdominal sepsis or injury remains to be established. High-quality clinical trials are warranted to determine the role of mediators in intra-abdominal sepsis and injury.


Rangifer tarandus). In central Canada, the ranges of two caribou subspecies (barren-ground/woodland caribou) and two woodland caribou ecotypes  overlap. Our objectives were to reconstruct the evolutionary history of the eastern migratory ecotype and to assess the potential role of introgression in ecotype evolution. STRUCTURE analyses identified five higher order groups . The evolutionary history of the eastern migratory ecotype was best explained by an early genetic introgression from barren-ground into a woodland caribou lineage during the Late Pleistocene and subsequent divergence of the eastern migratory ecotype during the Holocene. These results are consistent with the retreat of the Laurentide ice sheet and the colonization of the Hudson Bay coastal areas subsequent to the establishment of forest tundra vegetation approximately 7000 years ago. This historical reconstruction of the eastern migratory ecotype further supports its current classification as a conservation unit, specifically a Designatable Unit, under Canada’s Species at Risk Act. These findings have implications for other sub-specific contact zones for caribou and other North American species in conservation unit delineation.Understanding the evolutionary history of contemporary animal groups is essential for conservation and management of endangered species like caribou ( Similarly, the two sub-specific lineages . The assessment of the goodness-of-fit of a model parameter posterior combination was tested via the model-checking option in DIYABC v. 2.0.4 [a–c).We ran approximately 3 million simulations and compared scenarios by estimating posterior probabilities using the logistic regression . For modv. 2.0.4 . Runs we3.3.1In total, 1360 unique genotypes  were identified. Samples with at least eight out of 10 successfully scored loci were used in subsequent analysis (missing data in the microsatellite dataset was 2.9%). However, one locus (BMS1788) was taken out of further analysis because it showed mutational patterns that are inconsistent with 1 bp shifts.icro-Checker tests nor tests for deviations from HWE and LD revealed any significant evidence that specific populations showed null alleles, deviated from HWE (9/333 cases significant after Bonferroni correction), or showed signs of LD (none significant after Bonferroni correction). Therefore, we retained nine loci for the subsequent analyses to examine phylogeographical structuring rather than delineate fine-scale local structure that might also exist but is beyond the scope of this work.Neither MK=2, corresponded to the subspecies level distinguishing barren-ground from all other populations . The second level, at K=6 , revealed population genetic structure at the ecotype and within-ecotype level, identifying barren-ground, the eastern migratory ecotype, including the Pen Island herd and coastal region in Ontario, as structured groups. Further, the boreal caribou ecotype consisted of four populations : (1) and (2) two populations in western Manitoba, (3) one population in southwestern Manitoba and (4) one population including eastern Manitoba and Ontario mainland. For further analysis, the two populations found in western Manitoba were combined because only a single sampling site with fairly low sampling size (N=23) showed high membership coefficients for that cluster with additional individuals being highly admixed. It is currently unknown whether that cluster is biologically meaningful and the pattern requires further investigation with more local sampling. Since the focus here was to understand the evolutionary origin of the eastern migratory ecotype, western Manitoba was considered one population with potential substructure. All subsequent analyses were carried out with the five major groups identified in STRUCTURE. Further, a population mean q bar plot  revealed that individuals of the eastern migratory ecotype had higher average membership assignments to their focal population. Likewise, Ontario/eastern Manitoba showed a different genetic signature including higher average membership coefficients to a different population cluster. Hence, the eastern migratory ecotype and the Ontario/eastern Manitoba population were treated as different populations in succeeding analyses. Additionally, a sorted q bar plot  confirmed the presence of highly assigned individuals to each of the identified populations indicating that, albeit high admixture levels in Ontario, two distinct clusters exist there (i.e. eastern migratory ecotype and Ontario/eastern Manitoba)  .Figure Lastly, the phylogeographical pattern in BMS1788 is consistent with results in that a preliminary assessment of allele variation supports a phylogeographical pattern separating western Manitoba and Ontario. An AMOVA on microsatellite data revealed lower differentiation values that nevertheless were highly significant; therefore, supporting the results found in the STRUCTURE analysis.Since barren-ground (B haplotypes) and woodland caribou (A haplotypes) clearly differed in mtDNA control region sequences ,12,13, iA hierarchical AMOVA (mtDNA) revealed that