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The objective of this trial was to evaluate how broilers responded to Aspergillus flavus strains that are toxigenic and atoxigenic. The study included four treatments in a 2 × 2 factorial design, with six replicates of 10 birds each. As a result of this study measuring feed intake (FI), weight gain (WG), feed conversion ratio (FCR), crude protein, ether extract, and crude fibre, the interaction was insignificant between the toxigenic and atoxigenic diets (P > 0.05). Consumption of toxigenic aflatoxin B1-500 ppb diet decreased FI and WG but increased FCR, and cost to produce live broiler weight (P < 0.05) compared to the control diets. The addition of atoxigenic strains to contaminated diets significantly offset (P < 0.05) the effects. Diets with or without 500 ppb toxigenic and atoxigenic A. flavus did not affect the relative weight g/100gBW of pancreas, gizzard and bursa of Fabricius. Dietary inclusion of 500 ppb toxigenic Aspergillus spp. increased the relative weight (P < 0.05) of the kidney, liver, spleen and thymus while atoxigenic dietary addition reduced the relative weight of the same organs (P < 0.05). Dietary inclusion of toxigenic and atoxigenic Aspergillus spp. did not significantly affect the haematological parameters measured (P < 0.05). Dietary inclusion of 500 ppb toxigenic Aspergillus elevated the urea, creatine, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) in the serum of the broilers (P < 0.05). A decrease was observed when atox igenic A. flavus was used in the intervention for urea, creatinine and AST (P < 0.05), whereas an insignificant reduction was observed for ALT and ALP (P ≤ 0.05). This study concluded that dietary atoxigenic strain improved broiler performance, digestibility, and blood parameters. [ABSTRACT FROM AUTHOR]

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Objectives Most human infections caused by Vibrio spp. do not warrant antimicrobial treatment but in severe cases, targeted antimicrobial treatment can be lifesaving. For Vibrio spp. standardized antimicrobial susceptibility testing (AST) guidelines with EUCAST methodology are lacking. In this study, we aimed to produce data suitable for EUCAST to establish clinical MIC breakpoints and zone diameter correlates for Vibrio spp. Methods An intercontinental collection (N = 524) comprising five important Vibrio spp. (V. alginolyticus , V. cholerae , V. fluvialis , V. parahaemolyticus and V. vulnificus) was organized. All isolates were subjected to broth microdilution (BMD) against 11 antimicrobial agents according to ISO 20776-1 using unsupplemented Mueller–Hinton broth on freeze-dried Sensititre panels (Thermo Scientific, UK), and most isolates (n = 371) were also tested with disc diffusion according to EUCAST methodology for non-fastidious organisms. Results Aggregated results were used to generate MIC and zone diameter distributions and to prepare graphs of MIC-zone diameter correlation. Based on these results, the EUCAST Steering Committee determined clinical susceptible (S) and resistant (R) MIC (mg/L) breakpoints (S≤/R>) for the five Vibrio spp. for piperacillin/tazobactam (1/1), cefotaxime (0.25/0.25), ceftazidime (1/1), meropenem (0.5/0.5), ciprofloxacin (0.25/0.25), levofloxacin (0.25/0.25), azithromycin (4/4), doxycycline (0.5/0.5) and trimethoprim/sulfamethoxazole (0.25/0.25). The corresponding zone diameter breakpoints were identified. Conclusions We demonstrated the validity of using standard BMD and EUCAST disc diffusion methodology for AST of five Vibrio spp. and generated suitable data to allow EUCAST to determine clinical MIC and zone diameter breakpoints for five pathogenic Vibrio spp. including both non-toxigenic and toxigenic V. cholerae. [ABSTRACT FROM AUTHOR]

Chickpea, Cicer arietinum L., is a nutrient rich crop that is widely cultivated and consumed in Pakistan. However, chickpea is highly prone to fungal growth leading to contamination with aflatoxins, the most potent carcinogen found in nature. In this study, fifty chickpea seed samples were collected from the local markets of the Punjab, Pakistan, to evaluate their nutritional quality, fungal and AFB1 contamination. Proximate analysis suggested that chickpea seeds contained 5.5–6.93% moisture, 62.24–63.24% carbohydrates, 22.75–23.44% protein, 4.99–5.4% fat, 5.62–5.84% fiber and 2.92–3.16% ash. Morphological identification techniques revealed fourteen fungal species belonging to six fungal genera from which Aspergillus flavus was the leading contaminant. AFB1 analysis revealed that sixty-two percent samples were contaminated with AFB1. All the AFB1 positive samples contained AFB1 level more than 2 ppb and 12.9% samples contain AFB1 level more than 20 ppb, exceeded the maximum limit (ML) assigned by EU and USA (FDA & FAO) respectively. The results of the present studies reported that chickpea is a highly contaminated commodity in terms of fungi and AFB1 that's why further investigations and monitoring are required to reduce the fungal and AFB1 contamination. These baseline data are an initial step in the effort to deal with this significant food safety issue. [ABSTRACT FROM AUTHOR]

Toxigenic Aspergillus flavus is a primary producer of aflatoxin in Indonesia, and its presence can lead to the contamination of agricultural commodities. This contamination poses a risk to export-targeted commodities, potentially resulting in their rejection. Therefore, this study aims to characterize the molecular profile of native A. flavus isolated from several Indonesian agricultural products, with a major focus on its toxigenicity and toxin production. A total of 18 A. flavus collections were isolated from nutmeg, ground peanut, cacao, coffee bean, corn, white pepper, and soil peanut plantation. Species identification was carried out using molecular and morphological approaches. The toxigenicity of isolates was characterized based on the amplification of aflatoxin gene clusters, while toxin production was assessed through growth simulation on a 10% coconut broth media followed by HPLC quantification. The result showed that all isolates were confirmed as A. flavus based on the morphological and sequence analysis of the ITS region. A total of 11 isolates (61%) were confirmed as toxigenic and produced 1-2 types of aflatoxin, in varying concentrations of high, moderate, or low levels of AFB1. High levels of AFB1 produced by seven isolates namely BIO3313, BIO33212, BIO3361, BIO33404, BIO3338, BIO3352, and BIO3344, had concentration levels ranging from 76.78 to 2241.06 µg/kg, while three isolates (BIO3314, BIO3312, and BIO3381) produced AFB1 below 1 µg/kg. Twenty-nine pairs of aflatoxin gene-specific sequences were successfully amplified as a single band, while some produced non-specific patterns in several low toxigenic and non-toxigenic isolates. Based on the results, it was concluded that completed gene clusters and variations of gene deletion were observed in both toxigenic and non-toxigenic isolates. However, no specific target gene could effectively distinguish the two groups. Two non-toxigenic isolates namely BIO3393 and BIO33403 exhibited a large deletion and could be potential candidates for biocontrol agents.

