Non-toxigenic Strains Research Articles

Conversion of a recA-Mediated Non-toxigenic Vibrio cholerae O1 Strain to a Toxigenic Strain Using Chitin-Induced Transformation.

Toxigenic Vibrio cholerae strains, including strains in serogroups O1 and O139 associated with the clinical disease cholera, are ubiquitous in aquatic reservoirs, including fresh, estuarine, and marine environments. Humans acquire cholera by consuming water and/or food contaminated with the microorganism. The genome of toxigenic V. cholerae harbors a cholera-toxin producing prophage (CT-prophage) encoding genes that promote expression of cholera toxin. The CT-prophage in V. cholerae is flanked by two satellite prophages, RS1 and TLC. Using cell surface appendages (TCP and/or MSHA pili), V. cholerae can sequentially acquire TLC, RS1, and CTX phages by transduction; the genome of each of these phages ultimately integrates into V. cholerae’s genome in a site-specific manner. Here, we showed that a non-toxigenic V. cholerae O1 biotype El Tor strain, lacking the entire RS1-CTX-TLC prophage complex (designated as RCT: R for RS1, C for CTX and T for TLC prophage, respectively), was able to acquire RCT from donor genomic DNA (gDNA) of a wild-type V. cholerae strain (E7946) via chitin-induced transformation. Moreover, we demonstrated that a chitin-induced transformant (designated as AAS111) harboring RCT was capable of producing cholera toxin. We also showed that recA, rather than xerC and xerD recombinases, promoted the acquisition of RCT from donor gDNA by the recipient non-toxigenic V. cholerae strain. Our data document the existence of an alternative pathway by which a non-toxigenic V. cholerae O1 strain can transform to a toxigenic strain by using chitin induction. As chitin is an abundant natural carbon source in aquatic reservoirs where V. cholerae is present, chitin-induced transformation may be an important driver in the emergence of new toxigenic V. cholerae strains.

A nationwide study of molecular epidemiology and antimicrobial susceptibility of Clostridioides difficile in South Korea

The molecular epidemiology and antimicrobial resistance of Clostridioides difficile were studied in South Korea in 2017 as part of a National Surveillance System. From February to May 2017, all non-duplicate isolates of C. difficile were recovered from patients who were suspected to have C. difficile infection and collected from 6 referral hospitals representing the 6 regions in South Korea. We performed PCRs for the toxin gene, PCR ribotyping, multilocus sequence typing (MLST), antimicrobial susceptibility testing by agar dilution according to the recommendations of the CLSI and detection of antimicrobial resistance genes such as ermB, catD, tetM, vanZ and nimR by PCR. Of 331 C. difficile isolates, 257 (77.6%) were toxigenic and the prevalence of strains producing binary toxin (CDT) was 5.1% (13/257). A total of 52 different ribotype (RT) patterns were found. RT018 was the most common (25.1% of all isolates), and RT014/020, RT002 and RT012 were also common. RT010 was most common non-toxigenic strain. MLST analysis of randomly selected 72 C. difficile isolates identified 46 sequence types (STs), of which three were new and not in the PubMLST library. There was a good correlation between MLST and RT as following: ST1 (RT027), ST8 (RT002), ST11 (RT078), ST17 (RT018), ST35 (RT046), ST37 (RT017), ST42 (RT106), ST53 (RT103), ST81 (RT369), and ST99 (RT070). All toxigenic isolates were susceptible to metronidazole and vancomycin (MIC ≤ 2 mg/L). For rifaximin, 24% of toxigenic isolates were resistant. Of randomly selected 106 toxigenic isolates, resistance rates for ampicillin, cefotetan, clindamycin, imipenem, chloramphenicol, tetracycline, and moxifloxacin were 48%, 46%, 64%, 54%, 0%, 6% and 52% respectively and frequencies of various resistance genes were 62.3% for ermB, 0.9% catD and 10.4% tetM. RTs018, 002, 017 and 369 showed high MICs to various antimicrobial agents and multi-drug resistance was common also.

177 The Presence of Clostridioides difficile Strains in the Gut by Next Generation Shotgun Sequencing: Innocent Bystander or Villain?

