Virulence Gene Profiles Research Articles

Zoonotic potential and antimicrobial resistance of Escherichia spp. in urban crows in Japan-first detection of E. marmotae and E. ruysiae

Little is known about the prevalence of antimicrobial-resistant bacteria and pathogenic Escherichia coli in crows (carrion and jungle crows). We studied the phylogeny, virulence and antimicrobial resistance gene profiles of crow E. coli isolates to investigate their zoonotic potential and molecular epidemiology. During the winter of 2021–2022, 34 putative E. coli isolates were recovered from 27 of the 65 fresh fecal samples collected in urban areas. Three strains of the B1-O88:H8-ST446-fimH54 lineage, classified as extraintestinal pathogenic E. coli (ExPEC) and necrotoxigenic E. coli type 2, were colistin-resistant and harbored mcr-1.1-carrying IncI2 plasmids. The blaCTX-M-55 was identified in a multidrug-resistant B1-O non-typeable:H23-ST224-fimH39 strain. In phylogroup B2, two lineages of O6:H1-ST73-fimH30 and O6:H5-ST83-fimH21 were classified as ExPEC, uropathogenic E. coli, and necrotoxigenic E. coli type 1 (O6:H5-ST83-fimH21), and contained several virulence genes associated with avian pathogenic E. coli. Noteworthy is that three isolates, identified as E. coli by MALDI-TOF MS, were confirmed to be two Escherichia marmotae (cryptic clade V) and one Escherichia ruysiae (cryptic clade III) based on ANI and dDDH analyses. Our results provide the first evidence of these new species in crows. E. marmotae and E. ruysiae isolates in this study were classified as ExPEC and contained the enteroaggregative E. coli heat-stable toxin 1 gene. In addition, these two E. marmotae isolates displayed a close genetic relationship with human isolates associated with septicemia. This study provides the first insights into the prevalence and zoonotic significance of Escherichia spp. in urban crows in Japan, posing a significant risk for their transmission to humans.

Prevalence, virulence gene profile and antibiogram of Campylobacter jejuni from fresh vegetables in Mymensingh, Bangladesh

This study aimed to investigate Campylobacter jejuni, a major cause of food-borne bacterial infections worldwide, in fresh vegetables from five upzillas (Mymensingh, Trishal, Bhaluka, Muktagacha, and Fulbaria) in the Mymensingh district between July 2020 and April 2023. Using cultural, biochemical, and molecular techniques, 100 fresh vegetable samples (including tomato, carrot, cucumber, green chili, and coriander) were examined for C. jejuni. The isolates were further tested for virulence genes and antimicrobial susceptibility. Out of the 100 samples, 23% were confirmed as C. jejuni, by 16S rRNA gene-based polymerase chain reaction and all were found to be virulent with cytolethal distending toxins (cdtA, cdtB and cdtC genes). Antibiotic susceptibility testing revealed resistance to amoxicillin (47.83%), tetracycline (43.48%), and streptomycin (39.13%) among the isolates. However, ceftriaxone and ciprofloxacin were effective against 47.83% and 43.48% of the isolates, respectively. Moreover, 52.17% of the isolates were sensitive to erythromycin. Alarmingly, 34.78% of the C. jejuni isolates exhibited multidrug resistance (MDR) with eight different antibiotic resistance patterns, including four MDR patterns. These findings highlight the presence of virulent and antibiotic-resistant C. jejuni in fresh vegetables, emphasizing the need for monitoring and control to ensure food safety and public health issues. Asian Australas. J. Food Saf. Secur. 2023, 7 (2), 62-72

Diversity of Streptococcus spp. and genomic characteristics of Streptococcus uberis isolated from clinical mastitis of cattle in Bangladesh.

