Biodiversity and antifouling activity of fungi associated with two soft corals from the South China Sea.

Bacteria in corals have been studied in detail in the past decades. However, the biodiversity and bioactivity of fungi in corals are still poorly understood. This study investigated the biodiversity and antifouling activity of fungi in soft corals Cladiella krempfi and Sarcophyton tortuosum from the South China Sea. A high diverse and abundant fungal community was found in the two soft corals. Furthermore, five isolates shared 83-95% similarity with their closest relatives, indicating that they might be novel species in genera Phaeoshaeria and Mucor. In addition, approximately 50% of the representative isolates exhibited distinct antifouling activity. In particular, isolates Fungal sp. SCAU132 and Fungal sp. SCAU133 displayed very strong antifouling activity against Bugula neritina, suggesting they can provide a potential resource for further investigation on isolation of novel antifouling metabolites. To our knowledge, this study is the first report to investigate the biodiversity and antifouling activity of fungi in C. krempfi and S. tortuosum.

Tracking the relative concentration between Bacteroidales DNA markers and culturable Escherichia coli in fecally polluted subtropical seawater: potential use in differentiating fresh and aged pollution.

Routine water quality monitoring practices based on the enumeration of culturable Escherichia coli provides no information about the source or age of fecal pollution. An emerging strategy is to use culturable E. coli and the DNA markers of Bacteroidales complementarily for microbial source tracking. In this study, we consistently observed in seawater microcosms of 3 different conditions that culturable E. coli decayed faster (T99 = 1.14 - 4.29 days) than Bacteroidales DNA markers did (T99 = 1.81 - 200.23 days). Concomitantly, the relative concentration between Bacteroidales DNA markers and culturable E. coli increased over time in all treatments. Particularly, the increase during the early stage of the experiments (before T99 of E. coli was reached) was faster than during the later stage (after T99 of E. coli was attained). We propose that the tracking of the relative concentration between Bacteroidales DNA markers and culturable E. coli provides an opportunity to differentiate a pollution that is relatively fresh from one that has aged. This method, upon further investigation and validation, could be useful in episodic pollution events where the surge of E. coli concentration causes noncompliance to the single sample maximum criterion that mandates high frequency follow-up monitoring.

Sperm proteome of Mytilus galloprovincialis: Insights into the evolution of fertilization proteins in marine mussels.

Cataloging the sperm proteome of an animal can improve our understanding of its sperm-egg interaction and speciation, but such data are available for only a few free-spawning invertebrates. This study aimed to identify the sperm proteome of Mytilus galloprovincialis, a free-spawning marine mussel. We integrated public transcriptome datasets by de novo assembly, and applied SDS-PAGE coupled LC-MS/MS analysis to profile the sperm proteome, resulting in the identification of 550 proteins. Comparing the homologous sperm protein coding genes between M. galloprovincialis and its closely related species M. edulis revealed that fertilization proteins have the highest mean nonsynonymous substitution rate (Ka/Ks = 0.62) among 11 functional groups, consistent with previous reports of positive selection of several fertilization proteins in Mytilus. Moreover, 78 sperm proteins in different functional groups have Ka/Ks values > 0.5, indicating the presence of many candidate sperm proteins for further analysis of rapid interspecific divergence. The MS data are available in ProteomeXchange with the identifier PXD001665.

Differential utility of the Bacteroidales DNA and RNA markers in the tiered approach for microbial source tracking in subtropical seawater.

Source tracking of fecal pollution is an emerging component in water quality monitoring. It may be implemented in a tiered approach involving Escherichia coli and/or Enterococcus spp. as the standard fecal indicator bacteria (FIB) and the 16S rRNA gene markers of Bacteroidales as source identifiers. The relative population dynamics of the source identifiers and the FIB may strongly influence the implementation of such approach. Currently, the relative performance of DNA and RNA as detection targets of Bacteroidales markers in the tiered approach is not known. We compared the decay of the DNA and RNA of the total (AllBac) and ruminant specific (CF128) Bacteroidales markers with those of the FIB in seawater spiked with cattle feces. Four treatments of light and oxygen availability simulating the subtropical seawater of Hong Kong were tested. All Bacteroidales markers decayed significantly slower than the FIB in all treatments. Nonetheless, the concentrations of the DNA and RNA markers and E. coli correlated significantly in normoxic seawater independent of light availability, and in hypoxic seawater only under light. In hypoxic seawater without light, the concentrations of RNA but not DNA markers correlated with that of E. coli. Generally, the correlations between Enterococcus spp. and Bacteroidales were insignificant. These results suggest that either DNA or RNA markers may complement E. coli in the tiered approach for normoxic or hypoxic seawater under light. When light is absent, either DNA or RNA markers may serve for normoxic seawater, but only the RNA markers are suitable for hypoxic seawater.