INTRODUCTION: Pathogenic Clostridioides difficile (C. difficile) is the most common cause of nosocomial infection in the United States. However, the prevalence of C. difficile colonization in the general population is poorly understood. We sought to determine the presence and nature of various C. difficile strains colonizing a representative sample of 17 asymptomatic adult volunteers, consisting of 10 healthy and 7 stable Crohn's patients. METHODS: Next-generation (NGS) shotgun sequencing was performed on fecal samples. Following fecal collection, DNA was extracted from stool samples, before being quantitated and normalized for downstream library fabrication utilizing shotgun methodology. Prepared and indexed libraries were subsequently pooled and sequenced on the Illumina NextSeq 550 System. Sample FASTQ files were analyzed with a computational tool (Kraken) profiling the microbial communities from metagenomic sequencing data with species level resolution. Individual microbiome profiles were then analyzed for the presence of C. difficile and specific categorical strains (Figure 1). RESULTS: All 17/17 subjects (100%) were found to possess ≥1 strain of C. difficile (Figure 1). The ATCC 43255 strain was the most common strain (94.1% 16/17), followed by QCD-63q42 (88.2% 15/17), and then M120 and QCD-37x79 strains (both identified in 10/17 [58.8%] individuals). CONCLUSION: In our representative sample of 17 adult volunteers, all (100%) were found to possess at least one C. difficile strain in their gut, with many colonized by several strains. While it is recognized that some C. difficile strains are pathogenic, our findings indicate that various non-pathogenic C. difficile species make up an important component of the commensal gut microbiome, and may perhaps play a protective role. Although symptomatic toxigenic CDI is a clear indication for therapy, C. difficile colonization with non-toxigenic strains is not believed to be a direct precursor for CDI. These findings demonstrate our need for awareness that numerous strains of C. difficile exist and the relevance of sequencing (a) prior to hospitalization or antibiotic treatment to help predict those at risk of CDI, and (b) after treatment to identify any loss of potentially protective components of our microbiome.

Inhibition of growth and ochratoxin A production in Aspergillus species by fungi isolated from coffee beans.

Ochratoxin A (OTA) is a mycotoxin found in several agricultural commodities. Produced by Aspergillus spp., it is nephrotoxic and hepatotoxic and can be carcinogenic. Preventive measures are preventing fungal growth and OTA production. In this study, fungal strains (Rhizopus oryzae, Lichtheimia ramosa, Aspergillus westerdijkiae, Aspergillus niger, Aspergillus tamarii, Aspergillus sp., and Aspergillus fumigatus) isolated from coffee beans were identified for their abilities to inhibit the growth of Aspergillus ochraceus, Aspergillus westerdijkiae, Aspergillus carbonarius, and Aspergillus niger, and OTA production. All fungi strains tested were able to inhibit growth of the four Aspergillus species and OTA production, where A. niger showed the best results in both tests. L. ramosa showed the lowest growth-reducing potential, while the other fungal strains had a growth-reducing potential higher than 70% against all Aspergillus species tested. Regarding OTA production, L. ramosa and Aspergillus sp. completely inhibited the mycotoxin production by A. ochraceus and non-toxigenic strain A. niger completely inhibited OTA production by A. niger. Our findings indicate that the strains tested can be used as an alternative means to control growth of OTA-producing fungi and production of the mycotoxin in coffee beans.

Using PCR in place of GDH as the first line assay in a two-step CDI testing algorithm – evidence of help not hindrance from samples processed at clinical microbiology laboratories in Wales

