Extracellular vesicles of Bacteroides fragilis regulated macrophage polarization through promoted Sema7a expression.

Abnormal activation of macrophage and gut Bacteroides fragilis (BF) are the important induction factors in the occurrence of type 2 diabetes (T2D) and vascular complications. However, it remains unknown whether BF involves in macrophage polarization. In this study, we found that BF extracellular vesicles (EV) can be uptaken by macrophage. BF-EV promote macrophage M1/M2 polarization significantly, and increase Sting expression significantly. Bioinformatics analysis found that Sema7a is an important gene involving in macrophage polarization. The expression of Sema7a can be induced by BF-EV and can be inhibited after C-176 treated. The inhibition expression of Sema7a prevent BF-EV to induce macrophage polarization. Further analysis reveals that there is no direct interaction between Sting and Sema7a, but Sgpl1 can interact with Sting or Sema7a. BF-EV promote the expression of Sgpl1, which the phenomenon can be inhibited after C-176 treated. Importantly, overexpression of Sgpl1 reversed the effect of C-176 for Sema7a expression, while inhibit Sema7a expression has limitation influence for Sting and Sgpl1 expression. In conclusion, this study confirms that Sting-Sgpl1-Sema7a is a key mechanism by which BF-EV regulates macrophage polarization.

Bacteroides fragilis expresses three proteins similar to Porphyromonas gingivalis HmuY: Hemophore-like proteins differentially evolved to participate in heme acquisition in oral and gut microbiomes.

Oral and gut microbiomes are important for the maintenance of homeostasis in the human body. Altered or disturbed mutualism between their members results in dysbiosis with local injury and subsequent systemic diseases. The high bacterial density causes intense competition among microbiome residents to acquire nutrients, including iron and heme, the latter of high importance for heme auxotrophic members of the Bacteroidetes phylum. Our main hypothesis is that the heme acquisition mechanism, with the leading role played by a novel HmuY family of hemophore-like proteins, can be used to fulfill nutritional requirements and increase virulence. We characterized HmuY homologs expressed by Bacteroides fragilis and compared their properties with the first representative of this family, the HmuY protein of Porphyromonas gingivalis. In contrast to other Bacteroidetes members, B. fragilis produces three HmuY homologs (Bfr proteins). All bfr transcripts were produced at higher levels in bacteria starved of iron and heme (fold change increase ~60, ~90, and ~70 for bfrA, bfrB, and bfrC, respectively). X-ray protein crystallography showed that B. fragilis Bfr proteins are structurally similar to P. gingivalis HmuY and to other homologs, except for differences in the potential heme-binding pockets. BfrA binds heme, mesoheme, and deuteroheme, but preferentially under reducing conditions, using Met175 and Met146 to coordinate heme iron. BfrB binds iron-free protoporphyrin IX and coproporphyrin III, whereas BfrC does not bind porphyrins. HmuY is capable of heme sequestration from BfrA, which might increase the ability of P. gingivalis to cause dysbiosis also in the gut microbiome.

Association between colorectal cancer, the frequency of Bacteroides fragilis, and the level of mismatch repair genes expression in the biopsy samples of Iranian patients.

BACKGROUND: Deficient DNA mismatch repair (MMR) can cause microsatellite instability (MSI) and is more common in colorectal cancer (CRC) patients. Understanding the carcinogenic mechanism of bacteria and their impact on cancer cells is crucial. Bacteroides fragilis (B. fragilis) has been identified as a potential promoter of tumorigenesis through the alteration of signaling pathways. This study aims to assess the expression levels of msh2, msh6, mlh1, and the relative frequency of B. fragilis in biopsy samples from CRC patients. MATERIALS AND METHODS: Based on the sequence of mlh1, msh2, and msh6 genes, B. fragilis specific 16srRNA and bacterial universal 16srRNA specific primers were selected, and the expression levels of the target genes were analyzed using the Real-Time PCR method. RESULTS: Significant increases in the expression levels of mlh1, msh2, and msh6 genes were observed in the cancer group. Additionally, the expression of these MMR genes showed a significant elevation in samples positive for B. fragilis presence. The relative frequency of B. fragilis in the cancer group demonstrated a significant rise compared to the control group. CONCLUSION: The findings suggest a potential correlation between the abundance of B. fragilis and alterations in the expression of MMR genes. Since these genes can play a role in modifying colon cancer, investigating microbial characteristics and gene expression changes in CRC could offer a viable solution for CRC diagnosis.

