Spatial Characteristics of Colonic Mucosa-Associated Gut Microbiota in Humans.

Limited data exist on the spatial distribution of the colonic bacteria in humans. We collected the colonic biopsies from five segments of 27 polyp-free adults and collected feces from 13 of them. We sequenced the V4 region of the bacterial 16S rRNA gene using the MiSeq platform. The sequencing data were assigned to the amplicon sequence variant (ASV) using SILVA. Biodiversity and the relative abundance of the ASV were compared across the colonic segments and between the rectal and fecal samples. Bacterial functional capacity was assessed using Tax4fun. Each individual had a unique bacterial community composition (Weighted Bray-Curtis P value = 0.001). There were no significant differences in richness, evenness, community composition, and the taxonomic structure across the colon segments in all the samples. Firmicutes (47%), Bacteroidetes (39%), and Proteobacteria (6%) were the major phyla in all segments, followed by Verrucomicrobia, Fusobacteria, Desulfobacterota, and Actinobacteria. There were 15 genera with relative abundance > 1%, including Bacteroides, Faecalibacterium, Escherichia/Shigella, Sutterella, Akkermansia, Parabacteroides, Prevotella, Lachnoclostridium, Alistipes, Fusobacterium, Erysipelatoclostridium, and four Lachnospiraceae family members. Intra-individually, the community compositional dissimilarity was the greatest between the cecum and the rectum. There were significant differences in biodiversity and the taxonomic structure between the rectal and fecal bacteria. The bacterial community composition and structure were homogeneous across the large intestine in adults. The inter-individual variability of the bacteria was greater than inter-segment variability. The rectal and fecal bacteria differed in the community composition and structure.

Immunosuppressive Drug Levels in Liver Transplant Recipients: Impact in Decision Making.

To prevent rejection, liver transplant providers largely base their management decisions on their clinical impression and pharmacokinetics. Clinical impression relies on assessing graft function, liver enzymes, and biopsy. High immunosuppressive drug levels, although minimizing rejection, are related to significant side effects such as nephrotoxicity and metabolic syndrome, contributing to long-term morbidity and mortality. Similarly, levels that are lower than necessary can decrease the rate of side effects with a potential toll on rejection and graft survival. Herein, the authors present an update on immunosuppressive drug level monitoring and manipulation strategies according to different scenarios and time from transplant. They also provide a brief overview of next level immunosuppression monitoring strategies that aim to properly balance rejection rates with drug side effect profiles.

A multi-host approach for the systematic analysis of virulence factors in Cryptococcus neoformans.

A multi-host approach was followed to screen a library of 1201 signature-tagged deletion strains of Cryptococcus neoformans mutants to identify previously unknown virulence factors. The primary screen was performed using a Caenorhabditis elegans-C. neoformans infection assay. The hits among these strains were reconfirmed as less virulent than the wild type in the insect Galleria mellonella-C. neoformans infection assay. After this 2-stage screen, and to prioritize hits, we performed serial evaluations of the selected strains, using the C. elegans model. All hit strains identified through these studies were validated in a murine model of systemic cryptococcosis. Twelve strains were identified through a stepwise screening assay. Among them, 4 (CSN1201, SRE1, RDI1, and YLR243W) were previously discovered, providing proof of principle for this approach, while the role of the remaining 8 genes (CKS101, CNC5600, YOL003C, CND1850, MLH3, HAP502, MSL5, and CNA2580) were not previously described in cryptococcal virulence. The multi-host approach is an efficient method of studying the pathogenesis of C. neoformans. We used diverse model hosts, C. elegans, G. mellonella, and mice, with physiological differences and identified 12 genes associated with mammalian infection. Our approach may be suitable for large pathogenesis screens.

Risk factors for invasive fungal disease after allogeneic hematopoietic stem cell transplantation: a single center experience.