Streptococci are the major etiology in mastitis in dairy cattle, a cause of huge economic losses in the dairy industries. This study was aimed to determine the diversity of Streptococcus spp. isolated from clinical mastitis of cattle reared in Bangladesh. A total of 843 lactating cattle reared in four prominent dairy farms and one dairy community were purposively included in this study where 80 cattle were positive to clinical mastitis (CM) based on gross changes in the udder (redness, swelling, and sensitive udder) and/or milk (flakes and/or clots). Milk samples were collected from all the eighty cattle with clinical mastitis (CCM) and twenty five apparently healthy cattle (AHC). Samples were enriched in Luria Bertani broth (LB) and one hundred microliter of the enrichment culture was spread onto selective media for the isolation of Staphylococcus spp., Streptococcus spp., Enterococcus spp., Escherichia coli and Corynebacterium spp., the major pathogen associated with mastitis. Isolates recovered from culture were further confirmed by species specific PCR. Out of 105 samples examined 56.2% (59/105), 17.14% (18/105), 9.52% (10/105) and 22.9% (24/105) samples were positive for Staphylococcus, Streptococcus, Enterococcus faecalis and E. coli, respectively. This study was then directed to the determination of diversity of Streptococcus spp. through the sequencing of 16S rRNA. A total of eighteen of the samples from CCM (22.5%) but none from the AHC were positive for Streptococcus spp. by cultural and molecular examination. Sequencing and phylogenetic analysis of 16S rRNA identified 55.6, 33.3, 5.6 and 5.6% of the Streptococcus isolates as Streptococcus uberis, Streptococcus agalactiae, Streptococcus hyovaginalis and Streptococcus urinalis, respectively. Considering the high prevalence and worldwide increasing trend of S. uberis in mastitis, in-depth molecular characterization of S. uberis was performed through whole genome sequencing. Five of the S. uberis strain isolated in this study were subjected to WGS and on analysis two novel ST types of S. uberis were identified, indicating the presence of at least two different genotypes of S. uberis in the study areas. On virulence profiling, all the isolates harbored at least 35 virulence and putative virulence genes probably associated with intramammary infection (IMI) indicating all the S. uberis isolated in this study are potential mastitis pathogen. Overall findings suggest that Streptococcus encountered in bovine mastitis is diverse and S. uberis might be predominantly associated with CM in the study areas. The S. uberis genome carries an array of putative virulence factors that need to be investigated genotypically and phenotypically to identify a specific trait governing the virulence and fitness of this bacterium. Moreover, the genomic information could be used for the development of new genomic tools for virulence gene profiling of S. uberis.

Genomic typing and virulence gene profile analysis of Salmonella Derby from different sources

Salmonella enterica serovar Derby (S. Derby) is one of the most common Salmonella serovars which can infect poultry, swine, and humans. With the reduction of the sequencing cost and the improvement of sequencing technology, whole genome sequencing (WGS) has become an important method for bacterial determination, molecular investigation, and pathogenic tracing analysis. In this study, we investigated S. Derby isolates from different sources in China using in-silico multilocus sequence typing (MLST), core genome MLST (cgMLST) and whole genome MLST (wgMLST) analysis based on WGS. The results showed that 21 S. Derby strains were divided into 3 STs using MLST analysis, including ST40 (n = 19, accounting for 90.48%), ST71 (n = 1, accounting for 4.76%) and ST8016 (n = 1, accounting for 4.76%). cgMLST and wgMLST analysis categorized the tested strains into 13 cgSTs and 21 wgSTs, respectively. The minimum spanning trees of cgMLST and wgMLST both divided these strains into 3 clusters and 4 singletons. In addition, virulence gene profiles of S. Derby isolates were also analyzed, and a total of 174 virulence genes belonged to 8 categories were identified. In summary, we studied genomic typing, phylogenetic relationship and virulence gene profiles of S. Derby strains from different sources in China. These findings were beneficial for the epidemiology and pathogenesis of Salmonella.

Antimicrobial Resistance and Virulence Gene Profile of Clinical Staphylococcus aureus: A Multi-Center Study from Ethiopia.

Staphylococcus aureus causes a wide range of infections from mild skin and soft tissue to severe life-threatening bacteremia. The pathogenicity of S. aureus infections is related to various bacterial surface components and extracellular proteins such as toxic-shock syndrome (TSS) toxin and Panton-Valentine leukocidin (PVL). In this study we determine the antimicrobial resistance of isolated strains and their virulence genes in Ethiopia. A total of 190 archived S. aureus isolates from four Ethiopia Antimicrobial Resistance (AMR) Surveillance sites were analyzed. The identification of S. aureus was done by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF Biotyper) and antimicrobial susceptibility test (AST) was done using VITEK® 2. Multiplex PCR was used to detect mecA, mecC, pvl and spa genes and super-antigens (sea, seb, sec, seh and sej staphylococcal enterotoxins). A total of 172 isolates were confirmed as S. aureus, 9 (5.23%) were methicillin-resistant S. aureus (MRSA) and 163 (94.76%) were methicillin-susceptible S. aureus (MSSA). AST showed that 152 (88.4%) isolates were resistant to penicillin; 90 (52.32%) resistant to trimethoprim-sulfamethoxazole; and 45 (26.16%) resistant to tetracycline. A total of 66 (38.37%) isolates harbored at least one staphylococcal enterotoxin gene and 31 (46.96%) isolates had more than one. The most frequent enterotoxin gene encountered was seb 28 (16.28%). The TSST-1 gene was detected in 23 (13.37%). Presence of staphylococcal enterotoxin gene showed significant association with antibiotic resistance to cefoxitin, benzylpenicillin, oxacillin, erythromycin, clindamycin, tetracycline and SXT. The pvl gene was detected in 102 (59.3%) of isolates. Isolates from patients below 15 years of age showed significantly high numbers of pvl gene (P = 0.02). Presence of sej (P = 0.011) and TSST-1 (P <0.001) genes were associated with the presence of pvl gene. In this study, isolates were highly resistant to oral antibiotics and the pvl, seb, sea and TSST-1 genes were prevalent.

