Problems concerning representative sampling for possible contamination in a complex waste processing facility

Objective To explore the positive rate and clinical value of different clinical specimens. Methods a total of 40 960 samples were selected from the microbial inspection department of our hospital from January, 2015 to December, 2018, which were divided into 9 461 samples from January to December, 2015, 9 462 samples from January to December, 2016, 10 668 samples from January to December 2017, and 11 269 samples from January to December, 2018 according to different collection years. All the collected samples were tested in the laboratory of our hospital. The positive ratios of microbiological test in all the clinical specimens of different years were observed and compared. Results In 2015, there were 2 561 blood samples, with a positive rate of 8.0% (204 samples); 1 984 urine samples, with a positive rate of 16.0% (317 samples); 2 353 samples from respiratory tract, with a positive rate of 28.0% (658 samples); 1 741 feces samples, with a positive rate of 12.0% (209 samples); and 822 punctured fluid samples, with a positive rate of 18.9% (155 samples). In 2016, there were 2 602 blood samples, with a positive rate of 14.0% (364 samples); 2 011 urine samples, with a positive rate of 19.1% (385 samples); 2 401 samples from respiratory tract, with a positive rate of 33.1% (795 samples); 1 633 feces samples, with a positive rate of 15.1% (247 samples); and 815 punctured fluid samples, with a positive rate of 21.0% (171 samples). In 2017, there were 2 919 blood samples, with a positive rate of 17.1% (498 samples); 2 392 urine samples, with a positive rate of 21.1% (505 samples); 2 770 samples from respiratory tract, with a positive rate of 38.1% (1 056 samples); 1 845 feces samples, with a positive rate of 16.9% (311 samples); and 742 punctured fluid samples, with a positive rate of 23.5% (174 samples). In 2018, there were 2 996 blood samples, with a positive rate of 19.1% (571 samples); 2 461 urine samples, with a positive rate of 24.1% (593 samples); 2 985 samples from respiratory tract, with a positive rate of 45.9% (1 371 samples); 1 909 feces samples, with a positive rate of 17.8% (340 samples); and 918 punctured fluid samples, with a positive rate of 24.3% (223 samples). According to the analysis results, from 2015 to 2018, the numbers of blood, urine, respiratory, and fecal samples in the hospital increased year by year, with statistical differences between different years (P<0.05). From 2015 to 2018, the positive rates of blood, urine, respiratory, fecal, and puncture fluid samples in the hospital increased year by year, and the positive rates of different clinical samples were significantly different from year to year (P<0.05). Conclusion By analyzing the microbial test results of different periods and clinical specimens in the hospital, we can fully understand the prevalence of pathogenic bacteria in the hospital, and provide more references for clinical diagnosis and treatment. However, in the process of microbial examination of clinical specimens, the quality and accuracy of the examination must be ensured, so as to provide more accurate reference materials for clinical diagnosis and treatment. Key words: Specimen; Microbes; Positive rate; Inspection

Simple SummaryTestosterone is the main reproductive hormone in male vertebrates. Conventional methods to assess testosterone rely on invasive blood sampling procedures, which can induce pain and distress to the animals. Here we successfully validated a non-invasive method to determine testosterone levels by assessing testosterone metabolites (TMs) in excreta of mice. We investigated the effects of sex and daytime on the metabolism and the excretion of TMs and validated the applied EIA to measure TMs. Further, we assessed diurnal fluctuations in TM excretions in both sexes and across strains. We found that males excreted more radiolabeled TMs via their feces compared to females. TM excretion patterns did not differ between sexes but TM excretion occurred faster in urinary than fecal samples. Animals excreted TM faster during the night than during the day. Daytime had no effect on the formed TMs; however, males and females formed different TMs. As expected, males showed higher fecal TM levels than females. Males also showed diurnal fluctuations in their TM levels but we found no differences between mouse strains. Our non-invasive method to assess fecal TMs can be widely used and will benefit various research disciplines.Testosterone is the main reproductive hormone in male vertebrates and conventional methods to measure testosterone rely on invasive blood sampling procedures. Here, we aimed to establish a non-invasive alternative by assessing testosterone metabolites (TMs) in fecal and urinary samples in mice. We performed a radiometabolism study to determine the effects of daytime and sex on the metabolism and excretion pattern of radiolabeled TMs. We performed physiological and biological validations of the applied EIA to measure TMs and assessed diurnal fluctuations in TM excretions in male and female mice and across strains. We found that males excreted significantly more radiolabeled TMs via the feces (59%) compared to females (49.5%). TM excretion patterns differed significantly between urinary and fecal samples and were affected by the daytime of ³H-testosterone injection. Overall, TM excretion occurred faster in urinary than fecal samples. Peak excretion of fecal TMs occurred after 8 h when animals received the 3H-testosterone in the morning, or after 4 h when they received the 3H-testosterone injection in the evening. Daytime had no effect on the formed TMs; however, males and females formed different types of TMs. As expected, males showed higher fecal TM levels than females. Males also showed diurnal fluctuations in their TM levels but we found no differences in the TM levels of C57BL/6J and B6D2F1 hybrid males. Finally, we successfully validated our applied EIA (measuring 17β-hydroxyandrostane) by showing that hCG (human chorionic gonadotropin) administration increased TM levels, whereas castration reduced them. In conclusion, our EIA proved suitable for measuring fecal TMs in mice. Our non-invasive method to assess fecal TMs can be widely used in various research disciplines like animal behavior, reproduction, animal welfare, ecology, conservation, and biomedicine.