Aflatoxin contamination of the staples maize and groundnut is a concern for health and economic impacts across sub-Saharan Africa. The current study (i) determined aflatoxin levels in maize and groundnut collected at harvest in Burundi, (ii) characterized populations of Aspergillus section Flavi associated with the two crops, and (iii) assessed aflatoxin-producing potentials among the recovered fungi. A total of 120 groundnut and 380 maize samples were collected at harvest from eight and 16 provinces, respectively. Most of the groundnut (93%) and maize (87%) contained aflatoxin below the European Union threshold, 4 µg/kg. Morphological characterization of the recovered Aspergillus section Flavi fungi revealed that the L-morphotype of A. flavus was the predominant species. Aflatoxin production potentials of the L-morphotype isolates were evaluated in maize fermentations. Some isolates produced over 137,000 µg/kg aflatoxin B1. Thus, despite the relatively low aflatoxin levels at harvest, the association of both crops with highly toxigenic fungi poses significant risk of post-harvest aflatoxin contamination and suggests measures to mitigate aflatoxin contamination in Burundi should be developed. Over 55% of the L-morphotype A. flavus did not produce aflatoxins. These atoxigenic L-morphotype fungi were characterized using molecular markers. Several atoxigenic genotypes were detected across the country and could be used as biocontrol agents. The results from the current study hold promise for developing aflatoxin management strategies centered on biocontrol for use in Burundi to reduce aflatoxin contamination throughout the value chain. [ABSTRACT FROM AUTHOR]

Toxigenic members of Aspergillus flavus contaminate cereal grains, resulting in contamination by aflatoxin, a food safety hazard that causes hepatocellular carcinoma. This study identified probiotic strains as aflatoxin detoxifiers and investigated the changes to the grain amino acid concentrations during fermentation with probiotics in the presence of either A. flavus La 3228 (an aflatoxigenic strain) or A. flavus La 3279 (an atoxigenic strain). Generally, higher concentrations (p < 0.05) of amino acids were detected in the presence of toxigenic A. flavus La 3228 compared to the atoxigenic A. flavus La 3279. Compared to the control, 13/17 amino acids had elevated (p < 0.05) concentrations in the presence of the toxigenic A. flavus compared to the control, whereas in systems with the atoxigenic A. flavus 13/17 amino acids had similar (p > 0.05) concentrations to the control. There were interspecies and intraspecies differences in specific amino acid elevations or reductions among selected LAB and yeasts, respectively. Aflatoxins B1 and B2 were detoxified by Limosilactobacillus fermentum W310 (86% and 75%, respectively), Lactiplantibacillus plantarum M26 (62% and 63%, respectively), Candida tropicalis MY115 (60% and 77%, respectively), and Candida tropicalis YY25, (60% and 31%, respectively). Probiotics were useful detoxifiers; however, the extent of decontamination was species- and strain-dependent. Higher deviations in amino acid concentrations in the presence of toxigenic La 3228 compared to atoxigenic La 3279 suggests that the detoxifiers did not act by decreasing the metabolic activity of the toxigenic strain. [ABSTRACT FROM AUTHOR]

Vibrio cholerae remains a major public health threat worldwide, causing millions of cholera cases each year. Although much is known about the evolution and pathogenicity of the O1/O139 serogroups of V. cholerae, information is lacking on the molecular epidemiology of non‒O1/O139 strains isolated from patients who have diarrheal illnesses. We performed whole-genome sequence analysis and in vivo infections to investigate characteristics of V. cholerae O141 isolated from sporadic diarrheal cases in 4 countries. The strains formed a distinct phylogenetic clade distinguishable from other serogroups and a unique multilocus sequence type 42, but interstrain variation suggests that O141 isolates are not clonal. These isolates encode virulence factors including cholera toxin and the toxin-coregulated pilus, as well as a type 3 secretion system. They had widely variable capacities for intestinal colonization in the infant mouse model. We propose that O141 isolates comprise a distinct clade of V. cholerae non‒O1/O139, and their continued surveillance is warranted.

Aflatoxin contamination of the staples maize and groundnut is a concern for health and economic impacts across sub-Saharan Africa. The current study (i) determined aflatoxin levels in maize and groundnut collected at harvest in Burundi, (ii) characterized populations of Aspergillus section Flavi associated with the two crops, and (iii) assessed aflatoxin-producing potentials among the recovered fungi. A total of 120 groundnut and 380 maize samples were collected at harvest from eight and 16 provinces, respectively. Most of the groundnut (93%) and maize (87%) contained aflatoxin below the European Union threshold, 4 μg/kg. Morphological characterization of the recovered Aspergillus section Flavi fungi revealed that the L-morphotype of A. flavus was the predominant species. Aflatoxin production potentials of the L-morphotype isolates were evaluated in maize fermentations. Some isolates produced over 137,000 μg/kg aflatoxin B1. Thus, despite the relatively low aflatoxin levels at harvest, the association of both crops with highly toxigenic fungi poses significant risk of post-harvest aflatoxin contamination and suggests measures to mitigate aflatoxin contamination in Burundi should be developed. Over 55% of the L-morphotype A. flavus did not produce aflatoxins. These atoxigenic L-morphotype fungi were characterized using molecular markers. Several atoxigenic genotypes were detected across the country and could be used as biocontrol agents. The results from the current study hold promise for developing aflatoxin management strategies centered on biocontrol for use in Burundi to reduce aflatoxin contamination throughout the value chain.

ABSTRACT Vibrio cholerae O1 is the causative agent of cholera, a severe diarrheal disease which can cause death if left untreated. In this study, a collection of clinical and environmental V. cholerae serogroup O1 isolates from Australia (1977 to 1987) (from local cases and cases acquired through international travel) and publicly available international isolates were characterized for genotypic features (virulence genes, mobile genetic elements [MGEs], and antimicrobial resistance gene profiles). Whole-genome sequencing (WGS) was used to investigate and compare the genetic relatedness between the 44 Australian and nine travel-associated isolates and the 60 publicly available international V. cholerae sequences representing pre-seventh-pandemic (pre-7PET) isolates and different waves of 7PET isolates. In this study, 36 (81%) Australian clinical and aquatic isolates harbored the cholera toxin-producing genes located in the CTX bacteriophage region. All the Australian environmental and clinical isolates lacked the seventh-pandemic virulence-associated genomic islands (VSP-I and -II). In silico multilocus sequence typing (MLST) classified all nine internationally acquired isolates as sequence type 69 (ST69), 36 clinical and aquatic isolates as ST70, and eight isolates from Australia as ST71. Most of the nontoxigenic clinical and aquatic isolates of ST71 had diverse genetic variations compared to ST70 Australian strains. The antimicrobial resistance-associated genes gyrA, parC, and parE had no mutations in all the environmental and clinical isolates from Australia. The SXT genetic element and class 1 integron gene sequences were not detected in Australian strains. Moreover, in this study, a Bayesian evolutionary study suggests that two distinct lineages of ST71 (new set of strains) and ST70 strains were prevalent around similar times in Australia, in ~1973 and 1969. IMPORTANCE Australia has its own indigenous V. cholerae strains, both toxigenic and nontoxigenic, that are associated with disease. Exotic strains are also detected in Australian patients returning from overseas travel. The clinical and aquatic V. cholerae O1 toxin gene-positive isolates from Australia responsible for cases in 1977 to 1987 were linked to acquisition from Queensland waterways but until now had not been characterized genetically. It is important to determine the genetic relatedness of Australian strains to international strains to assist in understanding their origin. This is the first extensive study to provide sequences and genomic analysis focused on toxigenic O1 V. cholerae clinical and environmental strains from Australia and its possible evolutionary relationship with other publicly available pre-7PET and 7PET V. cholerae strains. It is important to understand the population genetics of Australian V. cholerae from a public health perspective to assist in devising control measures and management plans for reducing V. cholerae exposure in Australia, given previous Australian disease clusters.