Phylogeny, Virulence, and Antimicrobial Resistance Gene Profiles of Enterococcus faecium Isolated from Australian Feedlot Cattle and Their Significance to Public and Environmental Health.

The extent of similarity between E. faecium strains found in healthy feedlot beef cattle and those causing extraintestinal infections in humans is not yet fully understood. This study used whole-genome sequencing to analyse the antimicrobial resistance profile of E. faecium isolated from beef cattle (n = 59) at a single feedlot and compared them to previously reported Australian isolates obtained from pig (n = 60) and meat chicken caecal samples (n = 8), as well as human sepsis cases (n = 302). The E. faecium isolated from beef cattle and other food animal sources neither carried vanA/vanB responsible for vancomycin nor possessed gyrA/parC and liaR/liaS gene mutations associated with high-level fluoroquinolone and daptomycin resistance, respectively. A small proportion (7.6%) of human isolates clustered with beef cattle and pig isolates, including a few isolates belonging to the same sequence types ST22 (one beef cattle, one pig, and two human isolates), ST32 (eight beef cattle and one human isolate), and ST327 (two beef cattle and one human isolate), suggesting common origins. This provides further evidence that these clonal lineages may have broader host range but are unrelated to the typical hospital-adapted human strains belonging to clonal complex 17, significant proportions of which contain vanA/vanB and liaR/liaS. Additionally, none of the human isolates belonging to these STs contained resistance genes to WHO critically important antimicrobials. The results confirm that most E. faecium isolated from beef cattle in this study do not pose a significant risk for resistance to critically important antimicrobials and are not associated with current human septic infections.

Listeria monocytogenes Strains Persisting in a Meat Processing Plant in Central Italy: Use of Whole Genome Sequencing and In Vitro Adhesion and Invasion Assays to Decipher Their Virulence Potential.

In this study, we used both a WGS and an in vitro approach to study the virulence potential of nine Listeria monocytogenes (Lm) strains belonging to genetic clusters persisting in a meat processing plant in Central Italy. The studied clusters belonged to CC1-ST1, CC9-ST9, and CC218-ST2801. All the CC1 and CC218 strains presented the same accessory virulence genes (LIPI-3, gltA, gltB, and aut_IVb). CC1 and CC9 strains presented a gene profile similarity of 22.6% as well as CC9 and CC218 isolates. CC1 and CC218 showed a similarity of 45.2% of the same virulence profile. The hypervirulent strains of lineage I (CC1 and CC218) presented a greater ability to adhere and invade Caco-2 cells than hypovirulent ones (CC9). CC1 strains were significantly more adhesive and invasive compared with CC9 and CC218 strains, although these last CCs presented the same accessory virulence genes. No statistically significant difference was found comparing CC218 with CC9 strains. This study provided for the first time data on the in vitro adhesiveness and invasiveness of CC218-ST2801 and added more data on the virulence characteristics of CC1 and CC9. What we observed confirmed that the ability of Lm to adhere to and invade human cells in vitro is not always decipherable from its virulence gene profile.

Stress Resistance and Virulence Gene Profiles Associated with Phylogeny and Phenotypes of Escherichia coli from Cattle

Seven serogroups of E. coli (Top seven E. coli) are frequently implicated in foodborne outbreaks in North America, largely due to their carriage of Shiga toxin genes (stx). This study aimed to profile resistance genes and virulence factors (VF), and their potential association with phylogeny and phenotypes of Top seven E. coli originating from cattle in Canada. 155 Top seven E. coli isolates previously characterized for heat and acid resistance and biofilm-forming ability were whole-genome sequenced and analyzed for phylogeny, VF, and stress resistance genes. The 155 E. coli strains belonged to six phylogroups: A (n = 32), B1 (n = 93), C (n = 3), D (n = 11), E (n = 15), and G (n = 1). Different phylogroups were clearly separated on the core genome tree, with strains of the same serotype closely clustered. The carriage of stx and the transmissible locus of stress tolerance (tLST), the extreme heat resistance marker, was mutually exclusive, in 33 and 15 genomes, respectively. A novel O84:H2 strain carrying stx1a was also identified. In total, 70, 41, and 32 VF, stress resistance genes and antibiotic resistance genes were identified. The stress resistance genes included those for metal (n = 29), biocides/acid (n = 4), and heat (n = 8) resistance. All heat resistance genes and most metal-resistance genes that were differentially distributed among the phylogroups were exclusively in phylogroup A. VF were least and most present in phylogroups A and D, respectively. No specific genes associated with acid resistance or biofilm formation phenotypes were identified. VF were more abundant (P < 0.05) in the non–biofilm-forming population and acid-resistant population.