Effects of chlorination on the survival of sewage bacteria in seawater microcosms.

Chlorination is a commonly used disinfection method in sewage treatment process. However, resistant bacteria may survive chlorination and enter the receiving aquatic environment upon effluent discharge. There has been limited research on the effects of chlorination on bacterial survival in seawater. To address this knowledge gap, microcosm experiments were conducted to simulate the discharge of chlorinated effluents into coastal seawater. The results revealed that bacterial communities in seawater-based effluents survived better in seawater than those in freshwater-based effluents. High chlorine dosages could significantly reduce the viable bacterial populations and their chance of regrowth in seawater. Additionally, faecal indicator bacteria (FIB) that entered the viable but non-culturable (VBNC) state under chlorination tended to persist in the VBNC state without resuscitation during seawater incubation. Because of the prevalence of VBNC indicator bacteria, qPCR quantification of FIB was more effective than conventional culture-based methods in tracing viable pathogenic chlorine-resistant bacteria, although the correlation strength varied depending on the type of effluent. This study sheds light on how chlorine dosages and the intrinsic properties of effluents affect bacterial survival in seawater and highlights the potential and limitations of using FIB in monitoring the health risks associated with the discharge of chlorinated effluents.

Validation of an Integrated Genetic-Epigenetic Test for the Assessment of Coronary Heart Disease.

BACKGROUND: Coronary heart disease (CHD) is the leading cause of death in the world. Unfortunately, many of the key diagnostic tools for CHD are insensitive, invasive, and costly; require significant specialized infrastructure investments; and do not provide information to guide postdiagnosis therapy. In prior work using data from the Framingham Heart Study, we provided in silico evidence that integrated genetic-epigenetic tools may provide a new avenue for assessing CHD. METHODS AND RESULTS: In this communication, we use an improved machine learning approach and data from 2 additional cohorts, totaling 449 cases and 2067 controls, to develop a better model for ascertaining symptomatic CHD. Using the DNA from the 2 new cohorts, we translate and validate the in silico findings into an artificial intelligence-guided, clinically implementable method that uses input from 6 methylation-sensitive digital polymerase chain reaction and 10 genotyping assays. Using this method, the overall average area under the curve, sensitivity, and specificity in the 3 test cohorts is 82%, 79%, and 76%, respectively. Analysis of targeted cytosine-phospho-guanine loci shows that they map to key risk pathways involved in atherosclerosis that suggest specific therapeutic approaches. CONCLUSIONS: We conclude that this scalable integrated genetic-epigenetic approach is useful for the diagnosis of symptomatic CHD, performs favorably as compared with many existing methods, and may provide personalized insight to CHD therapy. Furthermore, given the dynamic nature of DNA methylation and the ease of methylation-sensitive digital polymerase chain reaction methodologies, these findings may pave a pathway for precision epigenetic approaches for monitoring CHD treatment response.

Improving the performance of an end-point PCR assay commonly used for the detection of Bacteroidales pertaining to cow feces.