Invasive fungal disease (IFD) is a major cause of morbidity and mortality after hematopoietic stem cell transplantation (HCT). We performed a retrospective review of 271 adults with a hematologic malignancy undergoing allogeneic HCT to determine the incidence of and risk factors for IFD and to examine the impact of IFD on nonrelapse mortality and overall survival. We defined IFD using standard criteria and selected proven and probable cases for analysis. Diagnoses in the study group included acute leukemia (42%), non-Hodgkin lymphoma (24%), myelodysplastic syndrome (15%), chronic lymphocytic leukemia (5%), and other hematologic disorders (14%). Conditioning included reduced-intensity (64%) and myeloablative (36%) regimens. Donor sources were HLA-matched sibling (60%), matched unrelated (20%), haploidentical (12%), and cord blood (8%). A total of 51 episodes of IFD were observed in 42 subjects (15%). Aspergillus spp (47%) was the most frequent causative organism, followed by Candida spp (43%). The majority of IFD cases (67%) were reported after day +100 post-HCT. In multivariate analysis, haploidentical donor transplantation (hazard ratio [HR], 3.82; 95% confidence interval [CI], 1.49-9.77; P = .005) and grade II-IV acute graft-versus-host disease (HR, 2.55; 95% CI, 1.07-6.10; P = .03) were risk factors for the development of IFD. Conversely, higher infused CD34(+) cell dose was associated with a lower risk of IFD (HR, 0.80; 95% CI, 0.68-0.94; P = .006, per 1 × 10(6) cells/kg increase in CD34(+) cell infusion). IFD-related mortality was 33.3%. Nonrelapse mortality was significantly higher in patients who developed IFD compared with those without IFD (P < .001, log-rank test). Patients with IFD had lower overall survival (5.8 months versus 76.1 months; P < .001, log-rank test). Further studies exploring strategies to increase the infused cell dose and determine adequate prophylaxis, especially against aspergillus, beyond day +100 are needed.

N-acetylglucosamine induces white-to-opaque switching and mating in Candida tropicalis, providing new insights into adaptation and fungal sexual evolution.

Pathogenic fungi are capable of switching between different phenotypes, each of which has a different biological advantage. In the most prevalent human fungal pathogen, Candida albicans, phenotypic transitions not only improve its adaptation to a continuously changing host microenvironment but also regulate sexual mating. In this report, we show that Candida tropicalis, another important human opportunistic pathogen, undergoes reversible and heritable phenotypic switching, referred to as the "white-opaque" transition. Here we show that N-acetylglucosamine (GlcNAc), an inducer of white-to-opaque switching in C. albicans, promotes opaque-cell formation and mating and also inhibits filamentation in a number of natural C. tropicalis strains. Our results suggest that host chemical signals may facilitate this phenotypic switching and mating of C. tropicalis, which had been previously thought to reproduce asexually. Overexpression of the C. tropicalis WOR1 gene in C. albicans induces opaque-cell formation. Additionally, an intermediate phase between white and opaque was observed in C. tropicalis, indicating that the switching could be tristable.

Polymerase chain reaction-based assays for the diagnosis of invasive fungal infections.

Currently accepted fungal diagnostic techniques, such as culture, biopsy, and serology, lack rapidity and efficiency. Newer diagnostic methods, such as polymerase chain reaction (PCR)-based assays, have the potential to improve fungal diagnostics in a faster, more sensitive, and specific manner. Preliminary data indicate that, when PCR-based fungal diagnostic assays guide antifungal therapy, they may lower patient mortality and decrease unnecessary antifungal treatment, improving treatment-associated costs and avoiding toxicity. Moreover, newer PCR techniques can identify antifungal resistance DNA loci, but the clinical correlation between those loci and clinical failure has to be studied further. In addition, future studies need to focus on the implementation of PCR techniques in clinical decision making and on combining them with other diagnostic tests. A consensus on the standardization of PCR techniques, along with validation from large prospective studies, is necessary to allow widespread adoption of these assays.

The effect of cumulative length of hospital stay on the antifungal resistance of Candida strains isolated from critically ill surgical patients.

