Comparative study of the gut microbial community structure of Spodoptera frugiperda and Spodoptera literal (Lepidoptera).

Background: Spodoptera frugiperda, the fall armyworm is a destructive invasive pest, and S. litura the tobacco cutworm, is a native species closely related to S. frugiperda. The gut microbiota plays a vital role in insect growth, development, metabolism and immune system. Research on the competition between invasive species and closely related native species has focused on differences in the adaptability of insects to the environment. Little is known about gut symbiotic microbe composition and its role in influencing competitive differences between these two insects. Methods: We used a culture-independent approach targeting the 16S rRNA gene of gut bacteria of 5th instar larvae of S. frugiperda and S. litura. Larvae were reared continuously on maize leaves for five generations. We analyzed the composition, abundance, diversity, and metabolic function of gut microbiomes of S. frugiperda and S. litura larvae. Results: Firmicutes, Proteobacteria, and Bacteroidetes were the dominant bacterial phyla in both species. Enterococcus, ZOR0006, Escherichia, Bacteroides, and Lactobacillus were the genera with the highest abundance in S. frugiperda. Enterococcus, Erysipelatoclostridium, ZOR0006, Enterobacter, and Bacteroides had the highest abundance in S. litura. According to α-diversity analysis, the gut bacterial diversity of S. frugiperda was significantly higher than that of S. litura. KEGG analysis showed 15 significant differences in metabolic pathways between S. frugiperda and S. litura gut bacteria, including transcription, cell growth and death, excretory system and circulatory system pathways. Conclusion: In the same habitat, the larvae of S. frugiperda and S. litura showed significant differences in gut bacterial diversity and community composition. Regarding the composition and function of gut bacteria, the invasive species S. frugiperda may have a competitive advantage over S. litura. This study provides a foundation for developing control strategies for S. frugiperda and S. litura.

Lactic acid bacteria modulate the CncC pathway to enhance resistance to ß-cypermethrin in the oriental fruit fly.

The gut microbiota of insects has been shown to regulate host detoxification enzymes. However, the potential regulatory mechanisms involved remain unknown. Here, we report that gut bacteria increase insecticide resistance by activating the cap "n" collar isoform-C (CncC) pathway through enzymatically generated reactive oxygen species (ROS) in Bactrocera dorsalis. We demonstrated that Enterococcus casseliflavus and Lactococcus lactis, two lactic acid-producing bacteria, increase the resistance of B. dorsalis to ß-cypermethrin by regulating cytochrome P450 (P450) enzymes and α-glutathione S-transferase (GST) activities. These gut symbionts also induced the expression of CncC and muscle aponeurosis fibromatosis. BdCncC knockdown led to a decrease in resistance caused by gut bacteria. Ingestion of the ROS scavenger vitamin C in resistant strain affected the expression of BdCncC/BdKeap1/BdMafK, resulting in reduced P450 and GST activity. Furthermore, feeding with E. casseliflavus or L. lactis showed that BdNOX5 increased ROS production, and BdNOX5 knockdown affected the expression of the BdCncC/BdMafK pathway and detoxification genes. Moreover, lactic acid feeding activated the ROS-associated regulation of P450 and GST activity. Collectively, our findings indicate that symbiotic gut bacteria modulate intestinal detoxification pathways by affecting physiological biochemistry, thus providing new insights into the involvement of insect gut microbes in the development of insecticide resistance.

Relationship between heart failure and intestinal inflammation in infants with congenital heart disease.

