A Distinct, Flocculent, Acidogenic Microbial Community Accompanies Methanogenic Granules in Anaerobic Digesters.

The formation of dense, well-settling methanogenic granules is essential for the operation of high-rate, up-flow anaerobic bioreactors used for wastewater treatment. Granule formation (granulation) mechanisms have been previously proposed, but an ecological understanding of granule formation is still lacking. Additionally, much of the current research on granulation only examines the start-up phase of bioreactor operation, rather than monitoring the fate of established granules and how new granules emerge over time. This paper, therefore, attempts to provide an insight into the microbial ecology of granule formation outside the start-up phase of bioreactor operation and develop an ecological granulation model. The microbial communities of granules actively undergoing growth, breakage, and reformation were examined, and an ecological granulation model was proposed. A distinct pregranular microbial community, with a high proportion of acidogenic organisms, such as the Streptococcaceae, was identified and suggested to have a role in initiating granulation by providing simpler substrates for the methanogenic and syntrophic communities which developed during granule growth. After initial granule formation, deterministic influences on microbial community assembly increased with granule size and indicated that microbial community succession was influenced by granule growth, leading to the formation of a stepwise ecological model for granulation. IMPORTANCE Complex microbial communities in engineered environments can aggregate to form surface-attached biofilms. Others form suspended biofilms, such as methanogenic granules. The formation of dense, methanogenic granules underpins the performance of high-rate, anaerobic bioreactors in industrial wastewater treatment. Granule formation (granulation) has been well studied from a physico-chemical perspective, but the ecological basis is poorly understood. We identified a distinct, flocculent, microbial community, which was present alongside granules, comprising primary consumers likely key in providing simpler substrates to granules. This flocculent community is understudied in anaerobic digestion and may initiate, or perpetuate, granule formation. We propose that it may be possible to influence bioreactor performance (e.g., to regulate volatile fatty acid concentrations) by manipulating this community. The patterns of microbial community diversity and assembly revealed by the study indicate that cycles of granule growth and breakage lead to overall diversification of the bioreactor meta-community, with implications for bioreactor process stability.

Dried Protein Structure Revealed at the Residue Level by Liquid-Observed Vapor Exchange NMR.

Water is key to protein structure and stability, yet the relationship between protein-water interactions and structure is poorly understood, in part because there are few techniques that permit the study of dehydrated protein structure at high resolution. Here, we describe liquid-observed vapor exchange (LOVE) NMR, a solution NMR-based method that provides residue-level information about the structure of dehydrated proteins. Using the model protein GB1, we show that LOVE NMR measurements reflect the fraction of the dried protein population trapped in a conformation where a given residue is protected from exchange with D2O vapor. Comparisons to solution hydrogen-deuterium exchange data affirm that the dried protein structure is strongly influenced by local solution stability and that the mechanism of dehydration protection exerted by the widely used protectant trehalose differs from its mechanism of stabilization in solution. Our results highlight the need for refined models of cosolute-mediated dehydration protection and demonstrate the ability of LOVE NMR to inform such models.

The influences of low protein diet on the intestinal microbiota of mice.

Recent research suggests that protein deficiency symptoms are influenced by the intestinal microbiota. We investigated the influence of low protein diet on composition of the intestinal microbiota through animal experiments. Specific pathogen-free (SPF) mice were fed one of four diets (3, 6, 9, or 12% protein) for 4 weeks (n = 5 per diet). Mice fed the 3% protein diet showed protein deficiency symptoms such as weight loss and low level of blood urea nitrogen concentration in their serum. The intestinal microbiota of mice in the 3% and 12% protein diet groups at day 0, 7, 14, 21 and 28 were investigated by 16S rRNA gene sequencing, which revealed differences in the microbiota. In the 3% protein diet group, a greater abundance of urease producing bacterial species was detected across the duration of the study. In the 12% diet protein group, increases of abundance of Streptococcaceae and Clostridiales families was detected. These results suggest that protein deficiency may be associated with shifts in intestinal microbiota.

Comparison of microbiomes in ulcerative and normal mucosa of recurrent aphthous stomatitis (RAS)-affected patients.