The fungus Aspergillus flavus infects corn, peanut, and cottonseed, and contaminates seeds with acutely poisonous and carcinogenic aflatoxin. Aflatoxin contamination is a perennial threat in tropical and subtropical climates. Nonaflatoxin-producing isolates (atoxigenic) are deployed in fields to mitigate aflatoxin contamination. The biocontrol competitively excludes toxigenic A. flavus via direct replacement and thigmoregulated (touch) toxin inhibition mechanisms. To understand the broad-spectrum toxin inhibition, toxigenic isolates representing different mating types and sclerotia sizes were individually cocultured with different atoxigenic biocontrol isolates. To determine whether more inhibitory isolates had a competitive advantage to displace or touch inhibit toxigenic isolates, biomass accumulation rates were determined for each isolate. Finally, to determine whether atoxigenic isolates could inhibit aflatoxin production without touch, atoxigenic isolates were grown separated from a single toxigenic isolate by a membrane. Atoxigenic isolates 17, Af36, and K49 had superior abilities to inhibit toxin production. Small (<400 µm) sclerotial, Mat1-1 isolates were not as completely inhibited as others by most atoxigenic isolates. As expected for both direct replacement and touch inhibition, the fastest-growing atoxigenic isolates inhibited aflatoxin production the most, except for atoxigenic Af36 and K49. Aflatoxin production was inhibited when toxigenic and atoxigenic isolates were grown separately, especially by slow-growing atoxigenic Af36 and K49. Additionally, fungus-free filtrates from atoxigenic cultures inhibited aflatoxin production. Toxin production inhibition without direct contact revealed secretion of diffusible chemicals as an additional biocontrol mechanism. Biocontrol formulations should be improved by identifying isolates with broad-spectrum, high-inhibition capabilities and production of secreted inhibitory chemicals. [ABSTRACT FROM AUTHOR]

Corynebacterium diphteriae is divided based on its ability to produce toxin. Toxigenic C. diphteriae is the type that has capacity to produce toxin and life threatening. This study was descriptive study in Corynebacterium diphteriae found in children less than 18 years old who were administered to National Infection Center Prof. Dr. Sulianti Saroso. We found 36 viable toxigenic C. diphteriae isolates, which 52.8% intermedius strain, 33.3% gravis strain, 8.3% mitis, and 5.6% non-strain. Most of the host (52,8%) were 60-144 months years old. Majority of the host had completed basic vaccination recommended by Indonesian Paediatrician Association while 75% of them hadn’t gotten the supplementary doses. Fever (48.5%) and odynophagia (54.5%) were experienced by toxigenic intermedius strain infection. Snoring (50%) and thick pseudomembran mostly found in gravis strain host while Bullneck sign found in intermedius strain infection. Complications like airway obstruction, chronic kidney disease, and myocarditis were found in host with toxigenic intermedius strain by 66.7%, 100%, and 75%. Death case were also mostly with this strain. Intermedius strain were mostly found in this study. Complications and mortality were also connected to intermedius strain infection.

Abstract Background The biological control mechanism of Aspergillus flavus (aflatoxigenic) strains in groundnuts with atoxigenic strains from the same species through competitive exclusion employed the use of endemic and well-adapted strains within the agro-ecological zones of Zimbabwe. The selected elite non-aflatoxigenic isolates of A. flavus native to Zimbabwe were evaluated for their capability to reduce aflatoxin contamination in groundnuts under laboratory conditions. Results Average reduction percentages in aflatoxin B concentration for the 2019 and 2020 set of experiments ranged from 91.6 ± 3.4 to 95.8 ± 3.1% and 90.29 ± 3.6% to 95.29 ± 4.1%, respectively. Levels of aflatoxin in the co-inoculation research experiments administered were significantly reduced in all the experimental units carried out. Treatment efficiencies of the tested isolates in this study at 4:1 and 2:1 ranged from 1.20 to 2.52 and from 1.02 to 1.21, respectively. The efficacy of the tested non-aflatoxigenic strains against the aflatoxigenic strain native to Zimbabwe (ZMW 0127) indicates that the non-aflatoxigenic isolates of A. flavus. have sound practical applications against vast communities of aflatoxin-producing fungi across all the agro-ecological zones in Zimbabwe. Conclusion The recognized non-aflatoxigenic isolates will be of an incentive as dynamic active ingredients in biocontrol formulations for the decrease in aflatoxins in groundnuts grown in Zimbabwe.

The study was carried out to enumerate fungal contamination, and toxigenicity of Aspergillus flavus strains on cacao beans during drying and storage. As many as 3500 g cacao beans during drying and storage were purchased from smallholder farmers on the plantation areas at Karo Regency, Northern Sumatera. The percentage of the beans contaminated by fungi was conducted using direct plating. Fungal populations on soil and beans were determined using dilution followed by pour plated in dichloran 18% glycerol agar (DG18) and Aspergillus flavus and parasiticus agar (AFPA). The mycological evaluation was carried out based on morphological characteristics. Results showed eighteen genera of soil fungi were isolated at the cacao plantation; genera of Aspergillus sp., A. niger, A. flavus, and Penicillium citrinum were the most important contaminants. Six species of the fungi were associated with contamination on cocoa beans during drying i.e. Aspergillus sp., Candida tropicalis, Saccharomyces sp., A. niger, Penicillium spp., and Fusarium spp. Whereas three fungal species were associated during storage i.e. A. niger, A. flavus, and P. citrinum. The percentage of cacao contaminated during drying and storage was dominated (>40%) by Aspergillus sp. Fusarium sp. A. niger, A. flavus, and P. citrinum, respectively. Among 21 strains of A. flavus, 3 strains (15%) were isolated from soil, and 18 strains (85%) were isolated from beans during storage. Among toxigenic A. flavus, both strain scaf6 isolated from soil and strain cbaf5 isolated from beans during storage were the highest aflatoxin producers (30.0 ppb). Preventing soil contamination during harvesting, drying, and storage of cacao beans was a prerequisite to minimise fungal contamination. [ABSTRACT FROM AUTHOR]

The goal is to study the features of the biological properties of Vibrio cholerae strains found in environmental objects (EO) on the territory of Rostov-on-Don from 1989 to 2018.Materials and methods. Bacteriological, molecular biological and statistical methods were used to study 73 strains of cholera vibrios O1, O139 and 1702 strains of nonO1/nonO139 serogroups isolated over a 30-year period.Results and discussion. It was found that in Rostovon-Don, 78.0 % of toxigenic strains were isolated from all isolated from environmental protection in Russia, among which 2 were genetically unchanged. The V. cholerae O1 classical strain and 72 V. cholerae El Tor strains were found, of which 70 % belonged to the Ogawa serovar, and 3 % of the nontoxigenic strains were susceptible to the classical phage. A “very weak direct relationship” was statistically revealed between the isolation of V. cholerae O1, O139 and V. cholerae nonO1/nonO139 strains, in which the dominant serogroups (O67, O76, O75, O53, O16) were identified, and 18 % of the strains showed atypical agglutinability. Revealing the biological characteristics of V. cholerae strains contributes to the optimization of monitoring studies.

Hand-dug wells are natural, inexpensive solution to widespread water scarcity; however, they could be contaminated with toxic algae. This study aimed to assess the physicochemical parameters and survey algal species in water from three hand-dug wells in Nassarawa Local Government area of Kano State (11°57'53"N; 8°30'50"E). Water samples were collected during May and June 2021. Temperature, pH, Total Dissolved Solids (TDS), Electrical Conductivity (EC), Biological Oxygen Demand (BOD) Dissolved oxygen (DO), Turbidity (TBDT), Nitrate and Phosphate were determined according to (APHA, 2005). Algal composition (org/L) and identification were by using the guide of Palmer (1980). Analyses of variance was used to determine significant differences between means at (p<0.05). All parameters were within permissible limits of WHO. Site B (lined with rocks) had the highest mean values of TDS (513.67mg/L), EC (596.70mg/L) and TBDT (4.60mg/L). Mean values for DO and BOD showed significant differences between sites. A total of 430 org/L algae were identified belonging to 13 genera consisting of Cyanophyta (51.1%), Chlorophyta (16.2%) and Bacillariophyta (32.7%). Site A (the oldest and unlined) was the most diverse. Site C (lined with concrete) had the highest total number of organisms (42.3%). Oscillatoria sp., Navicula sp., Phormidium sp and Dictyosphaerium sp. occurred in all sites. Nitszchia, Gomphonema and Fragillaria were the least abundant. Hand-dug wells should be protected and conserved to ensure a sustainable supply of water for various purposes. [ABSTRACT FROM AUTHOR]

