Aquatic microbial community is partially functionally redundant: Insights from an in situ reciprocal transplant experiment.

Microbial communities are considered to be functionally redundant, but few studies have tested this hypothesis empirically. In this study, we performed an in situ reciprocal transplant experiment on the surface and bottom waters of two lakes (Tsuei-Feng (T) and Yuan-Yang (Y)) with disparate trophic states and tracked changes in their microbial community composition and functions for 6 weeks using high-throughput sequencing and functional approaches. T lake's surface (Ts) and bottom (Tb) water active bacterial community (16S rRNA gene-transcript) was dominated by Actinobacteria, Bacteroidia, and Cyanobacteria, whereas Y lake's surface (Ys) and bottom (Yb) water had Gammaproteobacteria, Alphaproteobacteria, and Bacteroidia as the dominant classes. The community composition was resistant to changes in environmental conditions following the reciprocal transplant, but their functions tended to become similar to the incubating lakes' functional profiles. A significant linear positive relationship was observed between the microbial community and functional attributes (surface: R2 = 0.5065, p < 0.0001; bottom: R2 = 0.4592, p < 0.0001), though with varying scales of similarity (1-Bray Curtis distance), suggesting partial functional redundancy. Also, the entropy-based L-divergence measure identified high divergence in community composition (surface: 1.21 ± 0.54; bottom: 1.17 ± 0.51), and relatively low divergence in functional attributes (surface: 0.04 ± 0.01; bottom: 0.04 ± 0.01) in the two lakes' surface and bottom waters, providing further support for the presence of partial functional redundancy. This study enriches our understanding of community functional relationships and establishes the presence of partial functional redundancy in freshwater ecosystems.

Seasonal Distribution and Genotyping of Antibiotic Resistant Strains of ListeriaInnocua Isolated from A River Basin Categorized by ERIC-PCR.

Listeria innocua retains many conserved homologous domains with Listeria monocytogenes, which is a food-borne and water-borne diarrhea-causing bacterium. Studies of antimicrobial resistance in L. innocua showed that this microbe is more prone to acquire resistance than other bacteria in the genus Listeria. However, little is known about the seasonal population distribution and antimicrobial resistance patterns of L. innocua in natural water environments. The aims of the study were: (1) to investigate the occurrence of L. innocua isolates in a subtropical watershed and reconstruct the population structure and (2) to analyze the antibacterial resistance patterns of the identified L. innocua isolates according to ERIC type. A total of 288 water samples was collected from the Puzi River basin (23°28' N, 120°13' E) between March 2014 and March 2015, and 36 L. innocua isolates were recovered from 15 positive water samples. With regard to seasonal variation, L. innocua was only detected in the spring and summer. Eighteen enterobacterial repetitive intergenic consensus (ERIC)-PCR types were identified, and two genogroups with four subgroups were reconstructed in a minimum spanning tree. Isolates from different sampling areas that were located near each other were genetically different. All L. innocua isolates (including 41.7% of the multidrug-resistant (MDR) isolates) were resistant to oxacillin and showed high minimum inhibitory concentrations of tetracycline. These findings demonstrate the seasonal variations and differing geographical distributions of L. innocua in this subtropical water environment, as well as the existence of strong population structures and MDR and antimicrobial resistance patterns. Phylogenetic analysis based on ERIC-type showed that the Cluster A isolates were resistant to more antibiotics, and two types, ERIC8 and ERIC15 were multidrug resistant. The more commonly detected types, such as ERIC1 and ERIC12, were also more likely to be resistant to two or more antibiotics. Close monitoring of drug resistance in environmental L. innocua is warranted due to its potential for transferring antimicrobial resistance determinants to pathogenic Listeria.

Bacterial Community in Water and Air of Two Sub-Alpine Lakes in Taiwan.

Very few studies have attempted to profile the microbial communities in the air above freshwater bodies, such as lakes, even though freshwater sources are an important part of aquatic ecosystems and airborne bacteria are the most dispersible microorganisms on earth. In the present study, we investigated microbial communities in the waters of two high mountain sub-alpine montane lakes-located 21 km apart and with disparate trophic characteristics-and the air above them. Although bacteria in the lakes had locational differences, their community compositions remained constant over time. However, airborne bacterial communities were diverse and displayed spatial and temporal variance. Proteobacteria, Actinobacteria, Bacteroidetes, and Cyanobacteria were dominant in both lakes, with different relative abundances between lakes, and Parcubacteria (OD1) was dominant in air samples for all sampling times, except two. We also identified certain shared taxa between lake water and the air above it. The results obtained on these communities in the present study provide putative candidates to study how airborne communities shape lake water bacterial compositions and vice versa.