approximately 22% of the genetic variation was found between the STRUCTURE groups indicating clear genetic structure  and 3. Wa–c). As would be expected for the rates of change at each type of loci and the time frame under investigation, these results suggest that mtDNA is more reliable in estimating the more ancient admixture event, whereas the microsatellite data are superior in estimating the more recent divergence event . Since mtDNA has a lower mutation rate than microsatellites, haplotype resolution may simply be insufficient to fully capture the divergence of eastern migratory and Ontario/eastern Manitoba boreal caribou, and therefore microsatellite data are likely more suitable for the analysis of the more recent divergence event. Cornuet et al. [ABC  identified scenario 3 as the most likely evolutionary scenario based on posterior probability values, credible intervals  and logit et al.  pointed 4.Increased understanding of the evolutionary processes that lead to contemporary spatial genetic partitioning is critical for delineating units for conservation assessment and subsequent recovery plans and may enhance our ability to protect important adaptive potential of species at risk ,65,66. IUnder this scenario, barren-ground caribou likely migrated along the retreating ice sheet allowing for lineage mixing in northern Ontario and Manitoba. The eastern migratory ecotype diverged from this ancient introgressed population, leaving residual barren-ground-specific haplotypes in both the eastern migratory ecotype and Ontario/eastern Manitoba boreal caribou population. The lack of B haplotypes in western and southwestern Manitoba might be best explained by later arriving western (i.e. A2 lineage) caribou populations that migrated eastwards and consequently did not readily interbreed with barren-ground. The occurrence of B haplotypes in eastern migratory and the boreal ecotypes in Ontario and eastern Manitoba is possibly the result of the ancient introgression event identified in the ABC analysis. Alternatively, gene flow between the eastern migratory ecotype and boreal caribou populations led to distributions of a few B haplotypes.In western Canada, several studies ,13 detecet al. [McDevitt et al.  also suget al. ) may be Migratory behaviour has been shown to increase fitness through exposure to seasonal changes in predator avoidance ,72 and rRegardless of the source of migratory behaviour, lineage-specific diversity is important, both biologically and for conservation ranking because genetically, different populations in widespread species often follow independent evolutionary trajectories and therefore may respond differently to new or changing environments ,75. The The maintenance of significant phylogeographical structure found in this study also points to the potential importance of additional environmental factors and landscape features  in shaping of genetic diversity. Since environmental factors are likely to be diverse across the distribution range of widespread species, locally adapted populations will be spatially distributed as well. It has been proposed  that pro


Speech perception requires that sounds be transformed into speech-related objects with lexical and semantic meaning. It is unclear at what level in the auditory pathways this transformation emerges. Primary auditory cortex has been implicated in both representation of acoustic sound attributes and sound objects. While non-primary auditory cortex located on the posterolateral superior temporal gyrus (PLST) is clearly involved in acoustic-to-phonetic pre-lexical representations, it is unclear what role this region plays in auditory object formation. Additional data support the importance of prefrontal cortex in the formation of auditory objects, while other data would implicate this region in auditory object selection. To help clarify the respective roles of auditory and auditory-related cortex in the formation and selection of auditory objects, we examined high gamma activity simultaneously recorded directly from Heschl's gyrus (HG), PLST and prefrontal cortex, while subjects performed auditory semantic detection tasks. Subjects were patients undergoing evaluation for treatment of medically intractable epilepsy. We found that activity in posteromedial HG and early activity on PLST was robust to sound stimuli regardless of their context, and minimally modulated by tasks. Later activity on PLST could be strongly modulated by semantic context, but not by behavioral performance. Activity within prefrontal cortex also was related to semantic context, and did co-vary with behavior. We propose that activity in posteromedial HG and early activity on PLST primarily reflect the representation of spectrotemporal sound attributes. Later activity on PLST represents a pre-lexical processing stage and is an intermediate step in the formation of word objects. Activity in prefrontal cortex appears directly involved in word object selection. The roles of other auditory and auditory-related cortical areas in the formation of word objects remain to be explored. Speech perception requires that incoming sounds be transformed into word objects. It is unclear at what level in the auditory pathways this transformation occurs. Some data suggest that primary