BACKGROUND: Recurrent aphthous stomatitis (RAS) is the most common form of oral ulcerative disease, whose cause is still unknown. Researchers have found the association of many factors with the occurrence of RAS, and proposed oral bacterial infection could be a cause for this disease. METHODS: To investigate whether the occurrence of RAS is associated with oral bacterial infection, we performed high throughput sequencing analysis of bacterial samples collected from the normal oral mucosa and aphthous ulcers of 24 patients. RESULTS: Firmicutes, Proteobacteria and Bacteriodetes were the most abundant phyla in the microbiomes analysed. The alpha diversities of the oral mucosa and aphthous ulcer microbiomes were similar, suggesting a similar richness and diversity. The NMDS analysis showed the oral mucosa and aphthous ulcer microbiomes are significantly different. This suggestion is further supported by Anosim, MRPP, and Adonis analyses. More detailed comparison of the two groups of microbiomes suggested that the occurrence of RAS is significantly associated with the increase of Escherichia coli and Alloprevotella, as well as the decrease of Streptococcus. CONCLUSIONS: Considering E. coli is a very common intestinal bacterium, we propose that E. coli colonization could be a cause for RAS, and controlling E. coli colonization could help curing RAS.

Partial rpoB Gene Sequencing Identification and Probiotic Potential of Floricoccus penangensis ML061-4 Isolated from Assam Tea (Camellia sinensis var. assamica).

Assam tea or Miang is a local name of Camellia sinensis var. assamica in northern Thailand. By the local wisdom, Assam tea leaves are used as the raw material in tea fermentation to produce "Fermented Miang" consumed by people in northern Thailand and the countries nearby. In this study, twenty-eight bacterial isolates were obtained from Assam tea leaf samples collected from Nan province, Thailand. Bacterial isolates were identified within 6 genera including Bacillus, Floricoccus, Kocuria, Lysinibacillus, Micrococcus and Staphylococcus. Among these, the strain ML061-4 shared 100.0 and 99.4% similarity of 16S rRNA and rpoB gene sequence with F. penangensis JCM 31735T, respectively. This is the first discovery of F. penangensis in Thailand. F. penangensis ML061-4 exhibited probiotic characteristics including lactic acid production (9.19 ± 0.10 mg/ml), antibacterial activities (Escherichia coli ATCC 25922 and E. coli O157:H7 DMST 12743), acid and bile salt tolerance (71.1 and 54.9%, respectively), autoaggregation (97.0%), coaggregation (66.0% with E. coli O157:H7), cell surface hydrophobicity (90.0%), bacterial adhesion (82.9% with Lactobacillus plantarum FM03-1), competitive inhibition (17.8% with E. coli O157:H7) and competitive exclusion (34.9% with E. coli O157:H7). Overall, the data suggested that F. penangensis ML061-4 had a great potential to be a probiotic.

Novel Image Processing Method for Detecting Strep Throat (Streptococcal Pharyngitis) Using Smartphone.

In this paper, we propose a novel strep throat detection method using a smartphone with an add-on gadget. Our smartphone-based strep throat detection method is based on the use of camera and flashlight embedded in a smartphone. The proposed algorithm acquires throat image using a smartphone with a gadget, processes the acquired images using color transformation and color correction algorithms, and finally classifies streptococcal pharyngitis (or strep) throat from healthy throat using machine learning techniques. Our developed gadget was designed to minimize the reflection of light entering the camera sensor. The scope of this paper is confined to binary classification between strep and healthy throats. Specifically, we adopted k-fold validation technique for classification, which finds the best decision boundary from training and validation sets and applies the acquired best decision boundary to the test sets. Experimental results show that our proposed detection method detects strep throats with 93.75% accuracy, 88% specificity, and 87.5% sensitivity on average.

Alterations in Skin Microbiomes of Patients With Cirrhosis.