Background Guidance recommends the use of a two-stage algorithm for diagnosis of Clostridioides difficile infection (CDI) utilising either PCR or a glutamate dehydrogenase (GDH) assay as the first line screen. There is ongoing debate as to whether PCR is a hindrance due to fear of over-diagnosis. Methods Between December 2017 and March 2019 over 65,000 tests for CDI were undertaken in Wales. PCR (Serosep EntericBio C. difficile assay) was employed as the first line test for 36% of samples with the remainder tested using a GDH assay (Techlab C. diff ChekTM -60). Positive samples were tested using a toxin EIA (Techlab Tox A/B Quik Chekò). Culture (alcohol shock & CCEY; Oxoid) and capillary electrophoresis PCR ribotyping were undertaken at the UK Anaerobe Reference Unit (UKARU). Results The proportion of samples testing positive using PCR and GDH was 5.2% and 8.3% with a toxin EIA positive percentage of 30% and 25% respectively. Of these samples 73% (n=2543) of GDH and 80% (n=974) of PCR positive samples were referred for culture and ribotyping. No C. difficile was isolated from 15% of GDH and 10.5% of PCR positive specimens. Non-toxigenic strains were isolated from 6.6% of GDH and 0.2% of PCR positive samples and approximately equal proportions of both GDH and PCR positive samples were ribotyped (60% vs. 63% of 2125 samples). Conclusions There was no evidence of increased ascertainment of CDI using PCR. In fact, culture and ribotyping demonstrated an improved specificity for PCR that is helpful for accurate CDI diagnosis.

The TLCΦ satellite phage harbors a Xer recombination activation factor

The circular chromosomes of bacteria can be concatenated into dimers by homologous recombination. Dimers are solved by the addition of a cross-over at a specific chromosomal site, dif, by 2 related tyrosine recombinases, XerC and XerD. Each enzyme catalyzes the exchange of a specific pair of strands. Some plasmids exploit the Xer machinery for concatemer resolution. Other mobile elements exploit it to integrate into the genome of their host. Chromosome dimer resolution is initiated by XerD. The reaction is under the control of a cell-division protein, FtsK, which activates XerD by a direct contact. Most mobile elements exploit FtsK-independent Xer recombination reactions initiated by XerC. The only notable exception is the toxin-linked cryptic satellite phage of Vibrio cholerae, TLCΦ, which integrates into and excises from the dif site of the primary chromosome of its host by a reaction initiated by XerD. However, the reaction remains independent of FtsK. Here, we show that TLCΦ carries a Xer recombination activation factor, XafT. We demonstrate in vitro that XafT activates XerD catalysis. Correspondingly, we found that XafT specifically interacts with XerD. We further show that integrative mobile elements exploiting Xer (IMEXs) encoding a XafT-like protein are widespread in gamma- and beta-proteobacteria, including human, animal, and plant pathogens.

Ribotype variability of Clostridioides difficile strains in patients with hospital-acquired C. difficile infections, community-acquired C. difficile infections, and colonization with toxigenic and non-toxigenic strains of C. difficile

There have been few available data that presented a direct comparison between polymerase chain reaction ribotype (RT) distribution of Clostridioides difficile strains from C. difficile infection (CDI) and colonization. To understand the epidemiology of CDI in a hospital setting, we compared RTs of C. difficile strains from hospital-acquired CDI (HA-CDI) and toxigenic colonization and from community-acquired CDI (CA-CDI) and non-toxigenic colonization using the stool samples submitted for C. difficile cultures at an institution during 2009, 2012, and 2014. Overall, 721 C. difficile strains were identified from 607 patients. Among them, 450 (62.4%) were HA-CDI, 20 (2.8%) were CA-CDI, 126 (17.5%) were toxigenic colonization, and 125 (17.3%) were non-toxigenic colonization. RT018, RT017, RT002, RT015, and RT001 isolates were the most prevalent RTs in HA-CDI, and they comprised 74.9% of the total HA-CDI isolates but accounted for 60.4% of isolates from toxigenic colonization. In total, 32 strain compromising 18 RTs from HA-CDI (7.1%) were not seen among the toxigenic colonization group, and 3 RTs with 5 strains from toxigenic colonization were not seen among the HA-CDI group. The distribution of RTs was the most diverse in CA-CDI and the least diverse in HA-CDI. Although 5 RT strains, which were prevalent in HA-CDI, comprised 40% of CA-CDI, 5 isolates (25%) revealed unknown RTs, which were uncommon in HA-CDI or toxigenic colonization. In 12 patients with both episodes of CDI and toxigenic colonization, 8 had 2 isolates with different RTs and 4 had isolates with identical RTs. In conclusion, although RT017 and RT018 were the most common in HA-CDI and toxigenic colonization, C. difficile strains from toxigenic colonization were more diverse than those from HA-CDI.