auditory cortex principally represents acoustic sound attributes   generated during target detection tasks using both speech and non-speech stimuli. High gamma activity has been shown to be a sensitive and specific indicator of auditory cortical activation and has been successfully used to define organizational features of human auditory cortex  monitoring to identify potentially resectable seizure foci. The subjects were 38 (L258), 30 (L275), and 40 (L282) years old. All subjects were male, right-handed and left hemisphere language-dominant, as determined by intracarotid amytal (Wada) test results. Recordings were obtained from the left hemisphere in all three subjects. Research protocols were approved by the University of Iowa Institutional Review Board and the National Institutes of Health. Written informed consent was obtained from all subjects. Research participation did not interfere with acquisition of clinically required data, and subjects could rescind consent at any time without interrupting their clinical evaluation.All subjects underwent audiometric evaluation before the study, and none was found to have hearing deficits that should impact the findings presented in this study. Subjects L258 and L282 were native English speakers, and subject L275 was a native Bosnian speaker who learned German at the age of 10 and English at the age of 17. Neuropsychological testing of L258 was normal except for mild deficiencies in verbal working memory. Subject L275 had grossly intact conversational language comprehension, though formal neuropsychological testing showed non-localizing cognitive function deficits. Subject L282 had 13 years earlier undergone anterior temporal lobectomy that spared auditory cortex on the superior temporal gyrus. This subject was found to have mild deficits in verbal memory, fluency and naming. However, all three subjects had comparable performance in all experimental tasks both in terms of target detection accuracy and reaction times. This indicates that their performance of the tasks was not limited by any cognitive deficits identified during formal neuropsychological testing. Intracranial recordings revealed that auditory cortical areas were not epileptic foci in any subject.Experiments were carried out in a dedicated electrically-shielded suite in The University of Iowa General Clinical Research Center. The room was quiet, with lights dimmed. Subjects were awake and reclining in an armchair.http://librivox.org/) databases. Non-word syllables were excised from words using SoundForge 4.5 . A total of 20 unique exemplars of each syllable were used in each experiment: 14 spoken by different male and 6 by different female speakers. Additionally, the stimulus set included complex tones with fundamental frequencies of 125  and 250 Hz , approximating the average voice fundamental frequencies of male and female speakers, respectively. All stimuli were normalized to the same root-mean-square amplitude and edited to be 300 ms in duration using SoundForge with 5 ms rise-fall times. They were presented with an inter-stimulus interval chosen randomly within a Gaussian distribution  to reduce heterodyning in the recordings secondary to power line noise. Stimuli were delivered via insert earphones  that were integrated into custom-fit earmolds. Stimulus delivery was controlled using Presentation software .Experimental stimuli were consonant-vowel-consonant syllables [cat], [dog], [five], [ten], [red], [white], [res], and [tem] from TIMIT , speech stimuli spoken by female talkers, or words belonging to specific semantic categories such as animals or numbers. The subjects were instructed to use the index finger of their left hand  to push the response button whenever they heard a target sound. Prior to data collection, the subjects were presented with a random-sequence preview of stimuli to ensure that the sounds were presented at a comfortable level and that they understood the task requirements.ECoG recordings were simultaneously made from HG and lateral cortical surface using multicontact depth and subdural grid electrodes, respectively. Details of electrode implantation have been described previously, and more comprehensive details regarding recording, extraction and analysis of high gamma cortical activity are available for the interested reader  before electrode implantation to locate recording contacts. Two volumes were averaged to improve the signal-to-noise ratio of the MRI data sets and minimize the effects of movement artifact on image quality. Pre-implantation MRIs and post-implantation thin-sliced volumetric computerized tomography scans  were co-registered using a linear co-registration algorithm with six degrees of freedom . Collected ECoG data were amplified, filtered , digitized at a sampling rate of 2034.5 Hz, and stored for subsequent offline analysis. Behavioral responses to the target stimuli were recorded using a Microsoft SideWinder game controller. The timing of the button-press events was recorded and stored for analysis along with ECoG data.ECoG data obtained from each recording site were downsampled to a rate of 1000 Hz. To minimize contamination with power line noise, ECoG waveforms were de-noised using an adaptive notch filtering procedure  and target condition. Finally, p-values