Fluconazole is the first line of therapy for the management of candidiasis. However, fluconazole-resistant strains pose an emerging challenge in everyday clinical practice. In this study, we sought to determine whether cumulative length of hospital stay (CLOS) is a predictive factor for the acquisition of non-susceptible Candida strains to fluconazole. Thirty-three critically ill emergency surgery patients with 56 Candida isolates were enrolled in this prospective study. We divided our isolates according to their minimum inhibitory concentration (MIC) to fluconazole using 8 mcg/ml as a cutoff. We then compared the two groups with respect to basic demographics, antifungal agents prescribed, number of wide-spectrum antibiotics, duration of central venous catheter placement, elapsed time to positive culture, duration of prior hospital stay, and length of hospital stay. Non-susceptible fluconazole samples belonged to patients with a significantly longer prior hospital stay and a longer CLOS (P = 0.02 and 0.01, respectively). The difference between the 2 groups regarding non-albicans strains was statistically significant (P < 0.001). By fitting a non-parametric receiver-operating characteristics (ROC) curve into our analysis, a CLOS ≥ 29 days predicted the occurrence of non-susceptible strains with 90% sensitivity and 79.6% specificity (correct classification 81.5%). A CLOS ≥ 29 days is a strong predictor for the isolation of non-susceptible Candida isolates to fluconazole among critically ill emergency surgery patients. Clinicians should consider the duration of previous hospital stay when deciding on empiric antifungal therapy.

An Unusual Cause of Acute Pancreatitis in a Liver Transplant Recipient.

Posttransplant lymphoproliferative disorder (PTLD) in liver transplant recipients is relatively uncommon, with an estimated incidence of 1%-3%. Retrospective reviews of liver transplant recipients have mainly reported posttransplant lymphoproliferative disorder affecting the liver, gastrointestinal tract, or lymph nodes. In this case report, we describe a 45-y-old female with a history of deceased donor liver transplantation for autoimmune hepatitis who had recurrent hospital admissions for acute pancreatitis. Ultimately, imaging revealed numerous complex pancreatic and peripancreatic masses, appearing to originate from pancreatic lymphoid tissue. Tissue biopsy later confirmed monomorphic Epstein-Barr virus-negative large B-cell lymphoma. Overall, PTLD involving the pancreas after liver transplantation is incredibly rare. The patient's cumulative immunosuppression drug dose and time posttransplant were suspected to be her main risk factors, given that she had been exposed to several years of treatment with tacrolimus, azathioprine, mycophenolate mofetil, and prednisone. She was treated with rituximab monotherapy and later escalated to chemoimmunotherapy due to lack of response. PTLD involving the pancreas is an unusual cause of pancreatitis and should be considered in cases of recurrent pancreatitis in transplant recipients.

Pathogenesis of Alcoholic Liver Disease: An Update.

Apart from the classic knowledge that ethanol mediates its hepatotoxicity through its metabolism to acetaldehyde, a well-known hepatotoxic molecule, recent research has elucidated several key mechanisms that potentiate ethanol's damage to the liver parenchyma, such as generation of free radicals, activation of Kupffer cells, and alterations to the human bacterial and fungal microbiome. Genetic studies have suggested the role of PNPLA3 and TM6SF2 gene mutations in the progression of alcoholic liver disease.

Dietary Nutrients Involved in One-Carbon Metabolism and Colonic Mucosa-Associated Gut Microbiome in Individuals with an Endoscopically Normal Colon.

One carbon (1C) metabolism nutrients influence epigenetic regulation and they are supplied by diet and synthesized by gut microbiota. We examined the association between dietary consumption of methyl donors (methionine, betaine and choline) and B vitamins (folate, B2, B6, and B12) and the community composition and structure of the colonic mucosa-associated gut microbiota determined by 16S rRNA gene sequencing in 97 colonic biopsies of 35 men. We used the food frequency questionnaire to assess daily consumption of nutrients, and the UPARSE and SILVA databases for operational taxonomic unit classification. The difference in bacterial diversity and taxonomic relative abundance were compared between low versus high consumption of these nutrients. False discover rate (FDR) adjusted p value < 0.05 indicated statistical significance. The bacterial richness and composition differed significantly by the consumption of folate and B vitamins (p < 0.001). Compared with higher consumption, a lower consumption of these nutrients was associated with a lower abundance of Akkermansia (folate), Roseburia (vitamin B2), and Faecalibacterium (vitamins B2, B6, and B12) but a higher abundance of Erysipelatoclostridium (vitamin B2) (FDR p values < 0.05). The community composition and structure of the colonic bacteria differed significantly by dietary consumption of folate and B vitamins.