Retrospective surveillance of viable Bacillus cereus group contaminations in commercial food and feed vitamin B2 products sold on the Belgian market using whole-genome sequencing.

Bacillus cereus is a spore-forming bacterium that occurs as a contaminant in food and feed, occasionally resulting in food poisoning through the production of various toxins. In this study, we retrospectively characterized viable B. cereus sensu lato (s.l.) isolates originating from commercial vitamin B2 feed and food additives collected between 2016 and 2022 by the Belgian Federal Agency for the Safety of the Food Chain from products sold on the Belgian market. In total, 75 collected product samples were cultured on a general medium and, in case of bacterial growth, two isolates per product sample were collected and characterized using whole-genome sequencing (WGS) and subsequently characterized in terms of sequence type (ST), virulence gene profile, antimicrobial resistance (AMR) gene profile, plasmid content, and phylogenomic relationships. Viable B. cereus was identified in 18 of the 75 (24%) tested products, resulting in 36 WGS datasets, which were classified into eleven different STs, with ST165 (n = 10) and ST32 (n = 8) being the most common. All isolates carried multiple genes encoding virulence factors, including cytotoxin K-2 (52.78%) and cereulide (22.22%). Most isolates were predicted to be resistant to beta-lactam antibiotics (100%) and fosfomycin (88.89%), and a subset was predicted to be resistant to streptothricin (30.56%). Phylogenomic analysis revealed that some isolates obtained from different products were closely related or even identical indicating a likely common origin, whereas for some products the two isolates obtained did not show any close relationship to each other or other isolates found in other products. This study reveals that potentially pathogenic and drug-resistant B. cereus s.l. can be present in food and feed vitamin B2 additives that are commercially available, and that more research is warranted to assess whether their presence in these types of products poses a threat to consumers.

Evidence of Antibiotic Resistance and Virulence Factors in Environmental Isolates of Vibrio Species.

The outbreak of waterborne diseases such as cholera and non-cholera (vibriosis) is continuously increasing in the environment due to fecal and sewage discharge in water sources. Cholera and vibriosis are caused by different species of Vibrio genus which are responsible for acute diarrheal disease and soft tissue damage. Although incidences of cholera and vibriosis have been reported from the Vaishali district of Bihar, India, clinical or environmental strains have not been characterized in this region. Out of fifty environmental water samples, twelve different biochemical test results confirmed the presence of twenty Vibrio isolates. The isolates were found to belong to five different Vibrio species, namely V. proteolyticus, V. campbellii, V. nereis, V. cincinnatiensis, and V. harveyi. From the identified isolates, 65% and 45% isolates were found to be resistant to ampicillin and cephalexin, respectively. Additionally, two isolates were found to be resistant against six and four separately selected antibiotics. Furthermore, virulent hlyA and ompW genes were detected by PCR in two different isolates. Additionally, phage induction was also noticed in two different isolates which carry lysogenic phage in their genome. Overall, the results reported the identification of five different Vibrio species in environmental water samples. The isolates showed multiple antibacterial resistance, phage induction, and virulence gene profile in their genome.

The Slaughterhouse as Hotspot of CC1 and CC6 Listeria monocytogenes Strains with Hypervirulent Profiles in an Integrated Poultry Chain of Italy.