Bacteroidales are normal gut flora of warm-blooded animals. Since each host species carries a different diversity of Bacteroidales, the detection of host-associated gene markers of Bacteroidales has emerged as a promising tool for the tracking of the source of fecal pollution in aquatic ecosystems. To detect cow-associated Bacteroidales, a commonly used method has been an end-point PCR assay with the 16S rRNA genes primers CF128F (cow-associated) and Bac708R (all Bacteroidales). The PCR assay has demonstrated high rates of true-positive detection (i.e., high sensitivity) in all previous studies. However, the assay also had high rates of false-positive detection to the samples of non-target hosts in some cases (i.e., low specificity). In opposite to the reason many investigators have proposed, our results suggested that false detection was not necessarily due to the presence of the target sequence of CF128F in the feces of non-target hosts. Instead, we found sequences of non-target hosts having single internal mismatches with CF128F. Those mismatches were well tolerated in PCR, partly due to the universality of Bac708R. To improve the detection performance, we designed a novel primer CF592R (targeting the same clade of sequences as CF128F) to substitute Bac708R. The use of CF529R alleviated false detection and also led to a tenfold reduction in detection limit in the samples tested, compared to the use of Bac708R. Many other end-point PCR assays that detect the 16S rRNA genes in Bacteroidales also use a host-associated primer to couple with Bac708R, and low specificity or sensitivity has been reported. Based on our findings for CF128F, we suggest that the suitability of Bac708R in those PCR assays needs to be revisited.

Strategy to Evaluate Changes in Bacterial Community Profiles and Bacterial Pathogen Load Reduction After Sewage Disinfection.

Sewage effluent discharge is a major source of pathogenic contamination to the environment. The disinfection process is critical for the elimination of pathogens in sewage. In this study, we examined the impact of chlorine disinfection on the total, viable, and culturable populations of indicator bacteria, pathogens, and bacterial communities in two contrasting types of effluents (primarily treated saline and secondarily treated freshwater). Effluents collected bimonthly over 1 year were examined using cultivation, quantitative PCR (qPCR), and 16S rRNA gene amplicon sequencing coupled with or without propidium monoazide (PMA) treatment. The results showed that each type of effluent was characterized by a specific set of representative genera before disinfection. Salinity appeared to be the major abiotic factor associated with the differences in bacterial community compositions. The pathogen analysis pipeline revealed over 20 viable clinically important pathogenic species in the effluents. Although the bacterial communities differed markedly between the two types of effluents before disinfection, the species of pathogens persisting after disinfection were similar, many of them were members of Enterobacter and Vibrio. The relative abundances of all pathogens identified in the amplicon sequences were multiplied by the 16S rRNA gene copy numbers of total bacteria detected by PMA-qPCR to estimate their concentrations. Pathogens remained viable after disinfection reached 8 log10 16S rRNA copies ml-1 effluent. Meanwhile, around 80 % of the populations of three indicator bacteria including Escherichia coli, Enterococcus, and Bacteroidales were viable after disinfection, but over 99 % of the viable E. coli and Enterococcus were in the non-culturable state. We estimated the total pathogen load by adding the concentrations of all viable pathogens and examined their correlations with indicator bacteria of different types, physiological states, and effluents. The results showed that the PMA-qPCR measurement of E. coli is a reliable proxy of bacterial pathogen loads in both types of effluents. The utility of viable indicator bacteria as a biological index to assess the overall bacteriological hazards in effluents is discussed.

Genetic and Ecological Diversity of Escherichia coli and Cryptic Escherichia Clades in Subtropical Aquatic Environments.

Escherichia coli not only inhabit the large intestines of human and warm-blooded animals but could also persist in the external environment. However, current knowledge was largely based on host-associated strains. Moreover, cryptic Escherichia clades that were often misidentified as E. coli by conventional diagnostic methods were discovered. Failure to distinguish them from E. coli sensu stricto could lead to inaccurate conclusions about the population genetics of E. coli. Based on seven housekeeping genes, we determine the genetic and ecological diversity of E. coli and cryptic clades as they occupy aquatic habitats with different characteristics and human impact levels in subtropical Hong Kong. Contrary to previous reports, clade II was the most abundant cryptic lineage co-isolated with E. coli, being especially abundant in relatively pristine subtropical aquatic environments. The phylogenetically distinct cryptic clades and E. coli showed limited recombination and significant genetic divergence. Analyses indicated that these clade II strains were ecologically differentiated from typical E. coli; some may even represent novel environmental Escherichia clades that were closely related to the original clade II strains of fecal origins. E. coli of diverse origins exhibited clonality amidst divergent genotypes STs, echoing other studies in that recombination in housekeeping genes was insufficient to disrupt phylogenetic signals of the largely clonal E. coli. Notably, environmental E. coli were less diverse than fecal isolates despite contributing many new alleles and STs. Finally, we demonstrated that human activities influenced the distribution of E. coli and clade II in a small aquatic continuum. Moving from relatively pristine sites toward areas with higher human disturbance, the abundance of clade II isolates and new E. coli genotypes reduces, while E. coli bearing class I integrons and belonging to CCs of public health concern accumulates. Altogether, this work revealed the new genetic diversity of E. coli and cryptic clades embedded in selected subtropical aquatic habitats, especially relatively pristine sites, which will aid a more thorough understanding of the extent of their genetic and functional variations in relation to diverse habitats with varied conditions.