Non-toxigenic strains of Vibrio cholerae biovar El Tor, isolated in the territory of Russia: molecular-genetic peculiarities and pathogenic properties

Aim. Comparative analysis of the sequenced on our own effort genomes of non-toxigenic strains with different set of pathogenicity genes and assessment of their virulence on the model of intestinal inoculation of newborn rabbits. Materials and methods. Whole-genome DNA sequencing of 26 strains was carried out using semiconductor sequencing technology. Haemolysin and hemagglutinin-protease production was evaluated applying conventional methods. Virulence of the strains for newborn rabbits was determined through intraluminal inoculation at the dose of 107 CFU/ml. Results. On the basis of whole genome analysis of ctxA-tcpA+ and ctxA-tcpA- non-toxigenic strains, differences in composition and structure stability of mobile elements associated with pathogenicity have been identified. Significant differences in nucleotide sequences of hlyA, hapA, and rtxA genes, encoding production of additional pathogenicity factors, have also been detected. The paper provides the results of assessment of their phylogenetic relations. Experiments on animal models have confirmed the inability of non-toxigenic ctxA-tcpA+ and ctxA-tcpA- strains to cause the development of typical cholera infection. Conclusion. New data on the genome structure of different non-toxigenic strains and their phylogenetic relations have been obtained. Based on the results of inoculation of animal models with cells of non-toxigenic strains with the studied genome the inference is drawn on their inability to cause the development of typical cholera infection.

Insights to the Diphtheria Toxin Encoding Prophages amongst Clinical Isolates of Corynebacterium diphtheriae from India

AbstractDiphtheria is a dreadful disease caused by Corynebacterium diphtheriae. Lysogenised bacteriophages carrying toxin gene in C. diphtheriae can make the strain toxigenic. However, such phage disseminates the toxin genes to other strains when it undergoes lytic phase. As little is known about the phage diversity in C. diphtheriae in India, the present study was undertaken to investigate the prophages integrated into the genome of 29 clinical isolates of C. diphtheriae using whole-genome shotgun sequencing. Amongst these isolates, 27 were toxigenic, while 2 were non-toxigenic strains. Of the 27 toxigenic strains, all harbored known phages carrying toxin gene and two other phages with unknown function. However, the two non-toxin strains did not harbour any of the phages in the genome. It is imperative to devise prevention strategies that hinder the dissemination of toxin by prophages, as it may increase the complications of diphtheria post-immunisation.

Vibrio cholerae filamentation promotes chitin surface attachment at the expense of competition in biofilms

Collective behavior in spatially structured groups, or biofilms, is the norm among microbes in their natural environments. Though biofilm formation has been studied for decades, tracing the mechanistic and ecological links between individual cell morphologies and the emergent features of cell groups is still in its infancy. Here we use single-cell-resolution confocal microscopy to explore biofilms of the human pathogen Vibrio cholerae in conditions mimicking its marine habitat. Prior reports have noted the occurrence of cellular filamentation in V. cholerae, with variable propensity to filament among both toxigenic and nontoxigenic strains. Using a filamenting strain of V. cholerae O139, we show that cells with this morphotype gain a profound competitive advantage in colonizing and spreading on particles of chitin, the material many marine Vibrio species depend on for growth in seawater. Furthermore, filamentous cells can produce biofilms that are independent of primary secreted components of the V. cholerae biofilm matrix; instead, filamentous biofilm architectural strength appears to derive at least in part from the entangled mesh of cells themselves. The advantage gained by filamentous cells in early chitin colonization and growth is countered in long-term competition experiments with matrix-secreting V. cholerae variants, whose densely packed biofilm structures displace competitors from surfaces. Overall, our results reveal an alternative mode of biofilm architecture that is dependent on filamentous cell morphology and advantageous in environments with rapid chitin particle turnover. This insight provides an environmentally relevant example of how cell morphology can impact bacterial fitness.

Advanced Diagnostic Strategies for Clostridium difficile Infection (CDI).

