Antimicrobial Resistance, Virulence Determinants, and Biofilm Formation of Enterococcus Species From Ready-to-Eat Seafood

Enterococcus species form an important population of commensal bacteria and have been reported to possess numerous virulence factors considered significantly important in exacerbating diseases caused by them. The present study was designed to characterize antibiotic-resistant and virulent enterococci from ready-to-eat (RTE) seafood. A total of 720 RTE shrimp samples comprising sauced shrimp (n = 288), boiled shrimp (n = 216), and smoked shrimp (n = 216) obtained from open markets in Delta State, Nigeria, were assessed. Standard classical methods and polymerase chain reaction (PCR) were used in identifying the Enterococcus species. Potential virulence factors (β-hemolysis, gelatinase activity, S-layer, and biofilm formation) were assessed using standard procedures. The antibiotic susceptibility profile of the identified enterococci isolates was assayed using the Kirby–Bauer disc diffusion method. PCR was further used to screen selected antibiotic resistance and virulence genes. Prevalence of Enterococcus species from shrimp varieties is as follows: sauced, 26 (9.03%); boiled, 6 (2.78%); and smoked, 27 (12.50%), with an overall prevalence of 59 (8.19%) based on the occurrence of black hallow colonies after incubation. Enterococcus species detected include E. faecalis, 17 (28.8%); E. faecium, 29 (49.2%); E. gallinarum, 6 (10.2%); E. casseliflavus, 2 (3.4%); E. hirae, 3 (5.1%); and E. durans, 2 (3.4%). Biofilm occurrence among the shrimp varieties is as follows: 19/26 (73.1%) for sauced shrimps, 5/6 (83.3%) for boiled shrimps, and 16/27 (59.3%) for smoked shrimps. The phenotypic expression of the enterococci virulence revealed the following: S-layer, 59 (100%); gelatinase production, 19 (32.2%); and β-hemolysis, 21 (35.6%). An average of 3–11 virulence genes were detected in the Enterococcus species. The resistance profile of Enterococcus species is as follows: erythromycin, 29 (49.2%); vancomycin, 22 (37.3%); and tetracycline, 27 (45.8%). The frequency of occurrence of antibiotic resistance genes from the phenotypic resistant enterococci isolates to the macrolide, glycopeptide, and tetracycline antibiotics is as follows: ermA, 13/29 (44.8%); vanA, 14/22 (63.6%); tetA, 14/27 (51.9%); tetM, 15/27 (55.6%); ermB, 4/29 (13.8%); and vanB, 5/22 (22.7%). Findings from this study reveal the antibiotic resistance of enterococci strains of such species as E. durans, E. casseliflavus, E. gallinarum, and E. hirae. This study further revealed that RTE food products are reservoirs of potential virulent enterococci with antibiotic-resistant capabilities. This provides useful data for risk assessment and indicates that these foods may present a potential public health risk to consumers

Vibrio Fluvialis: An Unusual Enteric Pathogen of Increasing Public Health Concern

In developing countries, the fraction of treated wastewater effluents being discharged into watersheds have increased over the period of time, which have led to the deteriorations of the qualities of major rivers in developing nations. Consequently, high densities of disease causing bacteria in the watersheds are regularly reported including incidences of emerging Vibrio fluvialis. Vibrio fluvialis infection remains among those infectious diseases posing a potentially serious threat to public health. This paper addresses the epidemiology of this pathogen; pathogenesis of its disease; and its clinical manifestations in humans

Surveillance of invasive vibro species in discharged aqueous efflents of wastewater treatment plants in the Eastern Cape province of South Africa