OBJECTIVE: The association between heart failure (HF) and intestinal inflammation caused by a disturbed intestinal microbiota in infants with congenital heart disease (CHD) was investigated. METHODS: Twenty infants with HF and CHD who were admitted to our hospital between October 2021 and March 2022 were included in this study. Twenty age- and sex-matched infants without HF at our hospital were selected as the control group. Faecal samples were obtained from each participant and analysed by enzyme-linked immunoassay and 16 S rDNA sequencing to assess intestinal inflammatory factors and the microbiota. RESULTS: The levels of intestinal inflammatory factors, including IL-1ß, IL-4, IL-6, IL-17 A and TNF-α, were greatly increased, while the levels of IL-10 were significantly decreased in the HF group compared to the control group (p < 0.05). The intestinal microbial diversity of patients in the HF group was markedly lower than that in the control group (p < 0.05). The abundance of Enterococcus was significantly increased in the HF group compared to the control group (p < 0.05), but the abundance of Bifidobacterium was significantly decreased in the HF group compared to the control group (p < 0.05). The diversity of the intestinal microbiota was negatively correlated with the levels of IL-1ß, IL-4, IL-6 and TNF-α in the intestinal tract but was positively correlated with that of IL-10. The abundance of Enterococcus was positively associated with the levels of IL-1ß, IL-4, IL-6 and TNF-α in the intestinal tract but was negatively correlated with that of IL-10. NT-proBNP was positively associated with the levels of IL-1ß, IL-4, IL-6 and TNF-α in the HF group but was negatively correlated with that of IL-10. The heart function score was positively associated with the levels of IL-1ß, IL-4, IL-6 and TNF-α in the HF group but was negatively correlated with that of IL-10. CONCLUSIONS: Infants with CHD-related HF had a disordered intestinal microbiota, decreased diversity of intestinal microbes, increased levels of pathogenic bacteria and decreased levels of beneficial bacteria. The increased abundance of Enterococcus and the significant decrease in the diversity of the intestinal microbiota may exacerbate the intestinal inflammatory response, which may be associated with the progression of HF.

Complete genome sequence of cfr(B)-carrying Enterococcus raffinosus isolated from bile in a patient in Japan.

OBJECTIVES: Linezolid is an antibiotic used to treat infectious diseases caused by vancomycin-resistant enterococci and methicillin-resistant Staphylococcus aureus. Recently, Enterococcus Spp.-carrying mobile linezolid resistance genes were reported. Herein, we report the complete genome sequence of Enterococcus raffinosus JARB-HU0741, which was isolated from a bile sample of a patient in Japan on May 5, 2021, and carries a linezolid resistance gene, cfr(B). Nevertheless, this isolate was susceptible to linezolid. METHODS: Whole-genome sequencing was performed using HiSeq X FIVE (Illumina) and GridION (Oxford Nanopore Technologies). The sequence reads were assembled using Unicycler v0.4.8, and the complete genome was annotated using DFAST v1.2.18. Antimicrobial resistance genes were detected with Abricate v1.0.1, using the ResFinder database. The minimum inhibitory concentrations (MICs) were determined using broth microdilution and interpreted according to the guidelines of the Clinical and Laboratory Standards Institute. RESULTS: E. raffinosus JARB-HU0741 contained a 3 248 808-bp chromosome and a 1 156 277-bp megaplasmid. cfr(B) was present in the Tn6218-like transposon, which was inserted into a gene encoding a PRD domain-containing protein present in the megaplasmid, but the isolate was susceptible to linezolid (MIC, 0.5 µg/mL). The Tn6218-like transposon was similar to the Tn6218 of Clostridioides difficile Ox3196 and the Tn6218-like transposon of Enterococcus faecium UW11733; however, three genes encoding a topoisomerase, an S-adenosylmethionine-dependent methyltransferase, and a TetR family transcriptional regulator were present in the previous Tn6218- or Tn6218-like transposon. CONCLUSION: This is the first report of the complete genome sequence of E. raffinosus carrying cfr(B). E. raffinosus carrying cfr(B) without linezolid resistance poses a threat, as it could serve as a reservoir for mobile linezolid resistance genes.

Detection of faecal bacteria and antibiotic resistance genes in biofilms attached to plastics from human-impacted coastal areas.

Plastics have been proposed as vectors of bacteria as they act as a substrate for biofilms. In this study, we evaluated the abundance of faecal and marine bacteria and antibiotic resistance genes (ARGs) from biofilms adhered to marine plastics. Floating plastics and plastics from sediments were collected in coastal areas impacted by human faecal pollution in the northwestern Mediterranean Sea. Culture and/or molecular methods were used to quantify faecal indicators (E. coli, Enterococci and crAssphage), and the ARGs sulI, tetW and blaTEM and the 16S rRNA were detected by qPCR assays. Pseudomonas and Vibrio species and heterotrophic marine bacteria were also analysed via culture-based methods. Results showed that, plastic particles covered by bacterial biofilms, primarily consisted of marine bacteria including Vibrio spp. Some floating plastics had a low concentration of viable E. coli and Enterococci (42% and 67% of the plastics respectively). Considering the median area of the plastics, we detected an average of 68 cfu E. coli per item, while a higher concentration of E. coli was detected on individual plastic items, when compared with 100 ml of the surrounding water. Using qPCR, we quantified higher values of faecal indicators which included inactive and dead microorganisms, detecting up to 2.6 × 102 gc mm-2. The ARGs were detected in 67-88% of the floating plastics and in 29-57% of the sediment plastics with a concentration of up to 6.7 × 102 gc mm-2. Furthermore, enrichment of these genes was observed in biofilms compared with the surrounding water. These results show that floating plastics act as a conduit for both the attachment and transport of faecal microorganisms. In contrast, low presence of faecal indicators was detected in plastic from seafloor sediments. Therefore, although in low concentrations, faecal bacteria, and potential pathogens, were identified in marine plastics, further suggesting plastics act as a reservoir of pathogens and ARGs.