Clostridium difficile infection (CDI) is a leading cause of nosocomial intestinal diseases and produces considerable morbidity and mortality among inpatients globally. The increasing incidence of CDI, especially of its severe cases, creates a growing financial burden internationally. The pathogen responsible for CDI, C. difficile, is an anaerobic, spore-bearing Gram-positive bacillus that colonizes gastrointestinal tract in humans and animals. The bacterium can transfer from one host to another via the oral-fecal route and is consequently a public health concern in the community as well as hospitals. CDI also has implications for the personal hygiene of patients and medical workers. Classical risk factors for CDI include old age, long inpatient stay, and use of antibacterial agents. Clinical manifestations and a positive laboratory assay are both needed for an accurate diagnosis of CDI. Laboratory testing for C. difficile should distinguish between toxigenic and non-toxigenic strains, as well as asymptomatic colonization and the clinical manifestations of infection. Most stand-alone assays do not attain the desired sensitivity and specificity, and thus multistep algorithms and automated assays have emerged in recent years. Hospitals have adopted various assays, combining their conjunct advantages to enable rapid results, accurate diagnosis, and decreased expenditure. This strategy ultimately prevents misdiagnosis and over-diagnosis simultaneously. In this review, we describe common strategies for diagnosis of CDI and compare commercially available multistep and automatic assays.

Genomic analysis of a novel nontoxigenic Corynebacterium diphtheriae strain isolated from a cancer patient.

The genome of a novel nontoxigenic Corynebacterium diphtheriae, strain 5015, isolated from a patient with adenoid cystic carcinoma was sequenced and compared with 117 publically available genomes. This strain is phylogenetically distinct and lacks virulence genes encoding the toxin, BigA and Sdr-like adhesins. Strain 5015 possesses spaD-type and spaH-type pilus gene clusters with a loss of some gene functions, and 31 unique genes that need molecular characterization to understand their potential role in virulence characteristics.

Glyphosate in vitro removal and tolerance by Aspergillus oryzae in soil microcosms

Argentinian agriculture is based on a technological package that includes the application of significant amounts of pesticides. Glyphosate is the active ingredient of several glyphosate herbicides which are frequently used for weed control. Fungi can use glyphosate both as a nutrient and an energetic source. Therefore, the aim of the present study was to evaluate the in vitro glyphosate removal as the only source of phosphorus or nitrogen by two non-toxigenic Aspergillus oryzae strains. A. oryzae AM1 and AM2 were successfully able to use the herbicide both as phosphorus or nitrogen source. The degradation percentages were higher than 50% at the end of the incubation period (15 days) with glyphosate concentrations of 1 and 1.5 mM. In addition, AM1 strain was able to remove 57% of 10 mM of glyphosate at 0.99 of aW. Soil microcosm assay was performed in order to study the GP tolerance, permanence and competitiveness of A. oryzae AM1 in the presence of native mycota. A. oryzae count remained constant along the incubation period and was not significantly affected by the different glyphosate concentrations tested (10, 20 and 50 mM). This strain showed great in vitro removal ability, and it was also able to remain viable on soil microcosms contaminated with glyphosate. Therefore, these results encourage future studies in order to evaluate the ability of these strains to degrade glyphosate on soil and then promote them as potential bioremediation agents.

High prevalence and diversity of tcdA-negative and tcdB-positive, and non-toxigenic, Clostridium difficile in Thailand

Studies on the prevalence and diversity of Clostridium difficile in Thailand have been limited to those derived from a few tertiary hospitals in Central Thailand. In this study, 145 C. difficile isolates collected in 13 provinces in Thailand during 2006–2018 were characterized by ribotyping and detection of toxin genes. Minimum inhibitory concentrations of eight antimicrobial agents were determined also for all 100 C. difficile strains collected from 2006 until 2015. Of the 145 strains of C. difficile, 71 (49%) were non-toxigenic, 46 (32%) were toxin A-negative, toxin B-positive (A-B+) and 28 (19%) were A+B+. No binary toxin-positive strain was found. The most common ribotype (RT) was RT 017 (A-B+CDT-, 19%, 28/145). Besides RT 017, 20 novel non-toxigenic and A-B+ ribotyping profiles, which may be related to RT 017 by the similarity of ribotyping profile, were identified. All C. difficile strains remained susceptible to metronidazole and vancomycin, however, a slight increase in MIC for metronidazole was seen in both toxigenic and non-toxigenic strains (overall MIC50/90 0.25/0.25 mg/L during 2006–2010 compared to overall MIC50/90 1.0/2.0 mg/L during 2011–2015). There was a high rate of fluoroquinolone resistance among RT 017 strains (77%), but there was little resistance among non-toxigenic strains. These results suggest that RT 017 is endemic in Thailand, and that the misuse of fluoroquinolones may lead to outbreaks of RT 017 infection in this country. Further studies on non-toxigenic C. difficile are needed to understand whether they have a role in the pathogenesis of C. difficile infection in Asia.