Molecular characteristics of clinical methicillin-resistant Staphylococcus pseudintermedius harboring arginine catabolic mobile element (ACME) from dogs and cats.

This study aimed to investigate the presence of arginine catabolic mobile element (ACME) and its associated molecular characteristics in methicillin-resistant Staphylococcus pseudintermedius (MRSP). Among the 72 S. pseudintermedius recovered from various infection sites of dogs and cats, 52 (72.2%) were MRSP. ACME-arcA was detected commonly (69.2%) in these MRSP isolates, and was more frequently detected in those from the skin than from other body sites (P=0.047). There was a wide genetic diversity among the ACME-arcA-positive MRSP isolates, which comprised three SCCmec types (II-III, III and V) and 15 dru types with two predominant clusters (9a and 11a). Most MRSP isolates were multidrug-resistant. Since S. pseudintermedius could serve as a reservoir of ACME, further research on this putative virulence factor is recommended.

Class 1 Integrons and the Antiseptic Resistance Gene (qacEΔ1) in Municipal and Swine Slaughterhouse Wastewater Treatment Plants and Wastewater-Associated Methicillin-Resistant Staphylococcus aureus.

Class 1 integrons are mobile gene elements (MGEs) containing qacEΔ1 that are resistant to quaternary ammonium compound (QAC) disinfectants. This study compared the abundances of class 1 integrons and antiseptic resistance genes in municipal (M) and swine slaughterhouse (S) wastewater treatment plants (WWTPs) and investigated the presence of class 1 integrons and antiseptic resistance genes in methicillin-resistant Staphylococcus aureus (MRSA) isolated from wastewater samples. The abundances of intI1 and qacEΔ1 genes in 96 wastewater samples were quantified using real-time quantitative polymerase chain reaction (real-time qPCR), and 113 MRSA isolates recovered from the wastewater samples were detected class 1 integrons and linked antiseptic resistance genes (qacEΔ1), and minimum inhibitory concentrations (MICs) for QAC antiseptics. The intI1 and qacEΔ1 genes were detected in all the wastewater samples, and they were more abundant in S-WWTP samples than in M-WWTP samples. A higher percentage of MRSA isolates carried qacEΔ1 in MRSA from swine wastewater samples (62.8%) than in municipal MRSA (3.7%). All the MRSA isolates showed high MICs for antiseptic agents. This study provides important evidence regarding the abundances of intI1 and qacEΔ1 genes in municipal and swine slaughterhouse wastewater, and antiseptic-resistant MRSA strains were detected in swine slaughterhouse wastewater.

Spreading of ß-lactam resistance gene (mecA) and methicillin-resistant Staphylococcus aureus through municipal and swine slaughterhouse wastewaters.

Methicillin-resistant Staphylococcus aureus (MRSA) is a potential zoonotic agent. Municipal wastewater treatment plants (WWTPs) can be reservoirs for MRSA dissemination. It is unclear, however, whether MRSA and its ß-lactam resistance gene (mecA) can be spread from WWTPs that treat the wastewater of swine auction markets. The aims of the study were to compare (1) the abundance of the mecA gene in one municipal (M-) and one swine (S-) WWTP and (2) the genotypic and phenotypic characteristics of MRSA isolates from these two types of WWTPs. The concentrations of mecA gene from 96 wastewater samples were quantified using real-time quantitative polymerase chain reaction (real-time qPCR). One hundred and thirteen MRSA isolates were recovered and were characterized by antimicrobial susceptibility testing, minimum inhibitory concentrations (MICs), and staphylococcal cassette chromosome mec (SCCmec) typing. The mecA gene could be detected in all the wastewater samples. A high abundance of recovered mecA gene (2.6 × 10(1) to 1.9 × 10(4) gene copies µg(-1) of total DNA) in swine slaughterhouse wastewater implied a correspondingly high transferring/receiving potential. All MRSA isolates were multidrug resistant (MDR) and showed high MICs to different antimicrobials. The M-WWTP MRSA isolates harbored SCCmec II-IV and VII, whereas those from the S-WWTP harbored SCCmec V and IX. In conclusion, wastewater from swine slaughterhouses can make these slaughterhouses potential hotspots for the dissemination of mecA gene and MRSA, and the high MICs of MRSA from both WWTP origins may pose a health risk not only to workers but also to the general public.

Population deviation of piggery-associated methicillin-resistant Staphylococcus aureus based on mec-associated direct repeat unit analysis.