Objective: The aim of this work was to report the surveillance and dissemination of NDM-1 positive bacteria in a patient and ward environment. Methods: In 2010, during the therapy for a 51 years old patient, clinical and environmental samples were collected for carbapenem resistant bacterial culture, according to the clinical microbiological examination. Strains identification and antibiotic susceptibility were tested by VITEK Compact 2 system and E-test. The bla(NDM-1) was detected by PCR and analyzed by sequencing. Plasmids containing bla(NDM-1) were submitted to PFGE-S1 and Southern hybridization. Results: During hospitalization from October 1st to November 4th, nine strains were isolated from blood, sputum, urine, fecal, and ward ground samples. The Klebsiella oxytoca, Raoultella planticola, and Acinetobacter baumannii were isolated from blood sample. The Klebsiella pneumonia and Acinetobacter baumannii were isolated from sputum sample. An Acinetobacter lwoffii was isolated from urine sample. An Escherichia coli was isolated from fecal sample. And the Acinetobacter lwoffii and Acinetobacter spp. were isolated from ward ground. Four strains were NDM-1 positive, which were Raoultella planticola (RpNDM1) isolated from blood, Escherichia coli (EcNDM1) isolated from fecal, Acinetobacter lwoffii (AlDNM1) and Acinetobacter spp. (AsNDM1) isolated from ward ground. Four NDM-1 positive strains were resistant to Piperacillin, Piperacillin tazobactam, Cefepime, Ceftriaxone, Ceftazidime, Imipenem, Meropenem, and Ertapenem. Southern hybridization revealed that bla(NDM-1) were all located on plasmids in the four positive strains. Conclusion:bla(NDM-1) can transfer rapidly among different species, resulting in difficult to control and prevent. While isolating patient who is carrying NDM-1 positive strains, more attention should be paid to the disposal of patient's excreta, especially stool, should be paid more attention.

Boldenone (1,4-androstadiene-17-ol-3-one, Bol) has been the subject of a heated debate because of ongoing confusion about its endogenous or exogenous origin when detected in one of its forms in faecal or urine samples from cattle. An expert report was recently written on the presence and metabolism of Bol in various animal species. Androstadienedione (ADD) is a direct precursor of 17β-boldenone (βBol). It is a 3,17-dione; ßBol is a 17-ol-3-one. Not much is published on 1,4-androstadiene-3,17-diol, which is a 3,17-diol (ADL). If animals were exposed for a longer period to one of these analytes, a metabolic pathway would be initiated to eliminate these compounds. Similar to recent testosterone metabolism studies in the aquatic invertebrate Neomysis integer, ADD, ßBol and ADL could also be eliminated as hydroxymetabolites after exposure. The presence of 11-keto-steroids or 11-hydroxy-metabolites in faecal samples can interfere with a confirmation method by gas chromatography-negative chemical ionization mass spectrometry (GC-NCI-MS), after oxidation of corticosteroids with a double bond in the A-ring (e.g. prednisolone or its metabolite prednisone). The presence of androstadienetrione (ADT) in faecal samples of cattle has never been reported. The origin of its presence can be explained through different pathways, which are presented in this paper.