Vibrio infections remain a serious threat to public health. In the last decade, Vibrio disease outbreaks have created a painful awareness of the personal, economic, societal, and public health costs associated with the impact of contaminated water in the aquatic milieu. This study was therefore designed to assess the prevalence of Vibrio pathogens in the final effluents of wastewater treatment plants (WWTPs) in the Eastern Cape Province, as well as their abilities to survive the treatment processes of the activated sludge system either as free cells or as plankton-associated entities in relation to the physicochemical qualities of the effluents. Three wastewater treatment facilities were selected to represent typical urban, sub-urban and rural communities, and samples were collected monthly from August 2007 to July 2008 from the final effluent, discharge point, 500 meter upstream and downstream of the discharge points and analysed for physicochemical parameters, Vibrio pathogens prevalence and their antibiogram characteristics using both culture based and molecular techniques. Physicochemical parameters measured include pH, temperature, electrical conductivity, salinity, turbidity, total dissolved solid (TDS), dissolved oxygen (DO), chemical oxygen demand (COD), nitrate, nitrite and orthophosphate levels. Unacceptably high levels of the assayed parameters were observed in many cases for COD (<10 - 1180 mg/l), nitrate (0.08 - 13.14 mg NO3- as N/l), nitrite (0.06 - 6.78 mg NO2- as N/l), orthophosphate (0.07-4.81 mg PO43- as P/l), DO (1.24 - 11.22 mg/l) and turbidity (2.04 -159.06 NTU). Temperature, COD and nitrite varied significantly with season (P < 0.05), while pH, EC, salinity, TDS, COD, and nitrate all varied significantly with sampling site (P < 0.01; P < 0.05). In the rural wastewater treatment facility, free-living Vibrio densities varied from 0 to 3.45 × 101 cfu ml-1, while the plankton-associated Vibrio densities vary with plankton sizes as follows: 180 μm (0 – 4.50 × 103 cfu ml-1); 60 μm (0 – 4.86 × 103 cfu ml-1); 20 μm (0 – 1.9 × 105 cfu ml-1). The seasonal variations in the Vibrio densities in the 180 and 60 μm plankton size samples were significant (P < 0.05), while the 20 μm plankton size and free-living vibrios densities were not. Molecular confirmation of the presumptive vibrios isolates revealed V. fluvialis (36.5 percent), as the predominant species, followed by V. vulnificus (34.6 percent), and V. parahaemolyticus (23.1 percent), and V. metschnikovii (5.8 percent) (detected using only API 20 NE), suggesting high incidence of pathogenic Vibrio species in the final effluent of the wastewater facility. Correlation analysis suggested that the concentration of Vibrio species correlated negatively with salinity and temperature (P < 0.001 and P < 0.002 respectively) as well as with pH and turbidity (P < 0.001), in the final effluent. Population density of total Vibrio ranged from 2.1 × 101 to 4.36 × 104 cfu ml-1 and from 2.80 ×101 to 1.80 × 105 cfu ml-1 for the sub-urban and urban communities treatment facilities respectively. Vibrio species associated with 180 μm, 60 μm, and 20 μm plankton sizes, were observed at densities of 0 - 1.36 × 103 cfu ml-1, 0 - 8.40 × 102 cfu ml-1 and 0 - 6.80 × 102 cfu ml-1 respectively at the sub-urban community‘s WWTP. In the urban community, counts of culturable vibrios ranged from 0 - 2.80 × 102 cfu ml-1 (180 μm); 0 - 6.60 × 102 cfu ml-1 (60 μm) and 0 -1.80 × 103 cfu ml-1 (20 μm). Abundance of free-living Vibrio species varied between 0 and the orders of 102 and 103 cfu ml-1 in the sub-urban and urban communities WWTPs respectively. Molecular confirmation of the presumptive vibrios isolates revealed the presence of V. fluvialis (41.38 percent), V. vulnificus (34.48 percent), and V. parahaemolyticus (24.14 percent) in the sub-urban community effluents. In the urban community V. fluvialis (40 percent), V. vulnificus (36 percent), and V. parahaemolyticus (24 percent) were detected. There was no significant correlation between Vibrio abundance and season, either as free-living or plankton-associated entities, while Vibrio species abundance correlated positively with temperature (r = 0.565; P < 0.01), salinity and dissolved oxygen (P < 0.05). Turbidity and pH showed significant seasonal variation (P < 0.05) in both locations. The Vibrio strains showed the typical multi-antibiotic-resistance of an SXT element. They were resistant to sulfamethoxazole (Sul), trimethoprim (Tmp), cotrimoxazole (Cot), chloramphenicol (Chl) and streptomycin (Str), as well as other antibiotics such as ampicillin (Amp), penicillin (Pen), erythromycin (Ery), tetracycline (Tet), nalidixic acid (Nal), and gentamicin (Gen). The antibiotic resistance genes detected includes dfr18 and dfrA1 for trimethoprim; tetA, strB, floR, sul2 blaP1, for tetracycline, streptomycin, chloramphenicol, sulfamethoxazole and β-lactams respectively. A number of these genes were only recently described from clinical isolates, demonstrating genetic exchange between clinical and environmental Vibrio species. This study revealed that there was an adverse impact on the physicochemical characteristics of the receiving watershed as a result of the discharge of inadequately treated effluents from the wastewater treatment facilities. The occurrence of Vibrio species as plankton-associated entities confirms the role of plankton as potential reservoir for this pathogen. Also the treated final effluents are reservoirs of various antibiotics resistance genes. This could pose significant health and environmental risk to the biotic component of the environment including communities that rely on the receiving water for domestic purposes and may also affect the health status of the aquatic milieu in the receiving water. There is need for consistent monitoring programme by appropriate regulatory agencies to ensure compliance of the wastewater treatment facilities to regulatory effluent quality standards