Association between Bacteroides fragilis and Fusobacterium nucleatum infection and colorectal cancer in Vietnamese patients.

BACKGROUND: Colorectal cancer (CRC) is a significant global health concern, and understanding the role of specific bacterial infections in its development and progression is of increasing interest. This cross-sectional study investigated the associations between Bacteroides fragilis (B. fragilis) and Fusobacterium nucleatum (F. nucleatum) infections and Vietnamese CRC patients. METHODS: 192 patients with either polyps or CRC at varying stages were recruited from May 2017 to December 2020. Real-time PCR assessed infection rates and bacterial loads in CRC tissues. RESULTS: B. fragilis infection was notably higher in CRC tissues (51.6 %) than polyps (9.4 %), with a fivefold higher relative load. Positive associations were found in stages II and III, indicating a fivefold increase in CRC progression risk. F. nucleatum infection rates were significantly higher in CRC tissues (55.2 %) than in polyps (10.5 %). In stage II, the infection rate exceeded that in adjacent tissues. The relative load of F. nucleatum was higher in stage III than in stages I and II. Positive F. nucleatum patients had a 3.2 times higher risk of CRC progression. CONCLUSION: These findings suggest associations between loading of F. nucleatum or/and B. fragilis with the advanced stages of CRC.

Bacteroides fragilis spondylitis after suspected oral ulcer: a clinical case and comprehensive literature review.

This paper reports a case of Bacteroides fragilis induced spondylitis. Diagnosis was confirmed through blood culture and metagenomic sequencing of pus for pathogen detection. Due to persistent lumbar pain, surgical intervention became imperative, resulting in favorable postoperative outcomes. A detailed patient history revealed a severe episode of oral ulceration two weeks before symptom onset, although a direct link to the infection remained elusive. Leveraging insights from this case, we conducted a comprehensive literature review on B. fragilis spondylitis, elucidating clinical manifestations, diagnostic methodologies, and therapeutic strategies.

Novel and rare ß-lactamase genes of Bacteroides fragilis group species: Detection of the genes and characterization of their genetic backgrounds.

OBJECTIVES: This study screened the prevalence of rare ß-lactamase genes in Bacteroides fragilis group strains from clinical specimens and normal microbiota and examined the genetic properties of the strains carrying these genes. METHODS: blaHGD1, blaOXA347, cblA, crxA, and pbbA were detected by real-time polymerase chain reaction in collections of Bacteroides strains from clinical (n = 406) and fecal (n = 184) samples. To examine the genetic backgrounds of the samples, end-point PCR, FT-IR, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry were used. RESULTS: All B. uniformis isolates were positive for cblA in both collections. Although crxA was B. xylanisolvens-specific and associated with carbapenem resistance, it was only found in six fecal and three clinical B. xylanisolvens strains. Moreover, the crxA-positive strains were not clonal among B. xylanisolvens (contrary to cfiA in B. fragilis), implicating a rate of mobility or emergence by independent evolutionary events. The Phocaeicola (B.) vulgatus/P. dorei-specific gene blaHGD1 was detected among all P. vulgatus/P. dorei isolates from fecal (n = 36) and clinical (n = 26) samples. No blaOXA347-carrying isolate was found from European collections, but all US samples (n = 6) were positive. For three clinical isolates belonging to B. thetaiotaomicron (n = 2) and B. ovatus (n = 1), pbbA was detected on mobile genetic elements, and pbbA-positive strains displayed non-susceptibility to piperacillin or piperacillin/tazobactam phenotypically. CONCLUSIONS: Based on these observations, ß-lactamases produced by rare ß-lactamase genes in B. fragilis group strains should not be overlooked because they could encode important resistance phenotypes.

A randomized trial of Bacteroides fragilis 839 on preventing chemotherapy-induced myelosuppression and gastrointestinal adverse effects in breast cancer patients.