Piggery-associated methicillin-resistant Staphylococcus aureus (MRSA) is a potential zoonotic pathogen. We constructed the population structure and dynamics of staphylococcal chromosome cassette mec (SCCmec) in MRSA ST9 isolates from different geographical areas of Taiwan. A total of 140 MRSA (135 piggery and 5 human clinical) isolates from three populations located in western Taiwan (n=96 including the 5 clinical isolates), central eastern Taiwan (n=22), and Penghu Island (n=22) were collected and characterized by analysis of the mec-associated direct repeat unit (dru). Twenty-eight dru types (with 24 novel) and 15 dru-Clonal Complexes (CCs) were identified. The predominant novel dt12w type (48.6%) was widespread in all populations and may have a superior ability to transmit among populations. The minimum spanning network showed that at least two ancestral dru types (dt11a and dt12w) were identified, and the genetics between different populations could be differentiated. Temporal distributions of clone population dynamics estimated through the Bayesian skyline plot indicated a stable population with a long evolutionary history for MRSA ST9 in Taiwan. Findings indicating that some dru types are shared between piggery-associated and human-clinical MRSA ST9 suggest the occurrence of cross-species horizontal transmission of SCCmec.

Characteristics and virulence factors of livestock associated ST9 methicillin-resistant Staphylococcus aureus with a novel recombinant staphylocoagulase type.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) sequence type 9 (ST9) is a potential source of zoonotic infection for humans. In this study, we investigated and compared the virulence profiles of MRSA ST9 isolates from healthy swine and human clinical origins. A total of 152 MRSA ST9 isolates, including 147 LA-MRSA isolates and 5 human clinical isolates, were studied for the accessory gene regulator (agr) and 20 enterotoxin genes (se), exfoliatoxin gene, and tst gene. The evolutionary history of staphylocoagulase (SC) in Taiwan MRSA ST9 was reconstructed based on phylogenetic and population genetics. The predominant type of LA-MRSA ST9 isolates (78.9%) was agr-II that differed from the predominant agr-I human clinical MRSA strains in Taiwan and the LA-MRSA ST398 lineage from Europe. Forty-nine percent of the LA-MRSA ST9 isolates carried a combination of enterotoxin gene cluster-2 (egc-2, seg, sei, sem, sen, seo, and seu) and tst. In addition, the Taiwan LA-MRSA ST9 and the human clinical ST9-MRSA contained a novel SC XIc subtype and had a unique history of evolution indicating a recent common ancestor. These findings suggest a cross-species transmission of this emerging ST9-SC XIc MRSA between swine and human.

Molecular characterization and clonal genetic diversity of methicillin-resistant Staphylococcus aureus of pig origin in Taiwan.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is an emerging public health threat. The aims of this study were to identify nasal carriage of MRSA in finishing pigs from different geographical areas of Taiwan and to compare the genetic features of LA-MRSA with other countries. A total of 299 swine nasal samples were collected from 11 counties in western Taiwan with 220 identified MRSA isolates characterized by spa, SCCmec, MLST, pulsed-field gel electrophoresis (PFGE), antibiogram, and Panton-Valentine leukocidin (PVL) toxin genes. A widespread LA-MRSA clone, ST9-t899-PVL-negative, was detected as the predominant (91.6%) clone in Taiwan pigs, and a novel spa type t7616 was identified. Most isolates carried SCCmec type V but lacked ccrC gene (98.8%), a particular feature that was observed for the first time. The integration of minimum spanning trees (MST) based on a PFGE distance matrix and geographical information showed high genetic diversity among ST9-t899 isolates with three unique subpopulations identified in two counties that are nearby each other. Multidrug resistance (MDR) was prevalent in isolates from different counties with more than 80% resistance to erythromycin, ciprofloxacin, gentamicin, tetracycline, and clindamycin. In conclusion, this study reported the first identified LA-MRSA clone of MDR-ST9-t899-V lacking ccrC dominant in pig farms in western Taiwan that differed from ST398 in Europe and North America. Close monitoring of this MDR unique clone is warranted to survey its spread from livestock to humans.

Prevalence of enteroviruses in hot spring recreation areas of Taiwan.

Enteroviruses can be introduced into the water environment as a result of human activity. Contaminations within hot tubs, spas and public baths are also possible. We investigated the distribution of enteroviruses at six hot spring recreation areas throughout Taiwan. Spring water was collected from 34 sites and enteroviruses were detected in 13 (38.2%). The most frequently detected was coxsackievirus A2, followed by echovirus 11. Enterovirus 71 (EV 71) and porcine enterovirus 9 were detected once. Water quality indicators were not statistically associated with the occurrence of enteroviruses, although the enterovirus-positive samples were positive for a greater number of microbiological indicators and showed a link to pH and water temperature. The results confirm the ubiquity of enteroviruses in Taiwan spring recreation areas. Coxsackievirus A2, echovirus 11 and EV 71, the enteroviruses responsible for disease outbreaks identified at these sites, should be considered a potential public health threat in spring recreation areas of Taiwan.