In Europe, very few studies are available regarding the diversity of Listeria monocytogenes (L. monocytogenes) clonal complexes (CCs) and sequence types (ST) in poultry and on the related typing of isolates using whole genome sequencing (WGS). In this study, we used a WGS approach to type 122 L. monocytogenes strains isolated from chicken neck skin samples collected in two different slaughterhouses of an integrated Italian poultry company. The studied strains were classified into five CCs: CC1-ST1 (21.3%), CC6-ST6 (22.9%), CC9-ST9 (44.2%), CC121-ST121 (10.6%) and CC193-ST193 (0.8%). CC1 and CC6 strains presented a virulence gene profile composed of 60 virulence genes and including the Listeria Pathogenicity Island 3, aut_IVb, gltA and gltB. According to cgMLST and SNPs analysis, long-term persistent clusters belonging to CC1 and CC6 were found in one of the two slaughterhouses. The reasons mediating the persistence of these CCs (up to 20 months) remain to be elucidated, and may involve the presence and the expression of stress response and environmental adaptation genes including heavy metals resistance genes (cadAC, arsBC, CsoR-copA-copZ), multidrug efflux pumps (mrpABCEF, EmrB, mepA, bmrA, bmr3, norm), cold-shock tolerance (cspD) and biofilm-formation determinants (lmo0673, lmo2504, luxS, recO). These findings indicated a serious risk of poultry finished products contamination with hypervirulent L. monocytogenes clones and raised concern for the consumer health. In addition to the AMR genes norB, mprF, lin and fosX, ubiquitous in L. monocytogenes strains, we also identified parC for quinolones, msrA for macrolides and tetA for tetracyclines. Although the phenotypical expression of these AMR genes was not tested, none of them is known to confer resistance to the primary antibiotics used to treat listeriosis The obtained results increase the data on the L. monocytogenes clones circulating in Italy and in particular in the poultry chain.

The role of identified and characterized bacteriophage ZCEC13 in controlling pathogenic and multidrug-resistant Escherichia coli in wastewater: in vitro study

ABSTRACT The spread and development of Multi-Drug Resistant (MDR) bacteria in wastewater became beyond control and a global public health concern. The conventional disinfectants used in wastewater treatment methods have been becoming increasingly ineffective against a range of pathogenic and MDR bacteria. Bacteriophages are considered a novel approach to microbial control. Therefore, this study aims to explore the possibility of using phages against pathogenic and MDR Escherichia coli strains isolated from wastewater treatment plants. The wastewater samples were collected from two different treatment plants for E. coli isolation. The antibiotic sensitivity profile and occurrence of virulence and resistant genes were tested in 28 E. coli isolates. Phage ZCEC13 was selected based on its promising activity and host range to undergo identification and characterization. ZCEC13 was evaluated by transmission electron microscopy, genomic sequencing, in vitro lytic activity and tested for its stability under different conditions such as pH, Ultraviolet light exposure, and temperature. The results reported that ZCEC13 belongs to the Caudoviricetes class, with a high antibacterial dynamic. Phage ZCEC13 displayed high stability at different pH values ranging from 2 to 12, good tolerance to temperatures from −4 to 65°C, and high stability at UV exposure for 120 min. Respectively, the findings showed stability of the phage under several conditions and high efficiency in killing MDR bacteria isolated from the treatment plants. Further studies are encouraged to analyse the efficacy of phages as a microbial control agent in wastewater treatment plants.

Detection of multiple extensively-drug resistant hypervirulent Klebsiella pneumoniae clones from patients with ventilator-associated pneumonia in Egypt.

Introduction. Hypervirulent-K. pneumoniae (hvKP) is an evolving pathotype that is more virulent than the classical-K. pneumoniae (cKP) and causes serious fatal illnesses.Hypothesis/Gap Statement. Although there are few reports on hvKP isolated from Egyptian patients, the molecular characteristics and clonal relatedness of MDR-hvKP have not been adequately investigated.Aim. To investigate the microbiological and genetic characteristics as well as the epidemiology of hvKP induced ventilator-associated pneumonia (VAP).Methodology. A retrospective study of 59 K. pneumoniae inducing VAP was conducted at Assiut University Hospitals from November 2017 to January 2019. All K. pneumoniae were tested for resistance phenotype, capsular genotype (K1 and K2), virulence gene profile (c-rmpA, p-rmpA, iucA, kfu, iroB, iroN), and the presence of resistance genes (blaNDM-1, blaCTX-M-3-like, blaCTX-M-14-like). Clonal relatedness was assessed by Pulsed field gel electrophoresis (PFGE).Result. HvKP accounted for 89.8 % (53/59) of K. pneumoniae isolates with ~95 % exhibiting extensively-drug resistant (XDR) phenotype. Hypermucoviscous phenotype was detected in 19 (35.8 %) hvKP and K2 capsular gene was identified in 18 (33.9 %) of hvKP. Regarding the virulence genotype of hvKP strains, iucA was the most prevalent virulence gene (98.1%), while p-rmpA and kfu were detected in 75.4 and 52.8 % of hvKP strains, respectively. Resistance genes were highly prevalent in both cKP and hvKP with blaCTX-M-3-like being more prevalent in hvKP (100 % vs 94.3 % for blaNDM-1, 50 % vs 62.2 % for blaCTX-M-3-like and 83.3 % vs 69.8 % for blaCTX-M-14 -like, respectively). PFGE typing of 29 representative K. pneumoniae revealed 15 pulsotypes, with identical hvKP pulsotypes isolated from different ICUs at different times and several hvKP and cKP isolates belonged to the same pulsotype.Conclusion. This study highlights the dominance and clonal spread of XDR-hvKP strains at Assiut University Hospital in Egypt. Physicians should be aware of the increased risk of hvKP induced-VAP and support further epidemiologic studies.