BACKGROUND AND OBJECTIVES: To evaluate the potential benefits of Bacteroides fragilis 839 (BF839), a next-generation probiotics, in reducing myelosuppression and gastrointestinal toxicity associated with chemotherapy in breast cancer patient. METHODS AND STUDY DESIGN: 40 women with early breast cancer were randomly assigned to the BF839 (n=20) or placebo (n=20) during the administration of adjuvant chemotherapy (4 cycles of epirubicin 100mg/m2 and cyclophosphamide 600mg/m2). Myelosuppression and gastrointestinal adverse effects were monitored in both groups. RESULTS: Throughout the four treatment cycles, the percentage of patients experiencing myelosuppression was 42.5% in the BF839 group, significantly lower than the 66.3% observed in the control group (p=0.003). Two patients in the BF839 group and three patients in the placebo group received recombinant human granulocyte colony-stimulating factor (rhG-CSF) due to leuko-penia/neutropenia. When considering an ITT analysis, which included all patients regardless of rhG-CSF treatment, the BF839 group exhibited less reduction from baseline in white blood cells (-0.31±1.19 vs -1.15±0.77, p=0.012) and neutrophils (0.06±1.00 vs -0.84±0.85, p=0.004) compared to the placebo group. The difference became even more significant when excluding the patients who received rhG-CSF injections. Throughout the four treatment cycles, compared to the placebo group, the BF839 group had significantly lower rates of 3-4 grade nausea (35.0% vs 71.3%, p=0.001), vomiting (20.0% vs 45.0%, p=0.001), and diarrhea (15.0% vs 30.0%, p=0.023). CONCLUSIONS: These findings suggest that BF839 has the potential to effectively mitigate myelosuppression and gastrointestinal toxicity associated with chemotherapy in breast cancer patients.

Control the carcinogenic bacteria with new polysaccharides from agricultural wastes.

Agriculture activities industries produce a huge amount of waste every year. Agricultural wastes are a great source of natural polysaccharides characterized by accessibility, biocompatibility, and ease of modification. Finding new safe antibacterial agents has become one of the top priorities of health organizations worldwide. This priority emerged from the antibiotic resistance pathogenic bacteria hazard. Carcinogenic bacteria are one of the most dangerous antibiotic-resistant pathogenic bacteria. This study tries to investigate the antibacterial activity of polysaccharides from some agricultural wastes against carcinogenic bacteria related to gastrointestinal cancers. We determined the antibacterial activity (in terms of minimum inhibitory concentration (MIC)) and the biofilm reduction capacity. We studied the mechanism of the antibacterial activity by determining the effect of the MIC of the extracted polysaccharides on the plasma membrane permeability and the bacterial DNA content. All extracted polysaccharides showed effective antibacterial activity with low MICs ranging from 2 to 20 µg/mL. The barely straw polysaccharides showed the highest MIC (19.844 µg/mL) against Bacteroides fragilis, while the grape bagasse showed the lowest MIC (2.489 µg/mL) against Helicobacter pylori. The extracted polysaccharide showed high antibiofilm activity. Their capacity to reduce the formation of the pathogenic biofilm ranged from 75 to 95%. Regarding the antibacterial mechanism, the extracted polysaccharides showed destructive action on the DNA and the plasma membrane permeability. The bacterial DNA change percent after the treatment with the different polysaccharides ranged from 29% to -58%. The plasma membrane permeability increased by a high percentage, ranging from 92% to 123%. Agricultural waste polysaccharides are a promising antibacterial agent against antibiotic-resistant carcinogenic bacteria.

Phenotypic and genotypic identification of carbapenem resistance in Bacteroides fragilis clinical strains.

Bacteroides fragilis is an important etiological agent of serious infections in humans. Rapid methods, readily adaptable to use in medical laboratories, are needed to detect antibiotic resistance and decrease the likelihood of therapy failure. The aim of this study was to determine the prevalence of B. fragilis cfiA-positive isolates. The second purpose was to investigate the carbapenemase activity in B. fragilis strains by Carba NP test. In the study, 5.2% of B. fragilis isolates are phenotypically resistant to meropenem. The cfiA gene was identified in 6.1% of B. fragilis isolates. The MICs of meropenem were significantly higher in cfiA-positive strains. The presence of the cfiA gene along with the IS1186 was detected in one B. fragilis strain which was resistant to meropenem (MIC 1.5 mg/L). The Carba NP test results were positive for all the cfiA-positive strains, including those susceptible to carbapenems based on their MIC values. A review of the literature revealed that the rate of B. fragilis with the cfiA gene varies from 7.6 to 38.9% worldwide. Presented results are in line with the other European studies. Phenotypic testing with the Carba NP test, it seems to be a viable alternative for the cfiA gene detection in B. fragilis isolates. The positive result obtained is of greater clinical importance than the detection of the gene cfiA.