IntroductionThe zoonotic hepatitis E virus (HEV) genotype 3 (HEV-G3) has become a common cause of acute and chronic hepatitis among humans worldwide. In Israel, while HEV-3 sequences have previously been detected in sewage, only the non-zoonotic HEV-G1 genotype has been found in samples from human patients.AimIn this pilot study, we aimed to assess the status of HEV in a sample of the swine population and among swine farm workers in Israel.MethodsPig blood (n = 141) and faecal samples (n = 39), pig farm sewage samples (n = 8) and blood from farm workers (n = 24) were collected between February 2016 and October 2017. Anti-HEV IgG was detected using the Wantai assay. HEV RNA was analysed with the RealStar HEV kit. HEV open reading frame 1 fragments amplified from representative HEV RNA-positive samples were used for phylogenetic analysis.ResultsOverall prevalence of HEV antibodies in pigs was 75.9% (107/141). HEV RNA was detected in plasma (2.1%, 3/141), faecal (22.8%, 18/79) and pig sewage (4/8) samples. Pig and sewage-derived viral sequences clustered with previously identified human sewage HEV-G3 sequences. Most pig farms workers (23 of 24) were HEV-seropositive; none was viraemic or reported previous clinical signs.ConclusionsThis study showed that domestic pigs in Israel are infected with HEV-G3. The high HEV seropositivity of the farm workers together with the previous identification of this virus in human sewage suggests circulation to humans. The clinical impact of these findings on public health should be further explored.

At early stages of life, infant immature intestine is not fully developed, exposing the new-born to potential diseases. Compounds that can exert beneficial actions on the infant intestine are bioactive lactose-based oligosaccharides (LBOs). The natural source of LBOs is mother milk. When human milk is lacking, dietary supplementation with infant formula fortified with manufactured LBOs, such as galacto-oligosaccharides (GOS), is pursued. GOS have been shown to have several properties in common with HMOs. LBOs composition and intestinal fate is extensively described for humans, whereas they are hardly investigated for domestic animal. In this PhD thesis, composition of LBOs in equine and porcine colostrum were described and new structures were elucidated. The analysis were performed mainly using liquid chromatography and capillary electrophoresis techniques. High inter- and intra-individual variation were found for oligosaccharides present in equine and porcine milk. In vivo fermentation fate of porcine milk oligosaccharides (PMOs) was also described analysing PMOs as found in fecal samples of piglets. The results were correlated to existing literature on HMOs. Dietary oligosaccharides are partially present systemically, as suggested from HMO studies. GOS and PMOs in blood, urine and fecal samples from an in vivo feeding trial on piglet were described. Intact dietary oligosaccharides including GOS and milk oligosaccharides from the piglet diet were found in piglet blood and urine samples. All dietary oligosaccharides were fermented/absorbed in vivo, not being detectable in the piglet fecal samples. On the other hand, GOS in vitro fermentation by piglet inoculum delineate a unique fermentation profile regarding GOS size consumption compared to GOS in vitro fermentation by human fecal inoculum. Similar degradation profile regarding GOS linkage types was observed for GOS fermentation by piglet and human inocula.

The Metabolism Study of Oleraisoquinoline in Rats Using Ultrahigh- performance Liquid Chromatography-electrospray Coupled with Quadrupole Time-of-flight Mass Spectrometry and its Bioactivities

ObjectiveIn this study, metabolites that changed in the rumen fluid, urine and feces of dairy cows fed different feed ratios were investigated.MethodsEight Holstein cows were used in this study. Rumen fluid, urine, and feces were collected from the normal concentrate diet (NCD) (Italian ryegrass 80%: concentrate 20% in the total feed) and high concentrate diet (HCD) groups (20%: 80%) of dairy cows. Metabolite analysis was performed using proton nuclear magnetic resonance (NMR) identification, and statistical analysis was performed using Chenomx NMR software 8.4 and Metaboanalyst 4.0.ResultsThe two groups of rumen fluid and urine samples were separated, and samples from the same group were aggregated together. On the other hand, the feces samples were not separated and showed similar tendencies between the two groups. In total, 160, 177, and 188 metabolites were identified in the rumen fluid, urine, and feces, respectively. The differential metabolites with low and high concentrations were 15 and 49, 14 and 16, and 2 and 2 in the rumen fluid, urine, and feces samples, in the NCD group.ConclusionAs HCD is related to rumen microbial changes, research on different metabolites such as glucuronate, acetylsalicylate, histidine, and O-Acetylcarnitine, which are related to bacterial degradation and metabolism, will need to be carried out in future studies along with microbial analysis. In urine, the identified metabolites, such as gallate, syringate, and vanillate can provide insight into microbial, metabolic, and feed parameters that cause changes depending on the feed rate. Additionally, it is thought that they can be used as potential biomarkers for further research on subacute ruminal acidosis.