Vibrio cholerae remains a major public health threat worldwide, causing millions of cholera cases each year. Although much is known about the evolution and pathogenicity of the O1/O139 serogroups of V. cholerae, information is lacking on the molecular epidemiology of non‒O1/O139 strains isolated from patients who have diarrheal illnesses. We performed whole-genome sequence analysis and in vivo infections to investigate characteristics of V. cholerae O141 isolated from sporadic diarrheal cases in 4 countries. The strains formed a distinct phylogenetic clade distinguishable from other serogroups and a unique multilocus sequence type 42, but interstrain variation suggests that O141 isolates are not clonal. These isolates encode virulence factors including cholera toxin and the toxin-coregulated pilus, as well as a type 3 secretion system. They had widely variable capacities for intestinal colonization in the infant mouse model. We propose that O141 isolates comprise a distinct clade of V. cholerae non‒O1/O139, and their continued surveillance is warranted. [ABSTRACT FROM AUTHOR]

Cholera is a severe diarrheal disease caused by Vibrio cholerae, a natural inhabitant of brackish water. Effective control of cholera outbreaks depends on prompt detection of the pathogen from clinical specimens and tracking its source in the environment. Although the epidemiology of cholera is well studied, rapid detection of V. cholerae remains a challenge, and data on its abundance in environmental sources are limited. Here, we describe a sensitive molecular quantification assay by qPCR, which can be used on-site in low-resource settings on water without the need for DNA extraction. This newly optimized method exhibited 100% specificity for total V. cholerae as well as V. cholerae O1 and allowed detection of as few as three target CFU per reaction. The limit of detection is as low as 5 × 103 CFU/L of water after concentrating biomass from the sample. The ability to perform qPCR on water samples without DNA extraction, portable features of the equipment, stability of the reagents at 4 °C and user-friendly online software facilitate fast quantitative analysis of V. cholerae. These characteristics make this assay extremely useful for field research in resource-poor settings and could support continuous monitoring in cholera-endemic areas. [ABSTRACT FROM AUTHOR]

Background Diphtheria is a potentially fatal respiratory disease caused by toxigenic Corynebacterium diphtheriae. Although resistance to erythromycin has been recognized, β-lactam resistance in toxigenic diphtheria has not been described. Here, we report a case of fatal respiratory diphtheria caused by toxigenic C. diphtheriae resistant to penicillin and all other β-lactam antibiotics, and describe a novel mechanism of inducible carbapenem resistance associated with the acquisition of a mobile resistance element. Methods Long-read whole-genome sequencing was performed using Pacific Biosciences Single Molecule Real-Time sequencing to determine the genome sequence of C. diphtheriae BQ11 and the mechanism of β-lactam resistance. To investigate the phenotypic inducibility of meropenem resistance, short-read sequencing was performed using an Illumina NextSeq500 sequencer on the strain both with and without exposure to meropenem. Results BQ11 demonstrated high-level resistance to penicillin (benzylpenicillin minimum inhibitory concentration [MIC] ≥ 256 μg/ml), β-lactam/β-lactamase inhibitors and cephalosporins (amoxicillin/clavulanic acid MIC ≥ 256 μg/mL; ceftriaxone MIC ≥ 8 μg/L). Genomic analysis of BQ11 identified acquisition of a novel transposon carrying the penicillin-binding protein (PBP) Pbp2c, responsible for resistance to penicillin and cephalosporins. When strain BQ11 was exposed to meropenem, selective pressure drove amplification of the transposon in a tandem array and led to a corresponding change from a low-level to a high-level meropenem-resistant phenotype. Conclusions We have identified a novel mechanism of inducible antibiotic resistance whereby isolates that appear to be carbapenem susceptible on initial testing can develop in vivo resistance to carbapenems with repeated exposure. This phenomenon could have significant implications for the treatment of C. diphtheriae infection, and may lead to clinical failure. [ABSTRACT FROM AUTHOR]

Clostridioides difficile (C. difficile) infection (CDI) is the most common cause of antibiotic-associated diarrhea and one of the common infections in healthcare facilities. In recent decades, there has been an emerging threat of community-acquired CDI (CA-CDI). Environmental transmission of C. difficile in the community setting has become a major concern, and animals are an important reservoir for C. difficile causing human diseases. In this article, the molecular epidemiology of C. difficile in animals and recent evidences of zoonotic transfer to humans are reviewed based on an electronic search in the databases of PubMed and Google Scholar. C. difficile can be found in stool from diarrheal dogs and cats; therefore, household pets could be a potential source. C. difficile will threaten human health because hypervirulent C. difficile ribotype 078 strains have been found in retail chickens, pig farms, and slaughterhouses. Risk factors for fecal C. difficile carriage in animals include young age, dietary changes, and antibiotic abuse in domestic animals. With the advent of whole genome sequencing techniques, there will be more solid evidence indicating zoonotic transfer of C. difficile from animals to humans. [ABSTRACT FROM AUTHOR]

Background Asymptomatic C. difficile colonization is believed to predispose to subsequent C. difficile infection (CDI). While emerging insights into the role of the commensal microbiota in mediating colonization resistance against C. difficile have associated CDI with specific microbial components, corresponding prospectively collected data on colonization with C. difficile are largely unavailable. Methods C. difficile status was assessed by GDH EIA and real-time PCR targeting the toxin A (tcdA) and B (tcdB) genes. 16S V3 and V4 gene sequencing results from fecal samples of patients tested positive for C. difficile were analyzed by assessing alpha and beta diversity, LefSe, and the Piphillin functional inference approach to estimate functional capacity. Results 1506 patients were recruited into a prospective observational study (DRKS00005335) upon admission into one of five academic hospitals. 936 of them provided fecal samples on admission and at discharge and were thus available for longitudinal analysis. Upon hospital admission, 5.5% (83/1506) and 3.7% (56/1506) of patients were colonized with toxigenic (TCD) and non-toxigenic C. difficile (NTCD), respectively. During hospitalization, 1.7% (16/936) acquired TCD. Risk factors for acquisition of TCD included pre-existing lung diseases, lower GI endoscopy and antibiotics. Species protecting against hospital-related C. difficile acquisition included Gemmiger spp. Odoribacter splanchnicus, Ruminococcus bromii and other Ruminococcus spp. Metagenomic pathway analysis identified steroid biosynthesis as the most underrepresented metabolic pathway in patients who later acquire C. difficile colonization. Conclusions Gemmiger spp. Odoribacter splanchnicus, Ruminococcus bromii and other Ruminococci were associated with a decreased risk of C. difficile acquisition. Clinical Trials Registration DRKS00005335. [ABSTRACT FROM AUTHOR]

Toxin-producing Corynebacterium ulcerans, a causative agent of diphtheria in humans, was isolated from 53 hedgehogs in Belgium during the spring of 2020. Isolates showed low levels of acquired antimicrobial drug resistance. Strain diversity suggests emergence from an endemic situation. These findings stress the need for raising public awareness and improved wildlife disease surveillance. [ABSTRACT FROM AUTHOR]

This study was carried out to evaluate the prevalence of mycobiota and aflatoxin (AF) B1 contamination in sesame seeds from rainfed and irrigated zones of the Punjab, Pakistan. For this purpose, 100 sesame seed samples were collected directly from the fields of major sesame-producing areas in rainfed and irrigated zones. The agar plate method was used for isolation of mycobiota, and thin-layer chromatography was used to determine AFB1 concentrations. Seed samples were then stored for 12 months. After 12 months, the seeds were again analyzed for mycobiota and AFB1 for comparison. All samples were positive for fungal growth under fresh and stored conditions. Twenty-one fungal species of 10 genera were isolated. Aspergillus flavus was the most prevalent contaminant found in fresh and stored sesame seeds from rainfed and irrigated zones, followed by Aspergillus niger, Alternaria alternata, and Fusarium oxysporum. The least prevalent fungi were Aspergillus ochraceus and Cladosporium oxysporum. Analysis revealed that 92% of fresh and 99% of stored seed samples were contaminated with AFB1. In the rainfed zone, 88% of fresh and 100% of stored seed samples were contaminated with AFB1, with mean concentrations of 15.74 and 33.8 ppb, respectively. In the irrigated zone, 96% of fresh and 98% of stored seed samples were contaminated with AFB1, with mean concentrations of 20.5 and 27.56 ppb, respectively. AFB1 concentrations >20 ppb were found in 20% of fresh and 100% of stored seeds samples from the rainfed zone and in 28% of fresh and 60% of stored samples from the irrigated zone and thus were not fit for human consumption as per the maximum limit (20 ppb) assigned by the U.S. Food and Drug Administration and the Food and Agriculture Organization of the United Nations. This report is the first on the mycobiota and AFB1 contamination in sesame seeds from rainfed and irrigated zones of the Punjab, Pakistan. These baseline data are an initial step in the effort to deal with this significant food safety issue. Fresh and stored sesame seeds were positive for fungal growth. The leading contaminant in fresh and stored sesame seeds was Aspergillus flavus. Overall, 92% of fresh and 99% of stored seed samples were contaminated with AFB1. In the rainfed zone, 88% of fresh and 100% of stored samples were contaminated with AFB1. In the irrigated zone, 96% of fresh and 98% of stored samples were contaminated with AFB1. [ABSTRACT FROM AUTHOR]