Australian Group on Antimicrobial Resistance (AGAR) Australian Enterococcal Surveillance Outcome Program (AESOP).

Abstract: From 1 January to 31 December 2021, forty-eight institutions around Australia participated in the Australian Enterococcal Surveillance Outcome Programme (AESOP). The aim of AESOP 2021 was to determine the proportion of enterococcal bacteraemia isolates in Australia that were antimicrobial resistant, and to characterise the molecular epidemiology of the Enterococcus faecium isolates. Of the 1,297 unique episodes of enterococcal bacteraemia investigated, 94.4% were caused by either E. faecalis (54.1%) or E. faecium (40.3%). Ampicillin resistance was detected in one E. faecalis isolate and in 89.3% of E. faecium isolates. Vancomycin non-susceptibility was not detected in E. faecalis but was detected in 37.9% of E. faecium. Overall, 39.6% of E. faecium harboured the vanA and/or vanB genes. For the vanA/vanB positive E. faecium isolates, 35.8% harboured the vanA gene and 64.2% the vanB gene. Although the percentage of vancomycin-resistant E. faecium bacteraemia isolates was significantly lower than that reported in the 2020 AESOP report (presumably due to the COVID-19 elective surgery restrictions placed on hospitals), it remains substantially higher than that recorded in most European countries. Isolates of E. faecium consisted of 73 multi-locus sequence types (STs); 77.2% of isolates were classified into seven major STs each containing more than ten isolates. All major STs belonged to clonal cluster (CC) 17, a major hospital-adapted polyclonal E. faecium cluster. The major STs (ST17, ST1424, ST796, ST78, ST80, ST1421 and ST555) were found across most regions of Australia. The predominant ST was ST17 which was identified in all regions except the Northern Territory. Overall, 46.5% of isolates belonging to the seven major STs harboured the vanA or vanB gene. The AESOP 2021 has shown that enterococcal bacteraemia episodes in Australia are frequently caused by polyclonal ampicillin-resistant high-level gentamicin resistant vanA- or vanB-positive E. faecium which have limited treatment options.

Vagococcus allomyrinae sp. nov. and Enterococcus larvae sp. nov., isolated from larvae of Allomyrina dichotoma.

The taxonomic positions of two novel strains isolated from larvae of an insect (Allomyrina dichotoma) collected in Jeju, Republic of Korea, were determined by a polyphasic approach. Strain BWB3-3T was closely related to the type strain of Vagococcus salmoninarum, having 97.2 % 16S rRNA gene sequence similarity, whereas strain BWM-S5T formed an independent cluster within the genus Enterococcus in the 16S rRNA gene phylogeny and the closest relative was the type strain of Enterococcus canis (98.1 % sequence similarity). The core gene analysis supported the phylogenetic positions of the isolates revealed by 16S rRNA gene phylogeny. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain BWB3-3T and the type strain of V. salmoninarum were 73.2 and 20.0 %, respectively, whereas strain BWM-S5 T showed an ANI value of 70.9 % with the type strain of Enterococcus canis. The dDDH values between strain BWM-S5T and all the type strains of Enterococcus species were ≤25.1 %. On the basis of the results obtained here, the two isolates are considered to constitute two novel species of the family Enterococcaceae, for which the names Vagococcus allomyrineae sp. nov. and Enterococcus larvae sp. nov. are proposed, with the type strains BWB3-3T (=KCTC 43277T=CCM 9080T) and BWM-S5T (=KACC 22156T=CCM 9075T), respectively.