BACKGROUND & AIMS: Patients with cirrhosis have intestinal dysbiosis and are prone to itching and skin or soft-tissue infections. The skin microbiome, and its relationship with intestinal microbiome, have not been characterized. We investigated alterations in skin microbiota of patients with cirrhosis and their association with intestinal microbiota and modulators of itch. METHODS: We collected skin swabs at 7 sites and blood and stool samples from 20 healthy individuals (control subjects; mean age, 59 years) and 50 patients with cirrhosis (mean age, 61 years; mean model for end-stage disease score, 12; 20 with decompensation). Skin and stool samples were analyzed by 16s rRNA sequencing and serum samples were analyzed by liquid chromatography and mass spectrometry for levels of bile acids (BAs) and by an ELISA for autotaxin (an itch modulator). Participants were analyzed by the visual analog itch scale (VAS, 0-10,10 = maximum intensity). Data were compared between groups (cirrhosis vs control subjects, with vs without decompensation, VAS 5 or higher vs less than 5). Correlation networks between serum levels of BAs and skin microbiomes were compared between patients with cirrhosis with vs without itching. RESULTS: The composition of microbiomes at all skin sites differed between control subjects and patients with cirrhosis and between patients with compensated vs decompensated cirrhosis. Skin microbiomes of patients with cirrhosis (especially those with decompensation) contained a higher relative abundance of Gammaproteobacteria, Streptococaceae, and Staphylococcaceae, and fecal microbiomes contained a higher relative abundance of Gammaproteobacteria, than control subjects. These bacterial taxa were associated with serum levels of autotaxin and BAs, which were higher in patients with VAS scores ≥5. Based on network statistics, microbial and BA interactions at all sites were more complex in patients with greater levels of itching in the shin, the most common site of itch. CONCLUSIONS: We identified alterations in skin microbiome of patients with cirrhosis (in Gammaproteobacteria, Streptococcaceae, and Staphylococcaceae)-especially in patients with decompensation; fecal microbiomes of patients with cirrhosis had a higher relative abundance of Gammaproteobacteria than control subjects. These specific microbial taxa are associated with itching intensity and itch modulators, such as serum levels of BAs and autotaxin.

Addition of wood sawdust during the co-composting of sewage sludge and wheat straw influences seeds germination.

The objective of this study was to investigate the effect of addition of sawdust co-composted with sewage sludge and wheat straw on seeds germination. Two mixtures were piled and composted over 90 days. The first mixture (C1) was composed of sewage sludge and wheat straw, while the second mixture (C2) was composed of sewage sludge, wheat straw and wood sawdust. The results showed that the physicochemical parameters of both composts, i.e., temperature (> 55 °C in the thermophilic phase), moisture content (~ 30%). pH (6.73 for C1 and 7.19 for C2) and EC values (1.81 mS cm-1 for C1 and 1.32 mS cm-1 for C2) reached the required maturity standard. The values of C/N were below 12 indicating a high degree of maturity. Also, no bacterial pathogens were detected in the final composts. The concentration of total heavy metals has been reduced allowing the elimination of sewage sludge toxicity, confirmed by the germination index, which reached over 80%. Strong positive correlations were noticed between total Kjeldahl nitrogen of C2 and germination indexes of all the studied species. The obtained results indicate that the addition of wood sawdust increases the nitrogen content leading to slightly alkaline compost which influences seeds germination by reducing the phytotoxicity of sewage sludge.

Investigation of the viral and bacterial microbiota in intestinal samples from mink (Neovison vison) with pre-weaning diarrhea syndrome using next generation sequencing.

Pre-weaning diarrhea (PWD) in mink kits is a common multifactorial syndrome on commercial mink farms. Several potential pathogens such as astroviruses, caliciviruses, Escherichia coli and Staphylococcus delphini have been studied, but the etiology of the syndrome seems complex. In pooled samples from 38 diarrheic and 42 non-diarrheic litters, each comprising of intestinal contents from 2-3 mink kits from the same litter, the bacterial populations were studied using Illumina Next Generation Sequencing technology and targeted 16S amplicon sequencing. In addition, we used deep sequencing to determine and compare the viral intestinal content in 31 healthy non-diarrheic and 30 diarrheic pooled samples (2-3 mink kits from the same litter per pool). The results showed high variations in composition of the bacterial species between the pools. Enterococci, staphylococci and streptococci dominated in both diarrheic and non-diarrheic pools. However, enterococci accounted for 70% of the reads in the diarrheic group compared to 50% in the non-diarrheic group and this increase was at the expense of staphylococci and streptococci which together accounted for 45% and 17% of the reads in the non-diarrheic and diarrheic group, respectively. Moreover, in the diarrheic pools there were more reads assigned to Clostridia, Escherichia-Shigella and Enterobacter compared to the non-diarrheic pools. The taxonomically categorized sequences from the virome showed that the most prevalent viruses in all pools were caliciviruses and mamastroviruses (almost exclusively type 10). However, the numbers of reads assigned to caliciviruses were almost 3 times higher in the diarrheic pools compared the non-diarrheic pools and Sapporo-like caliciviruses were more abundant than the Norwalk-like caliciviruses. The results from this study have contributed to the insight into the changes in the intestinal microbiota associated with the PWD syndrome of mink.