Detecting Clostridioides (Clostridium) difficile using canine teams: What does the nose know?

SummaryBackgroundTrained canines are capable of detecting Clostridioides (Clostridium) difficile (CD) in the environment; however, the primary odour of interest on which the dogs alert is unclear.AimTo evaluate the inter-rater reliability of two canine detection teams for their ability to discriminate between scent pads containing CD-toxin-positive and -negative odours and their ability to discriminate between clostridial strains.MethodsDuring a six-month period, two canine teams were tested weekly for their ability to detect CD-toxin-positive odours and discriminate between these and -negative odours. To further determine the canines' discrimination capability, scent pads impregnated with odours from reference isolates representing common CD toxin types (including toxin-negative CD isolates) or from clinical isolates representing other clostridial species were used.ResultsA total of 264 samples were tested with an overall sensitivity of 94.7% (Team A) and 86.8% (Team B) and specificities of 96.9% and 98.7%, respectively. Inter-rater reliability was very good (Cohen's kappa 0.87). When challenged with toxin- and non-toxin-producing strains, the teams alerted on 96.3% of all CD isolate odours (including nontoxigenic strains) and 46.7% of closely related species.ConclusionsThe canine teams exhibited strong inter-rater reliability on both clinical faecal specimens and reference CD isolates (both toxin and non-toxin producing) but were challenged to discriminate between CD and closely related clostridial species. These findings strongly support the development of scent detection programmes provided dogs and their handlers are properly trained and used in the right context.

Non-toxigenic strain of Clostridioides difficile Z31 reduces the occurrence of C. difficile infection (CDI) in one-day-old piglets on a commercial pig farm

Neonatal porcine diarrhea (NPD) is a current problem on pig farms and is caused by several enteropathogens. Among them, Clostridioides difficile stands out due to its importance in piglets and zoonotic potential. A non-toxigenic strain of C. difficile (NTCD), named Z31, was previously tested in hamster and piglet experimental models as a strategy to prevent C. difficile infection (CDI). To evaluate the capacity of the strain Z31 to prevent CDI and NPD in one-day-old piglets on a commercial farm, 90 piglets from 16 litters received 1 × 106 spores of Z31 while 84 animals from the same litters served as controls. Animals were clinically evaluated, and fecal samples were collected 24 h after administration and submitted to A/B toxin detection and isolation of C. difficile. Stool samples were also submitted to rotavirus, Escherichia coli, and Clostridium perfringens detection. Administration of Z31 reduced the incidence of CDI in treated animals (7.8%) when compared to the control group (25.0%; P = 0.003). In animals that developed CDI, the intensity of diarrhea was lower in those that received Z31 than in the control group. Neonatal porcine diarrhea was reduced in treated animals when compared to untreated animals (P < 0.001). The present study suggests that Z31 can potentially be used to prevent CDI in piglets on commercial farms.

Evidence of transmission of Clostridium difficile in asymptomatic patients following admission screening in a tertiary care hospital.