Australian Group on Antimicrobial Resistance (AGAR) Australian Enterococcal Sepsis Outcome Programme (AESOP) Annual Report 2020.

From 1 January to 31 December 2020, forty-nine institutions around Australia participated in the Australian Enterococcal Sepsis Outcome Programme (AESOP). The aims of AESOP 2020 were to determine the proportion of enterococcal bacteraemia isolates in Australia that were antimicrobial-resistant, and to characterise the molecular epidemiology of the E. faecium isolates. Of the 1,230 unique episodes of enterococcal bacteraemia investigated, 93.9% were caused by either E. faecalis (54.2%) or E. faecium (39.7%). Ampicillin resistance was not detected in E. faecalis but was detected in 88.2% of E. faecium . Vancomycin non-susceptibility was detected in 0.2% of E. faecalis and 32.6% of E. faecium . Overall, 35.2% of E. faecium harboured vanA and/or vanB genes. For the vanA/B positive E. faecium isolates, 38.8% harboured the vanA gene, 60.6% the vanB gene, and 0.6% harboured both vanA and vanB . Although the percentage of E. faecium bacteraemia isolates was significantly lower than that detected in the 2019 AESOP (presumably due to the COVID-19 elective surgery restrictions placed on hospitals), it remains substantially higher than that recorded in most European countries. The E. faecium isolates detected consisted of 71 multilocus sequence types (STs), with 81.7% of these isolates classified into eight major STs each containing ten or more isolates. All major STs belonged to clonal cluster 17 (CC17), a major hospital-adapted polyclonal E. faecium cluster. The major STs (ST17, ST1424, ST80, ST796, ST78, ST1421, ST555 and ST117) were found across most regions of Australia. The predominant clone was ST17, which was identified in all regions except the Northern Territory. Overall, 40.9% of isolates belonging to the eight major STs harboured the vanA or vanB gene. The AESOP 2020 has shown enterococcal bacteraemia episodes in Australia are frequently caused by polyclonal ampicillin-resistant high-level gentamicin-resistant vanA - or vanB -positive E. faecium which have limited treatment options.

Characterization of Enterococcus spp. isolated from a fish farming environment in southern Brazil / Caracterização de Enterococcus spp. isolado de um ambiente de piscicultura no sul do Brasil

Abstract The aim of present study is to characterize the resistance and virulence profile of enterococci isolated from aquaculture excavated ponds and masonry tanks (6 samples) in southern Brazil. Samples were cultured in selective medium, 10 colonies were randomly selected from each sample, which were identified by MALDI-TOF and tested against 13 antimicrobials. The presence of resistance (tetL, tetM, tetS, ermB and msrC) and virulence (ace, esp, agg, cylA and gelE) genes were determined by PCR. A total of 79 enterococci were identified, and Entecococcus faecalis (44.3%) and E. casseliflavus (36.7%) were the most prevalent species isolated. Sixty-five strains (82.3%) were resistant to at least one of the antimicrobials tested, whereas 27 (34.2%) strains were multiresistant. The overall percentages of antimicrobial resistant isolates were: 58.2% to rifampicin, 40.5% to fluoroquinolones, 36.7% to erythromycin and 30.4% to tetracycline. The tetL and tetM genes were found in 57.7% of the tetracycline-resistant strains; and msrC in 31.01% of erythromycin-resistant strains. The most frequently detected virulence factors were ace and gelE genes. Although limited to a single farm, these data suggest that aquaculture may be a reservoir of resistant and virulent enterococci. This study is the first step towards enhancing our understandingof distribution, resistance and virulence profile in enterococci isolated from fish farming environments in the south Brazil.