Anti-vibrio potentials of acetone and aqueous leaf extracts of Ocimum gratissimum (Linn)

Purpose: To evaluate the anti-vibrio potentials of acetone and aqueous leaf extracts of Ocimum gratissimum and determine its relevance in the treatment of vibrios infection.Methods: The agar-well diffusion method was used for screening the extracts for their anti-vibrio activity. Broth micro-dilution assay was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the extracts. Time-kill assay was used to assess bactericidal and/or bacteriostatic activity.Results: The acetone extract showed activity against 47.5 % (19/40) of the test bacteria, while the aqueous extract had activity against 30 % (12/40). MIC and MBC values range for the acetone extract were 0.625 – 5.0 mg/mL and 2.5 – 10 mg/mL respectively. The range of MIC exhibited by the antibiotic (gentamicin) against the vibrios is 0.002 mg/mL and &gt;0.256 mg/mL. Significant reduction in the bacterial density was at 2 × MIC after a 4 h interaction period, while bacterial density after 6 and 8 h interactions with extract was highly bactericidal. Growth inhibition and efficacy of the crude acetone extract were observed to be both concentration- and time-dependent.Conclusion: The bacteriostatic and bactericidal activities observed for Ocimum gratissimum leaf suggest that the plant is a potential source of bioactive components that may be effective in the treatment of vibrios infections.Keywords: Ocimum gratissimum, Vibrios infection, Antibiotics, Multi-drug resistance, Minimum inhibitory concentration, Minimum bactericidal concentration, Time kill assa

Multiple antibiotics resistant among environmental isolates of Stenotrophomonas maltophilia

In this study we assessed the functionality of integrons, melanin-like pigment and biofilm formation on multidrug resistance among environmental isolates of Stenotrophomonas maltophilia . Marked resistances were noted against aztreonam (60%), cefepime (68%), ceftazidime (77%), ciprofloxacin (72%), gentamicin (65%), meropenem (75%), piperacillin/tazobactam (65%) in S. maltophilia. Ticarcillin/clavulanic acid (66%) and trimethoprim/sulfamethoxazole (75%) were the active antibiotics against S. maltophilia. Class 1 intergron was significantly detected in 56.3% (54/96) of S. maltophilia strains. Integron-positive strains were significantly resistant to cefepime (69%), ceftazidime (78%), ciprofloxacin (74%), gentamicin (65%), and meropenem (72%). Gene cassettes arrays within integrons were identified as aminoglycoside resistance genes aacA4, aadA2, aadB, aacC4, and aacA6\u2019-Ib; \u3b2- lactams resistance genes blaIMP, blaOXA, and blaCARB; chloramphenicol resistance genes cmlA and catB2; quaternary ammonium compound (QAC) resistance genes smr and qac; and multi-gene cassettes: smr/aacA4 and blaIMP/aac6-II/aadA5. High-pigment-producing S. maltophilia strains revealed significant correlation with resistance to cefepime, ceftazidime, ciprofloxacin, levofloxacin and piperacillin/ tazobactam. Biofilm formation was not significant with resistance to ciprofloxacin, levofloxacin, meropenem, ticarcillin/clavulanic and trimethoprim /sulfamethoxazole. Our findings characterize the significant roles of integrons, melanin-like pigment and biofilm formation in the multidrug resistance of S. maltophilia. The range of antibiotics resistance genes and mobile genetic elements found suggests that the organism could potentially act as a reservoir of drug resistance determinants in environmental and clinical settings, which is an issue of public health concern