Curcumin-mediated antimicrobial photodynamic therapy reduces the viability and vitality of infected dentin caries microcosms.

BACKGROUND: To our knowledge, there is a lack of evidence on the effect of Antimicrobial Photodynamic Therapy (aPDT) by the application of curcumin against complex biofilms of dental caries lesions. This study aimed to evaluate the viability, vitality, and acid metabolism of infected dentin caries microcosms treated with curcumin-mediated aPDT. METHODS: After microcosm biofilms growing anaerobically on bovine dentin disks immersed in McBain medium with 1% sucrose at 37 °C for 5 days, the biofilms were treated by the association of DMSO water solution or 600 µmol L-1 curcumin with 0, 37.5 or 75 J cm-2 blue LED (455 nm). Then, the colony-forming units (CFU) counts of total microorganisms, total streptococci, mutans streptococci, and total lactobacilli were determined by plating. The lactic acid concentration was analyzed by enzymatic spectrophotometry method, while the vitality of intact biofilms was evaluated by confocal laser scanning microscope (CLSM). Statistical analysis was performed by Kruskal Wallis and post-hoc Dunn's tests (P < 0.05). RESULTS: Curcumin alone did not affect the viability of microorganisms and the vitality of intact biofilms. However, 75 J cm-2 LED alone decreased the total microorganisms and total lactobacilli counts. The combination of curcumin and LED reduced significantly the counts of all microorganism groups and the vitality of intact biofilms. Differences were not observed between the lactic acid concentrations of distinct groups. CONCLUSIONS: Therefore, curcumin-mediated aPDT was effective in reducing the viability and the vitality of infected dentin caries microcosms, without interfering in their acidogenicity.

Explorative study on mucosal and major salivary secretion rates, caries and plaque microflora in head and neck cancer patients.

OBJECTIVE: To analyse mucosal and major salivary secretion rates, caries and plaque microflora in connection with treatment for cancer in the head and neck region. METHODS: Thirty-three patients were included and the number of teeth, filled surfaces and caries lesions registered. The labial and buccal gland secretion and stimulated whole salivary secretion rates were determined. Supragingival plaque microflora was analysed using cultivation technique. Data were collected pretreatment, during treatment and 6 months, 1 year and 2 years post-treatment. RESULTS: Two years post-treatment, 36% had new caries lesions, which had been restored. The labial secretion was comparable with pretreatment, while the buccal secretion was lower (P < .001). The stimulated secretion rate was lower compared with pretreatment (P < .001) and was ≤0.7 mL/minute for 50%. Growth of lactobacilli increased during treatment (P < .001) and remained increased (P < .001), while growth of mutans streptococci was decreased (P < .01) 2 years post-treatment. Growth of Candida increased over time and was higher 2 years post-treatment compared with pretreatment (P < .001) while growth of Prevotella was lower (P < .01). CONCLUSIONS: Two years post-treatment, the stimulated salivary secretion rate was substantially decreased, acid-tolerant lactobacilli and Candida increased, acid-sensitive microorganisms decreased, and the caries prevalence was low.

Discovery of a long-acting glucagon-like peptide-1 analog with enhanced aggregation propensity.