Prevalence of antimicrobial resistant genes in Bacteroides spp. isolated in Oita Prefecture, Japan.

INTRODUCTION: Bacteroides spp. are the most common anaerobic bacteria isolated from the human gastrointestinal tract. Several resistant genes are present in Bacteroides spp. However, most studies have focused on the prevalence of the cfiA gene in Bacteroides fragilis alone. We assessed the susceptibility to antimicrobial agents and the prevalence of cepA, cfiA, cfxA, ermF, nim, and tetQ genes in Bacteroides strains isolated from clinical specimens in our hospital. METHODS: We isolated 86 B. fragilis and 58 non-fragilis Bacteroides strains from human clinical specimens collected from January 2011 to November 2021. Resistance against piperacillin (PIPC), cefotaxime (CTX), cefepime (CFPM), meropenem (MEPM), clindamycin, and minocycline was determined. RESULTS: The resistant rates of penicillins and cephalosporins in non-fragilis isolates were significantly higher than those in B. fragilis isolates. In B. fragilis isolates, the resistant rates of PIPC, CTX, and CFPM in cfxA-positive isolates were significantly higher than those in cfxA-negative isolates (71% vs. 16%, 77% vs. 19%, and 77% vs. 30%, respectively). Thirteen B. fragilis isolates harbored the cfiA gene, two of which were resistant to MEPM. Six of the 13 cfiA-positive B. fragilis isolates were heterogeneously resistant to MEPM. CONCLUSION: It is important to evaluate the use of MEPM as empirical therapy for Bacteroides spp. infections, considering the emergence of carbapenem resistance during treatment, existence of MEPM-resistant strains, and heterogeneous resistance.

Molecular study on metronidazole resistance in Bacteroides fragilis group isolates from a South Indian tertiary care center.

OBJECTIVE: Bacteroides species are an important part of human intestinal microbiota. They can cause infections of significant mortality and morbidity when moved out of their niche in the gut. The cornerstone drug for prophylaxis and therapy, metronidazole, is exhibiting signs of resistance, which are frequently attributed to nitroimidazole (nim) resistance genes. The aim of this study was to use Epsilometer test (E-test) to assess the metronidazole susceptibility and conventional PCR methodology to map the distribution of nim genes in Bacteroides fragilis group (BFG) isolates. METHODS: MALDI-TOF MS was used to identify BFG isolates. Using the E-test methodology, metronidazole minimum inhibitory concentrations (MICs) were determined. The presence of nim genes in these isolates were checked by conventional PCR methodology. Sequencing was done on selected amplicons for determining the nim gene types. RESULTS: Bacteroides fragilis accounted for 55.3% of the total 273 BFG members identified. Of these, 196 (71.8%) were susceptible, 43 (15.8%) intermediate and 34 (12.5%) resistant to metronidazole as determined by the E-test. nim gene was present in 101 (37%) of the total 273 isolates. Out of the 34 phenotypically resistant isolates (MIC ≥32 µg/ml), 29 harboured nim gene (Chi-square test, p < 0.0000001) but nim gene was absent in 5 (14.7%) isolates. Also, nim gene was detected in 72 (30.1%) of the 239 isolates with susceptible and intermediate metronidazole MIC. Sequencing of 20 amplicons gave a nimE gene type. CONCLUSIONS: In view of the rising metronidazole resistance among BFG and its close association with nim genes, there is a need for implementing routine metronidazole susceptibility testing and more researches are needed to find the molecular basis of these nim genes.

High prevalence of cfiA positive Bacteroides fragilis isolates collected at a teaching hospital in Hohhot, China.