Development of Naphthalene Catabolism in Al and Fe contaminated soil

Metals have the potential to affect the biodegradation of polycyclic aromatic hydrocarbons (PAHs) in mixed contaminated soils. This investigation considered the effects of Al and Fe (50, 100, 250 and 500 mg/kg) on 14C-naphthalene biodegradation in soil over 63 d contact time. The presence of Al at low concentrations (50 and 100 mg/kg) stimulated (p &lt; 0.05) naphthalene biodegradation in fresh Al amendments; however, naphthalene mineralisation was significantly reduced (p &lt; 0.05) in 500 mg/kg aged Al amendment, but this reduction only occurred at 1 and 21 d in 500 mg/kg fresh Al. Naphthalene mineralisation was stimulated (p &lt; 0.05) in the highest Fe concentration (500 mg/kg) in both aged and fresh Fe amendments at 42 d. This suggests that Al has a greater impact than Fe on the development of PAH catabolism in soil. Studies on metal-PAH impact in soil facilitates the assessment of risk, hazard and bioremediation potential at sites contaminated with both contaminants

Evaluation of Antibiotic Resistant Gene in Abattoir Environment

Pseudomonas aeruginosa is a ubiquitous gram-negative pathogen with susceptibility to cause opportunistic infections in humans. Among the total of 75 isolated presumptive Pseudomonas aeruginosa, by cultural and morphology characteristic, 55 were identified as Pseudomonas aeruginosa by the specie-specific primer employing PCR assay which was more sensitive for confirmation of the isolates. Fifty-five Pseudomonas aeruginosa isolates were screened using PCR for the presence of integrons and associated resistance gene cassette. Thirty-four isolates harbored class 1 integrons (61.8%), of which 27 isolates (79.4%) carried gene cassettes. PCR was performed targeting the presence of these genes:- blaOXA 11(32.4%); blaIMP 5(14.7%); blaAMPC 5(14.7%); blaTEM 4(11.8%); and TetC 2(5.8%). Some of these genes were only recently described from clinical isolates, demonstrating genetic exchange between clinical and environmental Pseudomonas aeruginosa stains. Multi- resistance was observed in the isolates, revealing strong correlation between integron presence and multi-resistance. These results demonstrate that abattoir milieu is potential reservoirs of various antibiotics resistance genes, thus constituting a serious health risk to the communities dependent on the receiving water bodies

Antibiotic Synergy Interaction against Multidrug-Resistant Pseudomonas aeruginosa Isolated from an Abattoir Effluent Environment

Pseudomonas aeruginosa is an opportunistic pathogen in environmental waters with a high prevalence of multidrug resistance. In this study the synergistic efficacy of synergy antibiotic combinations in multidrug-resistant P. aeruginosa strains isolated from an abattoir effluent was investigated. Water samples were processed using membrane filtration; Pseudomonas was isolated with Pseudomonas Isolation Agar and confirmed using polymerase chain reaction with specie-specific primer. Susceptibility studies and in vitro synergy interaction testing were carried out, employing agar dilution and Etest procedure, respectively. Resistance was noted for clinically relevant antipseudomonal agents tested. Finding from antibiotic synergy interaction studies revealed that cefepime, imipenem, and meropenem combined with amikacin resulted in statistically significant (P < 0.0001) in vitro antibiotics synergy interaction, indicating the possible use of this regimen in treatment of pseudomonal infections

Polymyxin sensitivity/resistance cosmopolitan status, epidemiology and prevalence among O1/O139 and non-O1/non-O139 Vibrio cholerae : a meta-analysis