Molecular identification, antimicrobial resistance and virulence gene profiling of Staphylococcus spp. associated with bovine sub-clinical mastitis in Bangladesh

This study was conducted to investigate the diversity and antimicrobial resistance profiling of Staphylococcus species causing sub-clinical mastitis (SCM) in dairy herds in Bangladesh as well as putative risk factors associated with the infections. Individual quarter milk samples were collected from a total of 284 lactating cows from 30 dairy farms were screened by means of California mastitis test; 178 (62.7%) of them had at least of quarter affected by SCM. After conventional microbiological isolation procedures, PCR tests were used for Staphylococcus species identification and detection of antimicrobial resistance and virulence genes. S. chromogenes (65.7%) was the most predominant species followed by, S. epidermidis (20.2%), S. haemolyticus (19.1%), S. aureus (15.7%), and S. sciuri (5.6%). High levels of antimicrobial resistance to ampicillin and amoxicillin/clavulanic acid were observed in S. aureus (82.1% and 75%) and S. sciuri (80% and 70%), while resistance to cefepime was markedly higher in S. chromogenes (95.7%), S. haemolyticus (94.1%), and S. epidermidis (97.2%). Multidrug resistance isolates were identified in all five species. The mecA gene was detected in S. aureus (32.1%) and S. chromogenes (5.98%). In addition, 20% S. sciuri and 17.7% S. haemolyticus carried the cytotoxin (pvl) gene, while 14.3% S. aureus harbored the toxic shock syndrome toxin (tst) gene. Multivariable logistic regression analysis identified “Old aged” (OR [CI]: 3.5 [1–12.4]); “Early stage of lactation” (OR [CI]: 3.4 [1.2–9.7]) and, “Firm udder condition” (OR [CI]: 4.2 [1.2–14.6]) as risk factors associated with SCM caused by S. aureus, S. chromogenes, and S. haemolyticus, respectively. Moreover, “Use of antimicrobials” (OR [CI]: 10.4 [3.4–32.1] and “History of previous clinical mastitis” (OR [CI]: 4.9 [1.2–19.7] for the carriage of methicillin-resistant Staphylococcus spp.

Phenotypic and genotypic comparison of pathogenic group B Streptococcus isolated from human and cultured tilapia (Oreochromis species) in Malaysia

Group B Streptococcus (GBS) is a major cause of several infectious diseases in humans and fish. This study was conducted to compare human and fish-derived GBS in terms of their antimicrobial susceptibility, serotype, virulence and pili genes and sequence type (ST), and to determine whether there is a potential linkage of zoonotic transmission in Malaysia. GBS isolated from humans and fish had similar phenotypic characteristics and differed in virulence gene profile, antimicrobial susceptibility, serotype and sequence type. Fish GBS isolates had lower genetic diversity and higher antibiotic susceptibility than human isolates. We report a rare detection of the potentially fish-adapted ST283 in human GBS isolates. Both human and fish ST283 shared several phenotypic and genotypic features, including virulence and pilus genes and antimicrobial susceptibility, illustrating the value of monitoring GBS within the One Health scope. In this study, two human GBS ST283 isolates belonging to the variant common in fish hosts were identified, raising awareness of the zoonotic potential between the different species in Malaysia.

Genomics, biofilm formation and infection of bladder epithelial cells in potentially uropathogenic Escherichia coli (UPEC) from animal sources and human urinary tract infections (UTIs) further support food-borne transmission

