Variable freshwater influences on the abundance of Vibrio vulnificus in a tropical urban estuary

To better understand the controls on the opportunistic human pathogen Vibrio vulnificus in warm tropical waters, we conducted a year-long investigation in the Ala Wai Canal, a channelized estuary in Honolulu, HI. The abundance of V. vulnificus as determined by qPCR of the hemolysin gene (vvhA), varied spatially and temporally over four orders of magnitude (≤ 3 to 14,000 mL-1). Unlike in temperate and subtropical systems, temperatures were persistently warm (19–31°C) and explained little of the variability in V. vulnificus abundance. Salinity (1–36 ppt) had a significant, but non-linear, relationship with V. vulnificus abundance with highest abundances (> 2,500 mL-1) observed only at salinities from 7 to 22 ppt. V. vulnificus abundances were lower on average in the summer dry season when waters were warmer but more saline. Highest canal-wide average abundances were observed during a time of modest rainfall when moderate salinities and elevated concentrations of reduced nitrogen species and silica suggested a groundwater influence. Distinguishing the abundances of two genotypes of V. vulnificus (C-type and E-type) suggest that C-type strains, which are responsible for most human infections, were usually less abundant (25% on average), but their relative contribution was greater at higher salinities, suggesting a broader salinity tolerance. Generalized regression models suggested up to 67% of sample-to-sample variation in log-transformed V. vulnificus abundance was explained (n = 202) using the measured environmental variables, and up to 97% of the monthly variation in canal-wide average concentrations (n = 13) was explained with the best subset of four variables.

Characterization and Analysis of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) in Pandemic and Non-Pandemic Vibrio parahaemolyticus Isolates from Seafood Sources

Vibrio parahaemolyticus is one of the significant seafood-borne pathogens causing gastroenteritis in humans. Clustered regularly interspaced short palindromic repeats (CRISPR) are commonly detected in the genomes of V. parahaemolyticus and the polymorphism of CRISPR patterns has been applied as a genetic marker for tracking its evolution. In this work, a total of 15 pandemic and 36 non-pandemic V. parahaemolyticus isolates obtained from seafood between 2000 and 2012 were characterized based on hemolytic activity, antimicrobial susceptibility, and CRISPR elements. The results showed that 15/17 of the V. parahaemolyticus seafood isolates carrying the thermostable direct hemolysin gene (tdh+) were Kanagawa phenomenon (KP) positive. The Multiple Antibiotic Resistance (MAR) index ranged between 0.1 and 0.4, and 45% of the isolates have an MAR index ≥ 0.2. A total of 19 isolates were positive for CRISPR detection, including all tdh+ trh− isolates, two of tdh− trh+, and each of tdh+ trh+ and tdh− trh−. Four spacer types (Sp1 to Sp4) were identified, and CRISPR-positive isolates had at least one type of spacer homolog to the region of Vibrio alginolyticus megaplasmid. It is of interest that a specific CRISPR profile and spacer sequence type was observed with correlations to the hemolysin genotype (tdh/trh). Thus, these provide essential data on the exposure of foreign genetic elements and indicate shared ancestry within different genotypes of V. parahaemolyticus isolates.

Antibiofilm Activity of Phorbaketals from the Marine Sponge Phorbas sp. against Staphylococcus aureus

Biofilm formation by Staphylococcus aureus plays a critical role in the persistence of chronic infections due to its tolerance against antimicrobial agents. Here, we investigated the antibiofilm efficacy of six phorbaketals: phorbaketal A (1), phorbaketal A acetate (2), phorbaketal B (3), phorbaketal B acetate (4), phorbaketal C (5), and phorbaketal C acetate (6), isolated from the Korean marine sponge Phorbas sp. Of these six compounds, 3 and 5 were found to be effective inhibitors of biofilm formation by two S. aureus strains, which included a methicillin-resistant S. aureus. In addition, 3 also inhibited the production of staphyloxanthin, which protects microbes from reactive oxygen species generated by neutrophils and macrophages. Transcriptional analyses showed that 3 and 5 inhibited the expression of the biofilm-related hemolysin gene hla and the nuclease gene nuc1.