were corrected for multiple comparisons  using false discovery rate by controlling the false discovery rate following the method of Benjamini and Hochberg  and squared. The resultant high gamma power waveforms were smoothed using a moving average filter with a span of 25 ms, log-transformed, normalized to power in a pre-stimulus reference (250-50 ms prior to stimulus onset), and averaged across trials. To assess the presence and timing of task-related modulation of high gamma activity on representative cortical sites, single-trial high gamma ERBP was first averaged in 50 ms-wide consecutive windows to decrease the number of multiple comparisons. Next, for each window from 0–50 to 950–1000 ms, a two-sample one-tailed Hochberg  and BenjHochberg  with a tSD 25 Hz; female talkers: mean 202 Hz, SD 36 Hz). These differences represent a contribution in the high gamma responses of phase locking to the lower fundamental frequency of the male talkers within posteromedial HG [sites (a) through (d)]   relative to those with more prolonged VOTs . This effect was maximal in more central portions of HG compared to the observed effect of pitch on neural activity [sites (e), (f); Figure q < 0.01 at 750–800 ms after stimulus onset; middle and right columns of Figure Whereas activity along most of HG was strongly modulated by the acoustic attributes of the sounds, responses in the high gamma range were only weakly affected by the target detection tasks Figure . The lefA different pattern was observed within the most anterolateral portion of HG outside of presumed core cortex [site (h) in Figure More complex response profiles were observed on PLST Figures , 4 when p < 0.001, Mann-Whitney rank-sum tests).Responses to non-target words were also modulated by the specific task requirements. For instance, late high gamma activity to non-target words spoken by females was enhanced when the target detection tasks required words to be categorized relative to the task where complex tones were the targets . Therefore, it is reasonable to conclude that these differences between target and relevant non-target were not based solely on task difficulty. Importantly, increases in high gamma activity observed during either semantic categorization task began prior to the offset of the syllables, suggesting that these increases were not directly related to word classification, and likely were reflecting lower-level phonological processing, a prerequisite for semantic classification , and were marginally significant (q < 0.05) in the semantic categorization tasks on sites (d) and (e) (significance bars are not shown). Additionally, there was a trend for non-target words to elicit larger late responses during semantic categorization tasks compared to tone detection  did not significantly vary as a function of whether the subject responded rapidly or slowly or when the target was missed altogether Figure , where fAs expected from previous studies, activity within posteromedial HG was highly sensitive to the acoustic characteristics of speech , it was later than task-related modulations in all other regions studied. Thus, current findings do not support the premise that human primary auditory cortex is the location where auditory object formation occurs. In contrast, studies in primary auditory cortex of experimental animals have shown robust responses reflecting auditory object formation, task-related activity, and reward expectancy  studies , effects were not uniform and showed site-by-site variability. This variability may partly explain why task-related modulation on PLST was not seen in subject L258  that are likely important in long-range interactions between spatially disparate regions. As we continue investigation of these circuits, our conclusions will undoubtedly be refined and, hopefully, translationally relevant for the understanding of normal speech processing and its disfunction occurring in developmental language disorders, and acquired disorders such as stroke and normal aging.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Fathead minnow and zebrafish are among the most intensively studied fish species in environmental toxicogenomics. To aid the assessment and interpretation of subtle transcriptomic effects from treatment conditions of interest, better characterization and understanding are needed for natural variation in gene expression among fish individuals from lab cultures. Leveraging the transcriptomics data from a number of our toxicogenomics studies conducted over the years, we conducted a meta-analysis of nearly 600 microarrays generated from the ovary tissue of untreated, reproductively mature fathead minnow and zebrafish samples. As expected, there was considerable batch-to-batch transcriptomic variation; this “batch-effect” appeared to differentially impact subsets of fish transcriptomes in a nonsystematic way. Temporally more closely spaced batches tended to share a greater transcriptomic similarity among one another. The overall level of within-batch variation was quite low in fish ovary tissue, making it a suitable system for studying chemical stressors with subtle biological effects. The observed differences in the within-batch variability of gene expression, at the levels of both individual genes