DATA AVAILABLE STATEMENT : The datasets/information used for this study are avail- able within the manuscript and the supplementary mate- rials.Resistance/sensitivity to polymyxin-B (PB) antibiotic has been employed as one among other epidemiologically relevant biotyping-scheme for Vibrio cholerae into Classical/El Tor biotypes. However, recent studies have re- vealed some pitfalls bordering on PB-sensitivity/resistance (PBR/S) necessitating study. Current study assesses the PBR/S cosmopolitan prevalence, epidemiology/distribution among O1/O139 and nonO1/nonO139 V. cholerae strains. Relevant databases (Web of Science, Scopus and PubMed) were searched to retrieve data from environ- mental and clinical samples employing the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). Random-effect-model (REM) and common-effect-model (CEM) of meta-analysis was performed to de- termine prevalence of PBR/S V. cholerae strains, describe the cosmopolitan epidemiological potentials and biotype relevance. Heterogeneity was determined by meta-regression and subgroup analyses. The pooled analyzed iso- lates from articles (7290), with sensitive and resistance are 2219 (30.44%) and 5028 (69.56%). Among these PB- sensitive strains, more than 1944 (26.67%) were O1 strains, 132 (1.81%) were nonO1 strains while mis-reported Classical biotype were 2080 (28.53) respectively indicating potential spread of variant/dual biotype. A significant PB-resistance was observed in the models (CEM = 0.66, 95% CI [0.65; 0.68], p -value = 0.001; REM = 0.83 [0.74; 0.90], p = 0.001) as both models had a high level of heterogeneity ( I2 = 98.0%; 2 =33 = 1755 . 09 , = 2 . 4932 ). Egger test ( z = 5.4017, p < 0.0001) reveal publication bias by funnel plot asymmetry. The subgroup analysis for continents (Asia, Africa) and sources (acute diarrhea) revealed (98% CI (0.73; 0.93); 55% CI (0.20; 0.86)), and 92% CI (0.67; 0.98). The Epidemiological prevalence for El tor/variant/dual biotype showed 88% CI (0.78; 0.94) with O1 strains at 88% CI (0.78; 0.94). Such global prevalence, distribution/spread of phenotypes/genotypes ne- cessitates updating the decades-long biotype classification scheme. An antibiotic stewardship in the post antibiotic era is suggestive/recommended. Also, there is need for holistic monitoring/evaluation of clinical/epidemiological relevance of the disseminating strains in endemic localities.African German Network of Excellence in Science 2022.www.elsevier.com/locate/imjhttp://www.elsevier.com/locate/imjam2024Microbiology and Plant PathologySDG-03:Good heatlh and well-bein

IN VITRO ASSESSMENT OF ANTIOXIDANT, PHYTOCHEMICAL AND NUTRITIONAL PROPERTIES OF EXTRACTS FROM THE LEAVES OF OCIMUM GRATISSIMUM (LINN)

The antioxidant, phytochemical and nutritional properties of acetone, methanol and aqueous extracts of the leaves of Ocimum gratissimum (Linn) were investigated to evaluate the therapeutic and nutritional potential of the leaves of this plant. The antioxidant of the plant extracts were assessed against 1,1-Diphenyl-2-picrylhydrazyl (DPPH) and 2,2-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) and ferric reducing agent. Total phenolics, flavonoids, flavonols and proanthocyanidins were determined to assess their corresponding effect on the antioxidant activity of this plant. The extracts exhibited DPPH and ABTS.+ radical scavenging activities, which was comparable to butylated hydroxytoluene (BHT). The phytochemical screening revealed the presence of alkaloids, tannins, saponin, steroids, cardiacglycoside, flavonoid, terpenoids and phenol. The proximate analysis confirms that the leaves contain appreciable amount of ash, crude protein, lipids, fibre and carbohydrates. The macro and micro elements and constituents revealed that the leaves contain significant amount of sodium, potassium, calcium, magnesium, iron, zinc, phosphorus, copper, nitrogen, and manganese. This study shows that the leaf can be used as a therapeutic agent and justifies its application in folkloric medicine