Resumo O objetivo do estudo apresentado é caracterizar o perfil de resistência e virulência de enterococos isolados de viveiros escavados e tanques de alvenaria (6 amostras) de uma pisicultura no Sul do Brasil. As amostras foram cultivadas em meio seletivo, 10 colônias foram selecionadas aleatoriamente de cada amostra, que foram identificadas por MALDI-TOF e testadas contra 13 antimicrobianos. A presença de genes de resistência (tetL, tetM, tetS, ermB e msrC) e virulência (ace, esp, agg, cylA e gelE) foi determinada por PCR. Foram identificados 79 enterococos, sendo Entecococcus faecalis (44,3%) e E. casseliflavus (36,7%) as espécies mais frequentes isoladas. Sessenta e cinco cepas (82,3%) eram resistentes a pelo menos um dos antimicrobianos testados, enquanto 27 (34,2%) eram multirresistentes. As porcentagens gerais de isolados resistentes a antimicrobianos foram: 58,2% para rifampicina, 40,5% para fluoroquinolonas, 36,7% para eritromicina e 30,4% para tetraciclina. Os genes tetL e tetM foram encontrados em 57,7% das cepas resistentes à tetraciclina; e msrC em 31,01% das cepas resistentes à eritromicina. Os fatores de virulência mais comumente detectados foram ace e gelE. Embora limitados a uma única fazenda, esses dados indicam que a aquicultura pode ser uma fonte de enterococos resistentes e virulentos. Este estudo é o primeiro passo para melhorar nosso entendimento da distribuição, resistência e perfil de virulência em enterococos isolados de ambientes de piscicultura no sul do Brasil.

Bile Microbiota in Patients with Pigment Common Bile Duct Stones.

BACKGROUND: Common bile duct (CBD) stone is one of the most prevalent gastroenterological diseases, but the role played by biliary microbiota in the pathogenesis of CBD stones remains obscure. The aim of this study was to investigate the characteristics of the biliary tract core microbiome and its potential association with the formation of pigment stones. METHODS: Twenty-eight patients with biliary obstruction of various causes were enrolled. Thirteen had new-onset pigment CBD stone. Of the remaining 15, four had benign biliary stricture, four had gallbladder cancer, three had pancreatic cancer, 3 had distal CBD cancer, and one had hepatocellular carcinoma. Endoscopic retrograde cholangiopancreatography was used to collect bile samples for DNA extraction, 16S ribosomal RNA gene sequencing, and bile microbiota composition analysis. RESULTS: Proteobacteria (61.7%), Firmicutes (25.1%), Bacteroidetes (5%), Fusobacteria (4.6%), and Actinobacteria (2.6%) were the most dominant phyla in the bile of the 28 study subjects. A comparison between new-onset choledocholithiasis and other causes of biliary obstruction (controls) showed Enterococcus was found to be significantly abundant in the CBD stone group at the genus level (linear discriminant analysis score = 4.38; P = 0.03). However, no other significant compositional difference was observed. CONCLUSION: This study demonstrates an abundance of microbiota in bile juice and presents a biliary microbiome composition similar to that of duodenum. The study also shows Enterococcus was significantly abundant in the bile juice of patients with a brown pigment stone than in controls, which suggests Enterococcus may play an important role in the development of pigment stones.

Anti-Proliferative and Anti-Biofilm Potentials of Bacteriocins Produced by Non-Pathogenic Enterococcus sp.

The incidence of cancer is increasing worldwide; likewise, the emergence of antibiotic-resistant biofilm-forming pathogens has led to a tremendous increase in morbidity and mortality. This study aimed to evaluate the probiotic properties of bacteriocin-producing Enterococcus sp. with a focus on their anti-biofilm and anticancer activities. Three of 79 Enterococcus isolates (FM43, FM65, FM50) were identified as producers of broad-spectrum bioactive molecules and were molecularly characterized as Enterococcus faecium by 16S rRNA sequencing. Phenotypic and genotypic screening for potential virulence factors revealed no factors known to promote pathogenicity. Treatment with proteinase K resulted in diminished antimicrobial activity; PCR-based screening for bacteriocin genes suggested the presence of both entA and entB genes that encode enterocins A and B, respectively. Maximum antimicrobial activity was detected during the early stationary phase, while activity disappeared after 24 h in culture. Bacteriocins from these isolates were stable at high temperatures and over a wide range of pH. Interestingly, crude supernatants of Ent. faecium FM43 and Ent. faecium FM50 resulted in significant destruction (80% and 48%, respectively; P < 0.05) of Streptococcus mutans ATCC 25175-associated preformed biofilms. Moreover, in vitro cytotoxicity assays revealed that extracts from Ent. faecium isolates FM43, FM65, and FM50 inhibited Caco-2 cell proliferation by 76.9%, 70%, and 85.3%, respectively. Taken together, the multifunctional capabilities of the microbial-derived proteins identified in our study suggest potentially important roles as alternative treatments for biofilm-associated infections and cancer.