Antibody Response to Hepatitis B Virus Vaccine is Impaired in Patients With Inflammatory Bowel Disease on Infliximab Therapy.

Background: Studies have demonstrated an association between anti-TNF/immunomodulator agents used in inflammatory bowel disease (IBD) and impaired hepatitis B virus (HBV) vaccine immunogenicity, but little data exist on whether specific medication types affect protective HBsAb titers. Our aim was to analyze this association. Methods: This is a retrospective cohort study. Inclusion criteria: age ≥18, diagnosis of Crohn's disease (CD) or ulcerative colitis (UC), previous HBV vaccination series and/or ≥1 positive HBsAb, and record of IBD therapy in 6 months before titer level. Patients were stratified based upon medication exposures: anti-TNF, immunomodulator, combination anti-TNF and immunomodulatory, and a reference arm. Titer levels following vaccination and specific medication types given in the 6 months before titer were recorded. Seroprotection was defined as HBsAb ≥10 IU/l and ≥100 IU/l. Results: The study cohort (N = 391) was 70.8% white, 51.4% female and 64.2% had CD and 35.8% had UC. The mean age was 45.8 years. A significantly lower percentage of patients exposed to anti-TNF, immunomodulator or dual therapy had titers ≥10 (P < 0.01). Regarding specific medications, only patients exposed to infliximab (P < 0.01) were less likely to have titer levels ≥10, after controlling for other medication exposures, age at titer level, and interval time between vaccination/titer level. This was not found for patients exposed to adalimumab, methotrexate, 6-mercaptopurine, or azathioprine. Conclusions: Patients exposed to infliximab were significantly less likely to have protective HBsAb titer levels following vaccination, a trend not seen in patients on adalimumab. Efforts to vaccinate IBD patients against HBV before use of immunomodulators and anti-TNFs, infliximab specifically, and screen periodically thereafter must be reinforced.

The impact of antimicrobial resistance and aging in VAP outcomes: experience from a large tertiary care center.

BACKGROUND: Ventilator associated pneumonia (VAP) is a serious infection among patients in the intensive care unit (ICU). METHODS: We reviewed the medical charts of all patients admitted to the adult intensive care units of the Massachusetts General Hospital that went on to develop VAP during a five year period. RESULTS: 200 patients were included in the study of which 50 (25%) were infected with a multidrug resistant pathogen. Increased age, dialysis and late onset (≥ 5 days from admission) VAP were associated with increased incidence of resistance. Multidrug resistant bacteria (MDRB) isolation was associated with a significant increase in median length of ICU stay (19 vs. 16 days, p=0.02) and prolonged duration of mechanical ventilation (18 vs. 14 days, p=0.03), but did not impact overall mortality (HR 1.12, 95% CI 0.51-2.46, p=0.77). However, age (HR 1.04 95% CI 1.01-1.07, p=0.003) was an independent risk factor for mortality and age ≥ 65 years was associated with increased incidence of methicillin-resistant Staphylococcus aureus (MRSA) infections (OR 2.83, 95% CI 1.27-6.32, p=0.01). CONCLUSIONS: MDRB-related VAP is associated with prolonged ICU stay and mechanical ventilation. Interestingly, age ≥ 65 years is associated with MRSA VAP.

Nocardiosis of the central nervous system: experience from a general hospital and review of 84 cases from the literature.

Central nervous system (CNS) nocardiosis is a rare disease entity caused by the filamentous bacteria Nocardia species. We present a case series of 5 patients from our hospital and a review of the cases of CNS nocardiosis reported in the literature from January 2000 to December 2011. Our results indicate that CNS nocardiosis can occur in both immunocompromised and immunocompetent individuals and can be the result of prior pulmonary infection or can exist on its own. The most common predisposing factors are corticosteroid use (54% of patients) and organ transplantation (25%). Presentation of the disease is widely variable, and available diagnostic tests are far from perfect, often leading to delayed detection and initiation of treatment. The optimal therapeutic approach is still undetermined and depends on speciation, but lower mortality and relapse rates have been reported with a combination of targeted antimicrobial treatment including trimethoprim/sulfomethoxazole (TMP-SMX) for more than 6 months and neurosurgical intervention.