Patterns of Urinary and Fecal Steroid Excretion during the Ovarian Cycle and Pregnancy in the African Elephant (Loxodonta africana)

The previous studies have shown that tea polyphenols are metabolized by gut microbiota. This study investigated the effect of gut microbiota on the bioavailability, tissue levels, and degradation of tea polyphenols. Mice were treated with antibiotics (ampicillin/sulfamethoxazole/trimethoprim) in drinking water and the control mice received water for 11 days, and they were given an AIN93M diet enriched with 0.32% of Polyphenon E. The levels of catechins and their metabolites (if present) in the serum, liver, urine, and fecal samples were determined by high-performance liquid chromatography. The results showed that treatment with antibiotics significantly increased the levels of the major polyphenol, (-)-epigallocatechin-3-gallate (EGCG), in serum and liver samples. Antibiotics also raised the levels of some catechins in urine and fecal samples but decreased the levels of their metabolites. These results suggest that antibiotics eliminated gut microbes and increased the bioavailabilities of these tea catechins. In a second study, mice were given different concentrations of green tea infusions as the drinking fluid. The plasma levels of EGCG and (-)-epicatechin-3-gallate (ECG) at day 112 were significantly lower than those at day 5. The urine levels of EGCG and ECG increased in the first 4 or 5 days, and then decreased to much lower levels at day 23 and beyond. In contrast, the levels of (-)-epigallocatechin and (-)-epicatechin showed a trend of increase during the 112-day experiment, likely owing to microbial hydrolysis of EGCG and ECG. Both sets of experiments support the idea that the degradation of EGCG and ECG by gut microbiota decreases their bioavailabilities. © 2018 BioFactors, 2018.

Changes of Human Gut Microbiome Correlated with Metabolomics After Cranberry Juice Consumption in a Double-Blinded, Placebo Controlled, Crossover Study

Preclinical evidence suggests that short chain fatty acids (SCFAs) produced by gut microbiota may impact body composition and muscle growth. While aging is implicated in negative alterations to the gut microbiome, exercise may mitigate these changes. Limited human evidence indicates that resistance training (RT) does not appreciably alter the gut microbiome in older adults, and no human study has examined whether resistance training differentially alters the gut microbiome and associated SCFAs between younger and older individuals. Therefore, we examined whether 10 weeks of RT differentially altered fecal microbiota composition, fecal and circulating SCFAs, and serum markers associated with gastrointestinal integrity in two cohorts of adults. Fecal and serum samples were obtained from untrained younger (22 ± 2 years, n = 12) and older (58 ± 8 years, n = 12) participants prior to and following 10 weeks of supervised twice-weekly full-body RT. Outcome measures immediately before (PRE) and after the intervention (POST) included dual X-ray absorptiometry for body composition, ultrasound for vastus lateralis (VL) thickness, 16S rRNA gene sequencing fecal microbiome data, serum and fecal SCFAs measured by gas chromatography, and serum intestinal fatty acid-binding protein 2 (FABP2), lipopolysaccharide-binding protein (LBP), and leucine-rich alpha-2 glycoprotein (LRG-1) quantified by enzyme-linked immunosorbent assays. Main effects and interactions were measured by repeated measures analysis of variance (group × time; G × T) for all dependent variables, and Spearman correlations were used to explore relationships among changes in relevant outcomes. The intervention significantly increased VL thickness and lean body mass (p < 0.05) equally in both groups. Although group differences in microbiome beta diversity were identified, no effects of age, time, or their interaction were observed for the alpha diversity measures. Seven SCFAs were detected in the fecal samples, albeit no significant age, time, or interaction effects were evident. In serum, acetic acid was the only SCFA detected, with no significant age, time, or interaction effects. Serum LRG1 decreased for all participants (p = 0.007) with higher levels in younger adults (p = 0.015), but no G × T interactions were observed for this marker, serum FABP2, or LBP. No significant correlations were observed among RT-induced changes in muscle mass-related outcomes and changes in fecal microbiome diversity, total or individual SCFAs, or serum FABP2/LBP/LRG-1. These results highlight that 10 weeks of RT largely does not affect fecal microbiota, associated SCFAs, or select markers of gastrointestinal integrity in untrained younger or older adults.