The Gut Microbiome in Patients With Chronic Pancreatitis Is Characterized by Significant Dysbiosis and Overgrowth by Opportunistic Pathogens.

INTRODUCTION: Exocrine pancreatic function is a critical host factor in determining the intestinal microbiota composition. Diseases affecting the exocrine pancreas could therefore influence the gut microbiome. We investigated the changes in gut microbiota of patients with chronic pancreatitis (CP). METHODS: Patients with clinical and imaging evidence of CP (n = 51) were prospectively recruited and compared with twice the number of nonpancreatic disease controls matched for distribution in age, sex, body mass index, smoking, diabetes mellitus, and exocrine pancreatic function (stool elastase). From stool samples of these 153 subjects, DNA was extracted, and intestinal microbiota composition was determined by bacterial 16S ribosomal RNA gene sequencing. RESULTS: Patients with CP exhibited severely reduced microbial diversity (Shannon diversity index and Simpson diversity number, P < 0.001) with an increased abundance of facultative pathogenic organisms (P < 0.001) such as Enterococcus (q < 0.001), Streptococcus (q < 0.001), and Escherichia.Shigella (q = 0.002). The CP-associated changes were independent of exocrine pancreatic insufficiency. Short-chain fatty acid producers, considered protective for epithelia such as Faecalibacterium (q < 0.001), showed reduced abundance in patients with CP. Of 4 additional patients with CP previously treated with antibiotics (ceftriaxone and metronidazole), 3 patients were characterized by distinct Enterococcus overgrowth. DISCUSSION: CP is associated with marked gut microbiota dysbiosis, greatly reduced diversity, and increased abundance of opportunistic pathogens, specifically those previously isolated from infected pancreatic necrosis. Taxa with a potentially beneficial role in intestinal barrier function are depleted. These changes can increase the probability of complications from pancreatitis such as infected fluid collections or small intestinal bacterial overgrowth (see Graphical Abstract, Supplementary Digital Content 1, http://links.lww.com/CTG/A383).

Polymicrobial infections and microbial patterns in infected nonunions - a descriptive analysis of 42 cases.

BACKGROUND: Frequencies of polymicrobial infection and pathogens evidenced in course of infected nonunion treatment are largely unknown. Therefore, this study aims at investigating microbial patterns in infected nonunions. METHODS: Surgically treated patients with long bone infected nonunion admitted between January 2010 and March 2018 were included in the study. Microbiological culture and polymerase-chain-reaction results of tissue samples of initial and follow-up revision surgeries were assessed and compared with patient and treatment characteristics. RESULTS: Forty two patients with a mean age of 53.9 ± 17.7 years were included. In six patients (14.3%) polymicrobial infection was evident. A change of pathogens evidenced in course of the treatment occurred in 21 patients (50%). In 16 patients (38.1%) previously detected bacteria could be determined by microbial testing after further revision surgery. Staphylococcus aureus was most often detected (n = 34, 30.6%), followed by Enterococcus spp. (n = 25, 22.5%) and Staphylococcus epidermidis (n = 18, 16.2%). Five Staphylococcus aureus were resistant to methicillin (MRSA). In patients without polymicrobial infection or further germ detection in course of the treatment, 86.4% of the infections were due to Staphylococcus spp.. Infections due to Streptococcus spp. and gram-negative bacteria were only present in patients with polymicrobial infection and germ-change in course of the treatment. CONCLUSION: A low rate of polymicrobial infections was evidenced in the present study. Germ-change often occurs in course of revision surgeries. Prospective studies with more sensitive diagnostic tools are necessary to elucidate the therapeutical relevance of microbiological testing results for surgical as well as medical treatment in infected nonunions.

Whole Genome Sequence Analysis of the First Vancomycin-Resistant Enterococcus faecium Isolates from a Libyan Hospital in Tripoli.