Fusarium infection: report of 26 cases and review of 97 cases from the literature.

Fusarium species is a ubiquitous fungus that causes opportunistic infections. We present 26 cases of invasive fusariosis categorized according to the European Organization for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG) criteria of fungal infections. All cases (20 proven and 6 probable) were treated from January 2000 until January 2010. We also review 97 cases reported since 2000. The most important risk factors for invasive fusariosis in our patients were compromised immune system, specifically lung transplantation (n = 6) and hematologic malignancies (n = 5), and burns (n = 7 patients with skin fusariosis), while the most commonly infected site was the skin in 11 of 26 patients. The mortality rates among our patients with disseminated, skin, and pulmonary fusariosis were 50%, 40%, and 37.5%, respectively. Fusarium solani was the most frequent species, isolated from 49% of literature cases. Blood cultures were positive in 82% of both current study and literature patients with disseminated fusariosis, while the remaining 16% had 2 noncontiguous sites of infection but negative blood cultures. Surgical removal of focal lesions was effective in both current study and literature cases. Skin lesions in immunocompromised patients should raise the suspicion for skin or disseminated fusariosis. The combination of medical monotherapy with voriconazole or amphotericin B and surgery in such cases is highly suggested.

Update on the epidemiology and management of cryptococcal meningitis.

Despite recent improvements in the diagnosis and treatment of cryptococcosis, cryptococcal meningitis is responsible for > 600,000 deaths/year worldwide. The aim of this work is to provide an update on the developments in its epidemiology and management. Understanding the pathogenesis of Cryptococcus has improved, and new insights for the virulence of the fungus and the host response have enabled scientists to design new ways to confront this infection. Additionally, invertebrate model hosts have greatly facilitated the research in this field. Importantly, the epidemiology of Cryptococcus gattii has continued to evolve, and the emergence of this highly virulent species in immunocompetent populations, especially in Northwestern America and British Columbia, warrants increased awareness because delayed diagnosis and inappropriate antifungal therapy is associated with high mortality. Diagnosis remains a challenge, but new techniques for early and inexpensive identification of the pathogen are under development. Management can vary, based on the patient population (HIV-seropositive, organ transplant recipients or non-transplant/non-HIV). In most patients, amphotericin B with flucytosine continues to be the most appropriate induction therapy. However, in organ transplant recipients the use of liposomal amphotericin B improves mortality compared with deoxycholate amphotericin B. Also, the combination of amphotericin B with fluconazole seems to be a reasonable alternative, while fluconazole with flucytosine is superior to fluconazole monotherapy.

Central nervous system aspergillosis: a series of 14 cases from a general hospital and review of 123 cases from the literature.

Central nervous system (CNS) aspergillosis is a highly fatal infection. We review the clinical presentation, diagnosis, and outcome of this infection and present a case series of 14 consecutive patients with CNS aspergillosis admitted to Massachusetts General Hospital (MGH) from 2000 to 2011. We also review 123 cases reported in the literature during that time. We included only proven CNS aspergillosis cases conforming to the European Organization for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG) definitions of invasive fungal infections. In the MGH case series, neutropenia, hematologic malignancies, autoimmune diseases requiring steroid treatment, and solid organ transplantation were the predominant comorbid conditions. Notably, all MGH patients were immunosuppressed, and more than half (n = 8) had a history of previous brain injury, unrelated to their index hospitalization. For most MGH patients (11 of 14), the lung was the primary focus of aspergillosis, while 2 had paranasal sinus involvement, and 1 had primary Aspergillus discitis. Among reported cases, paranasal sinuses (27.6%) and the lung (26.8%) were the primary foci of infection, whereas 22% of those cases had no obvious primary organ involvement. Although a selection bias should be considered, especially among published cases, our findings suggest that patients who underwent neurosurgery had improved survival, with MGH and literature patients having 25% and 28.6% mortality, respectively, compared to 100% and 60.4%, respectively, among patients who received only medical treatment. Although this was not the case among MGH patients, CNS aspergillosis can affect patients without significant immune suppression, as indicated by the high number of reported immunocompetent cases. In conclusion, mortality among CNS aspergillosis patients remains high, and the infection may be more common among patients with previous brain pathology. When indicated, neurosurgical procedures may improve prognosis.