and pathways, were probably both technical and biological. This suggests that biological interpretation and prioritization of genes and pathways targeted by experimental conditions should take into account both their intrinsic variability and the size of induced transcriptional changes. There was significant conservation of both the genomes and transcriptomes between fathead minnow and zebrafish. The high degree of conservation offers promising opportunities in not only studying fish molecular responses to environmental stressors by a comparative biology approach, but also effective sharing of a large amount of existing public transcriptomics data for developing toxicogenomics applications. Among an organism’s responses to environmental perturbations, gene transcription could be regarded as one of the earliest molecular events along the genotype-phenotype continuum. Dynamic and transient in nature for most genes, variation in this transcriptional response and its complex regulatory mechanisms is believed to contribute to much of the phenotypic complexity across biota An organism’s transcriptome represents its full range of expressed gene transcripts regulated by a wide variety of molecular control mechanisms. At the molecular level, gene regulatory complexity is determined in part by the combinatorial nature of multiple cis-regulatory elements and trans-acting transcription factors It is, of course, also possible to introduce technical variation into transcriptomic data due to methodological differences across studies, for example, in RNA sample preparation, choice of profiling platforms, data processing approaches, and so on. Over the years, there have been extensive investigations into these technical issues and their impact on gene expression profiling by microarrays Among a wide range of applications of transcriptomics in various disciplines are human disease diagnostics/prognostics Danio rerio) arguably the most studied species in this diverse group. As a common biological model with extensive genome-level knowledge, the zebrafish is a logical species of choice for toxicogenomics work. The fathead minnow also is an attractive model species because it has been the dominant aquatic vertebrate test organism in regulatory toxicity testing for decades Pimephales promelas; PPR), based on samples across different gender-tissue-treatment conditions, variability among individuals was found to be very high, with a wide-ranging distribution among genes as well as molecular networks Fish in general are among the most commonly studied non-mammalian organisms in environmental toxicogenomics, with the fathead minnow and zebrafish  were exposed to a continuous flow of sand filtered, UV-sterilized, Lake Superior water  for 24, 48, or 96 h at the US Environmental Protection Agency (USEPA) laboratory in Duluth, Minnesota. At the end of each exposure period, fish were anesthetized in a buffered solution of tricaine methanesulfonate  and ovaries were collected and shipped overnight on dry ice to the USEPA laboratory in Cincinnati, Ohio. Total RNA isolated from selected tissue samples was then sent to Cogenics Corporation, an Agilent certified contract laboratory . Hybridization was conducted using a two-color protocol on Agilent zebrafish microarrays with 21K probes , followed by high-resolution scanning and image processing by Agilent Feature Extraction software. Eighty ovary controls for nine treatment conditions, all based on the design 015064, were included in this study.Reproductively mature fathead minnows (5–7 months old) reared at the US EPA laboratory in Duluth were exposed to LSW. All exposures were continuous flow-through exposures. Representative experimental designs for the experiments included as part of the current analysis have been detailed elsewhere www.r-project.org) primarily by using the Limma package Various analyses were conducted in the R environment  and variance partitioning http://www.is.titech.ac.jp/~shimo/prog/pvclust/).Between-batch variation was further evaluated by several approaches. To provide an empirical critical value to assess the statistical significance of DEG counts, a non-parametric distribution of the number of DEGs between batches was generated. Microarray sample labels were permuted according to the original number of replicates in each batch under the factor Experiment to generate 1000 simulated datasets, followed by identifying DEGs in paired comparisons between a selected common reference batch and each of the remaining batches in every dataset. Note that a different batch was selected as a common reference in the analysis of each simulated dataset. To determine if between-batch variation is biologically significant, the 100 DEGs with the highest F statistic p-values for between-batch variation under each factor were also combined and analyzed for possible enrichment in biological pathways by DAVID  http://regressit.com/index.html).Individuals within a batch are subjected to the same degree of technical variation, thus their differences should better reflect biological variability. We used two different approaches to evaluate within-batch variation. First, for a given factor, half the samples in each batch randomly assigned as “case” were compared to the remaining half