The purpose of the study was to investigate the molecular characteristics and genetic relatedness of the first reported cases of vancomycin-resistant enterococci (VRE) from the Tripoli Medical Center, Libya. In total, 43 VRE isolates were obtained from various clinical sites throughout the years 2013-2014, including 40 vanA-type and 2 vanB-type vancomycin-resistant Enterococcus faecium isolates and 1 vanC1-type Enterococcus gallinarum. Of the 42 E. faecium, 19 isolates were subjected to whole genome sequencing. Core genome multilocus sequence typing (cgMLST) analysis revealed three sequence clusters (SCs) of clonally related isolates, which were linked to different hospital wards. The first two VRE isolates, isolated early 2013 from patients in the medical intensive care unit, were grouped in SC1 (MLST [ST] 78, vanB) and differed in only 3 of 1423 cgMLST alleles. The SC2 (n = 16, special care baby unit, neonatal intensive care unit, pediatric surgery ward, and oncology ward) and SC3 (n = 1, antenatal ward) were all ST80 vanA-VRE, but the single SC3 isolate differed in 233 alleles compared with SC2. Within SC2, isolates differed in 1-23 alleles. Comparison with a larger database of E. faecium strains indicated that all isolates clustered within the previously defined hospital clade A1. A combination of Resfinder and mlplasmid analysis identified the presence of resistance genes on different plasmid predicted genetic elements among different SCs. In conclusion, this study documents the first isolates causing outbreaks with VRE in the Libyan health care system. Further surveillance efforts using molecular typing methods to monitor spread of multidrug-resistant bacteria in the Libyan health care system are urgently needed.

Distributions of enterococci and human-specific bacteriophages of enterococci in a tropical watershed.

The bacteriophages of E. faecalis strains AIM06 (DSM100702) and SR14 (DSM100701) have previously been validated as human-specific microbial source tracking (MST) markers in Thailand. In this study, their spatial and temporal distribution in a freshwater river was investigated for the first time (n = 48). The abundance of enterococci as a standard microbial water quality parameter was evaluated by both the qPCR detection assay with primers and a hydrolysis probe according to the US EPA Method 1611 and the US EPA Method 1600 membrane filtration culture method. AIM06 and SR14 phages were detected by a double layer agar assay and were present in 87.5% and 81.3% of all samples with a co-presence of 92.9% of phage-positive samples. After spiking the representative phages, the ranges of recovery efficiencies were 57.9-99.6% and 49.6-99.9% (n = 48) for AIM06 and SR14 phages, respectively. The absolute abundance of AIM06 and SR14 phages ranged from 0.25 to 221.94 and from 0.25 to 76.66 PFU/100 mL, respectively. Enterococci DNA copies and CFU were detected in all samples ranging from 3.24 to 6.32 log10 copies/100 mL and 100.00 to 1593 CFU/100 mL, respectively. Enterococci in the qPCR assay also showed a moderate correlation with the culture method. The AIM06 and SR14 phage results indicated continuing human faecal pollution along the river with no significant different levels among stations. Interestingly, the higher levels of enterococci in downstream stations for both the qPCR and culture methods along with the significant correlation with other faecal indicator organisms and non-human MST markers implied non-human faecal pollution. In conclusion, this study provides insightful information that could lead to effective water quality management and public health risk reduction from exposure to faecally-contaminated water.

Risks associated with enterococci as probiotics.

Probiotics are naturally occurring microorganisms that confer health benefits by altering host commensal microbiota, modulating immunity, enhancing intestinal barrier function, or altering pain perception. Enterococci are human and animal intestinal commensals that are used as probiotics and in food production. These microorganisms, however, express many virulence traits including cytolysin, proteases, aggregation substance, capsular polysaccharide, enterococcal surface protein, biofilm formation, extracellular superoxide, intestinal translocation, and resistance to innate immunity that can lead to serious hospital-acquired infections. In addition, enterococci are facile in acquiring antibiotic resistance genes to many clinically important antibiotics encoded on a wide variety of conjugative plasmids, transposons, and bacteriophages. The pathogenicity and disease burden caused by enterococci render them poor choices as probiotics. No large, randomized, placebo-controlled clinical trials have demonstrated the safety and efficacy of any enterococcal probiotic. As a result, no enterococcal probiotic has been approved by the United States Food and Drug Administration for the treatment, cure, or amelioration of human disease. In 2007, the European Food Safety Authority concluded that enterococci do not meet the standard for "Qualified Presumption of Safety". Enterococcal strains used or proposed for use as probiotics should be carefully screened for efficacy and safety.

Enterococcus: A Predictor of Ravaged Microbiota and Poor Prognosis after Allogeneic Hematopoietic Stem Cell Transplantation.