OBJECTIVES: Carbapenem-resistant Bacteroides fragilis has emerged globally and cfiA is the key underlying factor. However, the prevalence of cfiA-positive carbapenem-resistant B. fragilis varies among countries. Therefore, we investigated the prevalence of cfiA-positive B. fragilis clinical isolates in a tertiary hospital in China. METHODS: Carbapenem-resistant cfiA-positive B. fragilis isolates were identified using polymerase chain reaction. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to identify the characteristic mass spectra of cfiA-positive B. fragilis. RESULTS: The prevalence of cfiA among 153 B. fragilis isolates was 22.2% (34/153), when 20.6% (7/34) cfiA-positive B. fragilis strains were isolated from pediatric patients. Twenty-one carbapenem-resistant B. fragilis isolates were identified and were all positive with cfiA gene. Two characteristic peaks (4825 and 9642 Da) were identified using MALDI-TOF MS, and the sensitivity, specificity, and both the positive and negative predictive values of these two peaks were 100%. A new peak shift from 9627 Da for cfiA-negative isolates to 9642 Da for cfiA-positive isolates was observed. CONCLUSIONS: A high prevalence of cfiA was observed among B.fragilis isolates in this study, especially those isolated from pediatric patients. Characteristic MS spectra can accurately discriminate cfiA-positive and -negative B. fragilis isolates and can contribute to the rapid screening of cfiA-positive B. fragilis isolates in clinical laboratories.

Antimicrobial susceptibility testing of anaerobic bacteria causing bacteremia: A 13-year (2010-2022) retrospective study in a tertiary hospital.

Infections from anaerobic microorganisms result from breached mucosal barriers, posing a significant mortality risk. A retrospective study at Hospital Universitario La Paz (Madrid) from 2010 to 2022 analyzed 491 (6.17 %) anaerobic bacteremia cases out of 7956 significant bacteremia cases among 171,833 blood culture requests. Bacteroides fragilis was the most frequently isolated species (28.3 %), followed by Clostridium perfringens (13.6 %). B. fragilis showed good susceptibility to amoxicillin/ clavulanic acid (86 %), piperacillin/tazobactam (86 %), and metronidazole (87.7 %). In general, non-fragilis Bacteroides species showed low susceptibility to penicillin (7 %), amoxicillin (17.5 %), and clindamycin (64.9 %). Of our 13 non-perfringens Clostridium isolates, four exhibited resistance to penicillin and four showed resistance to clindamycin. Lactobacillus species were highly susceptible to antibiotics tested. Prevotella spp. showed low susceptibility to penicillin (20 %), amoxicillin (20 %), and clindamycin (40 %). The study contributes valuable data for monitoring and improving anaerobic bacteremia treatment.

Clinical significance of Bacteroides fragilis as a potential prognostic factor in colorectal cancer.

INTRODUCTION: Bacteroides fragilis (B. fragilis) is considered to act in an anti-inflammatory manner on the intestinal tract. On the contrary, enterotoxigenic B. fragilis (ETBF), a subtype of B. fragilis, produces an enterotoxin (BFT; B. fragilis toxin), leading to asymptomatic chronic infections and colonic tumor formation. However, the impact of B. fragilis and ETBF on the clinical outcome of colorectal cancer (CRC) remains unclear. We aim to assess whether their presence affects the outcome in patients with CRC after curative resection. METHODS: We obtained 197 pairs of matched formalin-fixed paraffin-embedded samples from cancerous and adjacent non-cancerous tissues of patients with pathological stage (pstage) II and III CRC after curative resection. The presence of B. fragilis and ETBF were estimated using real-time polymerase chain reaction, and recurrence-free survival (RFS) and overall survival (OS) of the patients were analyzed. RESULTS: 16S rRNA for B. fragilis and bft DNA were detected in 120 (60.9%) and 12 (6.1%) of the 197 patients, respectively. B. fragilis-positive patients had better RFS than B. fragilis-negative patients, although that was not statistically significant. In subgroup analysis, better outcomes on RFS were observed in the presence of B. fragilis in pstage II and left-sided CRC. The association of B. fragilis positivity on OS was accentuated in the depth of T4 subgroup. No significant differences were observed in RFS and OS between ETBF and non-toxigenic B. fragilis. CONCLUSIONS: Our findings suggest that the presence of B. fragilis is associated with better outcomes in patients with pstage II and III CRC after curative resection.

Characterization of mobile genetic elements in multidrug-resistant Bacteroides fragilis isolates from different hospitals in the Netherlands.