In the course of our search for new GLP-1 analogs, we screened a number of [Ser8]-GLP-1 analogs using the C-terminal helix 3 of the albumin binding domain 3 of protein G from bacterial Streptococcal G strain 148 (G148-ABD3) as appendage. Our efforts led to the discovery of [Ser8]-GLP-1 (7-35)-GVKALIDEILAA-NH2, peptide 6, as a long-acting GLP-1 analog with enhanced self-associated aggregation. Peptide 6 showed enhanced stability in rat and human plasma and an extended half-life of 5.4 h with good bioavailability in rats and subsequently prolonged therapeutic effects in diabetic mice. Analytical ultracentrifugation and TLC suggest that 6 remains oligomeric in the circulation, which accounts for its extended in vivo half-life. The present work shows the possible enhancement of medium-sized oligopeptides aggregation propensity and highlights the potential advantages of peptide aggregates for long-acting peptide drugs.

[Intestinal Microbiota in Migrating Barn Swallows around Osaka].

Migratory birds are considered as vectors of infectious diseases, owing to their potential for transmitting pathogens over large distances. The populations of barn swallow (Hirundo rustica) migrate from Southeast Asia to the Japanese mainland during spring and migrate back to Southeast Asia during autumn. This migratory population is estimated to comprise approximately hundreds to thousands of individuals per year. However, to date, not much is known about the gastrointestinal microbiota of the barn swallow. In this study, we characterized the fecal bacterial community in barn swallow. Using 16S rRNA gene metagenomic sequencing analysis, we examined the presence and composition of potentially pathogenic bacteria in the fecal samples, which were collected during spring season from Osaka. The number (±S.D.) of total bacteria was approximately 2.1(±3.4)×108 per gram of feces. In most samples, the bacterial community composition was dominated by families, such as Enterobacteriaceae, Pseudomonadaceae, Mycoplasmataceae, Enterococcaceae, Streptococcaceae, and Alcaligenaceae. However, no relationship was found between the bacterial community composition and geographical area in the fecal samples. Potentially pathogenic bacteria were detected at the rate of >0.1%, which included Pseudomonas spp., Escherichia/Shigella spp., Enterobacter spp., Yersinia spp., Mycoplasma spp., Enterococcus spp., Achromobacter spp., and Serratia spp. Our results suggested that barn swallow is instrumental in the transmission of these genera over large distances.

Intestinal Dysbiosis Secondary to Proton-Pump Inhibitor Use.

BACKGROUND: Gut dysbiosis associated with the use of proton-pump inhibitors (PPIs) has been found to lead to the occurrence of infectious and inflammatory adverse events. A longitudinal observational cohort study has demonstrated the heightened risk of death associated with PPI use. SUMMARY: We evaluated meta-analyses to determine the association between PPI use and infectious and inflammatory diseases. Meta-analyses showed that PPI use is a potential risk for the development of enteric infections caused by Clostridium difficile, as well as small intestinal bacterial overgrowth, spontaneous bacterial peritonitis, community-acquired pneumonia, hepatic encephalopathy, and adverse outcomes in inflammatory bowel disease. We also examined changes in the composition and function of the gut microbiota with the use of PPIs. PPI use significantly increased the presence of Streptococcaceae and Enterococcaceae, which are risk factors for C. difficile infection, and decreased that of Faecalibacterium, a commensal anti-inflammatory microorganism. Key Message: High-throughput, microbial 16S rRNA gene sequencing has allowed us to investigate the association between the gut microbiome and PPI use. Future prospective comparison studies are necessary to confirm this association, and to develop new strategies to prevent complications of PPI use.

Floricoccus tropicus gen. nov., sp. nov. and Floricoccus penangensis sp. nov. isolated from fresh flowers of durian tree and hibiscus.