assigned as “control” for DEGs during each of 250 permutations. Between-batch variation was controlled in these analyses by including the factor under consideration in the GLM to ensure the comparisons were made within a batch. This method essentially searched through samples’ possible membership assignments between the two classes to uncover the configuration where the number of DEGs was maximized. This DEG count served as an indirect measure of the variation among individuals. To be included in these permutations, each batch needed to contain a minimum of ten (zebrafish) or 12 (fathead minnow) samples, respectively, to ensure that there were at least 250 unique permutations and the maximum amount of data was utilized. Second, the coefficient of variation (CV) and intensity of individual probes were also calculated from multiple biological replicates in individual batches under the Experiment factor and averaged over all its batches. An assessment of these two simple metrics both within and between the two species, at the levels of entire transcriptome, KEGG (Kyoto Encyclopedia of Genes and Genomes) molecular pathways, and individual genes, would be informative of not only gene expression variability among individuals and relative biological contribution to this variability, but also the interspecific transcriptomic conservation as well. The simple linear regressions and validations of their underlying statistical assumptions were conducted for selected inter- and intraspecific comparisons of these two metrics using “RegressIt”, an Excel macro developed jointly at Duke University and University of Texas (ndenslow@ufl.edu) by TBLASTX so the latter with a greater sequence length could be used as queries for more successful mapping across species. The EST sequences were then mapped to the NCBI  nucleotide  and protein  databases by TBLASTX and BLASTX respectively, effectively associating fathead minnow probe IDs to their corresponding NCBI accession IDs. All three rounds of mapping had a minimum E-value cutoff of E−06. These fathead minnow IDs were then joined to a variety of zebrafish accession IDs prepared by the NCBI , and finally to Agilent probe annotations (https://earray.chem.agilent.com/earray/). In the end, a total of 9311 probes from probes from Agilent 019597 through 6617 common Entrez GeneIDs in NCBI (http://rest.kegg.jp/link/dre/pathway) by these GeneIDs. Before the pathway level the CV and intensity of those duplicated probes were consolidated first by Entrez GeneIDs.To allow interspecific comparison of within-batch variation, probes from Agilent design 019597 (fathead minnow) and 015064 (zebrafish) representing orthologous genes were identified through several successive steps of identification (ID) mapping. Probe sequences from Agilent 019597 were first mapped to their corresponding EST (Expressed Sequence Tag) target sequences  scripts developed by the authors based primarily on the R. In contrast to DEG determination by a regular t-test (or F-test), this software fits expression data of each gene into a linear model and generates a modified t-statistic (or F-statistic for multiple contrasts) using an empirical Bayesian and hierarchical modeling approach to adjust for unreliable variance estimates caused by small sample size The analytical procedures described above for both between- and within-batch variation were incorporated into several R  under the Experiment factor, dividing the samples into two classes with each containing four to 11 samples will result in 2.4×EThere have been a limited number of studies reporting transcriptomic variation among individuals within a population, which are similar in nature to the within-batch variation under consideration here. These studies were limited to several organisms sampled from wild or lab populations with different levels of heterozygosity. Most of them were based on utilizing technical replicates to various degrees to estimate variability. The range of variation by percentage of genes identified as DEGs is quite wide: 2 to 9% in nematode (whole organism) 2 = 0.46 and 0.49, The expression intensities of the orthologs and their variation within-batch differed considerably among the molecular pathways in the two species. For example, in both species, some of the pathways involved in basic cellular functions such as Ribosome, RNA Transport, Citrate Cycle, Cell Cycle, and RNA Polymerase tended to be less variable. Some of the metabolic pathways, on the other hand, were more variable in both species but involved in different functions. Many pathways involved in signal transduction were expressed on average at a fairly low level, while those participating in basic cellular functions like DNA replication and protein synthesis were highly expressed. Although this difference in the average variability of pathways could be explained to a considerable extent by an inverse correlation with average intensity , a large portion of the variance in the average CV remains unaccounted for by the average intensity at pathway level  and 019597 (fathead minnow) via their orthologs. A total of 9311 probes from Agilent 015064 were linked to 6950 probes from Agilent 019597 through 6617 common Entrez GeneIDs in NCBI.