OBJECTIVES: Five human clinical multidrug-resistant (MDR) Bacteroides fragilis isolates, including resistance to meropenem and metronidazole, were recovered at different hospitals in the Netherlands between 2014 and 2020 and sent to the anaerobic reference laboratory for full characterization. METHODS: Isolates were recovered from a variety of clinical specimens from patients with unrelated backgrounds. Long- and short-read sequencing was performed, followed by a hybrid assembly to study the presence of mobile genetic elements (MGEs) and antimicrobial resistance genes (ARGs). RESULTS: A cfxA gene was present on a transposon (Tn) similar to Tn4555 in two isolates. In two isolates a novel Tn was present with the cfxA gene. Four isolates harbored a nimE gene, located on a pBFS01_2 plasmid. One isolate contained a novel plasmid carrying a nimA gene with IS1168. The tetQ gene was present on novel conjugative transposons (CTns) belonging to the CTnDOT family. Two isolates harbored a novel plasmid with tetQ. Other ARGs in these isolates, but not on an MGE, were: cfiA, ermF, mef(EN2), and sul2. ARGs harboured differed between isolates and corresponded with the observed phenotypic resistance. CONCLUSIONS: Novel CTns, Tns, and plasmids were encountered in the five MDR B. fragilis isolates, complementing our knowledge on MDR and horizontal gene transfer in anaerobic bacteria.

Promotion of Colitis in B Cell-Deficient C57BL/6 Mice Infected with Enterotoxigenic Bacteroides fragilis.

Enterotoxigenic Bacteroides fragilis (ETBF) causes colitis and is implicated in inflammatory bowel diseases and colorectal cancer. The ETBF-secreted B. fragilis toxin (BFT) causes cleavage of the adherence junction, the E-cadherin, resulting in the large intestine showing IL-17A inflammation in wild-type (WT) mice. However, intestinal pathology by ETBF infection is not fully understood in B-cell-deficient mice. In this study, ETBF-mediated inflammation was characterized in B-cell-deficient mice (muMT). WT or muMT C57BL/6J mice were orally inoculated with ETBF and examined for intestinal inflammation. The indirect indicators for colitis (loss of body weight and cecum weight, as well as mortality) were increased in muMT mice compared to WT mice. Histopathology and inflammatory genes (Nos2, Il-1ß, Tnf-α, and Cxcl1) were elevated and persisted in the large intestine of muMT mice compared with WT mice during chronic ETBF infection. However, intestinal IL-17A expression was comparable between WT and muMT mice during infection. Consistently, flow cytometry analysis applied to the mesenteric lymph nodes showed a similar Th17 immune response in both WT and muMT mice. Despite elevated ETBF colonization, the ETBF-infected muMT mice showed no histopathology or inflammation in the small intestine. In conclusion, B cells play a protective role in ETBF-induced colitis, and IL-17A inflammation is not attributed to prompted colitis in B-cell-deficient mice. Our data support the fact that B cells are required to ameliorate ETBF infection-induced colitis in the host.

The Intestinal Commensal, Bacteroides fragilis, Modulates Host Responses to Viral Infection and Therapy: Lessons for Exploration during Mycobacterium tuberculosis Infection.

The gut microbiota has emerged as a critical player in host health. Bacteroides fragilis is a prominent member of the gut microbiota within the phyla Bacteroidetes. This commensal bacterium produces unique capsular polysaccharides processed by antigen-presenting cells and activates CD4+ T cells to secrete inflammatory cytokines. Indeed, due to their immunomodulatory functions, B. fragilis and its capsular polysaccharide-A (PSA) are arguably the most explored single commensal microbiota/symbiotic factor. B. fragilis/PSA has been shown to protect against colitis, encephalomyelitis, colorectal cancer, pulmonary inflammation, and asthma. Here, we review recent data on the immunomodulatory role of B. fragilis/PSA during viral infections and therapy, B. fragilis PSA's dual ability to mediate pro-and anti-inflammatory processes, and the potential for exploring this unique characteristic during intracellular bacterial infections such as with Mycobacterium tuberculosis. We also discuss the protective roles of single commensal-derived probiotic species, including B. fragilis in lung inflammation and respiratory infections that may provide essential cues for possible exploration of microbiota based/augmented therapies in tuberculosis (TB). Available data on the relationship between B. fragilis/PSA, the immune system, and disease suggest clinical relevance for developing B. fragilis into a next-generation probiotic or, possibly, the engineering of PSA into a potent carbohydrate-based vaccine.

A tale of two habitats: Bacteroides fragilis, a lethal pathogen and resident in the human gastrointestinal microbiome.