Intestinal flora plays an essential role in regulating immune responses. Changes in the gut flora are associated with poor prognosis after allogeneic hematopoietic stem cell transplantation (HSCT). We aimed to investigate the impact of diverse intestinal flora on survival after allogeneic HSCT. Using next-generation sequencing of the bacterial 16S ribosomal RNA (rRNA) gene, we found that the intestinal microbiota of patients undergoing allogeneic HSCT differed significantly from that of healthy controls. Furthermore, dysbiosis persisted for at least 1 year after transplantation. Interestingly, increased abundance of the genus Enterococcus detected by 16S rRNA sequencing as early as 1 month after transplantation was correlated with poor survival (overall survival at 2 years post-HSCT, 83.9% for patients with <1% relative abundance of Enterococcus and 47.6% for those with ≥1% relative abundance of Enterococcus), which was undetectable by conventional standard stool culture. These findings suggest that detection of Enterococcus by 16S rRNA analysis reflects compromised intestinal flora and may be a promising prognostic indicator.

Lactose drives Enterococcus expansion to promote graft-versus-host disease.

Disruption of intestinal microbial communities appears to underlie many human illnesses, but the mechanisms that promote this dysbiosis and its adverse consequences are poorly understood. In patients who received allogeneic hematopoietic cell transplantation (allo-HCT), we describe a high incidence of enterococcal expansion, which was associated with graft-versus-host disease (GVHD) and mortality. We found that Enterococcus also expands in the mouse gastrointestinal tract after allo-HCT and exacerbates disease severity in gnotobiotic models. Enterococcus growth is dependent on the disaccharide lactose, and dietary lactose depletion attenuates Enterococcus outgrowth and reduces the severity of GVHD in mice. Allo-HCT patients carrying lactose-nonabsorber genotypes showed compromised clearance of postantibiotic Enterococcus domination. We report lactose as a common nutrient that drives expansion of a commensal bacterium that exacerbates an intestinal and systemic inflammatory disease.

Enterotype-based Analysis of Gut Microbiota along the Conventional Adenoma-Carcinoma Colorectal Cancer Pathway.

The dysbiosis of human gut microbiota is strongly associated with the development of colorectal cancer (CRC). The dysbiotic features of the transition from advanced polyp to early-stage CRC are largely unknown. We performed a 16S rRNA gene sequencing and enterotype-based gut microbiota analysis study. In addition to Bacteroides- and Prevotella-dominated enterotypes, we identified an Escherichia-dominated enterotype. We found that the dysbiotic features of CRC were dissimilar in overall samples and especially Escherichia-dominated enterotype. Besides a higher abundance of Fusobacterium, Enterococcus, and Aeromonas in all CRC faecal microbiota, we found that the most notable characteristic of CRC faecal microbiota was a decreased abundance of potential beneficial butyrate-producing bacteria. Notably, Oscillospira was depleted in the transition from advanced adenoma to stage 0 CRC, whereas Haemophilus was depleted in the transition from stage 0 to early-stage CRC. We further identified 7 different CAGs by analysing bacterial clusters. The abundance of microbiota in cluster 3 significantly increased in the CRC group, whereas that of cluster 5 decreased. The abundance of both cluster 5 and cluster 7 decreased in the Escherichia-dominated enterotype of the CRC group. We present the first enterotype-based faecal microbiota analysis. The gut microbiota of colorectal neoplasms can be influenced by its enterotype.

Caracterização fenotípica e genotípica de Enterococcus spp. isolados de ambiente de processamento de queijo Minas Frescal / Phenotypic and genotypic characterization of Enterococcus spp. isolated from Minas Frescal cheese processing line

O objetivo deste trabalho foi identificar as fontes de contaminação de Enterococcus spp. em uma linha de processamento de queijo Minas Frescal e caracterizar estes micro-organismos quanto à capacidade proteolítica e lipolítica bem como quanto as características de patogenicidade. A partir dos resultados, pode-se verificar que os Enterococcus spp. estavam presentes em amostras de matéria-prima, ambiente e produto final, com destaque para o equipamento de ordenha que apresentou a maior contagem de Enterococcus spp. (5 log UFC/cm2) e a maçaneta que apresentou contaminação persistente ao longo das coletas. Aproximadamente 47% dos isolados apresentaram atividade proteolítica e lipolítica. Além disso, os isolados testados apresentaram resistência múltipla a antibióticos e possuíram múltiplos genes de virulência.