Abstract Background Diphtheria has been reported as an outbreak in some regions in Indonesia, most especially in East Java Province. Resistance to penicillin, erythromycin, and other antibiotics, single or multiple, has been reported in several studies. This study aims to evaluate the first-line antibiotic susceptibility pattern of toxigenic Corynebacterium diphtheriae isolates. Methods This descriptive observational study was performed from August to November 2018. C. diphtheriae isolates were collected from diphtheria patients and carriers in East Java from 2012 to 2017 and kept at the Balai Besar Laboratorium Kesehatan Daerah Surabaya or the Public Health Laboratory of Surabaya. Sample selection was done by random cluster sampling. The sensitivity test by E-test®of the five antibiotics (penicillin, oxacillin, erythromycin, azithromycin, and clarithromycin) was done to determine the minimum inhibitory concentration (MIC). The Clinical and Laboratory Standards Institute M45A (2015) Corynebacterium spp. for penicillin and erythromycin was used as standard. Results From 114 targeted isolates, 108 were viable and toxigenic. The E-test was performed on the viable isolates. The majority of the hosts were male (58.3%), with median (range) age of 6.5 (1–14) years. Half of the samples were from the 1 to 5-year-old age group. The isolates were acquired much more from patients (78.7%) than carriers (21.3%) and from pharyngeal swab (74.1%). Most of these isolates were from Madura Island (47.2%) and the northern and eastern parts of the province (horseshoe area). Mitis isolates were the major variant (76.9%). The susceptibility pattern of C. diphtheriae to erythromycin was better than that to penicillin. The E-test result for penicillin was 68.52% susceptible, 31.48% intermediate, and 0% resistant (MIC range, 256 μg/L) was 85.2% susceptible, 12% intermediate, and 2.8% resistant The MIC range for oxacillin was 1 to 96 μg/L, while for both azithromycin and clarithromycin were 256 μg/L. Conclusion The susceptibility rate of C. diphtheriae to erythromycin is higher than that to penicillin. The regular update of antibiotic selection to the national guidelines is recommended. The MIC reference standard to azithromycin and clarithromycin is also needed.

ABSTRACT: Bovine mastitis has a negative impact on milk production and can pose risks to public health. The present study aimed to evaluate the quality of bovine milk from small farms in the Botucatu/SP region. Somatic cell counts (SCC), identification of pathogens involved in mastitis, and sensitivity antimicrobial profile of staphylococci isolated were performed. The presence of enterotoxin encoding genes in isolates of staphylococci obtained from milk was investigated. Milk samples from individual mammary quarters of cows were submitted to the California mastitis test (CMT) and SCC. Of the 239 dairy cows from 21 dairy herds evaluated (mean = 11.4 animals/property), two cows (0.8%) presented clinical mastitis and 86 (35.9%) subclinical mastitis. Bacterial culture was performed in 177 quarter milk samples. Staphylococci were identified in 55 (31.1%), corynebacteria in 45 (25.4%), streptococci in 25 (14.1%) and coliforms in four (2.3%) milk samples. Average SCC from culture-positive samples was 1598x103 cells/mL, in case of staphylococci was 1362x103 cells/ml, streptococci was 2857x103 cells/mL, corynebacteria was 976x103 cells/mL and in the cases of coliforms 1161x103 cells/mL were obtained. Staphylococci showed a high sensitivity (>95%) to cephalothin, cotrimoxazole, enrofloxacin, and gentamicin, with a 41.2% resistance to penicillin and 11.8% to oxacillin. Both coagulase positive (CPS) and negative staphylococci (CNS) carried genes encoding enterotoxins in 21.6% of the first group and 41.9% in the second. The sea gene was the most detected 45.8% (n=24) between them, followed by seb with 29.2% and sec with 25.0%. The sed gene was not identified. We highlight the potential risk to public health in the possibility of strains of Staphylococcus spp. enterotoxin-producing genes that can cause staphylococcal food poisoning.

Bacillus cereus is rarely implicated when diarrheal cases in children are diagnosed in developing countries due to the lack of molecular methods to identify its enterotoxigenic genes. We report that out of 62 enterobacteria isolated from 70 stool samples collected from children hospitalized at the Mile 4 Hospital, Ebonyi State, Nigeria, 24 isolates were identified as B. cereus based on 16SrRNA gene sequence. The enterotoxins genes nheA and cytK2 were detected in 23 out of the 24 isolates, while hblC was detected in 19 isolates. B. cereus may be responsible for greater number of yearly incidences of acute childhood gastroenteritis in Nigeria., Competing Interests: Declaration of competing interest Authors Declare No Competing Interest., (Copyright © 2024 Indian Association of Medical Microbiologists. Published by Elsevier B.V. All rights reserved.)

Abstract Objective There is a lack of study in Corynebacterium diphtheriae isolates in Malaysia. The alarming surge of cases in year 2016 lead us to evaluate the local clinical C. diphtheriae strains in Malaysia. We conducted single nucleotide polymorphism phylogenetic analysis on the core and pan-genome as well as toxin and diphtheria toxin repressor (DtxR) genes of Malaysian C. diphtheriae isolates from the year 1986–2016. Results The comparison between core and pan-genomic comparison showed variation in the distribution of C. diphtheriae. The local isolates portrayed a heterogenous trait and a close relationship between Malaysia’s and Belarus’s, Africa’s and India’s strains were observed. A toxigenic C. diphtheriae clone was noted to be circulating in the Malaysian population for nearly 30 years and from our study, the non-toxigenic and toxigenic C. diphtheriae strains can be differentiated significantly into two large clusters, A and B respectively. Analysis against vaccine strain, PW8 portrayed that the amino acid composition of toxin and DtxR in Malaysia’s local strains are well-conserved and there was no functional defect noted. Hence, the change in efficacy of the currently used toxoid vaccine is unlikely to occur.

The genus Corynebacterium includes species of biotechnological, medical and veterinary importance. An atypical C. ulcerans strain, W25, was recently isolated from a case of necrotizing lymphadenitis in a wild boar. In this study, we have analysed the genome sequence of this strain and compared the phenotypic and virulence properties with other corynebacterial pathogens. Phylogenomic analyses revealed that strain W25 belongs to a novel species along with PO100/5 and KL1196. The latter strains were isolated from a pig and a roe deer, respectively; hence, this species appears to be associated to animals. The isolate W25 is likely a non-toxigenic tox gene bearing strain and may have compromised abilities to adhere to pharyngeal and laryngeal epithelial cells due to potential loss of the gene functions in spaBC and spaDEF pilus gene clusters. A number of corynebacterial virulence genes are present including pld encoding phospholipase D. Therefore, this strain may be able to cause severe invasive infections in animals and zoonotic infections in humans. [ABSTRACT FROM AUTHOR]

Cholera is a severe diarrheal disease caused by Vibrio cholerae, a natural inhabitant of brackish water. Effective control of cholera outbreaks depends on prompt detection of the pathogen from clinical specimens and tracking its source in the environment. Although the epidemiology of cholera is well studied, rapid detection of V. cholerae remains a challenge, and data on its abundance in environmental sources are limited. Here, we describe a sensitive molecular quantification assay by qPCR, which can be used on-site in low-resource settings on water without the need for DNA extraction. This newly optimized method exhibited 100% specificity for total V. cholerae as well as V. cholerae O1 and allowed detection of as few as three target CFU per reaction. The limit of detection is as low as 5 × 103 CFU/L of water after concentrating biomass from the sample. The ability to perform qPCR on water samples without DNA extraction, portable features of the equipment, stability of the reagents at 4 °C and user-friendly online software facilitate fast quantitative analysis of V. cholerae. These characteristics make this assay extremely useful for field research in resource-poor settings and could support continuous monitoring in cholera-endemic areas.