Bacteroides fragilis is an obligately anaerobic Gram-negative bacterium and a major colonizer of the human large colon where Bacteroides is a predominant genus. During the growth of an individual clonal population, an astonishing number of reversible DNA inversion events occur, driving within-strain diversity. Additionally, the B. fragilis pan-genome contains a large pool of diverse polysaccharide biosynthesis loci, DNA restriction/modification systems and polysaccharide utilization loci, which generates remarkable between-strain diversity. Diversity clearly contributes to the success of B. fragilis within its normal habitat of the gastrointestinal (GI) tract and during infection in the extra-intestinal host environment. Within the GI tract, B. fragilis is usually symbiotic, for example providing localized nutrients for the gut epithelium, but B. fragilis within the GI tract may not always be benign. Metalloprotease toxin production is strongly associated with colorectal cancer. B. fragilis is unique amongst bacteria; some strains export a protein >99 % structurally similar to human ubiquitin and antigenically cross-reactive, which suggests a link to autoimmune diseases. B. fragilis is not a primary invasive enteric pathogen; however, if colonic contents contaminate the extra-intestinal host environment, it successfully adapts to this new habitat and causes infection; classically peritoneal infection arising from rupture of an inflamed appendix or GI surgery, which if untreated, can progress to bacteraemia and death. In this review selected aspects of B. fragilis adaptation to the different habitats of the GI tract and the extra-intestinal host environment are considered, along with the considerable challenges faced when studying this highly variable bacterium.

Development and Validation of a Novel Anaerobic Carbapenem Inactivation Method (Ana-CIM) for the Detection of Carbapenemase Production in Bacteroides fragilis.

Antibiotic resistance, particularly to carbapenems, is of increasing concern in Bacteroides fragilis. Carbapenem resistance in B. fragilis is most often mediated by the activation of chromosomally encoded metallo-ß-lactamase cfiA by the presence of an upstream insertion sequence (IS). While traditional phenotypic susceptibility methods and molecular tests to detect carbapenem resistance in B. fragilis exist, they are not available in most clinical microbiology laboratory settings. Here, we describe the development of the anaerobic carbapenem inactivation method (Ana-CIM) for predicting carbapenemase production in B. fragilis based off the principles of the well-established modified carbapenem inactivation method (mCIM) for Enterobacterales and Pseudomonas aeruginosa. We also present the clinical validation and reproducibility of the Ana-CIM at three clinical laboratory sites (with 60 clinical isolates, 45% ertapenem resistant). Compared to ertapenem susceptibility by Etest interpreted by CLSI M100 Ed30, the Ana-CIM accurately detected carbapenem resistance in B. fragilis with categorical agreement (CA) of 87% (52/60) and 0% (0/21) very major error (VME), 11% (4/36) major error (ME), and 7% (4/60) minor error (mE) rates across all sites. Additionally, the Ana-CIM demonstrated high reproducibility with 5 clinical and 3 quality control (QC) isolates tested in triplicate with 3 commercial Mueller-Hinton media across all sites, with 93% (604/648) of replicates within a 2-mm zone size of the mode for each isolate. We conclude that the Ana-CIM can be readily deployed in clinical laboratories at a low cost for detection of carbapenemase-mediated resistance in B. fragilis.

Molecular recognition and binding of CcrA from Bacteroides fragilis with cefotaxime and ceftazidime by fluorescence spectra and molecular docking.

In search of new super-bacterial inhibitor agents, the recognition and binding mechanism of the B1 subclass MßL CcrA from Bacteroides fragilis with cefotaxime (CTX) and ceftazidime (CAZ) were studied using spectroscopy analysis and molecular docking. The results showed that the fluorescence quenching of CcrA induced by CTX and CAZ were all due to the complex formation, which belonged to static quenching and was forced by hydrogen bonds and Van der Waals forces, despite the greater binding ability of CTX with CcrA than CAZ. Upon recognizing CTX or CAZ, the CcrA opened its binding pocket by the microenvironmental and conformational of three loops changing to promote an induced-fit of the freshly introduced antibiotics. In addition, the whole antibiotic molecule ultimately entered the active pocket of CcrA with its original carbonate replaced by the carboxyl oxygen of the hexatomic ring adjacent to the ß-lactam ring in CTX or CAZ, forming a new tetrahedral coordination structure at the Zn2 site. Moreover, the difference in steric hindrance and electrostatic effects of the side chain affected the binding ability of the two antibiotics to the CcrA. This work showed the refined procedures of the antibiotics binding to CcrA and might provide useful information hint for the new strategy of developing the novel and innovative super-bacterial antibiotics.