Air insufflation of the stomach following laparoscopic pyloromyotomy may not detect perforation.

BACKGROUND: Undetected perforation during laparoscopic pyloromyotomy can be fatal. Detecting a perforation at the time of laparoscopic pyloromyotomy is difficult. The purpose of this study was to determine whether air insufflation of the stomach reliably detects perforation during laparoscopic pyloromyotomy. CASE REPORTS: Between 2007 and 2008, 71 patients underwent laparoscopic pyloromyotomy and 2 patients (3.3%) had perforation. Insufflating the stomach with air did not demonstrate the perforation in either case. Both perforations were detected by careful inspection of the myotomy; a small amount of mucus was seen at the perforation site. Both patients underwent open suture repair with an omental patch and had unremarkable postoperative courses. CONCLUSIONS: Air insufflation of the stomach during laparoscopic pyloromyotomy does not reliably rule out perforation. As with all procedures with potential complications, a high index of suspicion and careful inspection of the entire myotomy may help detect perforation.

How Does Resident Participation Alter the Outcome of Surgery for Pectus Excavatum?

BACKGROUND: General surgery resident participation in the operating room is critical in training the next generation of surgeons. As of yet, the impact of resident participation on outcomes of surgery for pectus excavatum and many complex subspecialty operations has not been well studied. METHODS: A multi-institutional retrospective study of patients undergoing operative repair for pectus excavatum was performed. All relevant data were analyzed (IRB 11144). RESULTS: Two hundred and fourteen patients underwent operative correction (195 Nuss, 19 Ravitch). There were 185 males. Average age at repair was 14.7 years with a Haller index of 4.5. Average surgery time was 144 minutes (57-255) for the Nuss procedure and 263 minutes (141-373) for the Ravitch procedure. The presence of a second pediatric surgeon reduced the surgery time from 170 to 135 minutes (p < 0.01) and the presence of residents increased the time from 129 to 155 minutes (p < 0.01) for the Nuss procedure. One hundred and fifty patients had a single bar and 57 patients had 2 bars (28%). Average length of stay was 4.96 days (3-11). Long-term follow-up averaged 1737 days (42-3894). There were few complications and no difference in complication rate or length of stay between groups. Ninety nine percent of patients deemed the repair excellent and no patients required revision. CONCLUSIONS: Resident participation increases operative time, but with no demonstrable effect on hospital stay or long-term outcomes. Complication rates are low regardless of operating team composition. Thus, continuing to allow resident involvement, especially in subspecialty operations such as the Nuss and Ravitch procedures, may be worthwhile for resident education and surgical experience.

Genetic Variation and Preliminary Indications of Divergent Niche Adaptation in Cryptic Clade II of Escherichia.

The evolution, habitat, and lifestyle of the cryptic clade II of Escherichia, which were first recovered at low frequency from non-human hosts and later from external environments, were poorly understood. Here, the genomes of selected strains were analyzed for preliminary indications of ecological differentiation within their population. We adopted the delta bitscore metrics to detect functional divergence of their orthologous genes and trained a random forest classifier to differentiate the genomes according to habitats (gastrointestinal vs external environment). Model was built with inclusion of other Escherichia genomes previously demonstrated to have exhibited genomic traits of adaptation to one of the habitats. Overall, gene degradation was more prominent in the gastrointestinal strains. The trained model correctly classified the genomes, identifying a set of predictor genes that were informative of habitat association. Functional divergence in many of these genes were reflective of ecological divergence. Accuracy of the trained model was confirmed by its correct prediction of the habitats of an independent set of strains with known habitat association. In summary, the cryptic clade II of Escherichia displayed genomic signatures that are consistent with divergent adaptation to gastrointestinal and external environments.

Correction: Regev, Y., et al. Molecular Identification and Characterization of Vibrio Species and Mycobacterium Species in Wild and Cultured Marine Fish from the Eastern Mediterranean Sea. Microorganisms 2020, 8, 863.