Aspergillus flavus is a phytopathogenic fungus able to produce aflatoxin B1 (AFB1), a carcinogenic mycotoxin that can contaminate several crops and food commodities. In A. flavus, two different kinds of strains can co-exist: toxigenic and non-toxigenic strains. Microbial-derived volatile organic compounds (mVOCs) emitted by toxigenic and non-toxigenic strains of A. flavus were analyzed by solid phase microextraction (SPME) coupled with gas chromatography–mass spectrometry (GC-MS) in a time-lapse experiment after inoculation. Among the 84 mVOCs emitted, 44 were previously listed in the scientific literature as specific to A. flavus, namely alcohols (2-methylbutan-1-ol, 3-methylbutan-1-ol, 2-methylpropan-1-ol), aldehydes (2-methylbutanal, 3-methylbutanal), hydrocarbons (toluene, styrene), furans (2,5-dimethylfuran), esters (ethyl 2-methylpropanoate, ethyl 2-methylbutyrate), and terpenes (epizonaren, trans-caryophyllene, valencene, α-copaene, β-himachalene, γ-cadinene, γ-muurolene, δ-cadinene). For the first time, other identified volatile compounds such as α-cadinol, cis-muurola-3,5-diene, α-isocomene, and β-selinene were identified as new mVOCs specific to the toxigenic A. flavus strain. Partial Least Square Analysis (PLSDA) showed a distinct pattern between mVOCs emitted by toxigenic and non-toxigenic A. flavus strains, mostly linked to the diversity of terpenes emitted by the toxigenic strains. In addition, the comparison between mVOCs of the toxigenic strain and its non-AFB1-producing mutant, coupled with a semi-quantification of the mVOCs, revealed a relationship between emitted terpenes (β-chamigrene, α-corocalene) and AFB1 production. This study provides evidence for the first time of mVOCs being linked to the toxigenic character of A. flavus strains, as well as terpenes being able to be correlated to the production of AFB1 due to the study of the mutant. This study could lead to the development of new techniques for the early detection and identification of toxigenic fungi.

Abstract Background Diphtheria remains a major public health concern with multiple recent outbreaks around the world. Moreover, invasive non-toxigenic strains have emerged globally causing severe infections. A diphtheria epidemic in the former Soviet Union in the 1990s resulted in ~5000 deaths. In this study, we analysed the genome sequences of a collection of 93 C. diphtheriae strains collected during and after this outbreak (1996 – 2014) in a former Soviet State, Belarus to understand the evolutionary dynamics and virulence capacities of these strains. Results C. diphtheriae strains from Belarus belong to ten sequence types (STs). Two major clones, non-toxigenic ST5 and toxigenic ST8, encompassed 76% of the isolates that are associated with sore throat and diphtheria in patients, respectively. Core genomic diversity is limited within outbreak-associated ST8 with relatively higher mutation rates (8.9 × 10−7 substitutions per strain per year) than ST5 (5.6 × 10−7 substitutions per strain per year) where most of the diversity was introduced by recombination. A variation in the virulence gene repertoire including the presence of tox gene is likely responsible for pathogenic differences between different strains. However, strains with similar virulence potential can cause disease in some individuals and remain asymptomatic in others. Eight synonymous single nucleotide polymorphisms were observed between the tox genes of the vaccine strain PW8 and other toxigenic strains of ST8, ST25, ST28, ST41 and non-toxigenic tox gene-bearing (NTTB) ST40 strains. A single nucleotide deletion at position 52 in the tox gene resulted in the frameshift in ST40 isolates, converting them into NTTB strains. Conclusions Non-toxigenic C. diphtheriae ST5 and toxigenic ST8 strains have been endemic in Belarus both during and after the epidemic in 1990s. A high vaccine coverage has effectively controlled diphtheria in Belarus; however, non-toxigenic strains continue to circulate in the population. Recombination is an important evolutionary force in shaping the genomic diversity in C. diphtheriae. However, the relative role of recombination and mutations in diversification varies between different clones.

Background:Clostridium difficile (CD) is a major cause of healthcare-associated infections and antibiotic-associated diarrhea in hospitalized patients worldwide. Carriers of toxigenic CD (tCD) have a higher risk of developing CD infections and can transmit CD to the environment and susceptible patients. However, little is known regarding the carriers and transmission of tCD in China.Methods: A multi-center cross-sectional study of tCD colonization (tCDC) was conducted from October 24 to 31, 2014, at 33 hospitals in Shanghai, China. Rectal swabs or stool samples were collected and tested, and the clinical and demographic status, epidemiological data, and blood parameters of 531 participants were recorded. The status of tCDC was defined by a positive result on the nucleic acid amplification test for the tcdA (toxin A), tcdB (toxin B), and cdtAB (toxin CDT) genes after positive bacterial culture.Results: The overall prevalence of CD colonization (CDC) was 19.02%, tCDC accounted for 92.08%, and A+B+CDT– was the dominant genotype (87.13%). The CD infection (CDI) prevalence was 1.51%. Potential tCDC-associated factors were admission to secondary grade hospitals, a body mass index

We report a fatal autochthonous diphtheria case in a migrant worker in Singapore. This case highlights the risk for individual cases in undervaccinated subpopulations, despite high vaccination coverage in the general population. Prompt implementation of public health measures and maintaining immunization coverage are critical to prevent reemergence of diphtheria.

Background: Clostridium difficile (CD) is a major cause of healthcare-associated infections and antibiotic-associated diarrhea in hospitalized patients worldwide. Carriers of toxigenic CD (tCD) have a higher risk of developing CD infections and can transmit CD to the environment and susceptible patients. However, little is known regarding the carriers and transmission of tCD in China. Methods: A multi-center cross-sectional study of tCD colonization (tCDC) was conducted from October 24 to 31, 2014, at 33 hospitals in Shanghai, China. Rectal swabs or stool samples were collected and tested, and the clinical and demographic status, epidemiological data, and blood parameters of 531 participants were recorded. The status of tCDC was defined by a positive result on the nucleic acid amplification test for the tcdA (toxin A), tcdB (toxin B), and cdtAB (toxin CDT) genes after positive bacterial culture. Results: The overall prevalence of CD colonization (CDC) was 19.02%, tCDC accounted for 92.08%, and A+B+CDT– was the dominant genotype (87.13%). The CD infection (CDI) prevalence was 1.51%. Potential tCDC-associated factors were admission to secondary grade hospitals, a body mass index <18.5, hospitalization during the previous 30 days, underlying diseases (including hypertension, diabetes mellitus, coronary heart disease, and respiratory failure), diarrhea during the previous 7 days, and exposure to fluoroquinolones or lansoprazole. Conclusions: This study reveals the prevalence of CDC and tCDC in Shanghai, elucidates several associated factors, contributes to the awareness of the current epidemiology in parts of eastern China and provides new insights for further study and infection control practices. [ABSTRACT FROM AUTHOR]