The Arctic is an accumulation zone of long-range transported pollution. In addition, local anthropogenic activities further contribute to regional pollution levels. The Svalbard reindeer (Rangifer tarandus platyrhynchus) is a suitable organism for studying and monitoring exposure to anthropogenic pollutants at the base of the terrestrial Arctic food web, and reindeer faeces have been promoted as non-invasive means of biomonitoring contaminant exposure. This study used HR-ICP-MS to analyse levels and composition of 16 elements in Svalbard reindeer faeces (n = 96) and soil (n = 9) from two locations on Svalbard, with the aim to assess whether local anthropogenic pollution influences element bioavailability. One of the sampling areas, the Nordenskiöld coast, is situated on the west coast of Spitsbergen, close to the Arctic Ocean and relatively far from local anthropogenic sources. The other sampling area, Adventdalen, is located further inland and close to Longyearbyen, the largest settlement of the archipelago. There was a significant difference in faecal elemental concentration and composition between the Adventdalen and Nordenskiöld coast reindeer populations. Elements of geogenic origin (e.g., Al, Cu and Fe) were found at higher levels in faeces from Adventdalen. In comparison, levels of Ca, Se and the toxic elements Cd and Pb were higher in faecal samples from the Nordenskiöld coast. The significantly higher levels of faecal Cd and Pb at Nordenskiöld coast may be due to marine input, dietary differences between the populations, or possible anthropogenic influence from the nearby settlement of Barentsburg. There was, however, a decoupling in elemental composition between faecal and soil samples, which may derive from a selective vegetational uptake of elements from the soil. The results suggest that reindeer are exposed to a range of elements and that faeces can be used to monitor the exposure to bioavailable environmental levels of both essential and toxic elements in terrestrial ecosystems.

Faecal microbiota transplantation (FMT) may contribute towards disease remission in ulcerative colitis (UC), but it is unknown which factors determine long-term effect of treatment. Here, we aimed to identify bacterial signatures associated with sustained remission. To this end, samples from healthy donors and UC patients-grouped into responders and non-responders at a primary end point (week 12) and further stratified by sustained clinical remission and relapse assessed at ⩾1-year follow-up were analysed, comparing the efficacy of FMT from either a healthy donor or autologous faeces. Microbiota composition was determined with a 16S rRNA gene-based phylogenetic microarray on faecal and mucosal samples, and functional profiles were predicted using PICRUSt with quantitative PCR verification of the butyrate production capacity; short-chain fatty acids were measured in faecal samples. At baseline, UC patients showed reduced amounts of bacterial groups from the Clostridium cluster XIVa, and significantly higher levels of Bacteroidetes as compared with donors. These differences were reduced after FMT mostly in responders. Sustained remission was associated with known butyrate producers and overall increased butyrate production capacity, while relapse was associated with Proteobacteria and Bacteroidetes. Ruminococcus gnavus was found at high levels in donors of failed FMT. A microbial ecosystem rich in Bacteroidetes and Proteobacteria and low in Clostridium clusters IV and XIVa observed in UC patients after FMT was predictive of poor sustained response, unless modified with a donor microbiota rich in specific members from the Clostridium clusters IV and XIVa. Additionally, sustained response was associated with restoration of the butyrate production capacity.

To prospectively examine the changes in microbiota within the urinary tract after transrectal prostate biopsy. Data, urine, and fecal samples prospectively collected from 30 patients before and after transrectal biopsy of the prostate. DNA was extracted from urine collected after a prostate massage before and after prostate biopsy, and from fecal samples collected before the biopsy. We sequenced DNA using the bacterial 16S rRNA high-throughput next-generation sequencing and analyzed changes in microbial profiles for taxonomy comparison between samples. Pre-biopsy urinary microbial profiles contained Lactobacillus and Staphylococcus bacteria. Post-biopsy urinary microbial profiles included lower levels of Lactobacillus and higher levels of Prevotella bacteria. Bacteroides bacteria were predominant in fecal samples. We identified two clustering patterns containing both pre- and post-biopsy urine samples. Cluster 1 had a urine cluster pattern that was distinct from fecal, whereas cluster 2 was similar to fecal. We observed two different modes of microbial changes, 11 patients had both of their urine (pre and post) samples associated with a particular cluster group, whereas others (n = 15) had movement between clusters 1 and 2 following the biopsy procedure. Four patient's post-biopsy urine microbial profiles clustered very tightly to the fecal microbial profile. We describe two models of change in the urinary tract microbiota after prostate biopsy using 16S RNA gene analysis. Further research to determine what controls changes in the urinary microbiota after prostate biopsy can help us understand why some patients are more susceptible to develop post-biopsy infections.