Escherichia coli is the main cause of urinary tract infections (UTI). While genomic comparison of specific clones recovered from animals, and human extraintestinal infections show high identity, studies demonstrating the uropathogenicity are lacking. In this study, comparative genomics combined with bladder-cell and biofilm formation assays, were performed for 31 E. coli of different origins: 7 from meat (poultry, beef, and pork); 2 from avian-farm environment; 12 from human uncomplicated UTI, uUTI; and 10 from human complicated UTI, cUTI. These isolates were selected based on their genetic uropathogenic (UPEC) status and phylogenetic background. In silico analysis revealed similar virulence-gene profiles, with flagella, type 1 and curli fimbriae, outer-membrane proteins (agn43, ompT, iha), and iron-uptake (iutA, entA, and fyuA) associated-traits as the most prevalent (>65%). In bladder-cell assays, moderate to strong values of association (83%, 60%, 77.8%) and invasion (0%, 70%, 55.5%) were exhibited by uUTI, cUTI, and animal-derived isolates, respectively. Of interest, uUTI isolates exhibited a significantly lower invasive capacity than cUTI isolates (p < 0.05). All isolates but one produced measurable biofilm. Notably, 1 turkey meat isolate O11:H6-F-ST457, and 2 cUTI isolates of the pandemic lineages O83:H42-F-ST1485-CC648 and O25b:H4-B2-ST131, showed strong association, invasion and biofilm formation. These isolates showed common carriage of type 1 fimbriae and csg operons, toxins (hlyF, tsh), iron uptake systems (iutA, entA, iroN), colicins, protectins (cvaC, iss, kpsM, traT), ompT, and malX. In summary, the similar in vitro behaviour found here for certain E. coli clones of animal origin would further reinforce the role of food-producing animals as a potential source of UPEC. Bladder-cell infection assays, combined with genomics, might be an alternative to in vivo virulence models to assess uropathogenicity.

Comparative study on genotypic properties of Pseudomonas aeruginosa isolated from subclinical mastitis in Türkiye

The results of antibacterial therapy in mastitis may be related to not only the antibiotic sensitivity of the etiological factors, but also to their abilities carrying integron and virulence genes. Integrons are mobile DNA element that can capture and carry genes, particularly those responsible for antibiotic resistance. Pseudomonas aeroginosa has numberless virulence factors which are contributed to bacterial invasion and toxicity. The aim of this study is to investigate antibiotic resistance, integron and virulence gene profiles of P. aeruginosa isolates obtained from cow milk with subclinical mastitis; further, was to evaluate the relationship between the antimicrobial resistance of bacteria with the of integron and virulence gene carrying. Material of the study is consisted with 32 (9.8%) of P. aeruginosa isolates obtained from 326 subclinical mastitic milk samples. After the bacterial identification was done by classical conventional methods, polymerase chain reaction (PCR) was used to verify the genus and species identifications of the isolates and to determine the integron and virulence gene profiles. Using disk diffusion method, antimicrobial resistance was determined to fifteen antibiotics against belonging to eleven antimicrobial families. The relationship the presence of integron and virulence genes associated with antibiotic resistance, further the relationship the presence of virulence genes and antimicrobial resistance was calculated with the Chi-Square (χ2) test. Ten of virulence genes (lasl, lasR, lasB, rhll, rhlR, rhlAB, plcH, plcN, ppyR, exoT) were found in all isolates whilst a virulence gene (aprA) was not present in any isolate. It was found that 34.4% of the isolates carried any integron gene. The results showed that the relationship is important between the presence of int genes and gentamicin, amikacin, tetracycline, cefoperazone, imipenem, aztreonam, ciprofloxacin, enrofloxacin resistance and exoU virulence gene presence. Also; there were also important associations between resistance to certain antibiotics and the presence of P. aeroginosa virulence genes: ciprofloxacin with lasA; netilmicin, cefoperazone and ciprofloxacin with pilB; netilmicin and ciprofloxacin with pelA; amikacin with algD, algU, algL; aztreonam with toxA; enrofloxacin with pvdA. All isolates obtained in this study showed multiple antibiotic resistance (MDR). In these cases, the presence of integron and some virulence genes could play a prominent role in the resistance of P. aeroginosa isolates to antimicrobial drugs. This study could be important in that it is the first study to show the presence of integrons and virulence genes in P. aeroginosa isolates originating from mastitic cow milk in Turkey. The presence of antibiotic-resistant P. aeroginosa strains in cattle farms may also pose a public health risk, as these bacteria can transmit their resistance genes to humans through food consumption.

High mortality risk of type III monomicrobial gram-negative necrotizing fasciitis: The role of extraintestinal pathogenic Escherichia coli (ExPEC) and Klebsiella pneumoniae