BackgroundClostridium difficile (CD) is the leading cause of infectious health-care associated diarrhea. However, little is known regarding CD carriage and transmission amongst asymptomatic colonizers. We evaluated carriage, characterized strains and examined epidemiologic linkages in asymptomatic colonized CD patients.MethodsRectal swabs from asymptomatic patients admitted to the general medicine ward from April 1-June 30 2012 were collected. PCR-confirmed CD colonies were ribotyped and characterized by Modified-Multi Locus Variable Number Tandem Repeat Analysis (MMLVA).Results1549-swabs were collected from 474-patients. Overall, 50/474(10.6%) were CD PCR-positive, 24/50 were colonized at admission, while 26/50 were first identified > = 72 hours after admission. Amongst the 50 CD PCR-positive patients, 90% were asymptomatically colonized and 80% of individuals carried toxigenic CD-strains, including ribotype-027 (5/45:11%). MMLVA revealed five-clusters involving 15-patients harboring toxigenic (4/5) and non-toxigenic CD strains (1/5). In two clusters, patients were CD positive on admission while in the other three clusters involving 10 patients, we observed CD transmission from asymptomatically colonized patients to 8 previously CD-negative patients.ConclusionsWe identified increasing rates of colonization during admission to medical wards. MMLVA typing effectively discriminated between strains and suggests that 20% of patients with CD colonization acquired their strain(s) from asymptomatically colonized individuals in hospital.

Fungi as industrial producers of food ingredients

Diverse ingredients (such as acidulants, enzymes, flavors, vitamins, colorants, and polyunsaturated fatty acids) used during food processing are obtained from fungi through industrial fermentation. Advances on laboratorial techniques for selection of non-toxigenic fungal strains, as well as the increased knowledge about biosynthesis regulation and the possibilities of genetic engineering allowed the achievement of higher yields of safe food grade products. These bioprocessing are encouraged by consumer awareness onto clean label ingredients, as well as interests toward sustainability. This review intends to show some food ingredients obtained from species of yeasts and molds as well as their use in food industries.

A study on clinical application of a novel reagent detecting toxigenic strains of Clostridium difficile

Objective: To evaluate the performances of a "ready-to-use" liquid chromogenic medium for detecting toxigenic strains of C. difficile from stool specimens, by comparing with the reference method of combined GeneXpert detection with MALDI-TOF MS identification. Methods: This study was a clinical application research. A total of 322 stool specimens from patients with diarrhea in Henan Provincial People's Hospital were detected by using this liquid chromogenic mediumor by combination of GeneXpert and MALDI-TOF MS identification after CCFA culture from March to November 2017. The reference method was combined GeneXpert detection with MALDI-TOF MS identification. Data were analyzed with kappa analysis by use of SPSS 13.0. In this study, the limit of detection and anti-interference capability of chromogenic medium were determined with standard strains. Results: This assay had a sensitivity of 92.86% (52/56), a specificity of 96.99% (258/266), a positive predictive value of 86.67% (52/60), and a negative predictive value of 98.47% (258/262). Furthermore, this assay was well consistent with the reference method (kappa=0.874, P<0.01). The limit of detection of this assay was 10(3) CFU/ml for toxigenic strain of ATCC9689, and there was no interference by non-toxigenic strain of ATCC43593 or Clostridium perfringens (ATCC13124) even.When the concentration of ATCC9689 was as low as 0.5 McFarland. Conclusions: It is worth expanding clinical research for this "ready-to-use" chromogenic medium and preparing for its popularization in primary medical institutions, due to its similar performance to the reference method for detecting toxigenic strains of C. difficile, simple operation and no need of anaerobic condition.

Evaluation of growth and sporulation of a non-toxigenic strain of Clostridioides difficile (Z31) and its shelf viability

The oral administration of non-toxigenic strains of Clostridioides difficile (NTCD) is currently showing promising results for the prevention of Clostridioides difficile infection (CDI) in humans and animals, and is being considered as a possible commercial product to be used in the near future. The aim of this work was to evaluate five culture media for the growth and sporulation of one NTCD (Z31) and evaluate the viability of a lyophilized spore solution of NTCD Z31 stored at 4°C or at 25°C for 2years. Reinforced clostridial medium (RCM) and brain heart infusion broth (BHI) provided the highest production of NTCD Z31 spores. In the first 6months of the storage of the lyophilized solution, a reduction in spore count of approximately 0.3Log10CFU/mL was observed; however, no further significant reduction in spore count was observed up to 24months. No difference in spore concentration was found between the two storage temperatures from 6 to 24months of storage. The present work showed BHI and RCM to be the best choices for the growth and sporulation of NTCD Z31 and suggested that the spores of NTCD Z31 are stable for up to 2years under both temperature conditions.