Genetic diversity of epidemic enterovirus 71 strains recovered from clinical and environmental samples in Taiwan.

Most enteroviruses excreted in human feces and urine are present in environmental water. In order to clarify the infection route of enterovirus, the detection of viruses in both clinical and environmental samples may contribute to understanding the mode of transmission of strains responsible for human infection. Thus, 21 epidemic enterovirus 71 strains from environmental water or stool samples were collected from HFMD children during 2005. Enterovirus genomic RNA was first amplified directly from clinical and environmental samples and then characterized by DNA sequencing and phylogenetic analysis. Results showed that these clinical strains share similar sequence identity (86.4-86.8%) to prototype BrCr based on the 5'-nontranslated region (NTR). However, environmental strains, except HME 77, share similar sequence identity (86.2-87.2%) to prototype BrCr. HME 77 showed higher sequence identity (90.1%). Results from phylogenetic analysis revealed that five environmental isolates were clustered as genogroup 3, which was closely related to a Taiwan outbreak in 1998 (AY055133). HME 77 was more closely related to a China epidemic isolate (AY895144), which belonged to genogroup 4. In contrast, all strains from clinical samples tested belonged to genogroup 3, which clustered with AY055133. In conclusion, there are two major epidemic clones (genogroups 3 and 4) prevalent in Taiwan since 2004 either in water or clinical patients.

Evaluation of enterovirus recovery in surface water by different adsorption and elution procedures.

The performance of six concentration method combinations and two quantitative analysis techniques were evaluated in terms of enterovirus recovery efficiencies by adjusting the pH and salt concentration of water samples. Of the six concentration method combinations, adsorption on nitrocellulose membranes followed by an acid rinse elution consistently gave the highest recovery efficiencies. In theory, an electropositive membrane should be the most appropriate technique for adsorption of electronegative viruses in pure water. However, it displayed the greatest loss in natural waters. For adsorption and elution procedures, both the electronegative membrane, accompanied by an acid rinse step, and the electropositive membrane, accompanied by a glycine rinse step, provided higher recovery efficiencies. MPN-RT-PCR, a statistically quantitative analysis method, performed more efficiently, in economic terms, but had a similar enterovirus recovery trend to real-time RT-PCR, which is the authoritative quantitative analysis method for nucleic acid.

Legionella prevalence in hot spring recreation areas of Taiwan.

Legionella is a bacterium ubiquitous to aquatic environments. Within the genus a few species are recognized as opportunistic potential human pathogens, especially the species Legionella pneumophila, which causes pneumonia legionellosis. Outbreaks of legionellosis are frequently reported by hotel guests and hospital patients, and are spread through inhaled aerosols of contaminated institutional water systems. Contaminations in hot tubs, spas and public baths are also possible. As a result, in this study, we investigated the distribution of Legionella at seven hot spring recreational areas throughout Taiwan. We gathered data on factors potentially associated with the pathogen's distribution, including environment, facility operation, and physical and microbiological water quality parameters. Spring water was collected from 91 sites and Legionella was detected in 21 (23%). The most frequently detected was L. pneumophila, followed by uncultured Legionella species, Legionella-like amoebal pathogen. Five species, L. bozemanii, L. dumoffi, L. feelei, L. lyticum and L. oakridgenesis, were all detected once. Legionella species were found in water temperatures ranging from 22 to 50 degrees C. Optimal pH appeared to be between 5.0 and 9.0. The prevalence of Legionella also coincided with the prevalence of indicator microorganisms. Legionella detection was not proportional to the frequency of cleaning. Results of this survey confirm the ubiquity of Legionella in Taiwan spring recreation areas. L. pneumophila, the organism responsible for the majority of legionellosis outbreaks, should be considered a potential public health threat in spa areas of Taiwan.

Using the flow cytometry to quantify the Giardia cysts and Cryptosporidium oocysts in water samples.

The flow cytometry (FC) has been used to detect Giardia cysts and Cryptosporidium oocysts quantitatively and instantaneously in this study. The experimental results showed that FC is potential to become a more precise method for the detection of Giardia and Cryptosporidium in water. This study also evaluated the staining efficiencies for three commercial antibodies. After staining Cryptosporidium oocysts with direct immunofluorescent antibodies in water samples, two populations were detected in the scatter-plots (FL1 versus SSC) of the FC. The Cryptosporidium oocysts and Giardia cysts are significantly separated from other particles while stained with direct immunofluorescent antibodies produced by Meridian Diagnostics and Waterborne Inc.