(XLSX)Click here for additional data file.S2 TableThe DAVID analysis of fathead minnow DEGs. The top 100 DEGs were combined from the between-batch variation under each factor for possible enrichment in biological pathways.(PDF)Click here for additional data file.S3 TableThe DAVID analysis of zebrafish DEGs. The top 100 DEGs were combined from the between-batch variation under each factor for possible enrichment in biological pathways.(PDF)Click here for additional data file.S4 TableInterspecific comparison of within-batch variation at the molecular pathway level. A total of 6617 orthologous genes were grouped into 144 out of the 162 KEGG pathways as of April, 2014. The intensity and CV were calculated by individual batches each containing multiple biological replicates under the Experiment factor, and then averaged over all batches. Only pathways with at least five orthologs were included. KEGG, Kyoto Encyclopedia of Genes and Genomes; DRE, zebrafish; PPR, fathead minnow.(XLSX)Click here for additional data file.S5 TableThe top 50 most variable fathead minnow (PPR) genes based on average within-batch coefficient of variation (CV) under the Experiment factor.(DOCX)Click here for additional data file.S6 TableThe top 50 most variable zebrafish (DRE) genes based on average within-batch coefficient of variation (CV) under the Experiment factor.(DOCX)Click here for additional data file.S1 FileAverage number of DEGs per pair of between-batch comparison in 1000 permutations. Fathead minnow  critical cutoffs were: 5%, 15; 1%, 27; 0.1%, 104. Zebrafish  critical cutoffs were: 5%, 418; 1%, 479; 0.1%, 594. Samples were grouped by the factor Experiment.(PDF)Click here for additional data file.S2 FileThe PCA plots of fathead minnow samples based on all the DEGs identified as between-batch variation. Samples were grouped by Experiment , RNA Date , RNA Person , Sampling Date , and Scan Date . Each figure was based on either the average gene intensity by individual batches (A) or the gene intensity of individual samples (B). DEGs were based on the simulated reference method.(PDF)Click here for additional data file.S3 FileThe PCA plots of zebrafish samples based on all the DEGs identified as between-batch variation. Samples were grouped by Experiment , RNA Date , RNA Person , Sampling Date , and Scan Date . Each figure was based on either the average gene intensity by individual batches (A) or the gene intensity of individual samples (B). DEGs were based on the simulated reference method.(PDF)Click here for additional data file.S4 FileThe dendrograms from resampling clustering of fathead minnow samples based on all the DEGs identified as between-batch variation. Samples were grouped by Experiment , RNA Date , RNA Person , Sampling Date , and Scan Date . Each figure was based on either the average gene intensity by individual batches (A) or the gene intensity of individual samples (B). DEGs were based on the simulated reference method.(PDF)Click here for additional data file.S5 FileThe dendrogram from resampling clustering of zebrafish samples based on all the DEGs identified as between-batch variation. Samples were grouped by Experiment , RNA Date , RNA Person , Sampling Date , and Scan Date . Each figure was based on either the average gene intensity by individual batches (A) or the gene intensity of individual samples (B). DEGs were based on the simulated reference method.(PDF)Click here for additional data file.S6 FileThe distribution of within-batch coefficients of variation (CV) of probe intensities. Fathead minnow  CVs were based on 15208 probes, and zebrafish  CVs were based on 21495 probes.(PDF)Click here for additional data file.


Maiasaura peeblesorum (Ornithopoda: Hadrosaurinae) exhibit exostoses. We show that these outgrowths are cases of biomechanically adaptive periosteal bone modeling resulting from overstrain on the tibia after a fibula fracture. Histological and biomechanical results are congruent with predictions derived from this hypothesis. Histologically, the outgrowths are constituted by radial fibrolamellar periosteal bone tissue formed at very high growth rates, as expected in a process of rapid strain equilibration response. These outgrowths show greater compactness at the periphery, where tensile and compressive biomechanical constraints are higher. Moreover, these outgrowths increase the maximum bending strength in the direction of the stresses derived from locomotion. They are located on the antero-lateral side of the tibia, as expected in a presumably bipedal one year old individual, and in the posterior position of the tibia, as expected in a presumably quadrupedal individual at least four years of age. These results reinforce myological evidence suggesting that Maiasaura underwent an ontogenetic shift from the primitive ornithischian bipedal condition when young to a derived quadrupedal posture when older.Predator confrontation or predator evasion frequently produces bone fractures in potential prey in the wild. Although there are reports of healed bone injuries and pathologies in non-avian dinosaurs, no previously published instances of biomechanically adaptive bone modeling exist. Two tibiae from an ontogenetic sample of fifty specimens of the herbivorous dinosaur  Maiasaura [Tyrannosaurus [Allosaurus [Maiasaura peeblesorum. This study is