We would like to change the authors' affiliation of paper [...].

Molecular Identification and Characterization of Vibrio Species and Mycobacterium Species in Wild and Cultured Marine Fish from the Eastern Mediterranean Sea.

In contrast to numerous documented pathogens and infectious diseases of aquaculture, there is a lack of baseline data and information regarding pathogenic agents' prevalence in wild marine fish populations. This study focused on two common fish pathogenic microorganisms, namely Mycobacterium species and Vibrio species, both of which are known to be major causes of fish loss, occasionally to the extent of being a limiting factor in fish production. Both microorganisms are known as zoonotic agents. In total, 210 wild marine indigenous and Lessepsian fish from four different species from the eastern Mediterranean Sea were sampled and tested for Vibrio species and Mycobacterium species during a two-year period (2016-2017). Using PCR with 16S rRNA primers, we detected different strain variations of Mycobacterium species and Vibrio species and, based on the sequencing results, the overall prevalence for Vibrio species in wild fish in 2016 was significantly higher compared to 2017. No significant difference was detected for Mycobacterium species prevalence in wild fish between 2016 and 2017. In addition, 72 gilthead seabream (Sparus aurata) from an Israeli offshore marine farm were also examined during the two-year period (2017-2018). The results suggest that Mycobacterium species prevalence was significantly higher in 2018, while in 2017 there was no positive results for Mycobacterium species. In addition, there was no significant difference between both years in regard to the prevalence of Vibrio species for maricultured fish. These results highlight the necessity of continuous molecular monitoring in order to evaluate the prevalence of pathogenic microorganisms in both wild and cultured fish populations.

Initial experience with laparoscopic Chait Trapdoor cecostomy catheter placement for the management of fecal incontinence in children: outcomes and lessons learned.

BACKGROUND: Antegrade enemas administered through a percutaneously placed Chait Trapdoor cecostomy catheter have resulted in a marked improvement in compliance and outcome of patients with fecal incontinence. The percutaneous technique, however, is a two-step procedure that is not performed under direct vision. This report presents the results and lessons learned from our experience with the laparoscopic approach to placement of Chait cecostomy catheters. METHODS: Retrospective review of patients who underwent laparoscopic placement of Chait cecostomy catheters from 1999 to 2008. Data collected included patient demographics, primary diagnosis, hospital stay, complications, follow-up duration and outcome. RESULTS: Seventeen patients, mean age 11.8 + or - 4.2 years (range 5-17), underwent laparoscopic Chait cecostomy catheter placement over a period of 8 years. Median follow-up was 46 + or - 21 months (range 4-67). The primary diagnosis was spina bifida in 82% of patients. There was one intraoperative complication, which consisted of tangential needle placement into the cecum, and required conversion to an open procedure. Mean hospital stay was 3.8 + or - 1.5 days (range 2-7). Emergency department visits related to Chait catheter complications were mainly due to catheter dislodgement and breakage. Long-term complications included accidental dislodgement of the catheter in seven patients (41%), mechanical failure of the catheter (breaks/leaks) in six patients (35%), hypertrophic granulation tissue in six patients (35%), wound infections at the catheter site in three patients (18%), complications related to the use of fasteners in two patients (12%) and ventirculoperitoneal (VP) shunt infection in two patients (11.8%). CONCLUSION: The laparoscopic approach to Chait cecostomy catheter placement is a simple and effective procedure. The rate of long term complications such as catheter dislodgement and mechanical failure, which are responsible for the majority of unplanned ED visits, may be decreased by routine yearly catheter exchanges. VP shunt infections are the most serious complications in this patient population consisting mostly of patients with spina bifida.

Draft Genome Sequences of 16 Strains of Escherichia Cryptic Clade II Isolated from Intertidal Sediment in Hong Kong.

The genus Escherichia includes several cryptic clades. Among them, the members of cryptic clade II have rarely been found, and their genome sequences remain largely uninvestigated. Here, we report the draft genome sequences of 16 strains of Escherichia cryptic clade II that were isolated from intertidal sediment in Hong Kong.

Minimally Invasive Repair of Pediatric Morgagni Hernias Using Transfascial Sutures with Extracorporeal Knot Tying.