ObjectivesThe aim of this study was to investigate the prognostic value of reclassified new type III monomicrobial gram-negative necrotizing fasciitis (NF) and the microbial factors associated with an increased risk of mortality. MethodsThis study included 235 NF cases treated at National Taiwan University Hospital. We compared the mortality risk of NF caused by different causal microorganisms and examined the bacterial virulence genes profile and antimicrobial susceptibility pattern associated with an increase in mortality risk. ResultsType III NF (n = 68) had a mortality risk two-fold higher than type I (polymicrobial, n = 64) or type II (monomicrobial gram-positive, n = 79) NF (42.6% vs 23.4% or 19.0%, P = 0.019 and 0.002, respectively). Mortality differed by causal microorganism (Escherichia coli [61.5%], Klebsiella pneumoniae [40.0%], Aeromonas hydrophila [37.5%], Vibrio vulnificus [25.0%], polymicrobial [23.4%], group A streptococci [16.7%], and Staphylococcus aureus [16.2%], in decreasing rank, P <0.001). Type III NF caused by E. coli, identified as extraintestinal pathogenic E. coli (ExPEC) via virulence gene analyses, was associated with a particularly high mortality risk (adjusted odds ratio: 6.51, P = 0.003) after adjusting for age and comorbidities. Some (38.5%/7.7%) of the E. coli strains were non-susceptible to third/fourth-generation cephalosporins but remained susceptible to carbapenems. ConclusionType III NF, especially cases caused by E. coli or K. pneumoniae, are associated with a comparatively higher mortality risk than type I or type II NF. Wound gram stain-based rapid diagnosis of type III NF may inform empirical antimicrobial therapy to include a carbapenem.

Virulence and antimicrobial resistance genes profiles of spa type t605 methicillin-susceptible Staphylococcus aureus isolated from subclinical bovine mastitis.

Staphylococcus aureus is one of the most common pathogens associated with mastitis in dairy herds worldwide. This study evaluated the profile of virulence and antimicrobial resistance genes of spa type t605 methicillin-susceptible Staphylococcus aureus isolated from subclinical bovine mastitis in São Paulo, Brazil. A total of 57 S. aureus strains were screened by conventional PCR (Polymerase Chain Reaction) for 49 virulence genes. The most prevalent virulence genes detected were icaD (94.7%), fib (93%), fnbA (82.5%), clfA (80.7%), bap (78.9%), clfB (73.7%), icaA (66.7%), see (64.9%), and sed (61.4%). The blaZ (94.7%), aac6'aph2' (15.8%), and ant4 (12.3%) genes were the most common antimicrobial resistance genes; however, mecA and mecC genes were not found. All methicillin-susceptible S. aureus (MSSA) strains were characterized through spa and agr typing. The spa type t605 was found in all isolates. By agr typing, the most prevalent were type II (56.1%). Antimicrobial resistance was determined by the disk diffusion method, and 93% showed resistance to at least one antibiotic. Penicillin resistance was the most prevalent (87.7%), followed by tetracycline (12.3%), oxacillin (10.5%), and gentamicin (10.5%) resistance. Our study confirmed the spa type t605 as endemic, carrying a wide variety of virulence factors and high-level penicillin resistance. The profile seems to be associated with the colonization of MSSA and its persistence in subclinical mastitis.

Surveillance of Vibrio parahaemolyticus pathogens recovered from ready-to-eat foods

This study examined the occurrence of V. parahaemolyticus from ready-to-eat (RTE) food in Delta State, Nigeria. It also characterized antibiotic resistance and virulence gene profile patterns to determine the associated health risk hazard. Food samples total of 380 were collected randomly and assessed for V. parahaemolyticus. V. parahaemolyticus isolates were characterized for their virulence and antibiogram potentials using a phenotypic and polymerase chain reaction (PCR) approach. A total of 42 (11.1%) samples were contaminated with V. parahaemolyticus. In 17/42 (40.5%) of the V. parahaemolyticus-positive samples, the densities were < 10 MPN/g. However, 19/42 (45.2%) and 6/42 (14.3%) of the samples had densities of 10 – 102 and > 102 MPN/g, respectively. A total of 67 V. parahaemolyticus isolates were identified using PCR; 54(80.6%) isolates were multidrug resistant. A total of 22 (32.8%), 39 (58.2%), and 67 (100%) of the V. parahaemolyticus harbored the tdh, trh, and tlh toxin genes, respectively. The T3SS1 gene (vcrD1) was detected in 67 (100%) of the isolates. The T3SS2α genes which were vcrD2, vopB2, and vopT were detected in 21 (31.3%), 11 (16.4%) and 30 (44.8%) of the isolates respectively. Some of the V. parahaemolytics strains harbored the orf8 gene 20 (29.9%), and a combination of orf8 + tdh genes 12 (17.9%), categorized as pandemic strains. The antibiotic resistance genes detected in this study include blaTEM 33 (49.3), tetM 19 (28.4), cmlA 32(47.8) and sul1 14 (20.9). The concentration levels and prevalence of V. parahaemolyticus in RTE foods indicate contamination of ready-to-eat foods, particularly street foods consumed in the Delta State of Nigeria, threatening public health and consumer safety.