Background: Diphtheria has been reported as an outbreak in some regions in Indonesia, most especially in East Java Province. Resistance to penicillin, erythromycin, and other antibiotics, single or multiple, has been reported in several studies. This study aims to evaluate the first-line antibiotic susceptibility pattern of toxigenic Corynebacterium diphtheriae isolates.Methods: This descriptive observational study was performed from August to November 2018. C. diphtheriae isolates were collected from diphtheria patients and carriers in East Java from 2012 to 2017 and kept at the Balai Besar Laboratorium Kesehatan Daerah Surabaya or the Public Health Laboratory of Surabaya. Sample selection was done by random cluster sampling. The sensitivity test by E-test®of the five antibiotics (penicillin, oxacillin, erythromycin, azithromycin, and clarithromycin) was done to determine the minimum inhibitory concentration (MIC). The Clinical and Laboratory Standards Institute M45A (2015) Corynebacterium spp. for penicillin and erythromycin was used as standard.Results: From 114 targeted isolates, 108 were viable and toxigenic. The E-test was performed on the viable isolates. The majority of the hosts were male (58.3%), with median (range) age of 6.5 (1-14) years. Half of the samples were from the 1 to 5-year-old age group. The isolates were acquired much more from patients (78.7%) than carriers (21.3%) and from pharyngeal swab (74.1%). Most of these isolates were from Madura Island (47.2%) and the northern and eastern parts of the province (horseshoe area). Mitis isolates were the major variant (76.9%). The susceptibility pattern of C. diphtheriae to erythromycin was better than that to penicillin. The E-test result for penicillin was 68.52% susceptible, 31.48% intermediate, and 0% resistant (MIC range, < 0.016 to 2 μg/L) and for erythromycin (MIC range, < 0.016 to > 256 μg/L) was 85.2% susceptible, 12% intermediate, and 2.8% resistant The MIC range for oxacillin was 1 to 96 μg/L, while for both azithromycin and clarithromycin were <  0.016 to > 256 μg/L.Conclusion: The susceptibility rate of C. diphtheriae to erythromycin is higher than that to penicillin. The regular update of antibiotic selection to the national guidelines is recommended. The MIC reference standard to azithromycin and clarithromycin is also needed. [ABSTRACT FROM AUTHOR]

Objectives: Concern exists regarding overdiagnosis of Clostridium difficile infection (CDI) via molecular modalities. We determined effects of a preanalytic order intervention on laboratory and CDI prevention measures in a multihospital system.Methods: Intervals before and following implementation of a CDI electronic order alert (relative to appropriate testing scenario) were assessed for C difficile test volume and positivity rate, hospital-onset CDI frequency, and hospital-onset C difficile standardized infection ratio (SIR). C difficile detection occurred by PCR throughout the study.Results: During the first half of 2015, testing volume was 1,578, with 88 hospital-onset CDIs. Following implementation, 18.9% and 56.8% reductions in volume and hospital-onset CDIs were realized, respectively, in the first half of 2017. Regression analysis revealed decreasing trends in PCR volume, positivity rate, hospital-onset CDI frequency, and SIR in larger facilities.Conclusions: Preanalytic considerations affect not only the microbiology laboratory but also hospital infection prevention in the context of CDI. [ABSTRACT FROM AUTHOR]

Objectives: Rates of Clostridioides (Clostridium) difficile infection (CDI) are higher in North Wales than elsewhere in the UK. We used WGS to investigate if this is due to increased healthcare-associated transmission from other cases.Methods: Healthcare and community C. difficile isolates from patients across North Wales (February-July 2015) from glutamate dehydrogenase (GDH)-positive faecal samples underwent WGS. Data from patient records, hospital management systems and national antimicrobial use surveillance were used.Results: Of the 499 GDH-positive samples, 338 (68%) were sequenced and 299 distinct infections/colonizations were identified, 229/299 (77%) with toxin genes. Only 39/229 (17%) toxigenic isolates were related within ≤2 SNPs to ≥1 infections/colonizations from a previously sampled patient, i.e. demonstrated evidence of possible transmission. Independent predictors of possible transmission included healthcare exposure in the last 12 weeks (P = 0.002, with rates varying by hospital), infection with MLST types ST-1 (ribotype 027) and ST-11 (predominantly ribotype 078) compared with all other toxigenic STs (P < 0.001), and cephalosporin exposure in the potential transmission recipient (P = 0.02). Adjusting for all these factors, there was no additional effect of ward workload (P = 0.54) or failure to meet cleaning targets (P = 0.25). Use of antimicrobials is higher in North Wales compared with England and the rest of Wales.Conclusions: Levels of transmission detected by WGS were comparable to previously described rates in endemic settings; other explanations, such as variations in antimicrobial use, are required to explain the high levels of CDI. Cephalosporins are a risk factor for infection with C. difficile from another infected or colonized case. [ABSTRACT FROM AUTHOR]

Background: Clostridioides difficile is a major cause of nosocomial infectious diarrhea in hospitalized patients throughout the world. Methods: A multiplex real-time PCR assay was developed and evaluated in comparison with toxigenic culture (TC) (as gold standard method) for direct detection of toxigenic C. difficile in fecal specimens. The multiplex realtime PCR assay simultaneously detected glutamate dehydrogenase (gluD), toxin A (tcdA), toxin B (tcdB), and binary toxin (cdtB) genes in stool samples. Results: The results of multiplex real-time PCR were compared to those of the TC method in 250 patients suspected of C. difficile infection. The prevalence of positive TC was 13.6%. Forty-two stool samples (16.8%) were determined to be gluD+ using multiplex real-time PCR. These included 35 (83.3%) toxigenic (32 tcdA+, tcdB+ and three tcdB+) and 7 (20.0%) were cdtB+. The multiplex real-time PCR assay had a sensitivity of 91.45%, specificity of 99.54%, and positive and negative predictive values of 97% and 98.6%, respectively, compared to the TC method for diagnosis of C. difficile. The analytical sensitivity of the multiplex real-time PCR assay was estimated to be 102 CFU/g of stools and 0.0200 pg of genomic DNA from culture. The analytical specificity was determined to be 100% by using enteric and non-C. difficile standard bacterial strains. Conclusions: The molecular method developed in the study was rapid, sensitive, and specific for detection of toxigenic C. difficile. It is applicable to be performed in clinical laboratories and correlated well with the results obtained by TC. [ABSTRACT FROM AUTHOR]

For the first time, experiments were performed using an ad hoc laboratory setup to study the mechanisms of the effects of low-intensity ultrasound on toxigenic cyanobacteria Synechocysctis sp. that prevail in the northwestern part of Russia. It is found that ultrasound triggers two stress-protective metabolic mechanisms of toxigenic cyanobacteria: (1) synthesis of polysaccharides and proteins to increase the thickness of cell walls and mucous membranes; (2) synthesis of toxins to suppress the external influence. Both mechanisms are extremely energy-consuming, depleting the vital resources of cyanobacteria and accelerating the process of cell death. [ABSTRACT FROM AUTHOR]

Epidemiology of diphtheria in the southwestern Indian Ocean is poorly documented. We analyzed 14 cases of infection with toxigenic Corynebacterium diphtheriae reported during 2007–2015 in Mayotte, a French department located in this region. Local control of diphtheria is needed to minimize the risk for importation of the bacterium into disease-free areas.