Three strains of Gram-staining-positive, coccus-shaped, lactic acid bacteria, designated as HibF3T, HibF2 and HibF5 were isolated from fresh flowers of hibiscus, and a fourth, DF1T, was isolated from fresh flowers of durian tree, in Penang, Malaysia. Taxonomic characterisation was performed by polyphasic analysis. Sequence similarities of the 16S rRNA gene and the housekeeping rpoA and pheS genes of these strains with their closely-related lactococcal and streptococcal relatives were 92-94, 78 and 81 %, respectively. The results of phylogenetic analysis indicated that strains DF1T, HibF2, HibF5 and HibF3T were clustered together but were clearly separated from species of the genera Streptococcus and Lactococcus, indicating that they represent members of a novel genus of the family Streptococcaceae. Calculation of average nucleotide identity (ANI) values between the genomes of DF1T and HibF3T yielded values of 92.50-92.93 %. ANI values below the cut-off value and distinctive chemotaxonomic characteristics supported the hypothesis that these strains represented two novel species. Major cellular fatty acids in DF1T, HibF2 and HibF5 were C18 : 1ω7c and C16 : 0, while C12 : 0 and C14 : 0 were also dominant, in addition to C18 : 1ω7c and C16 : 0, in HibF3T. A novel genus is proposed with the name Floricoccus gen. nov. which consists of two species, Floricoccus tropicus sp. nov as the type species, and Floricoccus penangensis sp. nov. The respective type strains are DF1T (=LMG 29833T=JCM 31733T) and HibF3T (=LMG 29831T=DSM 31735T).

Examination of the Microbial Spectrum in the Etiology of Erythema Nodosum: A Retrospective Descriptive Study.

Even though infections are the most common cause of erythema nodosum (EN), only certain microorganisms take the great interest such as streptococci in knowledge. Our aim was to examine the frequency and type of infections in EN, to determine the characteristics of patients with an infectious etiology, and to discuss the role of these microbes in EN pathology in the context of their interactions with humans. Charts of 81 patients with EN who were seen between 2003 and 2017 were retrospectively reviewed. Identified etiological factors were classified into three groups: infectious, noninfectious, and idiopathic. While there were no significant demographic and clinical differences between the infectious and idiopathic groups, systemic symptoms (p = 0.034) and the number of EN lesions (p = 0.016) were significantly lower; the mean erythrocyte sedimentation rate was significantly higher (p = 0.049), but the mean aspartate aminotransferase value was significantly lower in the infectious group compared to the noninfectious group (p = 0.019). Besides streptococci, many other microbes, including the ones living on and inside us, were identified in the etiology of EN. There is a need for large-scale prospective studies involving control groups for a better understanding of the microbial immunopathology of EN.

Successful Continuation of Pregnancy After Treatment of Group A Streptococci Sepsis.

BACKGROUND: Invasive group A streptococci infections in pregnancy have historically led to severe maternal and neonatal morbidity and mortality. We are reporting a rare and novel case of successful treatment of third-trimester group A streptococci infection with early, aggressive intervention and maintenance of the pregnancy to term. CASE: A 35 year old woman initially presented with fever, flu-like symptoms, and preterm contractions at 34 weeks of gestation. She demonstrated signs of early stages of septic shock, ultimately attributed to group A streptococci bacteremia. Early, aggressive intervention allowed the pregnancy to continue until 38 weeks of gestation with normal maternal and neonatal outcomes. CONCLUSION: Early and aggressive treatment of invasive group A streptococci infection during pregnancy can potentially avoid severe maternal and perinatal morbidity and mortality with a successful continuation of pregnancy.

Orbital Cellulitis in a Psoriatic Patient Treated With Adalimumab.

INTRODUCTION: Psoriatic patients on tumor necrosis factor alpha inhibitors (TNFi) may be at increased risk for upper respiratory tract infections, including chronic rhino-sinusitis (CRS). Rarely, CRS can progress to orbital cellulitis (OC), an emergent ophthalmic complication wherein respiratory flora from paranasal sinus disease intrude the retro-orbital space. MATERIALS AND METHODS: Single case report. RESULTS: We report the first case, to our knowledge, of an invasive sinusitis that rapidly evolved into OC in a patient receiving adalimumab treatment for plaque psoriasis and psoriatic arthritis. After TNFi withdrawal and appropriate medical and surgical intervention, the patient fully recovered. However, on resumption of TNFi therapy, symptoms of recalcitrant CRS returned. CONCLUSION: More investigation is needed to explore how TNFi might predispose to chronic, refractory rhino-sinusitis and subsequent progression to OC. Military physicians and other medical providers should be aware of this proposed new disease entity and the potential for rapidly evolving and invasive infections in immunocompromised patients. Screening and monitoring for chronic infectious disease, such as CRS before initiating and during TNFi therapy is warranted.