aimed at testing hypotheses on the proximal causation of these bone outgrowths.Intensive paleontological fieldwork over the last three decades has produced a rich collection of non-avian dinosaur fossils permitting detailed ontogenetic descriptions and paleobiological estimations of life history traits using bone histology . These losaurus ; 36% . Moreover, the topological position of these outgrowths is interpreted as evidence for an ontogenetic shift from a bipedal to a quadrupedal posture in Maiasaura.Bone periostitis is a common bone abnormality easily recognized by the expansion of diaphyseal contours (outgrowths) and involving an alteration in bone surface texture . Such feMaiasaura peeblesorum fossils collected from a rich, monodominant bonebed in the Campanian sediments of the Two Medicine Formation [Over more than thirty years, the Museum of the Rockies  has prepared disarticulated ormation . Fifty tMaiasaura tibiae. Thin section slides were processed using a Buehler Ecomet 4 variable speed grinder, using the following sequence of grit papers: 60, 180, 320, 600, and 800. Completed slides were analysed using a Nikon Optiphot-Pol polarizing microscope at either 10 X or 40 X total magnification, and photomicrographs were taken incrementally using a Nikon DS-Fi1 digital sight camera. The software package NIS-Elements BR 3.0 was used to create a single image from multiple photographs, so that composite images of the tibia thin sections have a mosaic appearance. Minimum age of Maiasaura individuals was determined by counting the number of annually deposited lines of arrested growth .Thin sections were prepared from 0.3 cm thick wafers of bone removed transversely from either side of the minimum diaphyseal circumference of wth , biomechanical ), and pMaiasaura peeblesorum analyzed contains two tibiae (MOR 005-T9 and MOR 005-T42) exhibiting exostoses  simulating the effects of a fracture in a zeugopodial bone (the ulna) produces increased strains in the adjacent zeugopodial element (the radius), promoting bone modeling (bone formation occurs over primary bone) and/or remodeling  –16. In aMaisaura tibia is mainly formed by laminar fibrolamellar bone tissue interrupted by lines of arrested growth  from a one year old specimen (MOR 005-T9) and a single layer in a larger tibia (90.5 cm length) from an immature individual at least four years of age (MOR 005-T42)  suggests an abrupt change in mechanical constraints compatible with fibula fracture. Always in the context of uniformitarianism, the finding of rapidly formed radial fibrolamellar bone tissue in the young pig radius after ulnar osteotomy [The cortex of wth Figs  and 3. F005-T42) . Accordisteotomy  is congrsteotomy  providesI), proportional to the bending strength of a long bone, is computed as:z δy is the cross-sectional area of a thin layer at a distance y from the neutral axis [The second moment of the area . Thus, ral axis , 20, andral axis ).Maiasaura tibiae would increase the diaphyseal second moment of the area . Experistrength . Data on torsion . Summari torsion ), wherea torsion ). Myolog posture . So we e posture  Fig 4C)Maiasauratetrapod ). This itetrapod , 26, 27.tetrapod , 29. Howtetrapod , 26). Thtetrapod . Moreovetetrapod . Periosttetrapod . In our Describing modified bone structure as resulting from modeling requires that scenarios regarding disease be ruled out. As noted, in MOR 005-T9 and T42, there is a collar of fibro-lamellar bone lying beneath the periosteal layer of the tibia. This can be observed grossly as an area of smooth bowed thickened bone on the diaphysis . AlthougMaiasaura specimens in this paper is the circumferential deposition of bone around the entire diaphysis that characterizes this condition. Additionally, both individuals lack evidence of the classic second phase of the disease in which the trabecular and endosteal bone becomes highly decreased in density [Perhaps the most compelling similarity to a pathologic condition is avian osteopetrosis –41. This density .In sum, the known pathologic scenarios fall short of explaining the bone histology observed in these two specimens. The authors grant that a scenario for a pathologic etiology for which there is no modern analog , 43 or aMaiasaura. In contrast, the hypothesis according to which these outgrowths are cases of biomechanically adaptive periosteal bone modeling after fibula fracture are strongly supported by histological and biomechanical data. The quantification of the frequency of (observed or inferred) bone fractures is important in paleontological population studies. Moreover, by documenting the differences in the compensating biomechanical responses in the tibiae between Maiasaura juveniles and sub-adults, our study independently supports the hypothesis that Maiasaura underwent an ontogenetic shift from bipedality to quadrupedality previously suggested by a myological analysis [Maiasaura, bipedality), whereas derived character states  did not develop until late-juvenile or sub-adult stages of growth, further implying the need to consider that small-bodied dinosaurs with unique combinations of shared and derived characteristics may in fact be immature morphs of derived taxa.Paleopathology hypotheses (periosteal reactive lesions , 32, 33,analysis . Evidenc