BACKGROUND: Morgagni hernias are rare, with a reported incidence of 2% to 5% of congenital diaphragmatic hernias. OBJECTIVES: To review a laparoscopic technique to repair Morgagni hernias in pediatric patients. METHODS: Retrospective chart review of pediatric patients who underwent minimally invasive repair of a Morgagni hernia from November 2009 to September 2017 within a defined population. RESULTS: During an 8-year period, 15 patients with Morgagni hernias were identified. Four patients with Morgagni hernias were excluded because they had open repairs. Eleven Morgagni hernias were repaired through a completely minimally invasive approach. Three repairs were completed using a soft-tissue patch (Gore-Tex patch, W L Gore & Associates Inc, Flagstaff, AZ). All minimally invasive repairs were completed with transfascial sutures using an endoscopic suturing device (Endo Close, Covidien/Medtronic, Fridley, MN) and 2-0 nonabsorbable synthetic sutures with extracorporeal knot tying. Median follow-up was 40 months (range = 2.6 months to 7.3 years). No patients had postoperative pectus excavatum defects. There were no recurrences. CONCLUSION: Morgagni hernias are amenable to minimally invasive repair with this simple technique. With large defects, synthetic patches should be used. Recurrences are rare, and morbidity is low.

A new technique in complex chest wall reconstruction: Open reduction and internal fixation.

BACKGROUND: Reconstruction of complex chest wall deformities is a surgical challenge. A new technique can improve long-term outcomes and result in high patient satisfaction. METHODS: A multicenter study was conducted on pediatric patients undergoing complex chest wall reconstruction between September 2015 and January 2018. The evolution of the technique using open reduction and internal fixation (ORIF) with SternaLock® and RibFix® to repair chest wall deformities is described. RESULTS: Seventeen patients underwent complex chest wall reconstruction with ORIF. Eight patients had severe or recurrent pectus excavatum, five patients had pectus carinatum, and four patients had complex chest wall fractures or other anomalies causing significant chronic pain. Up to three SternaLock® plates and four RibFix® plates were used for each procedure. Median length of hospital stay after surgery was four days. Median follow-up time was 12 months (range 2-30). There were no postsurgical complications. There was 100% patient satisfaction in postoperative recovery and cosmesis. CONCLUSION: ORIF using SternaLock® and RibFix® is an effective method of reconstructing complex chest wall deformities. This technique improves physical stability without the requirement of a subsequent surgery and enhances overall patient satisfaction. High volume centers should integrate this novel approach for challenging chest wall reconstruction. TYPE OF STUDY: Treatment study: case series. LEVEL OF EVIDENCE: Level IV.

The putative functions of lysogeny in mediating the survivorship of Escherichia coli in seawater and marine sediment.

Escherichia coli colonizes various body parts of animal hosts as a commensal and a pathogen. It can also persist in the external environment in the absence of fecal pollution. It remains unclear how this species has evolved to adapt to such contrasting habitats. Lysogeny plays pivotal roles in the diversification of the phenotypic and ecologic characters of E. coli as a symbiont. We hypothesized that lysogeny could also confer fitness to survival in the external environment. To test this hypothesis, we used the induced phages of an E. coli strain originating from marine sediment to infect a fecal E. coli strain to obtain an isogenic lysogen of the latter. The three strains were tested for survivorship in microcosms of seawater, marine sediment and sediment interstitial water as well as swimming motility, glycogen accumulation, biofilm formation, substrate utilization and stress resistance. The results indicate that lysogenic infection led to tractable changes in many of the ecophysiological attributes tested. Particularly, the lysogen had better survivorship in the microcosms and had a substrate utilization profile resembling the sediment strain more than the wild type fecal strain. Our findings provide new insights into the understanding of how E. coli survives in the natural environment.

Recent advances in molecular techniques for the detection of phylogenetic markers and functional genes in microbial communities.

The detection and analysis of nucleic acids extracted from microbial communities are the ultimate ways to determine the diversity and functional capability of microbial communities in the environments. However, it remains a challenge to use molecular techniques for unequivocal determination and quantification of microbial species composition and functional activities. Considerable efforts have been made to enhance the capability of molecular techniques. Here an update of the recent developments in molecular techniques for environmental microbiology is provided.