Staphylococcus aureus can produce staphylococcal enterotoxins (SEs) in foods and cause staphylococcal food poisoning. Milk and dairy products are foodstuffs among the most frequently associated with staphylococcal food poisoning. It is also relevant to consider that the transmission of S. aureus is possible either by direct contact with animals or through contaminated food as milk or cheese. The aim of this study was to determine the genetic lineages and the genotypes of antibiotic resistance and virulence factors of S. aureus isolates recovered of different dairy products in Tunisia. We collected 136 samples of various traditional and commercial dairy products in different areas in Tunisia. Bacterial isolates were identified by conventional methods (Gram staining, catalase test, oxidase test, DNase production, and ability to coagulate rabbit plasma). The antimicrobial resistance profile and the virulence gene content were investigated. Molecular typing was performed by spa-typing, agr-typing and pulse-field gel electrophoresis (PFGE). Twenty six (19.2%) of the total analyzed samples (N = 136) were contaminated by methicillin-sensitive S. aureus (MSSA). Antimicrobial sensitivity assays have shown that MSSA isolates exhibit a resistance rate of 73.1% to penicillin, 23.1% to tetracyclin, 19.2% to kanamycin, 15.4% to amikacin, and 3.9% to gentamicin. Furthermore, the following toxin encoding genes have been identified among MSSA: luk ED (24 isolates), luk M (2), see (3), sel (3), sea (2), sej (1), sec (1), seb (1), and tst (1). Results of molecular characterization and phylogenetic relations analysis revealed a considerable genetic diversity. Thirteen different spa-types were detected among the 26 isolated MSSA. S. aureus was attributed to the agr type I (16 isolates), II (2), III (2), and non-typables (6). Sixteen different electrophoretic profiles were revealed. Dairy products could constitute a reservoir of virulent and antibiotic-resistant Staphylococcus aureus. [ABSTRACT FROM AUTHOR]

Ginger and turmeric sold in the open markets and retail outlets in southern Nigeria were sampled between April and August, 2017. This period coincided with the first bimodal peak of the rainy season of the 2017 cropping season. Malt extract agar (MEA) and Dichloran 18% glycerol (DG18) media were used to isolate fungi from samples with or without surface sterilisation. Aspergillus spp isolated were examined for the production of orange-yellow pigmentation and blue fluorescence on the reverse side of the plate on CAM under UV light. Aflatoxin production by Aspergillus flavus on yeast extract sucrose (YES) was verified quantitatively using High Performance Liquid Chromatography (HPLC). Data showed that Fusarium, Penicillium and Aspergillus spp were the dominant fungal flora. Toxigenic isolates of A. flavus; AFg1, AFg3, AFt1, and AFt3 produced both orange-yellow pigmentation and blue fluorescence on CAM. The production of AFB1 and AFB2 on YES medium was confirmed using HPLC. The occurrence of toxigenic fungi indicates that there is a potential risk of mycotoxin contamination in ginger and turmeric consumed in southern Nigeria and problems can arise from contamination with aflatoxins. [ABSTRACT FROM AUTHOR]

Abstract Clostridioides (Clostridium) difficile and Bacillus cereus infections are frequently reported in human individually. However, co-infection of both pathogens in human is extremely rare. In the present study, we reported a case of human enteric disease caused by co-infection of C. difficile and B. cereus in Guizhou, China. The 16S rDNA sequencing result showed that C. difficile GMU1 and B. cereus GMU2 were most related to C. difficile ATCC 9689 and B. cereus ATCC 14579. The toxin genotype of C. difficile GMU1 and B. cereus GMU2 were tcdA + tcdB + tcdC + and bceT + nheA + nheB + nheC +, respectively. Cytotoxicity assay demonstrated that C. difficile GMU1 produced significantly higher toxin B compare to C. difficile 630 stain. In contrast, B. cereus GMU2 has comparable NheA toxin productivity compare to previous report. The antimicrobial susceptibility test showed that the combination of ampicillin and vancomycin was most efficient to inhibit both C. difficile GMU1 and B. cereus GMU2. Highlights • Report a rare case of toxin-producing B. cereus co-infected with C. difficile in China. • Analysis the physiological characteristics of co-infected C. difficile GMU1 and B. cereus GMU2. • Customize antimicrobial regimen (ampicillin and vancomycin) for the patient, which guided by antimicrobial susceptibility test. [ABSTRACT FROM AUTHOR]

Corynebacterium ulcerans is an important zoonotic pathogen which is causing diphtheria-like disease in humans globally. In this study, the genomes of three recently isolated C. ulcerans strains, 4940, 2590 and BR-AD 2649, respectively from an asymptomatic carrier, a patient with pharyngitis and a canine host, were sequenced to investigate their virulence potential. A comparative analysis was performed including the published genome sequences of 16 other C. ulcerans isolates. C. ulcerans strains belong to two lineages; 13 strains are grouped together in lineage 1, and six strains comprise lineage 2. Consistent with the zoonotic nature of C. ulcerans infections, isolates from both the human and canine hosts clustered in both the lineages. Most of the strains possessed spaDEF and spaBC gene clusters along with the virulence genes cpp , pld , cwlH , nanH , rpfI , tspA and vsp1 . The gene encoding Shiga-like toxin was only present in one strain, and 11 strains carried the tox gene encoding the diphtheria-like toxin. However, none of strains 4940, 2590 and BR-AD 2649 carried any toxin genes. These strains varied in the number of prophages in their genomes, which suggests that they play an important role in introducing diversity in C. ulcerans. The pan-genomic analyses revealed a variation in the number of membrane-associated and secreted proteins that may contribute to the variation in pathogenicity among different strains. [ABSTRACT FROM AUTHOR]

Introduction: Toxigenic Clostridium difficile is the major cause of antibiotic-associated diarrhea, colitis, and pseudomembranous colitis. The pathogenicity of C. difficile is related to toxins A&B. Children with cancer are at risk of developing C. difficile infection (CDI) due to increased exposure to antibiotics, immunosuppression, and longer hospital stays. Recently, due to higher sensitivity and specificity of nucleic acid amplification test (NAATs) compared to toxin enzyme immunoassays (EIAs), many laboratories are transitioning to NAATs for detection of CDI. We aimed to use a multiplex PCR to detect the C. difficile toxin genes tcdA, tcdB and tpi in stool of cancerous children. We also aimed to show the effects of chemotherapy regimens on the prevalence of C. difficile in these children. Methods: 105 fecal samples were collected from cancerous children who were hospitalized and undergoing chemotherapy. The presence of tcdA, tcdB, and tpi genes were tested. Results: C. difficile was identified in 17.14% of children and the detection rate of A-B+ strains was higher than A+B+ and A+B- strains. C. difficile was found in 17.8% of children who received single antibiotic (5/28 cases virulence genes were detected in 4 cases) and in 41.4% of patients who received more than one antibiotics (12/29 cases virulence genes were detected in 8 cases﴿. Conclusion: Multiplex PCR is a powerful technique for preliminary screening and rapid detection of C. difficile and its virulence genes in the stool of cancerous children. The prevalence of C. difficile in cases receiving several antibiotics was higher than those receiving single antibiotics.

The comparatively high cost of laboratory detection methods for Clostridium difficile infection (CDI) coupled to a low prevalence rate has resulted in testing algorithms that use cheaper and relatively sensitive screening methods, followed by more specific confirmatory methods. The aim of this prospectively-conducted study from two centres in the UK, and one in the Republic of Ireland was to determine the efficacy of the EntericBio ® realtime C. difficile Assay (EBCD) for the detection of toxigenic C. difficile in stool samples. The EBCD was compared to the in-use testing methods for Clostridium difficile (CD) detection in each centre. In the two UK centres, the EBCD was compared to the C.diff Quik Chek Complete ® kit (Techlab), and discrepancies were tested further using The Xpert ® C. difficile PCR assay (Cepheid) and PCR ribotyping after cultivation using the spore culture method, respectively. In the Irish centre, EBCD comparison was to an algorithm of C. DIFF CHEK™-60 test (Techlab) for screening followed by C. difficile Premier ™ Toxins A&B assay (Meridian Bioscience ® ) in the case of positive results; discrepancies were tested using the Xpert ® C. difficile PCR assay. In a retrospective analysis of data, a total of 947 stool samples were tested, of which eight (0.8%) proved inhibitory to the EBCD assay. Of the 939 valid tests conducted, reported sensitivities of the EBCD were 94.7%, 100% and 97.9%, respectively; specificities were 99.6%, 100% and 100%, respectively; positive predictive values were 94.7%, 100% and 100%, respectively, and negative predictive values were 99.6%, 100% and 99.8%, respectively. The CD positivity rates in the current study ranged between 6.6% and 8.2%. [ABSTRACT FROM AUTHOR]