Study of Hemolysin Gene “aspHS” and Its Phenotype in Aspergillus Fumigatus

AIM: The main goal of this study was to analysis the “aspHS” gene and its phenotype in A. fumigatus. METHODS: Fifty-three A. fumigatus strains, including environmental, clinical and reference isolates, were used in this research. PCR was carried out based on Asp-hemolysin gene sequence. Two restriction enzymes TagI and NcoI were employed for digestion of PCR products. RESULTS: PCR products of 180 and 450 bp were generated for all A. fumigatus isolates. Digestion of the aspHS gene 180 bp amplicons with TagI and 450 bp amplicons with TagI and NcoI produced the expected bands for most isolates. Hemolysin production of A. fumigatus isolates was evaluated on sheep blood agar (SBA). CONCLUSION: In conclusion, our results provide evidence hemolysin activity and analysis of aspHS gene of A. fumigatus. These data may be useful in early diagnosis of A. fumigatus infections.

Elizabethkingia anophelisresponse to iron stress: physiologic, genomic, and transcriptomic analyses

Elizabethkingia anophelisbacteria encounter fluxes of iron in the midgut of mosquitoes, where they live as symbionts. They also establish bacteremia with severe clinical manifestations in humans, and live in water service lines in hospitals. In this study, we investigated the global gene expression responses ofE. anophelisto iron fluxes in the midgut of femaleAnopheles stephensimosquitoes fed sucrose or blood, and in iron-poor or iron-rich culture conditions. Of 3,686 transcripts revealed by RNAseq technology, 218 were upregulated while 112 were down-regulated under iron-poor conditions. Most of these differentially expressed genes (DEGs) were enriched in functional groups assigned within “biological process,” “cell component” and “molecular function” categories.E. anophelispossessed 4 iron/heme acquisition systems. Hemolysin gene expression was significantly repressed when cells were grown under iron-rich or high temperature (37°C) conditions. Furthermore, hemolysin gene expression was down-regulated after a blood meal, indicating thatE. anopheliscells responded to excess iron and its associated physiological stress by limiting iron loading. By contrast, genes encoding respiratory chain proteins were up-regulated under iron-rich conditions, allowing these iron-containing proteins to chelate intracellular free iron.In vivostudies showed that growth ofE. anopheliscells increased 3-fold in blood-fed mosquitoes over those in sucrose-fed ones. Deletion of aerobactin synthesis genes led to impaired cell growth in both iron-rich and iron-poor media. Mutants showed more susceptibility to H2O2toxicity and less biofilm formation than did wild-type cells. Mosquitoes withE. anophelisexperimentally colonized in their guts produced more eggs than did those treated with erythromycin or left unmanipulated, as controls. Results reveal thatE. anophelisbacteria respond to varying iron concentration in the mosquito gut, harvest iron while fending off iron-associated stress, contribute to lysis of red blood cells, and positively influence mosquito host fecundity.

Prevalence and Antibiotic Resistance of Listeria monocytogenes Isolated from Ready-to-Eat Foods in Turkey

The aim of the present study was the determination of the prevalence and antibiotic resistance of L. monocytogenes in ready-to-eat (RTE) foods in Ankara, Turkey. In order to detect and isolate L. monocytogenes from 201 RTE food samples, the EN ISO 11290:1 method was used. All isolates were identified using the polymerase chain reaction. The strains were also confirmed by the detection of the hemolysin gene (hlyA). The overall prevalence of L. monocytogenes was 8.5% among the food samples. Seventeen L. monocytogenes strains were examined by the disk diffusion assay for their resistance to 23 antibiotics. All strains were susceptible to erythromycin, clarithromycin, streptomycin, gentamicin, vancomycin, imipenem, trimethoprim, and chloramphenicol, while all strains were resistant to nalidixic acid, ampicillin, penicillin G, linezolid, and clindamycin. The higher resistance was found against oxacillin (94.1%), kanamycin (76.5%), levofloxacin (70.6%), and teicoplanin (64.7%), followed by amoxicillin/clavulanic acid (53.0%), rifampicin (47.1%), and ciprofloxacin (35.3%). A lower incidence of resistance was observed against tetracycline (5.9%), meropenem (5.9%), and trimethoprim/sulfamethoxazole (17.7%). All isolates were multidrug resistant showing resistance to at least three antibiotic classes. High L. monocytogenes prevalence among analyzed RTE foods represents a high risk for public health. Our findings show a high prevalence of L. monocytogenes in RTE foods in Turkey. More effective control strategies for L. monocytogenes are needed to reduce both prevalence and resistance of L. monocytogenes in Turkish RTE foods.