Candida infection and colonization among trauma patients.

BACKGROUND: Data on Candida infection among critically ill trauma patients are limited and not recently updated. Here we study the epidemiology and economic impact of Candida and examine potential risk factors for Candida infection in this population. METHODS: In this 5-year retrospective study, all severely injured patients with ≥4 days of intensive care unit stay were included, with the primary outcome being Candida infection. We identified 3 distinct patient groups: 1) The Candida infection, 2) The Candida colonization and 3) the Candida-free group. All comparisons between groups with p-values ≤0.2 from the univariate analysis were entered into stepwise logistic regression to identify independent risk factors for candidiasis. RESULTS: 374 patients were included. Upon comparisons between groups, candidiasis patients received significantly more blood transfusions (p=0.013), antibiotics (p=0.005), and total parenteral nutrition (TPN) (p=0.004), had a longer duration of mechanical ventilation (MV) (p=0.008) and underwent more laparotomy procedures than Candida free patients (56.5% versus 16.4%; p<0.001). Surgical complications (13% versus 1.4%; p=0.013), injury of the upper (13% versus 0.9%; p=0.007) and lower gastrointestinal tract (8.7% versus 0.9%; p=0.048), and bacterial wound or intra-abdominal infections (17.4% versus 1.9%; p=0.004) were also more common in candidiasis patients. Upon multivariate analysis, patients receiving TPN had 7-fold higher odds for developing candidiasis (Odds ratio [OR]: 7.2; 95% Confidence interval [CI]: 2.6-19.4; p=0.0001). Other predisposing factors included laparotomy (OR: 3.8, 95% CI: 1.5-9.9; p=0.0057) and female gender (OR: 5.7; 95% CI: 2.1-15.6; p=0.0007). Average total hospital charges were higher for patients with Candida infection compared to patients with Candida colonization or without a positive Candida culture. CONCLUSIONS: TPN, laparotomy, and female gender independently predict the development of candidiasis among trauma patients. Severely injured women requiring laparotomy and TPN therapy should be carefully managed for the possibility of increased risk for candidiasis.

Candida infection and colonization among non-trauma emergency surgery patients.

BACKGROUND: Candida is a significant pathogen among critically ill patients. However, candidiasis among non-trauma emergency surgery (NTES) patients has not been previously investigated. Herein we describe the incidence of both colonization and infection from Candida and risk factors for invasive disease in this population. METHODS: For this retrospective single center study we included all NTES patients with ICU stay ≥4 days from May 1(st), 2002 to April 30(th), 2007. Patients were divided into 3 non-overlapping groups: 1) patients with Candida-infection, 2) patients with Candida colonization and 3) patients with negative Candida cultures. Groups were compared by univariate and multivariate analyses to identify significant risk factors for invasive candidiasis. RESULTS: Of all 289 eligible patients, 63 (21.7%) fulfilled the criteria for Candida infection and 110 (38%) were included in the Candida colonization group. Interestingly, from the 63 patients with invasive candidiasis, 25 (39.7%) were infected by a non-albicans species. Upon multivariate analyses, ventilator-associated pneumonia (VAP) (Odds Ratio [OR]: 2.34; 95%, Confidence Interval [CI]: 1.213-4.533, p = 0.0112), bacteremia (OR: 4.778; 95%CI: 1.519-15.029, p = 0.0075) and surgical complications (OR: 3.903; 95%CI: 1.335-11.412, p = 0.0129) were independent risk factors for the development of Candida infection. Candida infection and colonization were both found to correlate with approximately $40,000-100,000 mean additional costs). Interestingly, candidemia was associated with 63% all-cause mortality. For all other forms of candidiasis, mortality was not significantly different among groups. CONCLUSION: We found that Candida infection is alarmingly high among NTES patients with prolonged intensive care unit (ICU) stay. Surgical complications and bacterial infections (VAP and bacteraemia) were significantly correlated with the development of candidiasis. Candidiasis reached a rate of 21.7/100 discharges, which is significantly higher than most established high-risk populations for candidiasis. Future studies should review the need for antifungal prophylaxis on this population.